A method for the rapid, quantitative determination of 5-fluorocytosine (5-FC) in serum by high-performance liquid chromatography (HPLC) has been developed. After initially ultrafiltrating the serum, a portion was injected onto a cation exchange column. 5-FC was separated by using an ammonium-phosphate buffer as the mobile phase and detected by ultraviolet absorption at 254 nm. Quantitation of 5-FC was based on the linear relationship between peak area in the chromatograms and known concentrations of 5-FC in a set of serum standards (prepared by adding from 10 to 200 ,ig of 5-FC to 1-ml aliquots of pooled human serum). This method was compared with the standard microbiological method for 5-FC. Advantages of the HPLC method include: (i) Determination of 5-FC levels within 30 min; (ii) lack of interference from other antimicrobial drugs, particularly amphotericin B; (iii) more accurate determination of true 5-FC level, particularly at concentrations of <25 ug/ml or >100 ,tg/ml; and (iv) ease with which the assay may be automated for routine use.

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