A novel ion-pair RP-HPLC method for determination of five components in compound α-ketoacid tablets

Abstract

A sensitive and reliable ion-pair reversed-phase high-performance liquid chromatographic (RP-HPLC) method has been developed and validated for the simultaneous determination of D,L-α-hydroxymethionine calcium (HMACa), D,L-α-ketoisoleucine calcium (KILCa), α-ketovaline calcium (KVCa), α-ketoleucine calcium (KLCa) and α-ketophenylalanine calcium (KPACa) in the compound α-ketoacid tablets using tetrabutylammonium hydroxide as an ion-pair reagent. The analytes were separated on a C(18) column (250 mm × 4.6 mm, 5 μm) with the mobile phase of methanol-potassium dihydrogen phosphate buffer (pH 3.0; 50mM) (37:63, v/v) at a flow rate of 0.6 mL min(-1). The UV detection wavelength was set at 210 nm. Good linearity with correlation coefficients larger than 0.9990 (n=6) for all analytes were achieved. The average recoveries were within the range of 99.6-100.9%, and the RSDs of the results were within the acceptable limit of 2.0%, which showed that this method was accurate and precise. The limits of detection were 10.44, 5.94, 3.44, 3.60 and 1.63 ng mL(-1), and the limits of quantification were 34.80, 19.80, 10.32, 12.00 and 5.44 ng mL(-1) for HMACa, KILCa, KVCa, KLCa and KPACa, respectively. The method is simple and accurate for quality control of the compound α-ketoacid tablets.