bluuebunny/crossref_metadata_2025 huggingface dataset:
Gas-chromatographic analysis for therapeutic concentration of imipramine and disipramine in plasma, with use of a nitrogen detector.
Authors: Bailey D N , Jatlow P I
Published: January 2020

We describe a method for the simultaneous gas-chromatographic analysis of imipramine and desipramine (the active N-desmethyl metabolite) at therapeutic concentrations in human plasma, with use of a nitrogen detector. Imipramine is measured as the unchanged base and desipramine as its N-tribluoroacetyl derivative. Promazine is the internal standard. The lower limit of sensitivity is 5 mug/liter for each drug, and the within-day coefficients of variation for the determination of imipramine and desipramine, respectively, are 6.0% and 3.3%. We measured concentrations of these drugs both in patients receiving therapeutic doses of imipramine and in those who had ingested an overdose.
Reduced variation of tracer binding in digoxin radioimmunoassay by use of (125I)-labeled tyrosine-methyl-ester derivative: relation of thyroxine concentration to binding.
Authors: Kroening B H , Weintraub M
Published: January 2020

Between-sample variation in tracer binding in the 125I-labeled digoxin radioimmunoassay was investigated with two tracers, 3-O-succinyl-digoxigenin-[125I]-labeled tyrosine and [125I]-labeled tyrosine-methyl-ester-digoxin. Digoxin-free serum samples having various concentrations of thyroxine were assayed with both tracers. The percentage of tracer bound when the samples were assayed with the first-mentioned tracer was increased significantly for the low thyroxine groups when compared to the normal (P less than 0.001) or the high thyroxine groups (P less than 0.05). Little difference existed when the latter tracer was used. There was variation in tracer binding when serum from dogs dosed with thyrotropin was assayed with the first tracer, but there was little variation with the second. Tracer binding may be influenced by thyroxine-binding proteins. Variation in tracer binding appears to be reduced when [125I]-labeled tyrosine-methyl-ester-digoxin is used.
Long-term reproducibility of a new pH/blood-gas quality-control system compared to two other procedures.
Authors: Noonan D C , Komjathy Z L
Published: January 2020

The long-term precision and stability of a new quality-control system for blood pH and gas measurements are compared to that of tonometered bicarbonate solutions and serum-based preparations. The new system, consisting of gas-equilibrated bicarbonate solutions in glass ampuls, is shown to be as stable as the serum-based preparation, and as reproducible as either of the other methods. The new system, offering three discrete sets of control values, has certain advantages in the simultaneous quality of pH, carbon dioxide tension, and oxygen tension measurements.
Correlation of selected serum constitutents: 2. Consistency of intra-individual correlation values, means, and variances during four months.
Authors: Winkel P , Statland B E
Published: January 2020

We examined whether inter-individual differences in correlation coefficients previously found among subjects truly reflect consistent inter-individual differences or are time-related within an individual. The consitutents studied in this investigation were (a) the enzmes aspartate aminotransferase, alkaline phosphatase, and lactate dehydrogenase; and (b) the non=protein nitrogen-containing constituents urea, uric acid, and creatinine. Ten healthy women were each subjected to 15 venipunctures over a five-week period (Series I), and, after a two-month interval, were again subjected to 15 venipunctures over a second five-week period (Series II). Before statistical analysis, the data were corrected for the batch-to-batch (day-to-day) arnalytical variation. There was a signiificant (P less than .05) change in the covariance structure (variances or correlation coefficients, or both) between the two series in four of the 10 subjects for the combination of enzymes, and in three other subjects for the combination of nonprotein nitrogen constitutents. Although we found a significant (P lees than .05) average intra-individual variation in the mean values from series to series in the cases of the three enzymes and urea, the magnitude of the inter-series variation in means was relatively small. CV's were: alkaline phosphatase, 3.4%; lactate dehydrogenase, 2.3+; aspartate aminotransferase, 3.3%; urea, 5.0%; uric acid, 1.0%; and creatinine, 1.2%.
Kinetic determination of serum haptoblobin with a centrifugal analyzer.
Authors: Kickler T S , Gong P F , Johnson G F , Solomon H M
Published: January 2020

We describe a method for determining haptoglobin with a centrifugal analyzer that is based on haptoglobin combining stoichiometrically with hemoglobin to form a complex that has peroxidase-like activity proportional to the quantity of haptoglobin present. Under assay conditions, unbound hemoglobin exhibits only a small fraction of the total peroxidase activity. Activity is measured colorimetrically at 405 nm after reaction with o-dianisdine and ethyl hydrogen peroxide. The procedure is standardized by saturating aknown amount of hemoglobin with a serum whose hemoglobin binding capacity exceeds the amount of hemoglobin in the assay system. The mean and mean within-run precision of our method, determined by performing 17 replicate assays of both a pooled normal serum and a 10-fold dilution of the serum, was 1.13 g/liter (CV, 2.9%), and 106 mg/liter (cv, 5.8%), respectively. The 95 percentile estimate of the normal range by our method is 0.45-1.85 g/liter hemoglobin binding capacity. When results by our automated method were compared to those by a manual method [Scand. J. Clin. Lab. 2nvest. 18, 80 (1965)], the slope of the unweighted linear least-squares regression line was .970 the y-intercept 26 mg/liter, and the correlation coefficient .995.
A mechanism for the conversion of oxyhemoglobin to methemoglobin by nitrite.
Authors: Rodkey F L
Published: January 2020

Each mole of oxyhemoglobin iron converted to methemoglobin causes the oxidation of 1.5 mol of nitrite to nitrate and consumes 1 mol of protons. No oxygen is liberated. The overall reaction has two simultaneously occurring parts. In the beginning the rate-limiting reaction converting O2Hb to metHb is directly proportional to H+ and NO2- concentrations and is independent of metHb. The second portion accounts in major part for the stoichiometry and rate of the overall reaction. In this portion O2Hb tetramers and metHbNO2- are the reactants. Essentially no reaction takes place in the presence of CN-, which displaces nitrite from the metHbNO2-, nor in the presence of 0.5 mol/liter Nal, which converts the O2Hb to alphabeta-dimers. The autocatalytic nature of the overall reaction in the presence of excess nitrite is the result of metHb, which is formed in both parts of the reaction, associating with nitrite to increase the concentration of one reactant of the cyanide-sensitive part. The reaction rates at constant pH in excess nitrite are porportional to the product of the O2Hb concentration and the square of the metHb concentration. The rate increases up to about 66% conversion of O2Hb followed by a decrease as the O2Hb becomes limiting. The dissociation constant of metHbNO2- at 25 degrees C and pH = 6.4 was found to be 1.11+/-0.11 mmol/liter.
Relation of pH to fluorescence of serotonin, melatonin, and other indole compounds reacted with o-phthaldialdehyde.
Authors: de Leiva A , Schwartz S
Published: January 2020

We distinguished serotonin, malatonin, and other indole compounds by their markedly different fluorescence behavior when heated with o-phthaldialdehyde in different concentration of HCl, or after the subsequent addition of alkali. Fluorescence may increase, disappear, or change from blue to orange-red. Only melatonin developed significant fluorescence (at less than 0.5 mug/liter) when heated with the reagent in 50 mmol/liter HCl. Serotonin and 5-hydroxyindole compounds with a C-3 aliphatic chain lost their blue fluorescence when brought from strong to weakly acid and alkaline pH. Intense reddish fluorescence developed in strongly alkaline solution. C-5 methoxy derivatives, such as melatonin, maintained their blue fluorescence in alkaline solution. Differences in fluorescence color (and RF) also characterized these compounds on silica gel plates. The use of toluene decreased blank fluorescence, while sample preparation in dim light, as often recommended, resulted in diminished and less-stable fluorescence.
Enzymatic Rate Method for Measuring Cholesterol in Serum
Authors: Pesce Michael A , Bodourian Selma H
Published: January 2020

An enzymatic rate assay is described for measuring cholesterol in serum. Cholesterol is anzymed by mixing 5 µl of sample with a reagent consistaing of cholesterol esterase, cholesterol oxidase, catalase, acetylacetone, methanol. and hydroxypolyethoxydodecane in a ammonium phosphate of the dihydrolutidine product is measured at 37 °C and 405nm. The change in absorbance between 4 and 10 min is used to calculated the cholesterol concentrations by using simultaneously determined free cholesterol standards. The change is linearly ralated to cholesterol concentration up to 4 g/liter. Samples containing bilirubin up to 200 mg/liter, uric acid up to 200 mg/liter, and hemoglobin up to 1 g/liter, or certain drugs (clofibrate, phenobarbital, nicotinic acid, salicylate, Ketochol, and Ovral) gave no interference. Ascorbic acid added to serum caused a positive interference. Lipemic samples gave values that were slightly lower than did the method of Abell et al., used for comparison. Our kinetic assay, compared with the method of Abell et al., the enzymatic assay used with Abbott's Bichromatic Analyzer, and the Technicon SMA 12/60 enzymatic procedure gave correlation coefficient of 0.992, 0.985, and 0.986, respectively.
Sample stability: a suggested definition and method of determination.
Authors: Thiers R E , Wu G T , Reed A H , Oliver L K
Published: January 2020

We propose defining the stability of any chemical constituent of stored samples in terms that are quantitatively related to the precision of the measurement by which it is determined. We suggest that a constituent may be considered stable, for a stated period and under exactly defined conditions, when the average change in its measured value is less than a chosen number, K, of standard deviations of the data obtained by the measuring method over the concentration range in question. Based on this definition a technique utilizing a graphical truncated normal sequential test is presented as the appropriate experimental and statistical design for measuring stability. The statistical basis for the proposal is presented.
Analysis for indole compounds in urine by high-performance liquid chromatography with fluorometric detection.
Authors: Graffeo A P , Karger B L
Published: January 2020

We describe a chromatographic system involving a high-performance chemically-bonded reverse-phase column and fluorescence detection for measurement of indoles in urine. We controlled retention and selectivity by optimizing the methanol content and pH of the mobile phase. Six reference indoles were separated in less than 20 min; three 5-hydroxyindoles were eluted in less than 7 min. About 5-15 ng of aqueous solutions of these compounds can be detected. The combination of selectivity (from use of the chromatographic column) and fluorescence detection permitted analysis for five of the six indoles after a single urine-deproteinization step.
Improved assay for urinary 17-hydroxycorticosteroids.
Authors: Ariyoshi Y , Osawa Y
Published: January 2020

We describe a simple, rapid, reliable method for determining urinary 17-hydroxycorticosteroids. A neutral resin (Amberlite XAD-2), which is a non-ionic cross-linked polystyrene, is used to extract and concentrate the steroids, which are then quantitatively determined with the Porter-Silber reaction. Use of the resin eliminates the need for enzymatic hydrolysis and n-butanol extractions, thereby decreasing analysis time considerably; results can be obtained within 3 h of receipt of specimens. Most of the nonsteroidal Porter-Silber chromogens are removed, resulting in a method that is highly specific and sensitive, even at low concentrations (0.6 mg/liter), and so more accurate and reproducible than currently used methods.
Assessment of a serum separator device for obtaining serum specimens suitable for clinical analyses.
Authors: Laessig R H , Westgard J O , Carey R N , Hassemer D J , Schwartz T H , Feldbruegge D H
Published: January 2020

We have evaluated a serum separator device that is designed to effect better separation and isolation of serum from clotted blood. This device consists of an evacuated blood-drawing tube (Vacutainer) containing 1.5 ml of a semi-solid silicon polymer that forms a barrier between serum and clot during centrifugation. We compared test specimens with conventionally processed (Vacutainer) specimens as controls. Eighty-one chemical, toxicological, and immunological reactions, tests, or methods were studied. For several tests we observed changes that are statistically but not clinically significant. The one clinically significant change in serum chemistry values occurred in the case of lactate dehydrogenase, for which values were increased by 5-8% of the upper limit of normal.
Radioimmunoassay of plasma renin activity.
Authors: Fyhrquist F , Soveri P , Puutula L , Stenman U H
Published: January 2020

We describe a sensitive, simplified radioimmunoassay method for determination of plasma renin activity. Plasma was acidified to the optimal pH (6.0) of angiotensin l generation with the least possible dilution, by using a single addition of hydrochloric acid and the enzyme inhibitor hydroxyquinoline. Recovery of unlabeled angiotensin l added to plasma was 92-97%; that of monoiodinated angiotensin l exceeded 90%, indicating satisfactory protection from proteolytic enzymes. Plasma constituents interfered little with the radioimmunoassay. Bland values for plasma kept at 0 degrees C were 10.7 +/- 2.3 (mean +/- SD) percent of the activity values for samples kept at +37 degrees C (n equals 63). In the routine setting, 6.25 pg of angiotensin l or 10(-6) Goldblatt units of Standard Human Renin was detected. We report results of plasma renin activity measurements and a comparison with seven renin kits, and with bioassay for plasma renin activity.
Use of polyethylene glycol in separating bound from unbound ligand in radioimmunoassay of thyroxine.
Authors: Cheung M C , Slaunwhite W R
Published: January 2020

In most rapid radioimmunoassay methods, absorbents are used that disturb the equilibrium of the ligand-antibody interaction; thus the separation process must be rigidly timed. We show here that polyethylene glycol (mol wt. 6000) abolishes this constant. We measured gamma-globulin precipitation by polyethylene glycol (150g/liter) in a thyroxine radioimmunoassay system involving use of a sheep anti-thyroxine antiserum, quantitatively and qualitatively, with either normal human serum or completely precipitated at pH 6-9 at 4 and 25 degrees C, but incompletely if salt concentration was high (1 mol/liter). Specificity of gamma-globulin precipitation increased with increasing pH and temperature. All gamma-globulin-bound thyroxine was precipitated, and also some not bound to gamma-globulin. This was taken into account by including "blank" tubes (no antibody, but with normal sheep serum) in all assays. Because this reaction is predominantly entropic and temperature independent, the entire procedure can be done at 37 degrees C and room temperature without rigid timing. The assay requires 10 mul of serum, its reproducibility is 4-8% (CV) in the euthyroid and hyperthyroid range, and its accuracy is close to 100%.
Determination of apolipoprotein A and its constitutive A-I and A-II polypeptides by separate electroimmunoassays.
Authors: Curry M D , Alaupovic P , Suenram C A
Published: January 2020

Electroimmunoassays ("rocket" electrophoresis) are described for human serum apolipoprotein A and its constitutive A-I and A-II polypeptides. Purified lipoprotein A, A-I, and A-II were used to prepare monospecific antisera and to standardize assays. These specific, rapid (5-8 h), precise (the within-and between-assay coefficients of variations are 5 and 7%, respectively), and accurate (by gravimetry) assays are applicable to measurement of these polypeptides in whole serum and in various density classes of lipoproteins. Comparable results are obtained with intact and delipidized lipoproteins. Results correlated well with those obtained by radial immunodiffusion or radioimmunoassay. However, the present procedure is more rapid than the former and simpler than the latter immunoassay. Concentrations of A-I and A-II in the serum of normal men and women were similar (143 +/- 24 and 146 +/- 78 mg/dl, respectively, for A-I and 78 +/- 17 and 83 +/- 25 mg/dl for A-II). Subjects with type lla, llb, and IV hyperlipoproteinemias had similar concentrations of both polypeptides, while patients with type I disease, lecithin:cholesterol acyltransferase deficiency and LP-A deficiency had lowest concentrations of A-I (0.3-30 mg/dl) and A-II (11-20 mg/dl). The molar ratio of A-I/A-II in the serum and high-density lipoproteins was close to unity.
Evaluation of methods of coenzyme activation of erythrocyte enzymes for detection of deficiency of vitamins B1, B2, and B6.
Authors: Bayoumi R A , Rosalki S B
Published: January 2020

We describe optimized, ultraviolet spectrophotometric procedures for determination of erythrocyte transketolase, glutathione reductase, and aspartate aminotransferase activity, and their activation by their respective coenzymes--thiamine pyrophosphate, flavin-adenine dinucleotide, and pyridoxal-5-phosphate--as tests for vitamin B1, B2, and B6 deficiency. With these procedures we have investigated healthy subjects on normal and vitamin-supplemented diets, and a series of (mainly) alcoholic hospital in-patients. The enzyme procedures described have good precision and can be readily carried out in the routine laboratory. Abnormal transketolase activation correlated well with clinical evidence of vitamin B1 deficiency.
Method for monitoring plasma progesterone concentrations in pregnancy.
Authors: Winkel P , Gaede P , Lyngbye J
Published: January 2020

We present a time-series model for monitoring concentrations in plasma of hormones produced in the placenta, progesterone being chosen as an example. The model, which is based on the assumption that variations in plasma progesterone concentration in pregnant subjects mainly reflect variations in the growth rate of the placenta, was applied to eight series of progesterone values measured during pregnancy in eight subjects. In the model, which was found to fit the data, it is assumed that progesterone concentration is proportional to the size of the placenta and that the growth rate of the placenta varies at random, with a mean value alpha. The variation of alpha was of the same magnitude among and within the subjects. If the average of many subjects alpha values is used, a single subject may be used as her own reference, based on only one previous observation. When two observations are available, an individual's own alpha value may be estimated and used for the prediction. The predictive power of the new method was found to be far superior to the conventional method in which a single sample reference material is used. Furthermore, one need not know the gestational age in order to use the method.
Development of an aqueous temperature-indicating technique and its application to clinical laboratory instrumentation.
Authors: Bowie L , Esters F , Bolin J , Gochman N
Published: January 2020

We describe a new technique for monitoring intra-cuvette temperatures in temperature-controlled spectrophotometric cells by use of two solutions, one for which absorbance is highly sensitive to temperature and one for which absorbance is insensitive to temperature. The first solution, cresol red in tris(hydroxymethyl)aminomethane buffer, is used to construct an absorbance-vs.-temperature calibration curvae. The second solution, cresol red in sodium hydroxide, is used for optical calibration of the spectrophotometric system to be tested. The combination of these two procedures allows accurate estimation of temperature. We show how this technique is applied to temperature measurement in several types of laboratory instrumentation.
Expression of random analytical error as a precentage of the range of clinical interest.
Authors: Glick J H
Published: January 2020

I propose that random method error be expressed as a percentage of the range(s) of clinical interest (upper range limit minus lower range limit), in addition to its usual forms of presentation. The interpretation of this percent random error is simple because a given percentage error implies the same reliability, regardless of test or range of clinical interest. Two different upper limits for percent random analytical error are suggested, based on the medical application of the test and the statistical properties of various amounts of error. When the purpose of the test is to distinguish an upper medically defined limit from a lower medically defined limit, a maximum random error equivalent to 60% of the difference between the limits is proposed. If the purpose of the test is to monitor values in the neighborhood of a normal range or therapeutic range, an upper limit for random error equal to 20% of the difference between the limits of the range is suggested.
Immunofixation. I. General principles and application to agarose gel electrophoresis.
Authors: Ritchie R F , Smith R
Published: January 2020

Immunofixation offers the worker an economical means of physically locating a protein in an electrophoretic strip and is ideally suited to forensic medicine, genetic studies, or research. The method is as simple and economical as the commonly used one- or two-dimensional immunoelectrophoresis, yet yields considerably more information.
Fast, economical technique for separating serum from blood samples.
Authors: Ahluwalia G S , Kuczala Z J , Carlson L R
Published: January 2020

We evaluated the effectiveness with which polystyrene beads separate serum from blood. Paired blood samples were analyzed for 23 different serum constituents. One specimen of each pair was processed by the conventional separation method, the other by a single centrifugation with the polystyrene beads. Differences between the assay means were evaluated by the z-test, and the data pairs were compared by regression analysis. No statistically significant differences were found.
Evaluation of a new enzymatic kit for serum triglycerides.
Authors: Carter T , Martin P J , Wilding P , Goldberg D M
Published: January 2020

We evaluated a new enzymatic kit for serum triglycerides (triacylglycerols) in two laboratories. The procedure involved both enzymatic hydrolysis of the triglycerides and conversion of the glycerol, which was linked to an indicator reaction. There was excellent agreement with the mechanized procedure of Kessler and Lederer and with a manual method. The test requires 50 mul of serum and can be completed in about 50 min. Inter- and intra-laboratory variation, linearity, accuracy, and specimen storage were studied.
Improved specificity of serum albumin determination and estimation of "acute phase reactants" by use of the bromcresol green reaction.
Authors: Gustafsson J E
Published: January 2020

The reaction of serum samples with bromcresol green proceeds in two steps. Albumin is responsible for the faster (less than 1 min) reaction; the slower (30-min) reaction is a measure of "acute phase reactant(s)" in serum. Serum is simply mixed with bromcresol green reagent and the absorbance is measured twice, immediately and at 60 min. Albumin concentrations, determined from the absorbance at 0 min, correlate well with those determined by Laurell "rocket" immunoelectrophoresis; r = 0.95 with no certain deviation from unity for the slope and with a negligible difference at zero concentration. The slow reaction was expressed as deltaA% = 100 (deltaAs/deltaAv) where deltaAs and delta Av are the changes in absorbance between 60 and 0 min for the sample and a commercial control serum, respectively. The value for deltaA% correlates well with the percentage of alpha2-fraction as determined by electrophoresis on cellulose acetate, as well as with orosomucoid and ceruloplasmin, all of which are acute phase reactant(s). Whether these proteins or other acute phase reactant(s) actually cause the slow reaction has not yet been established.
Evaluation of lipase activity in serum by radial enzyme diffusion.
Authors: Goldberg J M , Pagast P
Published: January 2020

Results of determination of serum lipase by radial enzyme diffusion correlate well with those by a titrimetric reference method in the abnormal range. The specificity of the diffusion assay allows the differentiation of patients with pancreatic disease, even when the lipase activity of the serum is within the normal limits of the tritrimetric assay. Pancreatic lipase is not detectable by the diffusion assay in the serum of individuals who are free from pancreatic disease.
Technical pitfalls in measurement of venous plasma NH3 concentration.
Authors: Gerron G G , Ansley J D , Isaacs J W , Kutner M H , Rudman D
Published: January 2020

Measurement of venous plasma NH3 in normal subjects by the ion-exchange method of Forman [Clin. Chem. 10, 497 (1964)] in a hospital clinical laboratory gave a mean value of 640 mug/liter (range, 300-1320 mug/liter; intraassay, intra-individual, and inter-individual coefficients of variation, 8, 47, and 47%, respectively). The following conditions adversely affect the reproducibility of the test: pollution of laboratory atmosphere and glassware by NH3-containing detergents; smoking by patient or analyst; delay, turbulence, or use of heparin lock in venipuncture; delay or warming of plasma above degrees C before mixing it with resin; and delay in colorimetric analysis of resin eluate. When these sources of error were eliminated, the mean value for normals was reduced to 330 mug/liter, the range was narrowed to 220-470 mug/liter, and the above-mentioned CV's were 5, 16, and 17%, respectively. With the precautions cited, furthermore, the intra-assay and intra-individual CV's for fasting NH3 concentration in cirrhotic patients were similarly reduced. An NH3 tolerance test was done by administering a standard dose of NH4Cl to patients and measuring venous plasma NH3 at 0, 15, 30, 60, and 90 min; the NH3 tolerance was quantified from the area under the curve relating concentration to time (mug - min/liter). As measured in the clinical laboratory, NH3 tolerance of cirrhotic patients showed intra-assay and intra-individual CV's of 50 to 90%. When the tolerance tests were repeated in the same subjects with the laboratory precautions listed above, these CV's were reduced to 8-15%.
Analytical toxicology: Applications of element-selective electrolytic conductivity detection for gas chromatography.
Authors: Pape B E
Published: January 2020

We applied a commercially available microelectrolytic conductivity detector to toxicological problems of qualitative and quantitative analysis by gas chromatography. The detector can be used for the sensitive and selective detection of halogen-, nitrogen-, and sulfur-containing compounds. Relative response in different element-selective detector variables such as reaction gas, reaction catalyst, and furnace temperature can be used to further improve qualitative identification by gas chromatography.
Gas-chromatographic micro-scale procedure for theophylline, with use of a nitrogen-sensitive detector.
Authors: Least C J , Johnson G F , Solomon H M
Published: January 2020

We report a micro-scale procedure for determination of theophylline by use of gas chromatography with nitrogen-sensitive detection. The procedure requires only 10 mul of serum, plasma, or saliva. This sample size makes the determination especially appropriate for monitoring in a pediatric population. With the sample volume used, background interference is equivalent to about 0.1 mg/liter and 0.5 mg of theophylline per liter can easily be measured. The new method correlates well with our earlier flame ionization gas-chromatographic procedure that required 1 ml of sample [Clin. Chem. 21, 1038 (1975)]. The selectiviey of the nitrogen detector allows a simplified extraction procedure. Between-run precision (CV) is 2.8% at a theophylline concentration of 14.8 mg/liter.
Gas-chromatographic determination of 5-fluorocytosine in human serum.
Authors: Harding S A , Johnson G F , Solomon H M
Published: January 2020

We describe a sensitive and precise gas-chromatographic method, in which cytosine is used as the internal standard, for determination of an antifungal agent, 5-fluorocytosine, in serum. The trimethylsilyl derivative of this drug is well separated from the internal standard and from normal serum constituents. Amphotericin B does not interfere with the determination of 5-fluorocytosine. The lower limit of detection for 5-fluorocytosine is 1 mg/liter when 200 mul of serum is analyzed. Within-run precision (CV), established by analysis of 10 replicates, was 4.5% at a concentration of 19.9 mg/liter. Twenty-five serum samples were analyzed for 5-fluorocytosine by a microbiological assay and by the gas-chromatographic method. Mean value observed with the bioassay was 78.5 mg/liter and with our procedure was 69.4 mg/liter. When values for our assay were regressed against values for the bioassay, slope of the least-squares line was 0.85, intercept was 2.7 mg/liter, and r was 0.93.
Gas-chromatographic determination of an antifibrinolytic drug, epsilon-aminocaproic acid.
Authors: Keucher T R , Solow E B , Metaxas J , Campbell R L
Published: January 2020

We describe a modified method for assay of epsilon-aminocaproic acid. Serum or cerebrospinal fluid is deproteinized, followed by cation-exchange column-chromatography, and N-trifluoroacetyl-n-butyl derivatives of amino acids are formed and separated by gas chromatography. Tranexamic acid, a nonprotein amino acid, was used as an internal standard. The assay is sensitive and precise, and results correlate adequately with those obtained with an automated amino acid analyzer (ion-exchange chromatography).
Radioimmunoassay for clindamycin.
Authors: Gilbertson T J , Stryd R P
Published: January 2020

We describe a radioimmunoassay for measuring clindamycin in serum extracts that is more accurate than the microbiological assay because of the minimal response of the matabolite, N-demethylclindamycin. The assay is not affected by the presence of other antiobiotics. As described it is as sensitive as the microbiological assay, but can be made more sensitive.
Analysis of results of toxicological examinations performed by coroners' or medical examiners' laboratories in 2000 drug-involved deaths in nine major U. S. cities.
Authors: Dinovo E C , Gottschalk L A , McGuire F L , Birch H , Heiser J F
Published: January 2020

Data were collected on 2000 deaths in which psychoactive drugs were involved. The data were submitted by medical examiners or coroners in nine U. S. cities from their case files. The 2000 cases comprise a representative sample from each of these cities of deaths from psychoactive drugs between 1972 through 1974. This report details inter-city differences in methods and practices of the toxicological examination as well as the type and numbers of drugs reported. Even when the same analytical method was used in various cities, there were differences in extraction solvent and extraction pH. Of the 3909 drugs detected, 2945 were quantitated; the number of drugs quantitatively measured per case studied ranged from a low of 0.82 for New York to a high of 2.20 for Washington, D.C. The number of different drugs quantitatively measured varied from 16 for New York to 30 for San Francisco; however, New York qualitatively assayed for the presence of a total of 25 drugs. The number and type of drugs found per case varied. Methadone, for example, was found in 60% of the cases reported by New York and in 49% of the cases from Washington, D.C., but in only about 10% of cases reported by Philadelphia, Dallas, Miami, and San Francisco, and in less than 1% of Los Angeles and Cleveland cases; it was not reported by Chicago. Phenmetrazine-caused deaths were reported only by Dallas (one case) and Washington, D.C. (29 cases). From the data as a whole, information is presented for 33 drugs as to the concentration in physiological tissues and fluids. Analysis of single psychoactive drug cases and single-drug-plus-ethanol cases shows that, in the presence of ethanol, the toxic blood concentration of imipramine, amytriptyline, meprobamate, thioridazine, morphine, propoxyphene, methaqualone, and all barbiturates was decreased by an average of 50%.
Rapid micromethod for measuring anticonvulsant drugs in serum by high-performance liquid chromatography.
Authors: Soldin S J , Hill J G
Published: January 2020

We describe an assay system for measuring phenobarbital, diphenylhydantoin, primidone, ethosuximide, and carbamazepine in 25 mul of serum. The procedure involves precipitation of proteins with an acetonitrile solution containing cyheptamide as an internal standard, and reverse-phase chromatography on a 4 mm X 30 cm column containing "muBondapak C18." The anticonvulsants are eluted with an equivolume mixture of potassium phosphate buffer (10 mmol/liter, pH 8.0) and acetonitrile at a flow rate of 0.8 ml/min, detected by their absorbance at 200 nm, and quantitated by measuring peak areas. When measurement of primidone is not required, a 254-nm detector may be used. Each analysis requires 10 min. Optimum column temperature has been found to decrease with use. Analytical recoveries for the five drugs varied from 92% to 102% with good precision (coefficients of variation between 2.8% and 9.2% for therapeutic and toxic concentrations). The results obtained with this method compare favorably with results obtained by gas-liquid chromatography (correlation coefficients between 0.77 and 0.98). In over 1300 patients' samples analyzed to date, the only drugs known to have interfered with the assay are gentamicin, diazoxide, and mephobarbital.
Method for alcohol determination in biological liquids by sensing with a solid-state detector.
Authors: Dubowski K M
Published: January 2020

I describe a new method for quantitative alcohol determination in equilibrated headspace gas of biological liquids by means of a solid-state metal-oxide semiconductor (MOS) detector. After equilibration of the NaCl-saturated specimen at 40 degrees C or other convenient temperature in a closed vessel, a Taguchi MOS sensor is momentarily exposed to the headspace vapor. The resulting dc voltage change from the baseline conditions accurately reflects the alcohol concentration of the liquid specimen, and can be read in direct alcohol-concentration units. Calibration is linear to 3.00 g/liter (300 mg/dl). Values for analytical recovery ranged from 98 to 102%. Correlation of results for blood alcohol concentration by this method (yi) and an automated gas-chromatographic reference method (xi) was y, g/liter = 0.991x + 0.014, with r = 0.997 for 103 blood specimens having concentrations ranging from 0 to 2.89 g/liter (0-289 mg/dl), with a mean difference (xi - yi) - -6 mg/liter (0.6 mg/dl). The method is sensitive to 0.01 g/liter (1 mg/dl) and can be readily automated. An internal standard is not required.
Diazepam abuse: incidence, rapid screening, and confirming methods.
Authors: Rejent T A , Wahl K C
Published: January 2020

We studied 2500 patients suspected to be drug-overdose victims. Blood samples were quantitatively screened for the most commonly abused drugs, including diazepam. Of these, 61% had positive findings, including diazepam in about one of every four. A new rapid, simple, and quantitative gas-chromatographic method for simultaneous analysis of diazepam and sedatives (in two instruments) is described. A single extraction at low pH is used, preserving the balance of the sample to be used for confirming methods via ultraviolet spectrophotometry and thin-layer chromatography. Prevalance of other positive findings is also listed, and findings for diazepam are categorized by age.
Improved micro-radioimmunoassay of digoxin in serum, with use of 125I-labeled digoxin.
Authors: Calesnick B , Dinan A
Published: January 2020

A micro-radioimmunoassay of 125I-labeled digoxin is described in which 10-mul rather than 50-mul aliquots of sera are required. The method is a modification of a simplified, rapid, and accurate procedure, which is commercially available as a kit. There is excellent correlation between results by these two methods, in which lyophilized digoxin standards and insoluble antibody polymer are used. The digoxin concentration in capillary sera was not significantly different from that in the corresponding venous blood. It is clinically useful to monitor digoxin therapy in capillary sera, particularly in patients in whom blood sampling by venipuncture is inconvenient or difficult.
Determination of carbamazepine in blood or plasma by high-pressure liquid chromatography.
Authors: Kabra P M , Marton L J
Published: January 2020

We described a sensitive and precise high-pressure liquid-chromatographic method in which 5-(p-methylphenyl)-5-phenylhydantoin is used as the internal standard in determining carbamazepine in whole blood or plasma. Carbamazepine is well separated from normal blood constituents in less than 8 min, and other commonly used anticonvulsants do not interfere with the analysis. The sensitivity of this method is adequate to quantitate 0.25 mg of carbamazepine per liter in 2 ml of sample, and the lower limit of detection is 100 ng. Twenty specimens were analyzed by a gas-chromatographic method and by the present method; the resulting correlation coefficient was greater than .980.
Evaluation of adenosine 5'-monophosphate and fluoride as adenylate kinase inhibitors in the creatine kinase assay.
Authors: Rosano T G , Clayson K J , Strandjord P E
Published: January 2020

Adenylate kinase (EC 2.7.4.3) interferes positively in the serum creatine kinase (EC 2.7.3.2) assay when the rate of ATP production is monitored by a coupled enzyme system. A dual assay, measuring creatine kinase and adenylate kinase activity, was used to evaluate AMP and other possible adenylate kinase inhibitors that would permit specific measurement of creatine kinase activity in the presence of adenylate kinase. We found that AMP, routinely included in the creatine kinase assay system to inhibit adenylate kinase, partially inhibits both human serum creatine kinase and purified creatine kinase from rabbit muscle. The amount of creatine kinase inhibition is related directly to the AMP concentration and inversely to the substrate (ADP) concentration. We found that 25 mmol/liter of fluoride inhibits adenylate kinase without measurable effect on creatine kinase activity. We developed a serum creatine kinase assay including fluoride, and compared it with the dual assay system and with two commercial assay kits. Other halides or adenosine 2'-monophosphate did not selectively inhibit adenylate kinase.
Rate of decay and distribution volume of MB isoenzyme of creatine kinase, intravenously injected into the baboon.
Authors: Yasmineh W G , Pyle R B , Nicoloff D M
Published: January 2020

We determined the decay constant and distribution space of the MB isoenzyme of creatine kinase after intravenous injection of 440 to 720 U, in a group of seven adult baboons. The rate of decay of the isoenzyme was monoexponential, with a mean decay constant of 0.00175 min-1 (SD, 0.00019). The mean distribution volume obtained by extrapolation of the monoexponential curve to zero time was 5.22% (SD, 1.06) of body weight. We have determined these parameters with the purpose of using this isoenzyme in the baboon as an experimental model for measuring myocardial infarct size.
Direct use of ion-exchange papers in hemagglutination-inhibition tests for drug abuse.
Authors: Alexander G J
Published: January 2020

I modified a procedure for screening urine specimens for evidence of drug abuse, to permit direct use of commercially available ion-exchange papers in hemagglutination-inhibition assay for drug detection. Small circles punched out of the papers, which previously had been impregnated with drug-containing urine, were used in the detection step, leaving the remainder of the specimens on paper available for confirmatory analyses. The procedure is useful in screening for the presence of opiates, methadone, and (or) barbiturates.
Alkaline phosphatase. II. Conditions affecting determination of total activity in serum.
Authors: Van Belle H
Published: January 2020

Results of previous experiments on isolated purified isoenzymes of alkaline phosphatase from humans have been confirmed on sera containing relatively large activities of the different isoenzymes. The most remarkable finding is that activation by N-ethylaminoethanol is much more pronounced, in the case of the intestinal and placental isoenzymes, than is activation by diethanolamine. For several reasons, it is suggested that N-ethylaminoethanol is the buffer of choice, 0.1 mol/liter concentration for routine measurements and 1 mol/liter in those cases where the determination of the intestinal or placental isoenzymes is important. Mg2+ could be omitted because its addition increases the activity only marginally. Normal values for adults with use of 0.1 mol/liter N-ethylaminoethanol are 59 +/- 36 (2 SD) U/liter (n = 126).
Immunofixation electrophoretic techniques applied to identification of proteins in serum and cerebrospinal fluid.
Authors: Cawley L P , Minard B J , Tourtellotte W W , Ma B I , Chelle C
Published: January 2020

We describe the application of immunofixation staining of agarose-gel electrophoretograms in areas where its use in the clinical laboratory is appropriate. Immunofixation electrophoresis consists of an electrophoretic phase followed by a fixation phase in which antiserum is used to precipitate the protein. As long as the antibody is in slight excess or near equivalency, the antigen/antibody complex remains insoluble. The reaction can be detected by visual inspection in indirect light, by protein staining, or by use of antibodies labeled with fluorescein, enzyme, or isotope. In the method described here we primarily have used protein staining (Coomassie Blue) to accentuate the proteins fixed by antisera. All unreacted proteins are removed by pressing with filter paper and saline washing. In the clinical laboratory, this method expedites immunochemical evaluation of samples and may also supplement immunoelectrophoresis. It has been applied successfully in identifying small obscure monoclonal proteins in the serum and cerebrospinal fluid of patients with multiple sclerosis, subacute sclerosing panencephalitis, biclonal gammopathies, serum monoclonal light chains, and mobility shifts of certain proteins, particularly of the complement series. Immunofixation demonstrates that the protein bands present in spinal fluid from multiple sclerosis and subacute sclerosing panencephalitis patients are of the IgG class of immunoglobulins; and non-IgG protein, such as beta and gamma trace proteins, are not detected. We also comment on reverse immunofixation with labeled antigen as a branch of the procedure that allows detection of function of the immunoglobulins separated by electrophoresis.
Urinary organic acids in man. III. Quantitative ranges and patterns of excretion in a normal population.
Authors: Chalmers R A , Healy M J , Lawson A M , Hart J T , Watts R W
Published: January 2020

We studied the urinary excretion of organic acids by normal human subjects in an ambulant normal population, chosen without conscious bias. Quantitative ranges and frequency distribution patterns of excretion are reported, as a basis on which to compare results obtained for patients whose clinical condition suggests that their excretion values may be abnormal. The quantitative valuse we observed enable the relative significance of different urinary anionic metabolites to be assessed, including the recently identified aldonic and deoxyaldonic acids. The frequency distribution patterns observed are discussed briefly.
Purification of alpha1-fetoprotein.
Authors: Twomey S L , Sweet R V
Published: January 2020

Previously published methods for purifying alpha1-fetoprotein are inadequate because they either do not yield a completely pure product or they cause some denaturation. We present a method that does not have these serious disadvantages, and with which alpha1-fetoprotein was purified by sequential use of concanvalin A affinity-chromatography, preparative gel-electrophoresis, and immunoabsorption with anti-albumin antibody covalently coupled to Sepharose 4B. The purity of the product was monitored by discontinuous polyacrylamide-gel electrophoresis and counterimmunoelectrophoresis, both of which must be used to ascertain what proteins are present at each step of the purification.
Simplified, totally enzymatic method for determination of serum triglycerides with a centrifugal analyzer.
Authors: Grossman S H , Mollo E , Ertingshausen G
Published: January 2020

We describe a totally enzymatic method for determination of serum triglycerides (triacylglycerols) specifically adaptable to the CentrifiChem system. The method involves lipolysis with lipase from Rhizopus arrhizus alone and quantitation of the resulting glycerol with glycerol dehydrogenase in a kinetic, fixed-time mode. Hydrolysis by the lipase is complete, for concentrations up to at least 5.0 g/liter, in 10 min at room temperature. The unfavorable equilibrium for the oxidation of glycerol is overcome by increasing the pH and adding excess NAD+. Under these conditions the glycerol determination is linear to at least 4.0 g of glycerol per liter, as triglyceride. The test exhibits acceptable accuracy and precision, and results correlate well with those by an alternative totally enzymatic procedure. The present method is unaffected by phosphatase and a considerably simplified reagent is used.
Rapid-flow enthalpimetric determination of urea in serum, with use of an immobilized urease reactor.
Authors: Bowers L D , Canning L M , Sayers C N , Carr P W
Published: January 2020

We used a differential thermal detector in conjunction with an immobilized urease reactor to determine urea in serum. Samples (120 mul) are introduced into a flow stream and passed through an "adiabatic" column, which is packed with enough insolubilized urease to completely convert urea to ammonia and carbon dioxide. Measured temperature changes are directly proportional to the serum urea concentration. Urea in the presence of protein, bilirubin, and hemoglobin can thus be rapidly, simply, and inexpensively measured. Results correlate well with those obtained by the manual diacetyl monoxime and urease/indophenol methods.
Propoxyphene and norpropoxyphene concentrations in blood and tissues in cases of fatal overdose.
Authors: McBay A J
Published: January 2020

Propoxyphene and its major metabolite, norpropoxyphene, have been quantitated in tissue specimens obtained from autopsies of people suspected of dying from propoxyphene overdosage. Gas-chromatographic determination of both propoxyphene and norpropoxyphene is essential because the blood concentration of the parent drug should be about 1.0 mg/liter or greater to attribute a death to the drug. The metabolite concentration in blood may help to establish when the drug was ingested. Concentrations in the blood after high oral therapeutic doses are about 0.3 mg of propoxyphene per liter, and norpropoxyphene concentrations may be as high as 3 mg/liter. Methods of determining propoxyphene are discussed.
Purification and analysis of the purity of NADH.
Authors: Margolis S A , Howell B F , Schaffer R
Published: January 2020

We developed an analytical reverse-phase high-performance liquid chromatographic procedure for rapid assessment of the purity of NADH. The method completely separates adenosine monophosphate and adenosine diphosphoribose from NADH. By use of this analytical technique we found that preparative chromatography on DEAE-cellulose gives NADH that is free from adenine nucleotides as well as other impurities that commonly are present in NADH. The absorbance ratio at 260 and 340 nm of the purified NADH in 1.8 mmol/liter ammonium carbonate is 2.261 +/- 0.002 (+/- 1 SD).