Optimization of submerged culture conditions involving a developed fine powder solid seed for exopolysaccharide production by the medicinal mushroom Ganoderma lucidum

  • Sheng-Rong LiuEmail author
  • Wei-Rui Zhang


To facilitate Ganoderma lucidum submerged cultivation and achieve high productivity, four fine powder solid substrates incorporated with different nitrogen-rich supplements were utilized to grow the fungus and as solid seed for its submerged culture. Of the four solid seeds, the soybean meal solid seed gave the highest biomass (10.73 g/L) and exopolysaccharide (EPS) (1.22 g/L), higher than those (8.36 g/L biomass and 0.44 g/L EPS) obtained with mycelial liquid seed. The optimal level of soybean meal supplementation was 20% (w/w) for production of the solid seed. Following single factor experiments, levels of three selected process variables were optimized as: the moisture content of solid seed, 70%; inoculum size, 0.8 g/flask; and rotary speed, 160 rpm. These conditions were validated experimentally with improved EPS yield of 1.33 g/L. The developed solid seed can be conveniently used for G. lucidum submerged culture with improved EPS productivity.


Ganoderma lucidum Submerged culture Solid seed Exopolysaccharide Response surface methodology 



The authors would like to thank the financial support by the Program for Young and Middle School Teachers of Education Department of Fujian Province (Grant No. JA15558), Ningde Normal University (Grant No. 2013F22), and the Industry Guidance Program of the Department of Science and Technology of Fujian Province (Grant No. 2015N0032).

Compliance with ethical standards

Conflict of interest

The authors declare that there is no conflict of interest.


  1. Bae JT, Sinha J, Park JP, Song CH, Yun JW. Optimization of submerged culture conditions for exo-biopolymer production by Paecilomyces japonica. J. Microbiol. Biotechnol. 10: 482–487 (2000)Google Scholar
  2. Box GP, Behnken DW. Some new three level design for the study of quantitative variables. Technometrics 2: 456–475 (1960)CrossRefGoogle Scholar
  3. Chang MY, Tsai GJ, Houng JY. Optimization of medium composition for the submerged culture of Ganoderma lucidum by Taguchi array design and steepest ascent method. Enzyme Microb. Technol. 38: 407–414 (2006)CrossRefGoogle Scholar
  4. Dubois M, Gilles KA, Hamilton JK, Rebers PA, Smith F. Colorimetric method for determination of sugars and related substances. Anal. Chem. 28: 350–356 (1956)CrossRefGoogle Scholar
  5. Fang QH, Tang YJ, Zhong JJ. Significance of inoculation density control in production of polysaccharide and ganoderic acid by submerged culture of Ganoderma lucidum. Process Biochem. 37: 1375–1379 (2002)CrossRefGoogle Scholar
  6. Fang QH, Zhong JJ. Effect of initial pH on production of ganoderic acid and polysacchrides by submerged fermentation of Ganoderma lucidum. Process Biochem. 37: 769–774 (2002)CrossRefGoogle Scholar
  7. Feng J, Feng N, Zhang JS, Yang Y, Jia W, Lin CC. A new temperature control shifting strategy for enhanced triperpene production by Ganoderma lucidum G0119 based on submerged liquid fermentation. Appl. Biochem. Biotechnol. 180: 740–752 (2016)CrossRefGoogle Scholar
  8. Feng J, Zhang JS, Feng N, Yan MQ, Yang Y, Jia W, Lin CC. A novel Ganoderma lucidum G0119 fermentation strategy for enhanced triterpenes production by statistical process optimization and addition of oleic acid. Eng. Life. Sci. 17: 430–439 (2017)CrossRefGoogle Scholar
  9. Gao CJ, Wang ZY, Su TT, Zhang J, Yang XH. Optimisation of exopolysaccharide production by Gomphidius rutilus and its antioxidant activities in vitro. Carbohydr. Polym. 87: 2299–2305 (2012)CrossRefGoogle Scholar
  10. Jia J, Yang XF, Wu ZL, Zhang Q, Lin Z, Guo HT, Lin CSK, Wang JY, Wang YS. Optimization of fermentation medium for extracellular lipase production from Aspergillus niger using response surface methodology. Biomed Res. Int. 2015: 497462 (2015)Google Scholar
  11. Jong SC, Birmingham JM. Medicinal benefits of the mushroom Ganoderma. Adv. Appl. Microbiol. 37: 101–134 (1992)CrossRefGoogle Scholar
  12. Kim HM, Kim SW, Hwang HJ, Park MK, Mahmoud YAG, Choi JW, Yun JW. Influence of agitation intensity and aeration rate on production of antioxidative exopolysaccharides from submerged mycelial culture of Ganoderma resinaceum. J. Microbiol. Biotechnol. 16: 1240–1247 (2006a)Google Scholar
  13. Kim HM, Park MK, Yun JW. Culture pH affects exopolysaccharide production in submerged mycelial culture of Ganoderma lucidum. Appl. Biochem. Biotechnol. 134: 249–262 (2006b)CrossRefGoogle Scholar
  14. Le J, Hu SZ, Xu M. Optimization of submerged culture conditions for the production of mycelial biomass and exopolysaccharide by Pleurotus nebrodensis. Ann. Microbiol. 57: 389–393 (2007)CrossRefGoogle Scholar
  15. Lee KM, Lee SY, Lee HY. Biostage control of pH for improving exopolysaccharide production from mycelia of Ganoderma lucidum in an air-lift fermentor. J. Biosci. Bioeng. 88: 646–650 (1999)CrossRefGoogle Scholar
  16. Li HX, Lu ZM, Geng Y, Gong JS, Zhang XJ, Shi JS, Xu ZH, Ma YH. Efficient production of bioactive metabolites from Antrodia camphorata ATCC 200183 by asexual reproduction-based repeated batch fermentation. Bioresour. Technol. 194: 334–343 (2015)CrossRefGoogle Scholar
  17. Liu SR, Zhang WR. Hyperproduction of exopolysaccharides by submerged mycelial culture of Ganoderma lucidum using a solid seed grown in fine-powder of wheat bran and in vitro evaluation of the antioxidant activity of the exopolysaccharides produced. Food Sci. Biotechnol. 27: 1129–1136 (2018)CrossRefGoogle Scholar
  18. Lonsane BK, Ghildyal NP, Budiatman S, Ramakrishna SV. Engineering aspects of solid state fermentation. Enzyme Microb. Technol. 7: 258–265 (1985)CrossRefGoogle Scholar
  19. Mizuno T, Wang GY, Zhang J, Kawagishi H, Nishitoba T, Ti JX. Reishi, Ganoderma lucidum and Ganoderma tsugae: bioactive substances and medicinal effects. Food Rev. Int. 11: 151–166 (1995)CrossRefGoogle Scholar
  20. Montgomery DC. Design and analysis of experiments (2nd ED). J. Am. Stat. Assoc. 16: 241–242 (2000)Google Scholar
  21. Moonmoon M, Shelly NJ, Khan MA, Uddin MN, Hossain K, Tania M, Ahmed S. Effects of different levels of wheat bran, rice bran and maize powder supplementation with saw dust on the production of shiitake mushroom (Lentinus edodes (Berk.) Singer). Saudi J. Biol. Sci. 18: 323–328 (2011)CrossRefGoogle Scholar
  22. Naraian R, Sahu RK, Kumar S, Garg SK, Singh CS, Kanaujia RS. Influence of different nitrogen rich supplements during cultivation of Pleurotus florida on corn cob substrate. Environmentalist 29: 1–7 (2009)CrossRefGoogle Scholar
  23. Pandey A. Production of starch saccharifying enzyme (glucoamylase) in solid cultures. Starch 44: 75–77 (1992)CrossRefGoogle Scholar
  24. Postemsky PD, Delmastro SE, Curvetto NR. Effect of edible oils and Cu (II) on the biodegradation of rice by-products by Ganoderma lucidum mushroom. Int. Biodeter. Biodegr. 93: 25–32 (2014)CrossRefGoogle Scholar
  25. Reddy LVA, Wee YJ, Yun JS, Ryu HW. Optimization of alkaline protease production by batch culture of Bacillus sp. RKY3 through Plackett–Burman and response surface methodological approaches. Bioresour. Technol. 99: 2242–2249 (2008)CrossRefGoogle Scholar
  26. Rocky-Salimi K, Hamidi-Esfahani Z. Evaluation of the effect of particle size, aeration rate and harvest time on the production of cellulase by Trichoderma reesei QM9414 using response surface methodology. Food Bioprod. Process. 88: 61–66 (2010)CrossRefGoogle Scholar
  27. Shiao MS. Natural products of the medicinal fungus Ganoderma lucidum: occurrence, biological activities, and pharmacological functions. Chem. Res. 3: 172–180 (2003)Google Scholar
  28. Shu CH, Lung MY. Effect of pH on the production and molecular weight distribution of exopolysaccharide by Antrodia camphorata in batch cultures. Process Biochem. 39: 931–937 (2004)CrossRefGoogle Scholar
  29. Song CH, Yang BK, Ra KS, Shon DH, Park EJ, Go GI, Kim YH. Hepatoprotective effect of extracellular polymer produced by submerged culture of Ganoderma lucidum WK-003. J. Microbiol. Biotechnol. 8: 277–279 (1998)Google Scholar
  30. Tepwong P, Giri A, Ohshima T. Effect of mycelial morphology on ergothioneine production during liquid fermentation of Lentinula edodes. Mycoscience 53: 102–112 (2012)CrossRefGoogle Scholar
  31. Upadhyay M, Shrivastava B, Jain A, Kidwai M, Kumar S, Gomes J, Goswami DG, Panda AK, Kuhad RC. Production of ganoderic acid by Ganoderma lucidum RCKB-2010 and its therapeutic potential. Ann. Microbiol. 64: 839–846 (2014).CrossRefGoogle Scholar
  32. Vu VH, Pham TA, Kim K. Improvement of a fungal strain by repeated and sequential mutagenesis and optimization of solid-state fermentation for the hyper-production of raw-starch-digesting enzyme. J. Microbiol. Biotechnol. 20: 718–726 (2010)CrossRefGoogle Scholar
  33. Wang Z, Quan Y, Zhou F. Optimization of medium composition for exopolysacchride production by Phellinus nigricans. Carbohydr. Polym. 105: 200–206 (2014)CrossRefGoogle Scholar
  34. Wei ZH, Duan YY, Qian YQ, Guo XF, Li YJ, Jin SH, Zhou ZX, Shan SY, Wang CR, Chen XJ, Zheng YG, Zhong JJ. Screening of Ganoderma strains with high polysaccharides and ganoderic acid contents and optimization of the fermentation medium by statistical methods. Bioprocess Biosyst. Eng. 37: 1789–1797 (2014)CrossRefGoogle Scholar
  35. Yang FC, Ke YF, Kuo SS. Effect of fatty acids on the mycelial growth and polysaccharide formation by Ganoderma lucidum in shake flask cultures. Enzyme Microb. Technol. 27: 295–301 (2000)CrossRefGoogle Scholar
  36. Yang FC, Liau CB. The influence of environmental conditions on polysaccharide formation by Ganoderma lucidum in submerged cultures. Process Biochem. 33: 547–553 (1998).CrossRefGoogle Scholar
  37. Yang FC, Yang MJ, Cheng SH. A novel method to enhance the mycelia production of Ganoderma lucidum in submerged cultures by polymer additives and agitation strategies. J. Taiwan Inst. Chem. Eng. 40: 148–154 (2009)CrossRefGoogle Scholar
  38. Yang SQ, Xiong H, Yang HY, Yan QJ, Jiang ZQ. High-level production of ß-1,3-1,4-glucanase by Rhizomucor miehei under solid-state fermentation and its potential application in the brewing industry. J. Appl. Microbiol. 118: 84–91 (2014)CrossRefGoogle Scholar
  39. Yen HW, Chiang MH. Using the mycelium-covered cereals as an efficient inoculation method for rapamycin fermentation in a 15-L fermenter using Streptomyces hygroscopicus. Bioresour. Bioprocess 2: 43 (2015)CrossRefGoogle Scholar
  40. Zhao HZ, Wang J, Lu ZX. Optimization of process parameters of the Pholiota squarrosa extracellular polysaccharide by Box–Behnken statistical design. Carbohydr. Polym. 77: 677–680 (2009)CrossRefGoogle Scholar
  41. Zhu LW, Zhong JJ, Tang YJ. Significance of fungal elicitors on the production of ganoderic acid and Ganoderma polysaccharides by the submerged culture of medicinal mushroom Ganoderma lucidum. Process Biochem. 43: 1359–1370 (2008)CrossRefGoogle Scholar

Copyright information

© The Korean Society of Food Science and Technology 2018

Authors and Affiliations

  1. 1.College of Life ScienceNingde Normal UniversityNingdeChina
  2. 2.Fujian Higher Education Research Center for Local Biological Resources in Ningde CityNingdeChina

Personalised recommendations