Production of astaxanthin from cellulosic biomass sugars by mutants of the yeast Phaffia rhodozyma

Authors: Montanti, Justin; Nghiem, Nhuan; Johnston, David

Submitted to: Applied Biochemistry and Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/10/2011
Publication Date: 1/28/2011
Citation: Montanti, J.M., Nghiem, N.P., Johnston, D. 2011. Production of astaxanthin from cellulosic biomass sugars by mutants of the yeast Phaffia rhodozyma. Applied Biochemistry and Biotechnology. 164:655-665.

Interpretive Summary: Cellulosic ethanol is of great interest since corn ethanol production is limited. One major problem that keeps cellulosic ethanol technology from becoming economically feasible is the absence of high-value co-products. The cellulosic ethanol technology is much more complex than the technology that has been used for production of corn-based ethanol. Thus, construction of commercial cellulosic ethanol plants requires large capital investments. This requirement combined with the very narrow profit margin of fuel ethanol, which is a commodity chemical, makes commercial production of cellulosic ethanol unrealistic unless high-value co-products also are produced in the same facility using common equipment. In the Sustainable Biofuels and Co-products (SBC) Research Unit we have initiated a program on development of microorganisms and fermentation processes for production of high value-added co-products of cellulosic ethanol. Astaxanthin is one of the potential new co-products of fuel ethanol production. This carotenoid probably is best known for its role in giving the flesh of salmonids, shrimps, lobsters and crayfish the pinkish-red hue. In the marine environment, astaxanthin is acquired through ingestion of microalgae and phytoplankton, which are natural astaxanthin producers. However, since salmonids are unable to synthesize astaxanthin the farm-raised fish need to be fed this carotenoid through their artificial diets. Astaxanthin is a high-value specialty product. The selling price for astaxanthin in 2000 was estimated at ~$2,500/kg and that of 10% astaxanthin formulas in 2007 was listed at $250/kg. The world market for astaxanthin was predicted to reach over $250 million in 2009 and beyond. Recently astaxanthin was discovered to provide many human health benefits. These discoveries could lead to development of nutraceutical applications and significant expansion of the market for astaxanthin. In our laboratory we have developed several non-recombinant strains of the red yeast Phaffia rhodozyma capable of metabolizing the three major fermentable sugars that can be obtained from cellulosic feedstocks, which include glucose, xylose, and arabinose, for astaxanthin production. The results obtained in our experiments demonstrated that these strains could use clean sugar solutions prepared with purified laboratory-grade sugars as well as the actual sugar solutions made from two biomass feedstocks, namely barley straw and sugarcane bagasse. The result of this research effort is a fermentation process for production of a high value-added co-product of cellulosic ethanol. Improvement of the strains and the process is in progress together with the effort on development of microorganisms and processes for production of other potential co-products of cellulosic ethanol.

Technical Abstract: Astaxanthin is a carotenoid of high value to the aquaculture, nutraceutical, and pharmaceutical industries. Three mutant strains of the astaxanthin-producing yeast Phaffia rhodozyma, which were derived from the parent strain ATCC 24202 (UCD 67-210) and designated JTM166, JTM185, and SSM19, were tested for their capability of utilizing the major sugars found in cellulosic biomass, including glucose, xylose, and arabinose, for astaxanthin production. While all three strains were capable of metabolizing these sugars, both individually and in mixtures, JTM 185 demonstrated the greatest sugar utilization and astaxanthin production. The kinetics of sugar utilization was studied in fermenters using mixtures of glucose, xylose, and arabinose at varied concentrations. It was found that glucose was utilized preferentially, followed by xylose, and lastly, arabinose. The greatest astaxanthin production per total sugar consumption (0.21 mg/g) was observed when glucose was supplied in very low levels relative to xylose and arabinose; although the final astaxanthin concentration (8.3 mg/L) was lower than when glucose was supplied at higher concentrations. Hydrolysates produced from sugarcane bagasse and barley straw pretreated by the soaking in aqueous ammonia (SAA) method and hydrolyzed with the commercial cellulase enzyme product, Accellerase 1000, were used for astaxanthin production by the mutant strain JTM185. The organism was capable of metabolizing all of the sugars present in the hydrolysates from both biomass sources, and produced a similar amount of astaxanthin from both hydrolysates, although this amount was less than was produced from synthetic sugars.