{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,18]],"date-time":"2026-08-18T04:58:31Z","timestamp":1787029111611,"version":"build-2736575974"},"reference-count":60,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2026,5,11]],"date-time":"2026-05-11T00:00:00Z","timestamp":1778457600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Molecules"],"abstract":"<jats:p>This study investigates the extraction of perilla seed oil using an ultrasound\u2013ethanol pretreatment combined with aqueous enzymatic extraction (UEAEE). By comparing different pretreatment groups, it was found that the combination of ultrasound and ethanol pretreatment yielded the highest oil extraction efficiency from perilla seeds. Meanwhile, scanning electron microscopy (SEM) analysis further revealed pronounced structural disruption in seeds subjected to ultrasound\u2013ethanol pretreatment, including extensive cellular collapse and the formation of numerous irregular pores and fissures, which facilitated subsequent oil release. The parameters of aqueous enzymatic extraction (AEE) were optimized using a Box\u2013Behnken design. The optimal conditions were determined to be a hydrolysis time of 5 h, a reaction temperature of 52 \u00b0C, an enzyme concentration of 5%, and a liquid-to-material ratio of 9.5:1 (mL\/g). Compared with oils obtained by pressing extraction (PE) and Soxhlet extraction (SE), UEAEE-derived oil exhibited a higher proportion of unsaturated fatty acids, along with lower peroxide and Acid value (AV), indicating superior quality and a reduced susceptibility to rancidity. Moreover, UEAEE oil retained markedly higher levels of micronutrients (carotenoids: 1.23 mg\/kg; total flavonoids: 1.04 mg\/g; total phenols: 2.79 mg\/g) and demonstrated stronger antioxidant activity (DPPH: 80.60%). Overall, these results demonstrate that UEAEE is an efficient and environmentally friendly extraction method that supports the high-value utilization of perilla seed oil and aligns with modern green processing principles.<\/jats:p>","DOI":"10.3390\/molecules31101608","type":"journal-article","created":{"date-parts":[[2026,5,11]],"date-time":"2026-05-11T12:05:22Z","timestamp":1778501122000},"page":"1608","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Ultrasound\u2013Ethanol Pretreatment-Assisted Enzymatic Method: A Potential Method to Improve the Quality and Yield of Perilla Seed Oil"],"prefix":"10.3390","volume":"31","author":[{"given":"Jinhua","family":"Shao","sequence":"first","affiliation":[{"name":"College of Chemistry and Bioengineering, Hunan University of Science and Engineering, Yongzhou 425199, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yichun","family":"Zhu","sequence":"additional","affiliation":[{"name":"College of Chemistry and Bioengineering, Hunan University of Science and Engineering, Yongzhou 425199, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Miaomiao","family":"Chang","sequence":"additional","affiliation":[{"name":"College of Chemistry and Bioengineering, Hunan University of Science and Engineering, Yongzhou 425199, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shengmei","family":"Tang","sequence":"additional","affiliation":[{"name":"College of Chemistry and Bioengineering, Hunan University of Science and Engineering, Yongzhou 425199, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Weizhen","family":"Huang","sequence":"additional","affiliation":[{"name":"College of Chemistry and Bioengineering, Hunan University of Science and Engineering, Yongzhou 425199, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Liyan","family":"Jiang","sequence":"additional","affiliation":[{"name":"College of Chemistry and Bioengineering, Hunan University of Science and Engineering, Yongzhou 425199, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,5,11]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"e33281","DOI":"10.1016\/j.heliyon.2024.e33281","article-title":"A Comprehensive Review on Nutritional, Nutraceutical, and Industrial Perspectives of Perilla (Perilla frutscens L.) Seeds\u2014An Orphan Oilseed Crop","volume":"10","author":"Kaur","year":"2024","journal-title":"Heliyon"},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Guan, L., Zhu, L., Zhang, X., Han, Y., Wang, K., Ji, N., Yao, X., Zhou, Y., Li, B., and Chen, Q. (2024). Perilla Seed Oil and Protein: Composition, Health Benefits, and Potential Applications in Functional Foods. Molecules, 29.","DOI":"10.3390\/molecules29225258"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"111457","DOI":"10.1016\/j.lwt.2021.111457","article-title":"Study on Biological Activity of Perilla Seed Oil Extracted by Supercritical Carbon Dioxide","volume":"146","author":"Hao","year":"2021","journal-title":"LWT"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"3183","DOI":"10.1080\/87559129.2024.2347469","article-title":"Comprehensive Characterisation of Perilla Seed Oil for Potential Applications as a Bioactive Food Ingredient-Review","volume":"40","author":"Zhong","year":"2024","journal-title":"Food Rev. Int."},{"key":"ref_5","doi-asserted-by":"crossref","unstructured":"Li, M., Jiang, N., Guo, G., Lu, S., Li, Z., Mu, Y., Xia, X., Xu, Z., Hu, Y., and Xiang, X. (2024). Perilla Seed Oil: A Review of Health Effects, Encapsulation Strategies and Applications in Food. Foods, 13.","DOI":"10.3390\/foods13223615"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"1160","DOI":"10.1111\/1750-3841.15070","article-title":"Influence of Refining Processes on the Bioactive Composition, in Vitro Antioxidant Capacity, and Their Correlation of Perilla Seed Oil","volume":"85","author":"Pan","year":"2020","journal-title":"J. Food Sci."},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Huang, S., Nan, Y., Chen, G., Ning, N., Du, Y., Lu, D., Yang, Y., Meng, F., and Yuan, L. (2023). The Role and Mechanism of Perilla Frutescens in Cancer Treatment. Molecules, 28.","DOI":"10.3390\/molecules28155883"},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Kangwan, N., Pratchayasakul, W., Kongkaew, A., Pintha, K., Chattipakorn, N., and Chattipakorn, S.C. (2021). Perilla Seed Oil Alleviates Gut Dysbiosis, Intestinal Inflammation and Metabolic Disturbance in Obese-Insulin-Resistant Rats. Nutrients, 13.","DOI":"10.3390\/nu13093141"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"116048","DOI":"10.1016\/j.lwt.2024.116048","article-title":"A Novel Stepwise Microwave Hydrodistillation and Extraction Process for Separating Seed Oil and Essential Oil Simultaneously from Perilla Seeds","volume":"198","author":"Feng","year":"2024","journal-title":"LWT"},{"key":"ref_10","first-page":"76","article-title":"Research on the Ultrasonic Extraction Process and Physicochemical Properties of Perilla Seed Oil","volume":"35","author":"Li","year":"2019","journal-title":"For. Eng."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1590\/1678-457x.29116","article-title":"Ultrasound-Assisted Aqueous Enzymatic Extraction of Oil from Perilla Seeds and Determination of Its Physicochemical Properties, Fatty Acid Composition and Antioxidant Activity","volume":"37","author":"Li","year":"2017","journal-title":"Food Sci. Technol."},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Prommaban, A., Kuanchoom, R., Seepuan, N., and Chaiyana, W. (2021). Evaluation of Fatty Acid Compositions, Antioxidant, and Pharmacological Activities of Pumpkin (Cucurbita moschata) Seed Oil from Aqueous Enzymatic Extraction. Plants, 10.","DOI":"10.3390\/plants10081582"},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Tang, L., Wang, T., Xu, Y., He, X., Yan, A., Zhang, Z., Li, Y., and Chen, G. (2024). Research and Application Progress of Crude Oil Demulsification Technology. Processes, 12.","DOI":"10.3390\/pr12102292"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"9147926","DOI":"10.1155\/2017\/9147926","article-title":"Research on Crude Oil Demulsification Using the Combined Method of Ultrasound and Chemical Demulsifier","volume":"2017","author":"Yi","year":"2017","journal-title":"J. Chem."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"106766","DOI":"10.1016\/j.ultsonch.2024.106766","article-title":"Ultrasonic-Ethanol Pretreatment Assisted Aqueous Enzymatic Extraction of Hemp Seed Oil with Low \u03949-Thc","volume":"103","author":"Zhang","year":"2024","journal-title":"Ultrason. Sonochemistry"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"112405","DOI":"10.1016\/j.indcrop.2020.112405","article-title":"Combining Integrated Ultrasonic-Microwave Technique with Ethanol to Maximise Extraction of Green Coffee Oil from Arabica Coffee Beans","volume":"151","author":"Chen","year":"2020","journal-title":"Ind. Crops Prod."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"104315","DOI":"10.1016\/j.tifs.2023.104315","article-title":"Aqueous Enzymatic Extraction: A Green, Environmentally Friendly and Sustainable Oil Extraction Technology","volume":"144","author":"Gao","year":"2024","journal-title":"Trends Food Sci. Technol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"59","DOI":"10.1016\/j.procbio.2020.09.033","article-title":"Ultrasonic-assisted enzymolysis: Principle and applications","volume":"100","author":"Umego","year":"2021","journal-title":"Process Biochem."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"104933","DOI":"10.1016\/j.ultsonch.2019.104933","article-title":"Molecular aggregation and property changes of egg yolk low-density lipoprotein induced by ethanol and high-density ultrasound","volume":"63","author":"Xie","year":"2020","journal-title":"Ultrason. Sonochemistry"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"109307","DOI":"10.1016\/j.cep.2023.109307","article-title":"Enhanced castor seed oil extraction assisted by the synergistic effect of ultrasound and microwave: Impact on extraction effectiveness and oil quality","volume":"185","author":"Thakkar","year":"2023","journal-title":"Chem. Eng. Process.-Process Intensif."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"133010","DOI":"10.1016\/j.foodchem.2022.133010","article-title":"Effect of Microwave Pretreatment of Perilla Seeds on Minor Bioactive Components Content and Oxidative Stability of Oil","volume":"388","author":"Huang","year":"2022","journal-title":"Food Chem."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"513","DOI":"10.1002\/aocs.12793","article-title":"Effect of Infrared Roasting of Perilla Seeds on the Content of Bioactive Components and Antioxidant Capacity in Oil","volume":"101","author":"Huang","year":"2024","journal-title":"J. Am. Oil Chem. Soc."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"109026","DOI":"10.1016\/j.lwt.2020.109026","article-title":"Effect of Freeze-Thaw Pretreatment on Yield and Quality of Perilla Seed Oil","volume":"122","author":"Lee","year":"2020","journal-title":"LWT"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"543","DOI":"10.1016\/j.ultsonch.2005.09.009","article-title":"Ultrasound-Assisted Extraction of Anthraquinones from Roots of Morinda citrifolia","volume":"13","author":"Hemwimol","year":"2006","journal-title":"Ultrason. Sonochemistry"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"1222","DOI":"10.1007\/s13197-022-05359-7","article-title":"A Review on Application of Ultrasound and Ultrasound Assisted Technology for Seed Oil Extraction","volume":"60","author":"Thilakarathna","year":"2023","journal-title":"J. Food Sci. Technol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1826","DOI":"10.1080\/10408398.2019.1602823","article-title":"Combination of Emerging Technologies for the Extraction of Bioactive Compounds","volume":"60","author":"Wen","year":"2020","journal-title":"Crit. Rev. Food Sci. Nutr."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"105625","DOI":"10.1016\/j.ultsonch.2021.105625","article-title":"Review of Ultrasound Combinations with Hybrid and Innovative Techniques for Extraction and Processing of Food and Natural Products","volume":"76","author":"Khadhraoui","year":"2021","journal-title":"Ultrason. Sonochemistry"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"428","DOI":"10.1016\/j.ultsonch.2015.01.009","article-title":"Effect of Ultrasound Radiation Duration on Emulsification and Demulsification of Paraffin Oil and Surfactant Solution\/Brine Using Hele-Shaw Models","volume":"26","author":"Hamidi","year":"2015","journal-title":"Ultrason. Sonochemistry"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"104","DOI":"10.1007\/s42452-024-05714-7","article-title":"Optimization of Ultrasound-Assisted Extraction of Nigella Sativa Seed Oil for Enhancement of Yield and Antioxidant Activity","volume":"6","author":"Dar","year":"2024","journal-title":"Discov. Appl. Sci."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"e14192","DOI":"10.1111\/jfpe.14192","article-title":"Advances in Seed Oil Extraction Using Ultrasound Assisted Technology: A Comprehensive Review","volume":"46","author":"Junaid","year":"2023","journal-title":"J. Food Process Eng."},{"key":"ref_31","first-page":"177","article-title":"Effect of ultrasonic-assisted enzymatic extraction on the release of high-quality pecan nuts oil","volume":"48","author":"Wang","year":"2022","journal-title":"Food Ferment. Ind."},{"key":"ref_32","first-page":"2089","article-title":"Study on water enzymatic extraction of pine seed oil and enzymatic demulsification technology","volume":"36","author":"Zhang","year":"2024","journal-title":"Acta Agric. Zhejiangensis"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"402","DOI":"10.1016\/S0141-0229(96)80004-F","article-title":"Aqueous and Enzymatic Processes for Edible Oil Extraction","volume":"19","author":"Rosenthal","year":"1996","journal-title":"Enzyme Microb. Technol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"50","DOI":"10.1016\/j.chroma.2018.12.027","article-title":"A Method for Extracting Oil from Cherry Seed by Ultrasonic-Microwave Assisted Aqueous Enzymatic Process and Evaluation of Its Quality","volume":"1587","author":"Hu","year":"2019","journal-title":"J. Chromatogr. A"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"113811","DOI":"10.1016\/j.indcrop.2021.113811","article-title":"Aqueous Enzymatic Extraction of Ricinus Communis Seeds Oil Using Viscozyme L.","volume":"170","author":"Rodrigues","year":"2021","journal-title":"Ind. Crops Prod."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"6483","DOI":"10.1007\/s13399-024-05611-4","article-title":"Pomegranate Seed Oil Extraction by Cold Pressing, Microwave and Ultrasound Treatments","volume":"15","author":"Uyar","year":"2025","journal-title":"Biomass Convers. Biorefinery"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"1749","DOI":"10.5650\/jos.ess22119","article-title":"Efficient Ultrasonic-Assisted Aqueous Enzymatic Method for Pecan Nut Kernel Oil Extraction with Quality Analysis","volume":"71","author":"Liu","year":"2022","journal-title":"J. Oleo Sci."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"e12696","DOI":"10.1111\/jfpe.12696","article-title":"Optimization of Ultrasound-Assisted Enzymatic Extraction of Walnut Kernel Oil Using Response Surface Methodology","volume":"41","author":"Ghasemi","year":"2018","journal-title":"J. Food Process Eng."},{"key":"ref_39","first-page":"1643","article-title":"Research Progress in Non-thermal Processing Assisted Aqueous Enzymatic Extraction of Vegetable Oil","volume":"37","author":"Wang","year":"2023","journal-title":"J. Nucl. Agric. Sci."},{"key":"ref_40","first-page":"188","article-title":"Research Progress on the Effect of Interfacial Proteins on the Stability of Emulsion during Aqueous Enzymatic Oil Extraction","volume":"44","author":"Li","year":"2023","journal-title":"Food Sci."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"28442","DOI":"10.1038\/s41598-024-78180-1","article-title":"Optimization of Process Parameters of Catalytic Pyrolysis Using Natural Zeolite and Synthetic Zeolites on Yield of Plastic Oil through Response Surface Methodology","volume":"14","author":"Sudalaimuthu","year":"2024","journal-title":"Sci. Rep."},{"key":"ref_42","first-page":"97","article-title":"Process Optimization of Bitter Almond Oil Extraction by Cold Pressing Combined with AqueousEnzymatic Extraction and Structural Characterization of Protein in the Resulting Cake","volume":"44","author":"Wu","year":"2023","journal-title":"Food Res. Dev."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"1258","DOI":"10.1007\/s13197-019-04158-x","article-title":"Comparison and Analysis Characteristics of Flax, Perilla and Basil Seed Oils Cultivated in Iran","volume":"57","author":"Ezzatpanah","year":"2020","journal-title":"J. Food Sci. Technol."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"100977","DOI":"10.1016\/j.scp.2023.100977","article-title":"Study of the Optimization and Kinetics of the Surfactant-Induced Ultrasonic-Assisted Extraction of Perilla Seed Oil: Free Radical Scavenging Capacity and Physicochemical and Functional Characteristics","volume":"32","author":"Li","year":"2023","journal-title":"Sustain. Chem. Pharm."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"167","DOI":"10.5650\/jos.ess18141","article-title":"Effects of Cold-Press and Soxhlet Extraction Systems on Antioxidant Activity, Total Phenol Contents, Fatty Acids, and Tocopherol Contents of Walnut Kernel Oils","volume":"68","author":"Ahmed","year":"2019","journal-title":"J. Oleo Sci."},{"key":"ref_46","unstructured":"Jia, X. (2021). Nutrition Evaluation of Chongqing Perilla Seed and the Effect of Pre-Heat Treatmenton the Quality of Extracted Oil. [Master\u2019s Thesis, Southwest University]."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"309","DOI":"10.1016\/j.foodres.2018.03.006","article-title":"Enzyme Assisted Extraction of Biomolecules as an Approach to Novel Extraction Technology: A Review","volume":"108","author":"Nadar","year":"2018","journal-title":"Food Res. Int."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"4976","DOI":"10.1002\/fsn3.1796","article-title":"Optimization of Ultrasound-Assisted Solvent Extraction of Hemp (Cannabis sativa L.) Seed Oil Using Rsm: Evaluation of Oxidative Stability and Physicochemical Properties of Oil","volume":"8","author":"Dehghan","year":"2020","journal-title":"Food Sci. Nutr."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"105325","DOI":"10.1016\/j.ultsonch.2020.105325","article-title":"Ultrasound Assisted Extraction (Uae) of Bioactive Compounds from Fruit and Vegetable Processing by- Products: A Review","volume":"70","author":"Kumar","year":"2021","journal-title":"Ultrason. Sonochemistry"},{"key":"ref_50","unstructured":"(1993). Oilseeds-Determination of Moisture and Volatile Matter Content (Standard No. GB\/T 14489.1-2008)."},{"key":"ref_51","unstructured":"(2008). Oilseeds-Determination of Oil Content (Standard No. GB\/T 14488.1-2008)."},{"key":"ref_52","first-page":"619","article-title":"Extraction of Perilla Seed Oil by Aqueous Enzymatic Method","volume":"39","author":"Yuan","year":"2019","journal-title":"Bull. Bot. Res."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"139046","DOI":"10.1016\/j.foodchem.2024.139046","article-title":"The effect of extraction methods on the components and quality of Camellia oleifera oil: Focusing on the flavor and lipidomics","volume":"447","author":"Zeng","year":"2024","journal-title":"Food Chem."},{"key":"ref_54","unstructured":"(2016). Determination of Fatty Acids in Foods (Standard No. GB 5009.168-2016)."},{"key":"ref_55","unstructured":"(2016). Determination of Acid Value in Foods (Standard No. GB 5009.229-2016)."},{"key":"ref_56","unstructured":"(2023). Determination of Peroxide Value in Foods (Standard No. GB 5009.227-2023)."},{"key":"ref_57","first-page":"233","article-title":"Effects of Processing Techniques on the Quality Properties and Lipid Concomitants of Flaxseed Oil","volume":"41","author":"Yu","year":"2020","journal-title":"Food Sci."},{"key":"ref_58","first-page":"257","article-title":"Effect of Steam Explosion Pretreatment on the Quality and Flavor of Pressed Perilla Seed Oil","volume":"46","author":"Zeng","year":"2025","journal-title":"Food Sci."},{"key":"ref_59","first-page":"23","article-title":"Effect of cold pressing and aqueous enzymatic extraction methods on the active ingredients of Carya cathayensis oil","volume":"47","author":"Li","year":"2022","journal-title":"China Oils Fats"},{"key":"ref_60","first-page":"72","article-title":"Preparation and Performance Analysis of Manchurian Walnut Oil Microcapsules with Manchurian Walnut Protein Isolate as Wall Material","volume":"45","author":"Li","year":"2024","journal-title":"Food Sci."}],"container-title":["Molecules"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1420-3049\/31\/10\/1608\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,5,11]],"date-time":"2026-05-11T12:10:52Z","timestamp":1778501452000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1420-3049\/31\/10\/1608"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,5,11]]},"references-count":60,"journal-issue":{"issue":"10","published-online":{"date-parts":[[2026,5]]}},"alternative-id":["molecules31101608"],"URL":"https:\/\/doi.org\/10.3390\/molecules31101608","relation":{},"ISSN":["1420-3049"],"issn-type":[{"value":"1420-3049","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,5,11]]}}}