Headspace SPME GC/MS Analysis of Terpenes in Cannabis

By: Katherine K. Stenerson, Principal Scientist, Reporter US Volume 34.4

A rapid method to identify cannabis terpenes for forensic and organoleptic applications

Cannabis sativa (cannabis or marijuana) contains over 100 different terpenes and terpenoids, including mono, sesqui, di and tri, as well as other miscellaneous compounds of terpenoid orgin.1 Terpenes give the plant distinct organoleptic properties and produce characteristic aromas when the buds are heated or vaporized.2 Although the terpene profile does not necessarily indicate geographic origin of a cannabis sample, it can be used in forensic applications to determine the common source of different samples.3 In addition, different cannabis strains have been developed which have distinct aromas and flavors, a result of the differing amounts of specific terpenes present.4

Experimental

Dried cannabis sample was obtained courtesy of Dr. Hari H. Singh, Program Director at the Chemistry and Physiological Systems Research Branch of the United States National Institute on Drug Abuse at the National Institute of Health. Terpenes were isolated using headspace solid phase microextraction (SPME) followed by chromatographic separation on an Equity®-1 capillary GC column. Peak identifications were assigned using MS spectral matching against reference spectra in the Wiley and NIST libraries. Confirmatory identification was done based on retention index, which was calculated for the compounds identified in each sample using an n-alkane standard analyzed under the same GC conditions. This data was compared with published values and peak identifications were assigned.6,7 Final analytical conditions appear in Figure 1.

Figure 1. Headspace SPME-GC/MS Analysis of Dried Cannabis Sativa

 

Conditions
sample/matrix: 0.5 g dried, ground cannabis sativa
SPME fiber: 50/30 μm DVB/CAR/PDMS (Product No. 57298-U)
sample equilibration: 30 min, 40 °C
extraction: 20 min, headspace, 40 °C
desorption process: 3 min, 270 °C
fiber post bake: 3 min, 270 °C
column: Equity-1, 60 m × 0.25 mm I.D., 0.25 μm (Product No. 28047-U)
oven: 60 °C (2 min), 5 °C/min to 275 °C (5 min)
inj. temp.: 270 °C
detector: MSD
MSD interface: 300 °C
scan range: full scan, m/z 50-500
carrier gas: helium, 1 mL/min constant flow
liner: 0.75 mm ID SPME


Table 1.
Terpenes in Dried Cannabis Identified by MS Spectral Library Match and Retention Index

Peak
No.
RT
(min)

Name
Retention Index
(calculated)
Retention Index
(literature)
1 8.57 Hexanal
2 10.05 Hexene-1-ol
3 10.89 2-Heptanone
4 12.56 α-Thujene 928 938
5 12.86 α-Pinene + unknown 939 942
6 13.27 Camphene 953 954
7 13.69 6-Methyl-5-hepten-2-one 966 968
8 14.09 β-Pinene 979 981
9
14.27 β-Myrcene 984 986
10 15.09 Δ-3-Carene 1010 1015
11 15.20
α-Terpinene
1014
1012
12 15.29 Cymene 1018 1020
13 15.60 d-Limonene 1028 1030
14 16.42 γ-Terpinene 1056 1057
15 16.60 trans-Sabinene hydrate 1062 1078
16 16.72 cis-Linalool oxide 1066 1068
17 17.43 Linalool 1087 1092
18 18.04 d-Fenchyl alcohol 1107 1110
19 18.82 trans-Pinocarveol 1135 1134
20 19.59 Borneol L 1161 1164
21 19.81 1,8-Methandien-4-ol 1168 1173
22 19.81 p-Cymen-8-ol 1168 1172
23 19.92 Terpinene-4-ol 1172 1185
24 20.22 α-Terpineol 1181 1185
25 24.20 Piperitenone 1322 1320
26 24.76 Piperitenone oxide 1344 1352
27 25.85 α-Ylangene 1384 1373
28 25.97 α-Copaene 1388 1398
29 26.76 γ-Caryophyllene 1419 1403
30 27.01 α-Santalene 1429 1428
31 27.16 Caryophyllene 1435 1428
32 27.36 trans-α-Bergamotene + unknown 1443 1443
33 27.49
α-Guaiene
1448 1441
34 27.56 trans-β-Farnesene 1451 1446
35 27.98 Humulene 1467 1465
36 28.17 Alloaromadendrene 1475 1478
37 28.25 α-Curcumene 1478 1479
38 28.75 β-Selinene 1497 1487
39 28.97 α-Selinene 1507 1497
40 28.97 β-Bisobolene 1507 1506
41 29.13 α-Bulnesene 1514 1513
42 30.12 Selina-3,7(11)-diene 1556 1542
43 30.94 Caryophyllene oxide 1590 1595
44 31.50 Humulene oxide 1614 1599
45 32.48 Caryophylla-3,8(13)-dien-5-ol A 1658 1656

Results and Discussion

The terpenes identified in the cannabis sample (Figure 1) are indicated in Table 1. The profile was similar to those found previously in the analysis of dried cannabis.3,5 Early eluting peaks generally were monoterpenes and monoterpenoids. The later eluting peaks consisted of sequiterpenes and caryophyllene oxide, which is a sequiterpenoid. The most abundant terpene was caryophyllene. The predominance of this compound could be due to the specific strain of cannabis tested, and/or the nature of the sample tested, which was dried. Previous studies have shown the level of this compound to increase significantly relative to other terepenes and terpenoids with drying.3 Consequently, the levels of the more volatile monoterpenes and terpenoids would be expected to be less, and this was observed to some degree. Among the monoterpenes and terpenoids, the most abundant were α-pinene and limonene.

Materials

     

 References

  1. Turner, C.E.; Elsohly, E.G.; Boeren, E.G. Constituents of Cannabis Sativa L., XVIII. A Review of the Natural Constituents. J. Nat. Prod., 1980, 43, 169-234.
  2. Kemsley, J. Cannabis Safety. CEN, 2015, April 20, 27-28.
  3. Hood, L.V.S.; Barry, G.T. Headspace Volatiles of Marihuana and Hashish: Gas Chromatographic Analysis of Samples of Different Geographic Origin. J. Chrom. A., 1978, 166, 499-506.
  4. Rahn, B. Terpenes: The Flavors of Cannabis Aromatherapy. 2/12/2014. www.leafly.com/news, accessed September 18, 2015.
  5. Marchini, M.; Charvoz, C.; Dujourdy, L.; Baldovini, N. Multidimensional Analysis of Cannabis Volatile Constituents: Identification of 5,5-dimethyl- 1-vinylbicyclo[1.1.1]hexane as a Volatile Marker of Hashish, the Resin of Cannabis Sativa L. J. Chrom A., 2014, 1370, 200-215.
  6. Jennings, W.; Shibamoto, T. Qualitative Analysis of Flavor and Fragrance Volatiles by Glass Capillary Gas Chromatography, Academic Press, 1980.
  7. NIST Chemistry WebBook. www.webbook.nist.gov/chemistry.

 

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