Cascade valorization of okara by subcritical water hydrolysis: assessment of protein and amino acid profile in batch and semi-continuous systems

Cascade valorization of okara by subcritical water hydrolysis: assessment of protein and amino acid profile in batch and semi-continuous systems | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Cascade valorization of okara by subcritical water hydrolysis: assessment of protein and amino acid profile in batch and semi-continuous systems P. Barea, A. E. Illera, H. Candela, R. Melgosa, O. Benito-Román, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8653473/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 16 You are reading this latest preprint version Abstract This study assesses subcritical water hydrolysis (SWH) as a sustainable approach for protein recovery from okara, following isoflavone valorization. The initial extract, obtained at 120°C, contained over 1200 µg of isoflavones per gram of dry okara. The solid residue underwent a second protein extraction step in both batch and semicontinuous modes. The batch extraction was conducted at 180°C for 3 h at 5 MPa in a 0.5 L reactor. Under these conditions, total amino acid recovery reached 81 ± 3% (12.6 g/L), with 20 ± 1% of free amino acids, mainly glutamic acid, and a favourable profile of essential amino acids. These results were comparable to protein extraction from untreated okara, indicating that the prior isoflavone extraction did not hinder protein hydrolysis, but rather contributed positively within the cascade framework. Hydrolysis was also performed in a semi-continuous mode in a 2 L reactor, with a residence time of 2.4 min, in a two-stage temperature process (180°C and 270°C). The second stage yielded a significantly higher extraction than the first one, achieving a total 70.4% protein recovery (1.22 g/L) with 10% of free amino acids, with glutamic acid as the dominant one. Carbohydrate analysis revealed high hydrolysis yields for galactans, arabinans and xylans, but these exhibited lower thermal stability compared to proteins. Okara emerges as a promising by-product for a bio-cascade process, combining a mild first stage for isoflavone extraction with a more severe second stage for protein extraction/hydrolysis. Subcritical water okara isoflavones protein amino acid profile scaling up process Full Text Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Editorial decision: Revision requested 07 Mar, 2026 Reviews received at journal 05 Mar, 2026 Reviews received at journal 23 Feb, 2026 Reviews received at journal 09 Feb, 2026 Reviews received at journal 04 Feb, 2026 Reviewers agreed at journal 02 Feb, 2026 Reviews received at journal 01 Feb, 2026 Reviewers agreed at journal 30 Jan, 2026 Reviewers agreed at journal 29 Jan, 2026 Reviewers agreed at journal 28 Jan, 2026 Reviewers agreed at journal 28 Jan, 2026 Reviewers agreed at journal 28 Jan, 2026 Reviewers invited by journal 28 Jan, 2026 Editor assigned by journal 22 Jan, 2026 Submission checks completed at journal 21 Jan, 2026 First submitted to journal 20 Jan, 2026 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-8653473","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":582328860,"identity":"adc6d6e6-ac2d-4ffb-8063-c74690106aa5","order_by":0,"name":"P. Barea","email":"","orcid":"","institution":"University of Burgos","correspondingAuthor":false,"prefix":"","firstName":"P.","middleName":"","lastName":"Barea","suffix":""},{"id":582328861,"identity":"5b8f3c8a-8170-46f5-ab9b-13a0f701304d","order_by":1,"name":"A. E. Illera","email":"","orcid":"","institution":"University of Burgos","correspondingAuthor":false,"prefix":"","firstName":"A.","middleName":"E.","lastName":"Illera","suffix":""},{"id":582328864,"identity":"404e22cb-045c-4c5f-b94b-8eeca2902e03","order_by":2,"name":"H. Candela","email":"","orcid":"","institution":"University of Burgos","correspondingAuthor":false,"prefix":"","firstName":"H.","middleName":"","lastName":"Candela","suffix":""},{"id":582328865,"identity":"82916053-121c-4352-9c32-aeb2a68da327","order_by":3,"name":"R. Melgosa","email":"","orcid":"","institution":"University of Burgos","correspondingAuthor":false,"prefix":"","firstName":"R.","middleName":"","lastName":"Melgosa","suffix":""},{"id":582328866,"identity":"ea96e964-eb68-475d-98c2-95ea379cbcbf","order_by":4,"name":"O. Benito-Román","email":"","orcid":"","institution":"University of Burgos","correspondingAuthor":false,"prefix":"","firstName":"O.","middleName":"","lastName":"Benito-Román","suffix":""},{"id":582328867,"identity":"144bd548-43eb-48ca-b82c-d8f3fdee7bd5","order_by":5,"name":"M. T. Sanz","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAApklEQVRIiWNgGAWjYBADOQYJYpXyQGlj0rUkNhCtxZ699/GHnzvq0jfc7jFg+PCHGFt4jptJ9p5hy91w54wB48w2YrRIpLEx8Lbx5G64kWPAzNtAjBb5Z8wf/7ZJpBuAtPwhymESbAzSvG0GCWAtDGzEaDmTxiYt25ZgOPPOsYKDvcT4hb39GPPHt2118ny3mzc++EGMw1DAAVI1jIJRMApGwSjAAQB+DzHK/ZNk/QAAAABJRU5ErkJggg==","orcid":"","institution":"University of Burgos","correspondingAuthor":true,"prefix":"","firstName":"M.","middleName":"T.","lastName":"Sanz","suffix":""}],"badges":[],"createdAt":"2026-01-20 22:53:19","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-8653473/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-8653473/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":101751423,"identity":"e0d8ea7f-c3ad-48b7-a5b3-987f90f09b32","added_by":"auto","created_at":"2026-02-03 10:20:09","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":797471,"visible":true,"origin":"","legend":"","description":"","filename":"Paper2OKARAultima.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8653473/v1_covered_4f0b3eec-dcd1-4e4d-8f55-08e9211a7935.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Cascade valorization of okara by subcritical water hydrolysis: assessment of protein and amino acid profile in batch and semi-continuous systems","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"food-and-bioprocess-technology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"Learn more about [Food and Bioprocess Technology](https://www.springer.com/journal/11947)","snPcode":"11947","submissionUrl":"https://submission.nature.com/new-submission/11947/3","title":"Food and Bioprocess Technology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Subcritical water, okara, isoflavones, protein, amino acid profile, scaling up process","lastPublishedDoi":"10.21203/rs.3.rs-8653473/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8653473/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThis study assesses subcritical water hydrolysis (SWH) as a sustainable approach for protein recovery from okara, following isoflavone valorization. The initial extract, obtained at 120\u0026deg;C, contained over 1200 \u0026micro;g of isoflavones per gram of dry okara. The solid residue underwent a second protein extraction step in both batch and semicontinuous modes. The batch extraction was conducted at 180\u0026deg;C for 3 h at 5 MPa in a 0.5 L reactor. Under these conditions, total amino acid recovery reached 81\u0026thinsp;\u0026plusmn;\u0026thinsp;3% (12.6 g/L), with 20\u0026thinsp;\u0026plusmn;\u0026thinsp;1% of free amino acids, mainly glutamic acid, and a favourable profile of essential amino acids. These results were comparable to protein extraction from untreated okara, indicating that the prior isoflavone extraction did not hinder protein hydrolysis, but rather contributed positively within the cascade framework. Hydrolysis was also performed in a semi-continuous mode in a 2 L reactor, with a residence time of 2.4 min, in a two-stage temperature process (180\u0026deg;C and 270\u0026deg;C). The second stage yielded a significantly higher extraction than the first one, achieving a total 70.4% protein recovery (1.22 g/L) with 10% of free amino acids, with glutamic acid as the dominant one. Carbohydrate analysis revealed high hydrolysis yields for galactans, arabinans and xylans, but these exhibited lower thermal stability compared to proteins. Okara emerges as a promising by-product for a bio-cascade process, combining a mild first stage for isoflavone extraction with a more severe second stage for protein extraction/hydrolysis.\u003c/p\u003e","manuscriptTitle":"Cascade valorization of okara by subcritical water hydrolysis: assessment of protein and amino acid profile in batch and semi-continuous systems","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-01-30 02:55:48","doi":"10.21203/rs.3.rs-8653473/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2026-03-07T09:59:37+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-05T13:35:41+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-02-23T09:41:38+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-02-09T11:22:09+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-02-04T06:42:28+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"312743311802549137462331524081890618912","date":"2026-02-02T23:08:13+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-02-01T16:59:44+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"179823049614806470386510930902455403183","date":"2026-01-30T23:53:53+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"23395608185985582393637304790202644236","date":"2026-01-29T09:35:39+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"139931169495466150367882366611427438047","date":"2026-01-29T01:13:12+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"121524167784583675986786845783291747539","date":"2026-01-29T01:01:46+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"81001788242446977665266065472182992785","date":"2026-01-28T14:44:42+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-01-28T14:26:17+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-01-22T09:48:20+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2026-01-21T23:11:16+00:00","index":"","fulltext":""},{"type":"submitted","content":"Food and Bioprocess Technology","date":"2026-01-20T22:41:20+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"food-and-bioprocess-technology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"Learn more about [Food and Bioprocess Technology](https://www.springer.com/journal/11947)","snPcode":"11947","submissionUrl":"https://submission.nature.com/new-submission/11947/3","title":"Food and Bioprocess Technology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"83ae32f0-63ae-45bc-ba01-0fd354a9ff7b","owner":[],"postedDate":"January 30th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-05-05T01:38:20+00:00","versionOfRecord":[],"versionCreatedAt":"2026-01-30 02:55:48","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8653473","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8653473","identity":"rs-8653473","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}