CARBOHYDRATE ANALYSIS OF FERMENTATION BROTH BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY UTILIZING SOLID-PHASE EXTRACTION

Nutrient information concerning the specific needs in fermentation broth is crucial to the potency and production of an antibiotic. Nutrient analysis of carbohydrate metabolism by HPLC is hindered by an array of fermentation broth interferences. These interferences can be effectively removed by solid-phase extraction. The fermentation broth was diluted with an equal amount of water and vortexed. The sample mixture was eluted through an activated Acell QMA solid-phase cartridge. The resultant eluent was diluted with an equal amount of a acetonitrile-ethanol (75:25) mixture, vortexed and filtered. The sample extract was chromatographed on a YMC polyamine column using refractive index detection. The mobile phase consisted of acetonitrile-water (75:25) at a flow-rate of 1 ml/min. Under these conditions, glucose, fructose, sucrose and maltose were separated in less than 13 min with baseline resolution. This method is capable of monitoring the carbohydrate consumption and metabolism throughout the fermentation process.