{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,14]],"date-time":"2026-06-14T18:52:00Z","timestamp":1781463120847,"version":"3.54.1"},"reference-count":27,"publisher":"Oxford University Press (OUP)","issue":"2","license":[{"start":{"date-parts":[[2025,11,9]],"date-time":"2025-11-09T00:00:00Z","timestamp":1762646400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/academic.oup.com\/pages\/standard-publication-reuse-rights"}],"funder":[{"name":"National Science and Technology Council of the Republic of China","award":["NSTC 112-2320-B-037-027 - MY3; NSTC 114-2327-B-037-002-"],"award-info":[{"award-number":["NSTC 112-2320-B-037-027 - MY3; NSTC 114-2327-B-037-002-"]}]},{"name":"Kaohsiung Medical University Hospital Research Foundation","award":["KMUH113-3R69; KMUH112-2R81"],"award-info":[{"award-number":["KMUH113-3R69; KMUH112-2R81"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,4,22]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Ensuring analyte stability is essential for accurate forensic and clinical detection of sedative-type drugs. This study systematically evaluated the stability of 22 sedative-type drugs and metabolites in human urine under controlled conditions varying by pH (4.0, 7.0), temperature (25\u00b0C, 4\u00b0C, \u221220\u00b0C), and freeze\u2013thaw cycles (5 cycles), using a fully validated liquid chromatography\u2013tandem mass spectrometry (LC\u2013MS\/MS) method. While compounds such as midazolam, clobazam, and zolpidem remained highly stable, others\u2014including alprazolam, triazolam, and lorazepam\u2014exhibited notable degradation, particularly under acidic pH and elevated temperature. Flunitrazepam and clonazepam showed distinct degradation with the formation of 7-amino metabolites at neutral pH. Notably, this transformation occurred only in urine and not in phosphate-buffered saline, suggesting a urine-specific mechanism. These findings highlight the importance of compound-specific preservation strategies. In scenarios where analyte identity or sample pH cannot be verified promptly, immediate refrigeration or freezing (ideally at \u221220\u00b0C), along with minimizing freeze\u2013thaw cycles, is strongly recommended to preserve sample integrity and ensure reliable toxicological interpretation.<\/jats:p>","DOI":"10.1093\/jat\/bkaf100","type":"journal-article","created":{"date-parts":[[2025,11,7]],"date-time":"2025-11-07T13:03:20Z","timestamp":1762520600000},"source":"Crossref","is-referenced-by-count":1,"title":["Stability of 22 sedative-type drugs and metabolites in human urine under variable pH, temperature, and freeze\u2013thaw conditions"],"prefix":"10.1093","volume":"50","author":[{"given":"Feng-Shuo","family":"Yang","sequence":"first","affiliation":[{"name":"Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University , Kaohsiung 807,","place":["Taiwan"]},{"name":"Department of Medicinal and Applied Chemistry, Kaohsiung Medical University , Kaohsiung 807,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shu-Huei","family":"Jian","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University , Kaohsiung 807,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yi-Cheng","family":"Lee","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University , Kaohsiung 807,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yung-Sheng","family":"Lan","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University , Kaohsiung 807,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Li-Ping","family":"Tseng","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University , Kaohsiung 807,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yung-Hung","family":"Lee","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University , Kaohsiung 807,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0004-8337-3087","authenticated-orcid":false,"given":"Yi-Chen","family":"Chiu","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University , Kaohsiung 807,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0849-6149","authenticated-orcid":false,"given":"Yi-Ching","family":"Lin","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University , Kaohsiung 807,","place":["Taiwan"]},{"name":"Department of Laboratory Medicine, School of Medicine, College of Medicine, Kaohsiung Medical University , Kaohsiung 807,","place":["Taiwan"]},{"name":"Doctoral Degree Program of Toxicology, College of Pharmacy, Kaohsiung Medical University , Kaohsiung 807,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"286","published-online":{"date-parts":[[2025,11,9]]},"reference":[{"key":"2026042220274218500_bkaf100-B1","doi-asserted-by":"publisher","first-page":"594","DOI":"10.3390\/neurolint13040059","article-title":"Benzodiazepines: uses, dangers, and clinical considerations","volume":"13","author":"Edinoff","year":"2021","journal-title":"Neurol Int"},{"key":"2026042220274218500_bkaf100-B2","first-page":"214","article-title":"Benzodiazepine pharmacology and central nervous system-mediated effects","volume":"13","author":"Griffin","year":"2013","journal-title":"Ochsner J Summer"},{"key":"2026042220274218500_bkaf100-B3","doi-asserted-by":"crossref","first-page":"2170","DOI":"10.1001\/jama.1997.03550240060035","article-title":"Benzodiazepines and zolpidem for chronic insomnia: a meta-analysis of treatment efficacy","volume":"278","author":"Nowell","year":"1997","journal-title":"JAMA"},{"key":"2026042220274218500_bkaf100-B4","doi-asserted-by":"publisher","first-page":"155","DOI":"10.1007\/s13181-013-0292-0","article-title":"The clinical and forensic toxicology of Z-drugs","volume":"9","author":"Gunja","year":"2013","journal-title":"J Med Toxicol"},{"key":"2026042220274218500_bkaf100-B5","doi-asserted-by":"publisher","first-page":"1057","DOI":"10.1016\/s0731-7085(00)00393-9","article-title":"Stability of benzodiazepines in whole blood samples stored at varying temperatures","volume":"23","author":"El Mahjoub","year":"2000","journal-title":"J Pharm Biomed Anal"},{"key":"2026042220274218500_bkaf100-B6","doi-asserted-by":"crossref","first-page":"102","DOI":"10.1520\/JFS12243J","article-title":"Postmortem stability of benzodiazepines in blood and tissues","volume":"28","author":"Levine","year":"1983","journal-title":"J Forensic Sci"},{"key":"2026042220274218500_bkaf100-B7","doi-asserted-by":"publisher","first-page":"1505","DOI":"10.1007\/s00216-007-1267-2","article-title":"Stability of analytes in biosamples\u2014an important issue in clinical and forensic toxicology?","volume":"388","author":"Peters","year":"2007","journal-title":"Anal Bioanal Chem"},{"key":"2026042220274218500_bkaf100-B8","doi-asserted-by":"publisher","first-page":"75","DOI":"10.1016\/j.forsciint.2004.02.012","article-title":"Preanalytic aspects in postmortem toxicology","volume":"142","author":"Skopp","year":"2004","journal-title":"Forensic Sci Int"},{"key":"2026042220274218500_bkaf100-B9","doi-asserted-by":"publisher","first-page":"103","DOI":"10.1016\/j.forsciint.2012.02.005","article-title":"Degradation pathways of 4-methylmethcathinone in alkaline solution and stability of methcathinone analogs in various pH solutions","volume":"220","author":"Tsujikawa","year":"2012","journal-title":"Forensic Sci Int"},{"key":"2026042220274218500_bkaf100-B10","doi-asserted-by":"publisher","first-page":"486","DOI":"10.1093\/jat\/31.8.486","article-title":"Urine pH: the effects of time and temperature after collection","volume":"31","author":"Cook","year":"2007","journal-title":"J Anal Toxicol"},{"key":"2026042220274218500_bkaf100-B11","author":"Health UDo, Services H, Food","year":"2018"},{"key":"2026042220274218500_bkaf100-B12","doi-asserted-by":"publisher","first-page":"105","DOI":"10.1016\/S1355-0306(95)72641-7","article-title":"Stability of temazepam in blood","volume":"35","author":"al-Hadidi","year":"1995","journal-title":"Sci Justice"},{"key":"2026042220274218500_bkaf100-B13","doi-asserted-by":"publisher","first-page":"23","DOI":"10.1007\/978-1-4939-8823-5_3","article-title":"Analysis of benzodiazepines for drug-facilitated assaults and abuse settings (urine)","volume":"1872","author":"Drummer","year":"2019","journal-title":"Methods Mol Biol"},{"key":"2026042220274218500_bkaf100-B14","doi-asserted-by":"publisher","first-page":"101","DOI":"10.1016\/j.forsciint.2004.02.013","article-title":"Postmortem toxicology of drugs of abuse","volume":"142","author":"Drummer","year":"2004","journal-title":"Forensic Sci Int"},{"key":"2026042220274218500_bkaf100-B15","doi-asserted-by":"publisher","first-page":"101","DOI":"10.1016\/j.ijpharm.2004.12.013","article-title":"Accelerated aging: prediction of chemical stability of pharmaceuticals","volume":"293","author":"Waterman","year":"2005","journal-title":"Int J Pharm"},{"key":"2026042220274218500_bkaf100-B16","doi-asserted-by":"publisher","first-page":"4288","DOI":"10.1007\/s11356-018-2108-x","article-title":"Removal of pharmaceutically active compounds from synthetic and real aqueous mixtures and simultaneous disinfection by supported TiO(2)\/UV-A, H(2)O(2)\/UV-A, and TiO(2)\/H(2)O(2)\/UV-A processes","volume":"26","author":"Bosio","year":"2019","journal-title":"Environ Sci Pollut Res Int"},{"key":"2026042220274218500_bkaf100-B17","doi-asserted-by":"publisher","first-page":"1274","DOI":"10.1002\/jps.10141","article-title":"Isolation and structural elucidation of degradation products of alprazolam: photostability studies of alprazolam tablets","volume":"91","author":"Nudelman","year":"2002","journal-title":"J Pharm Sci"},{"key":"2026042220274218500_bkaf100-B18","doi-asserted-by":"publisher","first-page":"745","DOI":"10.1021\/js9504245","article-title":"2-(methylamino)-5-chlorobenzophenone imine: a novel product formed in base-catalyzed hydrolysis of diazepam","volume":"85","author":"Yang","year":"1996","journal-title":"J Pharm Sci"},{"key":"2026042220274218500_bkaf100-B19","doi-asserted-by":"publisher","first-page":"159","DOI":"10.1016\/j.forsciint.2011.04.005","article-title":"Stability of diazepam in blood samples at different storage conditions and in the presence of alcohol","volume":"215","author":"Atanasov","year":"2012","journal-title":"Forensic Sci Int"},{"key":"2026042220274218500_bkaf100-B20","doi-asserted-by":"publisher","first-page":"52","DOI":"10.1093\/jat\/bkr008","article-title":"Benzodiazepine stability in postmortem samples stored at different temperatures","volume":"36","author":"Melo","year":"2012","journal-title":"J Anal Toxicol"},{"key":"2026042220274218500_bkaf100-B21","doi-asserted-by":"publisher","first-page":"e569","DOI":"10.1097\/PCC.0000000000001687","article-title":"Evaluation of IV to enteral benzodiazepine conversion calculations in a pediatric intensive care setting","volume":"19","author":"Warrington","year":"2018","journal-title":"Pediatr Crit Care Med"},{"key":"2026042220274218500_bkaf100-B22","doi-asserted-by":"publisher","first-page":"109","DOI":"10.1080\/15287390490264767","article-title":"NADPH-cytochrome P-450 reductase is involved in flunitrazepam reductive metabolism in Hep G2 and Hep 3B cells","volume":"67","author":"Peng","year":"2004","journal-title":"J Toxicol Environ Health A"},{"key":"2026042220274218500_bkaf100-B23","doi-asserted-by":"publisher","DOI":"10.1093\/ijnp\/pyw083","article-title":"Optimization of clonazepam therapy adjusted to patient\u2019s CYP3A status and NAT2 genotype","volume":"19","author":"T\u00f3th","year":"2016","journal-title":"Int J Neuropsychopharmacol"},{"key":"2026042220274218500_bkaf100-B24","doi-asserted-by":"crossref","first-page":"85","DOI":"10.1016\/j.abb.2018.10.017","article-title":"Substrate selectivity of human aldehyde oxidase 1 in reduction of nitroaromatic drugs","volume":"659","author":"Ogiso","year":"2018","journal-title":"Arch Biochem Biophys"},{"key":"2026042220274218500_bkaf100-B25","doi-asserted-by":"publisher","first-page":"31","DOI":"10.1093\/jat\/30.1.31","article-title":"Determination of benzodiazepines in human urine using solid-phase extraction and high-performance liquid chromatography-electrospray ionization tandem mass spectrometry","volume":"30","author":"Hegstad","year":"2006","journal-title":"J Anal Toxicol"},{"key":"2026042220274218500_bkaf100-B26","doi-asserted-by":"publisher","first-page":"96","DOI":"10.1016\/j.bcp.2009.03.019","article-title":"Characterization of Escherichia coli nitroreductase NfsB in the metabolism of nitrobenzodiazepines","volume":"78","author":"LinWu","year":"2009","journal-title":"Biochem Pharmacol"},{"key":"2026042220274218500_bkaf100-B27","doi-asserted-by":"publisher","first-page":"196","DOI":"10.1093\/jat\/bkw113","article-title":"Stability of 21 cocaine, opioid and benzodiazepine drug analytes in spiked meconium at three temperatures","volume":"41","author":"Wu","year":"2017","journal-title":"J Anal Toxicol"}],"container-title":["Journal of Analytical Toxicology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/academic.oup.com\/jat\/advance-article-pdf\/doi\/10.1093\/jat\/bkaf100\/65249665\/bkaf100.pdf","content-type":"application\/pdf","content-version":"am","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/jat\/article-pdf\/50\/2\/bkaf100\/65249665\/bkaf100.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/jat\/article-pdf\/50\/2\/bkaf100\/65249665\/bkaf100.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,4,23]],"date-time":"2026-04-23T00:27:48Z","timestamp":1776904068000},"score":1,"resource":{"primary":{"URL":"https:\/\/academic.oup.com\/jat\/article\/doi\/10.1093\/jat\/bkaf100\/8317550"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,11,9]]},"references-count":27,"journal-issue":{"issue":"2","published-print":{"date-parts":[[2026,4,22]]}},"URL":"https:\/\/doi.org\/10.1093\/jat\/bkaf100","relation":{},"ISSN":["0146-4760","1945-2403"],"issn-type":[{"value":"0146-4760","type":"print"},{"value":"1945-2403","type":"electronic"}],"subject":[],"published-other":{"date-parts":[[2026,2]]},"published":{"date-parts":[[2025,11,9]]},"article-number":"bkaf100"}}