Extraction and Purification of Highly Active Astaxanthin from Corynebacterium glutamicum Fermentation Broth

The marine carotenoid astaxanthin is one of the strongest natural antioxidants and therefore is used in a broad range of applications such as cosmetics or nutraceuticals. To meet the growing market demand, the natural carotenoid producer Corynebacterium glutamicum has been engineered to produce astaxanthin by heterologous expression of genes from the marine bacterium Fulvimarina pelagi. To exploit this promising source of fermentative and natural astaxanthin, an efficient extraction process using ethanol was established in this study. Appropriate parameters for ethanol extraction were identified by screening ethanol concentration (62.5-97.5% v/v), temperature (30-70 °C) and biomass-to-solvent ratio (3.8-19.0 mg_CDW/mL_solvent). The results demonstrated that the optimal extraction conditions were: 90% ethanol, 60 °C, and a biomass-to-solvent ratio of 5.6 mg_CDW/mL_solvent. In total, 94% of the cellular astaxanthin was recovered and the oleoresin obtained contained 9.4 mg/g astaxanthin. With respect to other carotenoids, further purification of the oleoresin by column chromatography resulted in pure astaxanthin (100%, HPLC). In addition, a 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay showed similar activities compared to esterified astaxanthin from microalgae and a nine-fold higher antioxidative activity than synthetic astaxanthin.