Liquid Chromatographic Determination of Cyclosporine-A in Blood,with Chromosorb P Columns Used for Sample Purification

Abstract

A fast liquid-chromatographic determination of whole-blood cyclosporine A concenration is described. A sample preparation consisting of diethyl ether extraction and Chromosorb P column purification of extract, requires 1 mL of whole blood and takes 10 min of technical effort per sample. A reversed-phase C₁ column (5-μm particles) is used, with acetonitrile/methanol/water (20/45/35 by vol) as mobile phase. Cyclosporine A is quantified by absorbance at 214 nm, with cyclosporine D as internal standard. Chromatographic development is complete in 8 min. Linearity is verified by a five-point calibration curve in the range 50–900 μg/L, correlation coefficient r>0.998, y = 0.001x ? 0.053. Lower limit of sensitivity is 25 μg/L. Extraction efficiency was over 70%, accuracy varied between ?7.1% and +3.3%s, CVs were <5.8% within run, <7.5% between runs. No interference was observed from both endogenous compounds, and 33 drugs eventually co-administered during immunosuppression. Over 1,000 patient samples have been analysed by this method in our laboratory in about one year, without any sign of column deterioration.