AOAC 2019 CBD Analysis by UPLC-PDA

Introduction

In 2018, AOAC published a method to quantify 10 cannabinoids (CBDs) with a range of 0.500-10.0 µg/mL in cannabis. However, there are more than 100 CBDs isolated from cannabis in addition to the above 10 CBDs. Furthermore, CBDs have been legally and widely used for more and more products. Thus, the AOAC method may not meet the diverse needs of various CBD products. To address various needs, Dyad Labs has developed UPLC-PDA and UPLC-MS/MS methods for 16 major CBDs. In this poster, the fast, comprehensive and accurate quantitative UPLC-PDA assay for 16 CBDs is presented in different matrix. The UPLC-MS/MS method is presented in P-T-018 poster.

Methodology

Sample Preparation and Extraction:

An appropriate sample was weighed in a 50 mL centrifuge tube. 5 mL of DI water was added to hydrate the sample. 35 mL of methanol was then added to the sample to precipitate protein and other types of interferences. The supernatant is diluted if needed, and then cleaned up with a filter. Filtered sample is mixed with an equal volume of DI water to match the LC initial solvent before analysis on the instrument.

UPLC-PDA Conditions

UPLC system: Waters Acquity UPLC System including quaternary solvent manager, sample manager FTN, column manager, and PDA detector.

PDA detector:  Scanning range of 200 nm to 400 nm. Quantitation uses 220 nm as the channel.

Results and Discussions

Stability

In the AOAC method, the standard solutions are expensive, but only have 3 days of stability. In order to better understand the stability of CBD, we comprehensively investigated the stability of all CBDs with different parameters, including light, heat, and pH. The study indicated that CBDs are sensitive to light, heat and pH. A neutral condition with light protection and low temperature was adopted during sample preparation.

In order to establish longer stability, we also compared different solvents including water, methanol, acetonitrile and methanol: water (70:30 v/v). The data indicated that CBDs are most stable in a methanol:water (70:30 v/v) mixture. During validation, we established up to 7 days stability for the standard solutions at 4.00-40.0 ug/mL using methanol: water (70:30 v/v) as solvent in fridge. The extracted sample is stable for 7 days when stored in the fridge.

Sensitivity and Linearity

The curve range of 4.00-40.0 ug/mL was successfully validated. The regression is quadratic with 1/x as the weighing factor. The correlation coefficient R2 is > 0.995. The representative chromatograms of LLOQ and ULOQ were in Figure 5.

Accuracy and Precision

The accuracy and precision were investigated using a CBD oil sample and post-spiking CBDs in blank botanical and protein matrices at the lower, medium, and high regions of the established calibration curve range.

Conclusion

This is the first validated method for a fast, comprehensive, and accurate quantification of the 16 major CBDs in CBD oil, botanical, and protein matrix using UPLC-PDA .