Influence of hydrolysis, purification, and calibration method on furosine determination using ion-pair reversed-phase high-performance liquid chromatography

Abstract

The influence of HCl concentration (6M, 8M, and 10M) and the ratio of sample protein to acid (1 or 5 mg of protein per mL of acid) on furosine formation during sample hydrolysis is studied. The conditions that maximize furosine formation are 10M HCl in the ratio of 1 mg of protein to 1 mL of acid. Purification of the hydrolysate by solid-phase extraction is also considered by examining the effect of hydrolysate volume and volume of 3M HCl used to elute the furosine. Furosine quantitation is carried out using the standard additions and external standard methods. The results indicate that there is no interference by the sample matrix and that external calibration is adequate.