Development and validation of a high-throughput LC-MS/MS bioanalytical method for the simultaneous quantification of...

The removal of hemp from the definition of marijuana in the 2018 Agricultural Improvement Act has increased the number of cannabidiol-containing products available to consumers. Consequently, consumer use has also increased. Increased product availability and use drives the need for sensitive and specific analytical assays to measure the cannabidiol (CBD) and metabolites in patients, to establish dose-effect relationships and to gain knowledge of their pharmacokinetics. Here we describe the development and validation of a rapid high-throughput LC-MS/MS bioanalytical method for the quantification of cannabidiol and primary metabolites in human plasma to support an FDA-sponsored clinical study (NCT06192589). Sample preparation a single step protein precipitation followed by filtration through 96-well Phree™ Phospholipid removal plates. The method was validated over ranges of CBD: 1.95-500.00 ng mL^-1; 7-hydroxy-cannabidiol (7-OH-CBD): 3.91-1000.00 ng mL^-1; 7-carboxy-cannabidiol (7-COOH-CBD): 31.25-8000.00 ng mL^-1. There was no cross-analyte interference, injection carryover, or matrix effect observed with this method. Analyte recoveries were consistent across three QC levels ranging from 83.90 to 90.85 %. Inter-day Accuracy across four QC level of all three analytes ranged from 93.87 to 107.31 % while precision ranged from 1.03 to 14.33 %. These results and other outlined in this manuscript met acceptance criteria as outline the current M10 Guidance for Bioanalytical Method Validation and Study Sample Analysis.