β Scribed by H.J. Weder; M.H. Bickel
No coin nor oath required. For personal study only.
A sensitive method for the determination of imipramine and seven of its major metabolites in biological samples is described. The extraction procedures used are based on the partition properties which were determined for each metabolite. The extracted drug and its metabolites 'are in. some cases transformed into derivatives. They are finally separated and'determined by gas chromatography. The sensitivity reached is o.or-0.05 rug, 'except for z-hydroxy-desmethylimipramine (0.2-0.3 lug). Examples 'of analyses using tissues; bile, and liver perfusion medium are given.' > Several author&lo have described the determination of. imipramine and some of 'its metabolites in. biological materials. Thin layer chromatography~lends itself to the detection of ali known imipramine metabolites, though the' quantitative evaluation methodsll~ 12 are of limited accuracy. Thin layer chromatography followed by elution and U.V.-spectrophotometry proved not. sensitive enough for studies of imipramine metabolism. Gas-liquid chromatography (GLC) , due to its high sensitivity, proved to be thejmethod of choice'for obtaining accurate values in metabolic exper; iments. Gas. chromatography has been' used for the detection of imipramine and desipramine as early as rggr by GILLETTE et aZ. 13, More recently. gas chromatography has been used I for the: deten'nination of the' antidepressant dibenzepine and ,six metabolites by LEI~NER et al;14 and for phenothiazines by MCMARTIN AND' STREE~~~. .'
In 'addition to GLC ,procedures this paper also deals with the ,determination',of the partition coefficients of imipramine and' its metabolites between organic: solvents and aqueous buffers. Based on these values suitable extraction procedures have been developed ,for tissues, bile, and perfusion; medium. Simple methods. for the synthesis of derivatives to be used for the final gas chromatography are also described.
MATERIALS AND METHODS
'. : ..: Imipramine(IP) , desmethylimipramine (DMI) , desdimethylimipramine (DDMI) , 2-hydroxy&nipramine (2-OH-IP) ; 2-hydroxy-desmethylimipramine, (2-OH-DMI)', i '. J. Clbrimatog.,' '.
Isotonic phosphate bufferlo, pH 7.4, and ut-hexane or chloroform, respectively, were used, Previous to partition, of imipramine or its metabolites, the phases' were '. saturated with, respect to each'other by shaking for 5 min. 'The substance was then dissolved in ,the aqueous phase (5, ml) and shaken with the organic'phase (5, ml). in',a centrifuge tube (20 ml) for 30 min at zoo.' *The initial concentration in the, aqueous phase. and the final concentrations after equilibration were determined with-a Unicam' ,' ,U.V.-spectrophotometer SP 800.' The variation of the partition coefficients as a function of plf-I was determined by ,,using ,r,z-dichloroethanc, diethyl ether (peroxide-free) and tin-heptane as organic *phases, phosphate buffers (pH 5.0-7.0), borax buffers17 (pII 7.0-8.8) and glyciue buffersl* (pH g.o-r?r4) as aqueous phases.
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π SIMILAR VOLUMES
Encainide (ENC) and its metabolites O-demethylencainide (ODE), 3-methoxy-O-demethylencainide (MODE), N-demethylencainide (NDE) and bis-N,O-demethylencainide (NODE) have been measured by two HPLC procedures. The method of Mayol using a mu Porasil column with ethanol-water-methanesulphonic acid as mob