Long-Term Stability of 13 Fentanyl Analogs in Blood.J Anal Toxicol. 2021 May 19 [Online ahead of print]JA
Fentanyl analogs continue to play a major role in proliferating the opioid epidemic in the United States. With high rates of overdose deaths, forensic laboratories experience backlogs, which may lead to false negative results due to drug instability. To address this issue, a quantitative method was validated for fentanyl analogs (3-methylfentanyl, 4-ANPP, 4-fluoro-isobutyrylfentanyl (4-FIBF), acetylfentanyl, acrylfentanyl, butyrylfentanyl, carfentanil, cyclopropylfentanyl, fentanyl, furanylfentanyl, methoxyacetylfentanyl, p-fluorofentanyl, and valerylfentanyl) in blood using liquid chromatography-quadrupole-time-of-flight mass spectrometry (LC-QTOF-MS) and used to assess long-term stability under various temperature conditions (-20°C, 4°C, ~25°C, and 35°C) for 9 months. Authentic specimens were also analyzed 6 months apart for applicability to postmortem blood. Method validation resulted in calibration ranges of 1-100 ng/mL and limits of detection (LOD) of 0.5 ng/mL. Precision and bias were acceptable (within ±7.2%CV and ±15.2%, respectively). Matrix effects exhibited ion enhancement for all analytes, except carfentanil and 4-ANPP in low quality control (>25%). For long-term stability, fentanyl analogs (except acrylfentanyl) remained stable under room temperature and refrigerated conditions at low and high concentrations (81.3-112.5% target) for 9 months. While most fentanyl analogs remained stable frozen, degradation was observed after 2 weeks (4 freeze/thaw cycles). At elevated temperatures, most analytes were stable for 1 week (74.2-112.6% target). Acrylfentanyl was unstable after 24h under elevated (70% loss) and room temperatures (53-60% loss), 48-72h refrigerated (28-40% loss), and 4 weeks frozen (22% loss). In authentic bloods (n=7), initial furanylfentanyl (FuF) and 4-ANPP concentrations were 1.1-3.6 and 1.4-6.4 ng/mL, respectively. Percent loss of FuF and 4-ANPP over 6 months were 16.3-37.4% and 0.2-26.8%, respectively. Samples suspected to contain fentanyl analogs are recommended to be stored refrigerated or frozen with limited freeze/thaw cycles. Due to instability, in the event of an acrylfentanyl overdose, samples should be analyzed immediately or stored frozen with analysis within 1 month.