Validity of Triply Labelled Water Analysis for Energy Expenditure Measurements in Mice

Validity of Triply Labelled Water Analysis for Energy Expenditure Measurements in Mice | 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 Validity of Triply Labelled Water Analysis for Energy Expenditure Measurements in Mice Xing Wang, Dehuang Kong, Gertjan van Dijk, Harro Meijer This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-764218/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 9 You are reading this latest preprint version Abstract The Doubly Labelled Water (DLW) method is widely used to determine energy expenditure. In this work, we demonstrate the addition of the third stable isotope, 17 O, to turn it into Triply Labelled Water (TLW), using the three isotopes measurement of optical spectrometry. We performed TLW ( 2 H, 18 O and 17 O) measurements for the analysis of the CO 2 production (r CO2 ) of mice on different diets for the first time. Triply highly enriched water was injected into mice, and the isotope enrichments of the distilled blood samples of one initial and two finals were measured by an Off-Axis Integrated Cavity Output Spectroscopy instrument. We evaluated the impact of different calculation protocols and the values of evaporative water loss fraction. We found that the dilution space and turnover rates of 17 O and 18 O were equal for the same mice group, and that values of r CO2 calculated based on 18 O- 2 H, or on 17 O- 2 H agreed very well. This increases the reliability and redundancy of the measurements and it lowers the uncertainty in the calculated r CO2 to 3% when taking the average of two DLW methods. However, the TLW method overestimated the r CO2 compared to the indirect calorimetry measurements that we also performed, much more for the mice on a high-fat diet than for low-fat. We hypothesize an extra loss or exchange mechanism with a high fractionation for 2 H to explain this difference. Endocrinology & Metabolism Doubly Labelled Water (DLW) energy expenditure Triply Labelled Water (TLW) optical spectrometry Full Text Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Editorial decision: Major revision 18 Jan, 2022 Reviews received at journal 18 Jan, 2022 Reviews received at journal 14 Sep, 2021 Reviewers agreed at journal 09 Sep, 2021 Reviewers invited by journal 13 Aug, 2021 Editor assigned by journal 13 Aug, 2021 Editor invited by journal 13 Aug, 2021 Submission checks completed at journal 13 Aug, 2021 First submitted to journal 30 Jul, 2021 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. 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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-764218","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":45694908,"identity":"22c1c197-6f44-4534-ac0c-92237628f45f","order_by":0,"name":"Xing Wang","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAx0lEQVRIiWNgGAWjYDACZhiDvQFMyZCghecAhCLBOokEIrUYHOc9/IKxzS5PPvLtM8kvDHcIa5Fs5kuzYGxLLja8nW4mLcPwjLAWfmYeMwPGbcyJG2ensUlLMBwmrIUNoqU+cePMY0RqAdpi/IBx2+HE+RJsbJIfiNEi2cxjxpD473jiBp40ZmsGAyK0GJw/Y/zhw5nqxPntxxhv/qg4LEdQC8g74BgxOACMVh4DYjQAFX4AkfINDAyMP4jTMQpGwSgYBSMMAACKODNifppODgAAAABJRU5ErkJggg==","orcid":"","institution":"University of Groningen","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Xing","middleName":"","lastName":"Wang","suffix":""},{"id":45694909,"identity":"3ea6e7f6-8852-4934-85d9-902088b3205d","order_by":1,"name":"Dehuang Kong","email":"","orcid":"","institution":"University of Groningen","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Dehuang","middleName":"","lastName":"Kong","suffix":""},{"id":45694910,"identity":"2585c53f-2b40-466b-971c-9573b3c86a5a","order_by":2,"name":"Gertjan van Dijk","email":"","orcid":"","institution":"University of Groningen","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Gertjan","middleName":"van","lastName":"Dijk","suffix":""},{"id":45694911,"identity":"4d096ef6-a978-4d4c-81d8-5d260d4c0fee","order_by":3,"name":"Harro Meijer","email":"","orcid":"","institution":"University of Groningen","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Harro","middleName":"","lastName":"Meijer","suffix":""}],"badges":[],"createdAt":"2021-07-30 10:14:08","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-764218/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-764218/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":13668300,"identity":"99f91e09-a64d-4673-a401-2210676bc47c","added_by":"auto","created_at":"2021-09-17 10:56:33","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":437464,"visible":true,"origin":"","legend":"","description":"","filename":"TLWpaperXINGWANGnewsubmit2.pdf","url":"https://assets-eu.researchsquare.com/files/rs-764218/v1_covered.pdf"},{"id":12515372,"identity":"89a42ae8-1400-48e1-9368-49b4658ba79f","added_by":"auto","created_at":"2021-08-17 20:03:08","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":433915,"visible":true,"origin":"","legend":"","description":"","filename":"TLWpaperXINGWANGnewsubmit2.pdf","url":"https://assets-eu.researchsquare.com/files/rs-764218/v1_covered.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"\u003cp\u003eValidity of Triply Labelled Water Analysis for Energy Expenditure Measurements in Mice\u003c/p\u003e","fulltext":[{"header":"Full Text","content":"This preprint is available for \u003ca href='/article/rs-764218/latest.pdf' target='_blank'\u003edownload as a PDF\u003c/a\u003e."}],"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":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Doubly Labelled Water (DLW), energy expenditure, Triply Labelled Water (TLW), optical spectrometry","lastPublishedDoi":"10.21203/rs.3.rs-764218/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-764218/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThe Doubly Labelled Water (DLW) method is widely used to determine energy expenditure. In this work, we demonstrate the addition of the third stable isotope, \u003csup\u003e17\u003c/sup\u003eO, to turn it into Triply Labelled Water (TLW), using the three isotopes measurement of optical spectrometry. We performed TLW (\u003csup\u003e2\u003c/sup\u003eH, \u003csup\u003e18\u003c/sup\u003eO and\u003csup\u003e17\u003c/sup\u003eO) measurements for the analysis of the CO\u003csub\u003e2\u003c/sub\u003e production (r\u003csub\u003eCO2\u003c/sub\u003e) of mice on different diets for the first time. Triply highly enriched water was injected into mice, and the isotope enrichments of the distilled blood samples of one initial and two finals were measured by an Off-Axis Integrated Cavity Output Spectroscopy instrument. We evaluated the impact of different calculation protocols and the values of evaporative water loss fraction. We found that the dilution space and turnover rates of \u003csup\u003e17\u003c/sup\u003eO and \u003csup\u003e18\u003c/sup\u003eO were equal for the same mice group, and that values of r\u003csub\u003eCO2\u003c/sub\u003e calculated based on \u003csup\u003e18\u003c/sup\u003eO-\u003csup\u003e2\u003c/sup\u003eH, or on \u003csup\u003e17\u003c/sup\u003eO-\u003csup\u003e2\u003c/sup\u003eH agreed very well. This increases the reliability and redundancy of the measurements and it lowers the uncertainty in the calculated r\u003csub\u003eCO2\u003c/sub\u003e to 3% when taking the average of two DLW methods. However, the TLW method overestimated the r\u003csub\u003eCO2\u003c/sub\u003e compared to the indirect calorimetry measurements that we also performed, much more for the mice on a high-fat diet than for low-fat. We hypothesize an extra loss or exchange mechanism with a high fractionation for \u003csup\u003e2\u003c/sup\u003eH to explain this difference.\u003c/p\u003e","manuscriptTitle":"Validity of Triply Labelled Water Analysis for Energy Expenditure Measurements in Mice","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2021-08-17 20:03:02","doi":"10.21203/rs.3.rs-764218/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Major revision","date":"2022-01-19T03:51:54+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2022-01-18T17:11:37+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2021-09-14T22:34:03+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"03fb5691-b4a6-42f4-bc68-11232523a119","date":"2021-09-09T15:19:51+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2021-08-14T00:40:00+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2021-08-14T00:37:13+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2021-08-13T12:37:44+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2021-08-13T11:52:41+00:00","index":"","fulltext":""},{"type":"submitted","content":"Scientific Reports","date":"2021-07-30T10:05:37+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"8cd449cc-d828-4db6-bb5c-89592e48e196","owner":[],"postedDate":"August 17th, 2021","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[{"id":6507522,"name":"Endocrinology \u0026 Metabolism"}],"tags":[],"updatedAt":"2022-03-25T10:14:11+00:00","versionOfRecord":[],"versionCreatedAt":"2021-08-17 20:03:02","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-764218","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-764218","identity":"rs-764218","version":["v1"]},"buildId":"cBFmMYwuxLRRLfASyISRj","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}