Published: Journal of Analytical Toxicology, Volume 23, Number 3, May/June 1999, pp.222-224.

D⁹-Tetrahydrocannabivarin (D⁹-THCV) as a Marker for the Ingestion of Cannabis versus Marinol ®

To the Editor:

D⁹-THC (Figure 1, 1), the active ingredient in marijuana, has been approved by the FDA as an anti-emetic for cancer patients receiving chemotherapy and as an appetite stimulant for AIDS patients suffering from the wasting syndrome.
The procedures used in workplace drug testing and criminal justice system drug testing rely on the identification of the major D⁹-THC metabolite (11-nor-D⁹-THC-9-COOH, Figure 1, 2) in biological fluids, mainly in urine, to identify marijuana users.

Although the prescription drug Marinol is approved only for the two indications listed previously, off-label prescription could make it available to others, and the drug could be claimed as the reason for a positive test for marijuana. Therefore, the question is, could one determine if the D⁹-THC metabolite found in the urine of a given individual is the result of Marinol use only? That is, could one determine if marijuana (or other cannabis-related products) was used, with or without Marinol? This letter offers a scientific basis to the answer for this the question based on the identification of the metabolite of the C3 homologue of D⁹-THC in the urine of cannabis users.

The bulk active material used in the manufacture of Marinol is produced by a synthetic process that leads only to D⁹-THC with its C5 side chain. Other side products of the synthetic process such as D8-THC and CBN also contain a C5 side chain.

On the other hand, although D⁹-THC is the major cannabinoid in drug-type cannabis, other cannabinoids do exist at different levels (1). One such cannabinoid closely related to D⁹-THC that is found in most drug-type samples is the C3 homologue of D⁹-THC known as D⁹-tetrahydrocannabivarin (D⁹-THCV, Figure 1, 3). Being a homologue, this cannabinoid is expected to behave as D⁹-THC in every respect with differences related only to the side chain. Gill (3) described the isolation and characterization of D⁹-THCV and reported its mass spectral fragmentation to be parallel to that of D⁹-THC. The mass spectrum of D⁹-THCV from a cannabis sample was compared with that of D⁹-THC. In addition, to prove that these cannabinoid homologues would have similar fragmentation and ion ratios that differ only in masses, the mass spectrum of the C4 homologue of D⁹-THC (known also to be component of the cannabis plant [4]) was recorded and compared with the spectra of the C3 and C5 homologues. Figure 2 shows the mass spectra of the three homologues, which are essentially identical except for the differences in ion masses.

Now that the similarity in the mass spectral fragmentation of the parent cannbinoids is established, it is believed that D⁹-THCV would be metabolized in much the same way as D⁹-THC, resulting in 11-nor-D⁹-THCV-9-COOH (Figure 1, 4) that could be detected in the urine of subjects ingesting cannabis products containing D⁹-THCV (5). Again, being a lower homologue, the retention time of its metabolite derviative would be lower than that of the C5 homologue, but the ion ratios of the C3 homologue should be similar to the corresponding ion ratios of the C5 homologue. For example, ions at m/z 313, 357, and 372 are monitored for the methyl derivative of 11-nor-D⁹-THC-9-COOH with ion ratios of m/z 357/313 and 372/313 monitored for identification. The corresponding ions for the same derivative of the C3 homologue would be at m/z 285, 329, and 344, with the ion ratios 329/285 and 344/285 being in the same range as 357/313 and 372/313, respectively.

Figure 3 shows the ion chromatograms from the analysis of a urine specimen collected from a cannabis user. Table I shows the ions monitored, the expected ion ratios, and the ion ratio found. The peak at the shorter retention time relative to 11-nor-D⁹-THC-9-COOH (Figure 1, 2) with the ion masses 28 units less than 11-nor-D⁹-THC-9-COOH was determined to be that of 11-nor-D⁹-THCV-9-COOH (Figure 1, 4).

This approach was used to analyze a urine specimen, the donor of which had indicated on his consent form that he was using Marinol prescribed to him by his physician for back pain. The specimen showed high levels of both 11-nor-D⁹-THC-9-COOH and 11-nor-D⁹-THCV-9-COOH. Upon confronting the individual with the test results, he admitted use of marijuana.

Subsequently, we have synthesized the carboxy metabolite of THCV (Figure 1, 4) and established that the mass fragmentation of its derivative is identical to that of the C5 homologue as previously anticipated. Figure 4 shows the mass spectra of the methyl derivative of both metabolites. In addition, incubation of THCV with human hepatocytes resulted in the formation of the acid metabolite found to be identical to the synthetic standard.

It is therefore concluded that the detection of the D⁹-THCV metabolite in the urine of an individual would be a positive indication that marijuana (or a related product) had been ingested sometime (within days) prior to collection of the urine and that the presence of 11-nor-D⁹-THCV-9-COOH is a marker for such ingestion with or without Marinol. This is particularly true because analysis of Marinol showed no THCV, and analysis of plasma samples from subjects ingesting Marinol showed no D⁹-THCV-9-COOH. Details of the studies outlined in this letter will be the subject of upcoming communications.

Mahmoud A. ElSohly^1,2, Shixia Feng¹, Timothy P. Murphy¹, Samir A. Ross², Alison Nimrod², Zlatko Mehmedic², and Neil Fortner³
1ElSohly Laboratories, Inc., 5 Industrial Park Drive, Oxford, Mississippi 38655; 2National Center for the Development of Natural Products, University of Mississippi, University, Mississippi 38677; and 3PharmChem Laboratories, 1505 O’Brien Drive, Menlo Park, California 94025

References

1. R. Brenneisen and M. ElSohly. Chromatographic and spectroscopic profiles of cannabis of different origins: part 1.
   J. Forensic Sci. 33: 1385–1404 (1988).
2. C.E. Turner, K. Hadley, and P.S. Fetterman. Constituents of Cannabis sativa L., VI: propyl homologs in samples of known geographical origin. J. Pharm. Sci. 62: 1739–1741 (1973).
3. E.W. Gill. Propyl homologue of tetrahydrocannabinol: its isolation from cannabis, properties and synthesis. J. Chem. Soc. (C) 579–582 (1971).
4. D.J. Harvey. Characterization of the butyl homologues of delta-1-tetrahydrocannabinol, cannbinol, and cannabidiol in samples of cannabis by combined gas chromatography and mass spectrometry. J. Pharm. Pharmacol. 28: 280 (1976).
5. R. Foltz. Analysis of urine specimens for cannabinoids to distinguish the ingestion of Marinol® from smoked THC. Proceedings of the California Association of Toxicologists, November 1996, pp 28–31.

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