Quantitation of vancomycin and its crystalline degradation product (CDP-1) in human serum by high performance liquid chromatography.

The delayed clearance of vancomycin results in accumulation of vancomycin crystalline degradation product, CDP-1, in the bodies of renally impaired patients. The 2 isomers, CDP-1-M (major) and CDP-1-m (minor), of CDP-1 are antibiotically inactive but cross-react with some immunoassays that use polyclonal antibodies resulting in falsely elevated results. A high performance liquid chromatographic (HPLC) method was developed to quantitate vancomycin and CDP-1 in the serum of renal patients. After solid phase extraction of 200 microliters serum, the separation of vancomycin, the 2 isomers of CDP-1 and the internal standard (cefazolin) was accomplished by gradient HPLC on a reversed phase C18 column with detection at 210 nm. Linearity was established from 1 to 25 and 25 to 100 micrograms ml-1 vancomycin and 1 to 25 micrograms ml-1 CDP-1. Coefficients of variation for vancomycin and CDP-1 were 3.3-8.6% (n = 10) and 2.8-5.2% (n = 8).

A specific method for the measurement of tacrolimus in human whole blood by liquid chromatography/tandem mass spectrometry.

Tacrolimus (FK 506) measurement by immunoassays in clinical samples of organ transplant patients often lacks a specific reference method. A method combining liquid chromatography (LC) with tandem mass spectrometry (MS/MS) was developed to quantify tacrolimus in whole blood. Liquid-liquid extraction was performed on 1 ml of sample before narrow-bore LC/MS/MS analysis. Ascomycin was used as an internal standard. The standard curve was composed of seven points ranging from 1 to 50 micrograms/l (average r2 = 0.9999). Limits of detection and quantitation were 0.25 and 0.75 microgram/l, respectively. Imprecision was < 5% across the therapeutic range. Tacrolimus recovery averaged 62%. The most abundant metabolites detected in clinical samples were 13-O- and 15-O-demethyl tacrolimus. This method was used in a comparison study with a microparticle enzyme immunoassay (MEIA): MEIA = 1.03 LC/MS/MS -0.084 (microgram/l), (Sy/x = 1.43), r2 = 0.933. With its high sensitivity and specificity, this LC/MS/MS method presents a good reference method for immunoassay evaluation as well as a valuable tool for metabolism studies.