New Methods for the Comprehensive Analysis of Bioactive Compounds in Cannabis sativa L. (hemp)
Abstract
:1. Introduction
2. Results and Discussion
2.1. Extraction of Non-Psychoactive Cannabinoids and Flavonoids from Hemp Inflorescences
2.2. HPLC Methods for the Analysis of Non-Psychoactive Cannabinoids and Flavonoids in Hemp Extracts
2.3. Identification of Non-Psychoactive Cannabinoids and Flavonoids in Hemp Extracts
2.4. Validation Data of the HPLC Methods
2.5. Quantitative Analysis of Non-Psychoactive Cannabinoids in Hemp by HPLC-UV/DAD
2.6. Quantitative Analysis of Flavonoids and Related Compounds in Hemp by HPLC-UV/DAD
2.7. Extraction of Volatile Compounds from Hemp by HS-SPME
2.8. GC Methods for the Analysis of Volatile Compounds from Hemp
3. Materials and Methods
3.1. Chemicals and Solvents
3.2. Hemp Plant Material
3.3. Sample Preparation
3.3.1. Extraction of Non-Psychoactive Cannabinoids
3.3.2. Extraction of Flavonoids and Related Compounds
3.3.3. Extraction of Volatile Compounds
3.4. HPLC-UV/DAD Conditions
3.4.1. HPLC-UV/DAD Analysis of Non-Psychoactive Cannabinoids
3.4.2. HPLC-UV/DAD Analysis of Flavonoids and Related Compounds
3.5. HPLC-ESI-MS and MS2 Conditions
3.6. HPLC-UV/DAD Method Validation
3.7. GC-FID Analysis of Volatile Compounds
3.8. GC-MS Analysis of Volatile Compounds
3.9. Qualitative and Semi-Quantitative Analysis of Volatile Compounds
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Sample Availability: Samples of fibre-type hemp extracts are available from the authors. |
Peak Number | Compound | tR (min) | UV λmax (nm) | MS (m/z) a | MS2 (m/z) a | MS (m/z) b | MS2 (m/z) b |
---|---|---|---|---|---|---|---|
1 | CBDA | 13.8 | 225,269,307 | 359, 341 * | 219 (100), 261 (55), 285 (33), 232 (22) | 357 | 339 (100), 313 (9) |
2 | CBGA | 15.7 | 223,268,305 | 361, 343 * | 219 (100), 261 (49), 233 (19) | 359 | 341 (100), 359 (6) |
3 | CBG | 17.1 | 207,231sh,273 | 317 | 207 (100), 233 (40), 280 (10) | 315 | 271 (100), 204 (80), 247 (51), 297 (49) |
4 | CBD | 17.9 | 209,232sh,276 | 315 | 259 (100), 233 (26), 221 (24), 207 (24) | 313 | 245 (100), 210 (6) |
5 | CFL-B | 14.5 | 215,273,342 | 370 | 313 (100) | - | - |
6 | Canniprene | 16.9 | 204,280 | 343, 287 * | 255 (100), 227 (98), 269 (11) | 341 | 326 (100), 269 (84), 283 (43), 257 (12) |
7 | CFL-A | 21.1 | 214,274,342 | 437 | 313 (100) | - | - |
Compound | C1 | C2 | C3 | C4 | C5 | C6 |
---|---|---|---|---|---|---|
CBDA | 22.6 ± 2.5 | 4.1 ± 1.7 | 36.4 ± 1.0 | 4.1 ± 0.1 | 3.7 ± 0.2 | 17.6 ± 0.4 |
CBGA | 0.4 b | 0.2 ± 0.1 | 0.6 b | 0.2 b | <LOQ | 0.1 b |
CBG | <LOQ | 0.3 ± 0.1 | <LOQ | 0.3 b | <LOQ | <LOQ |
CBD | 1.9 ± 0.2 | 5.9 ± 0.4 | 6.3 ± 0.3 | 5.9 ± 0.1 | 8.6 ± 0.5 | 6.4 ± 0.4 |
Compound | C1 | C2 | C3 | C4 | C5 | C6 |
---|---|---|---|---|---|---|
CFL-A | 163.4 ± 2.4 | 109.4 ± 5.7 | 318.9 ± 5.8 | 59.9 ± 3.4 | 89.5 ± 6.7 | 140.6 ± 12.5 |
CFL-B | 100.9 ± 1.7 | 21.0 ± 2.6 | 412.2 ± 4.0 | 25.3 ± 0.2 | 61.4 ± 5.7 | 75.9 ± 6.6 |
Canniprene | <LOD | <LOD | <LOD | <LOD | <LOD | <LOD |
Peak Number | Compound | LRI Calc. | LRI Lit. c | % Relative Peak Area | |||||
---|---|---|---|---|---|---|---|---|---|
C1 | C2 | C3 | C4 | C5 | C6 | ||||
1 | α-Thujene | 932 | 911 | 3.7 ± 1.1 | 1.4 ± 1.0 | 0.2 b | 0.2 b | 0.5 ± 0.1 | - |
2 | α-Pinene | 939 | 932 | 20.1 ± 0.5 | 40.1 ± 2.3 | 26.7 b | 9.6 ± 0.4 | 15.9 ± 2.1 | 10.3 b |
3 | Camphene | 952 | 957 | 0.3 b | - | 0.5 b | 0.3 b | 0.3 b | 0.2 b |
4 | β-Pinene | 980 | 978 | 5.7 b | 9.3 ± 0.1 | 6.9 ± 0.1 | 2.9 ± 0.2 | 4.2 ± 0.3 | 3.7 b |
5 | β-Myrcene | 1000 | 992 | 28.7 ± 0.1 | 12.9 ± 0.1 | 36.9 ± 0.2 | 32.1 ± 3.3 | 6.9 ± 0.5 | 34.1 ± 1.1 |
6 | α-Phellandrene | 1009 | 1007 | 0.1 b | - | 0.4 b | 0.5 b | - | 0.3 b |
7 | Δ3-Carene | 1014 | 1010 | 1.3 b | 1.4 b | 0.5 b | 0.6 b | - | 0.3 b |
8 | α-Terpinene | 1020 | 1018 | - | - | 0.3 b | 0.4 b | - | 0.3 ± 0.1 |
9 | o-Cymene | 1022 | 1021 | 0.1 b | - | 0.1 b | 0.1 b | 1.3 ± 0.1 | - |
10 | p-Cymene | 1028 | 1026 | 0.2 b | - | 0.1 b | 0.5 ± 0.1 | 2.5 ± 0.1 | 0.8 ± 0.1 |
11 | Limonene | 1032 | 1035 | 2.4 b | 2.3 ± 0.1 | 1.6 b | 11.3 ± 0.5 | 3.5 ± 0.2 | 11.5 ± 0.3 |
12 | cis-Ocimene | 1042 | 1040 | 0.5 b | 0.7 b | 0.2 b | 0.6 ± 0.1 | 0.4 ± 0.1 | - |
13 | trans-Ocimene | 1053 | 1097 | 7.1 ± 0.3 | 0.6 b | 6.1 ± 0.2 | 5.2 ± 0.1 | 0.4 ± 0.1 | 0.5 b |
14 | γ-Terpinene | 1062 | 1062 | 0.2 b | - | 0.3 b | 0.4 b | 0.3 b | 0.5 b |
15 | α-Terpinolene | 1093 | 1088 | 3.9 ± 0.1 | - | 7.9 ± 0.1 | 12.0 ± 0.8 | 1.4 ± 0.1 | 4.9 ± 0.1 |
16 | Linalool | 1104 | 1104 | 0.6 b | - | 0.1 b | 0.3 b | 0.8 ± 0.1 | 0.4 b |
17 | 6-Camphenol | 1106 | 1110 | 0.3 b | - | - | 0.1 b | 1.2 ± 0.1 | - |
18 | neo-Alloocimene | 1133 | 1143 | 0.4 b | - | 0.1 b | 0.4 b | 1.1 ± 0.2 | - |
19 | Isopinocarveol | 1145 | 1139 | - | 0.6 b | - | - | 0.6 ± 0.1 | - |
20 | Terpinen-4-ol | 1181 | 1177 | 0.5 b | - | 0.2 b | 0.3 b | 3.3 ± 0.2 | 0.6 b |
21 | p-Cymen-8-ol | 1183 | 1181 | - | - | 0.1 b | - | 2.2 ± 0.2 | - |
22 | α-Terpineol | 1193 | 1195 | 0.1 b | - | - | 0.1 b | 0.7 ± 0.1 | 0.4 b |
23 | Eugenol | 1371 | 1373 | - | - | - | 0.3 ± 0.1 | - | - |
24 | α-Copaene | 1375 | 1376 | - | - | - | 0.8 ± 0.2 | - | 0.3 ± 0.1 |
25 | Geranyl acetate | 1379 | 1386 | - | - | - | 0.3 ± 0.1 | - | - |
26 | Ylangene | 1389 | 1406 | - | - | 0.1 b | - | - | - |
27 | Z,Z-α-Farnesene | 1415 | 1462 | 0.2 b | 0.7 b | - | 0.1 b | 1.2 b | 0.4 b |
28 | α-Santalene | 1423 | 1420 | - | - | 0.1 b | 0.1 b | 0.3 b | - |
29 | β-Caryopyllene | 1430 | 1428 | 13.4 ± 0.1 | 18.1 ± 0.5 | 5.2 ± 0.1 | 8.1 ± 2.0 | 22.6 ± 1.0 | 21.8 ± 0.7 |
30 | Z,E-α-Farnesene | 1437 | 1486 | - | - | - | 0.1 b | 2.2 b | - |
31 | α-Guaiene | 1444 | 1439 | 0.2 b | 0.5 b | 0.4 b | 0.9 ± 0.3 | - | - |
32 | Aromadendrene | 1446 | 1449 | 0.4 ± 0.1 | - | - | 0.3 ± 0.1 | - | - |
33 | β-Farnesene | 1460 | 1454 | 0.1 b | - | - | 1.8 ± 0.5 | 0.3 b | - |
34 | α-Humulene | 1464 | 1455 | 2.8 b | 4.6 ± 0.3 | 1.3 b | 2.1 ± 0.5 | 8.7 ± 0.6 | 5.6 ± 0.2 |
35 | Alloaromadendrene | 1471 | 1467 | 0.3 b | 0.6 b | 0.1 b | 0.3 ± 0.1 | 1.4 ± 0.2 | 0.2 b |
36 | Germacrene D | 1489 | 1480 | - | - | - | 0.1 b | 0.5 b | - |
37 | β-Selinene | 1493 | 1485 | 0.3 b | 0.6 b | - | 0.2 b | 0.5 ± 0.1 | 0.1 b |
38 | Valencene | 1496 | 1496 | 0.5 b | - | 0.1 b | 0.1 b | 1.4 ± 0.1 | 0.3 b |
39 | α-Selinene | 1505 | 1494 | 0.4 b | 0.4 b | 0.1 b | 0.1 b | 1.2 ± 0.1 | 0.2 b |
40 | γ-Cadinene | 1515 | 1514 | 0.3 b | - | - | 0.6 ± 0.2 | 0.4 b | - |
41 | δ-Cadinene | 1532 | 1530 | - | - | 0.1 b | 0.4 ± 0.1 | 0.4 b | - |
42 | α-Calacorene | 1547 | 1542 | - | - | - | 0.6 ± 0.1 | - | 0.2 b |
43 | Celina-3,7(11)-diene | 1554 | 1550 | - | - | 0.6 b | - | 0.3 b | - |
44 | Caryophyllene oxide | 1591 | 1583 | - | 0.6 b | 0.2 b | 0.1 b | 2.6 ± 0.4 | - |
Total | 94.8 ± 0.9 | 95.4 ± 1.0 | 97.5 ± 1.3 | 95.3 ± 1.1 | 91.5 ± 1.6 | 97.9 ± 0.7 |
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Pellati, F.; Brighenti, V.; Sperlea, J.; Marchetti, L.; Bertelli, D.; Benvenuti, S. New Methods for the Comprehensive Analysis of Bioactive Compounds in Cannabis sativa L. (hemp). Molecules 2018, 23, 2639. https://doi.org/10.3390/molecules23102639
Pellati F, Brighenti V, Sperlea J, Marchetti L, Bertelli D, Benvenuti S. New Methods for the Comprehensive Analysis of Bioactive Compounds in Cannabis sativa L. (hemp). Molecules. 2018; 23(10):2639. https://doi.org/10.3390/molecules23102639
Chicago/Turabian StylePellati, Federica, Virginia Brighenti, Johanna Sperlea, Lucia Marchetti, Davide Bertelli, and Stefania Benvenuti. 2018. "New Methods for the Comprehensive Analysis of Bioactive Compounds in Cannabis sativa L. (hemp)" Molecules 23, no. 10: 2639. https://doi.org/10.3390/molecules23102639