Determination of benzylpenicillin in plasma and lymph at the ng ml-1 level by reversed-phase liquid chromatography in combination with digital subtraction chromatography technique.
A method for the determination of benzylpenicillin (Pc-G) at very low levels in plasma and lymph is described. Detection at 325 nm of the mercuric mercaptide of benzylpenicillinic acid was made by liquid chromatography via a pre-column derivatization method. By using a digital subtraction chromatography technique, the bioanalytical method could be applied to different kinds of samples whether interfering peaks were present or not. Two different clean-up steps were used for two concentration ranges 0.1-100 micrograms and 1-1000 ng Pc-G ml-1; namely, precipitation of a 100-microliters sample with acetonitrile, or precipitation of a 1- or 2-ml sample followed by concentration via liquid-liquid extraction. The pre-column derivative of Pc-G was achieved using mercury(II)chloride in the presence of imidazole. The blank samples required for the digital subtraction chromatography technique were obtained by penicillinase treatment. Standard curves were made in the two concentration ranges. The relative standard deviation (RSD) at 5 ng Pc-G ml-1 plasma was 4.9% and at 5 micrograms Pc-G ml-1 lymph it was 2.8%. The stability of Pc-G, including the problems with non-sterile samples, was studied.
Wiese
B
B
Department of Pharmacology and Toxicology, Faculty of Veterinary Medicine, Swedish University of Agricultural Sciences, Uppsala.
Martin
K
K
D000818:Animals; D002417:Cattle; D002853:Chromatography, Liquid; D005260:Female; D008196:Lymph / Q000737:chemistry*; D010400:Penicillin G / Q000031:analogs & derivatives / Q000032:analysis* / Q000097:blood / Q000493:pharmacokinetics
C016262:benzylpenicillenic acid; D010400:Penicillin G