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Conventional fish-oil supply chains remain vulnerable to ecological pressure, compositional variability, and contaminants. Microbial production offers a controllable alternative, yet commercial feasibility depends on numerous factors beyond the strain yield. This review summarizes DHA-producing microorganisms and their metabolic pathways, including precursor supply, biosynthetic and storage pathways, and downstream separation and purification. Various separation strategies, ranging from crude lipid extraction to high-purity DHA, are evaluated in terms of recovery, selectivity, scale, cost, and oxidation risk. We discussed the synergistic mixed-fermentation and refining strategy that coproduces carotenoids, astaxanthin, and FAs to enhance the economic value. To achieve commercial competitiveness, a comprehensive approach combining metabolic engineering, fermentation optimization, and separation engineering is required to establish a microbial platform as an environmentally friendly and sustainable alternative to DHA from animal sources.<\/jats:p>","DOI":"10.1021\/acs.jafc.6c06093","type":"journal-article","created":{"date-parts":[[2026,10,5]],"date-time":"2026-10-05T15:02:49Z","timestamp":1791212569000},"page":"31290-31304","source":"Crossref","is-referenced-by-count":0,"title":["Sustainable Microbial Production and Application of\nDHA: From Laboratory to Shelf"],"prefix":"10.1021","volume":"74","author":[{"given":"Yi","family":"He","sequence":"first","affiliation":[{"name":"Nanjing Normal University , , ,","place":["Nanjing, China, 210023"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ying","family":"Dai","sequence":"additional","affiliation":[{"name":"Nanjing Normal University , , ,","place":["Nanjing, China, 210023"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wenhe","family":"Shen","sequence":"additional","affiliation":[{"name":"Nanjing Normal University , , ,","place":["Nanjing, China, 210023"]},{"name":"Zhihe Biotechnology (Changzhou) Co., LTD , ,","place":["Changzhou, China, 213125"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dongsheng","family":"Guo","sequence":"additional","affiliation":[{"name":"Nanjing Normal University , , ,","place":["Nanjing, China, 210023"]},{"name":"Zhihe Biotechnology (Changzhou) Co., LTD , ,","place":["Changzhou, China, 213125"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jie","family":"Zheng","sequence":"additional","affiliation":[{"name":"Nanjing Normal University , , ,","place":["Nanjing, China, 210023"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2699-3448","authenticated-orcid":true,"given":"Xing","family":"Zhang","sequence":"additional","affiliation":[{"name":"Nanjing Normal University , , ,","place":["Nanjing, China, 210023"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2026,10,5]]},"reference":[{"key":"2026101408093134900_cit1","doi-asserted-by":"publisher","first-page":"116802","DOI":"10.1016\/j.foodres.2025.116802","article-title":"Recent advances\nin DHA-containing phospholipids (PL-DHA): Sources, position-specific\neffects, metabolic pathways, and biological activities","volume":"218","author":"He","year":"2025","journal-title":"Food Res. Int."},{"issue":"4","key":"2026101408093134900_cit2","first-page":"675","article-title":"Docosahexaenoic\nacid and the brain-what is its role?","volume":"28","author":"Sinclair","year":"2019","journal-title":"Asia Pac.\nJ. Clin. Nutr."},{"issue":"23","key":"2026101408093134900_cit3","doi-asserted-by":"publisher","first-page":"11542","DOI":"10.3390\/ijms262311542","article-title":"The quantum brain: the untold story\nof docosahexaenoic acid\u2019s role in brain evolution, biophysics,\nand cognition","volume":"26","author":"Crawford","year":"2025","journal-title":"Int. J. Mol. Sci."},{"issue":"6","key":"2026101408093134900_cit4","doi-asserted-by":"publisher","first-page":"256","DOI":"10.3390\/md23060256","article-title":"A review on marine microbial docosahexaenoic\nacid production through\ncircular economy, fermentation engineering, and antioxidant technology","volume":"23","author":"Yin","year":"2025","journal-title":"Mar. Drugs"},{"issue":"9","key":"2026101408093134900_cit5","doi-asserted-by":"publisher","first-page":"310","DOI":"10.3390\/md16090310","article-title":"Transcriptomic\nprofiling and gene disruption revealed that two genes related to PUFAs\/DHA\nbiosynthesis may be essential for cell growth of Aurantiochytrium\nsp","volume":"16","author":"Liang","year":"2018","journal-title":"Mar. Drugs"},{"issue":"5","key":"2026101408093134900_cit6","doi-asserted-by":"publisher","first-page":"656","DOI":"10.1080\/07388551.2016.1213221","article-title":"Technological trends and market perspectives\nfor production of microbial oils rich in omega-3","volume":"37","author":"Finco","year":"2017","journal-title":"Crit. Rev. Biotechnol."},{"issue":"1","key":"2026101408093134900_cit7","doi-asserted-by":"publisher","first-page":"4055","DOI":"10.1038\/s41467-019-12025-8","article-title":"Polyunsaturated\nfatty acid production by Yarrowia lipolytica employing designed myxobacterial\nPUFA synthases","volume":"10","author":"Gemperlein","year":"2019","journal-title":"Nat. Commun."},{"key":"2026101408093134900_cit8","doi-asserted-by":"publisher","first-page":"380","DOI":"10.1016\/j.algal.2017.07.031","article-title":"De novo transcriptomic and metabolomic\nanalysis of docosahexaenoic\nacid (DHA)-producing Crypthecodinium cohnii during fed-batch fermentation","volume":"26","author":"Pei","year":"2017","journal-title":"Algal Res."},{"key":"2026101408093134900_cit9","doi-asserted-by":"publisher","first-page":"132292","DOI":"10.1016\/j.biortech.2025.132292","article-title":"Unveiling novel\noxygen-dependent docosahexaenoic acid biosynthesis pathway in Crypthecodinium\ncohnii via dual-function\u25b3 4-fatty acid desaturases","volume":"424","author":"Yao","year":"2025","journal-title":"Bioresour. 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Oleo Sci."},{"issue":"2","key":"2026101408093134900_cit12","doi-asserted-by":"publisher","first-page":"357","DOI":"10.1042\/BJ20040970","article-title":"Identification\nof two novel microalgal enzymes involved in the conversion of the\n\u03c93-fatty acid, eicosapentaenoic acid, into docosahexaenoic acid","volume":"384","author":"Pereira","year":"2004","journal-title":"Biochem. J."},{"issue":"9","key":"2026101408093134900_cit13","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0162861","article-title":"DHA production in Escherichia coli by expressing reconstituted key\ngenes of polyketide synthase pathway from marine bacteria","volume":"11","author":"Peng","year":"2016","journal-title":"PLoS One"},{"issue":"3","key":"2026101408093134900_cit14","doi-asserted-by":"publisher","first-page":"504","DOI":"10.1111\/tpj.12787","article-title":"Metabolism of acyl-lipids\nin Chlamydomonas\nreinhardtii","volume":"82","author":"Li-Beisson","year":"2015","journal-title":"Plant J."},{"issue":"5","key":"2026101408093134900_cit15","doi-asserted-by":"publisher","first-page":"509","DOI":"10.1080\/17597269.2022.2151451","article-title":"High yield production\nof lipid and carotenoids in a newly isolated Rhodotorula mucilaginosa\nby adapting process optimization approach","volume":"14","author":"Gedela","year":"2023","journal-title":"Biofuels,\nBioprod. Biorefin."},{"key":"2026101408093134900_cit16","doi-asserted-by":"publisher","first-page":"3","DOI":"10.1016\/j.ymben.2013.12.003","article-title":"Metabolic\nengineering\nof Phaeodactylum tricornutum for the enhanced accumulation of omega-3\nlong chain polyunsaturated fatty acids","volume":"22","author":"Hamilton","year":"2014","journal-title":"Metab.\nEng."},{"issue":"1","key":"2026101408093134900_cit17","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1155\/2014\/657414","article-title":"Production\nand enhancement of omega-3 fatty acid from\nMortierella alpina CFR-GV15: its food and therapeutic application","volume":"2014","author":"Vadivelan","year":"2014","journal-title":"BioMed Res. Int."},{"key":"2026101408093134900_cit18","doi-asserted-by":"publisher","first-page":"98","DOI":"10.1016\/j.ymben.2015.11.001","article-title":"Metabolic engineering of Pseudomonas putida for production\nof docosahexaenoic acid based on a myxobacterial PUFA synthase","volume":"33","author":"Gemperlein","year":"2016","journal-title":"Metab. Eng."},{"key":"2026101408093134900_cit19","doi-asserted-by":"publisher","first-page":"1104","DOI":"10.3389\/fmicb.2020.01104","article-title":"Bioconversion\nfrom docosahexaenoic acid to eicosapentaenoic acid in the marine bacterium\nShewanella livingstonensis Ac10","volume":"11","author":"Ogawa","year":"2020","journal-title":"Front. 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Int."},{"key":"2026101408093134900_cit23","doi-asserted-by":"publisher","first-page":"103127","DOI":"10.1016\/j.algal.2023.103127","article-title":"Adaptive evolution\nof Schizochytrium sp. under light and H2O2 condition to regulate its\nfatty acid and terpene biosynthesis","volume":"72","author":"Zhang","year":"2023","journal-title":"Algal Res."},{"issue":"1","key":"2026101408093134900_cit24","doi-asserted-by":"publisher","first-page":"263","DOI":"10.1007\/s10811-020-02304-z","article-title":"Evaluating\nlow-cost\nsubstrates for Crypthecodinium cohnii lipids and DHA production, by\nflow cytometry","volume":"33","author":"Taborda","year":"2021","journal-title":"J. Appl. Phycol."},{"key":"2026101408093134900_cit25","doi-asserted-by":"publisher","first-page":"88","DOI":"10.1016\/j.ymben.2018.10.004","article-title":"Rewiring metabolic\nnetwork by chemical\nmodulator based laboratory evolution doubles lipid production in Crypthecodinium\ncohnii","volume":"51","author":"Diao","year":"2019","journal-title":"Metab. Eng."},{"key":"2026101408093134900_cit26","doi-asserted-by":"publisher","first-page":"51","DOI":"10.1016\/j.biortech.2014.11.046","article-title":"A strategy for the highly efficient\nproduction of docosahexaenoic\nacid by Aurantiochytrium limacinum SR21 using glucose and glycerol\nas the mixed carbon sources","volume":"177","author":"Li","year":"2015","journal-title":"Bioresour. Technol."},{"key":"2026101408093134900_cit27","doi-asserted-by":"publisher","first-page":"75","DOI":"10.1016\/j.algal.2018.07.010","article-title":"Enhanced docosahexaenoic\nacid production\nby Crypthecodinium cohnii under combined stress in two-stage cultivation\nwith date syrup based medium","volume":"34","author":"Rumiani","year":"2018","journal-title":"Algal Res."},{"key":"2026101408093134900_cit28","doi-asserted-by":"publisher","first-page":"1041","DOI":"10.1002\/bbb.70100","article-title":"Insights from meta-analysis for efficient\ndocosahexaenoic acid production in thraustochytrids through advanced\ncultivation and fermentation strategies","volume":"20","author":"Da Costa","year":"2026","journal-title":"Biofuels,\nBioprod. Biorefin."},{"key":"2026101408093134900_cit29","doi-asserted-by":"publisher","first-page":"130532","DOI":"10.1016\/j.biortech.2024.130532","article-title":"Enhanced fatty acid storage combined\nwith the multi-factor optimization of fermentation for high-level\nproduction of docosahexaenoic acid in Schizochytrium sp","volume":"398","author":"Jia","year":"2024","journal-title":"Bioresour. Technol."},{"key":"2026101408093134900_cit30","doi-asserted-by":"publisher","first-page":"102868","DOI":"10.1016\/j.algal.2022.102868","article-title":"Transcriptome\nanalyses reveal the DHA enhancement mechanism in Schizochytrium limacinum\nLD11 mutant","volume":"67","author":"Chen","year":"2022","journal-title":"Algal Res."},{"key":"2026101408093134900_cit31","doi-asserted-by":"publisher","first-page":"107725","DOI":"10.1016\/j.biotechadv.2021.107725","article-title":"Biotechnological production of lipid\nand terpenoid from thraustochytrids","volume":"48","author":"Du","year":"2021","journal-title":"Biotechnol.\nAdv."},{"issue":"1","key":"2026101408093134900_cit32","doi-asserted-by":"publisher","first-page":"8909","DOI":"10.1038\/s41598-018-27309-0","article-title":"Optimization\nof culture conditions for enhanced growth, lipid and docosahexaenoic\nacid (DHA) production of Aurantiochytrium SW1 by response surface\nmethodology","volume":"8","author":"Nazir","year":"2018","journal-title":"Sci. Rep."},{"key":"2026101408093134900_cit33","doi-asserted-by":"publisher","first-page":"100688","DOI":"10.1016\/j.biteb.2021.100688","article-title":"Isolation and characterization of\ndocosahexaenoic acid-producing\nnovel strain Aurantiochytrium sp. ICTFD5: A sterol with vitamin d-cholecalciferol,\nand cellulase and lipase producing thraustochytrid","volume":"14","author":"Bagul","year":"2021","journal-title":"Bioresour. Technol."},{"key":"2026101408093134900_cit34","doi-asserted-by":"publisher","first-page":"915773","DOI":"10.3389\/fmicb.2022.915773","article-title":"Comparative transcriptomic\nand lipidomic analyses indicate that cold\nstress enhanced the production of the long C18\u2013C22 polyunsaturated\nfatty acids in Aurantiochytrium sp","volume":"13","author":"Song","year":"2022","journal-title":"Front. Microbiol."},{"key":"2026101408093134900_cit35","doi-asserted-by":"publisher","first-page":"102122","DOI":"10.1016\/j.bcab.2021.102122","article-title":"Evaluation\nof diverse\nbiochemical stimulants to enhance growth, lipid and docosahexaenoic\nacid (DHA) production of Aurantiochytrium Sp. ATCC PRA-276","volume":"36","author":"Hussain","year":"2021","journal-title":"Biocatal. Agric. Biotechnol."},{"issue":"3","key":"2026101408093134900_cit36","doi-asserted-by":"publisher","first-page":"207","DOI":"10.4067\/s0718-95162010000100002","article-title":"Microbial oils and fatty acids: effect\nof carbon source on docosahexaenoic acid (C22:6 n-3, DHA) production\nby thraustochytrid strains","volume":"10","author":"Shene","year":"2010","journal-title":"J. Soil Sci."},{"issue":"1","key":"2026101408093134900_cit37","doi-asserted-by":"publisher","first-page":"1378","DOI":"10.1038\/s42003-021-02857-w","article-title":"PUFA synthase-independent\nDHA synthesis pathway in Parietichytrium sp. and its modification\nto produce EPA and n-3DPA","volume":"4","author":"Ishibashi","year":"2021","journal-title":"Commun. Biol."},{"key":"2026101408093134900_cit38","doi-asserted-by":"publisher","first-page":"257","DOI":"10.1016\/j.biortech.2018.05.104","article-title":"Long-term\nculture of the marine dinoflagellate microalga Amphidinium carterae\nin an indoor LED-lighted raceway photobioreactor: Production of carotenoids\nand fatty acids","volume":"265","author":"Molina-Miras","year":"2018","journal-title":"Bioresour. Technol."},{"issue":"34","key":"2026101408093134900_cit39","doi-asserted-by":"publisher","first-page":"9667","DOI":"10.1021\/acs.jafc.9b02821","article-title":"Production of amphidinols and other bioproducts\nof interest by the marine microalga Amphidinium carterae unraveled\nby nuclear magnetic resonance metabolomics approach coupled to multivariate\ndata analysis","volume":"67","author":"Abreu","year":"2019","journal-title":"J. Agric. Food Chem."},{"issue":"4","key":"2026101408093134900_cit40","doi-asserted-by":"publisher","first-page":"218","DOI":"10.1038\/sj.jim.7000056","article-title":"Heterotrophic production\nof eicosapentaenoid acid by the diatom Nitzschia\nlaevis: Effects of silicate and glucose","volume":"25","author":"Wen","year":"2000","journal-title":"J.\nInd. Microbiol. Biotechnol."},{"issue":"4","key":"2026101408093134900_cit41","doi-asserted-by":"publisher","first-page":"874","DOI":"10.3390\/molecules23040874","article-title":"Fucoxanthin and polyunsaturated fatty acids co-extraction\nby a green process","volume":"23","author":"Delbrut","year":"2018","journal-title":"Molecules"},{"key":"2026101408093134900_cit42","doi-asserted-by":"publisher","first-page":"101506","DOI":"10.1016\/j.algal.2019.101506","article-title":"Betaine lipid and neutral\nlipid production\nunder nitrogen or phosphorus limitation in the marine microalga Tisochrysis\nlutea (Haptophyta)","volume":"40","author":"Huang","year":"2019","journal-title":"Algal Res."},{"issue":"1","key":"2026101408093134900_cit43","doi-asserted-by":"publisher","first-page":"130","DOI":"10.1186\/s40643-022-00612-5","article-title":"Simultaneous\nphotoautotrophic production of DHA and EPA by Tisochrysis lutea and\nMicrochloropsis salina in co-culture","volume":"9","author":"Thurn","year":"2022","journal-title":"Bio. Bioprocessing"},{"issue":"8","key":"2026101408093134900_cit44","doi-asserted-by":"publisher","first-page":"2237","DOI":"10.1021\/jf025952y","article-title":"Lipid class\ncomposition of the microalga\nPavlova lutheri: eicosapentaenoic and docosahexaenoic acids","volume":"51","author":"Meireles","year":"2003","journal-title":"J. Agric. Food Chem."},{"issue":"1","key":"2026101408093134900_cit45","doi-asserted-by":"publisher","first-page":"157","DOI":"10.1007\/s13213-013-0645-6","article-title":"Effects of photoperiod, salinity and pH on cell growth\nand lipid content of Pavlova lutheri","volume":"64","author":"Shah","year":"2014","journal-title":"Ann. Microbiol."},{"issue":"3\u20134","key":"2026101408093134900_cit46","doi-asserted-by":"publisher","first-page":"358","DOI":"10.1016\/j.enzmictec.2005.05.014","article-title":"Metabolic relationships\nbetween macro-and micronutrients, and the eicosapentaenoic acid and\ndocosahexaenoic acid contents of Pavlova lutheri","volume":"38","author":"Carvalho","year":"2006","journal-title":"Enzyme Microb. Technol."},{"issue":"12","key":"2026101408093134900_cit47","doi-asserted-by":"publisher","first-page":"2265","DOI":"10.1007\/s10529-012-1028-y","article-title":"Identification\nof a novel C22-\u0394 4-producing docosahexaenoic acid (DHA) specific\npolyunsaturated fatty acid desaturase gene from Isochrysis galbana\nand its expression in Saccharomyces cerevisiae","volume":"34","author":"Shi","year":"2012","journal-title":"Biotechnol. Lett."},{"issue":"6","key":"2026101408093134900_cit48","doi-asserted-by":"publisher","first-page":"1903","DOI":"10.1099\/00221287-148-6-1903","article-title":"Structure and regulation of the omega-3\npolyunsaturated\nfatty acid synthase genes from the deep-sea bacterium Photobacterium\nprofundum strain SS9","volume":"148","author":"Allen","year":"2002","journal-title":"Microbiology"},{"issue":"1","key":"2026101408093134900_cit49","doi-asserted-by":"publisher","first-page":"30","DOI":"10.1186\/s12934-016-0431-9","article-title":"Effect of\ncerulenin on fatty acid composition and gene\nexpression pattern of DHA-producing strain Colwellia psychrerythraea\nstrain 34H","volume":"15","author":"Wan","year":"2016","journal-title":"Microb. Cell Fact."},{"issue":"7","key":"2026101408093134900_cit50","doi-asserted-by":"publisher","first-page":"2859","DOI":"10.1007\/s00253-012-4529-7","article-title":"Growth condition\noptimization for\ndocosahexaenoic acid (DHA) production by Moritella marina MP-1","volume":"97","author":"Kautharapu","year":"2013","journal-title":"Appl. Microbiol. 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