Development and characterization of the α3ß4α5 nicotinic receptor cellular membrane affinity chromatography column and its application for on line screening of plant extracts.

The α3ß4α5 nAChR has been recently shown to be a useful target for smoking cessation pharmacotherapies. Herein, we report on the development and characterization of the α3ß4α5 nicotinic receptor column by frontal displacement chromatography. The binding affinity of the nicotine and minor alkaloids found in tobacco smoke condensates were determined for both the α3ß4 and α3ß4α5 nicotinic receptors. It was demonstrated that while no subtype selectivity was observed for nicotine and nornicotine, anabasine was selective for the α3ß4α5 nicotinic receptor. The non-competitive inhibitor binding site was also studied and it was demonstrated while mecamylamine was not selective between subtypes, buproprion showed subtype selectivity for the α3ß4 nicotinic receptor. The application of this methodology to complex mixtures was then carried out by screening aqueous-alcoholic solutions of targeted plant extracts, including Lycopodium clavatum L. (Lycopodiaceae) and Trigonella foenum graecum L. (Fabaceae) against both the α3ß4 and α3ß4α5 nAChRs.

Pharmacophore modelling of stereoselective binding to the human organic cation transporter (hOCT1).

BACKGROUND AND PURPOSE: The human organic cation transporter-1 (hOCT1) is a polyspecific transporter that plays a role in drug distribution, metabolism and excretion. Previous studies have demonstrated that hOCT1 binding can be stereoselective, but the mechanism for stereochemical recognition has not been described. The purpose of this study was to develop a pharmacophore model to describe stereoselective binding to hOCT1. EXPERIMENTAL APPROACH: A set of 22 compounds including 8 pairs of enantiomers and five pairs of diastereomers was used to develop a pharmacophore model. The pharmacophore modeling was carried out using Catalyst version 4.11 and HypoGen and was based upon the correlation of the structures and activities (K(i) values) of the compounds used in the study. KEY RESULTS: The resulting model contained a positive ion, hydrophobic and two hydrogen-bond acceptor interaction sites. The relative enantioselectivity of 8/8 enantiomeric pairs and diastereoselectivity of 5/5 diastereomers was described by mapping to a combination of at least 3 of the 4 functional feature sites of the model. CONCLUSIONS AND IMPLICATIONS: The pharmacophore model describes stereoselective interactions with hOCT1 at one of the binding sites on the molecule.

Determination of clemastine in human plasma by gas chromatography with nitrogen-phosphorus detection.

A method for the quantitative determination of clemastine in human plasma has been developed and validated. The assay uses gas chromatography with nitrogen-phosphorus detection and a HP-1 capillary column (25 mx0.22 mm, film thickness 0.33 mm) coated with dimethylpolysiloxane. Clemastine (with orphenadrine as internal standard) was isolated from human plasma using liquid-liquid extraction. A linear relationship was observed between 0.1 and 12.8 ng/ml using the peak area ratio of clemastine to orphenadrine with a correlation coefficient greater than 0.99 (the detection limit for clemastine was 0.06 ng/ml). The intra- and inter-day coefficients of variation were less than 11%. The developed method was used for the analysis of plasma samples from healthy volunteers (n = 19) to examine the pharmacokinetics of the antihistamine clemastine after single and multiple oral doses of clemastine fumarate.

Removal of methotrexate, leucovorin, and their metabolites by combined hemodialysis and hemoperfusion.

This article documents the case of a patient with severe renal failure immediately after having been given high-dose methotrexate; the patient was effectively treated with repeated hemodialysis, charcoal hemoperfusion, leucovorin, and thymidine. The methotrexate plasma concentration was reduced from 390 mumol/L to 7 mumol/L as a result of 24.5 hours of hemodialysis along with 39.5 hours of hemoperfusion. Although a rebound in the plasma methotrexate concentration occurred the first three times that hemodialysis and/or hemoperfusion was stopped, reinstitution of the procedure was always effective in further lowering methotrexate concentrations. The patient was subsequently managed with leucovorin and thymidine rescue. Simultaneous measurements before and after the hemodialysis-hemoperfusion apparatus and before and after the hemoperfusion device alone revealed a percent decrease in the concentration of d-leucovorin of 36% and 79%; 1-leucovorin, 82% and 75%; 5-methyltetrahydrofolate, 52% and 64%; methotrexate, 73% and 37%; and 7-hydroxymethotrexate, 21% and 24%, respectively. Gastrointestinal and hematologic toxicities were completely prevented, and serum creatinine normalized within 24 days.

Recent advances on endogenous Na+,K+-ATPase inhibitors: clinical investigation and purification.

Evidence exists which demonstrates the relationship between a Natriuretic Factor or Na+,K+-ATPase inhibitor and volemic expansion, both in man and animal. Patients having extracellular volume expansion have been studied for the effect of their plasma on erythrocytes 3H-ouabain binding. High levels of ouabain-like activity was found in plasma from acromegalic patients and patients with chronic renal failure. High levels were also observed in some hypertensive patients. A partial purification of such a compound was performed from urine of hypertensives. The partially purified compound inhibited to a greater extent the Na+,K+-ATPase semi-purified from dog kidney than that from sheep brain. The present data are consistent with the possible regulation of the activity or the secretion of plasma ouabain-like activity by extracellular volume.