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Enantioselective HPLC determination of R and S trimipramine in human serum using an octyldecylsilane column with beta-cyclodextrin as mobile phase additive and solid phase extraction



Enantioselective HPLC determination of R and S trimipramine in human serum using an octyldecylsilane column with beta-cyclodextrin as mobile phase additive and solid phase extraction



Journal of Liquid Chromatography & Related Technologies 20(19): 3107-3119



A stereospecific HPLC method was developed for the analysis of the enantiomers of trimipramine in human serum. The assay uses amitriptyline as the internal standard and a C-18 solid phase extraction column for serum sample clean-up. It is free of interference from desmethyl-trimipramine, 2-hydroxy-desmethyl-trimipramine and 2-hydroxy-trimipramine, the three major metabolites of trimipramine. Recoveries of 98.8% and 97.5% were obtained for the R and S enantiomers of trimipramine. respectively. Resolution of the enantiomers was obtained using an octyldecylsilane column with beta-cyclodextrin as the mobile phase additive. The composition of the mobile phase was 80:20 v/v aqueous 10 mM ammonium acetate buffer pH 4 (adjusted with acetic acid)- absolute ethanol containing 20 mM beta-cyclodextrin at a flow rate of 0.7 mL/min. Linear calibration curves were obtained in the 25-400 ng/mL range for each enantiomer in serum. The detection limit based on a S/N = 3 was 10ng/mL for each enantiomer in serum with UV detection at 220 nm. The limit of quantitation for each enantiomer was 25 ng/mL. Precision calculated as %RSD and accuracy calculated as % error were in the range 0.7-4.5% and 0.9-3.1%, respectively, for the R enantiomer and 0.7-5.1% and 0.4-4.4% respectively, for the S enantiomer. Separation of the three major metabolites of trimipramine was also investigated.

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