Analysis of cannabinoids and semi-synthetic cannabinoids in authentic breastmilk samples by liquid chromatography–tandem mass spectrometry

Marijuana (cannabis) is generally considered the most frequently misused substance during pregnancy. The prevalence in the use of either medical or non-medical marijuana for relief of pregnancy-related symptoms is increasing, as well as the use of cannabis-related products containing cannabidiol (CBD) and semi-synthetic cannabinoids (SSCs). Δ9-tetrahydrocannabinol (THC) and CBD are highly lipophilic substances and will readily pass into breastmilk upon ingestion. The solubility of THC and CBD in lipids poses significant analytical challenges in extracting and identifying these substances in breastmilk. The aim of this study was to develop a new and sensitive assay utilizing liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the detection of cannabinoids in breastmilk. The method was optimized to quantitate Δ8-THC, Δ9-THC, cannabigerol (CBG), CBD, and cannabidiolic acid (CBDA) and validated with the guidance of the American Academy of Forensic Sciences Standards Board (ASB) Standard 036. The assay was then used to analyze breastmilk samples (N = 57) collected postpartum from female patients enrolled in a study assessing use behaviors of medical marijuana, non-medical marijuana, and CBD. All analytes passed validation criteria. Calibration curves for all analytes ranged 0.5-400 ng/mL, with the LOD and LLOQ of the method set at the lowest calibrator concentration. Δ9-THC was quantitated in 19 samples (33.3%) with a concentration range of 0.5-291 ng/mL. Δ8-THC was detected in one sample (1.8%) at 0.8 ng/mL, while CBD was observed in 3 samples at a concentration <LLOQ, and quantitated in only one sample (1.8%) also at a concentration of 0.8 ng/mL. CBG was detected in 7 samples (12.2%) with a concentration range of 0.6-12.9 ng/mL, and at a concentration <LLOQ in 12 samples. This study presents a sensitive method for the analysis of cannabinoids in breastmilk to support the follow-up assessments of marijuana and CBD use during pregnancy and postpartum.