Determination of mannose-6-phosphate in pharmaceutical intermediates by high performance ion-exchange chromatography-electrochemical detection

Abstract

A novel method was developed for the determination of mannose-6-phosphate and the phosphate in pharmaceutical intermediates by high performance ion-exchange chromatography (IC) and electrochemical detection. The sample was dissolved with purified water, filtrated by a 0.22 microm nylon filter membrane, and then separated by ion-exchange chromatography. The separation was performed on an IonPac AS18 column (250 mm x4 mm) with an IonPac AG18 (50 mm x4 mm) as guard column, and 25 mmol/L KOH solution as the eluent at a flow rate of 1.0 mL/min. The detection was performed with electrochemical detectors (an integrated pulsed amperometric detector and a suppressed conductivity detector in line). Mannose-6-phosphate was selectively determined with an integrated pulsed amperometric detector first. Then, both of mannose-6-phosphate and phosphate were determined with a suppressed conductivity detector after the background conductance of KOH was suppressed with the ASRS suppressor. The injection volume was 25 microL. The external standard calibration curve was used for quantitative analysis. With an amperometric detector, the linear range of the method for mannose-6-phosphate were 0.06 - 10.0 mg/L (r = 0.9998). The recoveries were 92.1% - 103.1% with the relative standard deviations (RSDs) less than 3%. The detection limit was 0.02 mg/L for mannose-6-phosphate. The method is simple, effective, sensitive and selective. It can be used to determine the quality of pharmaceutical intermediates.