bluuebunny/crossref_metadata_2025 huggingface dataset:
Preparation of Urine Samples for Trace Metal Determination: A Study with Aluminium Analysis by Inductively Coupled Plasma Optical Emission Spectrometry
Authors: Burden T J , Whitehead M W , Thompson R P H , Powell J J
Published: February 2014

Urinary analysis of trace metals forms a significant role in clinical chemistry, but the optimal preparation and analysis of urine samples has not been investigated. Human urine is generally supersaturated with dissolved solids. Therefore, samples often precipitate following collection. X-ray microanalysis showed that this precipitate was predominantly rich in calcium and phosphorus but could include some trace metals from urine, potentially lowering their concentrations in solution. Hence, the precipitate must be fully redissolved for accurate analysis of trace metals in urine. Methods are emphasized for the best collection and preparation of urine samples for subsequent trace metal analysis; in this work inductively coupled plasma optical emission spectrometry (ICPOES) was used for the analysis of aluminium. For optimal accuracy, peak profiles were collected over 396.147 nm-396.157 nm. Urinary aluminium levels were investigated from 10 healthy volunteers and concentrations were obtained using either aqueous, pooled or individual urine-based standard curves. Since urine has a highly variable matrix, individual sample-based standards, which are unique to that particular sample, gave the most accurate results. However, where sample size is small or sample numbers are unfeasibly large, pooled sample-based standards give good approximations to within 15% and, with appropriate validation, other elements as internal standards could also be used for approximations. Aqueous standards should be avoided. Spike-recovery experiments confirmed these data since individual sample based standards showed optimal recovery [99.3 (4.4)%], while pooled sample-based standards were a close proxy [101.6 (9.2)%] but aqueous standards were inappropriate [137.4 (12.8)%]. Postprandial urinary aluminium levels of the 10 volunteers were [7.2 (3.7)μg/L] after analysis using individual sample-based standard curves.
Guidelines for the Interpretation of Analytical Toxicology Results and Unit of Measurement Conversion Factors
Authors: Flanagan R J
Published: February 2014

A data compilation has been published on the World Wide Web [ http://www.leeds.ac.uk/acb/annals ] which aims to assist in the interpretation of analytical toxicology results in individual patients. ‘Therapeutic’ or ‘normal’ plasma/whole blood/urine concentrations, as well as the concentrations associated with serious toxicity (if known), for some 700 analytes of toxicological interest are given together with (when possible) relative atomic or formula masses, and mass/amount and amount/mass concentration conversion factors. This paper gives background and supplementary information which should be borne in mind when providing interpretation or using the Web table. Supplementary tables listing (i) common therapeutic drug monitoring assays and guidelines for interpretation of results, (ii) emergency toxicology analyses which may influence active treatment, (iii) blood carboxyhaemoglobin saturation and clinical features of toxicity, (iv) information important when interpreting the results of toxicology investigations, (v) factors which may affect interpretation of analytical toxicology results, and (vi) conversion factors for volatile compounds are included.
A Simple Method for Carbohydrate-Deficient Transferrin Measurement in Patients with Alcohol Abuse and Hepato-Gastrointestinal Diseases
Authors: Cotton F , Adler M , Dumon J , Boeynaems J-M , Gulbis B
Published: February 2014

Carbohydrate-deficient transferrin (CDT) is known to be increased in alcohol abuse. Several methods were developed for its measurement (e.g. isoelectric focusing with Western blotting or immunofixation, anion-exchange chromatography followed by immunoassays). We describe a greatly simplified isoelectric focusing technique which does not require immunofixation. CDT results obtained with this method were compared to other biological markers of alcohol abuse, i.e. mean corpuscular volume (MCV), aspartate aminotransferase (ASAT) and γ-glutamyltransferase (GGT), in 55 patients distributed in three groups (i.e. healthy control subjects, control patients suffering from various hepato-gastrointestinal diseases and alcohol abusing patients). Sensitivity and specificity were 33 89%, 61–57%, 89–49% and 83–100% for MCV, ASAT, GGT and CDT, respectively. We conclude that our method is highly suitable for routine clinical use.
Determination of Free Fatty Acids in Human Bile by High-Performance Liquid Chromatography
Authors: Hori Yasushi , Nakamura Kazutoshi , Yamamoto Masaharu , Shimada Kenji , Nakadaira Hiroto , Shibuya Norio , Endoh Kazuo , Ogoshi Kazuei
Published: February 2014

We developed a high-performance liquid chromatography (HPLC) method for free fatty acids (FFAs) analysis in bile. In this method, FFAs were extracted from bile in a single step using an Isolute™ ODS cartridge, derivatized with 9-anthryldiazomethane (ADAM). ADAM was chosen because of its high reactivity with carboxylic acid at room temperature. Then, HPLC was used for separating and quantifying FFAs. This method proved to be simple and time-saving. The mean recovery of FFA added to human gallbladder bile was 97.6%, and the detection limit was 100–250 pg. Using this method, we determined FFA concentrations in the gallbladder bile of 11 gallstone patients. The mean concentration of total FFA was 0.61 (SD = 0.41) mmol/L, and there was wide variation in the individual FFAs.
Clinical and Biochemical Determinants of Plasma Lipid Peroxide Levels in Type 2 Diabetes
Authors: Lawrence J R , Campbell G R , Barrington H , Malcolm E A , Brennan G , Wiles D H , Paterson J R
Published: February 2014

The enhanced risk and increased severity of atheroma in diabetes is well recognized but, as yet, incompletely explained. A cross-sectional study of vascular disease risk factors in a group of type 2 diabetic patients from South West Scotland has revealed an association between glycaemic control, assessed by HbA1 level and plasma lipid peroxides measured by a specific high-performance liquid chromatography method. Duration of diabetes appeared to be a subsidiary contributor to lipid peroxidation.

We suggest this evidence supports the importance of glycaemic control in modulating glyco-oxidative mechanisms probably crucial to production of diabetic complications. Atherosclerosis prevention in diabetes may hinge on exemplary simultaneous control of both hyperglycaemia and hyperlipidaemia.
Apo A-I and Apo E Concentrations in Cerebrospinal Fluids of Patients with Acute Meningitis
Authors: Song Hua , Seishima Mitsuru , Saito Kuniaki , Maeda Satoshi , Takemura Masao , Noma Akio , Kondo Akira , Manabe Mitsuhisa , Urakami Katsuya , Nakashima Kenji
Published: February 2014

It has been demonstrated that apolipoproteins found in cerebrospinal fluid (CSF) play an important role in lipid metabolism in the central nervous system (CNS). Previously we reported that CSF apo A-I levels increased with the severity of neurological damage in poliovirus-infected macaques. In the present study, apo A-I was quantitatively analysed in CSF from patients with or without neurological diseases. In controls, CSF apo A-I level was significantly higher in males; 3·83 (0·40) mg/L, mean (SEM) ( n = 19) compared with females, 2·42 (0·26) mg/L ( n = 23, P < 0·05). CSF apo A-I concentrations in patients with acute meningitis increased at the active stage, 7·74 (1·78) mg/L ( n = 10), but returned to basal concentrations at the convalescent stage 2·72 (0·38) mg/L ( n = 10), while the CSF apo A-I level in patients with other neurological diseases remained in the same range as in controls. By contrast, CSF apo E was consistently elevated at either stage of acute meningitis. Furthermore, it was found that the levels of CSF apo A-I, but not of apo E, correlated positively with CSF albumin concentrations. These findings suggest that the CSF apo A-I and apo E have different origins and may play different roles in the lipoprotein metabolism in CNS.
A UK Survey of Laboratory-Based Gastrointestinal Investigations
Authors: Duncan A , Hill P G
Published: February 2014

A survey of hospital laboratory services has demonstrated marked deficiencies in the performance of gastrointestinal function tests. The repertoire of gastrointestinal investigations available varies widely between laboratories and, in general, analyses are performed infrequently. Most laboratories do not perform internal quality control, and inter-laboratory reproducibility of some analytes is very poor. A wide variety of protocols and reference ranges are in use, many of which are unevaluated. Some analytical methods and protocols in current use are outdated, with published improvements not being applied.
A Solid Phase Extraction Method for Determination of Nicotine in Serum and Urine by Isotope Dilution Gas Chromatography/Mass Spectrometry with Selected Ion Monitoring
Authors: Baskin Leland B , Anderson Rodger W , Charlson Joel R , Hurt Richard D , Lawson George M
Published: February 2014

A rapid method for measuring nicotine concentration in serum and urine is described. Deuterated nicotine is used as an internal standard. Nicotine and deuterated nicotine are extracted using a copolymeric-bonded phase silica column. The extract is analysed by gas chromatography coupled with mass spectrometry (GC/MS) operating in selected ion monitoring mode. The method has a lower limit of detection of approximately 2 μg/L and is linear to at least 2000 μg/L. Within-run percentage coefficients of variation (% CV) are < 4 in both assays over a nicotine concentration range of 10–2000 μg/L. Between-run %CV in the serum assay are 5.4, 5.2, 4.8 and 5.9, respectively, at nicotine concentrations of 10, 15, 25, and 50 μg/L. Between-run %CV in the urine assay are 5.9, 4.5, 2.7 and 5.2, respectively, at nicotine concentrations of 100, 250, 500, and 2000 μg/L. The absolute recovery of nicotine is 61±+6% (mean±SD) over the range of 10–250 μg/L. The assay has been used to measure serum nicotine concentrations and 24-h urinary excretion of nicotine to monitor the extent of replacement in subjects receiving transdermal nicotine therapy for smoking cessation.
Micro-Extraction of Chlordiazepoxide and its Primary Metabolites, Desmethylchlordiazepoxide and Demoxepam, from Plasma and Their Measurement by Liquid Chromatography
Authors: Garretty Deborah J , Wolff Kim , Hay Alastair W M
Published: February 2014

We have developed a micro-extraction procedure for the analysis of chlordiazepoxide and its two unique metabolites, demoxepam and desmethylchlordiazepoxide, in plasma, using liquid chromatography. The method is both reliable and sensitive for the quantitation of low plasma concentrations of these three compounds. The extraction procedure allows rapid sample processing, which, together with the small sample volume (100 μL), makes it ideal for routine sample handling. The limit of detection for the three compounds ranged from 0.075 to 0.125 mg/L and recovery of the three different benzodiazepines ranged from 87 to 94%. Within- and between-assay coefficients of variation ranged from 3.1–4.5% and from 4.7 to 7.6%, respectively.
The Incorporation of Lamotrigine into the Routine Automated Sequential Trace Enrichment of Dialysates Assay of Anticonvulsants
Authors: Higgins G , Hughes A , Dutton J , Roberts N B
Published: February 2014

We have extended automated sequential trace enrichment of dialysates and high performance liquid chromatography to include the simultaneous assay of lamotrigine with other anticonvulsant drugs: Phenobarbitone, carbamazepine and phenytoin. The procedure is reliable and precise (between-batch coefficients of variation < 2.8%) with no interference from other commonly prescribed drugs. Patients on lamotrigine presented with serum concentrations up to 67 μmol/L with no adverse effects, suggesting that the therapeutic range may need re-appraisal.
Multianalyte Serum Analysis Using Mid-Infrared Spectroscopy
Authors: Shaw R Anthony , Kotowich Steven , Leroux Michael , Mantsch Henry H
Published: February 2014

This study assesses the potential for using mid-infrared (mid-IR) spectroscopy of dried serum films as the basis for the simultaneous quantitation of eight serum analytes: Total protein, albumin, triglycerides, cholesterol, glucose, urea, creatinine and uric acid. Infrared transmission spectra were acquired for 300 serum samples, each analysed independently using accepted reference clinical chemical methods. Quantitation methods were based upon the infrared spectra and reference analyses for 200 specimens, and the models validated using the remaining 100 samples. Standard errors in the IR-predicted analyte levels (Sy/x) were 2.8 g/L (total protein), 2.2 g/L (albumin), 0.23 mmol/L (triglycerides), 0.28 mmol/L (cholesterol), 0.41 mmol/L (glucose) and 1.1 mmol/L for urea, with correlation coefficients (IR vs reference analyses) of 0.95 or better. The IR method emerged to be less suited for creatinine (S y/ x = μmol/L) and uric acid (S y/x = 140 μmol/L) due to the relatively low concentrations typical of these analytes.
Determination of Serum Physiological Concentration of Methylmalonic Acid by Gas Chromatography-Mass Spectrometry with Selected Ion Monitoring
Authors: Rifai Nader , Hagen Thilo , Bradley Loetta , Sakamoto Masayuki
Published: February 2014

We developed a sensitive assay for the rapid determination of serum methylmalonic acid concentration using capillary gas chromatography-mass spectrometry (GC/MS) with selected ion monitoring and a simple solid-phase extraction. The assay was linear up to 10 000 nmol/L and had a detection limit < 50 nmol/L, average recovery of 98% and between-day coefficient of variation at concentrations of 570 and 2206 nmol/L of 7.7% and 5.4%, respectively ( n = 25). Comparison with another validated GC/MS method using sera with a wide range of methylmalonic acid concentrations (94-2020 nmol/L) revealed a slope and intercept of 0.97 and 17 nmol/L, respectively ( n = 38). Methylmalonic acid concentrations determined by this assay in a group of apparently healthy individuals ranged from 64–331 nmol/L ( n = 81). We conclude that the method is ideally suited for the determination of methylmalonic acid at physiological concentrations in both clinical and research laboratories.
Distribution and Expression of Adrenomedullin in Human Gastrointestinal Tissue
Authors: Kitani Michitaka , Sakata Junichiro , Asada Yujiro , Kitamura Kazuo , Eto Tanenao
Published: February 2014

Adrenomedullin (AM) is a biologically active peptide recently isolated from phaeochromocytoma. We report here the distribution and characterization of immunoreactive AM and gene expression of AM in human gastrointestinal tissue. Using a sensitive radioimmunoassay system for the peptide, immunoreactive human AM was detected in the stomach, duodenum, jejunum, ileum and colon. The AM concentration of these tissues was about 0.4–0.8 pmol/g wet tissue. Reverse phase and gel filtration high-performance liquid chromatographies showed that most of the immunoreactive AM in stomach and jejunum was identical to authentic human AM. By northern blot analysis, human AM mRNA was found to be expressed ubiquitously in the human gastrointestinal tissues. Furthermore, an immunohistochemical study revealed that immunoreactive AM cells were present in the gastrointestinal glands. These results suggest that AM may play some role as a gastrointestinal hormone.
Clinical Usefulness of Bone Alkaline Phosphatase in Osteoporosis
Authors: Kyd P A , De Vooght K , Kerkhoff F , Thomas E , Fairney A
Published: February 2014

We have evaluated two commercial assays for serum bone specific alkaline phosphatase (bALP) as a marker of bone turnover in a group of 35 postmenopausal women with osteoporosis during their first year of treatment with the anti-resorptive drug, alendronate. The immunoradiometric assay (bALP-I) measured protein mass, whereas the immunocapture assay (bALP-E) measured activity.

Before treatment, bALP values with both methods were within the postmenopausal reference ranges. Treatment with alendronate produced a decrease in bALP after 3 months, reaching a plateau after 6 months: for bALP-I a mean change of −34%, ( P < 00001), bALP-E, −19% ( P < 0·001), and total ALP, −19% ( P < 00001). The decrease was significant in 53% (bALP-I) and 31% (bALP-E) of patients. The ratio of serum bALP-E/bALP-I in patients was not constant but rose after therapy with alendronate. Neither baseline bALP, nor the per cent change in bALP after 6 months, correlated with bone mineral density (BMD) change after 1 year at either lumbar spine or femoral neck. We conclude that bALP-I appeared to be a more sensitive marker of the suppression of bone turnover by alendronate than did bALP-E, but that neither was useful in the detection of osteoporosis, nor the prediction of individual BMD response to alendronate therapy.
Influence of Anti-Insulin Antibodies on Insulin Immunoassays in the Autoimmune Insulin Syndrome
Authors: Casesnoves Adela , Mauri Montserrat , Dominguez José R , Alfayate Rocío , Picó Antonio M
Published: February 2014

The autoimmune insulin syndrome (AIS) is a rare, benign syndrome characterized by hyperinsulinaemia and hypoglycaemia associated with the presence of autoantibodies to insulin in patients who have not been treated with insulin. We report here the case of a 52-year-old patient with recurrent attacks of severe postprandial hypoglycaemia and we also present the effect of anti-insulin antibodies on insulin immunoassays.

The patient was submitted to the following diagnostic tests: 5-h oral glucose tolerance test (OGTT), a prolonged 72-h fast and an insulin tolerance test (ITT). Serum glucose, total and free insulin, C-peptide, proinsulin, insulin antibodies and other autoantibodies were measured. Insulin concentrations were measured by two methods: a double antibody radioimmunoassay (RIA) and an immunoradiometric assay (IRMA).

Insulin concentration measured by RIA was extremely high in the OGTT and 72-h fast. In contrast, insulin concentrations measured by IRMA were between 120 and 888 pmol/L in the OGTT and between 37 and 133 pmol/L during the 72-h fast. Fasting free-insulin concentrations measured by RIA were between 2224 and 2669 pmol/L, whereas free-insulin concentrations measured by IRMA ranged between 93 and 237 pmol/L. Total insulin concentrations measured by RIA and IRMA were 57615 and 94021 pmol/L, respectively. The C-peptide concentrations were moderately high in the three tests. Serum insulin antibody concentrations were extremely high (62–71%), compared with less than 3% in normal serum samples. In conclusion, the high insulin concentrations measured by RIA were caused by insulin autoantibodies. However, insulin concentrations measured by IRMA were not influenced by them. We conclude that IRMA is the more accurate method for measuring insulin concentrations in such cases.
Carbohydrate-Deficient Glycoprotein Syndromes: Inborn Errors of Protein Glycosylation
Authors: Keir G , Winchester B G , Clayton P
Published: February 2014

The carbohydrate-deficient glycoprotein (CDG) syndromes (CDGS) are a series of autosomal recessive enzyme deficiencies which result in incomplete glycosylation of plasma proteins. CDGS types Ia and Ib have been related to deficiencies of phosphomannomutase and phosphomannose isomerase, respectively, while CDGS type II results from a deficiency of N-acetylglucosaminyltransferase II. Secondary CDG syndromes are associated with galactosaemia and hereditary fructose intolerance. The diagnosis of CDGS is most easily made by studying the glycoforms of suitable marker proteins using either electrophoresis or isoelectric focusing. This paper reviews the structure of the glycan chains of proteins and structural alterations in CDGS. It also outlines analytical techniques which are useful in the laboratory study of protein glycoforms and the diagnosis of CDGS.
Influence of Acetate and Bicarbonate Dialysate on Blood Short- and Long-Chain Acyl Carnitine in Adult Pyelonephritis Patients
Authors: Alhomida A S
Published: February 2014

The effect of acetate and bicarbonate haemodialysis (HD) on the concentrations of erythrocyte, whole-blood and plasma total carnitine (TC), free carnitine (FC), short- (SC) and long-chain acylcarnitine (LC) and acylcarnitine (AC) as well as the ratio of AC to FC was investigated in 30 healthy subjects (15 men and 15 women) and 27 patients (10 men and 17 women) with chronic pyelonephritis (CPN) undergoing chronic HD. Fourteen patients (5 men and 9 women) used acetate HD and the remainder (5 men and 8 women) used bicarbonate HD. The mean predialysis erythrocyte, whole-blood and plasma concentrations of TC, FC, SC, LC and AC as well as the ratio of AC to FC were not significantly different from those in healthy controls ( P > 0·05). However, after acetate or bicarbonate HD, a significant decrease in erythrocyte, whole-blood and plasma concentrations of TC, FC, SC, LC and AC were found, compared with either predialysis or healthy control values ( P < 0·001). Furthermore, the ratio of AC to FC was significantly higher following acetate HD as compared with either acetate or bicarbonate predialysis values ( P < 0·001). The observed variations in response between acetate and bicarbonate HD may be due to enhanced formation of acetyl-coenzyme A and fatty acid synthesis during acetate HD.
Analytical Error of Home Glucose Monitors: A Comparison of 18 Systems
Authors: Johnson Roger N , Baker John R
Published: February 2014

In a quality review of 18 home blood glucose monitors, we measured the imprecision and incidence of significant error of 12 colorimetric and six amperometric systems by using capillary blood specimens from patients attending diabetes clinics. Imprecision at the mean glucose concentration found in the respective blood specimens (about 9 mmol/L) gave coefficients of variation (CV) ranging from 5·2 to 22·8%. Eight monitors including five of amperometric design had a CV of less than 10%. The incidence of significant error (defined as the proportion of specimens differing in value by more than 15% from a reference hexokinase assay of glucose in capillary blood) varied from 6 to 76%. Among the eight monitors identified as being most precise, the majority produced results that differed markedly from the reference assay, underlining the need for a common approach to calibration of home glucose monitors.
Survey of Serum Potassium Reference Measurements
Authors: Van Nuwenborg J E , Thienpont L M , Stöckl D , Davies K W , Smith S C , Palicka V , Friedecký B , Beránek M , Kratochvíla J
Published: February 2014

We compared the quality of reference measurements for serum potassium in four reference laboratories from three different European countries, using a panel of 60 native patients' samples. The reference methods were based on either ion chromatography (one laboratory) or flame atomic emission spectrometry (three laboratories). Performance specifications for serum potassium measurements were defined as a maximum overall coefficient of variation (CV) of 1·5%, a maximum bias of 0·65% and a maximum total error of 3·0%. The overall imprecision for all laboratories was in the range of 0·7 to 1·3%, and was thus below the proposed specification of 1·5%. However, two laboratories reported 12 and 13 quadruplicates with CVs exceeding this limit. The mean bias (expressed as deviation from the overall mean of all laboratories) for all reference laboratories was < 0·65%. In the lower concentration range, however, one laboratory exceeded this limit. No laboratory measured samples with a total error above 3·0%. From these results, it can be concluded that the reference measurements, and, thus, also the reference methodologies, based on ion chromatography and flame atomic emission spectrometry were equivalent, and able to satisfy current analytical specifications for serum potassium measurements.
Serum and Tissue Antioxidant Capacity in Cervical Intraepithelial Neoplasia Investigated Using an Enhanced Chemiluminescent Reaction
Authors: Cope Graham , Thorpe Gary , Holder Roger , Luesley David , Jordan Joseph
Published: February 2014

Depleted antioxidant defence has been implicated in the pathogenesis of cervical neoplasia. We determined the systemic and local antioxidant status of women with this condition. Thirty-four women with varying grades of cervical intraepithelial neoplasia, 25 patients who had been treated successfully with diathermy loop excision, and 56 women who had no evidence of cervical abnormality acted as controls. Total antioxidant capacity of serum and protein-free serum, and of neat and protein-free homogenized cervical punch biopsies were determined using enhanced chemiluminescence. Mean serum antioxidant capacity of patients with current neoplasia and treated patients was not significantly different from that of controls. However, mean antioxidant capacity of homogenized cervical tissue from women with neoplasia was significantly lower than control means ( P < 0·005), while results for treated patients were intermediate between those from diseased and normal samples ( P < 0·05). The enhanced chemiluminescence technique has potential as a suitable method for measuring total antioxidant capacity of cervical tissue, and warrants further investigations using other tissue types. Significant antioxidant depletion in cervical intra-epithelial neoplasia appears to be confined to the local cervical mucosa.
Accuracy of Down's Syndrome Risks Produced in a Prenatal Screening Program
Authors: Spencer Kevin
Published: February 2014

Down's syndrome screening in the second trimester using a variety of combinations of maternal serum markers has become an established part of antenatal care over the past decade.1 The methodology of calculating Down's syndrome risk using statistical models2 based on the Gaussian distribution functions of the various markers used is often considered daunting, and many screening centres rely on specific screening computer software to undertake the relevant calculations. Such software is often a closed system which does not allow the user to check or change any of the parameters within the model. It is clear from a recent UK NEQAS survey of Down's syndrome screening laboratories (Spencer, Ellis, Seth, 1997, unpublished data) that many centres are unsure what method of calculating risk and what population parameters are used in their software, and how appropriate these are for the population being screened. This confusion has led to uncertainty over the reliability of the risk estimates produced in Down's syndrome screening programs.3
Three-Colour, One-Tube Flow Cytometric Method for Enumeration of Cd4+ and Cd8+ Lymphocytes
Authors: Janssen W C M , Hoffmann J J M L
Published: February 2014

We investigated the usefulness of a one-tube, three-colour flow cytometric method for enumerating CD4 + and CD8 + lymphocytes. This method does not use any control antibodies and we compared it to the standard methods using either control and CD14/CD45 antibodies or control antibodies only on 38 blood samples from healthy and human immunodeficiency virus-infected patients. The one-tube method showed good agreement with the other, more complicated methods and is therefore suitable for reliable enumeration of CD4+ and CD8 + T-lymphocytes.
Measurement of Phenylalanine and Tyrosine in Plasma by High-Performance Liquid Chromatography Using the Inherent Fluorescence of Aromatic Amino Acids
Authors: Allen K R , Degg T J , Rushworth P A , Smith M , Henderson M J
Published: February 2014

An isocratic high-performance liquid chromatography (HPLC) method is described using the natural fluorescence of phenylalanine and tyrosine compared with that of an internal standard N-methyl phenylalanine. Plasma precipitated with 6% perchloric acid was separated isocratically using a base-deactivated C18 column with 5% acetonitrile in water as the mobile phase. Fluorescent measurements at an excitation wavelength of 215 nm and emission 283 nm showed only three peaks for tyrosine, phenylalanine and the internal standard eluting within 9 min. Inter-batch coefficients of variation for phenylalanine were 2·9% and 1·8% at levels of 70 and 567 μmol/L, respectively, and 2·9% at a level of 63 μmol/L for tyrosine. The results for phenylalanine for this method showed a small mean positive bias (11 μmol/L) when compared with the target all-method means for UK National External Quality Assessment Scheme samples ( n = 31). The results for tyrosine showed a small positive mean bias (10 μmol/L) when compared with an ion-exchange chromatographic method ( n = 40). This method provides a quick and simple alternative to those using HPLC with pre- or post-column derivatization for monitoring patients with phenylketonuria. It is also less subject to interferences than HPLC methods using ultraviolet detection, particularly for the early eluting tyrosine peak.
A Kinetic Model of Mitochondrial Aspartate Aminotransferase Transmigration in Hepatobiliary Disorders
Authors: Numakami Kiyohiko , Aoki Yoshikazu , Ogawa Zensuke , Itoh Hajime
Published: February 2014

A hypothetical kinetic model of transmigration of mitochondrial aspartate aminotransferase (m-AST) from liver to blood and its elimination from blood was constructed and assessed for prediction by computer simulation. The elimination of m-AST from plasma in healthy rats followed first-order kinetics. Depletion of m-AST from the liver after induction of hepatobiliary disorders in rats with α-naphthylisothiocyanate (ANIT) also followed first-order kinetics. In contrast, the changes in metabolic rate in the liver after ANIT administration, as estimated from the plasma indocyanine green clearance, approximated to a second-order time function. Based on these data, the time-dependent change in plasma m-AST activity after ANIT administration was simulated by computer according to a hypothetical kinetic model. The result of computer simulation of the change in plasma m-AST coincided well with that obtained experimentally, suggesting that continuous measurements of m-AST may be helpful to the clinical diagnosis of liver injury.
Threshold Concentration of Unbound Bilirubin to Induce Neurological Deficits in a Patient with Type I Crigler—Najjar Syndrome
Authors: Ihara Hiroshi , Hashizume Naotaka , Shimizu Norikazu , Aoki Tsugutoshi
Published: February 2014

Based on the clinical course of a 16-year-old boy with type I Crigler-Najjar syndrome, we estimated the threshold concentration of unbound bilirubin, as assayed by the horseradish peroxidase method, that apparently induces toxicity to the brain. Before the age of 15, the patient did not manifest any neurological or behavioural dysfunction despite increased bilirubin in serum. The binding affinity and the binding capacity of the patient's serum albumin for bilirubin determined when he was about 14 years old were 108(mol/L)−1 and 1·01 to 1·04 mol/L, respectively. These values were nearly the same as those of normal controls reported in the literature. The total bilirubin binding capacity was greater than the patient's total bilirubin concentration, showing that his serum albumin was not saturated with bilirubin. The reserve bilirubin binding capacity (RBBC) was estimated to be 158 μmol/L and the unbound bilirubin concentration to be 15·1 nmol/L. Concentration of unbound bilirubin peaked at 21·7 nmol/L at the age of 15 years and 11 months, i.e. 2 months before the onset of difficulties in walking and speaking. At this time, the RBBC was estimated as −64 μmol/L. A peak concentration of total bilirubin, 811 μmol/L, was observed during the period of rapid loss of the ability to walk or speak. At the age of 16 years and 1 month the RBBC decreased to −98 μmol/L and the unbound bilirubin concentration to 18·8 nmol/L. Following phototherapy, the patient's neurological state returned to normal; he could speak and walk normally. At the age of 16 years and 2 months the RBBC returned to 105 μmol/L and unbound bilirubin decreased to 16·6 nmol/L. These results suggest that maintaining the concentration of unbound bilirubin at < 20 nmol/L and the total bilirubin concentration at lower than the binding capacity of serum albumin is important for prevention of neurological deficits in Crigler-Najjar syndrome. The upper limit of unbound bilirubin in such an older patient was nearly the same as that reported for newborns.
Diurnal Variations in Peripheral Insulin Resistance and Plasma Non-Esterified Fatty Acid Concentrations: A Possible Link?
Authors: Morgan L M , Aspostolakou F , Wright J , Gama R
Published: February 2014

Glucose tolerance becomes impaired towards the evening. Increased peripheral insulin resistance may be responsible, at least in part, for this effect. The mechanism for the diurnal variation in insulin sensitivity is undefined. It is, however, possible that variations in non-esterified fatty acids (NEFA) could contribute to this variation because NEFA have been implicated in the pathogenesis of insulin resistance. Therefore, we have investigated insulin sensitivity and plasma NEFA responses to insulin at 0830 h and 2030 h in nine healthy men by measuring arterialized plasma glucose and venous plasma NEFA concentrations during a short insulin tolerance test. The studies were standardized for a period of fasting, pre-test meal and exercise. Insulin sensitivity measured KITT was greater ( P < 0·05) in the morning [(20±7) × 10−3 mmol/L/min] than in the evening [(11·6 ± 2) × 10−3 mmol/L/min]. Fasting NEFA levels were lower ( P < 0·01) in the morning (373 ± 84 μmol/L) than in the evening (913±122 μmol/L). Following insulin, NEFA fell more slowly ( P < 0·01) in the morning (149 ± 26 μmol/L/15 min) than in the evening (491 ± 91 μmol/L/15 min). These results confirm diurnal variations in insulin sensitivity and plasma NEFA concentrations irrespective of feeding and exercise. We speculate that the relatively elevated plasma NEFA levels in the evening are the cause rather than the consequence of increased insulin resistance at this time.
Bloodspot Testosterone Assay Suitable for Study of Neonates and Monitoring of Children with Congenital Adrenal Hyperplasia
Authors: Walker J , Hughes I A , Wood P J
Published: February 2014

A bloodspot assay for testosterone has been developed using an antiserum raised against testosterone-3-carboxymethyloxime-bovine serum albumin (T-3 CMO-BSA) conjugate and 125I-T-3CMO-histamine tracer. The method has a detection limit of 0·4 nmol/L with between-batch precision of less than 15% for testosterone values between 0·9 and 2·0 nmol/L and less than 10% for values greater than 2·0 nmol/L. Recovery of 4 nmol/L testosterone spiked into whole blood samples from 16 women (with unspiked bloodspot testosterone concentrations ranging from 0·4 to 3·7 nmol/L) was 100·9%. Bloodspot testosterone concentration was measured in thirty 6–9 day old infants (19 boys, 11 girls) with an absolute range in boys of < 0·4–2·3 nmol/L and in girls of < 0·4 nmol/L. In preterm infants (12 boys, 12 girls) of varying birth gestation and postnatal age, levels were higher than in term infants, with boys showing a rise with increasing postnatal age. Daytime bloodspot profiles in four children on replacement therapy for congenital adrenal hyperplasia demonstrated consistently low testosterone levels in the presence of moderately elevated early morning 17-hydroxyprogesterone concentrations, thus indicating adequate treatment. The bloodspot testosterone assay can be applied to the study of androgen physiology in preterm neonates as well as for monitoring treatment in disorders such as congenital adrenal hyperplasia.
Lymphocyte Populations in Tuberculous Pleural Effusions
Authors: José M Esther San , Valdés Luis , Saavedra M Jesús , De Vega Jesus M , Alvarez David , Viñuela Juan , Penela Pedro , Valle Jose M , Seoane Rafael
Published: February 2014

Different systemic and local responses to mycobacterial antigens suggest an active compartmentalization of responsive lymphocytes to tubercular antigens. This fact, observed in pleuritic processes, raises doubts about the accuracy of information obtained in the study of cells taken solely from peripheral blood. For this reason we decided to study the concept of compartmentalization in 140 patients suffering from pleural effusions. Patients were classified into six groups according to the aetiology of the effusion: group I, tuberculous, n = 23; group II, paraneoplastic, n = 41; group III, metapneumonic empyematous, n = 5; group IV, transudate, n = 38; group V, miscellaneous exudate, n = 19; group VI, unknown aetiology, n = 14. In each group we studied the lymphocyte population by using flow cytometry with doubly fluorescent monoclonal antibodies: B [expressing human lymphocyte antigen (HLA)-DR on the surface], T (CD3 +), CD4 + and CD8 +, and the subpopulation of activated T lymphocytes (together expressing CD3 and HLA-DR on the surface) (CD3 + DR +). The study of these subpopulations in peripheral blood did not yield valuable results, but the CD3 + DR + population in pleural fluid demonstrated a diagnostic efficiency of 84% [positive predictive value (PPV) 51%, negative predictive value (NPV) 96%] at a cut-off value of 80.4 cells/mm3. The CD3 + DR + pleural fluid/peripheral blood ratio demonstrated an efficiency of 83% (PPV 50%, NPV 96%), and showed a statistically significant difference ( P < 0·02) with regard to all the diagnostic groups, with the exception of the paraneoplastic effusions. The lymphocytic subpopulations study confirms the concept of compartmentalization in tuberculous pleuritis, as shown by the greater number of activated T lymphocytes present in pleural fluid in comparison with peripheral blood in tuberculous pleuritis, a 98% efficiency of adenosine deaminase (ADA) determination in pleural fluid versus a 50% value in peripheral blood, predominance of helper cells (CD4 +) in pleural fluid and suppressor cells (CD8 +) in peripheral blood, a greater CD4 + /CD8 + ratio in pleural fluid than in peripheral blood, and a significant correlation of ADA-CD3 + DR + in pleural fluid, which does not occur in peripheral blood.
Evaluation of Detection Techniques and Laboratory Proficiency in Testing for Drugs of Abuse in Urine: An External Quality Assessment Scheme Using Clinically Realistic Urine Samples
Authors: Wilson J F , Smith B L
Published: February 2014

Techniques for the detection of drugs of abuse were compared using data from 43 urine samples distributed by the United Kingdom National External Quality Assessment Scheme. Drug mixtures simulated clinical specimens and contained drugs at the following median concentrations: amphetamine 1·6 mg/L (six samples), barbiturates 1·7 mg/L (six samples), benzoylecgonine 0·6 mg/L (eight samples), benzodiazepines 1·6 mg/L (four samples), cannabinoids 59 μg/L (four samples), methadone 1·7 mg/L (11 samples) and morphine 1·7 mg/L (seven samples). Significant lack of sensitivity was demonstrated for one or more analytes by all chromatographic techniques, including gas chromatography with mass spectrometry. Commercial immunoassay kits exhibited higher sensitivity, though some assays performed significantly less well than others. Significant non-specificity was also a greater problem for chromatographic techniques compared with immunoassays in samples containing therapeutic agents not structurally related to abused compounds. In the presence of therapeutic interferents structurally related to the test compounds, false positive reports for illicit materials increased from < 1% to > 6%. Use of confirmatory tests significantly reduced the incidence of both false negative and false positive reports.
Hypocholesterolaemia in a Hospital Population
Authors: Crook M A , Velauthar U , Moran L , Griffiths W
Published: February 2014

The purpose of this study was to observe the prevalence of hypocholesterolaemia in a hospital population and also the causes and clinical outcome of this condition. Fifty-seven patients were found with a plasma cholesterol of ≤3·0 mmol/L, which was less than 0·50% of all plasma cholesterol requests; there were 39 men and 18 women ( P<0·05, Chi-squared test). The mean age was 53·8 [21·3] (range 3–83 years). The mean plasma cholesterol concentration was 2·28 [0·56] mmol/L (1·16–3·0) and the mean triglyceride concentration was 1·58 [1·09] mmol/L (0·49–7·35).

There was a significant correlation between plasma cholesterol concentration and plasma albumin (Rs = 0·48, P < 0·01) and between plasma total protein concentration (Rs = 0·49, P < 0·01). However, there was no relationship between the concentrations of plasma cholesterol and triglyceride (Rs = −0·10, P > 0·05).

Eighteen per cent of patients with hypocholesterolaemia died during their hospitalization. Thirty-nine per cent of those who had a plasma cholesterol of ≤2·0 mmol/L died whereas 71% of those who had a plasma cholesterol concentration of < 1·5 mmol/L died. Hypocholesterolaemia was more commonly seen in the intensive care unit and in post-operative patients, those with malignancy, sepsis, acute myocardial infarction, those who had inflammatory bowel disease and diabetics on insulin. Hypocholesterolaemia may be a useful predictor of mortality in hospital patients.
Quantitative Analysis of Ribonucleoside Triphosphates in Human Lymphoid Cells by Micellar Electrokinetic Capillary Chromatography
Authors: Bolzonella C , Gusella M , Bononi A , Loregian A , Crepaldi G , Toso S , Palù G , Ferrazzi E
Published: February 2014

Micellar electrokinetic capillary chromatography (MECC) was applied to develop an analytical method for quantitation of ribonucleoside triphosphates (rNTPs) in human lymphoid cells obtained from patients with B-chronic lymphocytic leukaemia (B-CLL) and cutaneous lymphomas. The results of this analysis showed a significant depression of intracellular rNTPs in patients with B-CLL, compared with rNTPs of healthy controls. These data are in agreement with other studies in which rNTP separations were performed with traditional high-performance liquid chromatography. MECC has proved to be a useful tool for intracellular rNTPs determination, revealing possible new applications in the study of the metabolic state of human cells. In addition, this method can be useful in monitoring the effect of many drugs (antiviral, antineoplastic) which interfere with nucleotide metabolism.
The Clinical Role of Clinical Biochemists
Authors: Pearson M J
Published: February 2014

Project EVETSIN investigated the training of clinical biochemists and other professional groups in the UK National Health Service. One of the most significant professional changes (with a training implication) identified was the increasing importance of the clinical role of clinical biochemists. The skills involved and the processes by which trainee clinical biochemists learn them had not been described or analysed to a significant extent. Factors that influence this changing role, the nature of the skills, how they are acquired and maintained, and the associated knowledge base are presented and discussed in the context of the training of clinical biochemists in the UK.
Faecal Elastase 1: A Marker of Exocrine Pancreatic Insufficiency in Cystic Fibrosis
Authors: Phillips I J , Rowe D J , Dewar P , Connett G J
Published: February 2014

Pancreatic elastase 1 (E1), a digestive protease, is synthesized by the acinar cells of the pancreas. Using an enzyme-linked immunosorbent assay, we evaluated stool E1 levels in the following groups of patients:

Specimens submitted for occult blood examination from 20 adults, over 3 consecutive days, to assess the inter-day variability in E1 excretion. There were no symptoms suggestive of pancreatic insufficiency in this group. The mean E1 concentration over all samples was 457 μg E1/g stool (range 124–1683). The intra-assay variation was 6·4% ( n = 14) and the inter-assay variation was 8·8% ( n = 12). The mean intra-patient variation was 17%. Cystic fibrosis (CF) patients. Eight patients had E1 levels in the reference range (>200 μg E1/g stool). The remaining 25 patients had undetectable E1 levels. A control group of children presenting with unexplained bronchiectasis and/or recurrent respiratory infections and no symptoms of pancreatic dysfunction. The mean E1 concentration in the group was 519 μg E1/g stool (range 139–1941).

There was no significant difference in E1 concentrations between the two non-CF groups, nor between the pancreatic-sufficient CF patients when compared with both non-CF groups. There was a significant difference between the pancreatic-sufficient and -insufficient CF groups ( P < 0·001) using the Mann—Whitney U test.

All fifteen CF patients who were ΔF508 homozygotes had undetectable E1. It may be possible to relate CF genotype to the presence or absence of E1 and to the degree of pancreatic insufficiency. Measurement of faecal E1 in children with CF appears to differentiate them into a group of children with normal pancreatic function and a larger group with severe insufficiency.
An Unusual Alkaline Phosphatase Isoenzyme Associated with Gastric Carcinoma
Authors: Jenkins M A , Steer C B , Cheng L W H , Ratnaike S
Published: February 2014

We describe a 50-year-old man with a diagnosis of gastric carcinoma made on gastroscopy after X-rays of the thoracolumbar spine had revealed multiple lytic metastases. A bone marrow aspirate showed adenocarcinoma cells. Polyacrylamide gel electrophoresis incorporating wheat germ lectin was used to separate the serum alkaline phosphatase isoenzymes. Isoenzyme separation showed a markedly increased amount of bone isoenzyme, a normal amount of liver isoenzyme and a considerable amount of an intestinal-like isoenzyme running cathodic to the bone isoenzyme. There was also some immunoglobulin-complexed alkaline phosphatase, which, when digested, showed more of the intestinal-like isoenzyme. This was a variant alkaline phosphatase isoenzyme found in a patient with a gastric carcinoma with a super bone scan. There have been two previous reports of patients with a variant alkaline phosphatase isoenzyme and a super bone scan.
Serum Concentration and Renal Handling of 1,5-Anhydro-D-Glucitol in Patients with Chronic Renal Failure
Authors: Shimizu Hidekazu , Shouzu Akira , Nishikawa Mitsushige , Omoto Seitaro , Hayakawa Takashi , Miyake Yutaka , Yonemoto Toshinaga , Inada Mitsuo
Published: February 2014

We measured serum and urinary 1,5-anhydro-D-glucitol (1,5-AG) during a glucose tolerance test (GTT) in patients with chronic renal failure (CRF) and compared the fractional excretion of 1,5-AG (FEAG) with that of diabetes mellitus (DM) patients and healthy controls. The mean serum 1,5-AG in CRF patients [60 ± 23(SE) μmol/L] was significantly lower than in controls (155 ± 7 μmol/L) in spite of a normal glycaemia. The levels in the CRF group were similar to those in the DM group. During GTT, the blood glucose profile in the CRF group was not significantly different from that of the control group, and urinary glucose excretion was negligible. However, FEAG was significantly higher in CRF patients than in controls. These data suggest that serum 1,5-AG in patients with CRF decreases due to a decrease in 1,5-AG reabsorption, independently of glucose excretion, and that serum and/or urinary 1,5-AG can be a useful marker for renal tubular dysfunction because the 1,5-AG reabsorption system is more vulnerable than the glucose reabsorption system.