bluuebunny/crossref_metadata_2025 huggingface dataset:
Measurement of total mercury in biological specimens by cold vapor atomic fluorescence spectrometry
Authors: Winfield S A , Boyd N D , Vimy M J , Lorscheider F L
Published: January 2020

An ultrasensitive method for determining total mercury concentrations in biological specimens is a prerequisite for monitoring exposure to chronic low-dose levels of Hg vapor such as those from dental silver amalgam fillings. The clinical consequences of such doses are currently in question. We describe an adaptation of a two-stage gold amalgamation preconcentration step combined with cold vapor atomic fluorescence spectrometric detection for Hg. At Hg concentrations of 40 and 350 nmol/L, the within-day assay CVs were 5% and 3%, respectively; between-day assay CVs were 8% and 5%, respectively. Accuracy, as demonstrated by analytical recovery, ranged from 98% to 105%. The detection limit for the assay is 50 pmol/L, which is suitable for measuring total Hg concentrations in specimens of human urine, blood, and breast milk, and in monkey kidney cortex and feces, obtained from subjects with and without amalgam fillings.
Modification of the dextran-Mg2+ high-density lipoprotein cholesterol precipitation method for use with previously frozen plasma
Authors: McNamara J R , Huang C , Massov T , Leary E T , Warnick G R , Rubins H B , Robins S J , Schaefer E J
Published: January 2020

Although dextran-Mg2+ precipitation produces accurate and precise results for high-density lipoprotein (HDL) cholesterol in fresh plasma and serum, precipitation of frozen specimens with triglycerides > 2.26 mmol/L (> 200 mg/dL) is difficult. We developed a modification that dilutes thawed samples by 35% and increases dextran-Mg2+ reagent to 15% of sample volume. Standard precipitations were performed on 62 fresh EDTA-treated plasma specimens; supernatant solutions were analyzed fresh and after freezing. Standard and modified methods were also performed on thawed, paired plasmas. In specimens with triglycerides < or = 2.26 mmol/L, HDL cholesterol results for all methods were similar. For triglycerides > 2.26 mmol/L, however, bias and precision were significantly affected by freezing, and 38.5% of samples with standard precipitation required additional procedures to produce clear supernatant solutions. HDL cholesterol concentrations for thawed samples with standard precipitation were significantly greater than for fresh samples (P < 0.02), but those for the modified method were not different from fresh samples, and only one specimen required additional steps to produce a clear supernate.
Distribution of CII and CIII peptides in lipoprotein classes: methods and clinical significance
Authors: Graziani M S , Zanolla L , Righetti G , Marchetti C , Zanotto G , Lupo A
Published: January 2020

We describe a method for measuring apolipoprotein (apo) C distribution between apo B-containing lipoprotein (apo B-LP) and non-apo B-LP. The procedure requires the precipitation of apo B-LP, the redissolution of the pellet, and the quantification of C peptides in the redissolved pellet. The ratio of apo C in non-apo B-LP to apo C in apo B-LP has been calculated for both CII and CII (R-CII and R-CIII, respectively). R-CII (0.49 +/- 0.25) and R-CIII (0.84 +/- 0.54) in patients on maintenance dialysis are significantly lower than in the control group (1.14 +/- 0.57 and 1.45 +/- 0.92, respectively), indicating that hypertriglyceridemia in these patients results from a reduced catabolism of triglyceride-rich LP (TGRLP). Patients with coronary artery disease (CAD) show a distribution of C peptides no different from the control group. Analysis of covariance reveals that the patterns of R-CII and R-CIII are not entirely predictable from the serum concentration of triglycerides. This result seems to support the hypothesis that the underlying metabolic defects involving TGRLP in dialysis patients are not the same as those in patients with CAD.
Role of serotonin in the pathophysiology of depression: focus on the serotonin transporter
Authors: Owens M J , Nemeroff C B
Published: January 2020

Considerable evidence has accrued in the last two decades to support the hypothesis that alterations in serotonergic neuronal function in the central nervous system occur in patients with major depression. These findings include the following: (a) reduced cerebrospinal fluid (CSF) concentrations of 5-hydroxyindoleacetic acid (5-HIAA), the major metabolite of serotonin (5-HT) in drug-free depressed patients; (b) reduced concentrations of 5-HT and 5-HIAA in postmortem brain tissue of depressed and (or) suicidal patients; (c) decreased plasma tryptophan concentrations in depressed patients and a profound relapse in remitted depressed patients who have responded to a serotonergic antidepressant when brain tryptophan availability is reduced; (d) in general, all clinically efficacious antidepressants augment 5-HT neurotransmission following chronic treatment; (e) clinically efficacious antidepressant action by all inhibitors of 5-HT uptake; (f) increases in the density of 5-HT2 binding sites in postmortem brain tissue of depressed patients and suicide victims, as well as in platelets of drug-free depressed patients; (g) decreased number of 5-HT transporter (determined with [3H]imipramine or [3H]paroxetine) binding sites in postmortem brain tissue of suicide victims and depressed patients and in platelets of drug-free depressed patients. In our studies, this reduction in platelet 5-HT transporter binding is not due to prior antidepressant treatment of hypercortisolemia and is not observed in mania, Alzheimer disease, schizophrenia, panic disorder, fibromyalgia, or atypical depression. In a pilot study, this deficit predicted treatment response to an experimental antidepressant. These findings support the hypothesis that alterations in 5-HT neurons play a role in the pathophysiology of depression.
Plasma concentrations of gamma-aminobutyric acid (GABA) and mood disorders: a blood test for manic depressive disease?
Authors: Petty F
Published: January 2020

gamma-Aminobutyric acid (GABA), an inhibitory neurotransmitter that serves about one-third of brain neurons, is involved in the development of depression and in the treatment of depression and mania with pharmacological therapy. Brain activity of GABA may be conveniently measured in plasma, and changes in plasma concentrations of GABA reflect brain GABA activity. Plasma concentrations of GABA are significantly lower than control values in about one-third of patients with major depressive disorder; concentrations are also low in patients with mania and in bipolar patients who are depressed. These low concentrations of GABA appear to persist after recovery from depression and are not increased by treatments that improve depressive symptoms. Follow-up studies suggest that GABA concentrations remain relatively constant over at least 4 years. Additionally, preliminary data suggest that low plasma GABA is a familial marker of mood disorders in a subset of patients. Despite the difficulty of demonstrating that a particular biochemical measure is a true genetic trait marker for vulnerability for development of an illness, the accumulated data suggest that low plasma GABA may represent a biological marker of vulnerability for development of various mood disorders.
Brain mechanisms in manic depression
Authors: Carroll B J
Published: January 2020

Manic depressive illness (bipolar disorder) is the mood disorder classically considered to have a strong biological basis. During manic depressive cycles, patients show dramatic fluctuations of mood, energy, activity, information processing, and behaviors. Theories of brain function and mood disorders must deal with the case of bipolar disorder, not simply unipolar depression. Shifts in the nosologic concepts of how manic depression is related to other mood disorders are discussed in this overview, and the renewed adoption of the Kraepelinian "spectrum" concept is recommended. The variable clinical presentations of manic depressive illness are emphasized. New genetic mechanisms that must be considered as candidate factors in relation to this phenotypic heterogeneity are discussed. Finally, the correlation of clinical symptom clusters with brain systems is considered in the context of a three-component model of manic depression.
Quantification of lipoprotein(a) in plasma by assaying cholesterol in lectin-bound plasma fraction
Authors: Seman L J , Jenner J L , McNamara J R , Schaefer E J
Published: January 2020

Lipoprotein(a) [Lp(a)] is a low-density lipoprotein (LDL)-like particle in which apolipoprotein(a) [apo(a)] is disulfide-linked to apolipoprotein B (apoB). High concentrations of Lp(a) in plasma are associated with an increased risk of coronary heart disease (CHD). Lp(a) has traditionally been measured by immunoassay and expressed as total mass of Lp(a). Measuring Lp(a) by its cholesterol content will provide a way to directly compare Lp(a) with other lipoproteins that are measured by cholesterol. We have developed an assay to quantify Lp(a) by its cholesterol content [Lp(a)-C], using lectin affinity to isolate Lp(a) from other lipoproteins, and then measuring the cholesterol within the isolated fraction. We compared the Lp(a)-C assay with an ELISA for Lp(a) mass in 47 plasma samples from normotriglyceridemic, fasting individuals with high Lp(a) contents (mean +/- SD, 446 +/- 350 mg/L). The mean Lp(a)-C concentration was 110 +/- 89 mg/L and correlated very highly with Lp(a) mass (r = 0.9975). Lp(a)-C measurement is an alternative method to screen for this CHD risk factor.
Demonstration and minimization of serum interference in flow cytometric two-site immunoassays
Authors: Frengen J , Kierulf B , Schmid R , Lindmo T , Nustad K
Published: January 2020

The ability of serum factors to cross-link labeled mouse monoclonal antibody (mAb) of irrelevant specificity (mAb FN61, subclass IgG1) to different particle types coated with sheep IgG, bovine gamma-globulin, or mAb FN61 was measured simultaneously by flow cytometry. Significant interference with mAb FN61-coated particles was detected in 53 of 101 sera. Of the 30 sera showing the most pronounced interference, 23 were characterized by an even stronger cross-linking to particles coated with bovine gamma-globulin. These were designated type 1 sera. Seven sera, designated type 2, displayed a dominant interference with the mAb FN61-coated particles. The interference reaction in the two serum types was characterized by different kinetics, dependence on particle concentration, and response to blocking agents. The interference was minimized by addition of 500 micrograms of bovine gamma-globulin and 50 micrograms of mAb HH1 (IgG1) of irrelevant specificity per 10 microL of serum sample in a final assay volume of 100 microL.
Plasma homocyst(e)ine as a risk factor for early familial coronary artery disease
Authors: Wu L L , Wu J , Hunt S C , James B C , Vincent G M , Williams R R , Hopkins P N
Published: January 2020

We measured plasma homocyst(e)ine [H(e)] and other coronary risk factors in 266 patients with early coronary artery disease from 170 families in which two or more siblings were affected and in 168 unmatched controls. The mean H(e) concentration adjusted for significant correlates (serum creatinine, uric acid, and low-density lipoprotein cholesterol) was 12.0 mumol/L in proband cases compared with 10.1 mumol/L in controls (P = 0.0001). Many (17.6%) of the proband cases had H(e) concentrations exceeding the 95th percentile for the controls (relative odds = 4.9, P < 0.001). H(e) among cases was bimodally distributed even after adjustment for concentrations of plasma vitamins. Concordant high H(e) was seen in at least 10 (12%) of 85 families with two or more affected siblings. We conclude that a substantial proportion of early familial coronary artery disease is probably related to production of high concentrations of H(e) by one or more major genes.
International Federation of Clinical Chemistry standardization project for measurements of apolipoproteins A-I and B. IV. Comparability of apolipoprotein B values by use of International Reference Material
Authors: Marcovina S M , Albers J J , Kennedy H , Mei J V , Henderson L O , Hannon W H
Published: January 2020

We performed temporal and thermal stability studies on SP3-07, a liquid-stabilized reference material for apolipoprotein (apo) B, selected during the previous phase of the International Federation of Clinical Chemistry project on standardization of apolipoprotein measurements. Results indicate that SP3-07 stored at -70 degrees C has the long-term stability required for a reference material. We assigned an accuracy-based apo B value of 1.22 g/L to SP3-07, using a nephelometric method that was calibrated with freshly isolated low-density lipoprotein for which the apo B mass value was determined by a standardized sodium dodecyl sulfate-Lowry procedure. Using a common protocol, the study participants transferred the assigned mass value from SP3-07 to the individual calibrators of the analytical systems and measured the apo B concentration of 20 fresh-frozen samples obtained from individual donors and covering a clinically relevant range of apo B values. The among-laboratory CV on these samples, analyzed by 25 analytical systems, ranged from 3.1% to 6.7%. These results demonstrate the lack of matrix effects of SP3-07 and its ability to provide accurate and comparable apo B values in a variety of immunochemical methods. On the basis of the outcome of these studies, the World Health Organization has endorsed SP3-07 as the International Reference Material for Apolipoprotein B.
Method-dependent variations in the stability of apolipoprotein B in a stabilized liquid reference material
Authors: Mei J V , Powell M K , Henderson L O , Smith S J , Cooper G R , Marcovina S M , Hannon W H
Published: January 2020

Using accelerated Arrhenius-type short-term and long-term temporal studies, we evaluated the storage life of a stabilized, liquid-frozen reference material (SLRM) for human apolipoprotein B (apo B) developed by the International Federation of Clinical Chemistry. As measured by our candidate reference RIA, the concentrations of immunoreactive apo B in the SLRM showed pronounced degradation with exposure to increasing temperatures over time. The SLRM was stable for as long as 1 year when stored at - 70 degrees C, but its immunoreactive apo B declined by < 10% when stored at 4 degrees C for 10 months. Using radial immunodiffusion and an ELISA to assess the equivalency of measured mass for the accelerated thermal stability of the SLRM, we found a loss of immunoreactive apo B similar to that measured by RIA. Analyzing the same samples by liquid immunoprecipitation (nephelometry) resulted in the amount of apo B present being overestimated, especially in samples held for long periods. By using different immunological methods to evaluate this thermally aged SLRM, we demonstrated that its measured behavior varies depending on the method of quantitation.
15-Desmethyl FK-506 and 15,31-desmethyl FK-506 from human liver microsomes: isolation, identification (by fast atom bombardment mass spectrometry and NMR), and evaluation of in vitro immunosuppressive activity
Authors: Lhöest G J , Maton N , Latinne D , Laurent A , Verbeeck R K
Published: January 2020

15-Desmethyl FK-506 and 15,31-desmethyl FK-506 metabolites extracted from incubated human liver microsomes were separated by normal and reversed-phase HPLC. The metabolites were identified by fast atom bombardment/mass spectrometry and nuclear magnetic resonance spectroscopy. The in vitro 50% inhibitory concentration, tested against bidirectional mixed lymphocyte reactions, was 325 mg/L for 15,31-desmethyl FK-506 and 4 mg/L for 15-desmethyl FK-506, indicating that these products of FK-506 demethylation retain in vitro immunosuppressive activity.
Time-resolved immunofluorometric assay and specimen storage conditions for measuring urinary gonadotropins
Authors: Saketos M , Sharma N , Adel T , Raghuwanshi M , Santoro N
Published: January 2020

We optimized storage conditions and validated a sensitive immunofluorometric assay (IFMA) for urinary gonadotropins. Assay linearity and parallelism for luteinizing hormone (LH) and follicle-stimulating hormone (FSH) was observed to 0.04 IU/L. Urinary LH and FSH were unaffected by changes of osmolarity from 0.5 to 3.0 mOsm/kg, and from pH 4.5 to 10.5. Serum and urine measurements of the hormones correlated well over a wide range of values: for LH, R2 = 0.94, P < 0.01; for FSH, R2 = 0.71, P < 0.01 (n = 304). Preservation of urine with glycerol (70 mL/L) and storage at -20 degrees C yielded > 80% recovery of LH and FSH after 51 weeks; this was comparable with recovery for acetone extracts of urine. Untreated urine showed loss of activity by 4 weeks of storage. Preserving urine specimens with glycerol is a simple method of storage for longitudinal study and compares favorably with acetone extraction. IFMAs can measure urinary gonadotropins reproducibly over a wide range of pH and osmotic conditions.
Immunoassays for serum and urine myoglobin: myoglobin clearance assessed as a risk factor for acute renal failure
Authors: Wu A H , Laios I , Green S , Gornet T G , Wong S S , Parmley L , Tonnesen A S , Plaisier B , Orlando R
Published: January 2020

We compared four immunoassays for serum and urine myoglobin. Within-run CVs were 5-13%, with biases seen between assays. Myoglobin was stable for 1 month in serum and 12 days in urine when the pH was adjusted to between 8.0 and 9.5. Hemoglobin caused no interference. We assayed 91 pairs of serum and timed urine specimens from 41 patients admitted for acute trauma or rhabdomyolysis. Most were treated with mannitol and alkalinization. Upon initial presentations, 21 patients with either low serum myoglobin concentrations (< 400 micrograms/L) or high myoglobin clearances (> or = 4 mL/min) had normal creatinine clearances and no clinical evidence of renal disease. The remaining 20 had low myoglobin clearances. Seven were in rhabdomyolysis-induced acute renal failure, or subsequently developed this complication. We suggest that low myoglobin clearance may indicate a high risk for developing renal failure or may be an early marker for kidney dysfunction. Low myoglobin clearance may prove useful in indicating failure of prophylactic treatment to clear myoglobin.
Enzyme immunoassay for erythrocyte aldose reductase
Authors: Nishimura C , Hamada Y , Tachikawa T , Ishikawa T , Gui T , Tsubouchi J , Hotta N , Tanimoto T , Urakami T
Published: January 2020

This two-site immunoassay measures erythrocyte aldose reductase by using monoclonal and polyclonal antibodies to recombinant human enzyme. Total incubation time is 2.5 h, and the limit of detection is < 0.05 microgram/L. Analytical recovery tested with blood samples from healthy and diabetic individuals was 101-106%. Average CVs within and between assays were 3.7% and 4.8%, respectively. The enzyme content determined by this system correlated well with the activity of aldose reductase isolated from the same erythrocyte preparations. The amount of erythrocyte aldose reductase per milligram of hemoglobin was higher in women than in men (P < 0.001), but no significant correlation was observed between the amount of enzyme and the age of the individuals. This assay method should provide useful clinical information to optimize administration of aldose reductase inhibitors for effective prevention and treatment of diabetic complications.
Monitoring tricyclic antidepressant concentrations in serum by fluorescence polarization immunoassay compared with gas chromatography and HPLC
Authors: Rao M L , Staberock U , Baumann P , Hiemke C , Deister A , Cuendet C , Amey M , Härtter S , Kraemer M
Published: January 2020

The fluorescence polarization immunoassay (FPIA) developed by Abbott to diagnose intoxication with tricyclic antidepressants was adapted for therapeutic drug monitoring and validated with chromatograpic methods to investigate its potential for this use. We compared serum concentrations of tricyclic antidepressants in vivo and in vitro obtained by FPIA with those by gas chromatography and HPLC. For amitriptyline, imipramine, clomipramine, and doxepin, the detection limit of the FPIA was 72, 71, 64, and 72 nmol/L (approximately 20 micrograms/L), respectively; that by gas chromatography was 18, 18, and 16 nmol/L (approximately 5 micrograms/L) for amitriptyline, imipramine and clomipramine, respectively; with HPLC the lower limit of detection for doxepin was 36 nmol/L (10 micrograms/L). The intra- and interassay CVs ranged from 3% to 6%. In patients being treated with amitriptyline, imipramine, clomipramine, and doxepin, at steady-state the correlation coefficients between FPIA and GC/HPLC results for split samples were 0.95, 0.92, 0.90 and 0.70, respectively. However, the slopes were close to unity only for amitriptyline and doxepin, being 0.6 for imipramine and 1.9 for clomipramine.
High-resolution 1H-NMR spectroscopy of blood plasma for metabolic studies
Authors: Wevers R A , Engelke U , Heerschap A
Published: January 2020

Although spin-echo techniques are often used to obtain 1H-NMR spectra of serum or plasma samples, they do not provide reliable quantitative analyses of metabolites. We present a standardized procedure, optimized for sensitivity, for using single-pulse 1H-NMR spectroscopy to analyze deproteinized plasma. The detection limit for various metabolites ranges between 2 and 40 mumol/L. The method allows quantitative analysis of many compounds of interest in studies of inborn errors of metabolism, including betaine and dimethylglycine, which cannot be measured easily with other techniques. For lactate, tyrosine, threonine, and alanine, we obtained results that correlated well with those obtained by established techniques. We also present a library containing resonance positions of 38 compounds occurring in plasma samples in health and disease, including 14 as-yet-unidentified resonances. As an example of the diagnostic power of the technique we show a spectrum of a plasma sample from a patient with 5-oxoprolinuria (pyroglutamic aciduria; McKusick 266130), an enzymatic defect in glutathione biosynthesis.
A simple enzyme-linked immunosorbent assay of human osteocalcin
Authors: Rosenquist C , Bonde M , Fledelius C , Qvist P
Published: January 2020

A heterologous ELISA for measurement of osteocalcin (bone Gla-protein) is described, involving biotinylated bovine osteocalcin and polyclonal antibodies. The log-linear range was 2.3-37.5 micrograms/L. Between-run (total) and within-run CVs (n = 10) were 5.7-6.4 and 5.9-6.1%, respectively; analytical recoveries ranged from 92% to 108%. Comparison of our method (x) with an RIA (y) yielded y = 1.10 x -0.01 microgram/L, Sylx = 1.4 micrograms/L, r = 0.958 (n = 167). Serum osteocalcin in healthy premenopausal women (n = 29) was 8.7 +/- 3.3 micrograms/L (mean +/- SD) and 11.8 +/- 4.5 micrograms/L in early-postmenopausal women (n = 24). The assay was evaluated in a double-blind placebo-controlled study of healthy early-postmenopausal women, treated for 12 months with either (a) estrogen valerate plus medroxyprogesterone acetate (n = 18), (b) 17 beta-estradiol and desogestrel (n = 22), or (c) placebo (n = 17). Serum osteocalcin decreased significantly (P < 0.001) with either therapy, but increased (P < 0.05) with placebo.
New decision limits and quality-control material for detecting human chorionic gonadotropin misuse in sports
Authors: Laidler P , Cowan D A , Hider R C , Kicman A T
Published: January 2020

Male athletes may administer human chorionic gonadotropin (hCG) to enhance performance. Despite imposing a ban on hCG doping, the International Olympic Committee has no recommended procedure to confirm the presence of hCG. Our aim was to establish a limit for hCG in urine above which a sample may be considered positive. We measured hCG concentrations in urine samples from 1400 men with the Serono MAIAclone IRMA. Statistical evaluation of the results gave a "far outside" value [75th percentile + (3 x interquartile range)] of 5.0 IU/L; greater values are extremely unusual. Immunoreactive material in urine samples from a reference group of 120 noncompeting individuals was concentrated about sevenfold by centrifugal ultrafiltration; all these concentrated samples gave assay values < 5.0 IU/L. To ensure with the greatest possible degree of certainty that no false-positive result is reported, we propose a decision limit of 10 IU/L in nonconcentrated ultrafiltered samples. We also prepared quality-control material to contain hCG at 10 IU/L. This material provides a decision limit above which, after confirmatory procedures, a sample should be considered positive. This should help establish comparability of results among laboratories testing for sports drugs.
Mechanisms of carcinogenesis and individual susceptibility to cancer
Authors: Venitt S
Published: January 2020

The population can be divided into four groups, or "oncodemes," depending on the relative contributions of environment and genetics to their risk of cancer. These oncodemes are: 1) background (random mutations in normal people); 2) environmental (environmental carcinogens acting on normal people); 3) environmental/genetic (environmental carcinogens acting on genetic susceptibility); and 4) genetic, with genetic susceptibility being more important than environmental exposure. Most cancer probably occurs in oncodemes 2 and 3. However, the contribution that genetic susceptibility to cancer makes to the total cancer burden is unknown. Genetic susceptibility may be important in occupational settings, where exposure to carcinogens is presumed to be restricted to concentrations believed to confer "acceptable" risks. This approach takes no account of individual susceptibility. The range over which metabolic polymorphisms exert their effects suggests that differences in pharmacogenetics between individuals may be important in occupational carcinogenesis. If this range were extrapolated to risk of human disease, at a given exposure one person might be 10-200 times more sensitive than another. Several metabolic polymorphisms have been linked to susceptibility to cancer. For some of these polymorphisms, genes have been cloned and genotypic tests based on use of the polymerase chain reaction have been developed. Limited data suggest that the effects of these polymorphisms on human cancer are complex. The genotypic testing of individuals occupationally exposed to carcinogens may provide firmer information on the relative contributions of nature and nurture on chemically induced cancer. The use to which such information is put demands wide debate.
Assessment of copper nutritional status
Authors: Milne D B
Published: January 2020

Despite increased interest in the role of copper deficiency in clinical problems and an increased understanding of the physiological roles of copper, the diagnosis of a marginal deficiency has not been perfected. The use of non-standardized procedures and the effects of factors other than copper nutriture have impeded identification of the "ideal" indicator of copper nutritional status in adult humans. The specific activity of copper enzymes, or of copper-containing enzymes in blood cells, such as erythrocyte superoxide dismutase and platelet or leukocyte cytochrome c oxidase, may be a better indicator of metabolically active copper stores than the serum concentration of copper or ceruloplasmin, because the enzyme activities are sensitive to changes in copper stores and are not as sensitive to factors not related to copper nutriture. A single index, such as serum copper concentration, is inadequate for assessing the total body copper nutriture of an individual and must be supported by corroborating evidence.
Pentane and isoprene in expired air from humans: gas-chromatographic analysis of single breath
Authors: Mendis S , Sobotka P A , Euler D E
Published: January 2020

Both pentane and isoprene are excreted in human breath. Although pentane is considered an index of lipid peroxidation, the significance of isoprene is unknown. Having a similar boiling point, these two hydrocarbons are difficult to separate by gas chromatography. We separated pentane from isoprene on both a Poraplot Q and a Poraplot U column, injecting single-breath samples directly into a gas chromatograph. The breath samples were pressurized to 800 mmHg to increase the amount of sample volume delivered to the column. In a group of 43 healthy volunteers, the concentrations of end-expiratory pentane and isoprene were 0.57 +/- 0.3 and 7.05 +/- 3.53 nmol/L, respectively. There was a significant linear correlation (r = 0.57, P < 0.0001) between age and pentane concentration in expired air; isoprene showed no correlation with age or pentane concentrations. The age-related increase in pentane production suggests that oxidative stress may play a role in the aging process in humans. The method described should allow for rapid, inexpensive, serial measurement of expired pentane and isoprene.
Time-resolved immunofluorometric assay of trypsin-2 complexed with alpha 1-antitrypsin in serum
Authors: Hedström J , Leinonen J , Sainio V , Stenman U H
Published: January 2020

We developed a sensitive time-resolved immunofluorometric assay (IFMA) for trypsin-2 complexed with alpha 1-antitrypsin (AAT). We used a trypsin-2-specific monoclonal antibody on the solid phase and a europium-labeled polyclonal antibody to AAT as tracer. The detection limit is 0.05 microgram/L and the range of linearity extends to 100 micrograms/L. We compared the clinical utility of the trypsin-2-AAT assay with that of free trypsinogen-2 and amylase in serum by studying 120 healthy subjects, 29 patients with acute pancreatitis, 11 with extrahepatic biliary obstruction, and 34 with acute abdominal disorders of extrapancreatic origin. In patients with acute pancreatitis the median concentration of trypsin-2-AAT in serum was 59-fold that in healthy controls, 42-fold that in patients with biliary obstruction, and 33-fold that in patients with acute abdominal disorders of extrapancreatic origin. These differences are greater than those for trypsinogen-2 (19-, 20-, and 28-fold, respectively) and amylase (5.4-, 6.5-, and 5.4-fold, respectively). Compared with the assays of free trypsinogen-2 and amylase, our assay of trypsin-2-AAT improved the clinical specificity for acute pancreatitis by eliminating false-positive results in our control groups. Increased concentrations of trypsin-2-AAT and trypsinogen-2 were also observed in patients with chronic renal failure undergoing dialysis.
Nanotechnology: Basic Concepts and Definitions
Authors: Kaehler Ted
Published: January 2020

Molecular nanotechnology is an engineering discipline in which the goal is to build devices and structures that have every atom in the proper place. By means of this general purpose material-processing technology it will be possible to build almost any rigid, covalently bonded structure. Identical parts will be truly identical, enabling energy conversion and computation systems to have extremely high performance and reliability. Assembly will be done by mechanosynthesis, the process of holding two reactive molecules in contact with each other in a controlled orientation. Synthesis will be done on nanoscale assembly lines called molecular mills, where systems of moving belts will press individual molecules together and catalyze 106 reactions/s per station. High-performance mechanical computers will use moving rods to block the motion of other rods. The most important physical limit will be radiation damage. Without redundancy, a subsystem lasting 100 years will be limited to 106 nm3.
Effect of red wine ingestion on the antioxidant capacity of serum
Authors: Whitehead T P , Robinson D , Allaway S , Syms J , Hale A
Published: January 2020

Aerobic metabolism in biological systems produces reactive oxygen species, and defense against such prooxidants requires antioxidant activity, e.g., predominantly vitamins C and E in serum. Recently, flavonoids (polyphenols occurring widely in plants) have been investigated in vitro for their antioxidant activity; whether they are absorbed after ingestion is not clear. Using a chemiluminescent assay of serum antioxidant capacity (SAOC), we have studied the effects in normal individuals of ingesting red wine, white wine, and high doses of vitamin C. In nine subjects who ingested 300 mL of red wine, the mean SAOC was increased by 18% after 1 h and by 11% at 2 h. The same amount of white wine produced 4% and 7% increases, respectively. The ingestion of 1000 mg (5.7 mmol) of ascorbic acid by four subjects increased their mean SAOC by 22% at 1 h and by 29% at 2 h. An in vitro comparison of red wine, white wine, and various fruit juices showed the high antioxidant capacity of red wine in addition to its ability to increase the antioxidant capacity of serum in vivo. The antioxidant effects of various flavonoids and other polyphenols were also studied.
Rapid diagnosis of maple syrup urine disease in blood spots from newborns by tandem mass spectrometry
Authors: Chace D H , Hillman S L , Millington D S , Kahler S G , Roe C R , Naylor E W
Published: January 2020

We report a new method for the diagnosis of maple syrup urine disease (MSUD) from dried blood spots on newborn screening cards based on tandem mass spectrometry (MS-MS). The mean +/- SD concentration of Leu plus Ile in normal newborns was 151 +/- 47 mumol/L (n = 1096); for Val, 131 +/- 58 mumol/L (n = 791). SDs were lower when the concentrations of these amino acids were expressed relative to that of Phe. The mean ratio for Leu + Ile to Phe was 2.5 +/- 0.49; for Val to Phe, 2.18 +/- 0.51. These results compare well with values previously reported in the literature. With these criteria, samples from a collection categorized by a bacterial inhibition assay as normal or falsely positive for MSUD were normal by MS-MS [(Leu + Ile): Phe < 5.0]. Samples from confirmed MSUD patients were categorized as abnormal [(Leu+Ile): Phe > 9.0] by MS-MS.
Solubility of silver sulfadiazine in physiological media and relevance to treatment of thermal burns with silver sulfadiazine cream
Authors: Tsipouras N , Rix C J , Brady P H
Published: January 2020

Silver sulfadiazine cream has been a standard treatment for burns over the past two decades. Although many studies have described the phenomenon of silver absorption from burn wounds treated with silver sulfadiazine, they failed to examine the chemistry underlying the absorption process: Silver chloride was assumed to form at the burn wound and absorption of silver was believed to be negligible. Here we have developed chemical model systems to investigate the interactions of silver sulfadiazine and silver chloride in direct contact with synthetic serum electrolyte solution (SSES), with SSES plus endogenous ligands or beef blood plasma, and with human serum. The results indicate that silver absorption from an acute burn site can be significant, because human serum is capable of solubilizing silver. This finding is of concern, given the potential for silver toxicity as a direct consequence of applying silver sulfadiazine to extensive burn wounds.
Intestinal strontium absorption: from bioavailability to validation of a simple test representative for intestinal calcium absorption
Authors: Sips A J , van der Vijgh W J , Barto R , Netelenbos J C
Published: January 2020

Calcium absorption tests have rarely been validated for being representative for absolute bioavailability (true absorption) or for intraindividual variation. Therefore, we investigated the reproducibility of the absolute bioavailability of strontium chloride, a marker for intestinal calcium absorption, in healthy male volunteers (n = 8) by measuring the area under the plasma strontium concentration-time curve after oral and intravenous administration of strontium. Subsequently, we selected a simple test variable as being representative of absolute bioavailability. The mean absolute bioavailability (+/- SD) was 25% +/- 7%. The best test variable appeared to be the fractional absorption at 240 min (Fc240) after oral intake, which demonstrated the highest correlation with absolute bioavailability (r = 0.66). The intraindividual variations of the data for this variable and for the absolute bioavailability are similar to those described for various absorption tests based on the use of calcium isotopes. Thus, the Fc240 of strontium offers the potential of a simple clinical test for use as a measure of intestinal calcium absorption and its modulation.
Serum bilirubin and risk of ischemic heart disease in middle-aged British men
Authors: Breimer L H , Wannamethee G , Ebrahim S , Shaper A G
Published: January 2020

The possibility that low concentrations of serum bilirubin may be associated with increased risk of ischemic heart disease has been examined in a prospective study of 7685 middle-aged British men. During 11.5 years there were 737 major ischemic heart disease (IHD) events. A U-shaped relationship was observed between serum bilirubin and risk of IHD. Low bilirubin was associated with several cardiovascular risk factors, in particular smoking, low concentrations of high-density lipoprotein cholesterol, low forced expiratory volume in 1 s, and low serum albumin. The U-shaped relationship persisted even after adjusting for several risk factors. Compared with men in the lowest fifth of the distribution (bilirubin < 7 mumol/L), those in the middle range (8-9 mumol/L) showed a 30% reduction in relative risk [RR = 0.68 (95% confidence intervals 0.51-0.89)] in IHD, whereas men in the top fifth (> 12 mumol/L) showed similar risk to the lowest fifth [RR = 0.99 (95% confidence intervals 0.73-1.34)], which persisted after exclusion of men with bilirubin > 17 mumol/L. The significance of this U-shaped relationship is unclear, but it could be interpreted as support for the role of endogenous antioxidants in the etiology of IHD.
Composition of interstitial fluid
Authors: Fogh-Andersen N , Altura B M , Altura B T , Siggaard-Andersen O
Published: January 2020

In several previous experiments to determine the composition of interstitial fluid, the results varied depending on the collecting technique, and the electrolyte concentrations differed from those of a hypothetical ultrafiltrate of plasma. In our approach, since a change of position from standing to supine is accompanied by hemodilution with interstitial fluid, we used the changes in hematocrit and composition of plasma in 20 subjects before and after lying down to calculate the composition of added interstitial fluid. The estimated protein concentration was 20.6 g/L, and the concentrations of total calcium and magnesium were low, in accord with a lower concentration of protein-bound calcium and magnesium. The activity of free cations was also lower, in agreement with a Donnan equilibrium potential of 1 mV across the endothelium. The concentration of leukocytes and platelets decreased according to the hemodilution, implying no escape or mobilization of these elements.
Monoclonal antibody assay for measuring bone-specific alkaline phosphatase activity in serum
Authors: Gomez B , Ardakani S , Ju J , Jenkins D , Cerelli M J , Daniloff G Y , Kung V T
Published: January 2020

Alkaline phosphatase (ALP) is present in human serum in the form of several isoenzymes. The two major circulating ALP isoenzymes, bone and liver, are difficult to distinguish because they are the products of a single gene and differ only by posttranslational glycosylation. Quantitative measurement of bone ALP (BAP) activity in serum can provide an index for the rate of bone formation. Furthermore, increased BAP activity in serum is indicative of bone disorders. We describe a method in which serum samples are added to a microtiter plate coated with monoclonal anti-BAP antibody and incubated 3 h at room temperature. After the unbound materials are washed off, the bound BAP activity is measured by adding p-nitrophenyl phosphate substrate. The assay demonstrated no cross-reactivity to intestinal or placental ALP and only 3-8% cross-reactivity to liver ALP. The intraassay (n = 21) CVs were 3.9-5.9%, and interassay (n = 8) CVs were 4.4-7.0%. Comparisons of the assay (y) with an IRMA (x) and a wheat germ agglutinin precipitation method (x') gave regression equations of y = 1.32x-6.4, r = 0.99, and y = 1.41x' + 4.8, r = 0.99. The assay detected increased BAP in sera from patients with osteoporosis, Paget disease, osteomalacia, or primary hyperparathyroidism.
Effect of aging on susceptibility of low-density lipoproteins to oxidation
Authors: Schmuck A , Fuller C J , Devaraj S , Jialal I
Published: January 2020

According to the Adult Treatment Panel of the National Cholesterol Education Program, age is a major risk factor for heart disease. To assess the relation between age and LDL oxidizability, we studied copper-mediated LDL oxidation in 13 healthy elderly subjects (> 59 years) and 13 sex-matched healthy young controls (< 30 years). Total and LDL-cholesterol concentrations were increased in elderly subjects. The time course of copper-mediated LDL oxidation showed no significant differences between the two groups as assessed by formation of conjugated dienes, lipid peroxides, and apolipoprotein B fluorescence. Kinetics of LDL oxidation as quantified by lag time, oxidation rate, and maximal oxidation were not significantly different between the elderly and young groups. Although the concentrations of 16:0, 18:0, 18:1, 18:3, and 20:4 and total polyunsaturated fatty acids were significantly higher in the elderly group, LDL fatty acid concentrations were similar in both groups. Lipid-standardized alpha-tocopherol, beta-carotene, and ascorbate concentrations were not significantly different between the two groups. The findings of the present study suggest that in the healthy elderly, LDL oxidation may not be a crucial mediator for atherogenesis.
Serum concentrations of apolipoprotein A-I, apolipoprotein B, and lipoprotein(a) in a population sample
Authors: Leino A , Impivaara O , Kaitsaari M , Järvisalo J
Published: January 2020

Serum concentrations of apolipoprotein (apo) A-I, apo B, and lipoprotein(a) [Lp(a)] were studied with respect to age and sex in a Finnish population sample of 575 subjects (286 men and 289 women), ages 27-67 years. Apo A-I and apo B were measured with an immunoturbidimetric method calibrated against WHO International Reference Materials. Lp(a) was measured by RIA. Apo A-I and apo B concentrations were almost normally distributed (apo A-I: mean 1.38 g/L vs median 1.34 g/L for men, and 1.58 g/L vs 1.55 g/L for women; apo B: mean 1.21 g/L vs median 1.20 g/L for men and 1.09 g/L vs 1.05 g/L for women). The distribution of Lp(a) was remarkably skewed (mean 190 mg/L vs median 86 mg/L for men, and 169 mg/L vs 85 mg/L for women). The 95% intervals for apo A-I were 1.09-1.84 g/L for men and 1.06-2.28 g/L for women; for apo B, they were 0.63-1.88 g/L and 0.56-1.82 g/L, respectively. Apo A-I concentrations appeared to be unrelated to age, whereas apo B and Lp(a) concentrations were age-dependent. Cutoff values based on the 90th percentile for apo B and the 10th percentile for apo A-I are proposed for identifying subjects at increased risk of coronary heart disease.
Detection of prostate-specific antigen mRNA by reverse transcription polymerase chain reaction and time-resolved fluorometry
Authors: Galvan B , Christopoulos T K , Diamandis E P
Published: January 2020

We have developed a time-resolved fluorometric hybridization assay for detecting prostate-specific antigen (PSA) mRNA amplified by reverse transcription polymerase chain reaction. During PCR, digoxigenin-11-dUTP is incorporated into the amplified product. An oligonucleotide internal to the primers is used as a specific probe, being biotinylated and captured on streptavidin-coated microtiter wells. Denatured PCR product hybridizes with the probe, and the hybrids are detected with an alkaline phosphatase-labeled antidigoxigenin antibody. We used the phosphate ester of fluorosalicylic acid as the substrate. The fluorosalicylate produced forms a highly fluorescent ternary complex with Tb(3+)-EDTA, which we can measure by time-resolved fluorometry. A signal-to-background ratio of 10 was obtained when 160 PSA cDNA molecules were present in the preamplification sample. Also, mRNA corresponding to one LNCaP cell in the presence of 10(6) PSA-negative cells can be detected (signal-to-background ratio of 3.1). Samples containing 100, 1000, and 50,000 LNCaP cells gave CVs of 12.4%, 4.9%, and 6.8%, respectively (n = 10).
Quantitative determination of high-, low-, and very-low-density lipoproteins and lipoprotein(a) by agarose gel electrophoresis and enzymatic cholesterol staining
Authors: Nauck M , Winkler K , März W , Wieland H
Published: January 2020

Quantification of lipoprotein cholesterol was performed by enzymatic staining of cholesterol in a new agarose gel electrophoresis method that allows the separation of LDL, VLDL, HDL, and lipoprotein(a) [Lp(a)]. Lp(a) shows an electrophoretic mobility clearly distinct from VLDL and HDL. The total CVs of lipoprotein cholesterol varied between 2.7% and 3.9% for LDL, 7.8% and 23.2% for VLDL, 5.2% and 9.5% for HDL, and 6.8% and 16.4% for Lp(a). Comparison of LDL-, VLDL-, and HDL-cholesterol concentrations with the results of a combined ultracentrifugation and precipitation technique gave correlation coefficients of 0.961, 0.947, and 0.918, respectively; comparison of Lp(a)-cholesterol values with those of a nephelometric Lp(a) assay gave r = 0.906. The new electrophoretic assay has several advantages: It allows the quantification of Lp(a)-cholesterol; VLDL-cholesterol is not affected by Lp(a)-cholesterol; and the LDL-cholesterol fraction does not contain Lp(a)-cholesterol, as happens with LDL-cholesterol determined by ultracentrifugation and precipitation.
Intracellular magnesium content of mononuclear blood cells and granulocytes isolated from leukemic, infected, and granulocyte colony-stimulating factor-treated patients
Authors: Loun B , Astles R , Copeland K R , Sedor F A
Published: January 2020

The intracellular magnesium (Mg) concentration of granulocytes and mononuclear blood cells (MBCs) was determined in cells isolated from patients with several disorders. The mean (+/-SD) Mg content of MBCs isolated from patients diagnosed with lymphocytic leukemia, myelocytic leukemia, or infection; from patients treated with granulocyte colony-stimulating factor (G-CSF); and from healthy volunteers (control group) was 2.3 (+/-0.6), 3.3 (+/-0.5), 4.1 (+/-0.8), 3.9 (+/-0.4), and 3.9 (+/-0.6) fmol/cell, respectively. The Mg content of MBCs isolated from patients with lymphocytic and myelocytic leukemia, but not those from patients with infection or receiving G-CSF treatment, were significantly lower (P < 0.001) than those from the control subjects. The mean Mg concentration of granulocytes obtained from lymphocytic leukemia, myelocytic leukemia, infection, and G-CSF patients and from the control group was 3.2 (+/-0.9), 3.4 (+/-0.5), 3.8 (+/-0.6), 4.5 (+/-0.6), and 4.6 (+/-0.6) fmol/cell, respectively. Granulocytes isolated from leukemic and infectious patients yielded lower intracellular Mg concentrations (P < 0.005) than those from patients receiving G-CSF and the control group. This study demonstrates that intracellular Mg content is altered in several pathological states. Several factors, including depleted Mg stores or altered intracellular Mg binding sites, could be responsible for these changes. Apparently, intracellular Mg content may be of use in assessing total body Mg status.