HPLC-MS/MS Method for Measuring Cotinine and Trans-3'-Hydroxycotinine in Bronchoalveolar Lavage Fluid of Patients with e-Cigarette, or Vaping, Product Use-Associated Lung Injury (EVALI).

In 2019, nearly 3000 U.S. residents developed severe lung injury associated with recent use of e-cigarette or vaping products. The Centers for Disease Control and Prevention (CDC) responded to the outbreak, which was formally defined as e-cigarette, or vaping, product use-associated lung injury (EVALI). CDC Laboratory rapidly developed assays to analyze potentially harmful and addictive substances in bronchoalveolar lavage (BAL) fluid collected from EVALI case patients. This report describes the development and validation of a high-throughput isotope dilution HPLC-MS/MS method for measuring two nicotine biomarkers, cotinine (COT) and trans-3'-hydroxycotinine (HCT), in BAL fluid samples. COT and HCT are the major metabolites of nicotine, the addictive alkaloid presents in tobacco products. This method had good specificity and sensitivity. The limit of detection is 0.033 and 0.0165 ng/mL for COT and HCT, respectively, using only 200 µL of sample volume. The within-run and between-run precision was 2-10%. The overall accuracy, calculated from recovery in three different sample matrices spiked at three concentrations, was 94.8% and 93.6% for COT and HCT, respectively. This novel HPLC-MS/MS method was utilized to characterize recent tobacco exposure in EVALI case patients. This method is useful for characterizing tobacco exposure that may be related to acute and chronic lung injury. Keyword: e-cigarette, vaping, EVALI, nicotine, hydroxycotinine, cotinine.