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The fully automated sample preparation procedure is based on salting\u2010out assisted liquid\u2013liquid extraction (SALLE) combined with high\u2010efficiency LC\u2010MS\/MS methods for both screening and confirmation of 37 drugs and incorporates novel features enabling direct injection of acetonitrile extracts into an innovative chromatographic system. The methods' drug panel includes opioids, benzodiazepines, benzodiazepine\u2010like drugs, cannabinoids, and stimulants. A full method validation was performed using OF\/buffer from Greiner Bio\u2010ONE International and Quantisal saliva collection devices. The validation included assessments of linearity, sensitivity, precision, accuracy, extraction recovery, matrix effects, process efficiency, stability, and carryover. All compounds demonstrated linearity across the concentration range 1\u201325\u2009ng\/mL, with\n                    <jats:italic>R<\/jats:italic>\n                    <jats:sup>2<\/jats:sup>\n                    \u2009\u2265\u20090.99. Both methods' limit of detection ranged between 0.001 and 0.03\u2009ng\/mL, and the limit of quantification ranged between 0.02 and 0.09\u2009ng\/mL. Precision was \u2264\u200914.8% for screening and \u2264\u20098.5% for the confirmation method. Accuracy was \u00b1\u200913.6% for screening and \u00b1\u20098.7% (except at 0.5 and 1\u2009ng\/mL, where it was \u00b1\u200925.3% and \u00b1\u200917.6%, respectively) for the confirmation method. Extraction recoveries ranged from 40.0% to 95.1%, except for hydromorphone (27.4%) and morphine (34.4%). Although matrix effects were observed for a large number of compounds to varying degrees, they were largely compensated for by the use of deuterium\u2010 and\n                    <jats:sup>13<\/jats:sup>\n                    C\u2010labeled internal standards (IS). IS\u2010corrected overall process efficiency ranged from 100.7% to 119.1% with precision (CV%)\u2009\u2264\u200910.8% for all compounds. Spiked calibrators and QC samples in OF were stable in autosampler for up to 72\u2009h and in the freezer for 3\u2009days. Methanol working solutions were stable for 6\u2009months. No significant carryover was observed. The methods have been successfully implemented in the routine analysis of approximately &gt;\u20091000 samples per month since March 2024.\n                  <\/jats:p>","DOI":"10.1002\/jms.5132","type":"journal-article","created":{"date-parts":[[2025,4,8]],"date-time":"2025-04-08T07:27:57Z","timestamp":1744097277000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":9,"title":["Automated and Fully Validated High\u2010Throughput LC\u2010MS\/MS Assay for Analyzing Multiple Drugs of Abuse in Oral Fluids Using Novel Features in Sample Preparation and Chromatographic Conditions"],"prefix":"10.1002","volume":"60","author":[{"given":"Joakim","family":"Tan","sequence":"first","affiliation":[{"name":"Department of Clinical Pharmacology, Laboratory of Drugs of Abuse Analysis Karolinska University Hospital  Stockholm Sweden"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alexia","family":"Rylski","sequence":"additional","affiliation":[{"name":"Department of Clinical Pharmacology, Laboratory of Drugs of Abuse Analysis Karolinska University Hospital  Stockholm Sweden"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anders","family":"Bergqvist","sequence":"additional","affiliation":[{"name":"Department of Clinical Pharmacology, Laboratory of Drugs of Abuse Analysis Karolinska University Hospital  Stockholm Sweden"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0007-6713-7852","authenticated-orcid":false,"given":"Niclas\u00a0Nikolai","family":"Stephanson","sequence":"additional","affiliation":[{"name":"Department of Clinical Pharmacology, Laboratory of Drugs of Abuse Analysis Karolinska University Hospital  Stockholm Sweden"},{"name":"Karolinska Institute  Stockholm Sweden"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2025,4,8]]},"reference":[{"key":"e_1_2_7_2_1","doi-asserted-by":"publisher","DOI":"10.1016\/s0378\u20104347(97)00514\u20108"},{"key":"e_1_2_7_3_1","doi-asserted-by":"publisher","DOI":"10.1007\/978-1-61779-934-1_13"},{"key":"e_1_2_7_4_1","doi-asserted-by":"publisher","DOI":"10.1373\/clinchem.2015.248146"},{"key":"e_1_2_7_5_1","doi-asserted-by":"publisher","DOI":"10.1385\/FSMP:2:2:75"},{"key":"e_1_2_7_6_1","doi-asserted-by":"publisher","DOI":"10.1002\/dta.1683"},{"key":"e_1_2_7_7_1","doi-asserted-by":"crossref","unstructured":"\u201cThe Challenges of LC\u2010MS\/MS Analysis of Opiates and Opioids in Urine \u201dhttps:\/\/doi.org\/10.4155\/bio.13.244.","DOI":"10.4155\/bio.13.244"},{"key":"e_1_2_7_8_1","doi-asserted-by":"crossref","unstructured":"\u201cAdvanced Urine Toxicology Testing \u201d accessed 2024\u201007\u201001 https:\/\/doi.org\/10.1080\/10550887.2010.509277?needAccess=true.","DOI":"10.1080\/10550887.2010.509277"},{"key":"e_1_2_7_9_1","doi-asserted-by":"publisher","DOI":"10.1002\/dta.1486"},{"key":"e_1_2_7_10_1","unstructured":"\u201cUNODC World Drug Report 2024: Harms of World Drug Problem Continue to Mount Amid Expansions in Drug Use and Markets \u201d United Nations: Office on Drugs and Crime.www.unodc.org\/unodc\/en\/press\/releases\/2024\/June\/unodc\u2010world\u2010drug\u2010report\u20102024_\u2010harms\u2010of\u2010world\u2010drug\u2010problem\u2010continue\u2010to\u2010mount\u2010amid\u2010expansions\u2010in\u2010drug\u2010use\u2010and\u2010markets.html. 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