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In this study, we describe a targeted and validated method that combines benzoyl chloride derivatization with reversed-phase ultrahigh-performance liquid chromatography tandem mass spectrometry (RP-UHPLC\/MS\/MS) for the quantitative analysis of the human serum lipidome. In total, 450 lipid species from 19 lipid subclasses were identified based on a combination of multiple reaction monitoring transitions, retention dependencies, dilution series, and derivatization tags. The developed methodology increases the sensitivity for most investigated lipid classes in comparison to conventional methods, but the highest improvement was observed for monoacylglycerols, diacylglycerols, sphingoid bases, and free sterols. The method\u2019s accuracy was confirmed using NIST SRM 1950, as the determined concentrations were in agreement with the consensus values from ring trials. Lipidomic profiling of clinical samples revealed a significant dysregulation of lipid metabolism in pancreatic cancer patients compared to healthy controls. Key findings included the upregulation of most monoacylglycerols and sphingosine, and a pronounced downregulation of sphingolipids with very long saturated\n                    <jats:italic>N<\/jats:italic>\n                    -acyl chains and phospholipids containing fatty acyl compositions 18:2 and 20:4. This targeted approach is consistent with the trends previously seen with other methods and also provides new findings and more detailed structural insights into metabolic alterations in pancreatic cancer.\n                  <\/jats:p>","DOI":"10.1007\/s00216-025-06151-0","type":"journal-article","created":{"date-parts":[[2025,10,6]],"date-time":"2025-10-06T06:20:50Z","timestamp":1759731650000},"page":"733-745","update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["Benzoyl chloride derivatization improves selectivity and sensitivity of lipidomic quantitation in human serum of pancreatic cancer patients using RP-UHPLC\/MS\/MS"],"prefix":"10.1007","volume":"418","author":[{"given":"Ond\u0159ej","family":"Peterka","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zuzana","family":"L\u00e1sko","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Robert","family":"Jir\u00e1sko","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Petra","family":"Peroutkov\u00e1","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anna","family":"Taylor","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Beatrice","family":"Moheln\u00edkov\u00e1-Ducho\u0148ov\u00e1","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Irena","family":"Kozub\u00edkov\u00e1","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Martin","family":"Love\u010dek","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bohuslav","family":"Melichar","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3978-1249","authenticated-orcid":false,"given":"Michal","family":"Hol\u010dapek","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2025,10,6]]},"reference":[{"key":"6151_CR1","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkad896","author":"MJ Conroy","year":"2024","unstructured":"Conroy MJ, Andrews RM, Andrews S, Cockayne L, Dennis EA, Fahy E, Gaud C, Griffiths WJ, Jukes G, Kolchin M, Mendivelso K, Lopez-Clavijo AF, Ready C, Subramaniam S, O\u2019Donnell VB. 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Concordant inter-laboratory derived concentrations of ceramides in human plasma reference materials via authentic standards. Nat Commun. 2024. https:\/\/doi.org\/10.1038\/s41467-024-52087-x.","journal-title":"Nat Commun"},{"key":"6151_CR40","doi-asserted-by":"publisher","DOI":"10.1038\/s42255-019-0094-z","author":"G Liebisch","year":"2019","unstructured":"Liebisch G, Ahrends R, Arita M, Arita M, Bowden JA, Ejsing CS, Griffiths WJ, Hol\u010dapek M, K\u00f6feler H, Mitchell TW, Wenk MR, Ekroos K, Initiative LS. Lipidomics needs more standardization. Nat Metab. 2019. https:\/\/doi.org\/10.1038\/s42255-019-0094-z.","journal-title":"Nat Metab"},{"key":"6151_CR41","doi-asserted-by":"publisher","DOI":"10.1016\/j.bbamem.2006.05.027","author":"T Kolter","year":"2006","unstructured":"Kolter T, Sandhoff K. Sphingolipid metabolism diseases. Biochimica et Biophysica Acta (BBA). 2006. https:\/\/doi.org\/10.1016\/j.bbamem.2006.05.027.","journal-title":"Biochimica et Biophysica Acta (BBA)"}],"container-title":["Analytical and Bioanalytical Chemistry"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1007\/s00216-025-06151-0.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/article\/10.1007\/s00216-025-06151-0","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1007\/s00216-025-06151-0.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,1,8]],"date-time":"2026-01-08T13:36:15Z","timestamp":1767879375000},"score":1,"resource":{"primary":{"URL":"https:\/\/link.springer.com\/10.1007\/s00216-025-06151-0"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,10,6]]},"references-count":41,"journal-issue":{"issue":"2","published-print":{"date-parts":[[2026,1]]}},"alternative-id":["6151"],"URL":"https:\/\/doi.org\/10.1007\/s00216-025-06151-0","relation":{},"ISSN":["1618-2642","1618-2650"],"issn-type":[{"value":"1618-2642","type":"print"},{"value":"1618-2650","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,10,6]]},"assertion":[{"value":"29 July 2025","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"19 September 2025","order":2,"name":"revised","label":"Revised","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"22 September 2025","order":3,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"6 October 2025","order":4,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Declarations"}},{"value":"This study was carried out in accordance with the Declaration of Helsinki. Informed consent was obtained from all volunteers, and the ethical committee approved the collection of blood samples.","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Ethical approval"}},{"value":"M.H. is a member of ABC\u2019s International Advisory Board but was not involved in the peer review of this paper. All other authors declare no conflict of interest.","order":3,"name":"Ethics","group":{"name":"EthicsHeading","label":"Conflict of interest"}}]}}