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Biomedical subjects

L M Shaw

Publications and source records attributed to L M Shaw.

172 records · Page 10Linked to original sources
PubMed1975

A systematic approach to enzyme assay optimization illustrated by aminotransferase assays.

We have developed a systematic approach to optimization of reagent concentrations for assays of alanine aminotransferase and aspartate aminotransferase: (a) Michaelis constants describing the initial-velocity kinetics of the coupled enzyme reactions were evaluated by a nonlinear least-squares fit of the appropriate equation to measured enzyme activities. Activities of more than 50 normal and pathological sera were measured at 30 degrees C. (b) These kinetic equations are used to calculate the set of reagent amino- and keto-acid concentrations that all yield a selected fraction of the theoretical maximum enzyme velocity. An optimal pair is determined by defining an additional criterion, such as minimal reagent cost or minimal concentration to Km ratio. (c) The optimum amounts of reagent NADH and coupling enzyme, being a function of desired pre-incubation and measurement intervals, maximum aminotransferase activity to be measured, and endogenous keto-acid concentration, are determined by computer simulation. An approximate relationship and an exact method for computing assay lag time are presented, along with experimentally measured endogenous keto-acid concentrations in serum. All procedures may be applied to other enzyme assays if appropriately modified.

Alanine Transaminase↗
PubMedDate not supplied

Routine monitoring of cyclosporine in whole blood and in kidney tissue using high performance liquid chromatography.

A simple isocratic reversed-phase high pressure liquid chromatographic procedure for the specific estimation of cyclosporine in 1 mL of whole blood using cyclosproin D as internal standard is described. The chromatographic conditions chosen afford a high degree of resolution and sensitivity, a prolonged analytical column life (four months), and complete analysis of ether extracts within 8 min. The limit of detection is 10 micrograms/L. Thirty-five patient samples can be analyzed in a single batch with a turnaround time of 7 hr. This method has been in use in the authors' therapeutic drug monitoring laboratory for nearly two years. It can be simply adapted for the analysis of cyclosporine in kidney samples as a possible aid in the diagnosis of therapeutic failure or toxicity in renal allograft recipients. Examples are given to illustrate the usefulness of frequent monitoring of cyclosporine in stabilizing graft function after organ transplantation.

Adult↗
PubMedDate not supplied

Colchicine poisoning: report of a fatal case with body fluid analysis by GC/MS and histopathologic examination of postmortem tissues.

A suicide caused by oral ingestion of colchicine is reported. Postmortem examination revealed circulating Pelger-Huet polymorphonuclear leukocytes and numerous mitotic and chromatin bodies in tissues with rapid cell turnover. Colchicine was identified by gas chromatography/mass spectrometry (GC/MS) of an organic extract of a urine specimen obtained about 36 h after the patient's original hospitalization. The clinical history and pathology of this rarely encountered intoxication are correlated with previous reports, and the rapid detection of colchicine by GC/MS is discussed.

Adult↗
PubMedDate not supplied

A new centrifugal filtration device for free drug separation.

A new centrifugal membrane filtration device (Syva Corporation) has been evaluated to determine its capabilities for separation of free phenytoin for analysis. The device is a test tube-size cylinder with two compartments separated by the membrane. In serum samples from 70 patients at the Hospital of the University of Pennsylvania, free phenytoin was prepared by the new device and by equilibrium dialysis. Levels were assayed by gas chromatography and enzyme immunoassay with good agreement at all phenytoin levels. Although pH has a significant effect on the binding of some drugs to serum proteins, phenytoin binding increased to only a small extent as pH was increased from 7.0 to 7.8 (85-88.5% bound). Centrifugal filtration with this device is a reliable, fast, and easy way to prepare free drug from serum and does not include the dilution artifact inherent in equilibrium dialysis.

Centrifugation↗
PubMedDate not supplied

Metabolic pathways of WR-2721 (ethyol, amifostine) in the BALB/c mouse.

This study investigated the metabolism of the radio- and chemoprotector compound, WR-2721 [amifostine; s-2-(3- aminopropylamino)ethylphosphorothioate], in the Balb/c mouse. The latter was selected for these studies because considerable radiation protection data have been published for this mouse strain using the WR-2721 dose, route of administration, and optimal time for protection following intraperitoneal injection used herein. It is known that protection requires conversion of the parent drug to its free thiol metabolite, WR-1065, in cultured cells. Because it is possible that metabolites of WR-1065 could be involved in protection and because thiols are metabolically very reactive molecules, we investigated the metabolism of WR-2721 using electrochemical detection-HPLC methods. The following are the major findings in this study: 1) WR-2721 drug was rapidly cleared from the bloodstream. Blood concentration of the parent drug decreased 10-fold 30 min after administration from the maximal observed value at 5 min 2) WR-1065 rapidly appeared in the perchloric acid (PCA)-soluble fraction of normal solid tissues. The highest WR-1065 concentrations in liver and kidney were 965 and 2195 mumol/kg, respectively, 10 min after parent drug administration, whereas for heart and small intestine the highest values were 739 and 410 mumol/kg at 30 min. 3) WR-1065 accumulated in the PCA-soluble fraction of two experimental tumors at a lower rate than for the other tissues.(ABSTRACT TRUNCATED AT 250 WORDS)

Amifostine↗
PubMedDate not supplied

Is the volume of distribution of digoxin reduced in patients with renal dysfunction? Determining digoxin pharmacokinetics by fluorescence polarization immunoassay.

STUDY OBJECTIVE: To determine digoxin pharmacokinetics in subjects with different degrees of renal function using fluorescence polarization immunoassay (FPIA), which is associated with less interference from digoxin-like immunoreactive substances (DLIS) than radioimmunoassay. SETTING: University hospital clinical research center. PARTICIPANTS: Eighteen subjects (mean age 44 yrs) with different degrees of renal function: group 1, creatinine clearance (Clcr) below 10 ml/minute; group 2, Clcr 10-50 ml/minute; and group 3, Clcr greater than 50 ml/minute (6 patients in each group). INTERVENTION: Over 5-7 days, 15 serum samples were collected after a single intravenous dose of digoxin 7 or 10 micrograms/kg actual body weight (WT) for serum concentration measurements by FPIA. Two-compartment pharmacokinetic parameters (zero-time intercept of the concentration-time curve of the initial distribution phase [A], zero-time intercept of the concentration-time curve of the terminal elimination phase [B], initial distribution phase constant [alpha], terminal elimination rate constant [beta], volume of distribution in the central compartment [Vc] and at steady state [Vss], total body clearance [Cl], mean residence time [MRT], area under the concentration-time curve [AUC]) were determined using a nonlinear least squares regression program. MEASUREMENTS AND MAIN RESULTS: No significant differences were found among groups for A, B, alpha, beta, beta-half-life Vc/WT, MRT, AUC, and Cl/WT. Significant differences were observed in Vss/WT (4.8 +/- 1.0, 6.6 +/- 0.5, 6.4 +/- 0.7 L/kg) between group 1 versus group 2 and group 1 versus group 3 (p < 0.01). Measured Clcr was correlated with Cl (r2 = 0.40, p < 0.01), Cl/WT (r2 = 0.29, p < 0.05), Vss (r2 = 0.35, p = 0.01), and Vss/WT (r2 = 0.24, p < 0.05). CONCLUSION: This study confirmed that Vss is smaller in patients with chronic renal failure (Clcr < 10 ml/min) than those without chronic renal failure. Therefore, previous recommendations that lower digoxin loading doses should be administered in patients with renal failure are applicable to digoxin serum concentration monitoring using FPIA.

Adolescent↗