{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,7]],"date-time":"2026-10-07T15:00:12Z","timestamp":1791385212234,"version":"4.3.3"},"reference-count":41,"publisher":"American Chemical Society (ACS)","issue":"10","license":[{"start":{"date-parts":[[2026,9,16]],"date-time":"2026-09-16T00:00:00Z","timestamp":1789516800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000054","name":"National Cancer Institute","doi-asserted-by":"publisher","award":["R00CA273517"],"award-info":[{"award-number":["R00CA273517"]}],"id":[{"id":"10.13039\/100000054","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100001368","name":"V Foundation for Cancer Research","doi-asserted-by":"publisher","award":["V2024-009"],"award-info":[{"award-number":["V2024-009"]}],"id":[{"id":"10.13039\/100001368","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000057","name":"National Institute of General Medical Sciences","doi-asserted-by":"publisher","award":["R35GM156596"],"award-info":[{"award-number":["R35GM156596"]}],"id":[{"id":"10.13039\/100000057","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,10,7]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Coenzyme A is an essential cofactor synthesized from pantothenate, which carries activated acyl groups in cellular metabolic reactions. Coenzyme A participates in catabolism of carbohydrates, fats, and amino acids; biosynthesis of fatty acids, cholesterol, and heme; and protein modification, including acetylation and 4-phosphopantetheinylation. Despite CoA\u2019s critical functions, the regulation of CoA levels and the rate of CoA synthesis in different cell types and disease states are not well understood. Acyl-CoA species are challenging to measure due to instability, poor ionization, and the wide range of biochemical properties conferred by different acyl chain lengths; moreover, most current methods cannot measure CoA isotopic labeling, required to quantify the CoA synthesis rate or to measure concentration using isotope-labeled internal standards. Here, we describe a method to quantify the concentration and isotopic labeling of total CoA, defined as the sum of CoASH plus all acyl-CoA species. Acyl-CoA species are hydrolyzed using sodium hydroxide to remove acyl chains; then, CoA is derivatized with N-ethylmaleimide (NEM). Following protein precipitation and solid-phase extraction, samples are analyzed by liquid chromatography-mass spectrometry. This method is linear in a wide range that captures mouse tissue CoA levels, with accuracy within 15% error and precision below 15% relative standard deviation for standards and tissue samples. We applied this method to measure total CoA concentration in five mouse tissues and the total CoA synthesis rate in mouse liver via infusion of 13C\u201315N-pantothenate. Overall, this method offers a tractable approach to measure total CoA concentration and synthesis rate in health and disease.<\/jats:p>","DOI":"10.1021\/jasms.6c00206","type":"journal-article","created":{"date-parts":[[2026,9,16]],"date-time":"2026-09-16T17:02:00Z","timestamp":1789578120000},"page":"2453-2462","source":"Crossref","is-referenced-by-count":0,"title":["A Liquid Chromatography-Mass\nSpectrometry Method to\nQuantify Total Coenzyme A Concentration and Isotopic Labeling"],"prefix":"10.1021","volume":"37","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-1284-6914","authenticated-orcid":true,"given":"Amelia L.","family":"Taylor","sequence":"first","affiliation":[{"name":"University of Pennsylvania , , 3600 Civic Center Blvd Room 16-111 , , ,","place":["Philadelphia, Pennsylvania, United States, 19104"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5643-8747","authenticated-orcid":true,"given":"Nathaniel W.","family":"Snyder","sequence":"additional","affiliation":[{"name":"Lewis Katz School of Medicine at Temple University , , 3500 N Broad St. , , ,","place":["Philadelphia, Pennsylvania, United States, 19104"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Caroline R.","family":"Bartman","sequence":"additional","affiliation":[{"name":"University of Pennsylvania , , 3600 Civic Center Blvd Room 16-111 , , ,","place":["Philadelphia, Pennsylvania, United States, 19104"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2026,9,16]]},"reference":[{"issue":"8","key":"2026100710070684100_cit1","doi-asserted-by":"publisher","first-page":"1031","DOI":"10.3390\/brainsci11081031","article-title":"Coenzyme a Biochemistry:\nFrom Neurodevelopment to Neurodegeneration","volume":"11","author":"Mignani","year":"2021","journal-title":"Brain Sci."},{"issue":"7921","key":"2026100710070684100_cit2","doi-asserted-by":"publisher","first-page":"192","DOI":"10.1038\/s41586-022-04984-8","article-title":"PI3K Drives the de\nNovo Synthesis of Coenzyme A from Vitamin B5","volume":"608","author":"Dibble","year":"2022","journal-title":"Nature"},{"issue":"6","key":"2026100710070684100_cit3","doi-asserted-by":"publisher","first-page":"1008","DOI":"10.1038\/s42255-024-01059-y","article-title":"Coenzyme A Biosynthesis:\nMechanisms\nof Regulation, Function and Disease","volume":"6","author":"Barritt","year":"2024","journal-title":"Nat. Metab."},{"issue":"4","key":"2026100710070684100_cit4","doi-asserted-by":"publisher","first-page":"567","DOI":"10.1016\/j.molcel.2018.06.039","article-title":"ACP Acylation Is an Acetyl-CoA-Dependent\nModification Required for Electron Transport Chain Assembly","volume":"71","author":"Van Vranken","year":"2018","journal-title":"Mol. Cell"},{"issue":"1","key":"2026100710070684100_cit5","doi-asserted-by":"publisher","first-page":"396","DOI":"10.1038\/s41392-022-01245-y","article-title":"Protein acylation:\nmechanisms, biological functions\nand therapeutic targets","volume":"7","author":"Shang","year":"2022","journal-title":"Signal Transduction\nTargeted Ther."},{"issue":"14","key":"2026100710070684100_cit6","doi-asserted-by":"publisher","first-page":"2489","DOI":"10.1042\/BCJ20170129","article-title":"Protein CoAlation: a redox-regulated\nprotein modification by coenzyme A in mammalian cells","volume":"474","author":"Tsuchiya","year":"2017","journal-title":"Biochem. J."},{"issue":"6","key":"2026100710070684100_cit7","doi-asserted-by":"publisher","first-page":"5951","DOI":"10.3390\/ijms24065951","article-title":"Inherited Disorders\nof Coenzyme A Biosynthesis: Models, Mechanisms, and Treatments","volume":"24","author":"Cavestro","year":"2023","journal-title":"Int. J. Mol. Sci."},{"issue":"4","key":"2026100710070684100_cit8","doi-asserted-by":"publisher","first-page":"1043","DOI":"10.1042\/BST20140094","article-title":"Role of CoA and acetyl-CoA in regulating\ncardiac fatty acid and glucose oxidation","volume":"42","author":"Abo Alrob","year":"2014","journal-title":"Biochem.\nSoc. Trans."},{"issue":"2","key":"2026100710070684100_cit9","doi-asserted-by":"publisher","first-page":"334","DOI":"10.1158\/0008-5472.CAN-19-1161","article-title":"CoA Synthase\n(COASY) Mediates Radiation Resistance via PI3K Signaling in Rectal\nCancer","volume":"80","author":"Ferrandon","year":"2020","journal-title":"Cancer Res."},{"issue":"11","key":"2026100710070684100_cit10","doi-asserted-by":"publisher","first-page":"1870","DOI":"10.1038\/s42255-023-00915-7","article-title":"Vitamin B5\nSupports MYC Oncogenic Metabolism and Tumor Progression in Breast\nCancer","volume":"5","author":"Kreuzaler","year":"2023","journal-title":"Nat. Metab."},{"issue":"23","key":"2026100710070684100_cit11","doi-asserted-by":"publisher","first-page":"12902","DOI":"10.1021\/acs.analchem.7b03659","article-title":"Comprehensive\nAnalysis of Short-, Medium-, and Long-Chain Acyl-Coenzyme A by Online\nTwo-Dimensional Liquid Chromatography\/Mass Spectrometry","volume":"89","author":"Wang","year":"2017","journal-title":"Anal. Chem."},{"issue":"4","key":"2026100710070684100_cit12","doi-asserted-by":"publisher","first-page":"823","DOI":"10.1016\/j.cmet.2017.03.006","article-title":"A Class of Reactive\nAcyl-CoA Species Reveals the Non-enzymatic Origins of Protein Acylation","volume":"25","author":"Wagner","year":"2017","journal-title":"Cell Metab."},{"issue":"1","key":"2026100710070684100_cit13","doi-asserted-by":"publisher","first-page":"158","DOI":"10.1016\/j.bbrc.2010.10.009","article-title":"Coenzyme A\nand Its Thioester Pools in Fasted and Fed Rat Tissues","volume":"402","author":"Tokutake","year":"2010","journal-title":"Biochem. Biophys. Res. Commun."},{"issue":"2","key":"2026100710070684100_cit14","doi-asserted-by":"publisher","first-page":"192","DOI":"10.3109\/10253890.2015.1137558","article-title":"Decreased\nHepatic Contents of Coenzyme A Molecular Species in Mice after Subchronic\nMild Social Defeat Stress","volume":"19","author":"Kubota","year":"2016","journal-title":"Stress"},{"issue":"3","key":"2026100710070684100_cit15","doi-asserted-by":"publisher","first-page":"550","DOI":"10.1042\/bj0930550","article-title":"Variations in Tissue\nContents of Coenzyme A Thio Esters\nand Possible Metabolic Implications","volume":"93","author":"Tubbs","year":"1964","journal-title":"Biochem.\nJ."},{"issue":"5","key":"2026100710070684100_cit16","doi-asserted-by":"publisher","first-page":"634","DOI":"10.1007\/BF00841821","article-title":"Changes in the Content and Structure\nof the Liver Coenzyme a Reserves in Diabetic (Db\/Db) Mice during Nicotinamide\nTherapy","volume":"103","author":"Obrosova","year":"1987","journal-title":"Bull. Exp. Biol. Med."},{"issue":"2","key":"2026100710070684100_cit17","doi-asserted-by":"publisher","first-page":"325","DOI":"10.1016\/0003-2697(83)90305-6","article-title":"Radioisotopic\nAssay of Picomolar Amounts of Coenzyme A","volume":"134","author":"Rabier","year":"1983","journal-title":"Anal.\nBiochem."},{"issue":"3","key":"2026100710070684100_cit18","doi-asserted-by":"publisher","first-page":"291","DOI":"10.1016\/j.chembiol.2007.01.013","article-title":"Chemical Knockout\nof Pantothenate\nKinase Reveals the Metabolic and Genetic Program Responsible for Hepatic\nCoenzyme A Homeostasis","volume":"14","author":"Zhang","year":"2007","journal-title":"Chem. Biol."},{"issue":"1","key":"2026100710070684100_cit19","doi-asserted-by":"publisher","first-page":"94","DOI":"10.1016\/0003-2697(88)90015-2","article-title":"A Radioisotopic Assay\nof Picomolar Concentrations of Coenzyme A in\nLiver Tissue","volume":"168","author":"Knights","year":"1988","journal-title":"Anal. Biochem."},{"key":"2026100710070684100_cit20","doi-asserted-by":"publisher","first-page":"151","DOI":"10.3791\/60182","article-title":"Quantification\nof Coenzyme A in Cells and Tissues","author":"Frank","year":"2019","journal-title":"J. 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Genet. Metab."},{"issue":"1","key":"2026100710070684100_cit24","doi-asserted-by":"publisher","first-page":"102745","DOI":"10.1016\/j.jbc.2022.102745","article-title":"NUDT7 Regulates Total\nHepatic CoA Levels and the Composition of the Intestinal Bile Acid\nPool in Male Mice Fed a Western Diet","volume":"299","author":"Vickers","year":"2023","journal-title":"J. Biol.\nChem."},{"issue":"4","key":"2026100710070684100_cit25","doi-asserted-by":"publisher","first-page":"350","DOI":"10.1016\/j.ymgme.2017.02.001","article-title":"Excess Coenzyme A Reduces\nSkeletal Muscle Performance and Strength in Mice Overexpressing Human\nPANK2","volume":"120","author":"Corbin","year":"2017","journal-title":"Mol. Genet. Metab."},{"key":"2026100710070684100_cit26","first-page":"1","volume-title":"Guidance for Industry:\nbioanalytical Method Validation, No. FDA-2013-D-1020","author":"US Food and Drug Administration.","year":"2018"},{"key":"2026100710070684100_cit27","doi-asserted-by":"publisher","first-page":"59","DOI":"10.1016\/j.ab.2014.12.014","article-title":"Production\nof Stable Isotope-Labeled Acyl-Coenzyme\nA Thioesters by Yeast Stable Isotope Labeling by Essential Nutrients\nin Cell Culture","volume":"474","author":"Snyder","year":"2015","journal-title":"Anal. Biochem."},{"issue":"1","key":"2026100710070684100_cit28","doi-asserted-by":"publisher","first-page":"14.39.1","DOI":"10.1002\/cpph.2","article-title":"Genetically\nEngineered Mouse Models of Pancreatic Cancer: The KPC Model (LSL-KrasG12D\/+;LSL-Trp53R172H\/+;Pdx-1-Cre),\nIts Variants and Their Application in Immuno-Oncology Drug Discovery","volume":"73","author":"Lee","year":"2016","journal-title":"Curr. Protoc. Pharmacol."},{"issue":"4","key":"2026100710070684100_cit29","doi-asserted-by":"publisher","first-page":"1447","DOI":"10.1021\/acs.jproteome.9b00640","article-title":"Alzheimer\u2019s Disease Metabolomics Consortium.\nSkyline for Small Molecules: A Unifying Software Package for Quantitative\nMetabolomics","volume":"19","author":"Adams","year":"2020","journal-title":"J. Proteome Res."},{"key":"2026100710070684100_cit30","doi-asserted-by":"publisher","first-page":"301","DOI":"10.1007\/978-1-4939-9236-2_19","article-title":"El-MAVEN: A Fast, Robust,\nand User-Friendly Mass Spectrometry Data Processing Engine for Metabolomics","volume":"1978","author":"Agrawal","year":"2019","journal-title":"Methods Mol. Biol."},{"key":"2026100710070684100_cit31","first-page":"No. 1","volume-title":"Liquid\nChromatography-Mass Spectrometry Methods; Approved Guidelines (C62-A)","author":"Clinical and Laboratory Standards Institute.","year":"2014"},{"issue":"1","key":"2026100710070684100_cit32","doi-asserted-by":"publisher","first-page":"100","DOI":"10.3390\/molecules15010100","article-title":"Determination\nof coenzyme A (CoASH) in the presence\nof different thiols by using flow-injection with a UV\/Vis spectrophotometric\ndetector and potentiometric determination of CoASH using an iodide\nISE","volume":"15","author":"Giljanovic\u0301\u0327","year":"2010","journal-title":"Molecules"},{"issue":"10","key":"2026100710070684100_cit33","doi-asserted-by":"publisher","first-page":"1403","DOI":"10.1038\/s41374-021-00631-4","article-title":"AccuCor2:\nisotope natural abundance\ncorrection for dual-isotope tracer experiments","volume":"101","author":"Wang","year":"2021","journal-title":"Lab. Invest."},{"key":"2026100710070684100_cit34","doi-asserted-by":"publisher","first-page":"202508","DOI":"10.70401\/fos.2025.0010","article-title":"Best practices for\ncysteine analysis","volume":"2","author":"Choudhury","year":"2025","journal-title":"Ferroptosis Oxid Stress"},{"issue":"1","key":"2026100710070684100_cit35","doi-asserted-by":"publisher","first-page":"75","DOI":"10.1021\/tx034127o","article-title":"In Vitro Reactivity\nof Carboxylic Acid-CoA Thioesters with Glutathione","volume":"17","author":"Sidenius","year":"2004","journal-title":"Chem. Res. Toxicol."},{"issue":"3","key":"2026100710070684100_cit36","doi-asserted-by":"publisher","first-page":"346","DOI":"10.1203\/00006450-200203000-00014","article-title":"CoASH and CoASSG\nLevels in Lungs of Hyperoxic Rats\nas Potential Biomarkers of Intramitochondrial Oxidant Stresses","volume":"51","author":"O\u2019donovan","year":"2002","journal-title":"Pediatr. Res."},{"issue":"2","key":"2026100710070684100_cit37","doi-asserted-by":"publisher","first-page":"275","DOI":"10.1016\/j.ab.2008.02.022","article-title":"Novel Isolation\nProcedure for Short-, Medium-, and Long-Chain Acyl-Coenzyme A Esters\nfrom Tissue","volume":"376","author":"Minkler","year":"2008","journal-title":"Anal. Biochem."},{"issue":"10","key":"2026100710070684100_cit38","doi-asserted-by":"publisher","first-page":"2520","DOI":"10.1021\/ja01106a522","article-title":"The Preparation of\nS-Succinyl Coenzyme A","volume":"75","author":"Simon","year":"1953","journal-title":"J. Am. Chem. Soc."},{"issue":"9","key":"2026100710070684100_cit39","doi-asserted-by":"publisher","first-page":"1388","DOI":"10.3390\/molecules22091388","article-title":"Determination\nof Coenzyme A and Acetyl-Coenzyme A in Biological Samples Using HPLC\nwith UV Detection","volume":"22","author":"Shurubor","year":"2017","journal-title":"Molecules"},{"issue":"19","key":"2026100710070684100_cit41","doi-asserted-by":"publisher","first-page":"14957","DOI":"10.3390\/ijms241914957","article-title":"Multi-Method\nQuantification of Acetyl-Coenzyme A and Further Acyl-Coenzyme A Species\nin Normal and Ischemic Rat Liver","volume":"24","author":"Tokarska-Schlattner","year":"2023","journal-title":"Int. J. Mol.\nSci."},{"issue":"7","key":"2026100710070684100_cit42","doi-asserted-by":"publisher","first-page":"896","DOI":"10.1038\/s42255-021-00419-2","article-title":"Quantitative flux analysis in mammals","volume":"3","author":"Bartman","year":"2021","journal-title":"Nat. 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