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Abstract: Dietary consumption of eicosapentaenoic acid (EPA) offers diverse health benefits, such as the regulation of blood triglycerides and the prevention of cardiovascular diseases. EPA is naturally synthesized by Schizochytrium sp.; however, its low production level limits its potential for industrial application. The goal of this study was to increase EPA productivity in Schizochytrium sp. by gas‒liquid-phase plasma (GLPP) mutagenesis combined with a high-throughput screening method. First, a diverse array of mutants was generated through GLPP mutagenesis. Next, the mutants with elevated EPA productivity were identified through near-infrared spectroscopy (NIRS). Notably, the M7-25 mutant demonstrated the highest and most consistent EPA production. After the culture medium was optimized, the EPA titer increased from 0.45 to 1.70 g/L. Finally, a cofermentation strategy using ammonia and glucose feeding was employed, and the EPA titer reached 2.08 g/L in a 7-L fermenter. This study reports the highest EPA titer achieved in Schizochytrium sp. via mutagenesis to date, highlighting its great market potential for industrial production.

气液相等离子体诱变技术结合高通量筛选提升裂殖壶菌中二十碳五烯酸产量的应用

[1]AuchterlonieNA, BescobyGH, 2021. Global market for the long-chain omega-3 fatty acids EPA and DHA and their regulation. In: García-Moreno PJ, Jacobsen C, Moltke Sørensen AD, et al. (Eds.), Omega-3 Delivery Systems. Academic Press, London, p.79-106.

[2]BazinetRP, MetherelAH, ChenCT, et al., 2020. Brain eicosapentaenoic acid metabolism as a lead for novel therapeutics in major depression. Brain Behav Immun, 85:21-28.

[3]BhattDL, StegPG, MillerM, et al., 2019. Effects of icosapent ethyl on total ischemic events: from REDUCE-IT. J Am Coll Cardiol, 73(22):2791-2802.

[4]CalderPC, 2013. n-3 Fatty acids, inflammation and immunity: new mechanisms to explain old actions. Proc Nutr Soc, 72(3):326-336.

[5]ChatterjeeN, WalkerGC, 2017. Mechanisms of DNA damage, repair, and mutagenesis. Environ Mol Mutagen, 58(5):235-263.

[6]ChenZL, YangLH, HeSJ, et al., 2023. Development of a green fermentation strategy with resource cycle for the docosahexaenoic acid production by Schizochytrium sp. Bioresour Technol, 385:129434.

[7]DerwenskusF, WeickertS, LewandowskiI, et al., 2020. Economic evaluation of up- and downstream scenarios for the co-production of fucoxanthin and eicosapentaenoic acid with P. tricornutum using flat-panel airlift photobioreactors with artificial light. Algal Res, 51:102078.

[8]DoaeiS, GholamiS, RastgooS, et al., 2021. The effect of omega-3 fatty acid supplementation on clinical and biochemical parameters of critically ill patients with COVID-19: a randomized clinical trial. J Transl Med, 19:128.

[9]DongW, ZhangYQ, ZhangB, et al., 2013. Rapid prediction of fatty acid composition of vegetable oil by Raman spectroscopy coupled with least squares support vector machines. J Raman Spectrosc, 44(12):1739-1745.

[10]DuF, WangYZ, XuYS, et al., 2021. Biotechnological production of lipid and terpenoid from Thraustochytrids. Biotechnol Adv, 48:107725.

[11]GolayPA, MoulinJ, 2016. Determination of labeled fatty acids content in milk products, infant formula, and adult/pediatric nutritional formula by capillary gas chromatography: collaborative study, final action 2012.13. J AOAC Int, 99(1):210-222.

[12]González-MartínMI, EscuredoO, Hernández-JiménezM, et al., 2021. Prediction of stable isotopes and fatty acids in subcutaneous fat of Iberian pigs by means of NIR: a comparison between benchtop and portable systems. Talanta, 224:121817.

[13]GuptaA, WilkensS, AdcockJL, et al., 2013. Pollen baiting facilitates the isolation of marine Thraustochytrids with potential in omega-3 and biodiesel production. J Ind Microbiol Biotechnol, 40(11):1231-1240.

[14]HamiltonHA, NewtonR, AuchterlonieNA, et al., 2020. Systems approach to quantify the global omega-3 fatty acid cycle. Nat Food, 1:59-62.

[15]JiaYL, GengSS, DuF, et al., 2022. Progress of metabolic engineering for the production of eicosapentaenoic acid. Crit Rev Biotechnol, 42(6):838-855.

[16]JiangY, FanKW, WongRTY, et al., 2004. Fatty acid composition and squalene content of the marine microalga Schizochytrium mangrovei. J Agric Food Chem, 52(5):1196-1200.

[17]KarunathilakaSR, ChoiSH, MossobaMM, et al., 2019. Rapid classification and quantification of marine oil omega-3 supplements using ATR-FTIR, FT-NIR and chemometrics. J Food Compos Anal, 77:9-19.

[18]KhanSU, LoneAN, KhanMS, et al., 2021. Effect of omega-3 fatty acids on cardiovascular outcomes: a systematic review and meta-analysis. eClinicalMedicine, 38:100997.

[19]KoláčkováP, RůžičkováG, GregorT, et al., 2015. Quick method (FT-NIR) for the determination of oil and major fatty acids content in whole achenes of milk thistle (Silybum marianum (L.) Gaertn.). J Sci Food Agric, 95(11):2264-2270.

[20]KosmopoulosA, BhattDL, MeglisG, et al., 2021. A randomized trial of icosapent ethyl in ambulatory patients with COVID-19. iScience, 24(9):103040.

[21]Lamon-FavaS, SoJ, MischoulonD, et al., 2021. Dose- and time-dependent increase in circulating anti-inflammatory and pro-resolving lipid mediators following eicosapentaenoic acid supplementation in patients with major depressive disorder and chronic inflammation. Prostaglandins Leukot Essent Fatty Acids, 164:102219.

[22]LeeTH, ChangJS, WangHY, 2013. Rapid and in vivo quantification of cellular lipids in Chlorella vulgaris using near-infrared Raman spectrometry. Anal Chem, 85(4):2155-2160.

[23]LingXP, GuoJ, LiuXT, et al., 2015. Impact of carbon and nitrogen feeding strategy on high production of biomass and docosahexaenoic acid (DHA) by Schizochytrium sp. LU310. Bioresour Technol, 184:139-147.

[24]LiuL, BaiMH, ZhangS, et al., 2021. ARTP mutagenesis of Schizochytrium sp. PKU#Mn4 and clethodim-based mutant screening for enhanced docosahexaenoic acid accumulation. Mar Drugs, 19(10):564.

[25]LiuYY, TianYY, CaiWZ, et al., 2021. DHA/EPA-enriched phosphatidylcholine suppresses tumor growth and metastasis via activating peroxisome proliferator-activated receptor γ in Lewis lung cancer mice. J Agric Food Chem, 69(2):676-685.

[26]MorabitY, HasanMI, WhalleyRD, et al., 2021. A review of the gas and liquid phase interactions in low-temperature plasma jets used for biomedical applications. Eur Phys J D, 75:32.

[27]OuY, LiYQ, FengSS, et al., 2023. Transcriptome analysis reveals an eicosapentaenoic acid accumulation mechanism in a Schizochytrium sp. mutant. Microbiol Spectr, 11(3):e0013023.

[28]PengZL, ZhangC, YanL, et al., 2020. EPA is more effective than DHA to improve depression-like behavior, glia cell dysfunction and hippcampal apoptosis signaling in a chronic stress-induced rat model of depression. Int J Mol Sci, 21(5):1769.

[29]Preston MasonR, 2019. New insights into mechanisms of action for omega-3 fatty acids in atherothrombotic cardiovascular disease. Curr Atheroscler Rep, 21:2.

[30]RatledgeC, HopkinsS, 2006. Applications and safety of microbial oils in food. In: Gunstone FD (Ed.), Modifying Lipids for Use in Food. Woodhead Publishing, Cambridge, p.567-586.

[31]ReisMG, AgnewM, JacobN, et al., 2022. Comparative evaluation of miniaturized and conventional NIR spectrophotometer for estimation of fatty acids in cheeses. Spectrochim Acta Part A Mol Biomol Spectrosc, 279:121433.

[32]RenLJ, FengY, LiJ, et al., 2013. Impact of phosphate concentration on docosahexaenoic acid production and related enzyme activities in fermentation of Schizochytrium sp. Bioprocess Biosyst Eng, 36(9):1177-1183.

[33]RosenbergSM, 2013. Reverse mutation. In: Maloy S, Hughes K (Eds.), Brenner’s Encyclopedia of Genetics, 2nd Ed. Academic Press, Amsterdam, p.220-221.

[34]ShenJ, ZhangH, XuZM, et al., 2019. Preferential production of reactive species and bactericidal efficacy of gas-liquid plasma discharge. Chem Eng J, 362:402-412.

[35]SheneC, ParedesP, VergaraD, et al., 2020. Antarctic Thraustochytrids: producers of long-chain omega-3 polyunsaturated fatty acids. MicrobiologyOpen, 9:e00950.

[36]SunXM, RenLJ, BiZQ, et al., 2018a. Adaptive evolution of microalgae Schizochytrium sp. under high salinity stress to alleviate oxidative damage and improve lipid biosynthesis. Bioresour Technol, 267:438-444.

[37]SunXM, RenLJ, BiZQ, et al., 2018b. Development of a cooperative two-factor adaptive-evolution method to enhance lipid production and prevent lipid peroxidation in Schizochytrium sp. Biotechnol Biofuels, 11:65.

[38]TsegayG, AmmareY, MesfinS, 2023. Development of non-destructive NIRS models to predict oil and major fatty acid contents of Ethiopian sesame. J Food Compos Anal, 115:104908.

[39]WangQY, LiZG, MaZH, et al., 2014. Real time monitoring of multiple components in wine fermentation using an on-line auto-calibration Raman spectroscopy. Sens Actuators B Chem, 202:426-432.

[40]WangS, WanWJ, WangZJ, et al., 2021. A two-stage adaptive laboratory evolution strategy to enhance docosahexaenoic acid synthesis in oleaginous thraustochytrid. Front Nutr, 8:795491.

[41]WeiXY, WangYZ, LiuXN, et al., 2023. Metabolic analysis of Schizochytrium sp. mutants with high EPA content achieved with ARTP mutagenesis screening. Bioprocess Biosyst Eng, 46(6):893-901.

[42]XiaoBP, LiYY, LinYQ, et al., 2022. Eicosapentaenoic acid (EPA) exhibits antioxidant activity via mitochondrial modulation. Food Chem, 373:131389.

[43]YinFW, ZhangYT, JiangJY, et al., 2019. Efficient docosahexaenoic acid production by Schizochytrium sp. via a two-phase pH control strategy using ammonia and citric acid as pH regulators. Process Biochem, 77:1-7.

[44]YinHW, LiuYY, YueH, et al., 2022. DHA- and EPA-enriched phosphatidylcholine suppress human lung carcinoma 95D cells metastasis via activating the peroxisome proliferator-activated receptor γ. Nutrients, 14(21):4675.

[45]YuHW, LiuHZ, WangQ, et al., 2020. Evaluation of portable and benchtop NIR for classification of high oleic acid peanuts and fatty acid quantitation. LWT, 128:109398.

[46]YueXH, ChenWC, WangZM, et al., 2019. Lipid distribution pattern and transcriptomic insights revealed the potential mechanism of docosahexaenoic acid traffics in Schizochytrium sp. A-2. J Agric Food Chem, 67(34):9683-9693.

[47]ZengL, BiYQ, GuoPF, et al., 2021. Metabolic analysis of Schizochytrium mutants with high DHA content achieved with ARTP mutagenesis combined with iodoacetic acid and dehydroepiandrosterone screening. Front Bioeng Biotechnol, 9:738052.

[48]ZengWZ, GuoLK, XuS, et al., 2020. High-throughput screening technology in industrial biotechnology. Trends Biotechnol, 38(8):888-906.

[49]ZhuYF, HuangXY, HanT, et al., 2025. Improvement of neutral protease activity of Bacillus amyloliquefaciens LX-6 by combined ribosome engineering and medium optimization. J Zhejiang Univ-Sci B (Biomed & Biotechnol), 26(8):805-812.