An efficient multi-stage fermentation strategy for the production of microbial oil rich in arachidonic acid in Mortierella alpina

Abstract

Background

Fungal morphology and aeration play a significant role in the growth process of Mortierella alpina. The production of microbial oil rich in arachidonic acid (ARA) in M. alpina was enhanced by using a multi-stage fermentation strategy which combined fed-batch culture with precise control of aeration and agitation rates at proper times.

Results

The fermentation period was divided into four stages according to the cultivation characteristics of M. alpina. The dissolved oxygen concentration was well suited for ARA biosynthesis. Moreover, the ultimate dry cell weight (DCW), lipid, and ARA yields obtained using this strategy reached 41.4, 22.2, 13.5 g/L, respectively. The respective values represent 14.8, 25.8, and 7.8% improvements over traditional fed-batch fermentation processes.

Conclusions

This strategy provides promising control insights for the mass production of ARA-rich oil on an industrial scale. Pellet-like fungal morphology was transformed into rice-shaped particles which were beneficial for oxygen transfer and thus highly suitable for biomass accumulation.

Keywords

Arachidonic acid / Mortierella alpina / Multi-stage fermentation / Aeration / Agitation / Morphology

References

[1] Gao MJ, Wang C, Zheng ZY, Zhu L, Zhan XB, Lin CC. Improving arachidonic acid fermentation by Mortierella alpina through multistage temperature and aeration rate control in bioreactor. Prep Biochem Biotechnol, 2016, 46: 360-367.

[2] Hamanaka T, Higashiyama K, Fujikawa S, Park EY. Mycelial pellet intrastructure and visualization of mycelia and intracellular lipid in a culture of Mortierella alpina. Appl Microbiol Biotechnol, 2001, 56: 233-238.

[3] Higashiyama K, Murakami K, Tsujimura H, Matsumoto N, Fujikawa S. Effects of dissolved oxygen on the morphology of an arachidonic acid production by Mortierella alpina 1S-4. Biotechnol Bioeng, 1999, 63: 442-448.

[4] Higashiyama K, Fujikawa S, Park EY, Shimizu S. Production of arachidonic acid by Mortierella fungi. Biotechnol Bioprocess Eng, 2002, 7: 252-262.

[5] Hwang BH, Kim JW, Park CY, Park CS, Kim YS, Ryu YW. High-level production of arachidonic acid by fed-batch culture of Mortierella alpina using NH4OH as a nitrogen source and pH control. Biotechnol Lett, 2005, 27: 731-735.

[6] Ji XJ, Ren LJ, Nie ZK, Huang H, Ouyang PK. Fungal arachidonic acid-rich oil: research, development and industrialization. Crit Rev Biotechnol, 2014, 34: 197-214.

[7] Ji XJ, Zhang AH, Nie ZK, Wu WJ, Ren LJ, Huang H. Efficient arachidonic acid-rich oil production by Mortierella alpina through a repeated fed-batch fermentation strategy. Bioresour Technol, 2014, 170: 356-360.

[8] Jin MJ, Huang H, Xiao AH, Zhang K, Liu X, Li S, Peng C. A novel two-step fermentation process for improved arachidonic acid production by Mortierella alpina. Biotechnol Lett, 2008, 30: 1087-1091.

[9] Koike Y, Cai HJ, Higashiyama K, Fujikawa S, Park EY. Effect of consumed carbon to nitrogen ratio of mycelial morphology and arachidonic acid production in cultures of Mortierella alpina. J Biosci Bioeng, 2001, 91: 382-389.

[10] Ling XP, Zeng SY, Chen CX, Liu XT, Lu YH. Enhanced arachidonic acid production using a bioreactor culture of Mortierella alpina with a combined organic nitrogen source. Bioresour Bioprocess, 2016, 3: 43.

[11] Lu JM, Peng C, Ji XJ, You JY, Cong LL, Ouyang PK, Huang H. Fermentation characteristics of Mortierella alpina in response to different nitrogen sources. Appl Biochem Biotechnol, 2011, 164: 979-990.

[12] Nie ZK, Ji XJ, Shang JS, Zhang AH, Ren LJ, Huang H. Arachidonic acid-rich oil production by Mortierella alpina with different gas distributors. Bioprocess Biosyst Eng, 2014, 37: 1127-1132.

[13] Park EY, Koike Y, Higashiyama K, Fujikawa S, Okabe M. Effect of nitrogen source on mycelial morphology and arachidonic acid production in cultures of Mortierella alpina. J Biosci Bioeng, 1999, 88: 61-67.

[14] Park EY, Koike Y, Cai HJ, Higashiyama K, Fujikawa S. Morphological diversity of Mortierella alpina: effect of consumed carbon to nitrogen ratio in flask culture. Bioprocess Biosyst Eng, 2001, 6: 161-166.

[15] Peng C, Huang H, Ji X, Liu X, Ren LJ, Yu W, You JY, Lu JM. Effects of n-hexadecane concentration and a two-stage oxygen supply control strategy on arachidonic acid production by Mortierella alpina ME-1. Chem Eng Technol, 2010, 33: 692-697.

[16] Ren LJ, Huang H, Xiao AH, Lian M, Jin LJ, Ji XJ. Enhanced docosahexaenoic acid production by reinforcing acetyl-CoA and NADPH supply in Schizochytrium sp. HX-308. Bioprocess Biosyst Eng, 2009, 32: 837-843.

[17] Ren LJ, Ji XJ, Huang H, Qu L, Feng Y, Tong QQ, Ouyang PK. Development of a stepwise aeration control strategy for efficient docosahexaenoic acid production by Schizochytrium sp. Appl Microbiol Biotechnol, 2010, 87: 1649-1656.

[18] Ryan AS, Zeller S, Nelson EB, Cohen Z, Ratledge C. Cohen Z, Ratledge C. Safety evaluation of single cell oils and the regulatory requirements for use as food ingredients. Single cell oils: microbial and algal oils, 2010, 2, Urbana: AOCS Press.

[19] Su GM, Jiao KL, Chang JY, Li Z, Guo XY, Sun Y, Zeng XH, Lu YH, Lin L. Enhancing total fatty acids and arachidonic acid production by the red microalgae Porphyridium purpureum. Bioresour Bioprocess, 2016, 3: 33.

[20] Tai C, Li S, Xu Q, Ying H, Huang H, Ouyang PK. Chitosan production from hemicellulose hydrolysate of corn straw: impact of degradation products on Rhizopus oryzae growth and chitosan fermentation. Lett Appl Microbiol, 2010, 51: 278-284.

[21] Totani N, Someya K, Oba K. Kyle DJ, Ratledge C. Industrial production of arachidonic acid by Mortierella. Industrial applications of single cell oils, 1992, Urbana: AOCS Press.

[22] Ward OP, Singh A. Omega-3/6 fatty acids: alternative sources of production. Process Biochem, 2005, 40: 3627-3652.

[23] Wynn JP, Ratledge C. Shahidi F. Oils from microorganisms. Bailey’s industrial oil and fat products, 2005, 6, New York: Wiley, 121-153.

[24] Xu Q, Li S, Fu Y, Tai C, Huang H. Two-stage utilization of corn straw by Rhizopus oryzae for fumaric acid production. Bioresour Technol, 2010, 101: 6262-6264.

[25] Zhang AH, Ji XJ, Wu WJ, Ren LJ, Yu YD, Huang H. Lipid fraction and intracellular metabolite analysis reveal the mechanism of arachidonic acid-rich oil accumulation in the aging process of Mortierella alpina. J Agric Food Chem, 2015, 63: 9812-9819.

[26] Zhu M, Yu LJ, Li W, Zhou PP, Li CY. Optimization of arachidonic acid production by fed-batch culture of Mortierella alpina based on dynamic analysis. Enzyme Microb Technol, 2006, 38: 735-740.

Funding

National Natural Science Foundation of China(21225626)

Natural Science Foundation of Jiangsu Province(BK20131405)