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High-performance liquid chromatographic methods have been developed for the determination of ampicillin (ABPC), amoxicillin (AMPC) and ciclacillin (ACPC) in serum and urine. The methods involve acetylation of these aminopenicillins with acetic anhydride in aqueous solutions (pH 9.0) at ambient temperature for 3 min followed by reaction with 2 M 1,2,4-triazole and 10-3 M mercury(l1) chloride in solution (pH 9.0) at 60 "C for 10 min. The resulting products were separated on a C18 column following stabilisation in an eluent containing sodium thiosulphate. They were detected at 328 nm for ABPC and AMPC and 327 nm for ACPC.The methods have been applied to assays of these aminopenicillins in serum and urine. The procedures, which permit the accurate determination of aminopenicillin concentrations in serum down to 0.05 pg ml-1 for AMPC and 0.1 pg ml-1 for ABPC and ACPC, are specific to intact penicillins without interference from the corresponding penicilloates. At an aminopenicillin concentration of 1 pg ml-1 in serum, the within-and between-run precisions were 0.97-3.51 % and 2.07-3.55%, respectively.

A new internal-surface reversed-phase (ISRP) silica support has been designed for direct injection analysis of drugs in biological fluids by liquid chromatography. The support, prepared by using a new enzyme, polymyxin acylase, has N-octanoylaminopropyl phases, bound only to the internal surfaces of the porous silica, and N-(2,3-dihydroxypropyl)-aminopropyl phases on the external surfaces in order to be nonadsorptive to proteins. The average pore diameter of the prepared ISRP silica support was 50 A, which is small enough to exclude macromolecules such as serum proteins from the pores. The new ISRP support can be used for the direct injection analysis of hydrophilic and hydrophobic drugs in serum or plasma without destructive accumulation of proteins over the eluent pH range of 3 to 7. The recovery of drugs from serum was almost 100%, regardless of the difference in their protein bindings.

A new beta-cyclodextrin (CD) bonded silica has been designed for direct injection analysis of drug enantiomers in serum for liquid chromatography. The new beta-CD bonded silica is synthesized by three steps: introduction of a 3-glycidoxypropyl phase, introduction of a beta-CD carbamate-bonded phase, and hydrolysis of the oxirane ring to a diol phase. Although the recovery of serum proteins from only a beta-CD bonded silica (having no diol phase) was poor, they were completely recovered from a mixed functional silica having beta-CD and diol phases. Direct injection analysis of drug enantiomers in serum can be attained with the mixed functional silica support over the eluent pH range of 3-7. The recovery of racemic drugs from serum was almost 100%.

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