Development of a Liquid Chromatography-Tandem Mass Spectrometry Method for the Simultaneous Determination of Four Cannabinoids in Umbilical Cord Tissue

Abstract

In utero exposure to marijuana may cause various short- and long-term health problems, such as stillbirth, low birth weight and decreased cognitive function. Detection of in utero marijuana exposure with a relatively new specimen type, umbilical cord tissue, can be used to plan treatment and guide social management. In this study, a liquid chromatography-tandem mass spectrometry (LC-MS-MS) assay was developed for the simultaneous identification of four cannabinoids in umbilical cord tissue, including ∆9-tetrahydrocannabinol (THC), 11-nor-9-carboxy-∆9--THC (THC-COOH), 11-hydroxy-∆9-THC (11-OH-THC) and cannabinol (CBN). Within- and between-run imprecision, accuracy, linearity, sensitivity, carryover, recovery, matrix effects and specificity were evaluated using drug-free umbilical cord tissue spiked with non-deuterated and deuterated standards. Calibration curves were reproducible and linear (r > 0.995) for all four analytes in the range of 0.2 ng/g lower limit of quantitation (LLOQ) and 30 ng/g upper limit of quantitation (ULOQ). Total imprecisions (% coefficient of variation) were 7.8% (THC), 13.3% (THC-COOH), 11.8% (11-OH-THC) and 10.6% (CBN) at low QC (n = 15, 0.25 ng/g), and were 7.2% (THC), 10.0% (THC-COOH), 9.5% (11-OH-THC) and 5.8% (CBN) at high QC (n = 15, 4 ng/g), respectively. No interfering substances were identified, and no carryover was observed. The average accuracies (N = 25) were 94-95%. The average recoveries observed for THC, THC-COOH, 11-OH-THC and CBN were 74, 82, 58 and 86%, respectively. By analyzing authentic clinical specimens that had been previously tested for cannabinoids by enzyme-linked immunoassay, positive and negative result agreements were 100 and 53.8%. In summary, the presented method can be used for the assessment of in utero exposure to four common cannabinoids.

Figure 1.

(A) Representative LC separation and extracted ion chromatograms of (B) THC, (C) THC-COOH, (D) 11-OH-THC and (E) CBN at 4 ng/g.

Figure 2.

Concentration distribution of (A) THC, (B) THC-COOH, (C) 11-OH-THC and (D) CBN in authentic samples. Neg, negative tests by ELISA; P, positive tests by ELISA; HP, high positive tests by ELISA. ‘---’ indicates cutoffs used in the LC–MS-MS method.

Figure 3.

The cannabinoid patterns in cord detected by LC–MS-MS. ‘---‘ indicates median.