Patent Publication Number: US-2015065366-A1

Title: Biomarkers for Bladder Cancer and Methods Using the Same

Description:
CROSS-REFERENCE TO RELATED APPLICATIONS 
     This application claims the benefit of U.S. Provisional Patent Application No. 61/558,688, filed Nov. 11, 2011, and of U.S. Provisional Patent Application No. 61/692,738, filed Aug. 24, 2012, the entire contents of both of which are hereby incorporated herein by reference. 
    
    
     FIELD 
     The invention generally relates to biomarkers for bladder cancer and methods based on the same biomarkers. 
     BACKGROUND 
     In the US, more than 90% of bladder cancer (BCA) cases are transitional cell carcinomas (TCC), also referred to as urothelial carcinomas (UC). Approximately 70% of newly diagnosed TCC/UC patients have non-muscle invasive bladder cancer (NMIBC) tumors (i.e. T0a, T1 and CIS). The management of NMIBC patients involves the removal of visible tumors by transurethral resection of bladder tumor (TURB-T) and active surveillance for tumor recurrence as to minimize the risk of cancer progression. 
     Cystoscopy is considered the gold standard for diagnosis of bladder cancer and for monitoring patients with non-muscle invasive bladder cancer (NMIBC). The main limitations of this technique are the inability to visualize some areas of the urothelium and the difficulty to visualize carcinoma in situ (CIS) tumors. In both cases, the presence of tumors may be missed either due to tumor location in the upper urinary tract or because of the relatively normal appearance of the tumor in visible light cystoscopy. The detection of CIS has recently benefited from the introduction of fluorescent dyes injected intravesically before the cystoscopic examination. Although the rate of detection is increased, it requires a longer procedure (incubation of dyes after intravesical injection) and it is not yet used in the US on a routine basis. 
     Often, a cytology examination that can aid in the detection of bladder tumors not visible or poorly visible by cystoscopy is performed. Cytology has been used in routine clinical practice for more than 60 years. However, cytology is a complex method that has a high inter-operator variability. It is noteworthy that cytology is not a laboratory test but a consultation; an interpretation of the morphological features of exfoliated urothelial cells is assessed by each pathologist. Nevertheless, cytology has enjoyed the reputation of having a very high specificity and a great sensitivity for high grade tumors (i.e. TaG3, T1/G3 and CIS). 
     However, there is evidence that cytology performs poorly with low grade tumors (i.e. TaG1/G2) and the notion of high performance of cytology in high grade tumors has recently been challenged. For example, a study by the Mayo Clinic (n=75) showed that the overall sensitivity of cytology was 58% for all tumor types, 47% for Ta, only 78% for CIS and 60% for pT1-pT4). By comparison, the fluorescent in situ hybridization (FISH) analysis on the very same Mayo Clinic sample set had an overall sensitivity of 81%, with 65% for Ta, 100% for CIS and 95% for T1-T4 tumors (Halling K. et al. (2000) A comparison of cytology and fluorescence in situ hybridization for the detection of urothelial carcinoma. J. Urol. 164; 1768). 
     In another example, a different study (n=668) looked at the FDA-approved NMP22 test as an aid to cystoscopy for the assessment of recurrence in a series of consecutive patients with a history of bladder cancer at different institutions (Grossman H. B. et al. (2006) Surveillance for recurrent bladder cancer using a point-of-care proteomic assay.  JAMA  295; 299-305). Again, the study highlighted that cytology did not perform as well as previously thought in high grade tumors. Despite a better sensitivity of NMP22 (49.5%) compared to that of cytology (12.2%), the positive predictive value (PPV) of both tests was essentially the same at 41.5% highlighting the striking advantage cytology has in terms of specificity (99% for cytology, 87% for NMP22). In addition, a published review of several studies assessing the sensitivity/specificity of cytology re-affirmed the high specificity of cytology (0.99 with 95% CI of [0.83-0.997]) and its relatively poor sensitivity 0.34 (95% CI of [0.20-0.53]) (Lotan Y. and Roehrborn C. G. (2003) Sensitivity and specificity of commonly available bladder tumor markers versus cytology: results of a comprehensive literature review and meta-analysis.  Urology  61; 109-118.). 
     Nevertheless, cystoscopy with or without use of urine cytology is the current standard of care for diagnosis of bladder cancer in hematuria/dysuria patients and assessment of recurrence in NMIBC patients. However, cytology assessment can often be inconclusive and not fulfill its intended goal to aid in the diagnosis of bladder tumor. Also, a negative cytology result does not preclude the presence of a tumor (especially low stage/low grade tumor) given the low sensitivity of the cytology assessment. Furthermore, despite its low sensitivity, cytology has become the reference test against which all new tests are being compared. 
     Because of the limitations of cytology and the invasive nature of cystoscopy, there has been a search for biomarkers to provide a clinically useful non-invasive tool to detect bladder tumors while reducing costs associated with surveillance of NMIBC patients. There is a clinical need for a novel, non-invasive diagnostic test to aid cystoscopy and cytology for the initial diagnosis of bladder cancer and to aid in the detection of recurrent bladder cancer tumors in NMIBC patients. 
     Several FDA-approved urine-based markers such as Bladder Tumor Antigen, ImmunoCyt, Nuclear Matrix Protein-22, and Fluorescent In Situ Hybridization are available for that purpose. None of these tests rely on metabolite or biochemical biomarkers. Many of these tests have good sensitivity but inadequate specificity, which would lead to too many false-positive results if used in routine clinical practice. So far, the National Comprehensive Cancer Network (NCCN) Guidelines do not recommend the use of these tests outside the experimental protocol setting. 
     A urine-based test with a specificity equivalent to that of cytology and a sensitivity significantly superior to that of cytology would significantly impact clinical practice when used in conjunction with cystoscopy and/or cytology by improving the rate of bladder tumor detection while minimizing the number of false positive results. Such biomarkers could be used to aid the initial diagnosis of bladder cancer in symptomatic patients without a history of bladder cancer as well as aid in the assessment of bladder cancer recurrence. The biomarkers could be used in, for example, a urine test that quantitatively measures a panel of biomarker metabolites whose levels, when used with a specific algorithm, are indicative of the presence or absence of intravesical bladder tumors in a patient and aid in the initial diagnosis of bladder cancer in a population of patients with symptoms consistent with bladder cancer (i.e. hematuria/dysuria) and in the detection of bladder tumor recurrence in a population of patients with a history of NMIBC. Further, said biomarkers may be used in combination with a specific algorithm to form a diagnostic test that is indicative of tumor grade and stage. 
     SUMMARY 
     In one aspect, the present invention provides a method of diagnosing whether a subject has bladder cancer, comprising analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers for bladder cancer in the sample, where the one or more biomarkers are selected from Tables 1, 5, 7, 9, 11 and/or 13 and comparing the level(s) of the one or more biomarkers in the sample to bladder cancer-positive and/or bladder cancer-negative reference levels of the one or more biomarkers in order to diagnose whether the subject has bladder cancer. 
     In another aspect, the present invention also provides a method of determining whether a subject is predisposed to developing bladder cancer, comprising analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers for bladder cancer in the sample, where the one or more biomarkers are selected from Tables 1, 5, 7, 9, 11 and/or 13; and comparing the level(s) of the one or more biomarkers in the sample to bladder cancer-positive and/or bladder cancer-negative reference levels of the one or more biomarkers in order to determine whether the subject is predisposed to developing bladder cancer. 
     In yet another aspect, the invention provides a method of monitoring progression/regression of bladder cancer in a subject comprising analyzing a first biological sample from a subject to determine the level(s) of one or more biomarkers for bladder cancer in the sample, where the one or more biomarkers are selected from Tables 1, 5, 7, 9, 11 and/or 13 and the first sample is obtained from the subject at a first time point; analyzing a second biological sample from a subject to determine the level(s) of the one or more biomarkers, where the second sample is obtained from the subject at a second time point; and comparing the level(s) of one or more biomarkers in the first sample to the level(s) of the one or more biomarkers in the second sample in order to monitor the progression/regression of bladder cancer in the subject. 
     In a further aspect, the invention provides a method of distinguishing bladder cancer from other urological cancers (e.g., kidney cancer, prostate cancer), comprising analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers for bladder cancer in the sample where the one or more biomarkers are selected from Tables 1, 5, 7, 9, 11 and/or 13 and comparing the level(s) of the one or more biomarkers in the sample to bladder cancer-positive and/or bladder cancer-negative reference levels of the one or more biomarkers in order to distinguish bladder cancer from other urological cancers. 
     In another aspect, the present invention provides a method of determining whether a subject has a recurrence bladder cancer comprising analyzing, from a subject with a history of bladder cancer a biological sample to determine the level(s) of one or more biomarkers for bladder cancer selected from Tables 1, 5, 7, 9, 11 and/or 13; and comparing the level(s) of the one or more biomarkers in the sample to (a) bladder cancer-positive reference levels of the one or more biomarkers, and/or (b) bladder cancer-negative reference levels of the one or more biomarkers. 
     In another aspect, the present invention also provides a method of determining the stage of bladder cancer, comprising analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers for bladder cancer stage in the sample, where the one or more biomarkers are selected from Tables 5 and/or 9; and comparing the level(s) of the one or more biomarkers in the sample to high stage bladder cancer and/or low stage bladder cancer reference levels of the one or more biomarkers in order to determine the stage of the subject&#39;s bladder cancer. 
     In another aspect, the present invention provides a method of assessing the efficacy of a composition for treating bladder cancer comprising analyzing, from a subject having bladder cancer and currently or previously being treated with the composition, a biological sample to determine the level(s) of one or more biomarkers for bladder cancer selected from Tables 1, 5, 7, 9, 11 and/or 13; and comparing the level(s) of the one or more biomarkers in the sample to (a) levels of the one or more biomarkers in a previously-taken biological sample from the subject, where the previously-taken biological sample was obtained from the subject before being treated with the composition, (b) bladder cancer-positive reference levels of the one or more biomarkers, and/or (c) bladder cancer-negative reference levels of the one or more biomarkers. 
     In another aspect, the present invention provides a method for assessing the efficacy of a composition in treating bladder cancer, comprising analyzing a first biological sample from a subject to determine the level(s) of one or more biomarkers for bladder cancer selected from Tables 1, 5, 7, 9, 11 and/or 13, the first sample obtained from the subject at a first time point; administering the composition to the subject; analyzing a second biological sample from the subject to determine the level(s) of the one or more biomarkers, the second sample obtained from the subject at a second time point after administration of the composition; comparing the level(s) of one or more biomarkers in the first sample to the level(s) of the one or more biomarkers in the second sample in order to assess the efficacy of the composition for treating bladder cancer. 
     In yet another aspect, the invention provides a method of assessing the relative efficacy of two or more compositions for treating bladder cancer comprising analyzing, from a first subject having bladder cancer and currently or previously being treated with a first composition, a first biological sample to determine the level(s) of one or more biomarkers selected from Tables 1, 5, 7, 9, 11 and/or 13; analyzing, from a second subject having bladder cancer and currently or previously being treated with a second composition, a second biological sample to determine the level(s) of the one or more biomarkers; and comparing the level(s) of one or more biomarkers in the first sample to the level(s) of the one or more biomarkers in the second sample in order to assess the relative efficacy of the first and second compositions for treating bladder cancer. 
     In another aspect, the present invention provides a method for screening a composition for activity in modulating one or more biomarkers of bladder cancer, comprising contacting one or more cells with a composition; analyzing at least a portion of the one or more cells or a biological sample associated with the cells to determine the level(s) of one or more biomarkers of bladder cancer selected from Tables 1, 5, 7, 9, 11 and/or 13; and comparing the level(s) of the one or more biomarkers with predetermined standard levels for the biomarkers to determine whether the composition modulated the level(s) of the one or more biomarkers. 
     In a further aspect, the present invention provides a method for identifying a potential drug target for bladder cancer comprising identifying one or more biochemical pathways associated with one or more biomarkers for bladder cancer selected from Tables 1, 5, 7, 9, 11 and/or 13; and identifying a protein affecting at least one of the one or more identified biochemical pathways, the protein being a potential drug target for bladder cancer. 
     In yet another aspect, the invention provides a method for treating a subject having bladder cancer comprising administering to the subject an effective amount of one or more biomarkers selected from Tables 1, 5, 7, 9, 11 and/or 13 that are decreased in subjects having bladder cancer. 
    
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         FIG. 1  shows osmolality-normalized abundance ratios for exemplary metabolites between bladder cancer patients (TCC) and case control subjects. 
         FIG. 2  is a graphical illustration of feature-selected principal components analysis (PCA) using osmolality-normalized data separated subjects in this study. Arbitrary cutoff lines are drawn to illustrate that these metabolic abundance profiles can separate patients into groups with both high Negative Predictive Value (NPV) (PC1&lt;-1) and high Positive Predictive Value (PPV) (PC1&gt;1). The individuals with intermediate values (−1&lt;PC1&lt;1) could not be classified using this computational approach. 
         FIG. 3  is a graphical illustration of feature-selected hierarchical clustering (Pearson&#39;s correlation) using osmolality-normalized values separated subjects in this study. Three distinct metabolic classes were identified, one containing 100% control (TCC-free) individuals, one containing 100% bladder cancer (TCC) cases, and an intermediate case containing 33% controls and 67% TCC cases. 
         FIG. 4  is a graphical illustration of the Receiver Operator Characteristic (ROC) curve using the five exemplary biomarkers for bladder cancer as discussed in Example 7. 
         FIG. 5  is a graphical illustration of a ROC curve generated using seven exemplary biomarkers to distinguish bladder cancer from non-cancer, as discussed in Example 7. 
         FIG. 6  illustrates a comparison of AUC results obtained using the ridge model with multiple biomarkers to distinguish BCA from non-cancer, as discussed in Example 7. 
         FIG. 7  is a graphical illustration of a ROC curve generated using ridge logistic regression analysis to distinguish bladder cancer from hematuria, as discussed in Example 7. 
         FIG. 8  illustrates a comparison of AUC results obtained using the ridge model with multiple biomarkers to distinguish BCA from hematuria, as discussed in Example 7. 
         FIG. 9  is a graphical illustration of the Tricarboxylic Acid Cycle (TCA) and box plots of the levels of the biomarker metabolites measured in control individuals (left) and bladder cancer patients (right). The y-axis values indicate the scaled intensity of the biomarker. The top and bottom of the shaded box represent the 75 th  and 25 th  percentile, respectively. The top and bottom bars (“whiskers”) represent the entire spread of the data points for each compound and group, excluding “extreme” points, which are indicated with circles. The “+” indicates the mean value and the solid line indicates the median value. 
         FIG. 10  is a graphical illustration of biochemical pathways and box plots of metabolites that are indicative of activity of glycolysis, branched chain amino acid catabolism and fatty acid oxidation. The box plot on the left is the levels measured in control individuals and the box plot on the right is the levels measured in bladder cancer (TCC) patients. The y-axis values indicate the scaled intensity of the biomarker. The top and bottom of the shaded box represent the 75 th  and 25 th  percentile, respectively. The top and bottom bars (“whiskers”) represent the entire spread of the data points for each compound and group, excluding “extreme” points, which are indicated with circles. The “+” indicates the mean value and the solid line indicates the median value. 
     
    
    
     DETAILED DESCRIPTION 
     Currently available tests approved by the FDA are based on either protein or DNA techniques. The biochemical constituents in urine are commonly thought to be subject to dramatic variability both between individuals and within an individual over time. This variability has served as a barrier for examination of the constituents for their diagnostic prowess. The finding that many urine metabolites differentiate subjects having bladder cancer from subjects that do not have bladder cancer is novel and the fact that some are apparently produced while others are consumed from the urine minimizes the need for external normalizers of these data. The specific metabolites that are identified in the urine of a bladder cancer patient are in large part unexpected based on data published for other cancers (especially renal cancer). Likewise, using a similar approach, novel biomarkers have been identified in tissue samples from patients with bladder cancer. 
     The present invention relates to biomarkers of bladder cancer, methods for diagnosis or aiding in diagnosis of bladder cancer, methods of distinguishing bladder cancer from other urological cancers (e.g., prostate cancer, kidney cancer), methods of determining or aiding in determining predisposition to bladder cancer, methods of monitoring progression/regression of bladder cancer, methods of determining recurrence of bladder cancer, methods of staging bladder cancer, methods of assessing efficacy of compositions for treating bladder cancer, methods of screening compositions for activity in modulating biomarkers of bladder cancer, methods of identifying potential drug targets of bladder cancer, methods of treating bladder cancer, as well as other methods based on biomarkers of bladder cancer. Prior to describing this invention in further detail, however, the following terms will first be defined. 
     DEFINITIONS 
     “Biomarker” means a compound, preferably a metabolite, that is differentially present (i.e., increased or decreased) in a biological sample from a subject or a group of subjects having a first phenotype (e.g., having a disease) as compared to a biological sample from a subject or group of subjects having a second phenotype (e.g., not having the disease). A biomarker may be differentially present at any level, but is generally present at a level that is increased by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, by at least 95%, by at least 100%, by at least 110%, by at least 120%, by at least 130%, by at least 140%, by at least 150%, or more; or is generally present at a level that is decreased by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, by at least 95%, or by 100% (i.e., absent). A biomarker is preferably differentially present at a level that is statistically significant (i.e., a p-value less than 0.05 and/or a q-value of less than 0.10 as determined using either Welch&#39;s T-test or Wilcoxon&#39;s rank-sum Test). 
     The “level” of one or more biomarkers means the absolute or relative amount or concentration of the biomarker in the sample. 
     “Sample” or “biological sample” means biological material isolated from a subject. The biological sample may contain any biological material suitable for detecting the desired biomarkers, and may comprise cellular and/or non-cellular material from the subject. The sample can be isolated from any suitable biological tissue or fluid such as, for example, bladder tissue, blood, blood plasma, urine, or cerebral spinal fluid (CSF). 
     “Subject” means any animal, but is preferably a mammal, such as, for example, a human, monkey, mouse, rabbit or rat. 
     A “reference level” of a biomarker means a level of the biomarker that is indicative of a particular disease state, phenotype, or lack thereof, as well as combinations of disease states, phenotypes, or lack thereof. A “positive” reference level of a biomarker means a level that is indicative of a particular disease state or phenotype. A “negative” reference level of a biomarker means a level that is indicative of a lack of a particular disease state or phenotype. For example, a “bladder cancer-positive reference level” of a biomarker means a level of a biomarker that is indicative of a positive diagnosis of bladder cancer in a subject, and a “bladder cancer-negative reference level” of a biomarker means a level of a biomarker that is indicative of a negative diagnosis of bladder cancer in a subject. A “reference level” of a biomarker may be an absolute or relative amount or concentration of the biomarker, a presence or absence of the biomarker, a range of amount or concentration of the biomarker, a minimum and/or maximum amount or concentration of the biomarker, a mean amount or concentration of the biomarker, and/or a median amount or concentration of the biomarker; and, in addition, “reference levels” of combinations of biomarkers may also be ratios of absolute or relative amounts or concentrations of two or more biomarkers with respect to each other. Appropriate positive and negative reference levels of biomarkers for a particular disease state, phenotype, or lack thereof may be determined by measuring levels of desired biomarkers in one or more appropriate subjects, and such reference levels may be tailored to specific populations of subjects (e.g., a reference level may be age-matched so that comparisons may be made between biomarker levels in samples from subjects of a certain age and reference levels for a particular disease state, phenotype, or lack thereof in a certain age group). Such reference levels may also be tailored to specific techniques that are used to measure levels of biomarkers in biological samples (e.g., LC-MS, GC-MS, etc.), where the levels of biomarkers may differ based on the specific technique that is used. 
     “Non-biomarker compound” means a compound that is not differentially present in a biological sample from a subject or a group of subjects having a first phenotype (e.g., having a first disease) as compared to a biological sample from a subject or group of subjects having a second phenotype (e.g., not having the first disease). Such non-biomarker compounds may, however, be biomarkers in a biological sample from a subject or a group of subjects having a third phenotype (e.g., having a second disease) as compared to the first phenotype (e.g., having the first disease) or the second phenotype (e.g., not having the first disease). 
     “Metabolite”, or “small molecule”, means organic and inorganic molecules which are present in a cell. The term does not include large macromolecules, such as large proteins (e.g., proteins with molecular weights over 2,000, 3,000, 4,000, 5,000, 6,000, 7,000, 8,000, 9,000, or 10,000), large nucleic acids (e.g., nucleic acids with molecular weights of over 2,000, 3,000, 4,000, 5,000, 6,000, 7,000, 8,000, 9,000, or 10,000), or large polysaccharides (e.g., polysaccharides with a molecular weights of over 2,000, 3,000, 4,000, 5,000, 6,000, 7,000, 8,000, 9,000, or 10,000). The small molecules of the cell are generally found free in solution in the cytoplasm or in other organelles, such as the mitochondria, where they form a pool of intermediates which can be metabolized further or used to generate large molecules, called macromolecules. The term “small molecules” includes signaling molecules and intermediates in the chemical reactions that transform energy derived from food into usable forms. Examples of small molecules include sugars, fatty acids, amino acids, nucleotides, intermediates formed during cellular processes, and other small molecules found within the cell. 
     “Metabolic profile”, or “small molecule profile”, means a complete or partial inventory of small molecules within a targeted cell, tissue, organ, organism, or fraction thereof (e.g., cellular compartment). The inventory may include the quantity and/or type of small molecules present. The “small molecule profile” may be determined using a single technique or multiple different techniques. 
     “Metabolome” means all of the small molecules present in a given organism. 
     “Bladder cancer” (BCA) or “transitional cell carcinoma” (TCC) refers to a disease in which cancer develops in the bladder. As used herein both BCA and TCC are used interchangeably to indicate bladder cancer. 
     “Staging” of bladder cancer refers to an indication of how far the bladder tumor has spread. The tumor stage is used to select treatment options and to estimate a patient&#39;s prognosis. Bladder tumor staging ranges from T0 (no evidence of primary tumor, least advanced) to T4 (tumor has spread beyond fatty tissue surrounding the bladder into nearby organs, most advanced). Early stages of bladder cancer can also be characterized as carcinoma in situ (CIS) meaning that cells are abnormally proliferating but are still contained within the bladder. “Low stage” or “lower stage” bladder cancer refers to bladder cancer tumors, including malignant tumors with lower potential for recurrence, progression, invasion and/or metastasis (i.e. bladder cancer that is considered to be less aggressive). Cancer tumors that are confined to the bladder (i.e. non-muscle invasive bladder cancer, NMIBC) are considered to be less aggressive bladder cancer. “High stage” or “higher stage” bladder cancer refers to a bladder cancer tumor that is more likely to recur and/or progress and/or become invasive in a subject, including malignant tumors with higher potential for metastasis (bladder cancer that is considered to be more aggressive). Cancer tumors that are not confined to the bladder (i.e. muscle-invasive bladder cancer) are considered to be more aggressive bladder cancer. 
     “History of bladder cancer” refers to patients that previously had bladder cancer. 
     “Prostate cancer” (PCA) refers to a disease in which cancer develops in the prostate. 
     “Kidney Cancer” or “renal cell carcinoma” (RCC) refers to a disease in which cancer develops in the kidney. 
     “Urological Cancer” (UCA) refers to a disease in which cancer develops in the bladder, kidney and/or prostate. 
     “Hematuria” refers to a condition in which blood is present in the urine. 
     “Cytology” refers to an FDA-approved procedure that is part of the standard of care and used alongside, or as a reflex to, cystoscopy for the detection of recurrence or the diagnosis of bladder cancer. It identifies tumor cells based on morphologic characteristics. It is not a test per se but a pathology consultation based on urinary samples. The procedure is complex and requires expertise and care in sample collection to provide a correct assessment. Historically, the performance of cytology was described as extremely good with high-grade tumors but more recent studies have challenged that perception. On the other hand, all studies are in general agreement regarding the low sensitivity of cytology in low grade, low stage tumors (the bulk of the NMIBC tumors). Its two main assets are a long history of use in clinical practice (entrenched) and very high specificity (evaluated to be anywhere between 90 and 100% with many studies putting it at 99%). This provides the cytology consultation a great positive predictive value. This procedure is the one against which all other tests are currently evaluated, either for the purpose of replacing or aiding the cytology assessment. 
     “BCA Score” is a measure or indicator of bladder cancer severity, which is based on the bladder cancer biomarkers and algorithms described herein. A BCA Score will enable a physician to place a patient on a spectrum of bladder cancer severity from normal (i.e., no bladder cancer) to high (e.g., high stage or more aggressive bladder cancer). One of ordinary skill in the art will understand that the BCA Score can have multiple uses in the diagnosis and treatment of bladder cancer. For example, a BCA Score may also be used to distinguish low stage bladder cancer from high stage bladder cancer, and to monitor the progression and/or regression of bladder cancer. 
     I. Biomarkers 
     The bladder cancer biomarkers described herein were discovered using metabolomic profiling techniques. Such metabolomic profiling techniques are described in more detail in the Examples set forth below as well as in U.S. Pat. Nos. 7,005,255, 7,329,489; 7,550,258; 7,550,260; 7,553,616; 7,635,556; 7,682,783; 7,682,784; 7,910,301; 6,947,453; 7,433,787; 7,561,975; 7,884,318, the entire contents of which are hereby incorporated herein by reference. 
     Generally, metabolic profiles were determined for biological samples from human subjects that were positive for bladder cancer or samples from human subjects that were bladder cancer-negative (control cases). Exemplary controls include cancer-negative, healthy subject; cancer-negative, hematuria subject; bladder cancer negative, cancer subject. The metabolic profile for biological samples from a subject having bladder cancer was compared to the metabolic profile for biological samples from one or more other groups of subjects. Those molecules differentially present, including those molecules differentially present at a level that is statistically significant, in the metabolic profile of samples positive for bladder cancer as compared to another group (e.g., bladder cancer-negative samples) were identified as biomarkers to distinguish those groups. 
     The biomarkers are discussed in more detail herein. The biomarkers that were discovered correspond with biomarkers for distinguishing subjects having bladder cancer vs. control subjects not diagnosed with bladder cancer (see Tables 1, 5, 7, 9, 11 and/or 13). 
     Metabolic profiles were also determined for biological samples from human subjects diagnosed with high stage bladder cancer or human subjects diagnosed with low stage bladder cancer. The metabolic profile for biological samples from a subject having high stage bladder cancer was compared to the metabolic profile for biological samples from subjects with low stage bladder cancer. Those small molecules differentially present, including those small molecules differentially present at a level that is statistically significant, in the metabolic profile of samples from subjects with high stage bladder cancer as compared to another group (e.g., subjects not diagnosed with high stage bladder cancer) were identified as biomarkers to distinguish those groups. 
     The biomarkers are discussed in more detail herein. The biomarkers that were discovered correspond with biomarkers for distinguishing subjects having high stage bladder cancer vs. subjects having low stage bladder cancer (see Tables 5 and 9). 
     II. Methods 
     A. Diagnosis of Bladder Cancer 
     The identification of biomarkers for bladder cancer allows for the diagnosis of (or for aiding in the diagnosis of) bladder cancer in subjects presenting with one or more symptoms consistent with the presence of bladder cancer and includes the initial diagnosis of bladder cancer in a subject not previously identified as having bladder cancer and diagnosis of recurrence of bladder cancer in a subject previously treated for bladder cancer. A method of diagnosing (or aiding in diagnosing) whether a subject has bladder cancer comprises (1) analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers of bladder cancer in the sample and (2) comparing the level(s) of the one or more biomarkers in the sample to bladder cancer-positive and/or bladder cancer-negative reference levels of the one or more biomarkers in order to diagnose (or aid in the diagnosis of) whether the subject has bladder cancer. The one or more biomarkers that are used are selected from Tables 1, 5, 7, 9, 11 and/or 13 and combinations thereof. When such a method is used to aid in the diagnosis of bladder cancer, the results of the method may be used along with other methods (or the results thereof) useful in the clinical determination of whether a subject has bladder cancer. 
     Any suitable method may be used to analyze the biological sample in order to determine the level(s) of the one or more biomarkers in the sample. Suitable methods include chromatography (e.g., HPLC, gas chromatography, liquid chromatography), mass spectrometry (e.g., MS, MS-MS), enzyme-linked immunosorbent assay (ELISA), antibody linkage, other immunochemical techniques, and combinations thereof. Further, the level(s) of the one or more biomarkers may be measured indirectly, for example, by using an assay that measures the level of a compound (or compounds) that correlates with the level of the biomarker(s) that are desired to be measured. 
     The levels of one or more of the biomarkers of Tables 1, 5, 7, 9, 11 and/or 13 may be determined in the methods of diagnosing and methods of aiding in diagnosing whether a subject has bladder cancer. For example, one or more of the following biomarkers may be used alone or in combination to diagnose or aid in diagnosing bladder cancer: lactate, palmitoyl sphingomyelin, choline phosphate, succinate, adenosine, 1,2-propanediol, adipate, anserine, 3-hydroxybutyrate (BHBA), pyridoxate, acetylcarnitine, 2-hydroxybutyrate (AHB), kynurenine, tyramine, adenosine 5′-monophosphate (AMP), 3-hydroxyphenylacetate, 2-hydroxyhippurate (salicylurate), 3-indoxyl-sulfate, phenylacetylglutamine, p-cresol-sulfate, 3-hydroxyhippurate, itaconate methylenesuccinate, cortisol, isobutyrylglycine, gluconate, xanthurenate, gulono 1,4-lactone, cinnamoylglycine, 2-oxindole-3-acetate, alpha-CEHC-glucuronide, catechol-sulfate, gamma-glutamylphenylalanine, 2-isopropylmalate, 4-hydroxyphenylacetate, isovalerylglycine, carnitine, tartarate, 6-phosphogluconate, stearoyl sphingomyelin, myo-inositol, glucose, 3-(4-hydroxyphenyl)lactate, 1-linoleoylglycerol (1-monolinolein), pro-hydroxy-pro, gamma-glutamylglutamate, creatine, 5,6-dihydrouracil, docosadienoate (22:2n6), phenyllactate (PLA), propionlycarnitine, isoleucylproline, N2-methylguanosine, eicosapentanenoate (EPA 20:5n3), 5-methylthioadenosine (MTA), alpha-glutamyllysine, 3-phosphoglycerate, 6-keto prostaglandin F1alpha, docosatrienoate (22:3n3), 2-palmitoleoylglycerophosphocholine, 1-stearoylglycerophosphoinositol, 1-palmitoylglycerophosphoinositol, scyllo-inositol, dihomo-linoleate (20:2n6), 3-phosphoserine, docosapentaenoate (n6 DPA 22:5n6), 1-palmitoylglycerol and (1-monopalmitin). Additionally, for example, the level(s) of one biomarker, two or more biomarkers, three or more biomarkers, four or more biomarkers, five or more biomarkers, six or more biomarkers, seven or more biomarkers, eight or more biomarkers, nine or more biomarkers, ten or more biomarkers, etc., including a combination of all of the biomarkers in Tables 1, 5, 7, 9, 11 and/or 13 and any fraction thereof, may be determined and used in such methods. Determining levels of combinations of the biomarkers may allow greater sensitivity and specificity in diagnosing bladder cancer and aiding in the diagnosis of bladder cancer. For example, ratios of the levels of certain biomarkers (and non-biomarker compounds) in biological samples may allow greater sensitivity and specificity in diagnosing bladder cancer and aiding in the diagnosis of bladder cancer. 
     One or more biomarkers that are specific for diagnosing bladder cancer (or aiding in diagnosing bladder cancer) in a certain type of sample (e.g., urine sample or tissue plasma sample) may also be used. For example, when the biological sample is urine, one or more biomarkers listed in Tables 1, 5, 11 and/or 13, or any combination thereof, may be used to diagnose (or aid in diagnosing) whether a subject has bladder cancer. When the sample is bladder tissue, one or more biomarkers selected from Tables 7 and/or 9 may be used to diagnose (or aid in diagnosing) whether a subject has bladder cancer. 
     After the level(s) of the one or more biomarkers in the sample are determined, the level(s) are compared to bladder cancer-positive and/or bladder cancer-negative reference levels to aid in diagnosing or to diagnose whether the subject has bladder cancer. Levels of the one or more biomarkers in a sample matching the bladder cancer-positive reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of a diagnosis of bladder cancer in the subject. Levels of the one or more biomarkers in a sample matching the bladder cancer-negative reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of a diagnosis of no bladder cancer in the subject. In addition, levels of the one or more biomarkers that are differentially present (especially at a level that is statistically significant) in the sample as compared to bladder cancer-negative reference levels are indicative of a diagnosis of bladder cancer in the subject. Levels of the one or more biomarkers that are differentially present (especially at a level that is statistically significant) in the sample as compared to bladder cancer-positive reference levels are indicative of a diagnosis of no bladder cancer in the subject. 
     The level(s) of the one or more biomarkers may be compared to bladder cancer-positive and/or bladder cancer-negative reference levels using various techniques, including a simple comparison (e.g., a manual comparison) of the level(s) of the one or more biomarkers in the biological sample to bladder cancer-positive and/or bladder cancer-negative reference levels. The level(s) of the one or more biomarkers in the biological sample may also be compared to bladder cancer-positive and/or bladder cancer-negative reference levels using one or more statistical analyses (e.g., t-test, Welch&#39;s T-test, Wilcoxon&#39;s rank sum test, Random Forest, T-score, Z-score) or using a mathematical model (e.g., algorithm, statistical model). 
     For example, a mathematical model comprising a single algorithm or multiple algorithms may be used to determine whether a subject has bladder cancer. A mathematical model may also be used to distinguish between bladder cancer stages. An exemplary mathematical model may use the measured levels of any number of biomarkers (for example, 2, 3, 5, 7, 9, etc.) from a subject to determine, using an algorithm or a series of algorithms based on mathematical relationships between the levels of the measured biomarkers, whether a subject has bladder cancer, whether bladder cancer is progressing or regressing in a subject, whether a subject has high stage or low stage bladder cancer, etc. 
     The results of the method may be used along with other methods (or the results thereof) useful in the diagnosis of bladder cancer in a subject. 
     In one aspect, the biomarkers provided herein can be used to provide a physician with a BCA Score indicating the existence and/or severity of bladder cancer in a subject. The score is based upon clinically significantly changed reference level(s) for a biomarker and/or combination of biomarkers. The reference level can be derived from an algorithm. The BCA Score can be used to place the subject in a severity range of bladder cancer from normal (i.e. no bladder cancer) to high. The BCA Score can be used in multiple ways: for example, disease progression, regression, or remission can be monitored by periodic determination and monitoring of the BCA Score; response to therapeutic intervention can be determined by monitoring the BCA Score; and drug efficacy can be evaluated using the BCA Score. 
     Methods for determining a subject&#39;s BCA Score may be performed using one or more of the bladder cancer biomarkers identified in Tables 1, 5, 7, 9, 11 and/or 13 in a biological sample. The method may comprise comparing the level(s) of the one or more bladder cancer biomarkers in the sample to bladder cancer reference levels of the one or more biomarkers in order to determine the subject&#39;s BCA score. The method may employ any number of markers selected from those listed in Tables 1, 5, 7, 9, 11 and/or 13, including 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more markers. Multiple biomarkers may be correlated with bladder cancer, by any method, including statistical methods such as regression analysis. 
     After the level(s) of the one or more biomarker(s) is determined, the level(s) may be compared to bladder cancer reference level(s) or reference curves of the one or more biomarker(s) to determine a rating for each of the one or more biomarker(s) in the sample. The rating(s) may be aggregated using any algorithm to create a score, for example, a BCA score, for the subject. The algorithm may take into account any factors relating to bladder cancer including the number of biomarkers, the correlation of the biomarkers to bladder cancer, etc. 
     Additionally, in one embodiment, the biomarkers provided herein to diagnose or aid in the diagnosis of bladder cancer may be used to distinguish bladder cancer from hematuria in subjects presenting with hematuria. A method of distinguishing bladder cancer from hematuria in a subject comprises (1) analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers of bladder cancer in the sample and (2) comparing the level(s) of the one or more biomarkers in the sample to bladder cancer-positive and/or bladder cancer-negative reference levels of the one or more biomarkers in order to distinguish bladder cancer from hematuria. The one or more biomarkers that are used are selected from Tables 1, 5, 7, 9, 11 and/or 13. For example, one or more of the following biomarkers may be used alone or in any combination to distinguish bladder cancer from hematuria: xanthurenate, isovalerylglycine, 2-hydroxybutyrate (AHB), 4-hydroxyhippurate, gluconate, gulono 1,4-lactone, 3-hydroxyhippurate, tartarate, 2-oxindole-3-acetate, isobutyrylglycine, catechol-sulfate, phenylacetylglutamine, succinate, 3-hydroxybutyrate (BHBA), cinnamoylglycine, isobutyrylcarnitine, 3-hydroxyphenylacetate, 3-indoxyl-sulfate, sorbose, 2-5-furandicarboxylic acid, methyl-4-hydroxybenzoate, 2-isopropylmalate, adenosine 5′-monophosphate (AMP), 2-methylbutyrylglycine, palmitoyl-sphingomyelin, phenylpropionylglycine, beta-hydroxypyruvate, tyramine, 3-methylcrotonylglycine, carnosine, fructose, lactate, choline phosphate, adenosine, 1,2-propanediol, adipate, anserine, pyridoxate, acetylcarnitine, and kynurenine. When such a method is used to distinguish bladder cancer from hematuria, the results of the method may be used along with other methods (or the results thereof) useful in the clinical determination of distinguishing bladder cancer from hematuria. 
     In another embodiment, the biomarkers provided herein to diagnose or aid in the diagnosis of bladder cancer may be used to distinguish bladder cancer from other urological cancers. A method of distinguishing bladder cancer from other urological cancers in a subject comprises (1) analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers of bladder cancer in the sample and (2) comparing the level(s) of the one or more biomarkers in the sample to bladder cancer-positive and/or bladder cancer-negative reference levels of the one or more biomarkers in order to distinguish bladder cancer from other urological cancers. The one or more biomarkers that are used are selected from Tables 1 and/or 11. For example, one or more of the following biomarkers may be used alone or in any combination to distinguish bladder cancer from other urological cancers: imidazole-propionate, 3-indoxyl-sulfate, phenylacetylglycine, lactate, choline, methyl-indole-3-acetate, beta-alanine, palmitoyl-sphingomyelin, 2-hydroxyisobutyrate, succinate, 4-androsten-3beta-17beta-diol-disulfate-2,4-hydroxyphenylacetate, glycerol, uracil, gulono 1,4-lactone, phenol sulfate, dimethylarginine (ADMA+SDMA), cyclo-gly-pro, sucrose, adenosine, serine, azelate (nonanedioate), threonine, pregnanediol-3-glucuronide, ethanolamine, gluconate, N6-methyladenosine, N-methyl-proline, glycine, and glucose 6-phosphate (G6P), choline phosphate, 1,2-propanediol, adipate, anserine, 3-hydroxybutyrate (BHBA), pyridoxate, acetylcarnitine, 2-hydroxybutyrate, kynurenine, tyramine and xanthurenate. When such a method is used to distinguish bladder cancer from other urological cancers, the results of the method may be used along with other methods (or the results thereof) useful in the clinical determination of distinguishing bladder cancer from other urological cancers. 
     B. Methods of Determining Predisposition to Bladder Cancer 
     The identification of biomarkers for bladder cancer also allows for the determination of whether a subject having no symptoms of bladder cancer is predisposed to developing bladder cancer. A method of determining whether a subject having no symptoms of bladder cancer is predisposed to developing bladder cancer comprises (1) analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers listed in Tables 1, 5, 7, 9, 11 and/or 13 in the sample and (2) comparing the level(s) of the one or more biomarkers in the sample to bladder cancer-positive and/or bladder cancer-negative reference levels of the one or more biomarkers in order to determine whether the subject is predisposed to developing bladder cancer. The results of the method may be used along with other methods (or the results thereof) useful in the clinical determination of whether a subject is predisposed to developing bladder cancer. 
     As described above in connection with methods of diagnosing (or aiding in the diagnosis of) bladder cancer, any suitable method may be used to analyze the biological sample in order to determine the level(s) of the one or more biomarkers in the sample. 
     As with the methods of diagnosing (or aiding in the diagnosis of) bladder cancer described above, the level(s) of one biomarker, two or more biomarkers, three or more biomarkers, four or more biomarkers, five or more biomarkers, six or more biomarkers, seven or more biomarkers, eight or more biomarkers, nine or more biomarkers, ten or more biomarkers, etc., including a combination of all of the biomarkers in Tables 1, 5, 7, 9, 11 and/or 13 or any fraction thereof, may be determined and used in methods of determining whether a subject having no symptoms of bladder cancer is predisposed to developing bladder cancer. 
     After the level(s) of the one or more biomarkers in the sample are determined, the level(s) are compared to bladder cancer-positive and/or bladder cancer-negative reference levels in order to predict whether the subject is predisposed to developing bladder cancer. Levels of the one or more biomarkers in a sample matching the bladder cancer-positive reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of the subject being predisposed to developing bladder cancer. Levels of the one or more biomarkers in a sample matching the bladder cancer-negative reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of the subject not being predisposed to developing bladder cancer. In addition, levels of the one or more biomarkers that are differentially present (especially at a level that is statistically significant) in the sample as compared to bladder cancer-negative reference levels are indicative of the subject being predisposed to developing bladder cancer. Levels of the one or more biomarkers that are differentially present (especially at a level that is statistically significant) in the sample as compared to bladder cancer-positive reference levels are indicative of the subject not being predisposed to developing bladder cancer. 
     Furthermore, it may also be possible to determine reference levels specific to assessing whether or not a subject that does not have bladder cancer is predisposed to developing bladder cancer. For example, it may be possible to determine reference levels of the biomarkers for assessing different degrees of risk (e.g., low, medium, high) in a subject for developing bladder cancer. Such reference levels could be used for comparison to the levels of the one or more biomarkers in a biological sample from a subject. 
     As with the methods described above, the level(s) of the one or more biomarkers may be compared to bladder cancer-positive and/or bladder cancer-negative reference levels using various techniques, including a simple comparison, one or more statistical analyses, and combinations thereof. 
     As with the methods of diagnosing (or aiding in diagnosing) whether a subject has bladder cancer, the methods of determining whether a subject having no symptoms of bladder cancer is predisposed to developing bladder cancer may further comprise analyzing the biological sample to determine the level(s) of one or more non-biomarker compounds. 
     C. Methods of Monitoring Progression/Regression of Bladder Cancer 
     The identification of biomarkers for bladder cancer also allows for monitoring progression/regression of bladder cancer in a subject. A method of monitoring the progression/regression of bladder cancer in a subject comprises (1) analyzing a first biological sample from a subject to determine the level(s) of one or more biomarkers for bladder cancer selected from Tables 1, 5, 7, 9, 11 and/or 13 the first sample obtained from the subject at a first time point, (2) analyzing a second biological sample from a subject to determine the level(s) of the one or more biomarkers, the second sample obtained from the subject at a second time point, and (3) comparing the level(s) of one or more biomarkers in the first sample to the level(s) of the one or more biomarkers in the second sample in order to monitor the progression/regression of bladder cancer in the subject. For example, one or more of the following biomarkers may be used alone or in combination to monitor progression/regression of bladder cancer: 3-hydroxyphenylacetate, 3-hydroxyhippurate, 3-hydroxybutyrate (BHBA), isovalerylglycine, phenylacetylglutamine, pyridoxate, 2-5-furandicarboxylic acid, allantoin, pimelate (heptanedioate), lactate, adenosine 5′-monophosphate (AMP), catechol-sulfate, 2-hydroxybutyrate (AHB), isobutyrylglycine, 2-hydroxyhippurate (salicylurate), gluconate, imidazole-propionate, succinate, alpha-CEHC-glucuronide, 3-indoxyl-sulfate, 4-hydroxyphenylacetate, acetylcarnitine, xanthine, p-cresol-sulfate, tartarate, 4-hydroxyhippurate, 2-isopropylmalate, palmitoyl-sphingomyelin, adipate, and N(2)-furoyl-glycine, choline phosphate, adenosine, 1,2-propanediol, anserine, tyramine, xanthurenate, and kynurenine. The results of the method are indicative of the course of bladder cancer (i.e., progression or regression, if any change) in the subject. 
     The change (if any) in the level(s) of the one or more biomarkers over time may be indicative of progression or regression of bladder cancer in the subject. In order to characterize the course of bladder cancer in the subject, the level(s) of the one or more biomarkers in the first sample, the level(s) of the one or more biomarkers in the second sample, and/or the results of the comparison of the levels of the biomarkers in the first and second samples may be compared to bladder cancer-positive and bladder cancer-negative reference levels. If the comparisons indicate that the level(s) of the one or more biomarkers are increasing or decreasing over time (e.g., in the second sample as compared to the first sample) to become more similar to the bladder cancer-positive reference levels (or less similar to the bladder cancer-negative reference levels), then the results are indicative of bladder cancer progression. If the comparisons indicate that the level(s) of the one or more biomarkers are increasing or decreasing over time to become more similar to the bladder cancer-negative reference levels (or less similar to the bladder cancer-positive reference levels), then the results are indicative of bladder cancer regression. 
     In one embodiment, the assessment may be based on a BCA Score which is indicative of bladder cancer in the subject and which can be monitored over time. By comparing the BCA Score from a first time point sample to the BCA Score from at least a second time point sample, the progression or regression of bladder cancer can be determined. Such a method of monitoring the progression/regression of bladder cancer in a subject comprises (1) analyzing a first biological sample from a subject to determine a BCA score for the first sample obtained from the subject at a first time point, (2) analyzing a second biological sample from a subject to determine a second BCA score, the second sample obtained from the subject at a second time point, and (3) comparing the BCA score in the first sample to the BCA score in the second sample in order to monitor the progression/regression of bladder cancer in the subject. 
     The biomarkers and algorithms described herein may guide or assist a physician in deciding a treatment path, for example, whether to implement procedures such as surgical procedures (e.g., transurethral resection, radical cystectomy, segmental cystectomy), treat with drug therapy, or employ a watchful waiting approach. 
     As with the other methods described herein, the comparisons made in the methods of monitoring progression/regression of bladder cancer in a subject may be carried out using various techniques, including simple comparisons, one or more statistical analyses, mathematical models (algorithms) and combinations thereof. 
     The results of the method may be used along with other methods (or the results thereof) useful in the clinical monitoring of progression/regression of bladder cancer in a subject. 
     As described above in connection with methods of diagnosing (or aiding in the diagnosis of) bladder cancer, any suitable method may be used to analyze the biological samples in order to determine the level(s) of the one or more biomarkers in the samples. In addition, the level(s) one or more biomarkers, including a combination of all of the biomarkers in Tables 1, 5, 7, 9, 11 and/or 13 or any fraction thereof, may be determined and used in methods of monitoring progression/regression of bladder cancer in a subject. 
     Such methods could be conducted to monitor the course of bladder cancer in subjects having bladder cancer or could be used in subjects not having bladder cancer (e.g., subjects suspected of being predisposed to developing bladder cancer) in order to monitor levels of predisposition to bladder cancer. 
     D. Methods of Staging Bladder Cancer 
     The identification of biomarkers for bladder cancer also allows for the determination of bladder cancer stage of a subject. A method of determining the stage of bladder cancer comprises (1) analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers listed in Tables 5 and/or 9 in the sample and (2) comparing the level(s) of the one or more biomarkers in the sample to high stage bladder cancer and/or low stage bladder cancer reference levels of the one or more biomarkers in order to determine the stage of the subject&#39;s bladder cancer. The results of the method may be used along with other methods (or the results thereof) useful in the clinical determination of the stage of a subject&#39;s bladder cancer. 
     As described above in connection with methods of diagnosing (or aiding in the diagnosis of) bladder cancer, any suitable method may be used to analyze the biological sample in order to determine the level(s) of the one or more biomarkers in the sample. 
     The levels of one or more biomarkers listed in Tables 5 and 9 and combinations thereof may be determined in the methods of determining the stage of a subject&#39;s bladder cancer. For example, one or more of the following biomarkers may be used alone or in combination to determine the stage of bladder cancer: palmitoyl ethanolamide, palmitoyl sphingomyelin, thromboxane B2, bilirubin (Z,Z), adrenate (22:4n6), C-glycosyltryptophan, methyl-alpha-glucopyranoside, methylphosphate, 3-hydroxydecanoate, 3-hydroxyoctanoate, 4-hydroxyphenylpyruvate, N-acetylthreonine, 1-arachidonoylglycerophosphoinositol, 5,6-dihydrothymine, 2-hydroxypalmiate, coenzyme A, N-acetylserione, nicotinamide adenine dinucleotide (NAD+), docosatrienoate (22:3n3), glutathione reduced (GSH), prostaglandin A2, glutamine, glutamate gamma-methyl ester, docosapentaenoate (n6 DPA 22:5n6), glycochenodeoxycholate, hexanoylcarnitine, arachidonate (20:4n6), pro-hydroxy-pro, docosahexaenoate (DHA 22:6n3), laurylcarnitine, lactate, choline phosphate, succinate, adenosine, 1,2-propanediol, adipate, anserine, 3-hydroxybutyrate (BHBA), pyridoxate, acetylcarnitine, 2-hydroxybutyrate (AHB), kynurenine, tyramine and xanthurenate. Additionally, for example, the level(s) of one biomarker, two or more biomarkers, three or more biomarkers, four or more biomarkers, five or more biomarkers, six or more biomarkers, seven or more biomarkers, eight or more biomarkers, nine or more biomarkers, ten or more biomarkers, etc., including a combination of all of the biomarkers in Tables 5 and/or 9 or any fraction thereof, may be determined and used in methods of determining the stage of bladder cancer of a subject. 
     After the level(s) of the one or more biomarkers in the sample are determined, the level(s) are compared to low stage bladder cancer and/or high stage bladder cancer reference levels in order to determine the stage of bladder cancer of a subject. Levels of the one or more biomarkers in a sample matching the high stage bladder cancer reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of the subject having high stage bladder cancer. Levels of the one or more biomarkers in a sample matching the low stage bladder cancer reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of the subject having low stage bladder cancer. In addition, levels of the one or more biomarkers that are differentially present (especially at a level that is statistically significant) in the sample as compared to low stage bladder cancer reference levels are indicative of the subject not having low stage bladder cancer. Levels of the one or more biomarkers that are differentially present (especially at a level that is statistically significant) in the sample as compared to high stage bladder cancer reference levels are indicative of the subject not having high stage bladder cancer. 
     Studies were carried out to identify a set of biomarkers that can be used to determine the bladder cancer stage of a subject. In another embodiment, the biomarkers provided herein can be used to provide a physician with a BCA Score indicating the stage of bladder cancer in a subject. The score is based upon clinically significantly changed reference level(s) for a biomarker and/or combination of biomarkers. The reference level can be derived from an algorithm. The BCA Score can be used to determine the stage of bladder cancer in a subject from normal (i.e. no bladder cancer) to high stage bladder cancer. 
     The biomarkers and algorithms described herein may guide or assist a physician in deciding a treatment path, for example, whether to implement procedures such as surgical procedures (e.g., transurethral resection, radical cystectomy, segmental cystectomy), treat with drug therapy, or employ a watchful waiting approach. 
     As with the methods described above, the level(s) of the one or more biomarkers may be compared to high stage bladder cancer and/or low stage bladder cancer reference levels using various techniques, including a simple comparison, one or more statistical analyses, mathematical models (algorithms) and combinations thereof. 
     As with the methods of diagnosing (or aiding in diagnosing) whether a subject has bladder cancer, the methods of determining the stage of bladder cancer of a subject may further comprise analyzing the biological sample to determine the level(s) of one or more non-biomarker compounds. 
     E. Methods of Assessing Efficacy of Compositions for Treating Bladder Cancer 
     The identification of biomarkers for bladder cancer also allows for assessment of the efficacy of a composition for treating bladder cancer as well as the assessment of the relative efficacy of two or more compositions for treating bladder cancer. Such assessments may be used, for example, in efficacy studies as well as in lead selection of compositions for treating bladder cancer. 
     A method of assessing the efficacy of a composition for treating bladder cancer comprises (1) analyzing, from a subject having bladder cancer and currently or previously being treated with a composition, a biological sample to determine the level(s) of one or more biomarkers selected from Tables 1, 5, 7, 9, 11 and/or 13, and (2) comparing the level(s) of the one or more biomarkers in the sample to (a) level(s) of the one or more biomarkers in a previously-taken biological sample from the subject, wherein the previously-taken biological sample was obtained from the subject before being treated with the composition, (b) bladder cancer-positive reference levels of the one or more biomarkers, and (c) bladder cancer-negative reference levels of the one or more biomarkers. The results of the comparison are indicative of the efficacy of the composition for treating bladder cancer. 
     Thus, in order to characterize the efficacy of the composition for treating bladder cancer, the level(s) of the one or more biomarkers in the biological sample are compared to (1) bladder cancer-positive reference levels, (2) bladder cancer-negative reference levels, and (3) previous levels of the one or more biomarkers in the subject before treatment with the composition. 
     When comparing the level(s) of the one or more biomarkers in the biological sample (from a subject having bladder cancer and currently or previously being treated with a composition) to bladder cancer-positive reference levels and/or bladder cancer-negative reference levels, level(s) in the sample matching the bladder cancer-negative reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of the composition having efficacy for treating bladder cancer. Levels of the one or more biomarkers in the sample matching the bladder cancer-positive reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of the composition not having efficacy for treating bladder cancer. The comparisons may also indicate degrees of efficacy for treating bladder cancer based on the level(s) of the one or more biomarkers. 
     When the level(s) of the one or more biomarkers in the biological sample (from a subject having bladder cancer and currently or previously being treated with a composition) are compared to level(s) of the one or more biomarkers in a previously-taken biological sample from the subject before treatment with the composition, any changes in the level(s) of the one or more biomarkers are indicative of the efficacy of the composition for treating bladder cancer. That is, if the comparisons indicate that the level(s) of the one or more biomarkers have increased or decreased after treatment with the composition to become more similar to the bladder cancer-negative reference levels (or less similar to the bladder cancer-positive reference levels), then the results are indicative of the composition having efficacy for treating bladder cancer. If the comparisons indicate that the level(s) of the one or more biomarkers have not increased or decreased after treatment with the composition to become more similar to the bladder cancer-negative reference levels (or less similar to the bladder cancer-positive reference levels), then the results are indicative of the composition not having efficacy for treating bladder cancer. The comparisons may also indicate degrees of efficacy for treating bladder cancer based on the amount of changes observed in the level(s) of the one or more biomarkers after treatment. In order to help characterize such a comparison, the changes in the level(s) of the one or more biomarkers, the level(s) of the one or more biomarkers before treatment, and/or the level(s) of the one or more biomarkers in the subject currently or previously being treated with the composition may be compared to bladder cancer-positive reference levels, and/or to bladder cancer-negative reference levels. 
     Another method for assessing the efficacy of a composition in treating bladder cancer comprises (1) analyzing a first biological sample from a subject to determine the level(s) of one or more biomarkers selected from Tables 1, 5, 7, 9, 11 and/or 13, the first sample obtained from the subject at a first time point, (2) administering the composition to the subject, (3) analyzing a second biological sample from a subject to determine the level(s) of the one or more biomarkers, the second sample obtained from the subject at a second time point after administration of the composition, and (4) comparing the level(s) of one or more biomarkers in the first sample to the level(s) of the one or more biomarkers in the second sample in order to assess the efficacy of the composition for treating bladder cancer. As indicated above, if the comparison of the samples indicates that the level(s) of the one or more biomarkers have increased or decreased after administration of the composition to become more similar to the bladder cancer-negative reference levels, then the results are indicative of the composition having efficacy for treating bladder cancer. If the comparisons indicate that the level(s) of the one or more biomarkers have not increased or decreased after treatment with the composition to become more similar to the bladder cancer-negative reference levels (or less similar to the bladder cancer-positive reference levels) then the results are indicative of the composition not having efficacy for treating bladder cancer. The comparison may also indicate a degree of efficacy for treating bladder cancer based on the amount of changes observed in the level(s) of the one or more biomarkers after administration of the composition as discussed above. 
     A method of assessing the relative efficacy of two or more compositions for treating bladder cancer comprises (1) analyzing, from a first subject having bladder cancer and currently or previously being treated with a first composition, a first biological sample to determine the level(s) of one or more biomarkers selected from Tables 1, 5, 7, 9, 11 and/or 13, (2) analyzing, from a second subject having bladder cancer and currently or previously being treated with a second composition, a second biological sample to determine the level(s) of the one or more biomarkers, and (3) comparing the level(s) of one or more biomarkers in the first sample to the level(s) of the one or more biomarkers in the second sample in order to assess the relative efficacy of the first and second compositions for treating bladder cancer. The results are indicative of the relative efficacy of the two compositions, and the results (or the levels of the one or more biomarkers in the first sample and/or the level(s) of the one or more biomarkers in the second sample) may be compared to bladder cancer-positive reference levels, bladder cancer-negative reference levels to aid in characterizing the relative efficacy. 
     Each of the methods of assessing efficacy may be conducted on one or more subjects or one or more groups of subjects (e.g., a first group being treated with a first composition and a second group being treated with a second composition). 
     As with the other methods described herein, the comparisons made in the methods of assessing efficacy (or relative efficacy) of compositions for treating bladder cancer may be carried out using various techniques, including simple comparisons, one or more statistical analyses, and combinations thereof. An example of a technique that may be used is determining the BCA score for a subject. Any suitable method may be used to analyze the biological samples in order to determine the level(s) of the one or more biomarkers in the samples. In addition, the level(s) of one or more biomarkers, including a combination of all of the biomarkers in Tables 1, 5, 7, 9, 11 and/or 13 or any fraction thereof; may be determined and used in methods of assessing efficacy (or relative efficacy) of compositions for treating bladder cancer. 
     Finally, the methods of assessing efficacy (or relative efficacy) of one or more compositions for treating bladder cancer may further comprise analyzing the biological sample to determine the level(s) of one or more non-biomarker compounds. The non-biomarker compounds may then be compared to reference levels of non-biomarker compounds for subjects having (or not having) bladder cancer. 
     F. Methods of Screening a Composition for Activity in Modulating Biomarkers Associated with Bladder Cancer 
     The identification of biomarkers for bladder cancer also allows for the screening of compositions for activity in modulating biomarkers associated with bladder cancer, which may be useful in treating bladder cancer. Methods of screening compositions useful for treatment of bladder cancer comprise assaying test compositions for activity in modulating the levels of one or more biomarkers in Tables 1, 5, 7, 9, 11 and/or 13. Such screening assays may be conducted in vitro and/or in vivo, and may be in any form known in the art useful for assaying modulation of such biomarkers in the presence of a test composition such as, for example, cell culture assays, organ culture assays, and in vivo assays (e.g., assays involving animal models). 
     In one embodiment, a method for screening a composition for activity in modulating one or more biomarkers of bladder cancer comprises (1) contacting one or more cells with a composition, (2) analyzing at least a portion of the one or more cells or a biological sample associated with the cells to determine the level(s) of one or more biomarkers of bladder cancer selected from Tables 1, 5, 7, 9, 11 and/or 13; and (3) comparing the level(s) of the one or more biomarkers with predetermined standard levels for the one or more biomarkers to determine whether the composition modulated the level(s) of the one or more biomarkers. As discussed above, the cells may be contacted with the composition in vitro and/or in vivo. The predetermined standard levels for the one or more biomarkers may be the levels of the one or more biomarkers in the one or more cells in the absence of the composition. The predetermined standard levels for the one or more biomarkers may also be the level(s) of the one or more biomarkers in control cells not contacted with the composition. 
     In addition, the methods may further comprise analyzing at least a portion of the one or more cells or a biological sample associated with the cells to determine the level(s) of one or more non-biomarker compounds of bladder cancer. The levels of the non-biomarker compounds may then be compared to predetermined standard levels of the one or more non-biomarker compounds. 
     Any suitable method may be used to analyze at least a portion of the one or more cells or a biological sample associated with the cells in order to determine the level(s) of the one or more biomarkers (or levels of non-biomarker compounds). Suitable methods include chromatography (e.g., HPLC, gas chromatograph, liquid chromatography), mass spectrometry (e.g., MS, MS-MS), ELISA, antibody linkage, other immunochemical techniques, and combinations thereof. Further, the level(s) of the one or more biomarkers (or levels of non-biomarker compounds) may be measured indirectly, for example, by using an assay that measures the level of a compound (or compounds) that correlates with the level of the biomarker(s) (or non-biomarker compounds) that are desired to be measured. 
     G. Method of Identifying Potential Drug Targets 
     The identification of biomarkers for bladder cancer also allows for the identification of potential drug targets for bladder cancer. A method for identifying a potential drug target for bladder cancer comprises (1) identifying one or more biochemical pathways associated with one or more biomarkers for bladder cancer selected from Tables 1, 5, 7, 9, 11 and/or 13 and (2) identifying a protein (e.g., an enzyme) affecting at least one of the one or more identified biochemical pathways, the protein being a potential drug target for bladder cancer. 
     Another method for identifying a potential drug target for bladder cancer comprises (1) identifying one or more biochemical pathways associated with one or more biomarkers for bladder cancer selected from Tables 1, 5, 7, 9, 11 and/or 13 and one or more non-biomarker compounds of bladder cancer and (2) identifying a protein affecting at least one of the one or more identified biochemical pathways, the protein being a potential drug target for bladder cancer. 
     One or more biochemical pathways (e.g., biosynthetic and/or metabolic (catabolic) pathway) are identified that are associated with one or more biomarkers (or non-biomarker compounds). After the biochemical pathways are identified, one or more proteins affecting at least one of the pathways are identified. Preferably, those proteins affecting more than one of the pathways are identified. 
     A build-up of one metabolite (e.g., a pathway intermediate) may indicate the presence of a ‘block’ downstream of the metabolite and the block may result in a low/absent level of a downstream metabolite (e.g. product of a biosynthetic pathway). In a similar manner, the absence of a metabolite could indicate the presence of a ‘block’ in the pathway upstream of the metabolite resulting from inactive or non-functional enzyme(s) or from unavailability of biochemical intermediates that are required substrates to produce the product. Alternatively, an increase in the level of a metabolite could indicate a genetic mutation that produces an aberrant protein which results in the over-production and/or accumulation of a metabolite which then leads to an alteration of other related biochemical pathways and result in dysregulation of the normal flux through the pathway; further, the build-up of the biochemical intermediate metabolite may be toxic or may compromise the production of a necessary intermediate for a related pathway. It is possible that the relationship between pathways is currently unknown and this data could reveal such a relationship. 
     For example, the data indicates that metabolites in the biochemical pathways involving nitrogen excretion, amino acid metabolism, energy metabolism, oxidative stress, purine metabolism and bile acid metabolism are enriched in bladder cancer subjects. Further, polyamine levels are higher in cancer subjects, which indicates that the level and/or activity of the enzyme ornithine decarboxylase is increased. It is known that polyamines can act as mitotic agents and have been associated with free radical damage. These observations indicate that the pathways leading to the production of polyamines (or to any of the aberrant biomarkers) would provide a number of potential targets useful for drug discovery. 
     In another example, the data indicate that metabolites in the biochemical pathways involving lipid membrane metabolism, energy metabolism, Phase I and Phase II liver detoxification, and adenosine metabolism are enriched in bladder cancer subjects. Further, choline phosphate levels are higher in cancer subjects, which indicates that the level and/or activity of the sphingomyelinase enzymes are increased. These observations indicate that the pathways leading to the production of choline phosphate (or to any of the aberrant biomarkers) would provide a number of potential targets useful for drug discovery. 
     The proteins identified as potential drug targets may then be used to identify compositions that may be potential candidates for treating bladder cancer, including compositions for gene therapy. 
     H. Methods of Treating Bladder Cancer 
     The identification of biomarkers for bladder cancer also allows for the treatment of bladder cancer. For example, in order to treat a subject having bladder cancer, an effective amount of one or more bladder cancer biomarkers that are lowered in bladder cancer as compared to a healthy subject not having bladder cancer may be administered to the subject. The biomarkers that may be administered may comprise one or more of the biomarkers in Tables 1, 5, 7, 9, 11 and/or 13 that are decreased in bladder cancer. In some embodiments, the biomarkers that are administered are one or more biomarkers listed in Tables 1, 5, 7, 9, 11 and/or 13 that are decreased in bladder cancer and that have a p-value less than 0.10. In other embodiments, the biomarkers that are administered are one or biomarkers listed in Tables 1, 5, 7, 9, 11 and/or 13 that are decreased in bladder cancer by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, by at least 95%, or by 100% (i.e., absent). 
     In one example, sphingomyelinases that are present in the urine cleave sphingomyelin to form choline phosphate and creamide. Sphingomyelinase activity may be increased in bladder cancer subjects in order to process the abundance of sphingomyelin. When increased activity of an enzyme such as sphingomyelinase is associated with bladder cancer, administering an inhibitor for sphingomyelinase activity represents one possible method of treating bladder cancer. 
     III. Other Methods 
     Other methods of using the biomarkers discussed herein are also contemplated. For example, the methods described in U.S. Pat. No. 7,005,255, U.S. Pat. No. 7,329,489, U.S. Pat. No. 7,553,616, U.S. Pat. No. 7,550,260, U.S. Pat. No. 7,550,258, U.S. Pat. No. 7,635,556, U.S. patent application Ser. No. 11/728,826, U.S. patent application Ser. No. 12/463,690 and U.S. patent application Ser. No. 12/182,828 may be conducted using a small molecule profile comprising one or more of the biomarkers disclosed herein. 
     In any of the methods listed herein, the biomarkers that are used may be selected from those biomarkers in Tables 1, 5, 7, 9, 11 and/or 13 having p-values of less than 0.05. The biomarkers that are used in any of the methods described herein may also be selected from those biomarkers in Tables 1, 5, 7, 9, 11 and/or 13 that are decreased in bladder cancer (as compared to the control) or that are decreased in urological cancer (as compared to control) by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, by at least 95%, or by 100% (i.e., absent); and/or those biomarkers in Tables 1, 5, 7, 9, 11 and/or 13 that are increased in bladder cancer (as compared to the control or remission) or that are increased in remission (as compared to the control or bladder cancer) by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, by at least 95%, by at least 100%, by at least 110%, by at least 120%, by at least 130%, by at least 140%, by at least 150%, or more. 
     EXAMPLES 
     The invention will be further explained by the following illustrative examples that are intended to be non-limiting. 
     I. General Methods 
     A. Identification of Metabolic Profiles for Bladder Cancer 
     Each sample was analyzed to determine the concentration of several hundred metabolites. Analytical techniques such as GC-MS (gas chromatography-mass spectrometry) and LC-MS (liquid chromatography-mass spectrometry) were used to analyze the metabolites. Multiple aliquots were simultaneously, and in parallel, analyzed, and, after appropriate quality control (QC), the information derived from each analysis was recombined. Every sample was characterized according to several thousand characteristics, which ultimately amount to several hundred chemical species. The techniques used were able to identify novel and chemically unnamed compounds. 
     B. Statistical Analysis 
     The data was analyzed using T-tests to identify molecules (either known, named metabolites or unnamed metabolites) present at differential levels in a definable population or subpopulation (e.g., biomarkers for bladder cancer biological samples compared to control biological samples or compared to patients in remission from bladder cancer) useful for distinguishing between the definable populations (e.g., bladder cancer and control). Other molecules (either known, named metabolites or unnamed metabolites) in the definable population or subpopulation were also identified. 
     The data was also analyzed using one-way Analysis of Variance (ANOVA) contrasts to identify molecules (either known, named metabolites or unnamed metabolites) present at differential levels in a definable population or subpopulation (e.g., biomarkers for bladder cancer biological samples compared to control biological samples or compared to patients in remission from bladder cancer) useful for distinguishing between the definable populations (e.g., bladder cancer and control). ANOVA is a statistical model used to test that the means of multiple groups (≧2) are equal. The groups may be levels of a single variable (called a One Way ANOVA), or combinations of two, three or more variables (Two Way ANOVA, Three Way ANOVA, etc.). General variable effects are accessed via main effects and interaction terms. Contrasts, which test that a linear combination of the group means is equal to 0, can then be used to test more specific hypotheses. Unlike two sample t-tests, ANOVAs can handle repeated measurements/dependent observations. Other molecules (either known, named metabolites or unnamed metabolites) in the definable population or subpopulation were also identified. 
     Data was also analyzed using Random Forest Analysis. Random forests give an estimate of how well individuals in a new data set can be classified into existing groups. Random forest analysis creates a set of classification trees based on continual sampling of the experimental units and compounds. Then each observation is classified based on the majority votes from all the classification trees. In statistics, a classification tree classifies the observations into groups based on combinations of the variables (in this instance variables are metabolites or compounds). There are many variations on the algorithms used to create trees. A tree algorithm searches for the metabolite (compound) that provides the largest split between the two groups. This produces nodes. Then at each node, the metabolite that provides the best split is used and so on. If the node cannot be improved on, then it stops at that node and any observation in that node is classified as the majority group. 
     Random forests classify based on a large number (e.g. thousands) of trees. A subset of compounds and a subset of observations are used to create each tree. The observations used to create the tree are called the in-bag samples, and the remaining samples are called the out-of-bag samples. The classification tree is created from the in-bag samples, and the out-of-bag samples are predicted from this tree. To get the final classification for an observation, the “votes” for each group are counted based on the times it was an out-of-bag sample. For example, suppose observation 1 was classified as a “Control” by 2,000 trees, but classified as “Disease” by 3,000 trees. Using “majority wins” as the criterion, this sample is classified as “Disease.” 
     The results of the random forest are summarized in a confusion matrix. The rows correspond to the true grouping, and the columns correspond to the classification from the random forest. Thus, the diagonal elements indicate the correct classifications. A 50% error would occur by random chance for 2 groups, 66.67% error for three groups by random chance, etc. The “Out-of-Bag” (OOB) Error rate gives an estimate of how accurately new observations can be predicted using the random forest model (e.g., whether a sample is from a diseased subject or a control subject). 
     It is also of interest to see which variables are more “important” in the final classifications. The “importance plot” shows the top compounds ranked in terms of their importance. The mean decrease in accuracy measure is used to determine importance. The Mean Decrease Accuracy is computed as follows: For each tree in the random forest, the classification error based on the out-of-bag samples is computed. Then each variable (metabolite) is permuted, and the resulting error for each tree is computed. Then the average of the difference between the two errors is computed. Then this average is scaled by dividing by the standard deviation of these differences. The more important the variable, the higher the mean decrease accuracy. 
     Regression analysis was performed using the ridge logistic regression model. The ridge regression version of logistic regression puts a limit to the sum of the squared coefficients, i.e., if b1, b2, b3, etc are the coefficients for each metabolite, then ridge regression puts a limit on the sum of the squares of these (i.e., b1̂2+b2̂2+b3̂2+ . . . +bp̂2&lt;c). This bound forces many of the coefficients to drop to zero, hence this method also performs variable selection. 
     C. Biomarker Identification 
     Various peaks identified in the analyses (e.g. GC-MS, LC-MS, LC-MS-MS), including those identified as statistically significant, were subjected to a mass spectrometry based chemical identification process. 
     Example 1 
     Biomarkers for Bladder Cancer 
     Biomarkers were discovered by (1) analyzing urine samples from different groups of human subjects to determine the levels of metabolites in the samples and then (2) statistically analyzing the results to determine those metabolites that were differentially present in the two groups. 
     Two studies were carried out to identify biomarkers for bladder cancer. In study 1, 10 control urine samples that were collected from subjects that did not have bladder cancer, and 10 urine samples from subjects having bladder cancer (urothelial transitional cell carcinoma) were used for analysis. Age, race and gender were all tightly controlled to minimize the effects of confounding demographic-influenced variables. All subjects were Caucasian males. The average age of the bladder cancer cohort was 71.1 and the average age of the control cohort was 67.7. The paired t-test analysis p-value for age was 0.2 indicating that age was not significantly different between the two groups. 
     After the levels of metabolites were determined, the data was analyzed using univariate T-tests (i.e., Welch&#39;s T-test). As listed in Table 1 below, the analysis of named compounds resulted in the identification of biomarkers which were elevated in urine from bladder cancer patients compared to control subjects and biomarkers which were lower in urine from bladder cancer patients compared to control subjects. 
     Biomarkers were identified that were differentially present between urine samples from bladder cancer patients and control patients who were free of bladder cancer. Table 1, columns 1-3, list the identified biomarkers and includes, for each listed biomarker, the biochemical name of the biomarker, the fold change (FC) of the biomarker in cancer compared to non-cancer subjects (TCC/Control) which is the ratio of the mean level of the biomarker in cancer samples as compared to the control mean level, and the p-value determined in the statistical analysis of the data concerning the biomarkers (Table 1, columns 1-3). Column 10 of Table 1 lists the internal identifier for that biomarker compound in the in-house chemical library of authentic standards (CompID). Metabolites with an (*) indicate statistical significance (p≦0.1) in both the TCC/Control comparison (Study 1) and in the larger study described below (Study 2). Bold values indicate a fold change with a p-value of ≦0.1. Table 1 includes additional data, which is explained fully below. 
     
       
         
           
               
             
               
                 TABLE 1 
               
             
            
               
                   
               
               
                 Bladder Cancer Biomarkers in Urine 
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 TCC/Control 
                 BCA/Norm 
                 BCA/Hem 
                 BCA/RCC + 
                   
               
               
                   
                 (Study 1) 
                 (Study 2) 
                 (Study 2) 
                 PCA (Study 2) 
                 Comp 
               
            
           
           
               
               
               
               
               
               
               
               
               
               
            
               
                 Biochemical Name 
                 FC 
                 P-value 
                 FC 
                 P-value 
                 FC 
                 P-value 
                 FC 
                 P-value 
                 ID 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
               
            
               
                 anserine 
                   
                   
                 
                   0.23 
                 
                 0.0018 
                 
                   0.23 
                 
                 0.0001 
                 1.02 
                 0.7968 
                 15747 
               
               
                 pyridoxate (*) 
                 
                   0.3 
                 
                 0.0331 
                 
                   0.33 
                 
                 4.90E−05 
                 
                   0.5 
                 
                 0.0015 
                 0.91 
                 0.5014 
                 31555 
               
               
                 adipate 
                 1.72 
                 &gt;0.1 
                 
                   4.53 
                 
                 1.02E−05 
                 
                   4 
                 
                 0.0003 
                 1.07 
                 0.234 
                 21134 
               
               
                 xanthurenate (*) 
                 
                   0.56 
                 
                 0.0307 
                 
                   0.58 
                 
                 1.51E−09 
                 
                   0.69 
                 
                 1.74E−05 
                 0.89 
                 0.103 
                 15679 
               
               
                 1,2-propanediol 
                 1.83 
                 &gt;0.1 
                 
                   5.37 
                 
                 2.68E−07 
                 
                   5.95 
                 
                 0.0009 
                 
                   0.42 
                 
                 0.0016 
                 38002 
               
               
                 choline phosphate 
                   
                   
                 
                   6.35 
                 
                 3.81E−05 
                 
                   5.85 
                 
                 0.0004 
                 
                   4.54 
                 
                 2.74E−05 
                 34396 
               
               
                 acetylcarnitine 
                 0.66 
                 &gt;0.1 
                 
                   2.39 
                 
                 6.27E−06 
                 
                   2.45 
                 
                 2.09E−05 
                 0.99 
                 0.8071 
                 32198 
               
               
                 3-hydroxybutyrate (BHBA) (*) 
                 
                   3.19 
                 
                 0.0404 
                 
                   18.95 
                 
                 1.53E−08 
                 
                   19.58 
                 
                 2.15E−06 
                 0.54 
                 0.6446 
                 542 
               
               
                 palmitoyl sphingomyelin 
                   
                   
                 
                   10.24 
                 
                 3.32E−06 
                 
                   8 
                 
                 6.13E−05 
                 
                   5.29 
                 
                 3.69E−07 
                 37506 
               
               
                 tyramine 
                   
                   
                 
                   0.68 
                 
                 9.12E−06 
                 
                   0.56 
                 
                 1.28E−07 
                 1.02 
                 0.5284 
                 1603 
               
               
                 lactate 
                 1.93 
                 &gt;0.1 
                 
                   3.14 
                 
                 1.56E−11 
                 
                   1.41 
                 
                 0.0024 
                 
                   2.92 
                 
                 6.21E−09 
                 527 
               
               
                 2-isopropylmalate (*) 
                 
                   0.23 
                 
                 0.0678 
                 
                   0.29 
                 
                 1.25E−09 
                 
                   0.36 
                 
                 1.16E−06 
                 1.82 
                 0.1239 
                 15667 
               
               
                 isobutyrylglycine (*) 
                 
                   0.49 
                 
                 0.0362 
                 
                   0.61 
                 
                 4.81E−08 
                 
                   0.64 
                 
                 4.37E−06 
                 0.98 
                 0.4954 
                 35437 
               
               
                 L-urobilin (*) 
                 
                   13.62 
                 
                 0.0791 
                 
                   0.76 
                 
                 8.09E−05 
                 
                   0.62 
                 
                 0.0014 
                 1.01 
                 0.2537 
                 40173 
               
               
                 2-aminoadipate (*) 
                 
                   0.45 
                 
                 0.0532 
                 
                   0.65 
                 
                 0.0049 
                 
                   0.64 
                 
                 0.0032 
                 
                   1.02 
                 
                 0.0501 
                 6146 
               
               
                 sucralose (*) 
                 
                   7.96 
                 
                 0.053 
                 
                   0.4 
                 
                 0.0071 
                 
                   0.34 
                 
                 0.2694 
                 0.96 
                 0.7723 
                 36649 
               
               
                 N-acetylvaline (*) 
                 
                   0.78 
                 
                 0.0769 
                 
                   0.84 
                 
                 0.0079 
                 
                   0.84 
                 
                 0.0598 
                 
                   0.92 
                 
                 0.0814 
                 1591 
               
               
                 N-acetylisoleucine (*) 
                 
                   0.59 
                 
                 0.0898 
                 
                   0.81 
                 
                 0.014 
                 
                   0.81 
                 
                 0.0159 
                 0.96 
                 0.5669 
                 33967 
               
               
                 N1-Methyl-2-pyridone-5- 
                 
                   0.62 
                 
                 0.0612 
                 
                   0.91 
                 
                 0.015 
                 1.03 
                 0.8419 
                 1.27 
                 0.5826 
                 40469 
               
               
                 carboxamide (*) 
               
               
                 allantoin (*) 
                 
                   4.17 
                 
                 0.0348 
                 
                   0.59 
                 
                 0.034 
                 
                   0.66 
                 
                 0.0062 
                 1.28 
                 0.5641 
                 1107 
               
               
                 isobutyrylcarnitine (*) 
                 
                   0.58 
                 
                 0.0489 
                 
                   0.77 
                 
                 0.0002 
                 
                   0.85 
                 
                 0.0018 
                 1.26 
                 0.39 
                 33441 
               
               
                 xanthine (*) 
                 
                   0.19 
                 
                 0.0928 
                 
                   1.33 
                 
                 0.0006 
                 0.95 
                 0.2463 
                 1.09 
                 0.1774 
                 3147 
               
               
                 thymine (*) 
                 
                   0.68 
                 
                 0.0619 
                 
                   0.69 
                 
                 0.0033 
                 
                   0.7 
                 
                 0.002 
                 
                   0.64 
                 
                 0.0042 
                 604 
               
               
                 adenosine 5′-monophosphate 
                   
                   
                 
                   20.94 
                 
                 &lt;0.00001 
                 
                   9.89 
                 
                 2.16E−09 
                 4.82 
                 0.1116 
                 32342 
               
               
                 (AMP) 
               
               
                 3-hydroxyphenylacetate 
                 0.73 
                 &gt;0.1 
                 
                   0.28 
                 
                 3.00E−15 
                 
                   0.35 
                 
                 5.14E−08 
                 1.06 
                 0.3546 
                 1413 
               
               
                 2-hydroxyhippurate 
                 0.61 
                 &gt;0.1 
                 
                   0.13 
                 
                 2.83E−12 
                 
                   0.21 
                 
                 0.0004 
                 3.45 
                 0.2321 
                 18281 
               
               
                 (salicylurate) 
               
               
                 3-hydroxyhippurate 
                 0.61 
                 &gt;0.1 
                 
                   0.4 
                 
                 3.45E−12 
                 
                   0.53 
                 
                 1.42E−08 
                 1.67 
                 0.6012 
                 39600 
               
               
                 2-oxindole-3-acetate 
                 0.57 
                 &gt;0.1 
                 
                   0.46 
                 
                 2.04E−11 
                 
                   0.46 
                 
                 9.59E−10 
                 1.5 
                 0.2941 
                 40479 
               
               
                 phenylacetylglutamine 
                   
                   
                 
                   0.71 
                 
                 2.59E−11 
                 
                   0.69 
                 
                 7.00E−10 
                 
                   1.04 
                 
                 0.0636 
                 35126 
               
               
                 3-indoxyl sulfate 
                   
                   
                 
                   0.51 
                 
                 3.13E−11 
                 
                   0.56 
                 
                 5.47E−08 
                 
                   0.68 
                 
                 2.15E−06 
                 27672 
               
               
                 p-cresol sulfate 
                   
                   
                 
                   0.48 
                 
                 1.17E−10 
                 
                   0.61 
                 
                 7.40E−06 
                 0.92 
                 0.3052 
                 36103 
               
               
                 4-hydroxyphenylacetate 
                   
                   
                 
                   0.47 
                 
                 1.51E−09 
                 
                   0.49 
                 
                 5.34E−08 
                 
                   0.77 
                 
                 0.0012 
                 541 
               
               
                 2,3-dihydroxyisovalerate 
                 0.61 
                 &gt;0.1 
                 
                   0.27 
                 
                 1.28E−08 
                 
                   0.47 
                 
                 4.69E−05 
                 1.67 
                 0.2736 
                 38276 
               
               
                 catechol sulfate 
                 0.9 
                 &gt;0.1 
                 
                   0.65 
                 
                 4.50E−08 
                 
                   0.63 
                 
                 3.29E−07 
                 
                   1.85 
                 
                 0.0016 
                 35320 
               
               
                 gluconate 
                   
                   
                 
                   11.08 
                 
                 8.98E−08 
                 
                   11.59 
                 
                 3.32E−06 
                 
                   0.6 
                 
                 1.24E−06 
                 2913 
               
               
                 alpha-CEHC glucuronide 
                   
                   
                 
                   0.46 
                 
                 2.01E−07 
                 
                   0.72 
                 
                 0.0003 
                 
                   1.48 
                 
                 0.0862 
                 39346 
               
               
                 alpha-tocopherol 
                   
                   
                 
                   6.15 
                 
                 2.54E−07 
                 
                   5.31 
                 
                 4.61E−06 
                 
                   2.3 
                 
                 0.0007 
                 1561 
               
               
                 cinnamoylglycine 
                   
                   
                 
                   0.49 
                 
                 4.43E−07 
                 
                   0.47 
                 
                 1.09E−06 
                 1.35 
                 0.6862 
                 38637 
               
               
                 tartarate 
                   
                   
                 
                   0.24 
                 
                 2.58E−06 
                 
                   0.35 
                 
                 1.36E−05 
                 2.82 
                 0.8694 
                 15336 
               
               
                 phenylpropionylglycine 
                   
                   
                 
                   0.5 
                 
                 2.80E−06 
                 
                   0.47 
                 
                 1.38E−05 
                 1.1 
                 0.5694 
                 35434 
               
               
                 methyl-4-hydroxybenzoate 
                   
                   
                 
                   7.51 
                 
                 3.77E−06 
                 
                   8.88 
                 
                 5.16E−06 
                 
                   0.28 
                 
                 8.35E−07 
                 34386 
               
               
                 3,4-dihydroxyphenylacetate 
                 0.19 
                 &gt;0.1 
                 
                   0.46 
                 
                 3.99E−06 
                 
                   0.64 
                 
                 0.0001 
                 0.97 
                 0.786 
                 18296 
               
               
                 glucono-1,5-lactone 
                   
                   
                 
                   8.62 
                 
                 4.06E−06 
                 
                   5.88 
                 
                 0.0024 
                 1.08 
                 0.6635 
                 32355 
               
               
                 gamma- 
                 2.06 
                 &gt;0.1 
                 
                   1.49 
                 
                 7.92E−06 
                 1.17 
                 0.1496 
                 
                   1.18 
                 
                 0.0199 
                 33422 
               
               
                 glutamylphenylalanine 
               
               
                 isovalerylglycine 
                   
                   
                 
                   0.56 
                 
                 8.21E−06 
                 
                   0.49 
                 
                 5.16E−09 
                 0.91 
                 0.4253 
                 35107 
               
               
                 fructose 
                 0.69 
                 &gt;0.1 
                 
                   0.55 
                 
                 8.32E−06 
                 
                   0.51 
                 
                 5.49E−07 
                 1.49 
                 0.2161 
                 577 
               
               
                 sorbose 
                   
                   
                 
                   0.58 
                 
                 8.78E−06 
                 
                   0.42 
                 
                 1.90E−08 
                 
                   2.21 
                 
                 0.0573 
                 563 
               
               
                 guanidine 
                   
                   
                 
                   0.5 
                 
                 1.28E−05 
                 
                   0.53 
                 
                 0.0015 
                 0.87 
                 0.2724 
                 22287 
               
               
                 pimelate (heptanedioate) 
                 0.7 
                 &gt;0.1 
                 
                   0.51 
                 
                 1.69E−05 
                 
                   0.62 
                 
                 0.0005 
                 0.88 
                 0.3598 
                 15704 
               
               
                 hexanoylglycine 
                 1.47 
                 &gt;0.1 
                 
                   1.62 
                 
                 2.02E−05 
                 
                   1.71 
                 
                 0.0022 
                 0.69 
                 0.0029 
                 35436 
               
               
                 gamma-aminobutyrate 
                   
                   
                 
                   0.55 
                 
                 2.46E−05 
                 
                   0.68 
                 
                 0.0045 
                 1.1 
                 0.9728 
                 1416 
               
               
                 (GABA) 
               
               
                 N-(2-furoyl)glycine 
                 0.54 
                 &gt;0.1 
                 
                   0.53 
                 
                 3.23E−05 
                 
                   0.59 
                 
                 0.0001 
                 
                   2.71 
                 
                 0.0003 
                 31536 
               
               
                 glutathione, oxidized (GSSG) 
                   
                   
                 
                   2.25 
                 
                 3.43E−05 
                 
                   2.18 
                 
                 0.0003 
                 
                   2.11 
                 
                 0.0001 
                 38783 
               
               
                 itaconate 
                   
                   
                 
                   0.59 
                 
                 4.61E−05 
                 
                   0.73 
                 
                 0.0038 
                 0.8 
                 0.8293 
                 18373 
               
               
                 (methylenesuccinate) 
               
               
                 2,5-furandicarboxylic acid 
                   
                   
                 
                   0.57 
                 
                 6.18E−05 
                 
                   0.76 
                 
                 0.0002 
                 
                   1.98 
                 
                 0.0059 
                 40809 
               
               
                 2-methylhippurate 
                 0.7 
                 &gt;0.1 
                 
                   2.9 
                 
                 6.75E−05 
                 
                   2.24 
                 
                 0.0144 
                 1.85 
                 0.9824 
                 15670 
               
               
                 cystine 
                 1.44 
                 &gt;0.1 
                 
                   0.35 
                 
                 8.17E−05 
                 
                   0.46 
                 
                 0.0147 
                 0.62 
                 0.2776 
                 39512 
               
               
                 N-acetylphenylalanine 
                 0.73 
                 &gt;0.1 
                 
                   0.59 
                 
                 0.0001 
                 0.86 
                 0.2777 
                 
                   1.19 
                 
                 0.0145 
                 33950 
               
               
                 4-hydroxymandelate 
                   
                   
                 
                   0.72 
                 
                 0.0001 
                 
                   0.68 
                 
                 5.60E−05 
                 1.16 
                 0.8295 
                 1568 
               
               
                 pyridoxal 
                   
                   
                 
                   0.41 
                 
                 0.0001 
                 
                   0.48 
                 
                 0.0002 
                 1.02 
                 0.8261 
                 1651 
               
               
                 cortisone 
                   
                   
                 
                   1.34 
                 
                 0.0001 
                 
                   1.21 
                 
                 0.0254 
                 1.05 
                 0.9893 
                 1769 
               
               
                 riboflavin (Vitamin B2) 
                 0.36 
                 &gt;0.1 
                 
                   0.24 
                 
                 0.0002 
                 0.4 
                 0.1853 
                 0.96 
                 0.3165 
                 1827 
               
               
                 biliverdin 
                   
                   
                 
                   1.2 
                 
                 0.0002 
                 
                   1.18 
                 
                 0.0036 
                 
                   1.19 
                 
                 0.0004 
                 2137 
               
               
                 choline 
                   
                   
                 
                   1.4 
                 
                 0.0002 
                 1.18 
                 0.1933 
                 
                   1.57 
                 
                 4.94E−07 
                 15506 
               
               
                 2,4,6-trihydroxybenzoate 
                   
                   
                 
                   0.37 
                 
                 0.0002 
                 
                   0.6 
                 
                 0.0119 
                 1.74 
                 0.2432 
                 35892 
               
               
                 N-acetyltryptophan 
                 0.5 
                 &gt;0.1 
                 
                   0.48 
                 
                 0.0003 
                 0.82 
                 0.4342 
                 
                   1.43 
                 
                 0.0045 
                 33959 
               
               
                 galactinol 
                   
                   
                 
                   0.47 
                 
                 0.0003 
                 
                   0.67 
                 
                 0.0409 
                 1.13 
                 0.4772 
                 21034 
               
               
                 2-pyrrolidinone 
                   
                   
                 
                   0.57 
                 
                 0.0003 
                 
                   0.66 
                 
                 0.0066 
                 0.88 
                 0.4113 
                 31675 
               
               
                 phenylacetylglycine 
                   
                   
                 
                   0.58 
                 
                 0.0003 
                 
                   0.51 
                 
                 1.65E−06 
                 
                   1.99 
                 
                 3.43E−06 
                 33945 
               
               
                 4-hydroxy-2-oxoglutaric acid 
                   
                   
                 
                   2.68 
                 
                 0.0003 
                 
                   2.16 
                 
                 0.0198 
                 
                   0.57 
                 
                 0.001 
                 40062 
               
               
                 2-methylbutyrylglycine 
                 0.7 
                 &gt;0.1 
                 
                   0.68 
                 
                 0.0004 
                 
                   0.63 
                 
                 5.84E−06 
                 0.92 
                 0.3893 
                 31928 
               
               
                 1-methylhistidine 
                   
                   
                 
                   0.55 
                 
                 0.0004 
                 
                   0.61 
                 
                 0.0427 
                 0.94 
                 0.804 
                 30460 
               
               
                 3-methylcrotonylglycine 
                 0.62 
                 &gt;0.1 
                 
                   0.59 
                 
                 0.0005 
                 
                   0.58 
                 
                 1.72E−05 
                 
                   1.11 
                 
                 0.0712 
                 31940 
               
               
                 3-(3- 
                 0.64 
                 &gt;0.1 
                 
                   0.47 
                 
                 0.0005 
                 
                   0.57 
                 
                 0.001 
                 2.31 
                 0.1714 
                 35635 
               
               
                 hydroxyphenyl)propionate 
               
               
                 ribitol 
                   
                   
                 
                   0.7 
                 
                 0.0005 
                 
                   0.77 
                 
                 0.0008 
                 0.88 
                 0.1093 
                 15772 
               
               
                 guanidinoacetate 
                   
                   
                 
                   0.63 
                 
                 0.0006 
                 
                   0.5 
                 
                 0.0002 
                 1.06 
                 0.6981 
                 12359 
               
               
                 4-hydroxyhippurate 
                 0.89 
                 &gt;0.1 
                 
                   0.77 
                 
                 0.0007 
                 
                   0.6 
                 
                 8.54E−07 
                 0.88 
                 0.6039 
                 35527 
               
               
                 biotin 
                   
                   
                 
                   0.5 
                 
                 0.0008 
                 
                   0.74 
                 
                 0.0176 
                 1.05 
                 0.8124 
                 568 
               
               
                 adenosine 3′,5′-cyclic 
                   
                   
                 
                   0.79 
                 
                 0.0008 
                 
                   0.81 
                 
                 0.0011 
                 
                   0.78 
                 
                 0.0043 
                 2831 
               
               
                 monophosphate (cAMP) 
               
               
                 prostaglandin E2 
                   
                   
                 
                   1.37 
                 
                 0.0008 
                 
                   1.28 
                 
                 0.0199 
                 
                   1.28 
                 
                 0.0011 
                 7746 
               
               
                 sorbitol 
                 0.44 
                 &gt;0.1 
                 
                   0.22 
                 
                 0.001 
                 
                   0.77 
                 
                 0.0016 
                 0.48 
                 0.9192 
                 15053 
               
               
                 mesaconate (methylfumarate) 
                 0.78 
                 &gt;0.1 
                 
                   0.63 
                 
                 0.001 
                 
                   0.71 
                 
                 0.0838 
                 1.05 
                 0.4652 
                 18493 
               
               
                 N-acetyltyrosine 
                 0.55 
                 &gt;0.1 
                 
                   0.66 
                 
                 0.001 
                 0.97 
                 0.1054 
                 1.29 
                 0.2245 
                 32390 
               
               
                 lactose 
                   
                   
                 
                   0.52 
                 
                 0.0011 
                 
                   0.65 
                 
                 0.0065 
                 1 
                 0.695 
                 567 
               
               
                 1-(3-aminopropyl)-2- 
                   
                   
                 
                   1.6 
                 
                 0.0012 
                 
                   1.37 
                 
                 0.039 
                 
                   1.28 
                 
                 0.0897 
                 40506 
               
               
                 pyrrolidone 
               
               
                 glucosamine 
                 0.3 
                 &gt;0.1 
                 
                   0.46 
                 
                 0.0014 
                 
                   0.4 
                 
                 0.0045 
                 
                   1.16 
                 
                 0.0548 
                 18534 
               
               
                 3-hydroxysebacate 
                 2.61 
                 &gt;0.1 
                 
                   2.04 
                 
                 0.0014 
                 
                   2.06 
                 
                 0.0094 
                 1 
                 0.51 
                 31943 
               
               
                 7-methylguanine 
                   
                   
                 
                   1.22 
                 
                 0.0014 
                 1.1 
                 0.4843 
                 1.01 
                 0.7678 
                 35114 
               
               
                 5-aminovalerate 
                 2.17 
                 &gt;0.1 
                 
                   1.52 
                 
                 0.0015 
                 
                   1.41 
                 
                 0.001 
                 
                   3.2 
                 
                 0.0515 
                 18319 
               
               
                 mandelate 
                   
                   
                 
                   0.78 
                 
                 0.0016 
                 
                   0.79 
                 
                 0.0092 
                 1.02 
                 0.9228 
                 22160 
               
               
                 N-acetylserine 
                   
                   
                 
                   1.48 
                 
                 0.0016 
                 0.85 
                 0.6788 
                 1.17 
                 0.1978 
                 37076 
               
               
                 glutathione, reduced (GSH) 
                   
                   
                 
                   7.25 
                 
                 0.0018 
                 
                   6.62 
                 
                 0.0031 
                 
                   6.93 
                 
                 7.17E−05 
                 2127 
               
               
                 3-phosphoglycerate 
                   
                   
                 
                   1.05 
                 
                 0.002 
                 
                   1 
                 
                 0.0105 
                 1.75 
                 0.2037 
                 40264 
               
               
                 gulono-1,4-lactone 
                   
                   
                 
                   1.87 
                 
                 0.0021 
                 
                   1.85 
                 
                 0.0152 
                 
                   0.73 
                 
                 0.0002 
                 33454 
               
               
                 N-acetylproline 
                   
                   
                 
                   0.71 
                 
                 0.0021 
                 
                   0.69 
                 
                 0.0005 
                 1.07 
                 0.9292 
                 34387 
               
               
                 N-carbamoylaspartate 
                   
                   
                 
                   0.43 
                 
                 0.0022 
                 
                   0.68 
                 
                 0.0093 
                 1.16 
                 0.5083 
                 1594 
               
               
                 2-hydroxyadipate 
                   
                   
                 
                   0.77 
                 
                 0.0022 
                 
                   0.78 
                 
                 0.0052 
                 
                   0.83 
                 
                 0.0891 
                 31934 
               
               
                 N-methylglutamate 
                   
                   
                 
                   0.97 
                 
                 0.0024 
                 
                   0.73 
                 
                 0.0001 
                 1.59 
                 0.3923 
                 31532 
               
               
                 galactitol (dulcitol) 
                 0.78 
                 &gt;0.1 
                 
                   0.76 
                 
                 0.0025 
                 
                   0.74 
                 
                 0.0002 
                 1.05 
                 0.672 
                 1117 
               
               
                 3-methylxanthine 
                 1.26 
                 &gt;0.1 
                 
                   0.62 
                 
                 0.0028 
                 0.87 
                 0.5921 
                 1.22 
                 0.4832 
                 32445 
               
               
                 5-methyltetrahydrofolate 
                   
                   
                 
                   0.45 
                 
                 0.0028 
                 0.5 
                 0.1388 
                 0.98 
                 0.7745 
                 18330 
               
               
                 (5MeTHF) 
               
               
                 urate 
                   
                   
                 
                   1.18 
                 
                 0.0032 
                 1.02 
                 0.7136 
                 
                   1.15 
                 
                 0.0358 
                 1604 
               
               
                 5-acetylamino-6-amino-3- 
                   
                   
                 
                   0.49 
                 
                 0.0035 
                 1.01 
                 0.4408 
                 1.14 
                 0.5455 
                 34424 
               
               
                 methyluracil 
               
               
                 4-vinylphenol sulfate 
                   
                   
                 
                   0.76 
                 
                 0.0035 
                 
                   0.69 
                 
                 0.0113 
                 1.05 
                 0.9684 
                 36098 
               
               
                 gamma-glutamylvaline 
                   
                   
                 
                   0.76 
                 
                 0.0037 
                 
                   0.73 
                 
                 0.0006 
                 0.85 
                 0.1465 
                 32393 
               
               
                 allo-threonine 
                 0.79 
                 &gt;0.1 
                 
                   0.68 
                 
                 0.0038 
                 
                   0.71 
                 
                 0.0251 
                 
                   0.99 
                 
                 0.0301 
                 15142 
               
               
                 pyroglutamylglutamine 
                 0.71 
                 &gt;0.1 
                 
                   0.77 
                 
                 0.004 
                 0.86 
                 0.1656 
                 0.95 
                 0.1634 
                 22194 
               
               
                 sucrose 
                 0.69 
                 &gt;0.1 
                 
                   0.46 
                 
                 0.0041 
                 
                   0.48 
                 
                 0.0073 
                 
                   1.41 
                 
                 8.36E−06 
                 1519 
               
               
                 glycolithocholate sulfate 
                 1.24 
                 &gt;0.1 
                 
                   0.73 
                 
                 0.0041 
                 
                   0.65 
                 
                 0.0012 
                 
                   0.57 
                 
                 0.0007 
                 32620 
               
               
                 beta-hydroxypyruvate 
                   
                   
                 
                   1.79 
                 
                 0.0041 
                 
                   2.61 
                 
                 0.0013 
                 
                   0.88 
                 
                 0.0353 
                 15686 
               
               
                 1,6-anhydroglucose 
                 0.78 
                 &gt;0.1 
                 
                   0.68 
                 
                 0.0042 
                 
                   0.74 
                 
                 0.025 
                 
                   1.41 
                 
                 0.0148 
                 21049 
               
               
                 5-acetylamino-6-formylamino- 
                   
                   
                 
                   0.72 
                 
                 0.0042 
                 1.21 
                 0.9968 
                 1.32 
                 0.5771 
                 34401 
               
               
                 3-methyluracil 
               
               
                 3-hydroxyglutarate 
                   
                   
                 
                   0.7 
                 
                 0.0045 
                 
                   0.78 
                 
                 0.0209 
                 0.89 
                 0.2755 
                 36863 
               
               
                 ciliatine (2- 
                 1.72 
                 &gt;0.1 
                 
                   1.93 
                 
                 0.0046 
                 
                   0.22 
                 
                 0.004 
                 3.7 
                 0.1618 
                 15125 
               
               
                 aminoethylphosphonate) 
               
               
                 3-methyl-2-oxovalerate 
                   
                   
                 
                   1.65 
                 
                 0.0046 
                 1.12 
                 0.6122 
                 
                   0.77 
                 
                 0.0632 
                 15676 
               
               
                 aspartylaspartate 
                   
                   
                 
                   0.58 
                 
                 0.0048 
                 0.72 
                 0.1509 
                 0.76 
                 0.7205 
                 40671 
               
               
                 N-methyl proline 
                 1.77 
                 &gt;0.1 
                 
                   1.6 
                 
                 0.0049 
                 1.16 
                 0.3297 
                 
                   1.91 
                 
                 0.0015 
                 37431 
               
               
                 theobromine 
                 0.58 
                 &gt;0.1 
                 
                   0.64 
                 
                 0.0051 
                 0.87 
                 0.9734 
                 1.38 
                 0.1514 
                 18392 
               
               
                 N-acetylcysteine 
                   
                   
                 
                   0.66 
                 
                 0.0052 
                 
                   0.59 
                 
                 0.0063 
                 1.32 
                 0.2267 
                 1586 
               
               
                 5-hydroxyhexanoate 
                   
                   
                 
                   0.65 
                 
                 0.0056 
                 
                   0.7 
                 
                 0.0076 
                 1.01 
                 0.452 
                 31938 
               
               
                 dopamine 
                 0.37 
                 &gt;0.1 
                 
                   0.58 
                 
                 0.0063 
                 
                   0.82 
                 
                 0.0378 
                 1.17 
                 0.2153 
                 12130 
               
               
                 3-methylglutaconate 
                   
                   
                 
                   0.79 
                 
                 0.0064 
                 0.96 
                 0.1681 
                 1.09 
                 0.6266 
                 38667 
               
               
                 alanylalanine 
                   
                   
                 
                   0.74 
                 
                 0.0068 
                 0.94 
                 0.7303 
                 1.11 
                 0.4675 
                 15129 
               
               
                 taurolithocholate 3-sulfate 
                   
                   
                 
                   0.66 
                 
                 0.007 
                 
                   0.74 
                 
                 0.013 
                 
                   0.58 
                 
                 0.0083 
                 36850 
               
               
                 trans-aconitate 
                   
                   
                 
                   0.76 
                 
                 0.0071 
                 
                   0.83 
                 
                 0.0399 
                 1.02 
                 0.7384 
                 27741 
               
               
                 glycerol 
                   
                   
                 
                   3.83 
                 
                 0.0075 
                 
                   3.94 
                 
                 0.0041 
                 
                   0.26 
                 
                 4.44E−07 
                 15122 
               
               
                 sebacate (decanedioate) 
                 1.36 
                 &gt;0.1 
                 
                   4.08 
                 
                 0.008 
                 3.65 
                 0.1668 
                 
                   1.08 
                 
                 0.0422 
                 32398 
               
               
                 N-carbamoylsarcosine 
                   
                   
                 
                   0.86 
                 
                 0.008 
                 
                   0.85 
                 
                 0.0038 
                 
                   1.35 
                 
                 0.0149 
                 38696 
               
               
                 vanillate 
                   
                   
                 
                   0.96 
                 
                 0.0081 
                 
                   1.08 
                 
                 0.0093 
                 
                   3.35 
                 
                 0.0024 
                 35639 
               
               
                 ethanolamine 
                 0.74 
                 &gt;0.1 
                 
                   0.65 
                 
                 0.0088 
                 
                   0.65 
                 
                 0.0052 
                 
                   1.17 
                 
                 0.0002 
                 1497 
               
               
                 galactose 
                   
                   
                 
                   0.67 
                 
                 0.009 
                 0.82 
                 0.2799 
                 
                   1.4 
                 
                 0.0834 
                 12055 
               
               
                 5-hydroxyindoleacetate 
                   
                   
                 
                   0.66 
                 
                 0.0092 
                 
                   0.84 
                 
                 0.0845 
                 1 
                 0.6618 
                 437 
               
               
                 pyridoxine (Vitamin B6) 
                   
                   
                 
                   0.43 
                 
                 0.0098 
                 0.85 
                 0.3614 
                 1 
                 1 
                 608 
               
               
                 threitol 
                 1.45 
                 &gt;0.1 
                 
                   0.96 
                 
                 0.0115 
                 
                   0.84 
                 
                 0.001 
                 1.01 
                 0.4182 
                 35854 
               
               
                 Ac-Ser-Asp-Lys-Pro-OH 
                   
                   
                 
                   0.61 
                 
                 0.0121 
                 1.52 
                 0.2446 
                 1.07 
                 0.8753 
                 40707 
               
               
                 (SEQ ID N0: 1) 
               
               
                 scyllo-inositol 
                   
                   
                 
                   0.79 
                 
                 0.0131 
                 
                   0.92 
                 
                 0.0984 
                 
                   1.57 
                 
                 0.0261 
                 32379 
               
               
                 pyruvate 
                   
                   
                 
                   0.78 
                 
                 0.0136 
                 
                   0.85 
                 
                 0.0864 
                 1.03 
                 0.7803 
                 599 
               
               
                 4-methyl-2-oxopentanoate 
                   
                   
                 
                   1.67 
                 
                 0.0145 
                 1.32 
                 0.12 
                 0.91 
                 0.3146 
                 22116 
               
               
                 N2-acetyllysine 
                   
                   
                 
                   0.77 
                 
                 0.0149 
                 
                   0.8 
                 
                 0.0484 
                 
                   0.78 
                 
                 0.082 
                 36751 
               
               
                 3-hydroxypyridine 
                   
                   
                 
                   0.72 
                 
                 0.0163 
                 0.79 
                 0.1332 
                 
                   2 
                 
                 0.0002 
                 21169 
               
               
                 putrescine 
                   
                   
                 
                   1.22 
                 
                 0.0167 
                 
                   0.61 
                 
                 0.0104 
                 
                   3.63 
                 
                 0.0042 
                 1408 
               
               
                 1,7-dimethylurate 
                 1.55 
                 &gt;0.1 
                 
                   0.83 
                 
                 0.0175 
                 1.06 
                 0.8313 
                 1.23 
                 0.719 
                 34400 
               
               
                 1,3,7-trimethylurate 
                   
                   
                 
                   0.67 
                 
                 0.0177 
                 0.8 
                 0.285 
                 
                   1.98 
                 
                 0.016 
                 34404 
               
               
                 3-methylhistidine 
                 0.75 
                 &gt;0.1 
                 
                   0.67 
                 
                 0.0189 
                 
                   0.67 
                 
                 0.0779 
                 0.89 
                 0.107 
                 15677 
               
               
                 nicotinurate 
                   
                   
                 
                   9.19 
                 
                 0.0204 
                 
                   8.98 
                 
                 0.0846 
                 
                   9.27 
                 
                 0.0217 
                 35121 
               
               
                 1,5-anhydroglucitol (1,5-AG) 
                   
                   
                 
                   1.28 
                 
                 0.0207 
                 0.82 
                 0.4261 
                 
                   1.37 
                 
                 0.0912 
                 20675 
               
               
                 imidazole propionate 
                   
                   
                 
                   1.4 
                 
                 0.0207 
                 1.16 
                 0.3092 
                 
                   1.57 
                 
                 1.87E−05 
                 40730 
               
               
                 N6-acetyllysine 
                   
                   
                 
                   0.82 
                 
                 0.0208 
                 
                   0.81 
                 
                 0.0079 
                 0.92 
                 0.2837 
                 36752 
               
               
                 N-acetylhistidine 
                 0.79 
                 &gt;0.1 
                 
                   0.92 
                 
                 0.0213 
                 
                   0.78 
                 
                 7.36E−05 
                 
                   0.84 
                 
                 0.0122 
                 33946 
               
               
                 gamma-glutamyltyrosine 
                 1.62 
                 &gt;0.1 
                 
                   0.74 
                 
                 0.0219 
                 
                   0.73 
                 
                 0.0426 
                 1.06 
                 0.5743 
                 2734 
               
               
                 picolinate 
                 0.24 
                 &gt;0.1 
                 
                   0.81 
                 
                 0.022 
                 0.97 
                 0.7127 
                 
                   0.83 
                 
                 0.0052 
                 1512 
               
               
                 7-methylxanthine 
                 1.36 
                 &gt;0.1 
                 
                   0.68 
                 
                 0.023 
                 0.93 
                 0.8384 
                 1.21 
                 0.4077 
                 34390 
               
               
                 dihydroferulic acid 
                 0.67 
                 &gt;0.1 
                 
                   0.74 
                 
                 0.0243 
                 
                   0.43 
                 
                 0.0011 
                 2.23 
                 0.1095 
                 40481 
               
               
                 erythronate 
                   
                   
                 
                   0.84 
                 
                 0.0252 
                 
                   0.92 
                 
                 0.0896 
                 0.91 
                 0.3029 
                 33477 
               
               
                 glucose-6-phosphate (G6P) 
                   
                   
                 
                   1.69 
                 
                 0.0256 
                 1.48 
                 0.1935 
                 
                   1.99 
                 
                 0.0002 
                 31260 
               
               
                 glutarate (pentanedioate) 
                   
                   
                 
                   0.72 
                 
                 0.0267 
                 
                   0.81 
                 
                 0.053 
                 0.53 
                 0.1088 
                 396 
               
               
                 phosphoethanolamine 
                   
                   
                 
                   0.84 
                 
                 0.0298 
                 0.92 
                 0.1519 
                 1.15 
                 0.1527 
                 12102 
               
               
                 3-hydroxycinnamate (m- 
                   
                   
                 
                   0.66 
                 
                 0.0311 
                 0.72 
                 0.1246 
                 1.22 
                 0.9227 
                 20698 
               
               
                 coumarate) 
               
               
                 2,4-dioxo-1H-pyrimidine-5- 
                   
                   
                 
                   0.75 
                 
                 0.0311 
                 0.86 
                 0.2357 
                 1.02 
                 0.7427 
                 37444 
               
               
                 carboxylic acid 
               
               
                 carnosine 
                   
                   
                 
                   0.52 
                 
                 0.0321 
                 
                   0.33 
                 
                 9.79E−06 
                 1.23 
                 0.5621 
                 1768 
               
               
                 2-octenedioate 
                   
                   
                 
                   0.76 
                 
                 0.0322 
                 0.93 
                 0.4621 
                 0.78 
                 0.9907 
                 35120 
               
               
                 arabonate 
                   
                   
                 
                   0.84 
                 
                 0.0327 
                 
                   0.87 
                 
                 0.04 
                 1.11 
                 0.3652 
                 37516 
               
               
                 ascorbate (Vitamin C) 
                   
                   
                 
                   0.24 
                 
                 0.033 
                 0.78 
                 0.4416 
                 1.71 
                 0.7973 
                 1640 
               
               
                 abscisate 
                 0.78 
                 &gt;0.1 
                 
                   0.59 
                 
                 0.0331 
                 
                   0.57 
                 
                 0.0059 
                 1.6 
                 0.275 
                 21156 
               
               
                 4-hydroxybenzoate 
                   
                   
                 
                   0.77 
                 
                 0.034 
                 
                   0.74 
                 
                 0.0306 
                 0.83 
                 0.2701 
                 21133 
               
               
                 gamma-glutamylleucine 
                 1.59 
                 &gt;0.1 
                 
                   0.73 
                 
                 0.0364 
                 
                   0.7 
                 
                 0.0062 
                 0.92 
                 0.6214 
                 18369 
               
               
                 malate 
                 2.04 
                 &gt;0.1 
                 
                   1.15 
                 
                 0.0365 
                 0.91 
                 0.7515 
                 0.59 
                 0.5138 
                 1303 
               
               
                 3-methylglutarate 
                   
                   
                 
                   0.88 
                 
                 0.0368 
                 1.11 
                 0.559 
                 0.98 
                 0.1892 
                 1557 
               
               
                 2,3-butanediol 
                   
                   
                 
                   0.44 
                 
                 0.0373 
                 
                   0.58 
                 
                 0.0477 
                 
                   1.29 
                 
                 0.0935 
                 35691 
               
               
                 mannose 
                   
                   
                 
                   0.67 
                 
                 0.0385 
                 0.87 
                 0.1506 
                 1.29 
                 0.2013 
                 584 
               
               
                 threonate 
                 1.27 
                 &gt;0.1 
                 
                   0.69 
                 
                 0.0389 
                 0.94 
                 0.1532 
                 
                   0.8 
                 
                 0.0852 
                 27738 
               
               
                 3-hydroxymandelate 
                   
                   
                 
                   0.22 
                 
                 0.0389 
                 0.28 
                 0.5415 
                 0.99 
                 0.2189 
                 22112 
               
               
                 cystathionine 
                   
                   
                 
                   0.68 
                 
                 0.0404 
                 
                   0.61 
                 
                 0.0233 
                 1.17 
                 0.7165 
                 15705 
               
               
                 phenol sulfate 
                 0.61 
                 &gt;0.1 
                 
                   0.94 
                 
                 0.0436 
                 
                   0.8 
                 
                 0.0073 
                 
                   0.77 
                 
                 0.0043 
                 32553 
               
               
                 5-oxoproline 
                 1.2 
                 &gt;0.1 
                 
                   0.85 
                 
                 0.0439 
                 
                   0.85 
                 
                 0.02 
                 0.93 
                 0.7294 
                 1494 
               
               
                 deoxycholate 
                   
                   
                 
                   0.75 
                 
                 0.0467 
                 0.98 
                 0.3143 
                 1.18 
                 0.5303 
                 1114 
               
               
                 3-hydroxybenzoate 
                 0.6 
                 &gt;0.1 
                 
                   0.79 
                 
                 0.0472 
                 0.84 
                 0.4362 
                 
                   1.35 
                 
                 0.0099 
                 15673 
               
               
                 cis-aconitate 
                   
                   
                 
                   0.89 
                 
                 0.0479 
                 
                   0.85 
                 
                 0.0049 
                 0.93 
                 0.1774 
                 12025 
               
               
                 3-hydroxyproline 
                 0.66 
                 &gt;0.1 
                 
                   0.8 
                 
                 0.0482 
                 
                   0.83 
                 
                 0.0806 
                 
                   1.11 
                 
                 0.045 
                 38635 
               
               
                 ethyl glucuronide 
                 0.58 
                 &gt;0.1 
                 
                   0.24 
                 
                 0.049 
                 0.57 
                 0.8533 
                 
                   0.88 
                 
                 0.0556 
                 39603 
               
               
                 1-methylxanthine 
                 1.33 
                 &gt;0.1 
                 
                   1.11 
                 
                 0.0509 
                 1.22 
                 0.966 
                 1.86 
                 0.2526 
                 34389 
               
               
                 UDP-glucuronate 
                   
                   
                 
                   0.86 
                 
                 0.0526 
                 1.05 
                 0.5627 
                 1.19 
                 0.2159 
                 34377 
               
               
                 2-(4- 
                   
                   
                 
                   0.4 
                 
                 0.0536 
                 
                   0.3 
                 
                 0.0847 
                 1.59 
                 0.3248 
                 35632 
               
               
                 hydroxyphenyl)propionate 
               
               
                 hexanoylcarnitine 
                 1.21 
                 &gt;0.1 
                 
                   1.21 
                 
                 0.0543 
                 
                   1.33 
                 
                 0.0421 
                 
                   0.85 
                 
                 0.054 
                 32328 
               
               
                 gamma-CEHC 
                   
                   
                 
                   0.62 
                 
                 0.0559 
                 
                   0.56 
                 
                 0.0311 
                 
                   0.46 
                 
                 5.65E−05 
                 37462 
               
               
                 arabitol 
                   
                   
                 
                   0.84 
                 
                 0.0561 
                 
                   0.85 
                 
                 0.0354 
                 1.01 
                 0.9139 
                 38075 
               
               
                 phosphoenolpyruvate (PEP) 
                   
                   
                 
                   2.4 
                 
                 0.0574 
                 
                   2.58 
                 
                 0.0649 
                 
                   2.21 
                 
                 0.0166 
                 597 
               
               
                 oxalate (ethanedioate) 
                   
                   
                 
                   2.11 
                 
                 0.0601 
                 2 
                 0.1947 
                 1.34 
                 0.498 
                 20694 
               
               
                 4-ureidobutyrate 
                   
                   
                 
                   0.88 
                 
                 0.0627 
                 
                   0.85 
                 
                 0.0073 
                 1.08 
                 0.1402 
                 22118 
               
               
                 tiglyl carnitine 
                 0.79 
                 &gt;0.1 
                 
                   0.87 
                 
                 0.0637 
                 0.93 
                 0.1619 
                 0.91 
                 0.3428 
                 35428 
               
               
                 tigloylglycine 
                   
                   
                 
                   0.79 
                 
                 0.0655 
                 
                   0.77 
                 
                 0.0065 
                 0.87 
                 0.3945 
                 1598 
               
               
                 homocitrate 
                   
                   
                 
                   0.92 
                 
                 0.0664 
                 
                   0.94 
                 
                 0.0404 
                 0.92 
                 0.1273 
                 39601 
               
               
                 pinitol 
                   
                   
                 
                   0.82 
                 
                 0.0756 
                 
                   0.43 
                 
                 0.0342 
                 
                   3.85 
                 
                 0.0098 
                 37086 
               
               
                 pregnen-diol disulfate 
                   
                   
                 
                   1.03 
                 
                 0.0763 
                 1.03 
                 0.9366 
                 
                   0.69 
                 
                 0.0071 
                 32562 
               
               
                 3-hydroxyisobutyrate 
                 1.68 
                 &gt;0.1 
                 
                   0.91 
                 
                 0.0773 
                 
                   0.92 
                 
                 0.0787 
                 0.95 
                 0.8405 
                 1549 
               
               
                 gamma-glutamylisoleucine 
                   
                   
                 
                   0.89 
                 
                 0.078 
                 
                   0.83 
                 
                 0.0074 
                 0.98 
                 0.6295 
                 34456 
               
               
                 ectoine 
                   
                   
                 
                   0.73 
                 
                 0.081 
                 0.67 
                 0.1321 
                 1.01 
                 0.4766 
                 35651 
               
               
                 N6-methyladenosine 
                   
                   
                 
                   1.68 
                 
                 0.0812 
                 0.96 
                 0.8786 
                 
                   0.73 
                 
                 0.0023 
                 37114 
               
               
                 2-phenylglycine 
                   
                   
                 
                   1.62 
                 
                 0.0871 
                 
                   1.64 
                 
                 0.0636 
                 0.91 
                 0.1756 
                 37441 
               
               
                 xylonate 
                   
                   
                 
                   0.9 
                 
                 0.0888 
                 
                   0.89 
                 
                 0.0521 
                 1.02 
                 0.7659 
                 35638 
               
               
                 neopterin 
                   
                   
                 
                   1.17 
                 
                 0.0895 
                 1.14 
                 0.1775 
                 0.96 
                 0.8238 
                 35131 
               
               
                 2-ethylphenylsulfate 
                   
                   
                 
                   1.96 
                 
                 0.0921 
                 1.03 
                 0.9339 
                 1.59 
                 0.1895 
                 36847 
               
               
                 sulforaphane-N-acetyl- 
                   
                   
                 
                   0.79 
                 
                 0.0923 
                 0.82 
                 0.2954 
                 1.02 
                 0.9047 
                 40468 
               
               
                 cysteine 
               
               
                 uridine 
                   
                   
                 
                   1.37 
                 
                 0.0944 
                 1.08 
                 0.9525 
                 1.11 
                 0.7757 
                 606 
               
               
                 fucose 
                   
                   
                 
                   0.88 
                 
                 0.0955 
                 0.97 
                 0.3105 
                 0.85 
                 0.1996 
                 15821 
               
               
                 N-acetylalanine 
                   
                   
                 
                   0.82 
                 
                 0.0987 
                 0.87 
                 0.4399 
                 1.01 
                 0.9492 
                 1585 
               
               
                 N-acetylarginine 
                   
                   
                 
                   0.9 
                 
                 0.0999 
                 
                   0.85 
                 
                 0.0889 
                 
                   0.76 
                 
                 0.014 
                 33953 
               
               
                 anthranilate 
                   
                   
                 0.81 
                 0.1291 
                 0.9 
                 0.1911 
                 
                   0.6 
                 
                 0.0041 
                 4970 
               
               
                 nicotinate 
                 0.79 
                 &gt;0.1 
                 0.84 
                 0.1463 
                 1.1 
                 0.8109 
                 
                   1.87 
                 
                 0.0011 
                 1504 
               
               
                 cyclo(leu-pro) 
                   
                   
                 0.97 
                 0.1786 
                 0.92 
                 0.2661 
                 
                   1.84 
                 
                 0.0012 
                 37104 
               
               
                 azelate (nonanedioate) 
                 0.37 
                 &gt;0.1 
                 0.8 
                 0.1948 
                 
                   0.61 
                 
                 0.0011 
                 
                   1.49 
                 
                 0.0021 
                 18362 
               
               
                 cyclo(gly-pro) 
                   
                   
                 1.18 
                 0.2919 
                 1.09 
                 0.2333 
                 
                   1.01 
                 
                 0.0219 
                 37077 
               
               
                 decanoylcarnitine 
                   
                   
                 1.05 
                 0.313 
                 
                   1.25 
                 
                 0.043 
                 
                   0.6 
                 
                 0.0008 
                 33941 
               
               
                 5alpha-androstan- 
                   
                   
                 0.88 
                 0.3762 
                 0.83 
                 0.1841 
                 
                   0.58 
                 
                 0.0006 
                 37190 
               
               
                 3beta,17beta-diol disulfate 
               
               
                 dimethylarginine (SDMA + 
                   
                   
                 0.95 
                 0.4243 
                 
                   0.9 
                 
                 0.0826 
                 
                   0.81 
                 
                 0.0047 
                 36808 
               
               
                 ADMA) 
               
               
                 21-hydroxypregnenolone 
                   
                   
                 0.79 
                 0.4434 
                 
                   0.76 
                 
                 0.0265 
                 
                   0.66 
                 
                 0.0007 
                 37173 
               
               
                 disulfate 
               
               
                 2-hydroxyglutarate 
                 1.94 
                 &gt;0.1 
                 0.96 
                 0.4442 
                 0.87 
                 0.1767 
                 
                   0.66 
                 
                 0.0043 
                 37253 
               
               
                 methyl indole-3-acetate 
                   
                   
                 1.36 
                 0.4537 
                 1.28 
                 0.9621 
                 
                   0.3 
                 
                 9.80E−11 
                 1584 
               
               
                 trigonelline (N′- 
                 0.7 
                 &gt;0.1 
                 1 
                 0.4604 
                 1.16 
                 0.8334 
                 
                   1.66 
                 
                 0.0007 
                 32401 
               
               
                 methylnicotinate) 
               
               
                 caffeate 
                   
                   
                 0.82 
                 0.4951 
                 0.96 
                 0.3892 
                 
                   2.11 
                 
                 0.0014 
                 21177 
               
               
                 5-methylthioadenosine (MTA) 
                   
                   
                 1.07 
                 0.5048 
                 0.98 
                 0.9248 
                 
                   0.56 
                 
                 0.0002 
                 1419 
               
               
                 4-androsten-3beta,17beta- 
                 0.72 
                 &gt;0.1 
                 0.81 
                 0.6211 
                 0.75 
                 0.1013 
                 
                   0.58 
                 
                 9.36E−05 
                 37203 
               
               
                 diol disulfate 2 
               
               
                 2-hydroxyisobutyrate 
                   
                   
                 1.08 
                 0.6896 
                 1.15 
                 0.4164 
                 
                   0.69 
                 
                 3.47E−06 
                 22030 
               
               
                 Isobar: glucuronate, 
                   
                   
                 0.89 
                 0.7531 
                 0.98 
                 0.4203 
                 
                   1.24 
                 
                 0.0045 
                 33001 
               
               
                 galacturonate, 5-keto- 
               
               
                 gluconate 
               
               
                 androsterone sulfate 
                 0.62 
                 &gt;0.1 
                 0.83 
                 0.8126 
                 
                   0.67 
                 
                 0.0171 
                 
                   0.65 
                 
                 0.0035 
                 31591 
               
               
                 glycine 
                 4.87 
                 &gt;0.1 
                 1.13 
                 0.8498 
                 
                   0.75 
                 
                 0.0679 
                 
                   1.28 
                 
                 0.0005 
                 11777 
               
               
                 beta-alanine 
                   
                   
                 0.64 
                 0.8514 
                 0.7 
                 0.5112 
                 
                   2.39 
                 
                 4.66E−06 
                 55 
               
               
                 4-androsten-3beta,17beta- 
                 0.33 
                 &gt;0.1 
                 0.96 
                 0.9628 
                 0.83 
                 0.2519 
                 
                   0.62 
                 
                 0.0005 
                 37202 
               
               
                 diol disulfate 1 
               
               
                 pregnanediol-3-glucuronide 
                   
                   
                 0.9 
                 0.9963 
                 0.62 
                 0.1759 
                 
                   0.59 
                 
                 0.0115 
                 40708 
               
               
                 4-acetamidophenol 
                 0.24 
                 0.0092 
               
               
                 N-acetylglutamate 
                 2.54 
                 0.0161 
               
               
                 dehydroisoandrosterone 
                 0.5 
                 0.0166 
               
               
                 sulfate (DHEA-S) 
               
               
                 isocitrate 
                 2.05 
                 0.0214 
               
               
                 tetrahydrocortisone 
                 0.54 
                 0.0219 
               
               
                 4-acetaminophen sulfate 
                 0.34 
                 0.032 
               
               
                 glycerol 2-phosphate 
                 2.29 
                 0.0369 
               
               
                 3-sialyllactose 
                 1.49 
                 0.0375 
               
               
                 pyroglutamine 
                 0.54 
                 0.038 
               
               
                 2-methoxyacetaminophen 
                 0.34 
                 0.0471 
               
               
                 glucuronide 
               
               
                 glycoursodeoxycholate 
                 
                   0.56 
                 
                 0.0503 
               
               
                 thymol sulfate 
                 
                   0.51 
                 
                 0.0515 
               
               
                 dihydrobiopterin 
                 
                   0.54 
                 
                 0.062 
               
               
                 trimethylamine N-oxide 
                 
                   0.7 
                 
                 0.0681 
               
               
                 homovanillate (HVA) 
                 
                   0.16 
                 
                 0.0742 
               
               
                 isoleucine 
                 1.35 
                 &gt;0.1 
                 
                   1.41 
                 
                 0.0015 
                 1.23 
                 0.2564 
                 1.18 
                 0.1725 
                 1125 
               
               
                 cortisol 
                 0.78 
                 &gt;0.1 
                 
                   2.6 
                 
                 4.30E−08 
                 
                   1.7 
                 
                 0.0064 
                 1.11 
                 0.7214 
                 1712 
               
               
                 2-hydroxybutyrate (AHB) 
                   
                   
                 
                   2.96 
                 
                 6.72E−06 
                 
                   2.04 
                 
                 0.0004 
                 0.69 
                 0.4915 
                 21044 
               
               
                 succinate 
                   
                   
                 
                   0.65 
                 
                 5.09E−05 
                 
                   0.6 
                 
                 0.0002 
                 
                   0.62 
                 
                 0.0002 
                 1437 
               
               
                 glutamine 
                   
                   
                 
                   1.65 
                 
                 6.99E−05 
                 0.96 
                 0.5801 
                 1.3 
                 0.1086 
                 53 
               
               
                 adenosine 
                   
                   
                 
                   0.73 
                 
                 9.13E−05 
                 
                   0.7 
                 
                 5.99E−05 
                 
                   0.73 
                 
                 3.46E−05 
                 555 
               
               
                 kynurenine 
                 1.53 
                 &gt;0.1 
                 
                   2.17 
                 
                 0.0002 
                 
                   1.93 
                 
                 0.0717 
                 1.51 
                 0.2261 
                 15140 
               
               
                 carnitine 
                 0.69 
                 &gt;0.1 
                 
                   1.77 
                 
                 0.0003 
                 
                   1.13 
                 
                 0.0141 
                 1.17 
                 0.146 
                 15500 
               
               
                 creatine 
                   
                   
                 
                   0.31 
                 
                 0.001 
                 
                   0.35 
                 
                 0.0004 
                 1.06 
                 0.9435 
                 27718 
               
               
                 pantothenate 
                 0.78 
                 &gt;0.1 
                 
                   0.57 
                 
                 0.0016 
                 
                   0.71 
                 
                 0.0906 
                 0.89 
                 0.1792 
                 1508 
               
               
                 arginine 
                   
                   
                 
                   0.39 
                 
                 0.0016 
                 
                   0.61 
                 
                 0.0019 
                 
                   1.8 
                 
                 0.0062 
                 1638 
               
               
                 leucine 
                   
                   
                 
                   1.34 
                 
                 0.002 
                 1.19 
                 0.236 
                 1.06 
                 0.7535 
                 60 
               
               
                 valine 
                 0.78 
                 &gt;0.1 
                 
                   1.34 
                 
                 0.0031 
                 1.18 
                 0.2408 
                 1.11 
                 0.4582 
                 1649 
               
               
                 histidine 
                 0.76 
                 &gt;0.1 
                 
                   1.33 
                 
                 0.0032 
                 0.94 
                 0.4906 
                 1.06 
                 0.3121 
                 59 
               
               
                 tryptophan 
                 0.68 
                 &gt;0.1 
                 
                   1.32 
                 
                 0.0034 
                 1.04 
                 0.6898 
                 0.9 
                 0.6005 
                 54 
               
               
                 homoserine 
                   
                   
                 
                   0.92 
                 
                 0.0079 
                 
                   1.01 
                 
                 0.0164 
                 
                   1.84 
                 
                 0.0325 
                 23642 
               
               
                 uracil 
                 0.66 
                 &gt;0.1 
                 
                   0.78 
                 
                 0.023 
                 
                   0.69 
                 
                 0.0071 
                 
                   0.66 
                 
                 0.0002 
                 605 
               
               
                 indolelactate 
                 0.79 
                 &gt;0.1 
                 
                   0.78 
                 
                 0.0275 
                 
                   1 
                 
                 0.5425 
                 
                   1.33 
                 
                 0.0288 
                 18349 
               
               
                 sarcosine (N-Methylglycine) 
                 1.46 
                 &gt;0.1 
                 
                   0.79 
                 
                 0.0401 
                 
                   0.75 
                 
                 0.0205 
                 
                   1.19 
                 
                 0.0077 
                 1516 
               
               
                 lysine 
                 1.63 
                 &gt;0.1 
                 
                   0.65 
                 
                 0.0448 
                 
                   0.54 
                 
                 0.0523 
                 
                   1.06 
                 
                 0.0314 
                 1301 
               
               
                 asparagine 
                   
                   
                 
                   0.83 
                 
                 0.0448 
                 
                   0.73 
                 
                 0.0007 
                 
                   1.26 
                 
                 0.0361 
                 11398 
               
               
                 3-(4-hydroxyphenyl)lactate 
                   
                   
                 
                   0.74 
                 
                 0.0499 
                 1.3 
                 0.7506 
                 1.15 
                 0.2448 
                 32197 
               
               
                 taurine 
                 0.62 
                 &gt;0.1 
                 
                   1.7 
                 
                 0.0637 
                 1.35 
                 0.8004 
                 
                   1.5 
                 
                 0.0014 
                 2125 
               
               
                 citramalate 
                 1.43 
                 &gt;0.1 
                 
                   0.87 
                 
                 0.0766 
                 
                   0.89 
                 
                 0.0574 
                 0.96 
                 0.6852 
                 22158 
               
               
                 glycerophosphorylcholine 
                 1.99 
                 0.0129 
               
               
                 (GPC) 
               
               
                 trans-urocanate 
                 
                   0.71 
                 
                 0.0609 
               
               
                 caffeine 
                 1.63 
                 &gt;0.1 
                 
                   0.68 
                 
                 0.0967 
                 0.63 
                 0.1153 
                 
                   2.47 
                 
                 0.0053 
                 569 
               
               
                 glutamate 
                 2.26 
                 &gt;0.1 
                 1.6 
                 0.1089 
                 1.15 
                 0.7539 
                 
                   1.63 
                 
                 0.0001 
                 57 
               
               
                 alanine 
                 0.8 
                 &gt;0.1 
                 0.92 
                 0.1924 
                 
                   0.69 
                 
                 0.0003 
                 
                   1.46 
                 
                 8.99E−06 
                 1126 
               
               
                 aspartate 
                   
                   
                 1.26 
                 0.4825 
                 1.19 
                 0.6645 
                 
                   1.77 
                 
                 5.78E−05 
                 15996 
               
               
                 threonine 
                 0.79 
                 &gt;0.1 
                 1 
                 0.899 
                 0.81 
                 0.1268 
                 
                   1.26 
                 
                 0.0014 
                 1284 
               
               
                 serine 
                 0.77 
                 &gt;0.1 
                 0.99 
                 0.9642 
                 0.76 
                 0.2345 
                 
                   1.15 
                 
                 0.0065 
                 1648 
               
               
                   
               
            
           
         
       
     
     Examples of biomarker metabolites that exhibit abundance profiles that support their use as diagnostic biomarkers for bladder cancer include a combination of oncometabolites that are observed in other cancers (glycerol-2-phosphate, isocitrate, glycerophosphoryl choline (GPC), isobutyryl carnitine/glycine, xanthurenate) and metabolites that are novel to bladder cancer α-hydroxybutyrate, N-acetylglutamate).  FIG. 1  provides a graphical representation of the fold-change profile for the osmolality-normalized abundance ratios between TCC and case controls for selected exemplary biomarker metabolites. A similar graphical representation could be prepared for any of the biomarker metabolites listed in Table 1. 
     In Study 2, biomarkers were discovered by (1) analyzing urine samples collected from: 89 control subjects that did not have bladder cancer (Normal), 66 subjects having bladder cancer (BCA), 58 subjects having hematuria (Hem), 48 subjects having renal cell carcinoma (RCC), and 58 subjects having prostate cancer (PCA) to determine the levels of metabolites in the samples and then (2) statistically analyzing the results to determine those metabolites that were differentially present in the groups. 
     After the levels of metabolites were determined, the data were analyzed using one-way ANOVA contrasts. Three comparisons were used to identify biomarkers for bladder cancer: Bladder cancer vs. Normal, Bladder cancer vs. Hematuria and Bladder cancer vs. Renal cell carcinoma and Prostate cancer. As listed in Table 1, the analysis of named compounds resulted in the identification of biomarkers that are differentially present between a) bladder cancer and Normal (columns 4-5) b) bladder cancer and hematuria (columns 6-7 and/or c) bladder cancer and Renal cell carcinoma+Prostate cancer (columns 8-9). 
     Table 1 includes, for each biomarker, the biochemical name of the biomarker, the fold change (FC) of the biomarker in bladder cancer compared to non-bladder cancer subjects (BCA/Normal, BCA/Hematuria and BCA/RCC+PCA) which is the ratio of the mean level of the biomarker in bladder cancer samples as compared to the non-bladder cancer mean level, and the p-value determined in the statistical analysis of the data concerning the biomarkers. Column 10 of Table 1 lists the internal identifier for that biomarker compound in the in-house chemical library of authentic standards (CompID). Metabolites with an (*) indicate statistical significance in both studies described above. Bold values indicate a fold of change with a p-value of ≦0.1. 
     Example 2 
     Classification of Subjects Based on Urine Biomarkers in Statistical 
     Models 
     A. BCA Vs. Non-Cancer 
     A number of analytical approaches can be used to evaluate the utility of the identified biomarkers for the diagnosis of a patient&#39;s condition (for example, whether the patient has bladder cancer). Below, two simple approaches were used: principal components analysis and hierarchical clustering using Pearson correlation. 
     In one analytical approach, Principal Component Analysis was carried out to create a model to classify the subjects as Control (Non-cancer) or Bladder Cancer (TCC). The data used in the Principal Component Analysis model was the osmolality-normalized data obtained from urine samples in Study 1 of Example 1 (i.e., 10 control urine samples that were collected from subjects that did not have bladder cancer, and 10 urine samples from subjects having bladder cancer (urothelial transitional cell carcinoma)). 
     Using the Principal Component Analysis derived model, it was found that 7 of 10 control subject samples were correctly classified as control while 7 of 10 bladder cancer subject samples were correctly classified as bladder cancer based on the measured level of the biomarkers. The model determined intermediate values for some individuals. The individuals with intermediate values could not be separated into one of the two groups. The intermediate group consisted of 6 subjects, 3 of which were controls and 3 of which were bladder cancer patients. A graphical depiction of the PCA results is presented in  FIG. 2 . 
     In another statistical analysis, hierarchical clustering (Pearson&#39;s correlation) was used to classify the BCA and non-cancer control subjects using the osmolality-normalized biomarker values obtained for Study 1 (i.e., 10 control urine samples that were collected from subjects that did not have bladder cancer, and 10 urine samples from subjects having bladder cancer (urothelial transitional cell carcinoma)) in Example 1. This analysis resulted in the subjects being divided into three distinct groups. One group consisted of 100% control individuals, one group consisted of 100% bladder cancer patients and one group consisted of 33% controls and 67% bladder cancer patients.  FIG. 3  provides a graphical depiction of the results of the hierarchical clustering. 
     The results from the PCA and Hierarchical clustering models provided evidence for the existence of multiple metabolic types of bladder disease and/or bladder cancer that can be distinguished using urine biomarker metabolite levels. For example, the cancer patients identified in the intermediate group may have a less aggressive form of bladder cancer or may be at an earlier stage of cancer. Distinguishing between types of cancer (e.g., less vs. more aggressive) and stage of cancer may be valuable information to a doctor determining a course of treatment. 
     In another analysis, the biomarkers identified in Example 1 were evaluated using Random Forest analysis to classify subjects as Normal or as having BCA. Urine samples from 66 BCA subjects and 89 Normal subjects (those subjects not diagnosed with BCA or other urological cancer) were used in this analysis. 
     Random Forest results show that the samples were classified with 84% prediction accuracy. The Confusion Matrix presented in Table 2 shows the number of samples predicted for each classification and the actual in each group (BCA or Normal). The “Out-of-Bag” (OOB) Error rate gives an estimate of how accurately new observations can be predicted using the Random Forest model (e.g., whether a sample is from a bladder cancer subject or a normal subject). The OOB error from this Random Forest was approximately 16%, and the model estimated that, when used on a new set of subjects, the identity of normal subjects could be predicted correctly 87% of the time and bladder cancer subjects could be predicted 80% of the time. 
     
       
         
           
               
             
               
                 TABLE 2 
               
             
            
               
                   
               
               
                 Results of Random Forest: Bladder cancer vs. Normal 
               
            
           
           
               
               
               
               
            
               
                   
                   
                 Predicted Group 
                 class. 
               
            
           
           
               
               
               
               
            
               
                   
                 BCA 
                 Normal 
                 Error 
               
               
                   
                   
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Actual 
                 BCA 
                 53 
                 13 
                 0.19697 
               
               
                   
                 Group 
                 Normal 
                 12 
                 77 
                 0.134832 
               
               
                   
                   
               
            
           
         
       
     
     Based on the OOB Error rate of 16%, the Random Forest model that was created predicted whether a sample was from an individual with bladder cancer with about 84% accuracy based on the levels of the biomarkers measured in samples from the subjects. Exemplary biomarkers for distinguishing the groups are adenosine 5′-monophosphate (AMP), 3-hydroxyphenylacetate, 2-hydroxyhippurate (salicylurate), 3-indoxyl-sulfate, phenylacetylglutamine, p-cresol-sulfate, 3-hydroxyhippurate, lactate, itaconate methylenesuccinate, cortisol, isobutyrylglycine, gluconate, xanthurenate, gulono 1,4-lactone, 3-hydroxybutyrate (BHBA), cinnamoylglycine, 2-oxindole-3-acetate, 2-hydroxybutyrate (AHB), 1-2-propanediol, alpha-CEHC-glucuronide, palmitoyl-sphingomyelin, catechol-sulfate, gamma-glutamylphenylalanine, 2-isopropylmalate, succinate, 4-hydroxyphenylacetate, pyridoxate, isovalerylglycine, carnitine, and tartarate. 
     The Random Forest analysis demonstrated that by using the biomarkers, BCA subjects were distinguished from Normal subjects with 80% sensitivity, 87% specificity, 82% PPV and 86% NPV. 
     B. BCA Vs. Other Urological Cancers 
     The biomarkers in Table 1 were used to create a statistical model to classify the subjects as having BCA or another urological cancer. Using Random Forest analysis the biomarkers were used in a mathematical model to classify subjects as having BCA or having either PCA or RCC. Urine samples from 66 BCA subjects and 106 subjects with PCA or RCC were used in this analysis. 
     Random Forest results show that the samples were classified with 83% prediction accuracy. The Confusion Matrix presented in Table 3 shows the number of samples predicted for each classification and the actual in each group (BCA or PCA+RCC). The “Out-of-Bag” (OOB) Error rate gives an estimate of how accurately new observations can be predicted using the Random Forest model (e.g., whether a sample is from a bladder cancer subject or subject with PCA or RCC). The OOB error from this Random Forest was approximately 17%, and the model estimated that, when used on a new set of subjects, the identity of BCA subjects could be predicted correctly 85% of the time and PCA+RCC subjects could be predicted 82% of the time. 
     
       
         
           
               
             
               
                 TABLE 3 
               
             
            
               
                   
               
               
                 Results of Random Forest: Bladder cancer vs. PCA + RCC 
               
            
           
           
               
               
               
               
            
               
                   
                   
                 Predicted Group 
                 class. 
               
            
           
           
               
               
               
               
            
               
                   
                 BCA 
                 PCA + RCC 
                 Error 
               
               
                   
                   
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Actual 
                 BCA 
                 56 
                 10 
                 0.151515 
               
               
                   
                 Group 
                 PCA + RCC 
                 19 
                 87 
                 0.179245 
               
               
                   
                   
               
            
           
         
       
     
     Based on the OOB Error rate of 17%, the Random Forest model that was created predicted whether a sample was from an individual with bladder cancer with about 83% accuracy based on the levels of the biomarkers measured in samples from the subjects. Exemplary biomarkers for distinguishing the groups are imidazole-propionate, 3-indoxyl-sulfate, phenylacetylglycine, lactate, choline, methyl-indole-3-acetate, beta-alanine, palmitoyl-sphingomyelin, 2-hydroxyisobutyrate, succinate, 4-androsten-3beta-17beta-diol-disulfate-2,4-hydroxyphenylacetate, glycerol, uracil, gulono 1,4-lactone, phenol sulfate, dimethylarginine (ADMA+SDMA), cyclo-gly-pro, sucrose, adenosine, serine, azelate (nonanedioate), threonine, pregnanediol-3-glucuronide, ethanolamine, gluconate, N6-methyladenosine, N-methy proline, glycine, glucose 6-phosphate (G6P). 
     The Random Forest results demonstrated that by using the biomarkers, BCA subjects were distinguished from PCA+RCC subjects, with 85% sensitivity, 82% specificity, 75% PPV, and 90% NPV. 
     C. BCA Vs. Hematuria 
     The biomarkers in Table 1 were used to create a statistical model to classify the subjects as having BCA or hematuria. Using Random Forest analysis the biomarkers were used in a mathematical model to classify subjects as having BCA or hematuria. Urine samples from 66 BCA and 58 hematuria patients were used in the analysis. 
     Random Forest results show that the samples were classified with 74% prediction accuracy. The Confusion Matrix presented in Table 4 shows the number of samples predicted for each classification and the actual in each group (BCA or Hematuria). The “Out-of-Bag” (OOB) Error rate gives an estimate of how accurately new observations can be predicted using the Random Forest model (e.g., whether a sample is from a bladder cancer subject or subject with hematuria). The OOB error from this Random Forest was approximately 26%, and the model estimated that, when used on a new set of subjects, the identity of BCA subjects could be predicted correctly 70% of the time and hematuria subjects could be predicted 79% of the time. 
     
       
         
           
               
             
               
                 TABLE 4 
               
             
            
               
                   
               
               
                 Results of Random Forest: Bladder cancer vs. Hematuria 
               
            
           
           
               
               
               
               
            
               
                   
                   
                 Predicted Group 
                 class. 
               
            
           
           
               
               
               
               
            
               
                   
                 BCA 
                 Hematuria 
                 Error 
               
               
                   
                   
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Actual 
                 BCA 
                 46 
                 20 
                 0.30303 
               
               
                   
                 Group 
                 Hematuria 
                 12 
                 46 
                 0.206897 
               
               
                   
                   
               
            
           
         
       
     
     Based on the OOB Error rate of 26%, the Random Forest model that was created predicted whether a sample was from an individual with bladder cancer with about 74% accuracy from analysis of the levels of the biomarkers measured in samples from the subject. Exemplary biomarkers for distinguishing the groups are isovalerylglycine, 2-hydroxybutyrate (AHB), 4-hydroxyhippurate, gluconate, gulono 1,4-lactone, 3-hydroxyhippurate, tartarate, 2-oxindole-3-acetate, isobutyrylglycine, catechol-sulfate, phenylacetylglutamine, succinate, 3-hydroxybutyrate (BHBA), cinnamoylglycine, isobutyrylcarnitine, 3-hydroxyphenylacetate, 3-indoxyl-sulfate, sorbose, 2-5-furandicarboxylic acid, methyl-4-hydroxybenzoate, 2-isopropylmalate, adenosine 5′-monophosphate (AMP), 2-methylbutyrylglycine, palmitoyl-sphingomyelin, phenylpropionylglycine, beta-hydroxypyruvate, tyramine, 3-methylcrotonylglycine, carnosine, fructose. 
     The Random Forest results demonstrated that by using the biomarkers, BCA subjects were distinguished from hematuria subjects, with 70% sensitivity, 79% specificity, 79% PPV, and 70% NPV. 
     Example 3 
     Biomarkers for Staging Bladder Cancer 
     Bladder cancer staging provides an indication of the extent of spreading of the bladder tumor. The tumor stage is used to select treatment options and to estimate a patient&#39;s prognosis. Bladder tumor staging ranges from T0 (no evidence of primary tumor, least advanced) to T4 (tumor has spread beyond fatty tissue surrounding the bladder into nearby organs, most advanced). Early stages of bladder cancer can also be characterized as carcinoma in situ (CIS) meaning that cells are abnormally proliferating but are still contained within the bladder. 
     To identify biomarkers of disease staging and/or progression, metabolomic analysis was carried out on urine samples from 21 subjects with Low stage BCA (CIS, T0, T1), 42 subjects with High stage BCA (T2-T4), and 89 normal subjects. After the levels of metabolites were determined, the data were analyzed using one-way ANOVA contrasts to identify biomarkers that differed between 1) Low stage bladder cancer compared to normal, 2) High stage bladder cancer compared to normal, and/or 3) Low stage bladder cancer compared to High stage bladder cancer. The identified biomarkers are listed in Table 5. 
     Table 5 includes, for each biomarker, the biochemical name of the biomarker, the fold change of the biomarker in 1) Low stage BCA compared to Normal 2) High stage BCA compared to normal 3) Low stage BCA compared to High stage BCA, and 4) bladder cancer compared to subjects with a history of bladder cancer (Example 4), and the p-value determined in the statistical analysis of the data concerning the biomarkers. Column 10 of Table 5 includes the internal identifier for the biomarker compound in the in-house chemical library of authentic standards (CompID). Bold values indicate a fold of change with a p-value of ≦0.1. 
     
       
         
           
               
             
               
                 TABLE 5 
               
             
            
               
                   
               
               
                 Biomarkers for bladder cancer staging and monitoring 
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 BCA 
                 BCA 
                 BCA Low/ 
                   
                   
               
               
                   
                 Low/Norm 
                 High/Norm 
                 BCA High 
                 BCA/HX 
                 Comp 
               
            
           
           
               
               
               
               
               
               
               
               
               
               
            
               
                 Biochemical Name 
                 FC 
                 P-value 
                 FC 
                 P-value 
                 FC 
                 P-value 
                 FC 
                 P-value 
                 ID 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
               
            
               
                 anserine 
                 
                   0.15 
                 
                 0.0096 
                 
                   0.28 
                 
                 0.0123 
                 0.52 
                 0.5492 
                 
                   0.14 
                 
                 0.0019 
                 15747 
               
               
                 pyridoxate 
                 
                   0.28 
                 
                 0.0039 
                 
                   0.35 
                 
                 0.0008 
                 0.81 
                 0.7945 
                 
                   0.3 
                 
                 9.14E−08 
                 31555 
               
               
                 adipate 
                 
                   3.15 
                 
                 0.0837 
                 
                   4.92 
                 
                 7.01E−06 
                 0.64 
                 0.1075 
                 
                   5.02 
                 
                 7.26E−08 
                 21134 
               
               
                 xanthurenate 
                 
                   0.61 
                 
                 0.0005 
                 
                   0.55 
                 
                 7.86E−09 
                 1.11 
                 0.3588 
                 
                   0.66 
                 
                 6.49E−06 
                 15679 
               
               
                 1,2-propanediol 
                 
                   5.93 
                 
                 0.0025 
                 
                   4.89 
                 
                 1.16E−06 
                 1.21 
                 0.4904 
                 
                   3.11 
                 
                 4.06E−05 
                 38002 
               
               
                 choline phosphate 
                 
                   9.74 
                 
                 8.26E−05 
                 
                   5.06 
                 
                 0.0013 
                 1.92 
                 0.179 
                 
                   4.99 
                 
                 0.0022 
                 34396 
               
               
                 acetylcarnitine 
                 
                   2.12 
                 
                 0.0006 
                 
                   2.61 
                 
                 0.0002 
                 0.81 
                 0.6464 
                 
                   2.63 
                 
                 4.61E−07 
                 32198 
               
               
                 3-hydroxybutyrate 
                 
                   42.46 
                 
                 1.27E−05 
                 
                   8.35 
                 
                 3.43E−06 
                 5.08 
                 0.4761 
                 
                   24.27 
                 
                 1.09E−10 
                 542 
               
               
                 (BHBA) 
               
               
                 palmitoyl 
                 
                   8.81 
                 
                 0.0202 
                 
                   11.64 
                 
                 1.17E−06 
                 0.76 
                 0.1816 
                 
                   8.03 
                 
                 1.96E−08 
                 37506 
               
               
                 sphingomyelin 
               
               
                 tyramine 
                 
                   0.76 
                 
                 0.0054 
                 
                   0.64 
                 
                 3.42E−05 
                 1.19 
                 0.6949 
                 
                   0.76 
                 
                 0.003 
                 1603 
               
               
                 lactate 
                 
                   3.17 
                 
                 2.41E−08 
                 
                   3.3 
                 
                 2.47E−08 
                 0.96 
                 0.238 
                 
                   3.13 
                 
                 1.39E−10 
                 527 
               
               
                 3- 
                 
                   0.29 
                 
                 7.43E−06 
                 
                   0.48 
                 
                 6.36E−09 
                 0.59 
                 0.9735 
                 
                   0.31 
                 
                 0.00E+00 
                 39600 
               
               
                 hydroxyhippurate 
               
               
                 adenosine 5′- 
                 
                   13.64 
                 
                 2.37E−10 
                 
                   25.99 
                 
                 1.12E−13 
                 0.52 
                 0.607 
                 
                   11.4 
                 
                 3.00E−14 
                 32342 
               
               
                 monophosphate 
               
               
                 (AMP) 
               
               
                 3- 
                 
                   0.29 
                 
                 4.94E−06 
                 
                   0.27 
                 
                 1.33E−13 
                 1.05 
                 0.2467 
                 
                   0.37 
                 
                 2.74E−12 
                 1413 
               
               
                 hydroxyphenylacetate 
               
               
                 phenylacetylglutamine 
                 
                   0.78 
                 
                 0.006 
                 
                   0.71 
                 
                 3.83E−11 
                 
                   1.1 
                 
                 0.0252 
                 
                   0.7 
                 
                 3.42E−12 
                 35126 
               
               
                 2,5- 
                 
                   0.22 
                 
                 8.09E−06 
                 
                   0.77 
                 
                 0.0231 
                 
                   0.28 
                 
                 0.0126 
                 
                   0.21 
                 
                 9.90E−11 
                 40809 
               
               
                 furandicarboxylic 
               
               
                 acid 
               
               
                 3-indoxyl sulfate 
                 
                   0.52 
                 
                 0.0017 
                 
                   0.53 
                 
                 3.23E−10 
                 0.98 
                 0.1017 
                 
                   0.54 
                 
                 1.26E−10 
                 27672 
               
               
                 catechol sulfate 
                 
                   0.61 
                 
                 0.0013 
                 
                   0.7 
                 
                 6.38E−06 
                 0.88 
                 0.7984 
                 
                   0.62 
                 
                 3.24E−10 
                 35320 
               
               
                 N-(2-furoyl)glycine 
                 
                   0.6 
                 
                 0.0022 
                 
                   0.5 
                 
                 0.0007 
                 1.2 
                 0.6928 
                 
                   0.48 
                 
                 4.26E−10 
                 31536 
               
               
                 2- 
                 
                   0.21 
                 
                 4.35E−07 
                 
                   0.09 
                 
                 5.51E−09 
                 2.31 
                 0.6246 
                 
                   0.17 
                 
                 6.77E−10 
                 18281 
               
               
                 hydroxyhippurate 
               
               
                 (salicylurate) 
               
               
                 2-oxindole-3- 
                 
                   0.68 
                 
                 0.0067 
                 
                   0.36 
                 
                 7.30E−12 
                 
                   1.87 
                 
                 0.0142 
                 
                   0.54 
                 
                 1.17E−09 
                 40479 
               
               
                 acetate 
               
               
                 2-isopropylmalate 
                 
                   0.2 
                 
                 3.90E−05 
                 
                   0.29 
                 
                 2.75E−08 
                 0.71 
                 0.8474 
                 
                   0.34 
                 
                 1.52E−09 
                 15667 
               
               
                 fructose 
                 
                   0.45 
                 
                 0.0013 
                 
                   0.59 
                 
                 0.0002 
                 0.75 
                 0.7814 
                 
                   0.46 
                 
                 2.10E−09 
                 577 
               
               
                 alpha-CEHC 
                 
                   0.28 
                 
                 6.07E−05 
                 
                   0.57 
                 
                 2.97E−05 
                 0.49 
                 0.4721 
                 
                   0.31 
                 
                 2.17E−09 
                 39346 
               
               
                 glucuronide 
               
               
                 p-cresol sulfate 
                 
                   0.48 
                 
                 0.0112 
                 
                   0.5 
                 
                 5.51E−11 
                 
                   0.96 
                 
                 0.0168 
                 
                   0.53 
                 
                 3.50E−09 
                 36103 
               
               
                 2,3- 
                 
                   0.23 
                 
                 5.31E−06 
                 
                   0.3 
                 
                 7.52E−06 
                 0.78 
                 0.3135 
                 
                   0.38 
                 
                 8.49E−09 
                 38276 
               
               
                 dihydroxyisovalerate 
               
               
                 4- 
                 
                   0.59 
                 
                 0.0111 
                 
                   0.88 
                 
                 0.0101 
                 0.66 
                 0.6138 
                 
                   0.52 
                 
                 1.11E−08 
                 35527 
               
               
                 hydroxyhippurate 
               
               
                 isovalerylglycine 
                 
                   0.53 
                 
                 0.0272 
                 
                   0.55 
                 
                 2.91E−06 
                 0.97 
                 0.1912 
                 
                   0.53 
                 
                 1.32E−08 
                 35107 
               
               
                 isobutyrylglycine 
                 
                   0.86 
                 
                 0.0057 
                 
                   0.51 
                 
                 1.85E−07 
                 1.67 
                 0.2334 
                 
                   0.61 
                 
                 1.53E−08 
                 35437 
               
               
                 4- 
                 
                   0.5 
                 
                 0.0135 
                 
                   0.46 
                 
                 3.24E−10 
                 
                   1.09 
                 
                 0.0244 
                 
                   0.61 
                 
                 2.41E−08 
                 541 
               
               
                 hydroxyphenylacetate 
               
               
                 sorbose 
                 
                   0.39 
                 
                 0.0033 
                 
                   0.53 
                 
                 1.65E−05 
                 0.75 
                 0.7118 
                 
                   0.44 
                 
                 3.18E−08 
                 563 
               
               
                 pimelate 
                 
                   0.61 
                 
                 0.0657 
                 
                   0.47 
                 
                 1.88E−05 
                 1.29 
                 0.1757 
                 
                   0.55 
                 
                 1.15E−07 
                 15704 
               
               
                 (heptanedioate) 
               
               
                 2-hydroxybutyrate 
                 
                   5.12 
                 
                 3.40E−05 
                 
                   1.99 
                 
                 0.0012 
                 2.57 
                 0.1293 
                 
                   3.29 
                 
                 1.36E−07 
                 21044 
               
               
                 (AHB) 
               
               
                 3- 
                 
                   0.54 
                 
                 0.0167 
                 
                   0.62 
                 
                 0.002 
                 0.88 
                 0.9956 
                 
                   0.52 
                 
                 1.75E−07 
                 31940 
               
               
                 methylcrotonylglycine 
               
               
                 arginine 
                 
                   0.29 
                 
                 0.0127 
                 
                   0.45 
                 
                 0.011 
                 0.65 
                 0.6295 
                 
                   0.14 
                 
                 2.00E−07 
                 1638 
               
               
                 tartarate 
                 
                   0.04 
                 
                 1.36E−06 
                 
                   0.36 
                 
                 0.0023 
                 
                   0.1 
                 
                 0.0218 
                 
                   0.29 
                 
                 2.24E−07 
                 15336 
               
               
                 galactitol (dulcitol) 
                 
                   0.62 
                 
                 0.0013 
                 
                   0.81 
                 
                 0.0494 
                 0.77 
                 0.1209 
                 
                   0.61 
                 
                 2.31E−07 
                 1117 
               
               
                 allantoin 
                 0.58 
                 0.1251 
                 0.61 
                 0.1611 
                 0.94 
                 0.6809 
                 
                   0.47 
                 
                 2.39E−07 
                 1107 
               
               
                 3-(3- 
                 
                   0.34 
                 
                 0.0394 
                 
                   0.57 
                 
                 0.002 
                 0.59 
                 0.7634 
                 
                   0.27 
                 
                 2.42E−07 
                 35635 
               
               
                 hydroxyphenyl)propionate 
               
               
                 succinate 
                 
                   0.53 
                 
                 0.0013 
                 
                   0.65 
                 
                 0.0003 
                 0.81 
                 0.708 
                 
                   0.51 
                 
                 2.95E−07 
                 1437 
               
               
                 cinnamoylglycine 
                 
                   0.49 
                 
                 0.0225 
                 
                   0.5 
                 
                 7.09E−07 
                 0.99 
                 0.1486 
                 
                   0.4 
                 
                 1.08E−06 
                 38637 
               
               
                 gluconate 
                 
                   7.43 
                 
                 0.0201 
                 
                   12.6 
                 
                 4.81E−08 
                 
                   0.59 
                 
                 0.0767 
                 
                   9.04 
                 
                 1.58E−06 
                 2913 
               
               
                 glutathione, 
                 
                   1.88 
                 
                 0.0307 
                 
                   10.41 
                 
                 0.0038 
                 0.18 
                 0.9443 
                 
                   9.27 
                 
                 2.38E−06 
                 2127 
               
               
                 reduced (GSH) 
               
               
                 pyridoxal 
                 0.54 
                 0.2705 
                 
                   0.36 
                 
                 4.44E−05 
                 
                   1.51 
                 
                 0.0597 
                 
                   0.34 
                 
                 2.55E−06 
                 1651 
               
               
                 methyl-4- 
                 
                   5.72 
                 
                 0.0149 
                 
                   8.75 
                 
                 5.15E−06 
                 0.65 
                 0.3103 
                 
                   0.44 
                 
                 3.45E−06 
                 34386 
               
               
                 hydroxybenzoate 
               
               
                 phenylacetylglycine 
                 
                   0.53 
                 
                 0.0461 
                 
                   0.57 
                 
                 0.0003 
                 0.93 
                 0.4435 
                 
                   0.52 
                 
                 3.61E−06 
                 33945 
               
               
                 vanillate 
                 
                   0.46 
                 
                 0.0159 
                 
                   1.18 
                 
                 0.032 
                 0.39 
                 0.4904 
                 
                   0.78 
                 
                 5.18E−06 
                 35639 
               
               
                 lactose 
                 
                   0.5 
                 
                 0.0691 
                 
                   0.53 
                 
                 0.0018 
                 0.95 
                 0.5837 
                 
                   0.52 
                 
                 7.94E−06 
                 567 
               
               
                 cortisol 
                 
                   2.93 
                 
                 0.0004 
                 
                   2.48 
                 
                 4.45E−07 
                 1.18 
                 0.7168 
                 
                   1.94 
                 
                 1.00E−05 
                 1712 
               
               
                 3- 
                 
                   0.76 
                 
                 0.0841 
                 
                   1.26 
                 
                 0.0133 
                 0.6 
                 0.8659 
                 
                   0.87 
                 
                 1.32E−05 
                 40264 
               
               
                 phosphoglycerate 
               
               
                 alpha-tocopherol 
                 
                   3.36 
                 
                 0.0002 
                 
                   7.65 
                 
                 2.23E−05 
                 0.44 
                 0.6685 
                 
                   3.78 
                 
                 1.35E−05 
                 1561 
               
               
                 N-acetyltyrosine 
                 
                   0.67 
                 
                 0.0866 
                 
                   0.68 
                 
                 0.0019 
                 0.99 
                 0.5321 
                 
                   0.6 
                 
                 1.66E−05 
                 32390 
               
               
                 2- 
                 
                   0.66 
                 
                 0.0171 
                 
                   0.69 
                 
                 0.0012 
                 0.95 
                 0.908 
                 
                   0.65 
                 
                 1.66E−05 
                 31928 
               
               
                 methylbutyrylglycine 
               
               
                 N- 
                 
                   0.57 
                 
                 0.0073 
                 
                   0.61 
                 
                 0.0012 
                 0.94 
                 0.8632 
                 
                   0.5 
                 
                 1.74E−05 
                 33950 
               
               
                 acetylphenylalanine 
               
               
                 phenylpropionylglycine 
                 
                   0.47 
                 
                 0.0013 
                 
                   0.51 
                 
                 3.64E−05 
                 0.92 
                 0.9793 
                 
                   0.47 
                 
                 1.78E−05 
                 35434 
               
               
                 N-acetyltryptophan 
                 
                   0.54 
                 
                 0.0096 
                 
                   0.47 
                 
                 0.0036 
                 1.13 
                 0.7579 
                 
                   0.45 
                 
                 1.85E−05 
                 33959 
               
               
                 xanthine 
                 
                   1.55 
                 
                 0.0322 
                 
                   1.24 
                 
                 0.0019 
                 1.25 
                 0.8123 
                 
                   1.6 
                 
                 1.97E−05 
                 3147 
               
               
                 1,6- 
                 
                   0.47 
                 
                 0.0145 
                 
                   0.79 
                 
                 0.034 
                 0.6 
                 0.4613 
                 
                   0.45 
                 
                 2.20E−05 
                 21049 
               
               
                 anhydroglucose 
               
               
                 galactinol 
                 
                   0.45 
                 
                 0.036 
                 
                   0.48 
                 
                 0.0006 
                 0.93 
                 0.6031 
                 
                   0.48 
                 
                 2.80E−05 
                 21034 
               
               
                 hexanoylglycine 
                 
                   1.43 
                 
                 0.0156 
                 
                   1.69 
                 
                 0.0001 
                 0.85 
                 0.5966 
                 
                   1.88 
                 
                 2.86E−05 
                 35436 
               
               
                 azelate 
                 0.79 
                 0.2568 
                 0.8 
                 0.3391 
                 0.99 
                 0.7188 
                 
                   0.59 
                 
                 3.42E−05 
                 18362 
               
               
                 (nonanedioate) 
               
               
                 guanidine 
                 
                   0.55 
                 
                 0.0112 
                 
                   0.47 
                 
                 5.16E−05 
                 1.17 
                 0.5811 
                 
                   0.53 
                 
                 7.08E−05 
                 22287 
               
               
                 N-methylglutamate 
                 
                   0.71 
                 
                 0.0495 
                 
                   1.05 
                 
                 0.0015 
                 0.68 
                 0.6544 
                 
                   0.78 
                 
                 7.34E−05 
                 31532 
               
               
                 galactose 
                 
                   0.69 
                 
                 0.0372 
                 
                   0.69 
                 
                 0.0646 
                 0.99 
                 0.5489 
                 
                   0.51 
                 
                 7.39E−05 
                 12055 
               
               
                 mandelate 
                 
                   0.63 
                 
                 0.0094 
                 
                   0.88 
                 
                 0.0246 
                 0.71 
                 0.431 
                 
                   0.76 
                 
                 7.93E−05 
                 22160 
               
               
                 5-acetylamino-6- 
                 
                   0.42 
                 
                 0.0422 
                 
                   0.54 
                 
                 0.0276 
                 0.78 
                 0.7629 
                 
                   0.4 
                 
                 8.18E−05 
                 34424 
               
               
                 amino-3- 
               
               
                 methyluracil 
               
               
                 riboflavin (Vitamin 
                 
                   0.14 
                 
                 0.0004 
                 
                   0.29 
                 
                 0.0071 
                 0.5 
                 0.1843 
                 
                   0.18 
                 
                 8.95E−05 
                 1827 
               
               
                 B2) 
               
               
                 4- 
                 
                   0.57 
                 
                 0.0045 
                 
                   0.7 
                 
                 0.0003 
                 0.81 
                 0.9468 
                 
                   0.71 
                 
                 9.54E−05 
                 1568 
               
               
                 hydroxymandelate 
               
               
                 glutathione, 
                 1.09 
                 0.4356 
                 
                   2.92 
                 
                 2.26E−07 
                 
                   0.37 
                 
                 0.0031 
                 
                   2.14 
                 
                 9.65E−05 
                 38783 
               
               
                 oxidized (GSSG) 
               
               
                 prostaglandin E2 
                 
                   1.74 
                 
                 2.03E−06 
                 1.22 
                 0.1192 
                 
                   1.42 
                 
                 0.0011 
                 
                   1.41 
                 
                 9.79E−05 
                 7746 
               
               
                 cortisone 
                 
                   1.37 
                 
                 0.0047 
                 
                   1.38 
                 
                 0.0002 
                 0.99 
                 0.9283 
                 
                   1.4 
                 
                 0.0001 
                 1769 
               
               
                 biotin 
                 
                   0.4 
                 
                 0.0073 
                 
                   0.57 
                 
                 0.0185 
                 0.7 
                 0.4307 
                 
                   0.46 
                 
                 0.0001 
                 568 
               
               
                 dihydroferulic acid 
                 1.02 
                 0.3165 
                 
                   0.62 
                 
                 0.0234 
                 1.66 
                 0.4951 
                 
                   0.45 
                 
                 0.0001 
                 40481 
               
               
                 N-acetylproline 
                 
                   0.71 
                 
                 0.0589 
                 
                   0.71 
                 
                 0.006 
                 1 
                 0.8309 
                 
                   0.65 
                 
                 0.0002 
                 34387 
               
               
                 glucono-1,5- 
                 
                   5.2 
                 
                 0.0012 
                 
                   10.85 
                 
                 4.63E−05 
                 0.48 
                 0.9433 
                 
                   6.06 
                 
                 0.0002 
                 32355 
               
               
                 lactone 
               
               
                 3-hydroxysebacate 
                 
                   3.06 
                 
                 0.0123 
                 
                   1.61 
                 
                 0.0107 
                 1.9 
                 0.6255 
                 
                   2.31 
                 
                 0.0002 
                 31943 
               
               
                 pantothenate 
                 
                   0.42 
                 
                 0.0067 
                 
                   0.64 
                 
                 0.0191 
                 0.65 
                 0.4084 
                 
                   0.48 
                 
                 0.0002 
                 1508 
               
               
                 4-hydroxybenzoate 
                 0.68 
                 0.1172 
                 
                   0.82 
                 
                 0.0879 
                 0.82 
                 0.8207 
                 
                   0.55 
                 
                 0.0002 
                 21133 
               
               
                 3- 
                 0.58 
                 0.2213 
                 
                   0.73 
                 
                 0.0709 
                 0.8 
                 0.8757 
                 
                   0.46 
                 
                 0.0002 
                 20698 
               
               
                 hydroxycinnamate 
               
               
                 (m-coumarate) 
               
               
                 guanidinoacetate 
                 
                   0.85 
                 
                 0.1638 
                 
                   0.53 
                 
                 0.0004 
                 1.61 
                 0.2141 
                 
                   0.52 
                 
                 0.0003 
                 12359 
               
               
                 mesaconate 
                 
                   0.66 
                 
                 0.0305 
                 
                   0.63 
                 
                 0.0044 
                 1.05 
                 0.9721 
                 
                   0.64 
                 
                 0.0004 
                 18493 
               
               
                 (methylfumarate) 
               
               
                 4-methyl-2- 
                 
                   2.02 
                 
                 0.0219 
                 
                   1.52 
                 
                 0.1157 
                 1.33 
                 0.3254 
                 
                   1.94 
                 
                 0.0005 
                 22116 
               
               
                 oxopentanoate 
               
               
                 7-methylguanine 
                 
                   1.23 
                 
                 0.0471 
                 
                   1.23 
                 
                 0.0027 
                 1 
                 0.7612 
                 
                   1.32 
                 
                 0.0005 
                 35114 
               
               
                 imidazole 
                 
                   1.63 
                 
                 0.0283 
                 1.02 
                 0.1385 
                 1.6 
                 0.3388 
                 
                   1.48 
                 
                 0.0006 
                 40730 
               
               
                 propionate 
               
               
                 N-acetylcysteine 
                 0.8 
                 0.1625 
                 
                   0.61 
                 
                 0.0108 
                 1.31 
                 0.6004 
                 
                   0.61 
                 
                 0.0006 
                 1586 
               
               
                 alpha- 
                 1.38 
                 0.2943 
                 1.38 
                 0.1531 
                 1 
                 0.9607 
                 
                   1.48 
                 
                 0.0006 
                 528 
               
               
                 ketoglutarate 
               
               
                 adenosine 
                 
                   0.72 
                 
                 0.0176 
                 
                   0.72 
                 
                 9.45E−05 
                 1.01 
                 0.5505 
                 
                   0.82 
                 
                 0.0006 
                 555 
               
               
                 3-hydroxybenzoate 
                 0.83 
                 0.6258 
                 
                   0.79 
                 
                 0.0378 
                 1.05 
                 0.3102 
                 
                   0.66 
                 
                 0.0007 
                 15673 
               
               
                 sinapate 
                 0.6 
                 0.5402 
                 
                   0.63 
                 
                 0.0759 
                 0.95 
                 0.4906 
                 
                   0.45 
                 
                 0.0007 
                 21150 
               
               
                 N- 
                 
                   0.57 
                 
                 0.0372 
                 
                   0.37 
                 
                 0.0059 
                 1.54 
                 0.9664 
                 
                   0.52 
                 
                 0.0008 
                 1594 
               
               
                 carbamoylaspartate 
               
               
                 threitol 
                 0.9 
                 0.186 
                 
                   0.96 
                 
                 0.0065 
                 0.94 
                 0.4759 
                 
                   0.85 
                 
                 0.0008 
                 35854 
               
               
                 N- 
                 
                   0.79 
                 
                 0.0588 
                 
                   0.93 
                 
                 0.0666 
                 0.85 
                 0.6664 
                 
                   0.8 
                 
                 0.001 
                 38696 
               
               
                 carbamoylsarcosine 
               
               
                 sucrose 
                 
                   0.21 
                 
                 0.0014 
                 
                   0.58 
                 
                 0.0716 
                 0.37 
                 0.1005 
                 
                   0.42 
                 
                 0.001 
                 1519 
               
               
                 biliverdin 
                 1.05 
                 0.3876 
                 
                   1.29 
                 
                 8.37E−06 
                 
                   0.81 
                 
                 0.018 
                 
                   1.17 
                 
                 0.0011 
                 2137 
               
               
                 tryptophan 
                 1.26 
                 0.1227 
                 
                   1.35 
                 
                 0.0057 
                 0.93 
                 0.5886 
                 
                   1.29 
                 
                 0.0013 
                 54 
               
               
                 carnitine 
                 
                   1.92 
                 
                 0.0054 
                 
                   1.7 
                 
                 0.0031 
                 1.13 
                 0.6518 
                 
                   1.53 
                 
                 0.0013 
                 15500 
               
               
                 hexanoylcarnitine 
                 1.21 
                 0.1114 
                 1.23 
                 0.1105 
                 0.98 
                 0.7431 
                 
                   1.57 
                 
                 0.0017 
                 32328 
               
               
                 cytidine 
                 1 
                 0.8018 
                 0.76 
                 0.1284 
                 1.31 
                 0.4019 
                 
                   0.62 
                 
                 0.0017 
                 514 
               
               
                 trans-aconitate 
                 
                   0.72 
                 
                 0.0443 
                 
                   0.8 
                 
                 0.0426 
                 0.9 
                 0.6843 
                 
                   0.66 
                 
                 0.0018 
                 27741 
               
               
                 3,4- 
                 
                   0.56 
                 
                 0.0049 
                 
                   0.41 
                 
                 4.08E−05 
                 1.36 
                 0.7377 
                 
                   0.57 
                 
                 0.0019 
                 18296 
               
               
                 dihydroxyphenylacetate 
               
               
                 abscisate 
                 
                   0.35 
                 
                 0.0243 
                 
                   0.58 
                 
                 0.0826 
                 0.61 
                 0.4052 
                 
                   0.4 
                 
                 0.0019 
                 21156 
               
               
                 3-methyl-2- 
                 
                   2.24 
                 
                 0.0277 
                 
                   1.37 
                 
                 0.0293 
                 1.63 
                 0.6363 
                 
                   1.59 
                 
                 0.002 
                 15676 
               
               
                 oxovalerate 
               
               
                 4-hydroxy-2- 
                 
                   3.43 
                 
                 0.0025 
                 
                   2.36 
                 
                 0.0076 
                 1.45 
                 0.377 
                 
                   1.82 
                 
                 0.0021 
                 40062 
               
               
                 oxoglutaric acid 
               
               
                 decanoylcarnitine 
                 1.05 
                 0.2984 
                 1.06 
                 0.4857 
                 0.99 
                 0.6484 
                 
                   1.37 
                 
                 0.0021 
                 33941 
               
               
                 ciliatine (2- 
                 
                   3.98 
                 
                 0.02 
                 
                   0.99 
                 
                 0.028 
                 4 
                 0.5652 
                 
                   0.23 
                 
                 0.0022 
                 15125 
               
               
                 aminoethylphosphonate) 
               
               
                 3-hydroxypyridine 
                 0.66 
                 0.1359 
                 
                   0.79 
                 
                 0.0529 
                 0.83 
                 0.9971 
                 
                   0.72 
                 
                 0.0023 
                 21169 
               
               
                 xylonate 
                 
                   0.74 
                 
                 0.0609 
                 0.97 
                 0.2164 
                 0.76 
                 0.402 
                 
                   0.79 
                 
                 0.0025 
                 35638 
               
               
                 itaconate 
                 
                   0.47 
                 
                 0.0011 
                 
                   0.64 
                 
                 0.0007 
                 0.74 
                 0.5571 
                 
                   0.7 
                 
                 0.0027 
                 18373 
               
               
                 (methylenesuccinate) 
               
               
                 isoleucine 
                 
                   1.36 
                 
                 0.0327 
                 
                   1.47 
                 
                 0.0025 
                 0.92 
                 0.8564 
                 
                   1.36 
                 
                 0.0028 
                 1125 
               
               
                 5- 
                 
                   0.75 
                 
                 0.0769 
                 
                   0.62 
                 
                 0.0143 
                 1.21 
                 0.9099 
                 
                   0.71 
                 
                 0.0029 
                 31938 
               
               
                 hydroxyhexanoate 
               
               
                 4-vinylphenol 
                 
                   0.62 
                 
                 0.0204 
                 
                   0.87 
                 
                 0.0463 
                 0.71 
                 0.4759 
                 
                   0.67 
                 
                 0.0029 
                 36098 
               
               
                 sulfate 
               
               
                 hippurate 
                 1.01 
                 0.73 
                 0.97 
                 0.1615 
                 1.05 
                 0.5039 
                 
                   0.83 
                 
                 0.003 
                 15753 
               
               
                 threonate 
                 
                   0.53 
                 
                 0.0235 
                 0.75 
                 0.1883 
                 0.71 
                 0.2549 
                 
                   0.69 
                 
                 0.0033 
                 27738 
               
               
                 asparagine 
                 
                   0.71 
                 
                 0.0061 
                 0.9 
                 0.5505 
                 
                   0.79 
                 
                 0.038 
                 
                   0.78 
                 
                 0.0036 
                 11398 
               
               
                 leucine 
                 
                   1.26 
                 
                 0.0544 
                 
                   1.4 
                 
                 0.0031 
                 0.9 
                 0.7408 
                 
                   1.27 
                 
                 0.0046 
                 60 
               
               
                 4-ureidobutyrate 
                 0.85 
                 0.1552 
                 0.9 
                 0.1399 
                 0.95 
                 0.7976 
                 
                   0.86 
                 
                 0.0046 
                 22118 
               
               
                 cystine 
                 
                   0.36 
                 
                 0.0104 
                 
                   0.32 
                 
                 0.0003 
                 1.13 
                 0.8187 
                 
                   0.22 
                 
                 0.0048 
                 39512 
               
               
                 2-octenedioate 
                 0.83 
                 0.263 
                 
                   0.72 
                 
                 0.0359 
                 1.15 
                 0.6479 
                 
                   0.61 
                 
                 0.005 
                 35120 
               
               
                 tigloylglycine 
                 0.84 
                 0.5305 
                 
                   0.79 
                 
                 0.0708 
                 1.06 
                 0.485 
                 
                   0.73 
                 
                 0.0053 
                 1598 
               
               
                 1-methylhistidine 
                 
                   0.6 
                 
                 0.0038 
                 
                   0.52 
                 
                 0.006 
                 1.17 
                 0.4792 
                 
                   0.71 
                 
                 0.0055 
                 30460 
               
               
                 3-hydroxyproline 
                 0.99 
                 0.7365 
                 
                   0.7 
                 
                 0.0134 
                 1.43 
                 0.1524 
                 
                   0.66 
                 
                 0.0058 
                 38635 
               
               
                 L-urobilin 
                 
                   0.54 
                 
                 0.0307 
                 
                   0.92 
                 
                 0.0002 
                 0.59 
                 0.5146 
                 
                   0.78 
                 
                 0.0061 
                 40173 
               
               
                 2-pyrrolidinone 
                 
                   0.6 
                 
                 0.0333 
                 
                   0.54 
                 
                 0.0006 
                 1.11 
                 0.6345 
                 
                   0.71 
                 
                 0.0061 
                 31675 
               
               
                 N-acetylhistidine 
                 1.06 
                 0.6617 
                 
                   0.87 
                 
                 0.0146 
                 1.22 
                 0.1877 
                 
                   0.91 
                 
                 0.0062 
                 33946 
               
               
                 urate 
                 1.09 
                 0.2354 
                 
                   1.24 
                 
                 0.0014 
                 0.88 
                 0.2433 
                 
                   1.2 
                 
                 0.0062 
                 1604 
               
               
                 nicotinate 
                 0.78 
                 0.4081 
                 0.91 
                 0.3085 
                 0.86 
                 0.9702 
                 
                   0.78 
                 
                 0.0063 
                 1504 
               
               
                 mannose 
                 
                   0.42 
                 
                 0.0053 
                 0.81 
                 0.4314 
                 
                   0.52 
                 
                 0.0469 
                 
                   0.71 
                 
                 0.0068 
                 584 
               
               
                 arabonate 
                 
                   0.77 
                 
                 0.0942 
                 
                   0.87 
                 
                 0.08 
                 0.88 
                 0.7692 
                 
                   0.82 
                 
                 0.007 
                 37516 
               
               
                 5-aminovalerate 
                 
                   0.87 
                 
                 0.0362 
                 
                   1.85 
                 
                 0.0057 
                 0.47 
                 0.968 
                 
                   1.69 
                 
                 0.0073 
                 18319 
               
               
                 3-hydroxy-2- 
                 
                   2.5 
                 
                 0.0341 
                 1.14 
                 0.839 
                 
                   2.19 
                 
                 0.0747 
                 
                   1.72 
                 
                 0.0074 
                 32397 
               
               
                 ethylpropionate 
               
               
                 allo-threonine 
                 
                   0.61 
                 
                 0.0077 
                 
                   0.72 
                 
                 0.0356 
                 0.85 
                 0.3413 
                 
                   0.76 
                 
                 0.0085 
                 15142 
               
               
                 2-methylhippurate 
                 
                   2.32 
                 
                 0.0147 
                 
                   3.37 
                 
                 9.60E−05 
                 0.69 
                 0.5926 
                 
                   2.51 
                 
                 0.0088 
                 15670 
               
               
                 1,3,7- 
                 
                   0.53 
                 
                 0.0108 
                 0.77 
                 0.2809 
                 0.69 
                 0.1177 
                 
                   0.8 
                 
                 0.009 
                 34404 
               
               
                 trimethylurate 
               
               
                 5- 
                 
                   0.33 
                 
                 0.009 
                 
                   0.47 
                 
                 0.0126 
                 0.71 
                 0.5296 
                 
                   0.47 
                 
                 0.0093 
                 18330 
               
               
                 methyltetrahydrofolate 
               
               
                 (5MeTHF) 
               
               
                 octanoylcarnitine 
                 1.01 
                 0.2432 
                 1.08 
                 0.3015 
                 0.93 
                 0.7368 
                 
                   1.4 
                 
                 0.0097 
                 33936 
               
               
                 gamma- 
                 
                   0.67 
                 
                 0.0169 
                 
                   0.48 
                 
                 5.03E−05 
                 1.4 
                 0.4892 
                 
                   0.76 
                 
                 0.0098 
                 1416 
               
               
                 aminobutyrate 
               
               
                 (GABA) 
               
               
                 valine 
                 
                   1.35 
                 
                 0.0213 
                 
                   1.37 
                 
                 0.008 
                 0.98 
                 0.8163 
                 
                   1.24 
                 
                 0.0106 
                 1649 
               
               
                 scyllo-inositol 
                 
                   0.59 
                 
                 0.0342 
                 
                   0.87 
                 
                 0.039 
                 0.68 
                 0.633 
                 
                   0.75 
                 
                 0.011 
                 32379 
               
               
                 glutamine 
                 
                   1.61 
                 
                 0.0063 
                 
                   1.69 
                 
                 0.0005 
                 0.95 
                 0.9782 
                 
                   1.28 
                 
                 0.0113 
                 53 
               
               
                 hypoxanthine 
                 1.28 
                 0.8467 
                 0.97 
                 0.8964 
                 1.33 
                 0.9328 
                 
                   1.23 
                 
                 0.0122 
                 3127 
               
               
                 gamma- 
                 
                   1.29 
                 
                 0.0275 
                 
                   1.59 
                 
                 1.24E−05 
                 0.81 
                 0.2763 
                 
                   1.16 
                 
                 0.0123 
                 33422 
               
               
                 glutamylphenylalanine 
               
               
                 glycerol 
                 3.88 
                 0.2355 
                 
                   3.91 
                 
                 0.0056 
                 0.99 
                 0.3841 
                 
                   2.62 
                 
                 0.0125 
                 15122 
               
               
                 homoserine 
                 1.28 
                 0.1829 
                 
                   0.76 
                 
                 0.0108 
                 1.68 
                 0.5611 
                 
                   1.02 
                 
                 0.0127 
                 23642 
               
               
                 2-oxo-1- 
                 1.36 
                 0.9636 
                 1 
                 0.1468 
                 1.36 
                 0.29 
                 
                   0.93 
                 
                 0.013 
                 40452 
               
               
                 pyrrolidinepropionate 
               
               
                 creatine 
                 0.47 
                 0.3733 
                 
                   0.24 
                 
                 0.0005 
                 
                   1.95 
                 
                 0.0991 
                 
                   0.45 
                 
                 0.0133 
                 27718 
               
               
                 quinate 
                 0.73 
                 0.157 
                 0.91 
                 0.5956 
                 
                   0.8 
                 
                 0.0978 
                 
                   0.64 
                 
                 0.0134 
                 18335 
               
               
                 kynurenine 
                 
                   1.77 
                 
                 0.0116 
                 
                   2.46 
                 
                 0.0006 
                 0.72 
                 0.9028 
                 
                   1.71 
                 
                 0.0139 
                 15140 
               
               
                 3-methylxanthine 
                 
                   0.52 
                 
                 0.0038 
                 
                   0.67 
                 
                 0.0529 
                 0.78 
                 0.1998 
                 
                   0.78 
                 
                 0.0141 
                 32445 
               
               
                 beta- 
                 1.46 
                 0.3252 
                 
                   2 
                 
                 0.0012 
                 0.73 
                 0.1649 
                 
                   1.94 
                 
                 0.0143 
                 15686 
               
               
                 hydroxypyruvate 
               
               
                 maltose 
                 
                   7.21 
                 
                 0.016 
                 1.02 
                 0.9379 
                 
                   7.05 
                 
                 0.0328 
                 
                   5.1 
                 
                 0.0143 
                 15806 
               
               
                 bilirubin (E,E) 
                 1.03 
                 0.9642 
                 1.25 
                 0.1453 
                 0.82 
                 0.2885 
                 
                   1.31 
                 
                 0.0146 
                 32586 
               
               
                 1,7-dimethylurate 
                 
                   0.79 
                 
                 0.0412 
                 0.89 
                 0.1693 
                 0.89 
                 0.3717 
                 
                   0.87 
                 
                 0.0155 
                 34400 
               
               
                 phenol sulfate 
                 0.94 
                 0.1749 
                 0.99 
                 0.1722 
                 0.95 
                 0.7829 
                 
                   0.81 
                 
                 0.016 
                 32553 
               
               
                 2-hydroxyadipate 
                 
                   0.8 
                 
                 0.0538 
                 
                   0.78 
                 
                 0.0108 
                 1.02 
                 0.9699 
                 
                   0.86 
                 
                 0.0169 
                 31934 
               
               
                 isobutyrylcarnitine 
                 0.86 
                 0.2524 
                 
                   0.74 
                 
                 5.02E−05 
                 
                   1.17 
                 
                 0.0681 
                 
                   0.89 
                 
                 0.0175 
                 33441 
               
               
                 glycolithocholate 
                 0.62 
                 0.2926 
                 
                   0.83 
                 
                 0.0042 
                 0.75 
                 0.2897 
                 
                   0.94 
                 
                 0.0175 
                 32620 
               
               
                 sulfate 
               
               
                 cis-aconitate 
                 0.9 
                 0.2137 
                 
                   0.88 
                 
                 0.0723 
                 1.02 
                 0.8952 
                 
                   0.88 
                 
                 0.018 
                 12025 
               
               
                 nicotinurate 
                 26.73 
                 4.16E−06 
                 1.02 
                 0.8223 
                 
                   26.29 
                 
                 5.47E−05 
                 
                   9.46 
                 
                 0.0186 
                 35121 
               
               
                 N1-Methyl-2- 
                 1.24 
                 0.467 
                 
                   0.76 
                 
                 0.0089 
                 1.62 
                 0.2382 
                 
                   0.9 
                 
                 0.0191 
                 40469 
               
               
                 pyridone-5- 
               
               
                 carboxamide 
               
               
                 sebacate 
                 
                   8.81 
                 
                 0.0187 
                 
                   1.9 
                 
                 0.0677 
                 4.63 
                 0.3905 
                 
                   4.11 
                 
                 0.0192 
                 32398 
               
               
                 (decanedioate) 
               
               
                 gulono-1,4-lactone 
                 1.36 
                 0.8894 
                 
                   2.16 
                 
                 0.0001 
                 
                   0.63 
                 
                 0.0093 
                 
                   1.71 
                 
                 0.0192 
                 33454 
               
               
                 pipecolate 
                 0.58 
                 0.6011 
                 0.64 
                 0.3911 
                 0.91 
                 0.8995 
                 
                   0.38 
                 
                 0.0213 
                 1444 
               
               
                 2- 
                 1.18 
                 0.4755 
                 1.04 
                 0.9233 
                 1.13 
                 0.5616 
                 
                   1.24 
                 
                 0.0218 
                 22030 
               
               
                 hydroxyisobutyrate 
               
               
                 citramalate 
                 0.9 
                 0.388 
                 
                   0.79 
                 
                 0.0418 
                 1.15 
                 0.519 
                 
                   0.76 
                 
                 0.022 
                 22158 
               
               
                 diglycerol 
                 0.87 
                 0.5057 
                 0.99 
                 0.6501 
                 0.88 
                 0.7745 
                 
                   0.77 
                 
                 0.0253 
                 40700 
               
               
                 3-hydroxyglutarate 
                 
                   0.57 
                 
                 0.0022 
                 
                   0.77 
                 
                 0.0907 
                 0.75 
                 0.1091 
                 
                   0.78 
                 
                 0.0257 
                 36863 
               
               
                 guanosine 
                 
                   1.39 
                 
                 0.0482 
                 1.07 
                 0.979 
                 
                   1.3 
                 
                 0.0758 
                 
                   1.37 
                 
                 0.0258 
                 1573 
               
               
                 sorbitol 
                 
                   0.28 
                 
                 0.0223 
                 
                   0.19 
                 
                 0.0039 
                 1.44 
                 0.9596 
                 
                   0.83 
                 
                 0.0266 
                 15053 
               
               
                 glycylglycine 
                 0.79 
                 0.4324 
                 0.97 
                 0.2068 
                 0.82 
                 0.8632 
                 
                   0.88 
                 
                 0.0271 
                 21030 
               
               
                 glucosamine 
                 
                   0.47 
                 
                 0.0241 
                 
                   0.46 
                 
                 0.0086 
                 1.02 
                 0.8381 
                 
                   0.44 
                 
                 0.0276 
                 18534 
               
               
                 3-methylhistidine 
                 0.61 
                 0.2197 
                 
                   0.73 
                 
                 0.043 
                 0.84 
                 0.7596 
                 
                   0.72 
                 
                 0.0287 
                 15677 
               
               
                 lysine 
                 
                   0.64 
                 
                 0.04 
                 0.59 
                 0.2041 
                 1.07 
                 0.3279 
                 
                   0.36 
                 
                 0.0288 
                 1301 
               
               
                 ethanolamine 
                 0.74 
                 0.2141 
                 
                   0.62 
                 
                 0.0088 
                 1.19 
                 0.4755 
                 
                   0.71 
                 
                 0.0288 
                 1497 
               
               
                 cystathionine 
                 1.09 
                 0.5655 
                 
                   0.5 
                 
                 0.0291 
                 2.16 
                 0.3121 
                 
                   0.74 
                 
                 0.0289 
                 15705 
               
               
                 ethylmalonate 
                 0.99 
                 0.6578 
                 1.09 
                 0.454 
                 0.9 
                 0.9026 
                 
                   1.28 
                 
                 0.0299 
                 15765 
               
               
                 gamma- 
                 
                   0.67 
                 
                 0.0572 
                 0.76 
                 0.1375 
                 0.88 
                 0.4913 
                 
                   0.75 
                 
                 0.0306 
                 18369 
               
               
                 glutamylleucine 
               
               
                 taurolithocholate 3- 
                 0.61 
                 0.2158 
                 
                   0.72 
                 
                 0.0095 
                 0.85 
                 0.4838 
                 
                   0.74 
                 
                 0.0311 
                 36850 
               
               
                 sulfate 
               
               
                 carnosine 
                 0.47 
                 0.2097 
                 
                   0.58 
                 
                 0.0681 
                 0.81 
                 0.8881 
                 
                   0.39 
                 
                 0.0331 
                 1768 
               
               
                 N2-acetyllysine 
                 0.78 
                 0.1297 
                 
                   0.77 
                 
                 0.0374 
                 1.01 
                 0.933 
                 
                   0.78 
                 
                 0.0342 
                 36751 
               
               
                 o-cresol sulfate 
                 1.04 
                 0.9447 
                 1.76 
                 0.1645 
                 0.59 
                 0.2997 
                 
                   0.78 
                 
                 0.0345 
                 36845 
               
               
                 1-methylxanthine 
                 0.9 
                 0.1232 
                 1.28 
                 0.2826 
                 0.7 
                 0.5172 
                 
                   1.13 
                 
                 0.0355 
                 34389 
               
               
                 pyroglutamylglutamine 
                 
                   0.66 
                 
                 0.0252 
                 
                   0.85 
                 
                 0.0386 
                 0.78 
                 0.5586 
                 
                   0.84 
                 
                 0.0374 
                 22194 
               
               
                 trigonelline (N′- 
                 0.82 
                 0.3471 
                 1.14 
                 0.9231 
                 0.72 
                 0.3571 
                 
                   0.88 
                 
                 0.0389 
                 32401 
               
               
                 methylnicotinate) 
               
               
                 sarcosine (N- 
                 1.01 
                 0.2635 
                 
                   0.67 
                 
                 0.0321 
                 1.52 
                 0.6247 
                 
                   0.78 
                 
                 0.0391 
                 1516 
               
               
                 Methylglycine) 
               
               
                 5-oxoproline 
                 0.82 
                 0.1502 
                 0.88 
                 0.1331 
                 0.93 
                 0.7995 
                 
                   0.88 
                 
                 0.04 
                 1494 
               
               
                 alanylalanine 
                 
                   0.67 
                 
                 0.0015 
                 0.79 
                 0.1473 
                 
                   0.86 
                 
                 0.061 
                 
                   0.8 
                 
                 0.0424 
                 15129 
               
               
                 malate 
                 1.12 
                 0.1185 
                 
                   1.2 
                 
                 0.0849 
                 0.93 
                 0.8337 
                 
                   1.07 
                 
                 0.0424 
                 1303 
               
               
                 sulforaphane- 
                 0.95 
                 0.6503 
                 0.95 
                 0.557 
                 1 
                 1 
                 
                   0.52 
                 
                 0.043 
                 40451 
               
               
                 cysteine 
               
               
                 glycocholate 
                 0.93 
                 0.6745 
                 0.79 
                 0.1027 
                 1.17 
                 0.4452 
                 
                   0.72 
                 
                 0.0446 
                 18476 
               
               
                 aspartylaspartate 
                 
                   0.49 
                 
                 0.0243 
                 
                   0.64 
                 
                 0.0344 
                 0.77 
                 0.5722 
                 
                   0.7 
                 
                 0.0451 
                 40671 
               
               
                 uridine 
                 
                   1.82 
                 
                 0.0322 
                 1.2 
                 0.3079 
                 1.51 
                 0.2172 
                 
                   1.41 
                 
                 0.0451 
                 606 
               
               
                 putrescine 
                 
                   0.49 
                 
                 0.0437 
                 
                   1.64 
                 
                 0.0925 
                 0.3 
                 0.5128 
                 
                   1.25 
                 
                 0.0455 
                 1408 
               
               
                 5-acetylamino-6- 
                 
                   0.39 
                 
                 0.007 
                 0.93 
                 0.1302 
                 0.42 
                 0.1632 
                 
                   0.92 
                 
                 0.0458 
                 34401 
               
               
                 formylamino-3- 
               
               
                 methyluracil 
               
               
                 chiro-inositol 
                 0.13 
                 0.2427 
                 1.24 
                 0.3029 
                 
                   0.1 
                 
                 0.0751 
                 
                   0.93 
                 
                 0.0473 
                 37112 
               
               
                 homocitrate 
                 
                   0.85 
                 
                 0.0232 
                 0.98 
                 0.4061 
                 0.86 
                 0.1381 
                 
                   0.94 
                 
                 0.0481 
                 39601 
               
               
                 erythronate 
                 
                   0.74 
                 
                 0.0484 
                 0.9 
                 0.1185 
                 0.82 
                 0.4836 
                 
                   0.89 
                 
                 0.0488 
                 33477 
               
               
                 homovanillate 
                 1 
                 0.5864 
                 0.81 
                 0.4937 
                 1.24 
                 0.9884 
                 
                   0.69 
                 
                 0.0498 
                 38349 
               
               
                 sulfate 
               
               
                 sulforaphane-N- 
                 0.84 
                 0.4563 
                 
                   0.76 
                 
                 0.0923 
                 1.1 
                 0.6139 
                 
                   0.53 
                 
                 0.0503 
                 40468 
               
               
                 acetyl-cysteine 
               
               
                 3-sialyllactose 
                 
                   0.68 
                 
                 0.0088 
                 1.02 
                 0.7611 
                 0.66 
                 0.0301 
                 
                   0.93 
                 
                 0.0505 
                 40424 
               
               
                 isocitrate 
                 0.84 
                 0.3846 
                 1.1 
                 0.6156 
                 0.76 
                 0.6615 
                 
                   0.91 
                 
                 0.0513 
                 12110 
               
               
                 N-acetylalanine 
                 0.78 
                 0.3469 
                 0.87 
                 0.2196 
                 0.9 
                 0.9949 
                 
                   0.72 
                 
                 0.0539 
                 1585 
               
               
                 theobromine 
                 
                   0.52 
                 
                 0.0054 
                 
                   0.68 
                 
                 0.0876 
                 0.76 
                 0.1818 
                 
                   0.75 
                 
                 0.0546 
                 18392 
               
               
                 prolylglycine 
                 1.01 
                 0.5897 
                 0.84 
                 0.2269 
                 1.19 
                 0.7207 
                 
                   0.7 
                 
                 0.0552 
                 40703 
               
               
                 alanine 
                 0.99 
                 0.7335 
                 0.89 
                 0.1897 
                 1.12 
                 0.5417 
                 
                   0.86 
                 
                 0.0566 
                 1126 
               
               
                 vanillylmandelate 
                 0.76 
                 0.2496 
                 0.99 
                 0.9666 
                 0.77 
                 0.3097 
                 
                   0.85 
                 
                 0.0573 
                 1567 
               
               
                 (VMA) 
               
               
                 deoxycholate 
                 
                   0.6 
                 
                 0.0923 
                 0.85 
                 0.1919 
                 0.71 
                 0.5384 
                 
                   0.74 
                 
                 0.0578 
                 1114 
               
               
                 caffeine 
                 
                   0.85 
                 
                 0.0812 
                 0.63 
                 0.5035 
                 1.35 
                 0.2643 
                 
                   1.09 
                 
                 0.0589 
                 569 
               
               
                 3- 
                 1.4 
                 0.6154 
                 1.13 
                 0.9967 
                 1.24 
                 0.6463 
                 
                   0.99 
                 
                 0.0618 
                 36848 
               
               
                 ethylphenylsulfate 
               
               
                 2-aminoadipate 
                 
                   0.74 
                 
                 0.0552 
                 
                   0.6 
                 
                 0.0132 
                 1.23 
                 0.9985 
                 
                   0.84 
                 
                 0.066 
                 6146 
               
               
                 adenosine 3′,5′- 
                 
                   0.75 
                 
                 0.0118 
                 
                   0.82 
                 
                 0.0065 
                 0.91 
                 0.7057 
                 
                   0.96 
                 
                 0.0663 
                 2831 
               
               
                 cyclic 
               
               
                 monophosphate 
               
               
                 (cAMP) 
               
               
                 3- 
                 1.32 
                 0.6526 
                 
                   1.34 
                 
                 0.0741 
                 0.99 
                 0.3986 
                 
                   0.79 
                 
                 0.0669 
                 22110 
               
               
                 hydroxykynurenine 
               
               
                 N2- 
                 1.02 
                 0.785 
                 1.01 
                 0.5668 
                 1.02 
                 0.8779 
                 
                   1.19 
                 
                 0.0674 
                 35133 
               
               
                 methylguanosine 
               
               
                 homovanillate 
                 0.83 
                 0.4846 
                 
                   0.75 
                 
                 0.0956 
                 1.11 
                 0.5931 
                 
                   0.72 
                 
                 0.0682 
                 1101 
               
               
                 (HVA) 
               
               
                 N- 
                 0.99 
                 0.8679 
                 0.89 
                 0.3932 
                 1.11 
                 0.6546 
                 
                   0.84 
                 
                 0.0739 
                 33942 
               
               
                 acetylasparagine 
               
               
                 anthranilate 
                 
                   0.7 
                 
                 0.0526 
                 0.88 
                 0.4658 
                 0.8 
                 0.2109 
                 
                   0.73 
                 
                 0.0741 
                 4970 
               
               
                 kynurenate 
                 0.85 
                 0.2071 
                 0.95 
                 0.5016 
                 0.89 
                 0.4995 
                 
                   0.83 
                 
                 0.0749 
                 1417 
               
               
                 2,3-butanediol 
                 0.4 
                 0.1435 
                 
                   0.47 
                 
                 0.0988 
                 0.85 
                 0.8638 
                 
                   0.35 
                 
                 0.0762 
                 35691 
               
               
                 phosphoethanolamine 
                 
                   0.55 
                 
                 0.006 
                 1.02 
                 0.3123 
                 0.54 
                 0.0729 
                 
                   0.85 
                 
                 0.0763 
                 12102 
               
               
                 pyridoxine (Vitamin 
                 
                   0.43 
                 
                 0.0844 
                 
                   0.43 
                 
                 0.0255 
                 
                   1 
                 
                 1 
                 
                   0.77 
                 
                 0.0787 
                 608 
               
               
                 B6) 
               
               
                 3- 
                 0.88 
                 0.3112 
                 
                   0.77 
                 
                 0.0074 
                 1.14 
                 0.3349 
                 
                   0.88 
                 
                 0.08 
                 38667 
               
               
                 methylglutaconate 
               
               
                 arabinose 
                 
                   0.69 
                 
                 0.056 
                 0.94 
                 0.4466 
                 0.73 
                 0.2286 
                 
                   0.88 
                 
                 0.0813 
                 575 
               
               
                 indolelactate 
                 0.82 
                 0.2641 
                 
                   0.78 
                 
                 0.0483 
                 1.05 
                 0.71 
                 
                   0.81 
                 
                 0.0814 
                 18349 
               
               
                 pyroglutamylvaline 
                 1.03 
                 0.5813 
                 0.94 
                 0.3287 
                 1.1 
                 0.8541 
                 
                   0.9 
                 
                 0.0832 
                 32394 
               
               
                 1-(3-aminopropyl)- 
                 
                   1.55 
                 
                 0.0483 
                 
                   1.67 
                 
                 0.002 
                 0.93 
                 0.7071 
                 
                   1.23 
                 
                 0.0848 
                 40506 
               
               
                 2-pyrrolidone 
               
               
                 ascorbate (Vitamin 
                 
                   0.11 
                 
                 0.0501 
                 0.32 
                 0.1096 
                 0.35 
                 0.5094 
                 
                   0.32 
                 
                 0.0861 
                 1640 
               
               
                 C) 
               
               
                 glucose 
                 0.51 
                 0.4847 
                 0.39 
                 0.3482 
                 1.32 
                 0.9817 
                 
                   0.85 
                 
                 0.0864 
                 31263 
               
               
                 gamma- 
                 0.77 
                 0.1772 
                 
                   0.74 
                 
                 0.0381 
                 1.04 
                 0.8182 
                 
                   0.77 
                 
                 0.089 
                 2734 
               
               
                 glutamyltyrosine 
               
               
                 dehydroisoandrosterone 
                 1.32 
                 0.3166 
                 1.27 
                 0.7266 
                 1.04 
                 0.5064 
                 
                   0.77 
                 
                 0.0896 
                 32425 
               
               
                 sulfate 
               
               
                 (DHEA-S) 
               
               
                 caffeate 
                 0.79 
                 0.373 
                 0.86 
                 0.8299 
                 0.92 
                 0.5103 
                 
                   0.74 
                 
                 0.0902 
                 21177 
               
               
                 choline 
                 
                   1.26 
                 
                 0.0203 
                 
                   1.51 
                 
                 0.0005 
                 0.84 
                 0.7221 
                 
                   1.09 
                 
                 0.0911 
                 15506 
               
               
                 sucralose 
                 
                   0.22 
                 
                 0.019 
                 
                   0.52 
                 
                 0.065 
                 0.43 
                 0.4004 
                 
                   0.58 
                 
                 0.0915 
                 36649 
               
               
                 N-acetylserine 
                 
                   1.91 
                 
                 0.0022 
                 
                   1.31 
                 
                 0.0206 
                 1.45 
                 0.2446 
                 
                   1.16 
                 
                 0.0935 
                 37076 
               
               
                 arabitol 
                 0.83 
                 0.2061 
                 0.86 
                 0.1029 
                 0.96 
                 0.9963 
                 
                   0.9 
                 
                 0.097 
                 38075 
               
               
                 sulforaphane 
                 1.09 
                 0.5569 
                 0.76 
                 0.2705 
                 1.43 
                 0.1921 
                 
                   0.61 
                 
                 0.0971 
                 38697 
               
               
                 ribitol 
                 
                   0.7 
                 
                 0.0283 
                 
                   0.71 
                 
                 0.0015 
                 0.99 
                 0.8078 
                 0.89 
                 0.1277 
                 15772 
               
               
                 2,4,6- 
                 
                   0.36 
                 
                 0.0539 
                 
                   0.39 
                 
                 0.0006 
                 0.93 
                 0.5028 
                 0.86 
                 0.2894 
                 35892 
               
               
                 trihydroxybenzoate 
               
               
                 histidine 
                 
                   1.31 
                 
                 0.0899 
                 
                   1.37 
                 
                 0.0023 
                 0.95 
                 0.5434 
                 1.09 
                 0.3406 
                 59 
               
               
                   
               
            
           
         
       
     
     Example 4 
     Biomarkers for Monitoring Bladder Cancer 
     To identify biomarkers for monitoring bladder cancer, urine samples were collected from 119 subjects with a history of bladder cancer but no indication of bladder cancer at the time of urine collection (HX) and 66 bladder cancer subjects. Metabolomic analysis was performed. After the levels of metabolites were determined, the data were analyzed using one-way ANOVA contrasts to identify biomarkers that differed between patients with a history of bladder cancer and normal subjects. The biomarkers are listed in Table 5, columns 1, 8, 9. 
     The biomarkers in Table 5 were used to create a statistical model to classify the subjects into BCA or FIX groups. Random Forest analysis was used to classify subjects as having bladder cancer or a history of bladder cancer. 
     Random Forest results show that the samples were classified with 83% prediction accuracy. The Confusion Matrix presented in Table 6 shows the number of samples predicted for each classification and the actual in each group (BCA or HX). The “Out-of-Bag” (OOB) Error rate gives an estimate of how accurately new observations can be predicted using the Random Forest model (e.g., whether a sample is from a bladder cancer subject or a subject with a history of bladder cancer). The OOB error from this Random Forest was approximately 17%, and the model estimated that, when used on a new set of subjects, the identity of bladder cancer subjects could be predicted correctly 76% of the time and subjects with a history of bladder cancer could be predicted 87% of the time. 
     
       
         
           
               
             
               
                 TABLE 6 
               
             
            
               
                   
               
               
                 Results of Random Forest, Bladder Cancer 
               
               
                 vs. History of Bladder Cancer 
               
            
           
           
               
               
               
               
            
               
                   
                   
                 Predicted Group 
                 class. 
               
            
           
           
               
               
               
               
            
               
                   
                 BCA 
                 HX 
                 Error 
               
               
                   
                   
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Actual 
                 BCA 
                 50 
                 16 
                 0.242424 
               
               
                   
                 Group 
                 HX 
                 15 
                 104 
                 0.12605 
               
               
                   
                   
               
            
           
         
       
     
     Based on the OOB Error rate of 17%, the Random Forest model that was created predicted whether a sample was from an individual with bladder cancer with about 83% accuracy from analysis of the levels of the biomarkers measured in samples from the subject. Exemplary biomarkers for distinguishing the groups are 3-hydroxyphenylacetate, 3-hydroxyhippurate, 3-hydroxybutyrate (BHBA), isovalerylglycine, phenylacetylglutamine, pyridoxate, 2-5-furandicarboxylic acid, allantoin, pimelate (heptanedioate), lactate, adenosine 5′-monophosphate (AMP), catechol-sulfate, 2-hydroxybutyrate (AHB), isobutyrylglycine, 2-hydroxyhippurate (salicylurate), gluconate, imidazole-propionate, succinate, alpha-CEHC-glucoronide, 3-indoxyl-sulfate, 4-hydroxyphenylacetate, acetylcarnitine, xanthine, p-cresol-sulfate, tartarate, 4-hydroxyhippurate, 2-isopropylmalate, palmitoyl-sphingomyelin, adipate, and N(2)-furoyl-glycine. 
     The Random Forest results demonstrated that by using the biomarkers, BCA subjects were distinguished from HX subjects with a 76% sensitivity, 87% specificity, 77% PPV, and 87% NPV. 
     Example 5 
     Tissue Biomarkers for Bladder Cancer 
     Biomarkers were discovered by (1) analyzing tissue samples from different groups of human subjects to determine the levels of metabolites in the samples and then (2) statistically analyzing the results to determine those metabolites that are differentially present in the groups. 
     The samples used for the analysis were: 31 control (benign) samples and 98 bladder cancer (tumor). 
     After the levels of metabolites were determined, the data were analyzed using Welch&#39;s two sample t-tests. To identify biomarkers for bladder cancer, benign samples were compared to bladder cancer samples. As listed in Table 7 below, the analysis of named compounds resulted in the identification of biomarkers that are differentially present between bladder cancer and control tissue. 
     Table 7 includes, for each biomarker, the biochemical name of the biomarker, the fold change of the biomarker in bladder cancer compared to control samples (BCA/Control) which is the ratio of the mean level of the biomarker in bladder cancer samples as compared to the non-bladder cancer mean level, and the p-value determined in the statistical analysis of the data concerning the biomarkers. Columns 4-6 of Table 7 list the following: the internal identifier for that biomarker compound in the in-house chemical library of authentic standards (CompID); the identifier for that biomarker compound in the Kyoto Encyclopedia of Genes and Genomes (KEGG), if available; and the identifier for that biomarker compound in the Human Metabolome Database (HMDB), if available. 
     
       
         
           
               
             
               
                 TABLE 7 
               
             
            
               
                   
               
               
                 Tissue Biomarkers for Bladder Cancer 
               
            
           
           
               
               
               
            
               
                   
                 BCA/Control 
                   
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Fold of 
                   
                 Comp 
                   
                   
               
               
                 Biochemical Name 
                 Change 
                 p-value 
                 ID 
                 KEGG 
                 HMDB 
               
               
                   
               
            
           
           
               
               
               
               
               
               
            
               
                 3-hydroxybutyrate (BHBA) 
                 0.67 
                 0.0783 
                 542 
                 C01089 
                 HMDB00357 
               
               
                 tyramine 
                 17.53 
                 0.0163 
                 1603 
                 C00483 
                 HMDB00306 
               
               
                 acetylcarnitine 
                 0.81 
                 0.0008 
                 32198 
                 C02571 
                 HMDB00201 
               
               
                 gluconate 
                 0.26 
                 0.00E+00 
                 587 
                 C00257 
                 HMDB00625 
               
               
                 myo-inositol 
                 0.4 
                 1.66E−10 
                 19934 
                 C00137 
                 HMDB00211 
               
               
                 6-phosphogluconate 
                 0.26 
                 1.71E−09 
                 15449 
                 C00345 
                 HMDB01316 
               
               
                 glucose 
                 0.38 
                 7.51E−09 
                 20488 
                 C00031 
                 HMDB00122 
               
               
                 pro-hydroxy-pro 
                 2.48 
                 8.99E−09 
                 35127 
                   
                 HMDB06695 
               
               
                 5-methylthioadenosine (MTA) 
                 4.24 
                 2.45E−08 
                 1419 
                 C00170 
                 HMDB01173 
               
               
                 2-myristoylglycerophosphocholine 
                 3.14 
                 3.07E−08 
                 35681 
               
               
                 N2-methylguanosine 
                 2.15 
                 3.43E−08 
                 35133 
                   
                 HMDB05862 
               
               
                 6-keto prostaglandin F1alpha 
                 0.23 
                 4.09E−08 
                 20476 
                 C05961 
                 HMDB02886 
               
               
                 1-myristoylglycerophosphocholine 
                 3.92 
                 7.07E−08 
                 35626 
                   
                 HMDB10379 
               
               
                 scyllo-inositol 
                 0.32 
                 1.05E−07 
                 32379 
                 C06153 
                 HMDB06088 
               
               
                 docosadienoate (22:2n6) 
                 3.01 
                 1.20E−07 
                 32415 
                 C16533 
               
               
                 sphinganine 
                 4.41 
                 1.57E−07 
                 17769 
                 C00836 
                 HMDB00269 
               
               
                 erythronate 
                 2.53 
                 1.60E−07 
                 33477 
                   
                 HMDB00613 
               
               
                 stearoyl sphingomyelin 
                 0.34 
                 2.19E−07 
                 19503 
                 C00550 
                 HMDB01348 
               
               
                 alpha-glutamyllysine 
                 0.65 
                 2.37E−07 
                 40441 
                   
                 HMDB04207 
               
               
                 7-methylguanine 
                 2.25 
                 2.45E−07 
                 35114 
                 C02242 
                 HMDB00897 
               
               
                 eicosapentaenoate (EPA; 20:5n3) 
                 2.12 
                 3.10E−07 
                 18467 
                 C06428 
                 HMDB01999 
               
               
                 1-palmitoylglycerophosphoinositol 
                 3.35 
                 3.53E−07 
                 35305 
               
               
                 docosatrienoate (22:3n3) 
                 3.08 
                 4.19E−07 
                 32417 
                 C16534 
                 HMDB02823 
               
               
                 2-palmitoleoylglycerophosphocholine 
                 4.08 
                 4.58E−07 
                 35819 
               
               
                 valerylcarnitine 
                 3.1 
                 4.64E−07 
                 34406 
                   
                 HMDB13128 
               
               
                 N1-methylguanosine 
                 2.19 
                 5.89E−07 
                 31609 
                   
                 HMDB01563 
               
               
                 nonadecanoate (19:0) 
                 1.72 
                 6.28E−07 
                 1356 
                 C16535 
                 HMDB00772 
               
               
                 1-stearoylglycerophosphoinositol 
                 2.08 
                 6.47E−07 
                 19324 
               
               
                 gamma-glutamylglutamine 
                 0.59 
                 7.70E−07 
                 2730 
                   
                 HMDB11738 
               
               
                 17-methylstearate 
                 1.94 
                 7.88E−07 
                 38296 
               
               
                 5,6-dihydrouracil 
                 2.9 
                 1.01E−06 
                 1559 
                 C00429 
                 HMDB00076 
               
               
                 prostaglandin I2 
                 0.23 
                 1.13E−06 
                 32466 
                 C01312 
                 HMDB01335 
               
               
                 propionylcarnitine 
                 1.97 
                 1.15E−06 
                 32452 
                 C03017 
                 HMDB00824 
               
               
                 pseudouridine 
                 1.92 
                 1.18E−06 
                 33442 
                 C02067 
                 HMDB00767 
               
               
                 dihomo-linoleate (20:2n6) 
                 2.23 
                 1.31E−06 
                 17805 
                 C16525 
               
               
                 N2,N2-dimethylguanosine 
                 2.28 
                 1.31E−06 
                 35137 
                   
                 HMDB04824 
               
               
                 gamma-glutamylglutamate 
                 0.43 
                 1.42E−06 
                 36738 
               
               
                 1-linoleoylglycerol (1-monolinolein) 
                 2.95 
                 1.75E−06 
                 27447 
               
               
                 eicosenoate (20:1n9 or 11) 
                 2.12 
                 1.81E−06 
                 33587 
                   
                 HMDB02231 
               
               
                 5,6-dihydrothymine 
                 1.78 
                 2.13E−06 
                 1418 
                 C00906 
                 HMDB00079 
               
               
                 adrenate (22:4n6) 
                 2.03 
                 2.15E−06 
                 32980 
                 C16527 
                 HMDB02226 
               
               
                 2-palmitoleoylglycerophosphoethanolamine 
                 3.92 
                 2.21E−06 
                 34871 
               
               
                 1-eicosadienoylglycerophosphocholine 
                 2.57 
                 2.28E−06 
                 33871 
               
               
                 palmitoleate (16:1n7) 
                 1.81 
                 2.49E−06 
                 33447 
                 C08362 
                 HMDB03229 
               
               
                 cytidine 5′-diphosphocholine 
                 3.36 
                 2.95E−06 
                 34418 
               
               
                 myristate (14:0) 
                 1.36 
                 3.08E−06 
                 1365 
                 C06424 
                 HMDB00806 
               
               
                 dihydrobiopterin 
                 1.86 
                 3.17E−06 
                 35129 
                 C02953, 
                 HMDB00038 
               
               
                   
                   
                   
                   
                 C00268 
               
               
                 docosapentaenoate (n3 DPA; 22:5n3) 
                 2.06 
                 3.20E−06 
                 32504 
                 C16513 
                 HMDB01976 
               
               
                 2-palmitoylglycerol (2-monopalmitin) 
                 1.96 
                 3.25E−06 
                 33419 
               
               
                 2-oleoylglycerophosphocholine 
                 3.99 
                 3.61E−06 
                 35254 
               
               
                 cholate 
                 2.23 
                 3.65E−06 
                 22842 
                 C00695 
                 HMDB00619 
               
               
                 N-acetylneuraminate 
                 2.93 
                 4.39E−06 
                 1592 
                 C00270 
                 HMDB00230 
               
               
                 2-linoleoylglycerol (2-monolinolein) 
                 2.52 
                 4.91E−06 
                 32506 
                   
                 HMDB11538 
               
               
                 3-phosphoglycerate 
                 0.31 
                 5.03E−06 
                 40264 
                 C00597 
                 HMDB00807 
               
               
                 dihomo-linolenate (20:3n3 or n6) 
                 2.04 
                 5.74E−06 
                 35718 
                 C03242 
                 HMDB02925 
               
               
                 margarate (17:0) 
                 1.66 
                 5.95E−06 
                 1121 
                   
                 HMDB02259 
               
               
                 1-oleoylglycerophosphocholine 
                 3.88 
                 6.03E−06 
                 33960 
               
               
                 1-oleoylglycerophosphoethanolamine 
                 2.04 
                 6.09E−06 
                 35628 
                   
                 HMDB11506 
               
               
                 1-heptadecanoylglycerophosphocholine 
                 3.3 
                 6.24E−06 
                 33957 
                   
                 HMDB12108 
               
               
                 2-phosphoglycerate 
                 0.27 
                 6.54E−06 
                 35629 
                 C00631 
                 HMDB03391 
               
               
                 N1-methyladenosine 
                 1.88 
                 7.19E−06 
                 15650 
                 C02494 
                 HMDB03331 
               
               
                 1-methylimidazoleacetate 
                 0.46 
                 7.66E−06 
                 32350 
                 C05828 
                 HMDB02820 
               
               
                 deoxycarnitine 
                 1.74 
                 7.90E−06 
                 36747 
                 C01181 
                 HMDB01161 
               
               
                 1-palmitoylplasmenylethanolamine 
                 2.09 
                 8.13E−06 
                 39270 
               
               
                 docosapentaenoate (n6 DPA; 22:5n6) 
                 2.28 
                 8.28E−06 
                 37478 
                 C06429 
                 HMDB13123 
               
               
                 phytosphingosine 
                 4.05 
                 9.57E−06 
                 1510 
                 C12144 
                 HMDB04610 
               
               
                 3-phosphoserine 
                 0.27 
                 1.00E−05 
                 543 
                 C01005 
                 HMDB00272 
               
               
                 oleic ethanolamide 
                 2.77 
                 1.05E−05 
                 38102 
                   
                 HMDB02088 
               
               
                 1-linoleoylglycerophosphoethanolamine 
                 1.94 
                 1.08E−05 
                 32635 
                   
                 HMDB11507 
               
               
                 gamma-glutamylmethionine 
                 0.67 
                 1.15E−05 
                 37539 
               
               
                 N-acetylgalactosamine 
                 4.29 
                 1.16E−05 
                 2766 
                 C01074 
                 HMDB00835 
               
               
                 1-oleoylglycerophosphoserine 
                 1.94 
                 1.23E−05 
                 19260 
               
               
                 docosahexaenoate (DHA; 22:6n3) 
                 1.83 
                 1.23E−05 
                 19323 
                 C06429 
                 HMDB02183 
               
               
                 1-palmitoylglycerol (1-monopalmitin) 
                 1.87 
                 1.33E−05 
                 21127 
               
               
                 glucosamine 
                 4.42 
                 1.60E−05 
                 18534 
                 C00329 
                 HMDB01514 
               
               
                 cis-vaccenate (18:1n7) 
                 1.77 
                 1.62E−05 
                 33970 
                 C08367 
               
               
                 gamma-glutamylalanine 
                 0.59 
                 1.66E−05 
                 37063 
               
               
                 10-nonadecenoate (19:1n9) 
                 1.75 
                 2.06E−05 
                 33972 
               
               
                 4-hydroxyhippurate 
                 5.02 
                 2.13E−05 
                 35527 
               
               
                 4-hydroxyphenylpyruvate 
                 2.5 
                 2.25E−05 
                 1669 
                 C01179 
                 HMDB00707 
               
               
                 1-linoleoylglycerophosphocholine 
                 3.2 
                 2.37E−05 
                 34419 
                 C04100 
               
               
                 N-acetylthreonine 
                 1.53 
                 2.60E−05 
                 33939 
                 C01118 
               
               
                 VGAHAGEYGAEALER (SEQ ID NO: 2) 
                 0.39 
                 2.61E−05 
                 41219 
               
               
                 prostaglandin D2 
                 0.4 
                 2.81E−05 
                 7737 
                 C00696 
                 HMDB01403 
               
               
                 sphingosine 
                 3.41 
                 2.89E−05 
                 17747 
                 C00319 
                 HMDB00252 
               
               
                 quinolinate 
                 3.99 
                 3.12E−05 
                 1899 
                 C03722 
                 HMDB00232 
               
               
                 N-acetylglucosamine 
                 3.45 
                 3.87E−05 
                 15096 
                 C00140 
                 HMDB00215 
               
               
                 arachidate (20:0) 
                 1.83 
                 4.04E−05 
                 1118 
                 C06425 
                 HMDB02212 
               
               
                 1-oleoylglycerol (1-monoolein) 
                 1.94 
                 4.11E−05 
                 21184 
                   
                 HMDB11567 
               
               
                 trans-4-hydroxyproline 
                 2.12 
                 4.14E−05 
                 1366 
                 C01157 
                 HMDB00725 
               
               
                 inosine 
                 0.75 
                 4.40E−05 
                 1123 
               
               
                 coenzyme A 
                 3.07 
                 4.87E−05 
                 2936 
                 C00010 
                 HMDB01423 
               
               
                 3-indoxyl sulfate 
                 4.93 
                 5.08E−05 
                 27672 
                   
                 HMDB00682 
               
               
                 13-HODE + 9-HODE 
                 0.51 
                 5.40E−05 
                 37752 
               
               
                 10-heptadecenoate (17:1n7) 
                 1.69 
                 5.68E−05 
                 33971 
               
               
                 erythritol 
                 2.09 
                 5.86E−05 
                 20699 
                 C00503 
                 HMDB02994 
               
               
                 2′-deoxyinosine 
                 1.88 
                 8.05E−05 
                 15076 
                 C05512 
                 HMDB00071 
               
               
                 lignocerate (24:0) 
                 2.49 
                 8.07E−05 
                 1364 
                 C08320 
                 HMDB02003 
               
               
                 isoleucylproline 
                 1.53 
                 8.22E−05 
                 35418 
                   
                 HMDB11174 
               
               
                 methyl-alpha-glucopyranoside 
                 4.01 
                 8.44E−05 
                 20714 
                 C04942, 
               
               
                   
                   
                   
                   
                 C02603 
               
               
                 2-linoleoylglycerophosphocholine 
                 2.59 
                 8.87E−05 
                 35257 
               
               
                 creatine phosphate 
                 0.52 
                 9.07E−05 
                 33951 
                 C02305 
                 HMDB01511 
               
               
                 methionylvaline 
                 1.77 
                 9.41E−05 
                 40677 
               
               
                 hexadecanedioate 
                 0.53 
                 9.61E−05 
                 35678 
                   
                 HMDB00672 
               
               
                 guanosine 3′-monophosphate (3′-GMP) 
                 2.82 
                 9.95E−05 
                 39786 
               
               
                 1-palmitoleoylglycerophosphocholine 
                 2 
                 0.0001 
                 33230 
               
               
                 2-eicosatrienoylglycerophosphocholine 
                 2.69 
                 0.0001 
                 35884 
               
               
                 2-palmitoylglycerophosphocholine 
                 2.63 
                 0.0001 
                 35253 
               
               
                 Ac-Ser-Asp-Lys-Pro-OH (SEQ ID NO: 1) 
                 2.04 
                 0.0001 
                 40707 
               
               
                 ergothioneine 
                 1.78 
                 0.0001 
                 37459 
                 C05570 
                 HMDB03045 
               
               
                 nicotinamide ribonucleotide (NMN) 
                 0.29 
                 0.0001 
                 22152 
                 C00455 
                 HMDB00229 
               
               
                 octadecanedioate 
                 0.7 
                 0.0001 
                 36754 
                   
                 HMDB00782 
               
               
                 phenol sulfate 
                 3.45 
                 0.0001 
                 32553 
                 C02180 
               
               
                 1-palmitoylglycerophosphoethanolamine 
                 1.75 
                 0.0002 
                 35631 
                   
                 HMDB11503 
               
               
                 2′-deoxyguanosine 
                 1.6 
                 0.0002 
                 1411 
                 C00330 
                 HMDB00085 
               
               
                 4-hydroxyphenylacetate 
                 3.14 
                 0.0002 
                 541 
                 C00642 
                 HMDB00020 
               
               
                 adenosine 3′-monophosphate (3′-AMP) 
                 2.38 
                 0.0002 
                 35142 
                 C01367 
                 HMDB03540 
               
               
                 arachidonate (20:4n6) 
                 1.46 
                 0.0002 
                 1110 
                 C00219 
                 HMDB01043 
               
               
                 fucose 
                 2.32 
                 0.0002 
                 15821 
                 C00382 
                 HMDB00174 
               
               
                 glycyltyrosine 
                 0.63 
                 0.0002 
                 33958 
               
               
                 mannose 
                 0.81 
                 0.0002 
                 584 
                 C00159 
                 HMDB00169 
               
               
                 myristoleate (14:1n5) 
                 1.36 
                 0.0002 
                 32418 
                 C08322 
                 HMDB02000 
               
               
                 N-acetylglutamate 
                 1.91 
                 0.0002 
                 15720 
                 C00624 
                 HMDB01138 
               
               
                 phosphoenolpyruvate (PEP) 
                 0.26 
                 0.0002 
                 597 
                 C00074 
                 HMDB00263 
               
               
                 stearate (18:0) 
                 1.24 
                 0.0002 
                 1358 
                 C01530 
                 HMDB00827 
               
               
                 tetrahydrocortisone 
                 2.5 
                 0.0002 
                 38608 
                 HMDB00903 
                 HMDB00903 
               
               
                 UDP-glucuronate 
                 3.16 
                 0.0002 
                 2763 
                 C00167 
                 HMDB00935 
               
               
                 vanillylmandelate (VMA) 
                 2.76 
                 0.0002 
                 1567 
                 C05584 
                 HMDB00291 
               
               
                 15-methylpalmitate (isobar with 2- 
                 1.43 
                 0.0003 
                 38768 
               
               
                 methylpalmitate) 
               
               
                 3′-dephosphocoenzyme A 
                 2.65 
                 0.0003 
                 18289 
                 C00882 
                 HMDB01373 
               
               
                 glycerophosphoethanolamine 
                 3.53 
                 0.0003 
                 37455 
                 C01233 
                 HMDB00114 
               
               
                 1-pentadecanoylglycerophosphocholine 
                 2.17 
                 0.0004 
                 37418 
               
               
                 1-stearoylglycerol (1-monostearin) 
                 1.52 
                 0.0004 
                 21188 
                 D01947 
               
               
                 4-acetamidobutanoate 
                 1.98 
                 0.0004 
                 1558 
                 C02946 
                 HMDB03681 
               
               
                 galactose 
                 2.65 
                 0.0004 
                 12055 
                 C01582 
                 HMDB00143 
               
               
                 phenylpyruvate 
                 3 
                 0.0004 
                 566 
                 C00166 
                 HMDB00205 
               
               
                 stearoyl ethanolamide 
                 3.74 
                 0.0004 
                 38625 
               
               
                 uridine 
                 0.84 
                 0.0004 
                 606 
                 C00299 
                 HMDB00296 
               
               
                 1-arachidonoylglycerophosphocholine 
                 2.44 
                 0.0005 
                 33228 
                 C05208 
               
               
                 4-guanidinobutanoate 
                 2.02 
                 0.0005 
                 15681 
                 C01035 
                 HMDB03464 
               
               
                 1-arachidonoylglycerophosphoinositol 
                 1.59 
                 0.0006 
                 34214 
               
               
                 2-linoleoylglycerophosphoethanolamine 
                 2.16 
                 0.0006 
                 34666 
               
               
                 3-methoxytyrosine 
                 1.45 
                 0.0006 
                 12017 
                   
                 HMDB01434 
               
               
                 1-stearoylglycerophosphocholine 
                 2.68 
                 0.0007 
                 33961 
               
               
                 aspartylvaline 
                 1.68 
                 0.0007 
                 41373 
               
               
                 stearoylcarnitine 
                 2.32 
                 0.0007 
                 34409 
                   
                 HMDB00848 
               
               
                 5-oxoproline 
                 0.64 
                 0.0008 
                 1494 
                 C01879 
                 HMDB00267 
               
               
                 2-arachidonoylglycerophosphocholine 
                 2.49 
                 0.0009 
                 35256 
               
               
                 beta-alanine 
                 1.81 
                 0.0009 
                 55 
                 C00099 
                 HMDB00056 
               
               
                 alanylisoleucine 
                 1.65 
                 0.001 
                 37118 
               
               
                 cyclo(leu-gly) 
                 0.56 
                 0.001 
                 37078 
               
               
                 guanosine 
                 0.76 
                 0.001 
                 1573 
                 C00387 
                 HMDB00133 
               
               
                 putrescine 
                 1.46 
                 0.001 
                 1408 
                 C00134 
                 HMDB01414 
               
               
                 alpha-hydroxyisocaproate 
                 2.6 
                 0.0011 
                 22132 
                 C03264 
                 HMDB00746 
               
               
                 behenate (22:0) 
                 1.86 
                 0.0011 
                 12125 
                 C08281 
                 HMDB00944 
               
               
                 dimethylarginine (SDMA + ADMA) 
                 1.41 
                 0.0012 
                 36808 
                 C03626 
                 HMDB01539, 
               
               
                   
                   
                   
                   
                   
                 HMDB03334 
               
               
                 glycylglycine 
                 1.6 
                 0.0012 
                 21029 
                 C02037 
                 HMDB11733 
               
               
                 methylphosphate 
                 1.88 
                 0.0013 
                 37070 
               
               
                 pregnanediol-3-glucuronide 
                 4.54 
                 0.0013 
                 40708 
               
               
                 anthranilate 
                 1.59 
                 0.0014 
                 4970 
                 C00108 
                 HMDB01123 
               
               
                 aspartate-glutamate 
                 1.59 
                 0.0014 
                 37461 
               
               
                 ribitol 
                 1.82 
                 0.0014 
                 15772 
                 C00474 
                 HMDB00508 
               
               
                 1-palmitoylglycerophosphocholine 
                 2.26 
                 0.0015 
                 33955 
               
               
                 riboflavin (Vitamin B2) 
                 1.55 
                 0.0015 
                 1827 
                 C00255 
                 HMDB00244 
               
               
                 cysteinylglycine 
                 0.59 
                 0.0016 
                 35637 
                 C01419 
                 HMDB00078 
               
               
                 glycerol 2-phosphate 
                 2.02 
                 0.0017 
                 27728 
                 C02979, 
                 HMDB02520 
               
               
                   
                   
                   
                   
                 D01488 
               
               
                 phenylacetylglutamine 
                 3.69 
                 0.0017 
                 35126 
                 C05597 
                 HMDB06344 
               
               
                 2-arachidonoylglycerophosphoinositol 
                 1.7 
                 0.0018 
                 38077 
               
               
                 2-hydroxypalmitate 
                 1.77 
                 0.0018 
                 35675 
               
               
                 N-acetylmannosamine 
                 1.98 
                 0.0018 
                 15060 
                 C00140 
                 HMDB00835 
               
               
                 caprate (10:0) 
                 1.18 
                 0.0019 
                 1642 
                 C01571 
                 HMDB00511 
               
               
                 histidylleucine 
                 0.58 
                 0.002 
                 40061 
               
               
                 ornithine 
                 1.58 
                 0.002 
                 1493 
                 C00077 
                 HMDB03374 
               
               
                 phenylalanylserine 
                 1.56 
                 0.002 
                 40016 
               
               
                 tetradecanedioate 
                 0.59 
                 0.002 
                 35669 
                   
                 HMDB00872 
               
               
                 2-methylcitrate 
                 2.41 
                 0.0022 
                 37483 
                 C02225 
                 HMDB00379 
               
               
                 ethanolamine 
                 1.91 
                 0.0022 
                 1497 
                 C00189 
                 HMDB00149 
               
               
                 valylisoleucine 
                 1.52 
                 0.0022 
                 40050 
               
               
                 1-stearoylglycerophosphoethanolamine 
                 1.47 
                 0.0023 
                 34416 
                   
                 HMDB11130 
               
               
                 hydroxyisovaleroyl carnitine 
                 1.69 
                 0.0024 
                 35433 
               
               
                 uridine-2′,3′-cyclic monophosphate 
                 1.44 
                 0.0024 
                 37137 
                 C02355 
                 HMDB11640 
               
               
                 2-oleoylglycerophosphoserine 
                 1.8 
                 0.0025 
                 37948 
               
               
                 glycylisoleucine 
                 1.62 
                 0.0025 
                 36659 
               
               
                 2-methylbutyroylcarnitine 
                 2.06 
                 0.0026 
                 35431 
                   
                 HMDB00378 
               
               
                 5-HETE 
                 2.06 
                 0.0028 
                 37372 
               
               
                 alanylproline 
                 1.1 
                 0.0029 
                 37083 
               
               
                 valylalanine 
                 1.51 
                 0.0029 
                 41518 
               
               
                 N-acetylglucosamine 6-phosphate 
                 1.82 
                 0.003 
                 15107 
                 C00357 
                 HMDB02817 
               
               
                 1-methylurate 
                 2.67 
                 0.0032 
                 34395 
                   
                 HMDB03099 
               
               
                 2-oleoylglycerophosphoethanolamine 
                 2.28 
                 0.0032 
                 35683 
               
               
                 serylphenyalanine 
                 1.53 
                 0.0033 
                 40054 
               
               
                 3-aminoisobutyrate 
                 2.6 
                 0.0035 
                 1566 
                 C05145 
                 HMDB03911 
               
               
                 S-lactoylglutathione 
                 2.41 
                 0.0035 
                 15731 
                 C03451 
                 HMDB01066 
               
               
                 5-methyltetrahydrofolate (5MeTHF) 
                 1.77 
                 0.0036 
                 18330 
                 C00440 
                 HMDB01396 
               
               
                 2-palmitoylglycerophosphoethanolamine 
                 1.74 
                 0.0037 
                 35684 
               
               
                 imidazole propionate 
                 2.85 
                 0.0039 
                 40730 
                   
                 HMDB02271 
               
               
                 uridine monophosphate (5′ or 3′) 
                 2.86 
                 0.0041 
                 39879 
               
               
                 cysteine 
                 0.82 
                 0.0042 
                 31453 
                 C00097 
                 HMDB00574 
               
               
                 glutamate, gamma-methyl ester 
                 1.99 
                 0.0042 
                 33487 
               
               
                 1-methylxanthine 
                 1.92 
                 0.0046 
                 34389 
               
               
                 alanylphenylalanine 
                 1.33 
                 0.0046 
                 38679 
               
               
                 enterolactone 
                 1.79 
                 0.0049 
                 39626 
               
               
                 hexanoylglycine 
                 1.41 
                 0.0049 
                 35436 
                   
                 HMDB00701 
               
               
                 cysteine sulfinic acid 
                 0.43 
                 0.0052 
                 37443 
                 C00606 
                 HMDB00996 
               
               
                 glutaroyl carnitine 
                 2.07 
                 0.0052 
                 35439 
                   
                 HMDB13130 
               
               
                 naringenin 
                 1.6 
                 0.0053 
                 21182 
                 C00509 
                 HMDB02670 
               
               
                 inositol 1-phosphate (I1P) 
                 0.76 
                 0.0057 
                 1481 
                   
                 HMDB00213 
               
               
                 threonylphenylalanine 
                 1.31 
                 0.0058 
                 31530 
               
               
                 pyroglutamylvaline 
                 1.59 
                 0.006 
                 32394 
               
               
                 linoleate (18:2n6) 
                 1.29 
                 0.0061 
                 1105 
                 C01595 
                 HMDB00673 
               
               
                 pelargonate (9:0) 
                 1.16 
                 0.0062 
                 12035 
                 C01601 
                 HMDB00847 
               
               
                 valylglycine 
                 0.98 
                 0.0062 
                 40475 
               
               
                 palmitoylcarnitine 
                 1.99 
                 0.0064 
                 22189 
               
               
                 alanylmethionine 
                 1.36 
                 0.0067 
                 37065 
               
               
                 valylleucine 
                 1.66 
                 0.0069 
                 39994 
               
               
                 glucuronate 
                 2.29 
                 0.0073 
                 15443 
                 C00191 
                 HMDB00127 
               
               
                 threitol 
                 1.95 
                 0.0081 
                 35854 
                 C16884 
                 HMDB04136 
               
               
                 S-adenosylhomocysteine (SAH) 
                 1.69 
                 0.0092 
                 15948 
                 C00021 
                 HMDB00939 
               
               
                 xanthosine 
                 1.55 
                 0.0093 
                 15136 
                 C01762 
                 HMDB00299 
               
               
                 13,14-dihydroprostaglandin E1 
                 1.64 
                 0.0095 
                 19450 
                   
                 HMDB02689 
               
               
                 glycerol 3-phosphate (G3P) 
                 0.54 
                 0.0097 
                 15365 
                 C00093 
                 HMDB00126 
               
               
                 triethanolamine 
                 0.2 
                 0.0099 
                 22202 
                 C06771 
               
               
                 gamma-glutamyltyrosine 
                 0.8 
                 0.0101 
                 2734 
               
               
                 leucylleucine 
                 1.39 
                 0.0106 
                 36756 
                 C11332 
               
               
                 isoleucylglycine 
                 0.71 
                 0.0107 
                 40008 
               
               
                 pentadecanoate (15:0) 
                 1.26 
                 0.011 
                 1361 
                 C16537 
                 HMDB00826 
               
               
                 xylose 
                 1.94 
                 0.0111 
                 15835 
                 C00181 
                 HMDB00098 
               
               
                 xylitol 
                 1.76 
                 0.0112 
                 4966 
                 C00379 
                 HMDB00568 
               
               
                 guanidinoacetate 
                 2.31 
                 0.0113 
                 1480 
                 C00581 
                 HMDB00128 
               
               
                 lathosterol 
                 1.23 
                 0.0115 
                 39864 
                 C01189 
                 HMDB01170 
               
               
                 pinitol 
                 1.66 
                 0.0116 
                 37086 
                 C03844 
               
               
                 alanylleucine 
                 1.29 
                 0.0117 
                 37093 
               
               
                 aspartylleucine 
                 1.4 
                 0.0126 
                 40068 
               
               
                 3-hydroxysebacate 
                 2.34 
                 0.0127 
                 31943 
                   
                 HMDB00350 
               
               
                 cytidine-5′-diphosphoethanolamine 
                 1.84 
                 0.0138 
                 34410 
                 C00570 
                 HMDB01564 
               
               
                 cytidine-3′-monophosphate (3′-CMP) 
                 1.65 
                 0.014 
                 2959 
                 C05822 
               
               
                 chiro-inositol 
                 0.59 
                 0.0149 
                 37112 
               
               
                 2-stearoylglycerophosphocholine 
                 2.09 
                 0.015 
                 35255 
               
               
                 aspartyltryptophan 
                 1.23 
                 0.015 
                 41481 
               
               
                 valylvaline 
                 1.76 
                 0.0154 
                 40728 
               
               
                 linolenate [alpha or gamma; (18:3n3 or 6)] 
                 1.33 
                 0.0159 
                 34035 
                 C06427 
                 HMDB01388 
               
               
                 stachydrine 
                 1.61 
                 0.016 
                 34384 
                 C10172 
                 HMDB04827 
               
               
                 stearidonate (18:4n3) 
                 1.73 
                 0.0165 
                 33969 
                 C16300 
                 HMDB06547 
               
               
                 ribose 
                 2.2 
                 0.0166 
                 12080 
                 C00121 
                 HMDB00283 
               
               
                 adenosine 2′-monophosphate (2′-AMP) 
                 1.96 
                 0.0168 
                 36815 
                 C00946 
                 HMDB11617 
               
               
                 isoleucylglutamine 
                 1.27 
                 0.0187 
                 40019 
               
               
                 valylaspartate 
                 1.41 
                 0.0188 
                 40650 
               
               
                 glutathione, oxidized (GSSG) 
                 1.94 
                 0.0189 
                 21121 
                 C00127 
                 HMDB03337 
               
               
                 glycerol 
                 1.37 
                 0.0197 
                 15122 
                 C00116 
                 HMDB00131 
               
               
                 1,6-anhydroglucose 
                 1.89 
                 0.0198 
                 21049 
                   
                 HMDB00640 
               
               
                 galactosylsphingosine 
                 1.36 
                 0.0203 
                 40083 
                   
                 HMDB00648 
               
               
                 tyrosylglutamine 
                 1.57 
                 0.0205 
                 41459 
               
               
                 phenethylamine (isobar with 1- 
                 3.19 
                 0.021 
                 38763 
                 C02455, 
                 HMDB02017, 
               
               
                 phenylethanamine) 
                   
                   
                   
                 C05332 
                 HMDB12275 
               
               
                 bilirubin (Z,Z) 
                 0.7 
                 0.0212 
                 27716 
                 C00486 
                 HMDB00054 
               
               
                 fructose 
                 2.9 
                 0.0218 
                 577 
                 C00095 
                 HMDB00660 
               
               
                 prolylproline 
                 1.16 
                 0.0218 
                 40731 
               
               
                 lactate 
                 1.23 
                 0.0221 
                 527 
                 C00186 
                 HMDB00190 
               
               
                 leucylalanine 
                 1.41 
                 0.0232 
                 40010 
               
               
                 7-methylxanthine 
                 1.42 
                 0.0235 
                 34390 
                 C16353 
                 HMDB01991 
               
               
                 isoleucylphenylalanine 
                 1.33 
                 0.0237 
                 40067 
               
               
                 methionylthreonine 
                 0.52 
                 0.0237 
                 40679 
               
               
                 3-hydroxyhippurate 
                 4.71 
                 0.0238 
                 39600 
                   
                 HMDB06116 
               
               
                 glycylproline 
                 1.19 
                 0.0243 
                 22171 
                   
                 HMDB00721 
               
               
                 levulinate (4-oxovalerate) 
                 1.25 
                 0.0253 
                 22177 
                   
                 HMDB00720 
               
               
                 serylleucine 
                 1.32 
                 0.0263 
                 40066 
               
               
                 phenylalanylphenylalanine 
                 1.3 
                 0.0264 
                 38150 
               
               
                 aspartylphenylalanine 
                 1.24 
                 0.0302 
                 22175 
                   
                 HMDB00706 
               
               
                 flavin adenine dinucleotide (FAD) 
                 1.33 
                 0.0304 
                 2134 
                 C00016 
                 HMDB01248 
               
               
                 3-methyl-2-oxovalerate 
                 0.79 
                 0.0306 
                 15676 
                 C00671 
                 HMDB03736 
               
               
                 3-methylxanthine 
                 1.44 
                 0.0309 
                 32445 
                 C16357 
                 HMDB01886 
               
               
                 adenosine 5′-diphosphate (ADP) 
                 0.68 
                 0.0317 
                 3108 
                 C00008 
                 HMDB01341 
               
               
                 daidzein 
                 1.49 
                 0.0318 
                 32453 
                 C10208 
                 HMDB03312 
               
               
                 alanylalanine 
                 1.28 
                 0.0319 
                 15129 
                 C00993 
                 HMDB03459 
               
               
                 aspartylaspartate 
                 0.66 
                 0.0325 
                 40671 
               
               
                 5-methyluridine (ribothymidine) 
                 1.3 
                 0.0328 
                 35136 
                   
                 HMDB00884 
               
               
                 threonylleucine 
                 1.35 
                 0.0329 
                 40051 
               
               
                 oleoylcarnitine 
                 1.83 
                 0.0332 
                 35160 
                   
                 HMDB05065 
               
               
                 p-cresol sulfate 
                 1.75 
                 0.0339 
                 36103 
                 C01468 
               
               
                 C-glycosyltryptophan 
                 1.32 
                 0.0343 
                 32675 
               
               
                 N-acetylglycine 
                 0.86 
                 0.0369 
                 27710 
                   
                 HMDB00532 
               
               
                 8-iso-15-keto-prostaglandin E2 
                 2.08 
                 0.0373 
                 7758 
                 C04707 
                 HMDB02341 
               
               
                 phenylalanylleucine 
                 0.99 
                 0.0373 
                 40192 
               
               
                 N-acetylalanine 
                 0.86 
                 0.0398 
                 1585 
                 C02847 
                 HMDB00766 
               
               
                 orotate 
                 1.79 
                 0.0401 
                 1505 
                 C00295 
                 HMDB00226 
               
               
                 2-aminoadipate 
                 0.96 
                 0.0416 
                 6146 
                 C00956 
                 HMDB00510 
               
               
                 N-acetylputrescine 
                 1.37 
                 0.042 
                 37496 
                 C02714 
                 HMDB02064 
               
               
                 L-urobilin 
                 0.83 
                 0.0455 
                 40173 
                 C05793 
                 HMDB04159 
               
               
                 choline 
                 1.19 
                 0.0465 
                 15506 
               
               
                 21-hydroxypregnenolone disulfate 
                 3.98 
                 0.0466 
                 37173 
                 C05485 
                 HMDB04026 
               
               
                 N-methylhydantoin 
                 6.29 
                 0.0472 
                 40006 
                 C02565 
                 HMDB03646 
               
               
                 succinylcarnitine 
                 1.81 
                 0.0476 
                 37058 
               
               
                 tyrosylleucine 
                 1.06 
                 0.0499 
                 40031 
               
               
                 prolylglycine 
                 1.23 
                 0.0502 
                 40703 
               
               
                 pyroglutamine 
                 1.48 
                 0.051 
                 32672 
               
               
                 butyrylcarnitine 
                 1.41 
                 0.0533 
                 32412 
               
               
                 gamma-glutamylisoleucine 
                 1.22 
                 0.0552 
                 34456 
                   
                 HMDB11170 
               
               
                 bilirubin (E,E) 
                 0.73 
                 0.0563 
                 32586 
               
               
                 myristoylcarnitine 
                 1.45 
                 0.0575 
                 33952 
               
               
                 N-acetylmethionine 
                 1.36 
                 0.0575 
                 1589 
                 C02712 
                 HMDB11745 
               
               
                 2- 
                 1.42 
                 0.0589 
                 34875 
               
               
                 docosapentaenoylglycerophosphoethanola 
               
               
                 mine 
               
               
                 threonate 
                 1.35 
                 0.0589 
                 27738 
                 C01620 
                 HMDB00943 
               
               
                 N-acetylasparagine 
                 2.23 
                 0.0609 
                 33942 
                   
                 HMDB06028 
               
               
                 imidazole lactate 
                 1.61 
                 0.0675 
                 15716 
                 C05568 
                 HMDB02320 
               
               
                 isoleucylalanine 
                 1.23 
                 0.0685 
                 40046 
               
               
                 taurolithocholate 3-sulfate 
                 2.92 
                 0.0699 
                 36850 
                 C03642 
                 HMDB02580 
               
               
                 methionylleucine 
                 0.98 
                 0.0711 
                 40023 
               
               
                 tryptophan betaine 
                 1.59 
                 0.0731 
                 37097 
                 C09213 
               
               
                 2-docosahexaenoylglycerophosphocholine 
                 0.72 
                 0.0733 
                 35883 
               
               
                 guanosine 5′- monophosphate (5′-GMP) 
                 2.19 
                 0.0734 
                 2849 
               
               
                 maltotriose 
                 0.67 
                 0.0754 
                 27723 
                 C01835 
                 HMDB01262 
               
               
                 7,8-dihydroneopterin 
                 1.52 
                 0.0773 
                 15689 
                 C04895 
                 HMDB02275 
               
               
                 leucylglutamate 
                 1.21 
                 0.0775 
                 40021 
               
               
                 maltose 
                 0.82 
                 0.0775 
                 15806 
                 C00208 
                 HMDB00163 
               
               
                 allantoin 
                 2.4 
                 0.0794 
                 1107 
                 C02350 
                 HMDB00462 
               
               
                 sorbitol 
                 2.06 
                 0.0805 
                 15053 
                 C00794 
                 HMDB00247 
               
               
                 alpha-hydroxyisovalerate 
                 1.24 
                 0.0814 
                 33937 
                   
                 HMDB00407 
               
               
                 valylhistidine 
                 1.14 
                 0.0835 
                 40680 
               
               
                 8-iso-prostaglandin F1 alpha 
                 1.02 
                 0.0845 
                 7820 
                 C06475 
                 HMDB02685 
               
               
                 2-docosahexaenoylglycerophosphoethanolam 
                 1.74 
                 0.086 
                 34258 
               
               
                 ine 
               
               
                 pro-pro-pro 
                 1.37 
                 0.0874 
                 40654 
               
               
                 glycylserine 
                 1.13 
                 0.0974 
                 33940 
                   
                 HMDB00678 
               
               
                 isoleucylglutamate 
                 1.08 
                 0.0986 
                 40057 
               
               
                 phosphopantetheine 
                 1.51 
                 0.0989 
                 15504 
                 C01134 
                 HMDB01416 
               
               
                 3-(4-hydroxyphenyl)lactate 
                 1.89 
                 1.10E−07 
                 32197 
                 C03672 
                 HMDB00755 
               
               
                 creatine 
                 0.49 
                 8.77E−07 
                 27718 
                 C00300 
                 HMDB00064 
               
               
                 thymine 
                 3.24 
                 1.41E−06 
                 604 
                 C00178 
                 HMDB00262 
               
               
                 phenyllactate (PLA) 
                 2.24 
                 2.50E−06 
                 22130 
                 C05607 
                 HMDB00779 
               
               
                 S-adenosylmethionine (SAM) 
                 3.4 
                 8.15E−06 
                 15915 
               
               
                 glycerophosphorylcholine (GPC) 
                 3.2 
                 2.01E−05 
                 15990 
                 C00670 
                 HMDB00086 
               
               
                 taurine 
                 0.7 
                 4.29E−05 
                 2125 
                 C00245 
                 HMDB00251 
               
               
                 uracil 
                 1.96 
                 4.68E−05 
                 605 
                 C00106 
                 HMDB00300 
               
               
                 succinate 
                 3.7 
                 4.75E−05 
                 1437 
                 C00042 
                 HMDB00254 
               
               
                 oleate (18:1n9) 
                 1.67 
                 6.45E−05 
                 1359 
                 C00712 
                 HMDB00207 
               
               
                 kynurenine 
                 2.11 
                 0.0004 
                 15140 
                 C00328 
                 HMDB00684 
               
               
                 palmitate (16:0) 
                 1.22 
                 0.0007 
                 1336 
                 C00249 
                 HMDB00220 
               
               
                 proline 
                 1.35 
                 0.0007 
                 1898 
                 C00148 
                 HMDB00162 
               
               
                 xanthine 
                 1.65 
                 0.0011 
                 3147 
                 C00385 
                 HMDB00292 
               
               
                 homocysteine 
                 1.67 
                 0.0019 
                 40266 
                 C00155 
                 HMDB00742 
               
               
                 homoserine 
                 2.25 
                 0.0025 
                 23642 
                 C00263, 
                 HMDB00719 
               
               
                   
                   
                   
                   
                 C02926 
               
               
                 betaine 
                 1.35 
                 0.0039 
                 3141 
                   
                 HMDB00043 
               
               
                 histamine 
                 0.78 
                 0.0062 
                 1574 
                 C00388 
                 HMDB00870 
               
               
                 methionine 
                 0.84 
                 0.0079 
                 1302 
                 C00073 
                 HMDB00696 
               
               
                 histidine 
                 1.23 
                 0.008 
                 59 
                 C00135 
                 HMDB00177 
               
               
                 pyridoxate 
                 3.37 
                 0.0098 
                 31555 
                 C00847 
                 HMDB00017 
               
               
                 kynurenate 
                 2.48 
                 0.0109 
                 1417 
                 C01717 
                 HMDB00715 
               
               
                 citrulline 
                 1.45 
                 0.011 
                 2132 
                 C00327 
                 HMDB00904 
               
               
                 tryptophan 
                 1.29 
                 0.0118 
                 54 
                 C00078 
                 HMDB00929 
               
               
                 alanine 
                 1.28 
                 0.0168 
                 1126 
                 C00041 
                 HMDB00161 
               
               
                 2-hydroxybutyrate (AHB) 
                 0.82 
                 0.0201 
                 21044 
                 C05984 
                 HMDB00008 
               
               
                 laurate (12:0) 
                 1.11 
                 0.025 
                 1645 
                 C02679 
                 HMDB00638 
               
               
                 cytidine 5′-monophosphate (5′-CMP) 
                 1.56 
                 0.0253 
                 2372 
                 C00055 
                 HMDB00095 
               
               
                 indolelactate 
                 1.64 
                 0.0255 
                 18349 
                 C02043 
                 HMDB00671 
               
               
                 caffeine 
                 0.66 
                 0.0386 
                 569 
                 C07481 
                 HMDB01847 
               
               
                 hippurate 
                 3.1 
                 0.0485 
                 15753 
                 C01586 
                 HMDB00714 
               
               
                 threonine 
                 1.16 
                 0.0528 
                 1284 
                 C00188 
                 HMDB00167 
               
               
                 adenosine 
                 0.7 
                 0.064 
                 555 
                 C00212 
                 HMDB00050 
               
               
                 dimethylglycine 
                 1.6 
                 0.0784 
                 5086 
                 C01026 
                 HMDB00092 
               
               
                 asparagine 
                 1.26 
                 0.0804 
                 11398 
                 C00152 
                 HMDB00168 
               
               
                 cortisol 
                 0.81 
                 0.0908 
                 1712 
                 C00735 
                 HMDB00063 
               
               
                 valine 
                 1.12 
                 0.0976 
                 1649 
                 C00183 
                 HMDB00883 
               
               
                   
               
            
           
         
       
     
     The biomarkers were used to create a statistical model to classify subjects. The biomarkers were evaluated using Random Forest analysis to classify samples as Bladder cancer or control. The Random Forest results show that the samples were classified with 84% prediction accuracy. The confusion matrix presented in Table 8 shows the number of samples predicted for each classification and the actual in each group (BCA or Control). The “Out-of-Bag” (OOB) Error rate gives an estimate of how accurately new observations can be predicted using the Random Forest model (e.g., whether a sample is a BCA or a control sample). The OOB error was approximately 15%, and the model estimated that, when used on a new set of subjects, the identity of Bladder cancer subjects could be predicted 87% of the time and control subjects could be predicted correctly 77% of the time and as presented in Table 8. 
     
       
         
           
               
             
               
                 TABLE 8 
               
             
            
               
                   
               
               
                 Results of Random Forest, Bladder cancer vs. Control 
               
            
           
           
               
               
               
               
            
               
                   
                   
                 Predicted Group 
                 class. 
               
            
           
           
               
               
               
               
            
               
                   
                 BCA 
                 Control 
                 Error 
               
               
                   
                   
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Actual 
                 BCA 
                 85 
                 13 
                 0.1327 
               
               
                   
                 Group 
                 Control 
                 7 
                 24 
                 0.2258 
               
               
                   
                   
               
            
           
         
       
     
     Based on the OOB Error rate of 16%, the Random Forest model that was created predicted whether a sample was from an individual with cancer with about 85% accuracy by measuring the levels of the biomarkers in samples from the subject. Exemplary biomarkers for distinguishing the groups are gluconate, 6-phosphogluconate, stearoyl sphingomyelin, myo-inositol, glucose, 3-(4-hydroxyphenyl)lactate (HPLA), 1-linoleoylglycerol (1-monolinolein), pro-hydroxy-pro, gamma-glutamylglutamate, creatine, 5,6-dihydrouracil, docosadienoate (22:2n6), phenyllactate (PLA), propionylcarnitine, isoleucylproline, N2-methylguanosine, eicosapentaenoate (EPA 20:5n3), 5-methylthioadenosine (MTA), alpha-glutamyllysine, 3-phosphoglycerate, 6-keto prostaglandin F1alpha, docosatrienoate (22:3n3), 2-palmitoleoylglycerophosphocholine, 1-stearoylglycerophosphoinositol, 1-palmitoylglycerophosphoinositol, scyllo-inositol, dihomo-linoleate (20:2n6), 3-phosphoserine, docosapentaenoate (n6 DPA 22:5n6), and 1-palmitoylglycerol (1-monopalmitin). 
     The Random Forest results demonstrated that by using the biomarkers, Bladder cancer samples were distinguished from control samples with 87% sensitivity, 77% specificity, 92% PPV, and 65% NPV. 
     Example 6 
     Tissue Biomarkers for Staging Bladder Cancer 
     Bladder cancer staging provides an indication of how far the bladder tumor has spread. The tumor stage is used to select treatment options and to estimate a patient&#39;s prognosis. Bladder tumor staging ranges from T0 (no evidence of primary tumor, least advanced) to T4 (tumor has spread beyond fatty tissue surrounding the bladder into nearby organs, most advanced). 
     To identify biomarkers of disease staging and/or progression, metabolomic analysis was carried out on tissue samples from 17 subjects with Low stage BCA (T0a, T1), 31 subjects with High stage BCA (T2-T4), and 44 Benign (Control) tissue samples. After the levels of metabolites were determined, the data were analyzed using Welch&#39;s two sample t-tests to identify biomarkers that differed between 1) Low stage bladder cancer compared to High stage bladder cancer, 2) Low stage bladder cancer compared to control, and 3) High stage bladder cancer compared to control. The biomarkers are listed in Table 9. 
     Table 9 includes, for each biomarker, the biochemical name of the biomarker, the fold change (FC) of the biomarker in 1) High stage bladder cancer compared to Low stage bladder cancer (T2-T4/Toa-T1), 2) Low stage bladder cancer compared to benign (T0a-T1/Benign) 3) High stage bladder cancer compared to benign (T2-T4/Benign) and the p-value determined in the statistical analysis of the data concerning the biomarkers. Columns 8-10 of Table 9 list the following: the internal identifier for that biomarker compound in the in-house chemical library of authentic standards (CompID); the identifier for that biomarker compound in the Kyoto Encyclopedia of Genes and Genomes (KEGG), if available; and the identifier for that biomarker compound in the Human Metabolome Database (HMDB), if available. Bold values indicate a fold change with a p-value of ≦0.1. 
     
       
         
           
               
             
               
                 TABLE 9 
               
             
            
               
                   
               
               
                 Tissue Biomarkers for Staging Bladder Cancer 
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 T2-T4 
                 T0a-T1 
                 T2-T4 
                   
                   
               
               
                   
                 T0a-T1 
                 Benign 
                 Benign 
                 Comp 
               
            
           
           
               
               
               
               
               
               
               
               
               
               
            
               
                 Biochemical Name 
                 FC 
                 p-value 
                 FC 
                 p-value 
                 FC 
                 p-value 
                 ID 
                 KEGG 
                 HMDB 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
               
            
               
                 bilirubin (Z,Z) 
                 
                   4.05 
                 
                 1.12E−06 
                 
                   0.25 
                 
                 1.86E−07 
                 0.87 
                 0.555 
                 27716 
                 C00486 
                 HMDB00054 
               
               
                 palmitoyl ethanolamide 
                 
                   7.99 
                 
                 6.85E−06 
                 0.67 
                 0.3724 
                 
                   2.76 
                 
                 0.0215 
                 38165 
               
               
                 adrenate (22:4n6) 
                 
                   2.35 
                 
                 1.39E−05 
                 
                   1.23 
                 
                 0.0561 
                 
                   2.34 
                 
                 1.87E−08 
                 32980 
                 C16527 
                 HMDB02226 
               
               
                 3-hydroxyoctanoate 
                 
                   1.89 
                 
                 1.57E−05 
                 0.92 
                 0.3237 
                 
                   1.34 
                 
                 0.0043 
                 22001 
                   
                 HMDB01954 
               
               
                 palmitoyl sphingomyelin 
                 
                   1.77 
                 
                 2.27E−05 
                 
                   0.74 
                 
                 0.0066 
                 1.09 
                 0.1949 
                 37506 
               
               
                 thromboxane B2 
                 
                   3 
                 
                 2.86E−05 
                 
                   0.65 
                 
                 0.0008 
                 
                   1.56 
                 
                 0.064 
                 17807 
                 C05963 
                 HMDB03252 
               
               
                 2-hydroxypalmitate 
                 
                   3.06 
                 
                 4.66E−05 
                 0.87 
                 0.6284 
                 
                   1.8 
                 
                 0.0004 
                 35675 
               
               
                 4-hydroxyphenylpyruvate 
                 
                   3.78 
                 
                 6.79E−05 
                 0.79 
                 0.6912 
                 
                   2.92 
                 
                 2.51E−06 
                 1669 
                 C01179 
                 HMDB00707 
               
               
                 5,6-dihydrothymine 
                 
                   2.06 
                 
                 8.90E−05 
                 1.14 
                 0.1697 
                 
                   1.89 
                 
                 2.98E−06 
                 1418 
                 C00906 
                 HMDB00079 
               
               
                 methyl-alpha- 
                 
                   0.2 
                 
                 9.37E−05 
                 
                   7.73 
                 
                 2.12E−06 
                 
                   1.96 
                 
                 0.0711 
                 20714 
                 C04942, 
               
               
                 glucopyranoside 
                   
                   
                   
                   
                   
                   
                   
                 C02603 
               
               
                 C-glycosyltryptophan 
                 
                   1.78 
                 
                 0.0001 
                 0.88 
                 0.3573 
                 
                   1.36 
                 
                 0.0041 
                 32675 
               
               
                 cytosine-2′,3′-cyclic 
                 
                   2.95 
                 
                 0.0002 
                 
                   0.46 
                 
                 0.014 
                 1.11 
                 0.1497 
                 37465 
                 C02354 
                 HMDB11691 
               
               
                 monophosphate 
               
               
                 laurylcarnitine 
                 
                   2.3 
                 
                 0.0003 
                 
                   0.71 
                 
                 0.0744 
                 1.17 
                 0.1886 
                 34534 
                   
                 HMDB02250 
               
               
                 pro-hydroxy-pro 
                 
                   2.12 
                 
                 0.0004 
                 
                   1 
                 
                 0.6377 
                 
                   1.96 
                 
                 1.42E−07 
                 35127 
                   
                 HMDB06695 
               
               
                 docosatrienoate (22:3n3) 
                 
                   3.43 
                 
                 0.0006 
                 
                   1.21 
                 
                 0.0146 
                 
                   3.6 
                 
                 2.21E−07 
                 32417 
                 C16534 
                 HMDB02823 
               
               
                 prostaglandin E1 
                 
                   6.73 
                 
                 0.0007 
                 
                   0.5 
                 
                 0.0446 
                 
                   2.71 
                 
                 0.0067 
                 19391 
                 C04741 
                 HMDB01442 
               
               
                 5,6-dihydrouracil 
                 
                   2.7 
                 
                 0.0007 
                 
                   1.41 
                 
                 0.0612 
                 
                   3.56 
                 
                 1.72E−09 
                 1559 
                 C00429 
                 HMDB00076 
               
               
                 N-acetylthreonine 
                 
                   1.58 
                 
                 0.0007 
                 1.1 
                 0.1125 
                 
                   1.6 
                 
                 2.45E−05 
                 33939 
                 C01118 
               
               
                 methylphosphate 
                 
                   0.51 
                 
                 0.0008 
                 
                   2.89 
                 
                 1.49E−05 
                 
                   1.56 
                 
                 0.0967 
                 37070 
               
               
                 quinolinate 
                 
                   3.27 
                 
                 0.0008 
                 1.44 
                 0.608 
                 
                   4.16 
                 
                 1.85E−07 
                 1899 
                 C03722 
                 HMDB00232 
               
               
                 phenylalanylserine 
                 
                   0.33 
                 
                 0.001 
                 
                   3.24 
                 
                 3.76E−06 
                 1 
                 0.1789 
                 40016 
               
               
                 alpha-tocopherol 
                 
                   1.97 
                 
                 0.001 
                 0.64 
                 0.2036 
                 
                   1.23 
                 
                 0.0156 
                 1561 
                 C02477 
                 HMDB01893 
               
               
                 3-hydroxydecanoate 
                 
                   1.72 
                 
                 0.0011 
                 0.88 
                 0.1209 
                 
                   1.49 
                 
                 0.0002 
                 22053 
                   
                 HMDB02203 
               
               
                 6-keto prostaglandin 
                 
                   7.39 
                 
                 0.0014 
                 
                   0.33 
                 
                 5.49E−08 
                 
                   0.31 
                 
                 0.0002 
                 20476 
                 C05961 
                 HMDB02886 
               
               
                 F1 alpha 
               
               
                 4-hydroxyhippurate 
                 
                   6.28 
                 
                 0.0014 
                 0.35 
                 0.4262 
                 
                   1.7 
                 
                 0.0084 
                 35527 
               
               
                 docosapentaenoate (n6 
                 
                   2.32 
                 
                 0.0016 
                 
                   1.4 
                 
                 0.0473 
                 
                   2.73 
                 
                 2.65E−08 
                 37478 
                 C06429 
                 HMDB13123 
               
               
                 DPA; 22:5n6) 
               
               
                 pyroglutamylvaline 
                 
                   2.08 
                 
                 0.0016 
                 0.92 
                 0.4171 
                 
                   1.69 
                 
                 0.0045 
                 32394 
               
               
                 bilirubin (E,E) 
                 
                   2.4 
                 
                 0.0018 
                 
                   0.47 
                 
                 0.0005 
                 0.91 
                 0.6474 
                 32586 
               
               
                 glutamate, gamma- 
                 
                   0.44 
                 
                 0.0019 
                 
                   2.59 
                 
                 0.0003 
                 1.18 
                 0.6414 
                 33487 
               
               
                 methyl ester 
               
               
                 docosadienoate (22:2n6) 
                 
                   2.85 
                 
                 0.002 
                 
                   1.51 
                 
                 0.0035 
                 
                   3.32 
                 
                 2.84E−07 
                 32415 
                 C16533 
               
               
                 arachidonate (20:4n6) 
                 
                   1.51 
                 
                 0.0021 
                 
                   1.19 
                 
                 0.056 
                 
                   1.59 
                 
                 1.43E−06 
                 1110 
                 C00219 
                 HMDB01043 
               
               
                 prostaglandin I2 
                 
                   9.79 
                 
                 0.0022 
                 
                   0.32 
                 
                 1.03E−06 
                 
                   0.3 
                 
                 0.0005 
                 32466 
                 C01312 
                 HMDB01335 
               
               
                 prostaglandin A2 
                 
                   3.02 
                 
                 0.0022 
                 
                   0.7 
                 
                 0.0136 
                 1.78 
                 0.1505 
                 19761 
                 C05953 
                 HMDB02752 
               
               
                 coenzyme A 
                 
                   0.2 
                 
                 0.0024 
                 
                   3.42 
                 
                 5.88E−05 
                 0.9 
                 0.6424 
                 2936 
                 C00010 
                 HMDB01423 
               
               
                 nicotinamide adenine 
                 
                   0.42 
                 
                 0.0027 
                 1 
                 0.5185 
                 
                   0.51 
                 
                 0.0186 
                 31475 
                 C00004 
                 HMDB01487 
               
               
                 dinucleotide reduced 
               
               
                 (NADH) 
               
               
                 hydroxyurea 
                 
                   8.74 
                 
                 0.0029 
                 0.63 
                 0.1281 
                 2.41 
                 0.3576 
                 21031 
                 C07044 
               
               
                 phenylpyruvate 
                 
                   3.95 
                 
                 0.0032 
                 0.79 
                 0.8452 
                 
                   3.68 
                 
                 1.99E−05 
                 566 
                 C00166 
                 HMDB00205 
               
               
                 7-alpha-hydroxy-3-oxo-4- 
                 
                   2.05 
                 
                 0.0036 
                 
                   0.62 
                 
                 0.0006 
                 1.08 
                 0.9305 
                 36776 
                 C17337 
                 HMDB12458 
               
               
                 cholestenoate (7-Hoca) 
               
               
                 1- 
                 
                   1.94 
                 
                 0.0041 
                 0.83 
                 0.4037 
                 
                   1.62 
                 
                 0.0004 
                 34214 
               
               
                 arachidonoylglycerophosphoinositol 
               
               
                 prostaglandin B2 
                 
                   3.53 
                 
                 0.0042 
                 
                   0.74 
                 
                 0.0842 
                 
                   2.44 
                 
                 0.0178 
                 19499 
                 C05954 
                 HMDB04236 
               
               
                 anthranilate 
                 
                   1.96 
                 
                 0.0042 
                 1.11 
                 0.4537 
                 
                   1.94 
                 
                 2.07E−05 
                 4970 
                 C00108 
                 HMDB01123 
               
               
                 N-acetylserine 
                 
                   1.52 
                 
                 0.0048 
                 0.9 
                 0.6112 
                 
                   1.21 
                 
                 0.0785 
                 37076 
                   
                 HMDB02931 
               
               
                 3′-dephosphocoenzyme A 
                 
                   0.23 
                 
                 0.0058 
                 
                   3.07 
                 
                 0.0008 
                 0.91 
                 0.9151 
                 18289 
                 C00882 
                 HMDB01373 
               
               
                 piperine 
                 
                   0.65 
                 
                 0.006 
                 1 
                 0.1707 
                 0.88 
                 0.7443 
                 33935 
                 C03882 
               
               
                 15-HETE 
                 
                   3.05 
                 
                 0.0062 
                 
                   0.45 
                 
                 0.005 
                 2.03 
                 0.6307 
                 37538 
                 C04742 
                 HMDB02110 
               
               
                 stearoyl sphingomyelin 
                 
                   1.86 
                 
                 0.0063 
                 
                   0.35 
                 
                 2.29E−06 
                 
                   0.47 
                 
                 4.04E−05 
                 19503 
                 C00550 
                 HMDB01348 
               
               
                 prostaglandin E2 
                 
                   2.85 
                 
                 0.0063 
                 
                   0.88 
                 
                 0.084 
                 
                   2.25 
                 
                 0.0417 
                 7746 
                 C00584 
                 HMDB01220 
               
               
                 N-acetylmannosamine 
                 
                   2.33 
                 
                 0.0069 
                 0.92 
                 0.7686 
                 
                   1.9 
                 
                 0.0075 
                 15060 
                 C00140 
                 HMDB00835 
               
               
                 tetrahydrocortisone 
                 
                   3.76 
                 
                 0.007 
                 0.61 
                 0.2577 
                 
                   1.72 
                 
                 0.0724 
                 38608 
                 HMDB00903 
                 HMDB00903 
               
               
                 ADSGEGDFXAEGGGV 
                 
                   1.96 
                 
                 0.0074 
                 0.82 
                 0.1436 
                 1.12 
                 0.979 
                 33084 
               
               
                 R (SEQ ID NO: 3) 
               
               
                 nicotinamide adenine 
                 
                   0.27 
                 
                 0.0084 
                 1.8 
                 0.288 
                 
                   0.72 
                 
                 0.008 
                 5278 
                 C00003 
                 HMDB00902 
               
               
                 dinucleotide (NAD+) 
               
               
                 octanoylcarnitine 
                 
                   2.78 
                 
                 0.0088 
                 
                   0.6 
                 
                 0.0075 
                 1.29 
                 0.6915 
                 33936 
               
               
                 5-methylthioadenosine 
                 
                   0.43 
                 
                 0.0094 
                 
                   5.25 
                 
                 2.20E−06 
                 
                   2.21 
                 
                 0.0007 
                 1419 
                 C00170 
                 HMDB01173 
               
               
                 (MTA) 
               
               
                 cholesterol 
                 
                   1.16 
                 
                 0.0103 
                 1.07 
                 0.2063 
                 
                   1.16 
                 
                 0.0141 
                 63 
                 C00187 
                 HMDB00067 
               
               
                 urate 
                 
                   1.59 
                 
                 0.0106 
                 
                   0.86 
                 
                 0.0889 
                 1.22 
                 0.1074 
                 1604 
                 C00366 
                 HMDB00289 
               
               
                 flavin mononucleotide 
                 
                   1.59 
                 
                 0.0107 
                 0.8 
                 0.1668 
                 1.12 
                 0.2762 
                 15797 
                 C00061 
                 HMDB01520 
               
               
                 (FMN) 
               
               
                 quinate 
                 
                   2.83 
                 
                 0.011 
                 0.72 
                 0.1339 
                 1.29 
                 0.1417 
                 18335 
                 C00296 
                 HMDB03072 
               
               
                 N-(2-furoyl)glycine 
                 
                   2.71 
                 
                 0.0112 
                 
                   0.46 
                 
                 0.0209 
                 1.55 
                 0.5573 
                 31536 
                   
                 HMDB00439 
               
               
                 beta-tocopherol 
                 
                   1.84 
                 
                 0.0112 
                 
                   0.6 
                 
                 0.021 
                 1.12 
                 0.5734 
                 35702 
                 C14152 
                 HMDB06335 
               
               
                 stearate (18:0) 
                 
                   1.36 
                 
                 0.0112 
                 
                   1.08 
                 
                 0.0786 
                 
                   1.32 
                 
                 0.0003 
                 1358 
                 C01530 
                 HMDB00827 
               
               
                 hexanoylcarnitine 
                 
                   2.19 
                 
                 0.0113 
                 
                   0.63 
                 
                 0.0065 
                 1.2 
                 0.6129 
                 32328 
                 C01585 
                 HMDB00705 
               
               
                 valylserine 
                 
                   0.49 
                 
                 0.0115 
                 
                   1.4 
                 
                 0.0056 
                 0.7 
                 0.8669 
                 40716 
               
               
                 phytosphingosine 
                 
                   0.6 
                 
                 0.0132 
                 
                   3.08 
                 
                 1.96E−05 
                 1.61 
                 0.1345 
                 1510 
                 C12144 
                 HMDB04610 
               
               
                 prostaglandin D2 
                 
                   2.66 
                 
                 0.0139 
                 
                   0.3 
                 
                 4.89E−06 
                 
                   0.63 
                 
                 0.0032 
                 7737 
                 C00696 
                 HMDB01403 
               
               
                 cyclo(gly-phe) 
                 
                   0.52 
                 
                 0.0147 
                 
                   1.49 
                 
                 0.0453 
                 0.83 
                 0.9392 
                 37102 
               
               
                 glucose 1-phosphate 
                 
                   0.42 
                 
                 0.0153 
                 
                   1.96 
                 
                 0.017 
                 0.97 
                 0.4461 
                 33755 
                 C00103 
                 HMDB01586 
               
               
                 dihydrobiopterin 
                 
                   1.96 
                 
                 0.0159 
                 1.16 
                 0.2339 
                 
                   2.01 
                 
                 0.0002 
                 35129 
                 C02953, 
                 HMDB00038 
               
               
                   
                   
                   
                   
                   
                   
                   
                   
                 C00268 
               
               
                 adenosine 2′- 
                 
                   0.51 
                 
                 0.0166 
                 
                   2.62 
                 
                 0.001 
                 1.29 
                 0.4785 
                 36815 
                 C00946 
                 HMDB11617 
               
               
                 monophosphate (2′- 
               
               
                 AMP) 
               
               
                 eicosenoate (20:1n9 or 
                 
                   1.74 
                 
                 0.017 
                 
                   1.43 
                 
                 0.0112 
                 
                   2.27 
                 
                 2.16E−06 
                 33587 
                   
                 HMDB02231 
               
               
                 11) 
               
               
                 galactose 
                 
                   0.62 
                 
                 0.0184 
                 
                   3.09 
                 
                 0.0006 
                 
                   2.04 
                 
                 0.036 
                 12055 
                 C01582 
                 HMDB00143 
               
               
                 alpha-hydroxyisovalerate 
                 
                   1.46 
                 
                 0.0185 
                 1.16 
                 0.3385 
                 
                   1.42 
                 
                 0.0041 
                 33937 
                   
                 HMDB00407 
               
               
                 prolylleucine 
                 
                   0.49 
                 
                 0.0203 
                 
                   1.55 
                 
                 0.0328 
                 0.99 
                 0.7695 
                 31914 
               
               
                 ophthalmate 
                 
                   0.31 
                 
                 0.0225 
                 1.77 
                 0.2074 
                 
                   0.62 
                 
                 0.0374 
                 34592 
                   
                 HMDB05765 
               
               
                 phosphopantetheine 
                 
                   0.5 
                 
                 0.0237 
                 
                   1.71 
                 
                 0.0071 
                 0.87 
                 0.4497 
                 15504 
                 C01134 
                 HMDB01416 
               
               
                 glycocholate 
                 
                   1.89 
                 
                 0.0238 
                 2.05 
                 0.8174 
                 
                   1.73 
                 
                 0.0719 
                 18476 
                 C01921 
                 HMDB00138 
               
               
                 nonadecanoate (19:0) 
                 
                   1.53 
                 
                 0.0249 
                 
                   1.33 
                 
                 0.0108 
                 
                   1.68 
                 
                 7.02E−05 
                 1356 
                 C16535 
                 HMDB00772 
               
               
                 cystine 
                 
                   2.05 
                 
                 0.0284 
                 
                   0.4 
                 
                 0.0463 
                 0.89 
                 0.5401 
                 39512 
                 C00491 
                 HMDB00192 
               
               
                 docosahexaenoate 
                 
                   1.43 
                 
                 0.0288 
                 
                   1.56 
                 
                 0.0188 
                 
                   2.18 
                 
                 8.50E−08 
                 19323 
                 C06429 
                 HMDB02183 
               
               
                 (DHA; 22:6n3) 
               
               
                 sucrose 
                 
                   1.85 
                 
                 0.0298 
                 0.91 
                 0.6263 
                 3.1 
                 0.1134 
                 1519 
                 C00089 
                 HMDB00258 
               
               
                 biliverdin 
                 
                   1.6 
                 
                 0.0308 
                 
                   0.71 
                 
                 0.0222 
                 1.05 
                 0.6571 
                 2137 
                 C00500 
                 HMDB01008 
               
               
                 AICA ribonucleotide 
                 
                   0.48 
                 
                 0.0321 
                 
                   1.7 
                 
                 0.0342 
                 1.04 
                 0.4794 
                 38325 
               
               
                 pregnanediol-3- 
                 
                   3.46 
                 
                 0.0328 
                 0.81 
                 0.8372 
                 
                   2.04 
                 
                 0.0414 
                 40708 
               
               
                 glucuronide 
               
               
                 phenylalanylphenylalanine 
                 
                   1.78 
                 
                 0.0329 
                 1.09 
                 0.2419 
                 
                   1.45 
                 
                 0.014 
                 38150 
               
               
                 docosapentaenoate (n3 
                 
                   1.56 
                 
                 0.0332 
                 
                   1.59 
                 
                 0.0044 
                 
                   2.36 
                 
                 4.92E−07 
                 32504 
                 C16513 
                 HMDB01976 
               
               
                 DPA; 22:5n3) 
               
               
                 glycochenodeoxycholate 
                 
                   1.25 
                 
                 0.0336 
                 1.56 
                 0.2622 
                 1.49 
                 0.7427 
                 32346 
                 C05466 
                 HMDB00637 
               
               
                 valylhistidine 
                 
                   0.57 
                 
                 0.0337 
                 
                   1.6 
                 
                 0.0054 
                 0.84 
                 0.7998 
                 40680 
               
               
                 N-acetylputrescine 
                 
                   1.58 
                 
                 0.0352 
                 0.94 
                 0.9693 
                 1.23 
                 0.1251 
                 37496 
                 C02714 
                 HMDB02064 
               
               
                 gamma-tocopherol 
                 
                   1.57 
                 
                 0.0359 
                 
                   0.6 
                 
                 0.0301 
                 1.11 
                 0.6365 
                 33420 
                 C02483 
                 HMDB01492 
               
               
                 cytidine-3′- 
                 
                   2.01 
                 
                 0.0361 
                 1.11 
                 0.3741 
                 
                   1.68 
                 
                 0.0229 
                 2959 
                 C05822 
               
               
                 monophosphate (3′- 
               
               
                 CMP) 
               
               
                 5-HETE 
                 
                   2.36 
                 
                 0.0369 
                 1.17 
                 0.3259 
                 
                   2.54 
                 
                 0.0006 
                 37372 
               
               
                 2- 
                 
                   0.52 
                 
                 0.0374 
                 
                   2.29 
                 
                 0.0003 
                 
                   1.34 
                 
                 0.0883 
                 34666 
               
               
                 linoleoylglycerophosphoethanolamine 
               
               
                 maltotriose 
                 
                   0.66 
                 
                 0.0376 
                 0.74 
                 0.6695 
                 
                   0.42 
                 
                 0.0121 
                 27723 
                 C01835 
                 HMDB01262 
               
               
                 maltotetraose 
                 
                   0.72 
                 
                 0.0385 
                 0.85 
                 0.5162 
                 
                   0.52 
                 
                 0.0303 
                 15910 
                 C02052 
                 HMDB01296 
               
               
                 tryptophylasparagine 
                 
                   0.56 
                 
                 0.0387 
                 
                   1.52 
                 
                 0.0412 
                 0.87 
                 0.7366 
                 40661 
               
               
                 allantoin 
                 
                   3.32 
                 
                 0.0395 
                 1.04 
                 0.9406 
                 
                   2.53 
                 
                 0.0289 
                 1107 
                 C02350 
                 HMDB00462 
               
               
                 1- 
                 
                   0.72 
                 
                 0.0399 
                 
                   1.7 
                 
                 0.0005 
                 
                   1.41 
                 
                 0.0385 
                 34419 
                 C04100 
               
               
                 linoleoylglycerophosphocholine 
               
               
                 N-acetylglutamate 
                 
                   1.92 
                 
                 0.0401 
                 0.55 
                 0.9941 
                 
                   0.95 
                 
                 0.04 
                 15720 
                 C00624 
                 HMDB01138 
               
               
                 nicotinamide 
                 
                   0.38 
                 
                 0.0416 
                 
                   0.46 
                 
                 0.0212 
                 
                   0.23 
                 
                 1.50E−05 
                 22152 
                 C00455 
                 HMDB00229 
               
               
                 ribonucleotide (NMN) 
               
               
                 isovalerylcarnitine 
                 
                   2.93 
                 
                 0.0421 
                 0.86 
                 0.2523 
                 1.77 
                 0.3309 
                 34407 
                   
                 HMDB00688 
               
               
                 uridine monophosphate 
                 
                   0.33 
                 
                 0.0425 
                 
                   2.37 
                 
                 0.002 
                 0.97 
                 0.5172 
                 39879 
               
               
                 (5′ or 3′) 
               
               
                 ribose 
                 
                   0.61 
                 
                 0.0432 
                 
                   2.65 
                 
                 0.0035 
                 1.66 
                 0.3101 
                 12080 
                 C00121 
                 HMDB00283 
               
               
                 dihomo-linoleate 
                 
                   1.73 
                 
                 0.0447 
                 
                   1.57 
                 
                 0.0035 
                 
                   2.35 
                 
                 2.26E−06 
                 17805 
                 C16525 
               
               
                 (20:2n6) 
               
               
                 leucylarginine 
                 
                   0.74 
                 
                 0.0449 
                 0.87 
                 0.9852 
                 
                   0.7 
                 
                 0.0427 
                 40028 
               
               
                 glycerol 
                 
                   0.67 
                 
                 0.0453 
                 
                   1.63 
                 
                 0.0022 
                 1.19 
                 0.3141 
                 15122 
                 C00116 
                 HMDB00131 
               
               
                 maltopentaose 
                 
                   0.66 
                 
                 0.0454 
                 1.19 
                 0.3849 
                 
                   0.75 
                 
                 0.0678 
                 35163 
                 C06218 
                 HMDB12254 
               
               
                 N-acetylasparagine 
                 
                   2.75 
                 
                 0.0456 
                 0.95 
                 0.3805 
                 
                   2.1 
                 
                 0.0714 
                 33942 
                   
                 HMDB06028 
               
               
                 citrate 
                 
                   3.07 
                 
                 0.046 
                 0.23 
                 0.2687 
                 
                   0.74 
                 
                 0.0732 
                 1564 
                 C00158 
                 HMDB00094 
               
               
                 13-HODE + 9-HODE 
                 
                   1.56 
                 
                 0.0474 
                 
                   0.33 
                 
                 5.97E−06 
                 
                   0.65 
                 
                 0.0037 
                 37752 
               
               
                 uridine 
                 
                   0.7 
                 
                 0.0481 
                 1.01 
                 0.7099 
                 
                   0.82 
                 
                 0.0168 
                 606 
                 C00299 
                 HMDB00296 
               
               
                 1-stearoylglycerol (1- 
                 
                   1.36 
                 
                 0.0494 
                 
                   1.4 
                 
                 0.0113 
                 
                   1.6 
                 
                 0.0002 
                 21188 
                 D01947 
               
               
                 monostearin) 
               
               
                 cytidine-5′- 
                 
                   0.49 
                 
                 0.0524 
                 
                   2.53 
                 
                 0.0038 
                 1.27 
                 0.4346 
                 34410 
                 C00570 
                 HMDB01564 
               
               
                 diphosphoethanolamine 
               
               
                 2- 
                 
                   0.6 
                 
                 0.0532 
                 
                   2.49 
                 
                 0.0002 
                 
                   1.59 
                 
                 0.0287 
                 35257 
               
               
                 linoleoylglycerophosphocholine 
               
               
                 pyroglutamylglutamine 
                 
                   2.13 
                 
                 0.0539 
                 0.52 
                 0.1377 
                 1.28 
                 0.1097 
                 22194 
               
               
                 fructose-6-phosphate 
                 
                   2.9 
                 
                 0.0546 
                 
                   0.67 
                 
                 0.0851 
                 1.85 
                 0.3968 
                 12021 
                 C05345 
                 HMDB00124 
               
               
                 2-linoleoylglycerol (2- 
                 
                   0.62 
                 
                 0.0547 
                 
                   3.42 
                 
                 1.93E−07 
                 
                   1.94 
                 
                 0.0002 
                 32506 
                   
                 HMDB11538 
               
               
                 monolinolein) 
               
               
                 dihomo-linolenate 
                 
                   1.48 
                 
                 0.0573 
                 
                   1.71 
                 
                 0.0037 
                 
                   2.24 
                 
                 3.61E−07 
                 35718 
                 C03242 
                 HMDB02925 
               
               
                 (20:3n3 or n6) 
               
               
                 leu-leu-leu 
                 
                   2.78 
                 
                 0.0578 
                 
                   0.82 
                 
                 0.0908 
                 1.62 
                 0.5602 
                 40672 
               
               
                 androsterone sulfate 
                 
                   2.38 
                 
                 0.0585 
                 
                   0.53 
                 
                 0.0125 
                 1.11 
                 0.6683 
                 31591 
                 C00523 
                 HMDB02759 
               
               
                 dehydroisoandrosterone 
                 
                   2.08 
                 
                 0.0588 
                 
                   0.48 
                 
                 0.0047 
                 0.94 
                 0.4439 
                 32425 
                 C04555 
                 HMDB01032 
               
               
                 sulfate (DHEA-S) 
               
               
                 pregnen-diol disulfate 
                 
                   1.61 
                 
                 0.0592 
                 
                   0.51 
                 
                 0.0048 
                 0.95 
                 0.9966 
                 32562 
                 C05484 
                 HMDB04025 
               
               
                 3-hydroxyhippurate 
                 
                   8.42 
                 
                 0.0597 
                 0.69 
                 0.3145 
                 3.98 
                 0.26 
                 39600 
                   
                 HMDB06116 
               
               
                 2- 
                 
                   0.65 
                 
                 0.0608 
                 1.33 
                 0.1111 
                 0.84 
                 0.6281 
                 34656 
               
               
                 arachidonoylglycerophosphoethanolamine 
               
               
                 hexanoylglycine 
                 
                   1.61 
                 
                 0.0613 
                 0.92 
                 0.956 
                 1.17 
                 0.2758 
                 35436 
                   
                 HMDB00701 
               
               
                 creatine phosphate 
                 
                   12.62 
                 
                 0.062 
                 
                   0.33 
                 
                 0.0001 
                 
                   0.84 
                 
                 0.0004 
                 33951 
                 C02305 
                 HMDB01511 
               
               
                 N2,N2- 
                 
                   1.61 
                 
                 0.0662 
                 
                   1.38 
                 
                 0.0064 
                 
                   1.88 
                 
                 2.82E−05 
                 35137 
                   
                 HMDB04824 
               
               
                 dimethylguanosine 
               
               
                 1- 
                 
                   0.86 
                 
                 0.0677 
                 
                   2.09 
                 
                 5.74E−05 
                 
                   1.8 
                 
                 0.0087 
                 33960 
               
               
                 oleoylglycerophosphocholine 
               
               
                 maltose 
                 
                   0.71 
                 
                 0.0677 
                 0.8 
                 0.8895 
                 
                   0.55 
                 
                 0.0134 
                 15806 
                 C00208 
                 HMDB00163 
               
               
                 hexadecanedioate 
                 
                   1.36 
                 
                 0.0696 
                 
                   0.52 
                 
                 4.71E−05 
                 
                   0.63 
                 
                 0.0006 
                 35678 
                   
                 HMDB00672 
               
               
                 alanylvaline 
                 
                   2.1 
                 
                 0.0718 
                 1.01 
                 0.4229 
                 
                   1.53 
                 
                 0.0285 
                 37084 
               
               
                 1- 
                 
                   1.57 
                 
                 0.0738 
                 
                   0.48 
                 
                 0.0002 
                 
                   0.66 
                 
                 0.0094 
                 32350 
                 C05828 
                 HMDB02820 
               
               
                 methylimidazoleacetate 
               
               
                 1- 
                 
                   2.1 
                 
                 0.0745 
                 1.18 
                 0.2032 
                 
                   2.27 
                 
                 6.06E−06 
                 19260 
               
               
                 oleoylglycerophosphoserine 
               
               
                 1- 
                 
                   0.7 
                 
                 0.0746 
                 
                   1.6 
                 
                 0.0798 
                 1.33 
                 0.1879 
                 37418 
               
               
                 pentadecanoylglycerophosphocholine 
               
               
                 anserine 
                 
                   1.31 
                 
                 0.0749 
                 0.84 
                 0.8878 
                 0.72 
                 0.3447 
                 15747 
                 C01262 
                 HMDB00194 
               
               
                 isoleucylproline 
                 
                   0.69 
                 
                 0.075 
                 
                   1.87 
                 
                 2.63E−05 
                 
                   1.41 
                 
                 0.0008 
                 35418 
                   
                 HMDB11174 
               
               
                 tyrosylleucine 
                 
                   0.75 
                 
                 0.0751 
                 
                   1.71 
                 
                 0.0003 
                 1.06 
                 0.1271 
                 40031 
               
               
                 cinnamoylglycine 
                 
                   2 
                 
                 0.0754 
                 0.92 
                 0.8183 
                 1.35 
                 0.3202 
                 38637 
               
               
                 pseudouridine 
                 
                   1.57 
                 
                 0.0767 
                 1.1 
                 0.1259 
                 
                   1.58 
                 
                 0.0014 
                 33442 
                 C02067 
                 HMDB00767 
               
               
                 N6-acetyllysine 
                 
                   1.27 
                 
                 0.0775 
                 0.98 
                 0.1559 
                 
                   1.22 
                 
                 0.0281 
                 36752 
                 C02727 
                 HMDB00206 
               
               
                 erucamide 
                 
                   1.39 
                 
                 0.08 
                 0.98 
                 0.9235 
                 1.04 
                 0.5864 
                 41729 
               
               
                 galactosylsphingosine 
                 
                   1.51 
                 
                 0.081 
                 1 
                 0.9588 
                 
                   1.32 
                 
                 0.0553 
                 40083 
                   
                 HMDB00648 
               
               
                 pyrophosphate (PPi) 
                 
                   1.45 
                 
                 0.0817 
                 
                   0.26 
                 
                 0.0276 
                 0.3 
                 0.2252 
                 2078 
                 C00013 
                 HMDB00250 
               
               
                 pyruvate 
                 
                   0.48 
                 
                 0.0833 
                 1.82 
                 0.2579 
                 1.02 
                 0.4122 
                 599 
                 C00022 
                 HMDB00243 
               
               
                 2-palmitoylglycerol (2- 
                 
                   0.74 
                 
                 0.0844 
                 
                   2.15 
                 
                 5.60E−07 
                 
                   1.84 
                 
                 2.56E−05 
                 33419 
               
               
                 monopalmitin) 
               
               
                 pinitol 
                 
                   0.57 
                 
                 0.0855 
                 
                   1.63 
                 
                 0.0104 
                 1.01 
                 0.2576 
                 37086 
                 C03844 
               
               
                 2- 
                 
                   0.96 
                 
                 0.0871 
                 
                   1.16 
                 
                 0.0427 
                 1.07 
                 0.8636 
                 34875 
               
               
                 docosapentaenoylglycerophosphoethanolamine 
               
               
                 stachydrine 
                 
                   1.49 
                 
                 0.0878 
                 1.05 
                 0.7163 
                 
                   1.29 
                 
                 0.0146 
                 34384 
                 C10172 
                 HMDB04827 
               
               
                 tryptophan betaine 
                 
                   2.5 
                 
                 0.0895 
                 0.82 
                 0.4502 
                 1.54 
                 0.3282 
                 37097 
                 C09213 
               
               
                 levulinate (4-oxovalerate) 
                 
                   1.28 
                 
                 0.0896 
                 
                   1.18 
                 
                 0.0392 
                 
                   1.4 
                 
                 0.0004 
                 22177 
                   
                 HMDB00720 
               
               
                 isoleucylserine 
                 
                   0.57 
                 
                 0.0921 
                 
                   1.4 
                 
                 0.0586 
                 0.78 
                 0.8995 
                 40012 
               
               
                 2-hydroxystearate 
                 
                   1.38 
                 
                 0.093 
                 0.88 
                 0.3023 
                 0.9 
                 0.6961 
                 17945 
                 C03045 
               
               
                 isoleucylglycine 
                 
                   0.71 
                 
                 0.0954 
                 0.84 
                 0.4451 
                 
                   0.64 
                 
                 0.0051 
                 40008 
               
               
                 glycerate 
                 
                   0.67 
                 
                 0.0966 
                 
                   1.47 
                 
                 0.0724 
                 1.33 
                 0.6707 
                 1572 
                 C00258 
                 HMDB00139 
               
               
                 4-androsten- 
                 
                   1.62 
                 
                 0.0971 
                 
                   0.44 
                 
                 0.0196 
                 0.78 
                 0.6699 
                 37202 
                   
                 HMDB03818 
               
               
                 3beta,17beta-diol 
               
               
                 disulfate 1 
               
               
                 urea 
                 
                   1.64 
                 
                 0.1 
                 0.93 
                 0.8471 
                 1.24 
                 0.3327 
                 1670 
                 C00086 
                 HMDB00294 
               
               
                 sedoheptulose-7- 
                 0.39 
                 0.1008 
                 
                   2.12 
                 
                 0.0623 
                 1.16 
                 0.5952 
                 35649 
                 C05382 
                 HMDB01068 
               
               
                 phosphate 
               
               
                 threitol 
                 1.91 
                 0.1021 
                 0.76 
                 0.6423 
                 
                   1.28 
                 
                 0.0922 
                 35854 
                 C16884 
                 HMDB04136 
               
               
                 2- 
                 0.81 
                 0.105 
                 
                   1.82 
                 
                 0.0062 
                 1.45 
                 0.2221 
                 35683 
               
               
                 oleoylglycerophosphoethanolamine 
               
               
                 alpha-glutamyltyrosine 
                 1.52 
                 0.1051 
                 0.89 
                 0.8683 
                 
                   1.17 
                 
                 0.0601 
                 40033 
               
               
                 gamma- 
                 1.53 
                 0.1058 
                 
                   0.48 
                 
                 0.0001 
                 
                   0.74 
                 
                 0.0255 
                 2730 
                   
                 HMDB11738 
               
               
                 glutamylglutamine 
               
               
                 1- 
                 0.65 
                 0.1104 
                 
                   2.05 
                 
                 0.0014 
                 1.36 
                 0.1452 
                 33957 
                   
                 HMDB12108 
               
               
                 heptadecanoylglycerophosphocholine 
               
               
                 gamma- 
                 1.75 
                 0.1123 
                 
                   0.4 
                 
                 8.13E−05 
                 
                   0.6 
                 
                 0.0093 
                 36738 
               
               
                 glutamylglutamate 
               
               
                 17-methylstearate 
                 1.56 
                 0.1123 
                 
                   1.51 
                 
                 0.0019 
                 
                   1.96 
                 
                 8.51E−05 
                 38296 
               
               
                 hydroxyisovaleroyl 
                 1.6 
                 0.1161 
                 1.3 
                 0.3681 
                 
                   1.85 
                 
                 0.0002 
                 35433 
               
               
                 carnitine 
               
               
                 deoxycarnitine 
                 1.39 
                 0.1164 
                 
                   1.53 
                 
                 0.0004 
                 
                   1.75 
                 
                 3.59E−06 
                 36747 
                 C01181 
                 HMDB01161 
               
               
                 myo-inositol 
                 0.63 
                 0.1182 
                 
                   0.52 
                 
                 0.0008 
                 
                   0.44 
                 
                 4.85E−07 
                 19934 
                 C00137 
                 HMDB00211 
               
               
                 cholate 
                 2.11 
                 0.1206 
                 1.11 
                 0.4538 
                 
                   1.92 
                 
                 0.0152 
                 22842 
                 C00695 
                 HMDB00619 
               
               
                 valylaspartate 
                 0.77 
                 0.1216 
                 
                   1.69 
                 
                 0.0068 
                 1.28 
                 0.1109 
                 40650 
               
               
                 vanillylmandelate (VMA) 
                 2.51 
                 0.1271 
                 1.11 
                 0.7826 
                 
                   2.17 
                 
                 0.0346 
                 1567 
                 C05584 
                 HMDB00291 
               
               
                 4-hydroxyphenylacetate 
                 2.02 
                 0.1298 
                 1.48 
                 0.6131 
                 
                   2.4 
                 
                 0.0416 
                 541 
                 C00642 
                 HMDB00020 
               
               
                 2- 
                 0.85 
                 0.1301 
                 
                   2.81 
                 
                 4.63E−05 
                 
                   2.41 
                 
                 0.0054 
                 35254 
               
               
                 oleoylglycerophosphocholine 
               
               
                 gamma-glutamylalanine 
                 1.62 
                 0.1321 
                 
                   0.47 
                 
                 6.48E−06 
                 
                   0.75 
                 
                 0.009 
                 37063 
               
               
                 5-methyluridine 
                 0.74 
                 0.133 
                 
                   1.33 
                 
                 0.0566 
                 1.05 
                 0.4453 
                 35136 
                   
                 HMDB00884 
               
               
                 (ribothymidine) 
               
               
                 glycerophosphoethanolamine 
                 0.46 
                 0.1356 
                 
                   6.97 
                 
                 0.001 
                 
                   1.83 
                 
                 0.0199 
                 37455 
                 C01233 
                 HMDB00114 
               
               
                 cyclo(leu-gly) 
                 1.11 
                 0.1399 
                 
                   0.63 
                 
                 0.0017 
                 
                   0.66 
                 
                 0.004 
                 37078 
               
               
                 UDP-glucuronate 
                 0.43 
                 0.14 
                 
                   3.9 
                 
                 0.0005 
                 
                   1.66 
                 
                 0.0336 
                 2763 
                 C00167 
                 HMDB00935 
               
               
                 alpha-glutamyllysine 
                 1.48 
                 0.1418 
                 
                   0.54 
                 
                 0.0013 
                 
                   0.76 
                 
                 0.0134 
                 40441 
                   
                 HMDB04207 
               
               
                 5-oxoproline 
                 1.21 
                 0.1433 
                 
                   0.74 
                 
                 0.0456 
                 
                   0.76 
                 
                 0.0481 
                 1494 
                 C01879 
                 HMDB00267 
               
               
                 valylasparagine 
                 0.49 
                 0.1452 
                 
                   1.96 
                 
                 0.02 
                 1.02 
                 0.4282 
                 40727 
                 C00252 
                 HMDB02923 
               
               
                 2- 
                 0.85 
                 0.1466 
                 
                   1.74 
                 
                 0.063 
                 1.34 
                 0.6746 
                 34258 
               
               
                 docosahexaenoylglycerophosphoethanolamine 
               
               
                 octadecanedioate 
                 1.32 
                 0.1475 
                 
                   0.71 
                 
                 0.0073 
                 
                   0.84 
                 
                 0.0439 
                 36754 
                   
                 HMDB00782 
               
               
                 4-androsten- 
                 1.59 
                 0.1485 
                 
                   0.62 
                 
                 0.0162 
                 0.96 
                 0.6038 
                 37203 
                   
                 HMDB03818 
               
               
                 3beta,17beta-diol 
               
               
                 disulfate 2 
               
               
                 1- 
                 0.85 
                 0.1506 
                 
                   1.33 
                 
                 0.0108 
                 1.13 
                 0.1235 
                 33955 
               
               
                 palmitoylglycerophosphocholine 
               
               
                 aspartylaspartate 
                 1.12 
                 0.1506 
                 
                   0.69 
                 
                 0.0373 
                 0.7 
                 0.1316 
                 40671 
               
               
                 valylglycine 
                 0.8 
                 0.1508 
                 
                   1.27 
                 
                 0.0012 
                 
                   1 
                 
                 0.0088 
                 40475 
               
               
                 8-iso-15-keto- 
                 1.48 
                 0.1522 
                 1.78 
                 0.4758 
                 
                   2.33 
                 
                 0.0175 
                 7758 
                 C04707 
                 HMDB02341 
               
               
                 prostaglandin E2 
               
               
                 stearoyl ethanolamide 
                 5.03 
                 0.1536 
                 1.34 
                 0.1313 
                 
                   4.71 
                 
                 0.0067 
                 38625 
               
               
                 oleic ethanolamide 
                 1.99 
                 0.1551 
                 1.55 
                 0.1484 
                 
                   2.97 
                 
                 0.001 
                 38102 
                   
                 HMDB02088 
               
               
                 isoleucylalanine 
                 0.75 
                 0.1584 
                 
                   1.51 
                 
                 0.0172 
                 1.13 
                 0.2253 
                 40046 
               
               
                 3-dehydrocarnitine 
                 0.76 
                 0.1594 
                 
                   1.34 
                 
                 0.0475 
                 1.09 
                 0.5941 
                 32654 
               
               
                 glycerol 3-phosphate 
                 0.44 
                 0.1611 
                 1 
                 0.7146 
                 
                   0.45 
                 
                 0.0013 
                 15365 
                 C00093 
                 HMDB00126 
               
               
                 (G3P) 
               
               
                 cysteinylglycine 
                 0.68 
                 0.1653 
                 0.99 
                 0.8233 
                 
                   0.6 
                 
                 0.0012 
                 35637 
                 C01419 
                 HMDB00078 
               
               
                 inosine 
                 0.8 
                 0.1679 
                 0.87 
                 0.1343 
                 
                   0.72 
                 
                 0.0005 
                 1123 
               
               
                 scyllo-inositol 
                 0.72 
                 0.1718 
                 
                   0.43 
                 
                 0.0011 
                 
                   0.33 
                 
                 6.36E−07 
                 32379 
                 C06153 
                 HMDB06088 
               
               
                 erythronate 
                 1.54 
                 0.1718 
                 
                   1.61 
                 
                 0.0018 
                 
                   2.2 
                 
                 1.35E−05 
                 33477 
                   
                 HMDB00613 
               
               
                 gamma- 
                 1.26 
                 0.1733 
                 
                   1.22 
                 
                 0.0187 
                 
                   1.37 
                 
                 0.0133 
                 34456 
                   
                 HMDB11170 
               
               
                 glutamylisoleucine 
               
               
                 glutathione, reduced 
                 0.55 
                 0.1753 
                 
                   1.86 
                 
                 0.0715 
                 0.93 
                 0.547 
                 2127 
                 C00051 
                 HMDB00125 
               
               
                 (GSH) 
               
               
                 valylvaline 
                 0.86 
                 0.1756 
                 
                   1.83 
                 
                 0.0043 
                 1.32 
                 0.335 
                 40728 
               
               
                 ergothioneine 
                 1.56 
                 0.182 
                 
                   1.32 
                 
                 0.0192 
                 
                   1.85 
                 
                 0.0002 
                 37459 
                 C05570 
                 HMDB03045 
               
               
                 7-methylguanine 
                 1.52 
                 0.1864 
                 
                   1.27 
                 
                 0.0112 
                 
                   1.66 
                 
                 0.0022 
                 35114 
                 C02242 
                 HMDB00897 
               
               
                 2-aminoadipate 
                 1.28 
                 0.1903 
                 
                   0.79 
                 
                 0.0415 
                 0.93 
                 0.3644 
                 6146 
                 C00956 
                 HMDB00510 
               
               
                 valylisoleucine 
                 1.51 
                 0.1908 
                 
                   1.38 
                 
                 0.04 
                 
                   1.62 
                 
                 0.0053 
                 40050 
               
               
                 phosphoenolpyruvate 
                 3.1 
                 0.1912 
                 
                   0.28 
                 
                 0.0057 
                 
                   0.33 
                 
                 0.0271 
                 597 
                 C00074 
                 HMDB00263 
               
               
                 (PEP) 
               
               
                 S-adenosylhomocysteine 
                 0.72 
                 0.1917 
                 
                   1.91 
                 
                 0.0066 
                 1.29 
                 0.1645 
                 15948 
                 C00021 
                 HMDB00939 
               
               
                 (SAH) 
               
               
                 glycerol 2-phosphate 
                 0.57 
                 0.1918 
                 
                   3.49 
                 
                 0.0006 
                 
                   1.48 
                 
                 0.0186 
                 27728 
                 C02979, 
                 HMDB02520 
               
               
                   
                   
                   
                   
                   
                   
                   
                   
                 D01488 
               
               
                 succinylcarnitine 
                 0.71 
                 0.1934 
                 
                   2.04 
                 
                 0.0122 
                 1.19 
                 0.8095 
                 37058 
               
               
                 andro steroid 
                 1.42 
                 0.197 
                 0.85 
                 0.1191 
                 
                   1.65 
                 
                 0.099 
                 32792 
                 C04555 
                 HMDB02759 
               
               
                 monosulfate 2 
               
               
                 histidylleucine 
                 0.64 
                 0.2002 
                 0.87 
                 0.253 
                 
                   0.6 
                 
                 0.0044 
                 40061 
               
               
                 chiro-inositol 
                 2.41 
                 0.2017 
                 
                   0.53 
                 
                 0.0746 
                 1.13 
                 0.3671 
                 37112 
               
               
                 1- 
                 1.43 
                 0.2036 
                 
                   1.63 
                 
                 0.0236 
                 
                   1.91 
                 
                 6.88E−05 
                 19324 
               
               
                 stearoylglycerophosphoinositol 
               
               
                 1- 
                 1.02 
                 0.2058 
                 
                   1.61 
                 
                 0.0011 
                 
                   1.57 
                 
                 0.032 
                 33230 
               
               
                 palmitoleoylglycerophosphocholine 
               
               
                 trans-4-hydroxyproline 
                 0.83 
                 0.2064 
                 
                   1.89 
                 
                 0.0003 
                 
                   1.79 
                 
                 0.0002 
                 1366 
                 C01157 
                 HMDB00725 
               
               
                 linolenate [alpha or 
                 0.76 
                 0.2068 
                 
                   1.61 
                 
                 0.0019 
                 
                   1.32 
                 
                 0.0529 
                 34035 
                 C06427 
                 HMDB01388 
               
               
                 gamma; (18:3n3 or 6)] 
               
               
                 glycolithocholate sulfate 
                 1.37 
                 0.2111 
                 
                   0.53 
                 
                 0.0622 
                 0.69 
                 0.4016 
                 32620 
                 C11301 
                 HMDB02639 
               
               
                 glutaroyl carnitine 
                 1.58 
                 0.212 
                 1.24 
                 0.6159 
                 
                   1.69 
                 
                 0.0216 
                 35439 
                   
                 HMDB13130 
               
               
                 3-hydroxyisobutyrate 
                 1.15 
                 0.2212 
                 1.07 
                 0.8071 
                 
                   1.19 
                 
                 0.0607 
                 1549 
                 C06001 
                 HMDB00336 
               
               
                 threonate 
                 1 
                 0.226 
                 
                   1.51 
                 
                 0.0024 
                 
                   1.45 
                 
                 0.0068 
                 27738 
                 C01620 
                 HMDB00943 
               
               
                 2′-deoxyinosine 
                 0.62 
                 0.2299 
                 
                   1.72 
                 
                 0.0016 
                 
                   1.13 
                 
                 0.0135 
                 15076 
                 C05512 
                 HMDB00071 
               
               
                 behenate (22:0) 
                 1.5 
                 0.2331 
                 
                   1.46 
                 
                 0.0344 
                 
                   2 
                 
                 0.0009 
                 12125 
                 C08281 
                 HMDB00944 
               
               
                 isoleucylglutamine 
                 0.56 
                 0.2359 
                 
                   1.87 
                 
                 0.0017 
                 1.01 
                 0.1387 
                 40019 
               
               
                 dimethylarginine (SDMA + 
                 1.18 
                 0.2391 
                 
                   1.37 
                 
                 0.0007 
                 
                   1.47 
                 
                 0.0002 
                 36808 
                 C03626 
                 HMDB01539, 
               
               
                 ADMA) 
                   
                   
                   
                   
                   
                   
                   
                   
                 HMDB03334 
               
               
                 guanosine 5′- 
                 0.74 
                 0.2429 
                 
                   1.7 
                 
                 0.0224 
                 1.19 
                 0.8906 
                 2849 
               
               
                 monophosphate (5′- 
               
               
                 GMP) 
               
               
                 aspartylphenylalanine 
                 0.81 
                 0.2454 
                 
                   1.79 
                 
                 0.0127 
                 
                   1.25 
                 
                 0.0506 
                 22175 
                   
                 HMDB00706 
               
               
                 gamma-glutamylvaline 
                 1.17 
                 0.2475 
                 1.13 
                 0.1094 
                 
                   1.19 
                 
                 0.0766 
                 32393 
                   
                 HMDB11172 
               
               
                 valylalanine 
                 0.65 
                 0.2566 
                 
                   2.02 
                 
                 0.0039 
                 
                   1.34 
                 
                 0.0277 
                 41518 
               
               
                 eicosapentaenoate 
                 1.22 
                 0.26 
                 
                   1.96 
                 
                 1.32E−05 
                 
                   2.29 
                 
                 4.36E−08 
                 18467 
                 C06428 
                 HMDB01999 
               
               
                 (EPA; 20:5n3) 
               
               
                 cytidine 5′- 
                 0.7 
                 0.2628 
                 
                   4.26 
                 
                 0.0001 
                 
                   2.39 
                 
                 0.0001 
                 34418 
               
               
                 diphosphocholine 
               
               
                 xanthosine 
                 1.43 
                 0.2674 
                 1.15 
                 0.2515 
                 
                   1.59 
                 
                 0.0118 
                 15136 
                 C01762 
                 HMDB00299 
               
               
                 triethanolamine 
                 0.56 
                 0.2718 
                 0.67 
                 0.4135 
                 
                   0.11 
                 
                 0.0076 
                 22202 
                 C06771 
               
               
                 1-oleoylglycerol (1- 
                 0.69 
                 0.2742 
                 
                   2.53 
                 
                 4.22E−06 
                 
                   1.76 
                 
                 9.37E−05 
                 21184 
                   
                 HMDB11567 
               
               
                 monoolein) 
               
               
                 7,8-dihydroneopterin 
                 1.88 
                 0.2766 
                 1.52 
                 0.2489 
                 
                   1.73 
                 
                 0.088 
                 15689 
                 C04895 
                 HMDB02275 
               
               
                 L-urobilin 
                 1.48 
                 0.279 
                 
                   0.56 
                 
                 0.0307 
                 0.82 
                 0.3225 
                 40173 
                 C05793 
                 HMDB04159 
               
               
                 cis-vaccenate (18:1n7) 
                 1.28 
                 0.2854 
                 
                   1.52 
                 
                 0.0011 
                 
                   1.98 
                 
                 2.11E−06 
                 33970 
                 C08367 
               
               
                 linoleate (18:2n6) 
                 0.9 
                 0.2899 
                 
                   1.36 
                 
                 0.0053 
                 
                   1.31 
                 
                 0.0125 
                 1105 
                 C01595 
                 HMDB00673 
               
               
                 glutathione, oxidized 
                 0.89 
                 0.2905 
                 
                   1.87 
                 
                 0.021 
                 1.48 
                 0.4043 
                 27727 
                 C00127 
                 HMDB03337 
               
               
                 (GSSG) 
               
               
                 2-phosphoglycerate 
                 2.34 
                 0.3003 
                 
                   0.4 
                 
                 9.94E−05 
                 
                   0.49 
                 
                 0.0023 
                 35629 
                 C00631 
                 HMDB03391 
               
               
                 1- 
                 0.83 
                 0.3012 
                 
                   1.62 
                 
                 0.0121 
                 
                   1.46 
                 
                 0.0578 
                 33961 
               
               
                 stearoylglycerophosphocholine 
               
               
                 2-hydroxyglutarate 
                 0.09 
                 0.3026 
                 
                   5.53 
                 
                 0.03 
                 0.62 
                 0.6356 
                 37253 
                 C02630 
                 HMDB00606 
               
               
                 alanylisoleucine 
                 1.44 
                 0.3027 
                 
                   1.52 
                 
                 0.001 
                 
                   1.82 
                 
                 0.0004 
                 37118 
               
               
                 aspartylleucine 
                 0.73 
                 0.31 
                 
                   2.12 
                 
                 0.013 
                 
                   1.42 
                 
                 0.0292 
                 40068 
               
               
                 N-acetylmethionine 
                 0.82 
                 0.3113 
                 
                   1.53 
                 
                 0.0286 
                 1.29 
                 0.1216 
                 1589 
                 C02712 
                 HMDB11745 
               
               
                 1- 
                 0.94 
                 0.313 
                 
                   2.37 
                 
                 0.0005 
                 
                   2.35 
                 
                 0.001 
                 35626 
                   
                 HMDB10379 
               
               
                 myristoylglycerophosphocholine 
               
               
                 1-linoleoylglycerol (1- 
                 0.95 
                 0.3148 
                 
                   3.25 
                 
                 4.44E−06 
                 
                   2.43 
                 
                 8.20E−05 
                 27447 
               
               
                 monolinolein) 
               
               
                 acetylcarnitine 
                 1.12 
                 0.3168 
                 
                   0.8 
                 
                 0.028 
                 0.91 
                 0.3245 
                 32198 
                 C02571 
                 HMDB00201 
               
               
                 glycylvaline 
                 1.46 
                 0.3195 
                 1.21 
                 0.1363 
                 
                   1.42 
                 
                 0.0284 
                 18357 
               
               
                 guanosine 3′- 
                 2.14 
                 0.3271 
                 
                   1.33 
                 
                 0.0142 
                 
                   2.43 
                 
                 0.0017 
                 39786 
               
               
                 monophosphate (3′- 
               
               
                 GMP) 
               
               
                 isoleucylphenylalanine 
                 1.58 
                 0.3371 
                 
                   1.22 
                 
                 0.0327 
                 
                   1.59 
                 
                 0.0114 
                 40067 
               
               
                 alanylalanine 
                 1.12 
                 0.3408 
                 
                   1.28 
                 
                 0.0284 
                 1.13 
                 0.3332 
                 15129 
                 C00993 
                 HMDB03459 
               
               
                 2- 
                 1.02 
                 0.3409 
                 
                   1.7 
                 
                 0.0058 
                 1.61 
                 0.1103 
                 35256 
               
               
                 arachidonoylglycerophosphocholine 
               
               
                 1- 
                 0.86 
                 0.3417 
                 
                   1.61 
                 
                 0.0069 
                 
                   1.54 
                 
                 0.06 
                 33871 
               
               
                 eicosadienoylglycerophosphocholine 
               
               
                 N-acetylglucosamine 6- 
                 0.83 
                 0.3425 
                 
                   1.56 
                 
                 0.0735 
                 
                   1.63 
                 
                 0.072 
                 15107 
                 C00357 
                 HMDB02817 
               
               
                 phosphate 
               
               
                 5-methyltetrahydrofolate 
                 0.85 
                 0.3438 
                 
                   2.09 
                 
                 0.0031 
                 
                   1.32 
                 
                 0.09 
                 18330 
                 C00440 
                 HMDB01396 
               
               
                 (5MeTHF) 
               
               
                 choline 
                 0.93 
                 0.346 
                 
                   1.25 
                 
                 0.0182 
                 1.15 
                 0.1697 
                 15506 
               
               
                 1- 
                 0.82 
                 0.3501 
                 
                   1.65 
                 
                 0.0078 
                 
                   1.67 
                 
                 0.0002 
                 32635 
                   
                 HMDB11507 
               
               
                 linoleoylglycerophosphoethanolamine 
               
               
                 lignocerate (24:0) 
                 1.88 
                 0.3558 
                 
                   1.44 
                 
                 0.0037 
                 
                   2.11 
                 
                 0.0078 
                 1364 
                 C08320 
                 HMDB02003 
               
               
                 pro-pro-pro 
                 1.05 
                 0.3704 
                 
                   1.48 
                 
                 0.022 
                 1.38 
                 0.2308 
                 40654 
               
               
                 adenosine 5′- 
                 0.49 
                 0.3724 
                 1.11 
                 0.5931 
                 
                   0.65 
                 
                 0.0064 
                 3108 
                 C00008 
                 HMDB01341 
               
               
                 diphosphate (ADP) 
               
               
                 10-heptadecenoate 
                 0.77 
                 0.3737 
                 
                   2.02 
                 
                 0.0002 
                 
                   1.56 
                 
                 0.0003 
                 33971 
               
               
                 (17:1n7) 
               
               
                 3-methylhistidine 
                 0.92 
                 0.3757 
                 
                   1.71 
                 
                 0.0261 
                 
                   1.91 
                 
                 0.0487 
                 15677 
                 C01152 
                 HMDB00479 
               
               
                 cytidine 
                 0.74 
                 0.3849 
                 1.03 
                 0.7617 
                 
                   0.78 
                 
                 0.0656 
                 514 
                 C00475 
                 HMDB00089 
               
               
                 N1-methyladenosine 
                 1.31 
                 0.3878 
                 
                   1.3 
                 
                 0.0074 
                 
                   1.54 
                 
                 0.0039 
                 15650 
                 C02494 
                 HMDB03331 
               
               
                 2- 
                 1.17 
                 0.3882 
                 
                   1.52 
                 
                 0.0021 
                 
                   1.84 
                 
                 0.0245 
                 35253 
               
               
                 palmitoylglycerophosphocholine 
               
               
                 15-methylpalmitate 
                 0.83 
                 0.3891 
                 
                   1.51 
                 
                 0.0053 
                 
                   1.26 
                 
                 0.0526 
                 38768 
               
               
                 (isobar with 2- 
               
               
                 methylpalmitate) 
               
               
                 myristate (14:0) 
                 1.16 
                 0.3989 
                 
                   1.28 
                 
                 0.0007 
                 
                   1.42 
                 
                 3.65E−05 
                 1365 
                 C06424 
                 HMDB00806 
               
               
                 flavin adenine 
                 0.78 
                 0.4026 
                 
                   1.56 
                 
                 0.02 
                 1.19 
                 0.1501 
                 2134 
                 C00016 
                 HMDB01248 
               
               
                 dinucleotide (FAD) 
               
               
                 phenol sulfate 
                 1.43 
                 0.4062 
                 1.89 
                 0.2514 
                 
                   2.75 
                 
                 0.0015 
                 32553 
                 C02180 
               
               
                 4-acetamidobutanoate 
                 1.53 
                 0.4072 
                 1.15 
                 0.1937 
                 
                   1.56 
                 
                 0.0381 
                 1558 
                 C02946 
                 HMDB03681 
               
               
                 alanylmethionine 
                 1.01 
                 0.4138 
                 
                   1.37 
                 
                 0.0093 
                 
                   1.39 
                 
                 0.0099 
                 37065 
               
               
                 oleoylcarnitine 
                 0.82 
                 0.4167 
                 
                   1.52 
                 
                 0.06 
                 0.98 
                 0.445 
                 35160 
                   
                 HMDB05065 
               
               
                 imidazole lactate 
                 0.65 
                 0.421 
                 
                   2.04 
                 
                 0.0987 
                 
                   1.51 
                 
                 0.0899 
                 15716 
                 C05568 
                 HMDB02320 
               
               
                 Isobar: ribulose 5- 
                 1.47 
                 0.4238 
                 
                   0.84 
                 
                 0.0391 
                 1.12 
                 0.4419 
                 37288 
               
               
                 phosphate, xylulose 5- 
               
               
                 phosphate 
               
               
                 erythritol 
                 1.33 
                 0.426 
                 
                   1.42 
                 
                 0.0521 
                 
                   1.84 
                 
                 0.0013 
                 20699 
                 C00503 
                 HMDB02994 
               
               
                 2- 
                 1.2 
                 0.4274 
                 1.06 
                 0.6101 
                 
                   1.4 
                 
                 0.0359 
                 38077 
               
               
                 arachidonoylglycerophosphoinositol 
               
               
                 N-acetylneuraminate 
                 1.6 
                 0.4294 
                 
                   2.45 
                 
                 0.0006 
                 
                   2.91 
                 
                 0.0004 
                 1592 
                 C00270 
                 HMDB00230 
               
               
                 trigonelline (N′- 
                 2.14 
                 0.4298 
                 0.97 
                 0.5024 
                 
                   1.78 
                 
                 0.089 
                 32401 
                   
                 HMDB00875 
               
               
                 methylnicotinate) 
               
               
                 2- 
                 0.85 
                 0.4352 
                 
                   1.86 
                 
                 0.0017 
                 
                   1.63 
                 
                 0.0102 
                 35884 
               
               
                 eicosatrienoylglycerophosphocholine 
               
               
                 beta-alanine 
                 1.3 
                 0.4393 
                 
                   1.46 
                 
                 0.0114 
                 
                   1.81 
                 
                 0.0018 
                 55 
                 C00099 
                 HMDB00056 
               
               
                 2- 
                 1.31 
                 0.451 
                 
                   1.72 
                 
                 0.0014 
                 
                   2.1 
                 
                 0.0171 
                 34871 
               
               
                 palmitoleoylglycerophosphoethanolamine 
               
               
                 alanylphenylalanine 
                 1.6 
                 0.4517 
                 
                   1.21 
                 
                 0.0197 
                 
                   1.55 
                 
                 0.0056 
                 38679 
               
               
                 leucylasparagine 
                 0.76 
                 0.4523 
                 
                   1.33 
                 
                 0.088 
                 1.09 
                 0.1373 
                 40052 
               
               
                 gluconate 
                 0.58 
                 0.4532 
                 
                   0.32 
                 
                 0.0002 
                 
                   0.45 
                 
                 8.80E−06 
                 587 
                 C00257 
                 HMDB00625 
               
               
                 glycylphenylalanine 
                 0.84 
                 0.4546 
                 
                   1.41 
                 
                 0.0374 
                 
                   1.13 
                 
                 0.0941 
                 33954 
               
               
                 2-methylbutyroylcarnitine 
                 1.11 
                 0.4583 
                 
                   2.07 
                 
                 0.0526 
                 
                   2.04 
                 
                 0.0077 
                 35431 
                   
                 HMDB00378 
               
               
                 choline phosphate 
                 0.86 
                 0.4604 
                 1.18 
                 0.8829 
                 
                   0.88 
                 
                 0.0502 
                 34396 
               
               
                 glucose 
                 0.93 
                 0.4641 
                 
                   0.48 
                 
                 0.0019 
                 
                   0.48 
                 
                 3.14E−05 
                 20488 
                 C00031 
                 HMDB00122 
               
               
                 aspartyltryptophan 
                 0.71 
                 0.4643 
                 
                   1.63 
                 
                 0.0085 
                 
                   1.27 
                 
                 0.0103 
                 41481 
               
               
                 phenylalanylalanine 
                 0.76 
                 0.466 
                 
                   1.47 
                 
                 0.099 
                 1.05 
                 0.6334 
                 41374 
               
               
                 5-aminovalerate 
                 2.21 
                 0.4763 
                 1.36 
                 0.3667 
                 
                   1.96 
                 
                 0.029 
                 18319 
                 C00431 
                 HMDB03355 
               
               
                 fructose 
                 1.37 
                 0.4813 
                 1.64 
                 0.1072 
                 
                   2.45 
                 
                 0.0513 
                 577 
                 C00095 
                 HMDB00660 
               
               
                 pentadecanoate (15:0) 
                 1 
                 0.4886 
                 
                   1.18 
                 
                 0.0733 
                 1.21 
                 0.1124 
                 1361 
                 C16537 
                 HMDB00826 
               
               
                 1-methylurate 
                 1.49 
                 0.4915 
                 1.29 
                 0.2405 
                 
                   1.69 
                 
                 0.0343 
                 34395 
                   
                 HMDB03099 
               
               
                 10-nonadecenoate 
                 1.17 
                 0.4939 
                 
                   1.59 
                 
                 0.003 
                 
                   1.83 
                 
                 4.80E−05 
                 33972 
               
               
                 (19:1n9) 
               
               
                 imidazole propionate 
                 2.5 
                 0.4967 
                 
                   1.83 
                 
                 0.0054 
                 
                   2.72 
                 
                 0.0803 
                 40730 
                   
                 HMDB02271 
               
               
                 N2-methylguanosine 
                 1.09 
                 0.5055 
                 
                   1.68 
                 
                 2.47E−05 
                 
                   1.82 
                 
                 2.34E−05 
                 35133 
                   
                 HMDB05862 
               
               
                 VGAHAGEYGAEALER 
                 1.24 
                 0.506 
                 
                   0.23 
                 
                 5.76E−05 
                 
                   0.28 
                 
                 6.78E−05 
                 41219 
               
               
                 (SEQ ID NO: 2) 
               
               
                 sphingosine 
                 2.57 
                 0.5155 
                 
                   1.2 
                 
                 0.0013 
                 
                   2.35 
                 
                 0.0055 
                 17747 
                 C00319 
                 HMDB00252 
               
               
                 tyrosylglutamine 
                 1.34 
                 0.5183 
                 
                   1.48 
                 
                 0.0071 
                 
                   1.59 
                 
                 0.0394 
                 41459 
               
               
                 ornithine 
                 1.27 
                 0.5334 
                 
                   1.45 
                 
                 0.0158 
                 
                   1.55 
                 
                 0.0212 
                 1493 
                 C00077 
                 HMDB03374 
               
               
                 6-phosphogluconate 
                 1.16 
                 0.5364 
                 
                   0.35 
                 
                 2.65E−05 
                 
                   0.37 
                 
                 6.63E−06 
                 15442 
                 C00345 
                 HMDB01316 
               
               
                 3-methyl-2-oxovalerate 
                 0.91 
                 0.5413 
                 1.18 
                 0.9763 
                 
                   0.8 
                 
                 0.0333 
                 15676 
                 C00671 
                 HMDB03736 
               
               
                 prolylproline 
                 1.14 
                 0.5437 
                 
                   1.28 
                 
                 0.0111 
                 
                   1.35 
                 
                 0.0012 
                 40731 
               
               
                 palmitoleate (16:1n7) 
                 1.1 
                 0.5448 
                 
                   1.69 
                 
                 0.0007 
                 
                   1.83 
                 
                 3.16E−06 
                 33447 
                 C08362 
                 HMDB03229 
               
               
                 1-palmitoylglycerol (1- 
                 0.9 
                 0.545 
                 
                   2.07 
                 
                 9.38E−06 
                 
                   1.91 
                 
                 3.81E−05 
                 21127 
               
               
                 monopalmitin) 
               
               
                 guanosine 
                 1.4 
                 0.5542 
                 
                   0.7 
                 
                 0.0178 
                 
                   0.86 
                 
                 0.0521 
                 1573 
                 C00387 
                 HMDB00133 
               
               
                 stearoylcarnitine 
                 1.35 
                 0.5607 
                 
                   1.69 
                 
                 0.0023 
                 
                   2.01 
                 
                 0.0573 
                 34409 
                   
                 HMDB00848 
               
               
                 aspartylvaline 
                 1.32 
                 0.5646 
                 
                   1.89 
                 
                 0.0012 
                 
                   1.75 
                 
                 0.0046 
                 41373 
               
               
                 riboflavin (Vitamin B2) 
                 1.2 
                 0.5664 
                 
                   1.28 
                 
                 0.093 
                 
                   1.46 
                 
                 0.0043 
                 1827 
                 C00255 
                 HMDB00244 
               
               
                 phenylacetylglutamine 
                 2.23 
                 0.5724 
                 0.7 
                 0.6412 
                 
                   1.6 
                 
                 0.0852 
                 35126 
                 C05597 
                 HMDB06344 
               
               
                 1- 
                 0.84 
                 0.5738 
                 
                   2.02 
                 
                 0.0072 
                 
                   1.86 
                 
                 3.29E−05 
                 35628 
                   
                 HMDB11506 
               
               
                 oleoylglycerophosphoethanolamine 
               
               
                 S-methylcysteine 
                 0.81 
                 0.5819 
                 
                   1.44 
                 
                 0.0371 
                 1.11 
                 0.4491 
                 40262 
                   
                 HMDB02108 
               
               
                 caprylate (8:0) 
                 1.06 
                 0.5915 
                 1.08 
                 0.2217 
                 
                   1.28 
                 
                 0.0323 
                 32492 
                 C06423 
                 HMDB00482 
               
               
                 1- 
                 1.07 
                 0.5976 
                 
                   1.65 
                 
                 0.0431 
                 
                   1.69 
                 
                 0.0004 
                 35631 
                   
                 HMDB11503 
               
               
                 palmitoylglycerophosphoethanolamine 
               
               
                 prolylglycine 
                 1.03 
                 0.5991 
                 
                   1.23 
                 
                 0.0162 
                 
                   1.43 
                 
                 0.0124 
                 40703 
               
               
                 putrescine 
                 0.88 
                 0.6241 
                 
                   1.61 
                 
                 0.0059 
                 
                   1.23 
                 
                 0.0163 
                 1408 
                 C00134 
                 HMDB01414 
               
               
                 lactate 
                 1.01 
                 0.6253 
                 
                   1.24 
                 
                 0.034 
                 
                   1.17 
                 
                 0.0937 
                 527 
                 C00186 
                 HMDB00190 
               
               
                 pyroglutamine 
                 0.69 
                 0.6267 
                 
                   1.8 
                 
                 0.0425 
                 
                   1.43 
                 
                 0.0417 
                 32672 
               
               
                 stearidonate (18:4n3) 
                 0.5 
                 0.6281 
                 
                   2.76 
                 
                 0.0066 
                 
                   1.53 
                 
                 0.0105 
                 33969 
                 C16300 
                 HMDB06547 
               
               
                 2- 
                 1.4 
                 0.6282 
                 
                   1.75 
                 
                 0.0019 
                 
                   2.42 
                 
                 0.0011 
                 35681 
               
               
                 myristoylglycerophosphocholine 
               
               
                 1-methylhistamine 
                 0.93 
                 0.6288 
                 
                   1.69 
                 
                 0.061 
                 1.19 
                 0.1978 
                 32441 
                 C05127 
                 HMDB00898 
               
               
                 methionylthreonine 
                 1.11 
                 0.6352 
                 
                   0.5 
                 
                 0.0038 
                 
                   0.55 
                 
                 0.0095 
                 40679 
               
               
                 2- 
                 1.65 
                 0.6366 
                 
                   1.67 
                 
                 0.0001 
                 
                   2.22 
                 
                 0.0099 
                 35819 
               
               
                 palmitoleoylglycerophosphocholine 
               
               
                 adenylosuccinate 
                 1.18 
                 0.6373 
                 
                   1.41 
                 
                 0.0303 
                 1.64 
                 0.44 
                 18360 
                 C03794 
                 HMDB00536 
               
               
                 N-acetylgalactosamine 
                 2.03 
                 0.6402 
                 
                   2.11 
                 
                 0.0422 
                 
                   4.49 
                 
                 0.0003 
                 2766 
                 C01074 
                 HMDB00835 
               
               
                 N-acetyltryptophan 
                 0.06 
                 0.6466 
                 0.33 
                 0.1296 
                 
                   0.08 
                 
                 0.0499 
                 33959 
                 C03137 
               
               
                 adenosine 3′- 
                 1.67 
                 0.6584 
                 
                   1.37 
                 
                 0.0191 
                 
                   1.86 
                 
                 0.0043 
                 35142 
                 C01367 
                 HMDB03540 
               
               
                 monophosphate (3′- 
               
               
                 AMP) 
               
               
                 inositol 1-phosphate 
                 0.91 
                 0.6626 
                 0.86 
                 0.2269 
                 
                   0.83 
                 
                 0.0639 
                 1481 
                   
                 HMDB00213 
               
               
                 (I1P) 
               
               
                 uridine-2′,3′-cyclic 
                 0.95 
                 0.6677 
                 
                   1.36 
                 
                 0.0264 
                 
                   1.25 
                 
                 0.0349 
                 37137 
                 C02355 
                 HMDB11640 
               
               
                 monophosphate 
               
               
                 glucosamine 
                 1.34 
                 0.6692 
                 1.26 
                 0.487 
                 
                   1.79 
                 
                 0.0753 
                 18534 
                 C00329 
                 HMDB01514 
               
               
                 glucuronate 
                 2.09 
                 0.6736 
                 
                   1.48 
                 
                 0.0837 
                 
                   2.66 
                 
                 0.0077 
                 15443 
                 C00191 
                 HMDB00127 
               
               
                 N-acetyl-aspartyl- 
                 0.79 
                 0.6752 
                 
                   0.46 
                 
                 0.0177 
                 
                   0.47 
                 
                 0.0794 
                 35665 
                 C12270 
                 HMDB01067 
               
               
                 glutamate (NAAG) 
               
               
                 3-indoxyl sulfate 
                 1.75 
                 0.6784 
                 1.1 
                 0.2329 
                 
                   1.78 
                 
                 0.0354 
                 27672 
                   
                 HMDB00682 
               
               
                 2- 
                 0.97 
                 0.6785 
                 
                   1.7 
                 
                 0.0644 
                 
                   1.69 
                 
                 0.0075 
                 37948 
               
               
                 oleoylglycerophosphoserine 
               
               
                 phenylalanylaspartate 
                 1.18 
                 0.6827 
                 
                   1.2 
                 
                 0.0383 
                 
                   1.23 
                 
                 0.0206 
                 41419 
               
               
                 methionylvaline 
                 0.97 
                 0.6828 
                 
                   2.04 
                 
                 9.59E−05 
                 
                   1.6 
                 
                 0.0011 
                 40677 
               
               
                 ribitol 
                 0.81 
                 0.6833 
                 
                   2.2 
                 
                 0.0017 
                 
                   1.78 
                 
                 0.0222 
                 15772 
                 C00474 
                 HMDB00508 
               
               
                 mannose 
                 0.63 
                 0.6854 
                 
                   0.89 
                 
                 0.0329 
                 
                   0.86 
                 
                 0.0088 
                 584 
                 C00159 
                 HMDB00169 
               
               
                 myristoleate (14:1n5) 
                 0.96 
                 0.6895 
                 
                   1.38 
                 
                 0.0297 
                 
                   1.36 
                 
                 0.0002 
                 32418 
                 C08322 
                 HMDB02000 
               
               
                 alpha- 
                 1.45 
                 0.6939 
                 
                   2.52 
                 
                 0.0332 
                 
                   2.69 
                 
                 0.0019 
                 22132 
                 C03264 
                 HMDB00746 
               
               
                 hydroxyisocaproate 
               
               
                 caprate (10:0) 
                 0.98 
                 0.6955 
                 
                   1.2 
                 
                 0.002 
                 
                   1.19 
                 
                 0.0016 
                 1642 
                 C01571 
                 HMDB00511 
               
               
                 2- 
                 1.12 
                 0.6985 
                 0.66 
                 0.3253 
                 
                   0.86 
                 
                 0.0949 
                 35883 
               
               
                 docosahexaenoylglycerophosphocholine 
               
               
                 butyrylcarnitine 
                 1.2 
                 0.7012 
                 1.29 
                 0.2636 
                 
                   1.51 
                 
                 0.0363 
                 32412 
               
               
                 isoleucine 
                 1.04 
                 0.7107 
                 1.1 
                 0.1797 
                 
                   1.16 
                 
                 0.0502 
                 1125 
                 C00407 
                 HMDB00172 
               
               
                 serylleucine 
                 0.88 
                 0.7315 
                 
                   1.73 
                 
                 0.021 
                 
                   1.34 
                 
                 0.0483 
                 40066 
               
               
                 conjugated linoleate 
                 1.22 
                 0.7353 
                 1.22 
                 0.4409 
                 
                   1.45 
                 
                 0.079 
                 27404 
                 C04056 
                 HMDB03797 
               
               
                 (18:2n7; 9Z,11E) 
               
               
                 valerylcarnitine 
                 0.58 
                 0.7382 
                 
                   2.94 
                 
                 0.0227 
                 
                   1.71 
                 
                 0.002 
                 34406 
                   
                 HMDB13128 
               
               
                 aspartate-glutamate 
                 0.87 
                 0.7427 
                 
                   1.58 
                 
                 0.0186 
                 
                   1.58 
                 
                 0.0025 
                 37461 
               
               
                 xylitol 
                 0.92 
                 0.7464 
                 1.9 
                 0.151 
                 
                   1.47 
                 
                 0.0832 
                 4966 
                 C00379 
                 HMDB00568 
               
               
                 glycylglycine 
                 0.97 
                 0.7521 
                 
                   1.65 
                 
                 0.0029 
                 
                   1.55 
                 
                 0.0057 
                 21029 
                 C02037 
                 HMDB11733 
               
               
                 glycylisoleucine 
                 0.99 
                 0.762 
                 
                   2.03 
                 
                 0.0003 
                 
                   1.71 
                 
                 0.0016 
                 36659 
               
               
                 3-methoxytyrosine 
                 1.01 
                 0.7668 
                 
                   1.54 
                 
                 0.0061 
                 
                   1.44 
                 
                 0.0008 
                 12017 
                   
                 HMDB01434 
               
               
                 Ac-Ser-Asp-Lys-Pro-OH 
                 1.02 
                 0.775 
                 
                   1.99 
                 
                 0.0003 
                 
                   2.09 
                 
                 0.0006 
                 40707 
               
               
                 (SEQ ID NO: 1) 
               
               
                 leucylleucine 
                 1.74 
                 0.8142 
                 
                   1.26 
                 
                 0.0572 
                 
                   1.64 
                 
                 0.0175 
                 36756 
                 C11332 
               
               
                 phenylalanylleucine 
                 1.5 
                 0.8204 
                 
                   1.06 
                 
                 0.0084 
                 
                   1.23 
                 
                 0.0472 
                 40192 
               
               
                 methionylleucine 
                 1.41 
                 0.823 
                 
                   1.05 
                 
                 0.0397 
                 
                   1.26 
                 
                 0.0612 
                 40023 
               
               
                 threonylphenylalanine 
                 1.51 
                 0.8303 
                 
                   1.31 
                 
                 0.0028 
                 
                   1.61 
                 
                 0.0047 
                 31530 
               
               
                 glycylserine 
                 1.12 
                 0.834 
                 1.03 
                 0.2302 
                 
                   1.18 
                 
                 0.0967 
                 33940 
                   
                 HMDB00678 
               
               
                 pelargonate (9:0) 
                 1.05 
                 0.8373 
                 
                   1.19 
                 
                 0.0011 
                 
                   1.22 
                 
                 0.0004 
                 12035 
                 C01601 
                 HMDB00847 
               
               
                 3-phosphoserine 
                 0.81 
                 0.8409 
                 
                   0.41 
                 
                 0.0077 
                 
                   0.3 
                 
                 0.0002 
                 543 
                 C01005 
                 HMDB00272 
               
               
                 serylphenyalanine 
                 1.24 
                 0.8433 
                 
                   1.48 
                 
                 0.0044 
                 
                   1.53 
                 
                 0.0104 
                 40054 
               
               
                 threonylleucine 
                 1.12 
                 0.8447 
                 1.43 
                 0.134 
                 
                   1.39 
                 
                 0.0615 
                 40051 
               
               
                 margarate (17:0) 
                 1.01 
                 0.8449 
                 
                   1.6 
                 
                 0.0023 
                 
                   1.46 
                 
                 0.004 
                 1121 
                   
                 HMDB02259 
               
               
                 1- 
                 1.15 
                 0.849 
                 
                   2.74 
                 
                 0.0018 
                 
                   2.66 
                 
                 0.0008 
                 35305 
               
               
                 palmitoylglycerophosphoinositol 
               
               
                 leucylglutamate 
                 1.16 
                 0.8585 
                 
                   1.34 
                 
                 0.0386 
                 
                   1.37 
                 
                 0.0441 
                 40021 
               
               
                 arachidate (20:0) 
                 1.19 
                 0.8783 
                 
                   1.52 
                 
                 0.0009 
                 
                   1.68 
                 
                 0.0007 
                 1118 
                 C06425 
                 HMDB02212 
               
               
                 orotate 
                 1.17 
                 0.8788 
                 
                   1.75 
                 
                 0.0578 
                 
                   1.92 
                 
                 0.0316 
                 1505 
                 C00295 
                 HMDB00226 
               
               
                 tetradecanedioate 
                 1.08 
                 0.8975 
                 
                   0.63 
                 
                 0.0199 
                 
                   0.69 
                 
                 0.0195 
                 35669 
                   
                 HMDB00872 
               
               
                 glycylproline 
                 1.08 
                 0.9022 
                 
                   1.22 
                 
                 0.0457 
                 
                   1.27 
                 
                 0.0103 
                 22171 
                   
                 HMDB00721 
               
               
                 alanylleucine 
                 1.42 
                 0.9049 
                 
                   1.26 
                 
                 0.0623 
                 
                   1.45 
                 
                 0.0113 
                 37093 
               
               
                 ethanolamine 
                 0.88 
                 0.9065 
                 
                   2.24 
                 
                 0.0055 
                 
                   1.88 
                 
                 0.0172 
                 1497 
                 C00189 
                 HMDB00149 
               
               
                 3-aminoisobutyrate 
                 0.68 
                 0.9179 
                 
                   3.79 
                 
                 0.0063 
                 
                   2.77 
                 
                 0.0015 
                 1566 
                 C05145 
                 HMDB03911 
               
               
                 fucose 
                 1.06 
                 0.9198 
                 
                   2 
                 
                 0.039 
                 
                   2.04 
                 
                 0.0055 
                 15821 
                 C00382 
                 HMDB00174 
               
               
                 4-guanidinobutanoate 
                 1.01 
                 0.9202 
                 
                   1.77 
                 
                 0.04 
                 
                   1.51 
                 
                 0.0562 
                 15681 
                 C01035 
                 HMDB03464 
               
               
                 glycyltyrosine 
                 1.07 
                 0.9309 
                 
                   0.67 
                 
                 0.0566 
                 0.82 
                 0.3039 
                 33958 
               
               
                 valylleucine 
                 1.34 
                 0.9314 
                 
                   1.57 
                 
                 0.0749 
                 
                   1.75 
                 
                 0.0338 
                 39994 
               
               
                 N-acetylglucosamine 
                 1.41 
                 0.9342 
                 
                   2.59 
                 
                 0.0262 
                 
                   3.68 
                 
                 0.0011 
                 15096 
                 C00140 
                 HMDB00215 
               
               
                 1- 
                 1.02 
                 0.9409 
                 
                   1.32 
                 
                 0.096 
                 
                   1.48 
                 
                 0.0036 
                 34416 
                   
                 HMDB11130 
               
               
                 stearoylglycerophosphoethanolamine 
               
               
                 sorbitol 
                 0.95 
                 0.942 
                 1.46 
                 0.445 
                 
                   1.62 
                 
                 0.0692 
                 15053 
                 C00794 
                 HMDB00247 
               
               
                 3-phosphoglycerate 
                 2.1 
                 0.9427 
                 
                   0.4 
                 
                 0.003 
                 
                   0.57 
                 
                 0.0054 
                 40264 
                 C00597 
                 HMDB00807 
               
               
                 leucylalanine 
                 1.19 
                 0.9444 
                 
                   1.38 
                 
                 0.0546 
                 
                   1.46 
                 
                 0.0311 
                 40010 
               
               
                 1- 
                 0.95 
                 0.9474 
                 
                   2.19 
                 
                 0.0031 
                 
                   2.09 
                 
                 8.43E−06 
                 39270 
               
               
                 palmitoylplasmenylethanolamine 
               
               
                 cysteine sulfinic acid 
                 0.97 
                 0.9496 
                 
                   0.51 
                 
                 0.0195 
                 
                   0.54 
                 
                 0.0188 
                 37443 
                 C00606 
                 HMDB00996 
               
               
                 palmitoylcarnitine 
                 1.29 
                 0.9498 
                 
                   1.38 
                 
                 0.0421 
                 1.57 
                 0.1144 
                 22189 
               
               
                 propionylcarnitine 
                 0.93 
                 0.9519 
                 
                   1.73 
                 
                 0.0041 
                 
                   1.52 
                 
                 0.0004 
                 32452 
                 C03017 
                 HMDB00824 
               
               
                 alanylproline 
                 0.92 
                 0.9538 
                 
                   1.31 
                 
                 0.0153 
                 
                   1.14 
                 
                 0.0104 
                 37083 
               
               
                 gamma- 
                 1.01 
                 0.9711 
                 
                   0.74 
                 
                 0.0288 
                 
                   0.75 
                 
                 0.0158 
                 37539 
               
               
                 glutamylmethionine 
               
               
                 sphinganine 
                 1.61 
                 0.9746 
                 
                   2.24 
                 
                 2.63E−05 
                 
                   2.81 
                 
                 0.0014 
                 17769 
                 C00836 
                 HMD800269 
               
               
                 aspartyllysine 
                 1.1 
                 0.9932 
                 1.06 
                 0.2536 
                 
                   1.24 
                 
                 0.0879 
                 40682 
               
               
                 N1-methylguanosine 
                 1.08 
                 0.9989 
                 
                   1.86 
                 
                 3.71E−05 
                 
                   1.89 
                 
                 5.37E−06 
                 31609 
                   
                 HMDB01563 
               
               
                 2′-deoxyguanosine 
                 0.91 
                 0.9992 
                 
                   1.4 
                 
                 0.0761 
                 
                   1.33 
                 
                 0.0258 
                 1411 
                 C00330 
                 HMDB00085 
               
               
                 glycerophosphorylcholine 
                 
                   0.49 
                 
                 0.0119 
                 
                   5.67 
                 
                 8.11E−06 
                 
                   1.98 
                 
                 0.0035 
                 15990 
                 C00670 
                 HMDB00086 
               
               
                 (GPC) 
               
               
                 thymine 
                 0.97 
                 0.6081 
                 
                   2.87 
                 
                 3.51E−05 
                 
                   2.34 
                 
                 0.0002 
                 604 
                 C00178 
                 HMDB00262 
               
               
                 phenyllactate (PLA) 
                 1.62 
                 0.2874 
                 
                   1.48 
                 
                 5.77E−05 
                 
                   2.24 
                 
                 1.28E−05 
                 22130 
                 C05607 
                 HMDB00779 
               
               
                 S-adenosylmethionine 
                 
                   0.39 
                 
                 0.0083 
                 
                   4.96 
                 
                 6.70E−05 
                 
                   1.88 
                 
                 0.0051 
                 15915 
               
               
                 (SAM) 
               
               
                 succinate 
                 
                   0.56 
                 
                 0.0978 
                 
                   4.24 
                 
                 0.0001 
                 
                   2.25 
                 
                 0.0312 
                 1437 
                 C00042 
                 HMDB00254 
               
               
                 uracil 
                 0.97 
                 0.7512 
                 
                   1.93 
                 
                 0.0003 
                 
                   1.87 
                 
                 0.0003 
                 605 
                 C00106 
                 HMDB00300 
               
               
                 xanthine 
                 0.93 
                 0.3561 
                 
                   1.75 
                 
                 0.0005 
                 
                   1.48 
                 
                 0.0329 
                 3147 
                 C00385 
                 HMDB00292 
               
               
                 3-(4- 
                 
                   1.42 
                 
                 0.0534 
                 
                   1.44 
                 
                 0.0007 
                 
                   2 
                 
                 1.80E−07 
                 32197 
                 C03672 
                 HMDB00755 
               
               
                 hydroxyphenyl)lactate 
               
               
                 oleate (18:1n9) 
                 0.99 
                 0.8839 
                 
                   1.7 
                 
                 0.001 
                 
                   1.7 
                 
                 0.0004 
                 1359 
                 C00712 
                 HMDB00207 
               
               
                 proline 
                 1.08 
                 0.6856 
                 
                   1.32 
                 
                 0.0014 
                 
                   1.43 
                 
                 0.0003 
                 1898 
                 C00148 
                 HMDB00162 
               
               
                 threonine 
                 0.8 
                 0.039 
                 
                   1.33 
                 
                 0.0023 
                 
                   1.18 
                 
                 0.0389 
                 1284 
                 C00188 
                 HMDB00167 
               
               
                 taurine 
                 1.45 
                 0.1226 
                 
                   0.63 
                 
                 0.0034 
                 
                   0.81 
                 
                 0.07 
                 2125 
                 C00245 
                 HMDB00251 
               
               
                 creatine 
                 0.72 
                 0.152 
                 
                   0.65 
                 
                 0.0073 
                 
                   0.57 
                 
                 4.05E−05 
                 27718 
                 C00300 
                 HMDB00064 
               
               
                 alanine 
                 0.86 
                 0.3709 
                 
                   1.4 
                 
                 0.0074 
                 
                   1.25 
                 
                 0.0389 
                 1126 
                 C00041 
                 HMDB00161 
               
               
                 tryptophan 
                 1 
                 0.5997 
                 
                   1.32 
                 
                 0.009 
                 
                   1.32 
                 
                 0.0033 
                 54 
                 C00078 
                 HMDB00929 
               
               
                 hypoxanthine 
                 
                   0.8 
                 
                 0.1661 
                 
                   1.34 
                 
                 0.0151 
                 1.12 
                 0.3363 
                 3127 
                 C00262 
                 HMDB00157 
               
               
                 histidine 
                 1.07 
                 0.5483 
                 
                   1.21 
                 
                 0.0168 
                 
                   1.31 
                 
                 0.0016 
                 59 
                 C00135 
                 HMDB00177 
               
               
                 homoserine 
                 0.74 
                 0.4123 
                 
                   2.26 
                 
                 0.0201 
                 
                   1.69 
                 
                 0.0821 
                 23642 
                 C00263, 
                 HMDB00719 
               
               
                   
                   
                   
                   
                   
                   
                   
                   
                 C02926 
               
               
                 histamine 
                 1.26 
                 0.5813 
                 
                   0.66 
                 
                 0.0211 
                 
                   0.73 
                 
                 0.0446 
                 1574 
                 C00388 
                 HMDB00870 
               
               
                 cytidine 5′- 
                 0.94 
                 0.7367 
                 
                   1.63 
                 
                 0.0236 
                 1.29 
                 0.1305 
                 2372 
                 C00055 
                 HMDB00095 
               
               
                 monophosphate (5′- 
               
               
                 CMP) 
               
               
                 carnitine 
                 0.85 
                 0.2334 
                 
                   1.26 
                 
                 0.0257 
                 1.05 
                 0.8208 
                 15500 
               
               
                 laurate (12:0) 
                 1.05 
                 0.5526 
                 
                   1.14 
                 
                 0.0272 
                 
                   1.16 
                 
                 0.006 
                 1645 
                 C02679 
                 HMDB00638 
               
               
                 asparagine 
                 0.78 
                 0.2082 
                 
                   1.46 
                 
                 0.0284 
                 1.25 
                 0.1303 
                 11398 
                 C00152 
                 HMDB00168 
               
               
                 valine 
                 1.05 
                 0.6324 
                 
                   1.17 
                 
                 0.0335 
                 
                   1.21 
                 
                 0.0156 
                 1649 
                 C00183 
                 HMDB00883 
               
               
                 guanine 
                 
                   2.03 
                 
                 0.0245 
                 
                   0.91 
                 
                 0.0436 
                 
                   2.15 
                 
                 0.0243 
                 32352 
                 C00242 
                 HMDB00132 
               
               
                 spermine 
                 
                   8.42 
                 
                 0.0134 
                 
                   0.49 
                 
                 0.0444 
                 2.59 
                 0.4402 
                 603 
                 C00750 
                 HMDB01256 
               
               
                 2-aminobutyrate 
                 0.76 
                 0.3869 
                 
                   1.58 
                 
                 0.0462 
                 1.15 
                 0.5752 
                 1577 
                 C02261 
                 HMDB00650 
               
               
                 cortisol 
                 
                   1.3 
                 
                 0.031 
                 
                   0.85 
                 
                 0.0577 
                 0.97 
                 0.9206 
                 1712 
                 C00735 
                 HMDB00063 
               
               
                 glutamine 
                 
                   0.7 
                 
                 0.0043 
                 
                   1.21 
                 
                 0.0719 
                 1 
                 0.4768 
                 53 
                 C00064 
                 HMDB00641 
               
               
                 palmitate (16:0) 
                 
                   1.26 
                 
                 0.0897 
                 
                   1.09 
                 
                 0.0798 
                 
                   1.29 
                 
                 0.0013 
                 1336 
                 C00249 
                 HMDB00220 
               
               
                 kynurenine 
                 
                   2.17 
                 
                 0.0154 
                 
                   1.43 
                 
                 0.0799 
                 
                   2.5 
                 
                 2.98E−05 
                 15140 
                 C00328 
                 HMDB00684 
               
               
                 leucine 
                 0.98 
                 0.8158 
                 
                   1.16 
                 
                 0.0826 
                 
                   1.17 
                 
                 0.0517 
                 60 
                 C00123 
                 HMDB00687 
               
               
                 aspartate 
                 0.89 
                 0.4494 
                 
                   1.3 
                 
                 0.094 
                 1.2 
                 0.1402 
                 15996 
                 C00049 
                 HMDB00191 
               
               
                 serine 
                 0.95 
                 0.6493 
                 1.12 
                 0.1562 
                 1.11 
                 0.3047 
                 1648 
                 C00065 
                 HMDB03406 
               
               
                 citrulline 
                 1.26 
                 0.295 
                 1.24 
                 0.1813 
                 
                   1.68 
                 
                 0.0002 
                 2132 
                 C00327 
                 HMDB00904 
               
               
                 adenosine 
                 0.63 
                 0.1128 
                 0.73 
                 0.2946 
                 
                   0.5 
                 
                 0.0011 
                 555 
                 C00212 
                 HMDB00050 
               
               
                 trans-urocanate 
                 
                   1.73 
                 
                 0.0891 
                 0.92 
                 0.3308 
                 1 
                 0.7151 
                 607 
                 C00785 
                 HMDB00301 
               
               
                 homocysteine 
                 
                   2.22 
                 
                 0.0205 
                 0.82 
                 0.373 
                 
                   1.82 
                 
                 0.0012 
                 40266 
                 C00155 
                 HMDB00742 
               
               
                 betaine 
                 
                   1.43 
                 
                 0.0263 
                 1.06 
                 0.3738 
                 
                   1.37 
                 
                 0.0023 
                 3141 
                   
                 HMDB00043 
               
               
                 indolelactate 
                 
                   2.53 
                 
                 0.0014 
                 1.06 
                 0.6124 
                 
                   1.86 
                 
                 0.0043 
                 18349 
                 C02043 
                 HMDB00671 
               
               
                 kynurenate 
                 
                   2.67 
                 
                 0.0577 
                 0.95 
                 0.6436 
                 
                   1.86 
                 
                 0.0861 
                 1417 
                 C01717 
                 HMDB00715 
               
               
                 pipecolate 
                 
                   2.32 
                 
                 0.0246 
                 0.64 
                 0.6463 
                 
                   1.47 
                 
                 0.0247 
                 1444 
                 C00408 
                 HMDB00070 
               
               
                 beta-hydroxyisovalerate 
                 1.46 
                 0.1361 
                 1.07 
                 0.7015 
                 
                   1.38 
                 
                 0.0517 
                 12129 
                   
                 HMDB00754 
               
               
                 adenine 
                 0.53 
                 0.291 
                 1.4 
                 0.9174 
                 
                   0.74 
                 
                 0.0577 
                 554 
                 C00147 
                 HMDB00034 
               
               
                   
               
            
           
         
       
     
     The biomarkers were used to create a statistical model to classify subjects. The biomarkers in Table 9 were evaluated using Random Forest analysis to classify samples as low stage bladder cancer or high stage bladder cancer. The Random Forest results show that the samples were classified with 83% prediction accuracy. The confusion matrix presented in Table 10 shows the number of subjects predicted for each classification and the actual in each group (BCA High or BCA Low). The “Out-of-Bag” (OOB) Error rate gives an estimate of how accurately new observations can be predicted using the Random Forest model (e.g., whether a sample is from a subject with Low stage bladder cancer or a subject with High stage bladder cancer). The OOB error was approximately 17%, and the model estimated that, when used on a new set of subjects, the identity of High stage bladder cancer subjects could be predicted 84% of the time and Low stage bladder cancer subjects could be predicted correctly 82% of the time and as presented in Table 10. 
     
       
         
           
               
             
               
                 TABLE 10 
               
             
            
               
                   
               
               
                 Results of Random Forest, Low Stage BCA vs. High Stage BCA 
               
            
           
           
               
               
               
               
            
               
                   
                   
                 Predicted Group 
                 class. 
               
            
           
           
               
               
               
               
            
               
                   
                 BCA High 
                 BCA Low 
                 Error 
               
               
                   
                   
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Actual 
                 BCA High 
                 26 
                 5 
                 0.1613 
               
               
                   
                 Group 
                 BCA Low 
                 3 
                 14 
                 0.1765 
               
               
                   
                   
               
            
           
         
       
     
     Based on the OOB Error rate of 17%, the Random Forest model that was created predicted whether a sample was from an individual with RCC with about 83% accuracy by measuring the levels of the biomarkers in samples from the subject. Exemplary biomarkers for distinguishing the groups are palmitoyl ethanolamide, palmitoyl sphingomyelin, thromboxane B2, bilirubin (Z,Z), adrenate (22:4n6), C-glycosyltryptophan, methyl-alpha-glucopyranoside, methylphosphate, 3-hydroxydecanoate, 3-hydroxyoctanoate, 4-hydroxyphenylpyruvate, N-acetylthreonine, 1-arachidonoylglycerophosphoinositol (20:4), 5 6-dihydrothymine, 2-hydroxypalmitate, coenzyme A, N-acetylserine, nicotinamide adenine dinucleotide (NAD+), docosatrienoate (22:3n3), glutathione reduced (GSH), prostaglandin A2, glutamine, glutamate gamma-methyl ester, docosapentaenoate (n6 DPA 22:5n6), glycochenodeoxycholate, hexanoylcarnitine, arachidonate (20:4n6), pro-hydroxy-pro, docosahexaenoate (DHA 22:6n3), and laurylcarnitine. 
     The Random Forest results demonstrated that by using the biomarkers, RCC subjects were distinguished from normal subjects with 84% sensitivity, 82% specificity, 90% PPV, and 74% NPV. 
     Example 7 
     Biomarker Panels and Mathematical Models for Identifying Bladder Cancer 
     In another example, a panel of five exemplary biomarkers was selected to identify bladder cancer, the panel being selected from biomarkers identified in Tables 1 and/or 5. The biomarkers identified were present at levels that differed between BCA and each of the comparison groups of individuals (i.e., BCA compared to Normal, HX, Hematuria, RCC, and PCA). For example, lactate, palmitoyl sphingomyelin, choline phosphate, succinate and adenosine were significant biomarkers for distinguishing subjects with bladder cancer from normal, HX, hematuria, RCC and PCA subjects. All of the biomarker compounds used in these analyses were statistically significant (p&lt;0.05). Table 11 includes, for each listed biomarker, the biochemical name of the biomarker, the fold change of the biomarker in: 1) bladder cancer subjects compared to normal subjects (BCA/NORM), 2) bladder cancer subjects compared to subjects with a history of bladder cancer (BCA/HX), 3) bladder cancer subjects compared to subjects with Hematuria (BCA/HEM), 4) bladder cancer subjects compared to kidney cancer subjects (BCA/RCC), 5) bladder cancer subjects compared to prostate cancer subjects (BCA/PCA), and the p-value determined in the statistical analysis of the data concerning the biomarkers for BCA compared to Normal. 
     
       
         
           
               
             
               
                 TABLE 11 
               
             
            
               
                   
               
               
                 Biomarkers to Identify Bladder Cancer 
               
            
           
           
               
               
               
            
               
                   
                 Fold Change 
                 BCA/ 
               
            
           
           
               
               
               
               
               
               
               
            
               
                   
                 BCA/ 
                 BCA/ 
                 BCA/ 
                 BCA/ 
                 BCA/ 
                 NORM 
               
               
                 Biochemical 
                 NORM 
                 HX 
                 HEM 
                 RCC 
                 PCA 
                 p-value 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
            
               
                 choline phosphate 
                 6.35 
                 4.99 
                 5.85 
                 3.22 
                 7.7 
                 3.81E−05 
               
               
                 palmitoyl 
                 10.24 
                 8.03 
                 8 
                 3.79 
                 8.74 
                 3.32E−06 
               
               
                 sphingomyelin 
               
               
                 lactate 
                 3.14 
                 3.13 
                 1.41 
                 2.55 
                 3.41 
                 1.56E−11 
               
               
                 succinate 
                 0.65 
                 0.51 
                 0.6 
                 0.58 
                 0.66 
                 5.09E−05 
               
               
                 adenosine 
                 0.73 
                 0.82 
                 0.7 
                 0.68 
                 0.79 
                 9.13E−05 
               
               
                   
               
            
           
         
       
     
     Next, the biomarkers in Table 11 were used in a mathematical model based on ridge logistic regression analysis. The ridge regression method builds statistical models that are useful to evaluate the biomarker compounds that are associated with disease and to evaluate biomarker compounds useful to classify individuals as, for example, having BCA or not having BCA, having BCA or being Normal (not having cancer), having BCA or having hematuria, having BCA or having a history of BCA. Predictive performance (for example, the ability of the mathematical model to correctly classify samples as cancer or non-cancer) of the five biomarkers identified in Table 11 was determined using ridge logistic regression analysis. Table 12 shows the AUC for the five biomarkers for bladder cancer as compared to the permuted AUC (that is, the AUC for the null hypothesis). The mean of the permuted AUC represents the expected value of the AUC that would be obtained by chance alone. For all comparisons, the five biomarkers listed in Table 11 predicted bladder cancer with higher accuracy than achieved with five metabolites that do not have a true association for the comparison (i.e., five biomarkers selected at random). A graphical illustration of the resulting Receiver Operator Characteristic (ROC) Curve is presented in  FIG. 4 . 
     
       
         
           
               
             
               
                 TABLE 12 
               
             
            
               
                   
               
               
                 Predictive Performance of Biomarkers for Bladder Cancer 
               
            
           
           
               
               
               
               
            
               
                   
                   
                 Permuted Mean 
                 5 Biomaker 
               
               
                   
                 Comparisons 
                 AUC Ridge 
                 Ridge 
               
               
                   
                   
               
            
           
           
               
               
               
               
            
               
                   
                 BCA vs HX 
                 0.711 
                 0.821 
               
               
                   
                 BCA vs NORM 
                 0.724 
                 0.823 
               
               
                   
                 BCA vs All other groups 
                 0.674 
                 0.799 
               
               
                   
                 BCA vs HEM 
                 0.75 
                 0.791 
               
               
                   
                   
               
            
           
         
       
     
     In another example, a panel of seven exemplary biomarkers was selected to identify bladder cancer, the panel being selected from biomarkers identified in Tables 1 and/or 5. The biomarkers identified were present at levels that differed between BCA and each of the comparison groups of individuals (i.e., BCA compared to Normal, HX, Hematuria,) as illustrated in Table 13. For example, 1,2 propanediol, adipate, anserine, 3-hydroxybutyrate (BHBA), pyridoxate, acetylcarnitine and 2-hydroxybutyrate (AHB) were significant (p&lt;0.05) biomarkers for distinguishing subjects with bladder cancer from normal, HX, and hematuria subjects. All of the biomarker compounds used in these analyses were statistically significant (p&lt;0.05). Table 13 includes, for each listed biomarker, the biochemical name of the biomarker, the fold change of the biomarker in: 1) bladder cancer subjects compared to normal subjects (BCA/NORM), 2) bladder cancer subjects compared to subjects with a history of bladder cancer (BCA/HX), and 3) bladder cancer subjects compared to subjects with Hematuria (BCA/HEM). 
     
       
         
           
               
             
               
                 TABLE 13 
               
             
            
               
                   
               
               
                 Biomarkers to distinguish BCA from 
               
               
                 non-cancer (Hematuria, HX, Normal) 
               
            
           
           
               
               
               
               
            
               
                 Biomarker 
                 BCA/Normal 
                 BCA/HX 
                 BCA/Hematuria 
               
               
                   
               
            
           
           
               
               
               
               
            
               
                 1,2-propanediol 
                 5.37 
                 3.11 
                 5.95 
               
               
                 Adipate 
                 4.53 
                 5.02 
                 4 
               
               
                 Anserine 
                 0.23 
                 0.14 
                 0.23 
               
               
                 3-hydroxybutyrate (BHBA) 
                 18.95 
                 24.27 
                 19.58 
               
               
                 Pyridoxate 
                 0.33 
                 0.3 
                 0.5 
               
               
                 Acetylcarnitine 
                 2.39 
                 2.63 
                 2.45 
               
               
                 2-hydroxybutyrate (AHB) 
                 2.96 
                 3.29 
                 2.04 
               
               
                   
               
            
           
         
       
     
     Next, the biomarkers in Table 13 were used in a mathematical model based on ridge logistic regression analysis. The ridge regression method builds statistical models that are useful to evaluate the biomarker compounds that are associated with disease and to evaluate biomarker compounds useful to classify individuals as for example, having BCA or being Normal (not having cancer), having BCA or having hematuria, having BCA or having a history of BCA. Predictive performance (for example, the ability of the mathematical model to correctly classify samples as cancer or non-cancer) of the seven biomarkers identified in Table 13 was determined using ridge logistic regression analysis. The AUC for the seven biomarkers for bladder cancer was 0.849 [95% CI, 0.794-0.905]. A graphical illustration of the ROC Curve is presented in  FIG. 5 . For all comparisons, the seven biomarkers listed in Table 13 predicted bladder cancer with higher accuracy than achieved with five metabolites that do not have a true association for the comparison. 
     In another example, a panel of exemplary biomarkers was selected to identify bladder cancer subjects and non-bladder cancer subjects using the subset of five biomarkers listed in Table 11 and seven biomarkers listed in Table 13 in combination with one or more exemplary biomarkers identified in Tables 1 and/or 5. In this example, kynurenine was selected as the one exemplary biomarker from Tables 1 and/or 5 (kynurenine is in both Tables 1 and 5). Thus, the resulting panel of markers comprised the 13 listed metabolites: lactate, palmitoyl sphingomyelin, choline phosphate, succinate, adenosine, 1,2propanediol, adipate, anserine, 3-hydroyxbutyrate, pyridoxate, acetyl carnitine, AHB and kynurenine. 
     Next, the 13 biomarkers were used in a mathematical model based on ridge logistic regression analysis. The Ridge regression method was used to build statistical models useful to evaluate the biomarker compounds that are associated with disease and to evaluate biomarker compounds useful to classify individuals as for example, having BCA or not having cancer (i.e., Normal, hematuria, or history of BCA). Predictive performance of various combinations of the 13 biomarkers comprised of two or more biomarkers selected from the group comprised of lactate, palmitoyl sphingomyelin, choline phosphate, succinate, adenosine, 1,2propanediol, adipate, anserine, 3-hydroyxbutyrate, pyridoxate, acetyl carnitine, AHB or kynurenine was determined using ridge logistic regression analysis. The AUCs for the panels of biomarkers for bladder cancer ranged from 0.85 for a two biomarker model to 0.9 for models comprised of ten to twelve biomarkers. A graphical illustration of the AUC obtained for the panels with the Ridge Models is presented in  FIG. 6 . 
     In another example, a panel of eleven exemplary biomarkers was selected to identify bladder cancer or hematuria in a subject. In this example, the biomarker panel comprised tyramine, palmitoyl sphingomyelin, choline phosphate, adenosine, 1,2 propanediol, adipate, BHBA, acetyl carnitine, AHB, xanthurenate and succinate. Predictive performance (that is, the ability of the mathematical model to correctly classify samples as cancer or hematuria) of the eleven biomarkers was determined using ridge logistic regression analysis. The AUC for the eleven biomarkers was 0.886 [95% CI, 0.831-0.941]. A graphical illustration of the ROC Curve is presented in  FIG. 7 . For all comparisons, the eleven biomarkers predicted bladder cancer with higher accuracy than achieved with metabolites that do not have a true association for the comparison. 
     Next, the 11 biomarkers in were used in a mathematical model based on ridge logistic regression analysis. The ridge regression method builds statistical models useful to evaluate the biomarker compounds that are associated with disease and to evaluate biomarker compounds useful to classify individuals as for example, having BCA or hematuria. Predictive performance (that is, the ability of the mathematical model to correctly classify samples as cancer or hematuria) of various combinations of the eleven biomarkers comprised of two or more biomarkers selected from the group comprised of tyramine, palmitoyl sphingomyelin, choline phosphate, adenosine, 1,2 propanediol, adipate, BHBA, acetyl carnitine, AHB, xanthurenate and succinate was determined using ridge logistic regression analysis. The AUCs for the panels of biomarkers for bladder cancer ranged from 0.82 for a two biomarker model to 0.886 for models comprised of eight to twelve biomarkers. A graphical illustration of the AUC obtained for the panels with the Ridge Models is presented in  FIG. 8 . 
     Example 8 
     Algorithm to Monitor Bladder Cancer Progression/Regression 
     Using the biomarkers for bladder cancer, an algorithm can be developed to monitor bladder cancer progression/regression in subjects. The algorithm, based on a panel of metabolite biomarkers from Tables 1, 5, 7, 9, 11 and/or 13, when used on a new set of patients, would assess and monitor a patient&#39;s progression/regression of bladder cancer. Using the results of this biomarker algorithm, a medical oncologist can assess the risk-benefit of surgery (e.g., transurethral resection, radical cystectomy, or segmental cystectomy), drug treatment or a watchful waiting approach. 
     The biomarker algorithm can be used to monitor the levels of a panel of biomarkers for bladder cancer identified in Tables 1, 5, 7, 9, 11 and/or 13. 
     Example 9 
     Identification of Drug Targets and Drug Screens Using Said Targets 
     To identify drug targets for bladder cancer, 10 control urine samples collected from subjects that did not have bladder cancer, and 10 urine samples from subjects having bladder cancer (urothelial transitional cell carcinoma) were analyzed to determine the levels of metabolites in the samples, then the results were statistically analyzed using univariate T-tests (i.e., Welch&#39;s test) to determine those metabolites that were differentially present in the two groups, and then the metabolic pathways of the differentially present metabolites were analyzed in a biological context to identify associated metabolites, enzymes and/or proteins. 
     The metabolites, enzymes and/or proteins associated with the differentially present metabolites represent drug targets for bladder cancer. The levels of metabolites that are aberrant (higher or lower) in bladder cancer subjects relative to control (non-BCA) subjects can be modulated to bring them into the normal range, which can be therapeutic. Such metabolites or enzymes involved in the associated metabolic pathways and proteins involved in the transport within and between cells can provide targets for therapeutic agents. 
     For example, bladder cancer is associated with altered levels of biochemical intermediates in the tricarboxylic acid cycle (TCA) as well as biochemicals associated with all of the major ATP-producing pathways. In this example, subjects with bladder cancer were found to have altered TCA cycle intermediates, with a pronounced effect on isocitrate and its immediate downstream metabolites. Isocitrate levels were found to be statistically significantly higher in the urine of bladder cancer subjects. Thus, an agent that can modulate the levels of isocitrate in urine may be a therapeutic agent. For example, said agent may modulate isocitrate urine levels by decreasing the biosynthesis of isocitrate. Bladder cancer also had pronounced effects on TCA cycle intermediates between citrate and succinyl-coA, especially isocitrate, α-ketoglutarate and the two TCA α-ketoglutarate-derived metabolites 2-hydroxyglutarate and glutamate. These results are graphically depicted in  FIG. 9 , which illustrates the TCA cycle. The levels of the biochemicals that were measured in urine collected from control individuals and from bladder cancer patients are presented in box plots. 
     In addition to the TCA cycle, urine metabolite profiles from bladder cancer cases suggested that all major ATP-producing pathways were altered in bladder cancer. An increased lactate/pyruvate ratio suggested that there is a Warburg-like utilization of glucose in bladder cancer patients. The increased ketone body production suggested that there is increased fatty acid β-oxidation in these patients. Finally, the decreased abundance of branched chain acyl carnitines and acyl glycines indicated that this pathway is differentially engaged in bladder cancer patients. Metabolites that report on the activity of glycolysis, branched chain amino acid catabolism and fatty acid oxidation were all altered in bladder cancer cases compared to the control population. The branched chain acyl carnitines were shown as surrogates for the branched chain acyl CoA compounds. These changes are illustrated by the box plots presented in  FIG. 10 . 
     The identification of biomarkers for bladder cancer can be useful for screening therapeutic compounds. For example, isocitrate, α-ketoglutarate or any biomarker(s) aberrant in subjects having bladder cancer as identified in Tables 1, 5, 7, 9, 11, and 13 can be used in a variety of drug screening techniques. 
     One exemplary method of drug screening utilizes eukaryotic or prokaryotic host cells such as bladder cancer cells. In this prophetic example, cells are plated in 96-well plates. Test wells are incubated in the presence of test compounds from the NIH Clinical Collection Library (available from BioFocus DPI) at a final concentration of 50 μM. Negative control wells receive no addition or are incubated with a vehicle compound (e.g., DMSO) at a concentration equivalent to that present in some of the test compound solutions. After incubation for 24 hours, test compound solutions are removed and metabolites are extracted from cells, and isocitrate levels are measured as described in the General Methods section. Agents that lower the level of isocitrate in the cell are considered therapeutic. 
     While the invention has been described in detail and with reference to specific embodiments thereof, it will be apparent to one skilled in the art that various changes and modifications can be made without departing from the spirit and scope of the invention.