STEREOSPECIFIC DETERMINATION OF AMISULPRIDE, A NEW BENZAMIDE DERIVATIVE, IN HUMAN PLASMA AND URINE BY AUTOMATED SOLID-PHASE EXTRACTION AND LIQUID-CHROMATOGRAPHY ON A CHIRAL COLUMN - APPLICATION TO PHARMACOKINETICS

Amisulpride, a drug belonging to the benzamide series, demonstrates an tischizophrenic and antidepressant (antidysthymic) properties in man. For the pharmacokinetic studies of the racemic drug in man, a method o f determination based on solid-phase extraction (SPE) from plasma and HPLC on a Stereoselective column was developed. For this aim, one mill ilitre of plasma, after the addition of the internal standard, tiaprid e or metoclopramide, is diluted with a berate buffer at pH 9, then aut omatically loaded onto a SPE C-18 100-mg column. The column is washed with different solvents, then eluted with 0.5 ml of methanol. After ev aporation of the eluted fraction, the residue is reconstituted in 0.25 mi of eluent mixture. An aliquot is injected onto the HPLC column, a Chiralpak AS, equilibrated with an eluent mixture constituted by n-hex ane-ethanol, (67:33, v/v) containing 0.2% (v/v) of diethylamine (DEA) or n-heptane-ethanol, (70:29.8, v/v) containing 0.2% of DEA and connec ted to a UV detector set at 280 nm or to a fluorimetric detector set a t lambda(ex)=280 nm and lambda(em)=370 nm. The limit of quantitation ( LOQ) in human plasma is 2.5 ng ml(-1) for both S-(-)- and R-(+)-amisul pride isomers with both detection methods. The method has been demonst rated to be linear in the range 2.5-320 ng ml(-1) for both R-(+)- and S-(-)-amisulpride in human plasma with both UV and fluorescence detect ion. Absolute recovery of S-(-)- and R-(+)-amisulpride enantiomers fro m human plasma, as well as selectivity, precision and accuracy have be en demonstrated to be satisfactory for pharmacokinetics in man and equ ivalent for both the proposed methods that have been cross-validated o n real dosed human plasma samples. The methods have bees used for clin ical pharmacokinetic studies allowing pharmacokinetic parameters for a misulpride enantiomers in agreement with those obtained for the racema te to be obtained. After dilution with water, urinary samples from sub jects treated with-amisulpride racemate can be analysed according to t he method used for plasma.