A Novel Method for the Selective Recovery and Purification of γ-Polyglutamic Acid from Bacillus licheniformis Fermentation Broth

摘要

Microbially produced gamma-poly glutamic acid (γ-PGA) is a commercially important bio-polymer with many applications in biopharmaceutical, food, cosmetic and waste-water treatment industries. Owing to its increasing demand in various industries, production of γ-PGA is well documented in the literature, however very few methods have been reported for its recovery.In this paper, we report a novel method for the selective recovery and purification of γ-PGA from cell-free fermentation broth of Bacillus licheniformis. The cell-free fermentation broth was treated with divalent copper ions, resulting in the precipitation of γ-PGA, which was collected as a pellet by centrifugation. The pellet was resolubilized and dialyzed against de-ionized water to obtain the purified γ-PGA biopolymer. The efficiency and selectivity of γ-PGA recovery was compared with ethanol precipitation method. We found that 85% of the original γ-PGA content in the broth was recovered by copper sulfate-induced precipitation, compared to 82% recovery by ethanol precipitation method. Since ethanol is a commonly used solvent for protein precipitation, the purity of γ-PGA precipitate was analyzed by measuring proteins that co-precipitated with γ-PGA. Of the total proteins present in the broth, 48% proteins were found to be co-precipitated with γ-PGA by ethanol precipitation, whereas in copper sulfate-induced precipitation, only 3% of proteins were detected in the final purified γ-PGA, suggesting that copper sulfate-induced precipitation offers better selectivity than ethanol precipitation method. Total metal content analysis of the purified γ-PGA revealed the undetectable amount of copper ions, whereas other metal ions detected were in low concentration range. The purified γ-PGA was characterized using infrared spectroscopy.