title: High-performance liquid chromatographic assay of pirlimycin in human serum and urine using 9-fluorenylmethylchloroformate.
abstract: A reversed-phase high-performance liquid chromatographic (HPLC) assay method has been developed for determining pirlimycin in human serum and urine. The method involves chloroform extraction of pirlimycin free base followed by derivatization with 9-fluorenylmethylchloroformate to form a carbamate ester. The reaction is rapid, reproducible, and quantitative. 9-Fluorenylmethylchloroformate reacts with amines to form derivatives sensitive to both ultraviolet and fluorescence detection. Human serum and urine samples following 50-mg and 500-mg single oral doses of pirlimycin were analyzed. The samples were chromatographed on an RP-18 Spherisorb 5-micron, 250 X 4.6 mm I.D. reversed-phase HPLC column. The eluent for the serum assay was acetonitrile-water (58:42) containing 0.02% acetic acid, and for the urine assay was acetonitrile-methanol-tetrahydrofuran-water (48:2:1:49). Fluoranthene was used as an internal standard. The assay sensitivity by ultraviolet detection (lambda max = 264) was about 5 ng/ml and by fluorescence detection (lambda excitation = 270 nm, lambda emission = 300 nm) was 0.1 ng/ml. Statistical analysis indicates an average drug recovery of 101 +/- 4.2% from serum and 102.0 +/- 2.62% from urine.
mesh_terms: D055598:Chemical Phenomena; D002621:Chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D002855:Chromatography, Thin Layer; D002981:Clindamycin / Q000031:analogs & derivatives* / Q000032:analysis / Q000097:blood / Q000652:urine; D006801:Humans; D013997:Time Factors
chemical_list: D002981:Clindamycin; C034093:pirlimycin