title: A novel Method for the selective recovery and purification of gamma-polyglutamic acid from Bacillus licheniformis fermentation broth.
abstract: Microbially produced gamma-polyglutamic acid (gamma-PGA) is a commercially important biopolymer with many applications in biopharmaceutical, food, cosmetic and waste-water treatment industries. Owing to its increasing demand in various industries, production of gamma-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 gamma-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 gamma-PGA, which was collected as a pellet by centrifugation. The pellet was resolubilized and dialyzed against de-ionized water to obtain the purified gamma-PGA biopolymer. The efficiency and selectivity of gamma-PGA recovery was compared with ethanol precipitation method. We found that 85% of the original gamma-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 gamma-PGA precipitate was analyzed by measuring proteins that co-precipitated with gamma-PGA. Of the total proteins present in the broth, 48% proteins were found to be co-precipitated with gamma-PGA by ethanol precipitation, whereas in copper sulfate-induced precipitation, only 3% of proteins were detected in the final purified gamma-PGA, suggesting that copper sulfate-induced precipitation offers better selectivity than ethanol precipitation method. Total metal content analysis of the purified gamma-PGA revealed the undetectable amount of copper ions, whereas other metal ions detected were in low concentration range. The purified gamma-PGA was characterized using infrared spectroscopy.
authors.affiliation: Department of Chemical and Biochemical Engineering, Faculty of Engineering, The University of Western Ontario, London Ontario, Canada N6A 5B9.
authors.forename: Bhavik
authors.initials: B
authors.lastname: Manocha
authors.forename: Argyrios
authors.initials: A
authors.lastname: Margaritis
mesh_terms: D001407:Bacillus / Q000737:chemistry* / Q000378:metabolism; D011232:Chemical Precipitation; D002850:Chromatography, Gel; D019327:Copper Sulfate / Q000737:chemistry; D017077:Culture Media, Conditioned; D000431:Ethanol / Q000737:chemistry; D005285:Fermentation; D005612:Freeze Drying; D008970:Molecular Weight; D011099:Polyglutamic Acid / Q000737:chemistry / Q000302:isolation & purification*; D017550:Spectroscopy, Fourier Transform Infrared
chemical_list: D017077:Culture Media, Conditioned; D011099:Polyglutamic Acid; D000431:Ethanol; D019327:Copper Sulfate