Towards Development of a Rapid Approach for Differentiating Marijuana from Hemp—Insights from High-level Density Functional Theory Calculations

Direct analysis in real time – high-resolution mass spectrometry (DART-HRMS) has been shown to enable rapid analysis of C. sativa plant material to reveal chemical fingerprint signatures that readily show a characteristic peak at nominal m/z 315, indicative of the presence of THC. However, in the absence of sample pretreatment such as derivatization, this analysis does not enable differentiation of THC from other isomeric cannabinoids such as cannabinol (CBD), which occurs in high concentrations in hemp. Interestingly, an m/z value consistent with the presence of the protonated dimers [2M + H+] of THC and CBD was observed in the DART-HRMS spectra of purified THC and CBD reference standards (i.e., nominal m/z 629). To investigate the structures of these dimer complexes, high-level density functional theory (DFT) calculations were performed for various protonated dimer combinations (e.g., CBD••CBD; THC••THC; and CBD••THC). The computations revealed that the THC••THC protonated homodimer complex is more stable than the others by ~1 – 3 kcal/mole. However, an investigation of the effect of temperature on the population of dimer complexes revealed that with increasing temperature, the populations of the THC••THC and CBD••THC dimers decrease and become negligible at ≥200 K, while that of the CBD••CBD dimer increases. The results show that at the temperature at which the DART-HRMS analysis of C. sativa is performed, the protonated dimer peak at m/z 629 corresponds to CBD••CBD when CBD levels are high (i.e., in hemp). On the other hand, in marijuana samples (high THC content with negligible CBD content), no m/z 629 peak would be apparent in the DART-HRMS spectra. Subsequent DART-HRMS analysis of marijuana and hemp samples, so designated by the ≤0.3% THC (hemp) and >0.3% THC (marijuana) cutoffs, revealed this prediction to be true. The results show that m/z 629 could be used to readily and rapidly differentiate hemp and marijuana using DART-HRMS analysis.