Enhancement of co-production of lutein and protein in Chlorella sorokiniana FZU60 using different bioprocess operation strategies

Abstract

Co-production of multiple compounds is an efficient approach to enhance the economic feasibility of microalgae-based metabolites production. In this study, Chlorella sorokiniana FZU60 was cultivated under different bioprocess strategies to enhance the co-production of lutein and protein. Results showed that both lutein and protein content (7.72 and 538.06 mg/g, respectively) were highest at the onset of nitrogen deficiency under batch cultivation. Semi-batch III strategy, with 75% microalgal culture replacement by fresh medium, obtained similar content, productivity, and yield of lutein and protein as batch cultivation, demonstrating that it can be used for stable and continuous production. Fed-batch II strategy, feeding with 1/3 modified BG11 medium, achieved super-high lutein and protein yield (28.81 and 1592.77 mg/L, respectively), thus can be used for high-output production. Besides, two-stage strategy, combining light intensity shift and semi-batch cultivation, gained extremely high lutein and protein productivity (15.31 and 1080.41 mg/L/day, respectively), thereby is a good option for high-efficiency production. Moreover, the fed-batch II and two-stage strategy achieved high-quality lutein and protein, thus are promising for the co-production of lutein and protein in C. sorokiniana FZU60 for commercial application.

Keywords

Chlorella sorokiniana / Co-production / Lutein / Protein / Strategy

References

[1] Becker EW. Micro-algae as a source of protein. Biotechnol Adv, 2007, 25: 207-210.

[2] Bin Azmi AA, Chew KW, Chia WY, Mubashir M, Sankaran R, Lam MK, Lim JW, Ho YC, Show PL. Green bioprocessing of protein from Chlorella vulgaris microalgae towards circular bioeconomy. Bioresour Technol, 2021, 333: 12519.

[3] Chauton MS, Olsen Y, Vadstein O. Biomass production from the microalga Phaeodactylum tricornutum: Nutrient stress and chemical composition in exponential fed-batch cultures. Biomass Bioenergy, 2013, 58: 87-94.

[4] Chen C-Y, Lee P-J, Tan CH, Lo Y-C, Huang C-C, Show PL, Lin C-H, Chang J-S. Improving protein production of indigenous microalga Chlorella vulgaris FSP-E by photobioreactor design and cultivation strategies. Bioresour Technol, 2015, 10: 905-914.

[5] Chen JH, Chen CY, Hasunuma T, Kondo A, Chang CH, Ng IS, Chang JS. Enhancing lutein production with mixotrophic cultivation of Chlorella sorokiniana MB-1-M12 using different bioprocess operation strategies. Bioresour Technol, 2019, 278: 17-25.

[6] Crawford DW, Lipsen MS, Purdie DA, Lohan MC, Statham PJ, Whitney FA, Putland JN, Johnson WK, Sutherland N, Peterson TD. Influence of zinc and iron enrichments on phytoplankton growth in the northeastern subarctic Pacific. Limnol Oceanogr, 2003, 48: 1583-1600.

[7] Dineshkumar R, Subramanian G, Dash SK, Sen R. Development of an optimal light-feeding strategy coupled with semi-continuous reactor operation for simultaneous improvement of microalgal photosynthetic efficiency, lutein production and CO₂ sequestration. Biochem Eng J, 2016, 113: 47-56.

[8] Dixon C, Wilken LR. Green microalgae biomolecule separations and recovery. Bioresour Bioprocess, 2018, 5: 14.

[9] Ferreira M, Seixas P, Coutinho P, Fábregas J, Otero A. Effect of the nutritional status of semi-continuous microalgal cultures on the productivity and biochemical composition of Brachionus plicatilis. Mar Biotechnol, 2011, 13: 1074-1085.

[10] Fields FJ, Ostrand JT, Mayfield SP. Fed-batch mixotrophic cultivation of Chlamydomonas reinhardtii for high-density cultures. Algal Res, 2018, 33: 109-117.

[11] Fuentes-Grünewald C, Bayliss C, Zanain M, Pooley C, Scolamacchia M, Silkina A. Evaluation of batch and semi-continuous culture of Porphyridium purpureum in a photobioreactor in high latitudes using Fourier Transform Infrared spectroscopy for monitoring biomass composition and metabolites production. Bioresour Technol, 2015, 189: 357-363.

[12] Garcia-Canedo JC, Cristiani-Urbina E, Flores-Ortiz CM, Ponce-Noyola T, Esparza-Garcia F, Canizares-Villanueva RO. Batch and fed-batch culture of Scenedesmus incrassatulus: effect over biomass, carotenoid profile and concentration, photosynthetic efficiency and non-photochemical quenching. Algal Res, 2016, 13: 41-52.

[13] Geada P, Moreira C, Silva M, Nunes R, Madureira L, Rocha CMR, Pereira RN, Vicente AA, Teixeira JA. Algal proteins: production strategies and nutritional and functional properties. Bioresour Technol, 2021, 332: 125125.

[14] Ho SH, Chen CNN, Lai YY, Lu WB, Chang JS. Exploring the high lipid production potential of a thermotolerant microalga using statistical optimization and semi-continuous cultivation. Bioresour Technol, 2014, 163: 128-135.

[15] Ho SH, Xie YP, Chan MC, Liu CC, Chen CY, Lee DJ, Huang CC, Chang JS. Effects of nitrogen source availability and bioreactor operating strategies on lutein production with Scenedesmus obliquus FSP-3. Bioresour Technol, 2015, 184: 131-138.

[16] Joint WHO/FAO/UNU Expert Consultation. Haden A. Protein and amino acid requirements in human nutrition introduction. World Health Organization technical report, 2007, Geneva: WHO Press, 150.

[17] Kent M, Welladsen HM, Mangott A, Li Y, Kumar S. Nutritional evaluation of australian microalgae as potential human health supplements. PLoS ONE, 2015, 10: e0118985.

[18] Khoo KS, Chong YM, Chang WS, Yap JM, Foo SC, Khoiroh I, Lau PL, Chew KW, Ooi CW, Show PL. Permeabilization of Chlorella sorokiniana and extraction of lutein by distillable CO₂-based alkyl carbamate ionic liquids. Sep Purif Technol, 2021, 256: 117471.

[19] Koyande AK, Tanzil V, Dharan HM, Subramaniam M, Robert RN, Lau PL, Khoiroh I, Show PL. Integration of osmotic shock assisted liquid biphasic system for protein extraction from microalgae Chlorella vulgaris. Biochem Eng J, 2020, 157: 107532.

[20] Lee E, Jalalizadeh M, Zhang Q. Growth kinetic models for microalgae cultivation: A review. Algal Res, 2015, 12: 497-512.

[21] Levasseur W, Perre P, Pozzobon V. A review of high value-added molecules production by microalgae in light of the classification. Biotechnol Adv, 2020, 41: 107545.

[22] Li YL, Sun H, Wu T, Fu YL, He YJ, Mao XM, Chen F. Storage carbon metabolism of Isochrysis zhangjiangensis under different light intensities and its application for co-production of fucoxanthin and stearidonic acid. Bioresour Technol, 2019, 282: 94-102.

[23] Lin JH, Lee DJ, Chang JS. Lutein production from biomass: Marigold flowers versus microalgae. Bioresour Technol, 2015, 184: 421-428.

[24] Lu SH, Wang JX, Niu YH, Yang J, Zhou J, Yuan YJ. Metabolic profiling reveals growth related FAME productivity and quality of Chlorella sorokiniana with different inoculum sizes. Biotechnol Bioeng, 2012, 109: 1651-1662.

[25] Lu X, Sun H, Zhao WY, Cheng KW, Chen F, Liu B. A hetero-photoautotrophic two-stage cultivation process for production of fucoxanthin by the marine diatom Nitzschia laevis. Mar Drugs, 2018, 16: 219.

[26] Lu K, Zhao X, Ho SH, Ma R, Xie Y, Chen J. Biorefining and the functional properties of proteins from lipid and pigment extract residue of Chlorella pyrenoidosa. Mar Drugs, 2019, 17: 454.

[27] Ma R, Zhao X, Xie Y, Ho SH, Chen J. Enhancing lutein productivity of Chlamydomonas sp. via high-intensity light exposure with corresponding carotenogenic genes expression profiles. Bioresour Technol, 2018, 275: 416-420.

[28] Ma R, Wang B, Chua ET, Zhao X, Lu K, Ho SH, Shi X, Liu L, Xie Y, Lu Y, Chen J. Comprehensive utilization of marine microalgae for enhanced co-production of multiple compounds. Mar Drugs, 2020, 18: 467.

[29] Ma R, Zhao X, Ho SH, Shi X, Liu L, Xie Y, Chen J, Lu Y. Co-production of lutein and fatty acid in microalga Chlamydomonas sp. JSC4 in response to different temperatures with gene expression profiles. Algal Res, 2020, 47: 101821.

[30] Maximize Market Research PVT. LTD (2020) Global lutein market industry analysis and forecast (2020–2027)—by form, by source, by production process, by application and by geography. https://www.maximizemarketresearch.com/market-report/lutein-market/661/. Accessed Jan 2020

[31] Muys M, Sui YX, Schwaiger B, Lesueur C, Vandenheuvel D, Vermeir P, Vlaeminck SE. High variability in nutritional value and safety of commercially available Chlorella and Spirulina biomass indicates the need for smart production strategies. Bioresour Technol, 2019, 275: 247-257.

[32] Osorio JHM, Pinto G, Pollio A, Frunzo L, Lens PNL, Esposito G. Start-up of a nutrient removal system using Scenedesmus vacuolatus and Chlorella vulgaris biofilms. Bioresour Bioprocess, 2019, 6: 27.

[33] Plyusnina TY, Khruschev SS, Degtereva NS, Konyukhov IV, Solovchenko AE, Kouzmanova M, Goltsev VN, Riznichenko GY, Rubin AB. Special issue in honour of Prof. Reto J. Strasser—gradual changes in the photosynthetic apparatus triggered by nitrogen depletion during microalgae cultivation in photobioreactor. Photosynthetica, 2020, 58: 443-451.

[34] Prates DD, Duarte JH, Vendruscolo RG, Wagner R, Ballus CA, Oliveira WD, Godoy HT, Barcia MT, de Morais MG, Radmann EM, Costa JAV. Role of light emitting diode (LED) wavelengths on increase of protein productivity and free amino acid profile of Spirulina sp. cultures. Bioresour Technol, 2020, 306: 123184.

[35] Siaut M. Oil accumulation in the model green alga Chlamydomonas reinhardtii: characterization, variability between common laboratory strains and relationship with starch reserves. BMC Biotechnol, 2011, 11: 7.

[36] Soto-Sierra L, Wilken LR, Dixon CK. Aqueous enzymatic protein and lipid release from the microalgae Chlamydomonas reinhardtii. Bioresour Bioprocess, 2020, 7: 46.

[37] Sui Y, Muys M, Van de Waal DB, D'Adamo S, Vermeir P, Fernandes TV, Vlaeminck SE. Enhancement of co-production of nutritional protein and carotenoids in Dunaliella salina using a two-phase cultivation assisted by nitrogen level and light intensity. Bioresour Technol, 2019, 287: 121398.

[38] Sun Z, Li T, Zhou ZG, Jiang Y. Microalgae as a source of lutein: chemistry, biosynthesis, and carotenogenesis. Adv Biochem Eng Biotechnol, 2016, 153: 37-58.

[39] Sun Z, Wang X, Liu J. Screening of Isochrysis strains for simultaneous production of docosahexaenoic acid and fucoxanthin. Algal Res, 2019, 41: 101545.

[40] Tachihana S, Nagao N, Katayama T, Hirahara M, Yusoff FM, Banerjee S, Shariff M, Kurosawa N, Toda T, Furuya K. High productivity of eicosapentaenoic acid and fucoxanthin by a marine diatom Chaetoceros gracilis in a semi-continuous culture. Front Bioeng Biotechnol, 2020, 8: 602721.

[41] Tibbetts SM, Bjornsson WJ, McGinn PJ. Biochemical composition and amino acid profiles of Nannochloropsis granulata algal biomass before and after supercritical fluid CO₂ extraction at two processing temperatures. Anim Feed Sci Technol, 2015, 204: 62-71.

[42] Tong Z, Chi Z, Sheng J. A highly thermosensitive and permeable mutant of the marine yeast Cryptococcus aureus G7a potentially useful for single-cell protein production and its nutritive components. Mar Biotechnol, 2009, 11: 280-286.

[43] Vaquero I, Mogedas B, Ruiz-Dominguez MC, Vega JM, Vilchez C. Light-mediated lutein enrichment of an acid environment microalga. Algal Res, 2014, 6: 70-77.

[44] Vello V, Chu W-L, Lim P-E, Majid NA, Phang S-M. Metabolomic profiles of tropical Chlorella species in response to physiological changes during nitrogen deprivation. J Appl Phycol, 2018, 30: 3131-3151.

[45] Xie Y, Ho SH, Chen CN, Chen CY, Ng IS, Jing KJ, Chang JS, Lu Y. Phototrophic cultivation of a thermo-tolerant Desmodesmus sp. for lutein production: Effects of nitrate concentration, light intensity and fed-batch operation. Bioresour Technol, 2013, 144: 435-444.

[46] Xie Y, Li J, Ma R, Ho SH, Shi X, Liu L, Chen J. Bioprocess operation strategies with mixotrophy/photoinduction to enhance lutein production of microalga Chlorella sorokiniana FZU60. Bioresour Technol, 2019, 290: 121798.

[47] Xie Y, Lu K, Zhao X, Ma R, Chen J, Ho SH. Manipulating nutritional conditions and salinity-gradient stress for enhanced lutein production in marine microalga Chlamydomonas sp. Biotechnol J, 2019, 14: 1800380.

[48] Xie Y, Xiong X, Chen S. Challenges and potential in increasing lutein content in microalgae. Microorganisms, 2021, 9: 1068.

[49] Zhao YT, Yue CC, Geng SX, Ning DL, Ma T, Yu XY. Role of media composition in biomass and astaxanthin production of Haematococcus pluvialis under two-stage cultivation. Bioprocess Biosyst Eng, 2019, 42: 593-602.

[50] Zhao TT, Liu MX, Zhao TT, Chen AL, Zhang L, Liu H, Ding K, Xiao PY. Enhancement of lipid productivity in Chlorella pyrenoidosa by collecting cells at the maximum cell number in a two-stage culture strategy. Algal Res, 2021, 55: 102278.

Funding

demonstration project for innovative development of fuzhou's marine economy during the 13th five-year plan, china(FZHJ15)

natural science foundation of fujian province(No. 2020J01484)

education and research project for middle-aged and young teachers of the education department of fujian province(JAT200036)