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Analysis of cannabinoids in laser-microdissected trichomes of medicinal Cannabis sativa using LCMS and cryogenic NMR.

By July 22, 2013No Comments

pm2Analysis of cannabinoids in laser-microdissected trichomes of medicinalCannabis sativa using LCMS and cryogenic NMR.


Department of Technical Biochemistry, Technical University of Dortmund, Technische Biochemie, Dortmund, Germany.


Trichomes, especially the capitate-stalked glandular hairs, are well known as the main sites of cannabinoid and essential oil production of Cannabis sativa. In this study the distribution and density of various types of Cannabis sativa L. trichomes, have been investigated by scanning electron microscopy (SEM). Furthermore, glandular trichomes were isolated over the flowering period (8 weeks) by laser microdissection (LMD) and the cannabinoid profile analyzed by LCMS. Cannabinoids were detected in extracts of 25-143 collected cells of capitate-sessile and capitate stalked trichomes and separately in the gland (head) and the stem of the latter. Δ(9)-Tetrahydrocannabinolic acid [THCA (1)], cannabidiolic acid [CBDA (2)], and cannabigerolic acid [CBGA (3)] were identified as most-abundant compounds in all analyzed samples while their decarboxylated derivatives, Δ(9)-tetrahydrocannabinol [THC (4)], cannabidiol [CBD (5)], and cannabigerol [CBG (6)], co-detected in all samples, were present at significantly lower levels. Cannabichromene [CBC (8)] along with cannabinol (CBN (9)) were identified as minor compounds only in the samples of intact capitate-stalked trichomes and their heads harvested from 8-week old plants. Cryogenic nuclear magnetic resonance spectroscopy (NMR) was used to confirm the occurrence of major cannabinoids, THCA (1) and CBDA (2), in capitate-stalked and capitate-sessile trichomes. Cryogenic NMR enabled the additional identification of cannabichromenic acid [CBCA (7)] in the dissected trichomes, which was not possible by LCMS as standard was not available. The hereby documented detection of metabolites in the stems of capitate-stalked trichomes indicates a complex biosynthesis and localization over the trichome cells forming the glandular secretion unit.
Copyright © 2012 Elsevier Ltd. All rights reserved.

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Graphical abstract

LCMS and cryogenic NMR analysis of cannabinoids in laser dissected glandular trichomes from Cannabis sativa L.

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► Specific cannabis trichomes and their individual parts were isolated using LMD. ► Nine cannabinoids were analyzed in low dissected samples with LCMS and cryogenic NMR. ► The first report of cannabinoids detection in the stem of capitate-stalked trichomes. ► The use of LMD in the dissection does not influence labile cannabinoids acid. ► Stems of capitate-stalked trichomes might play a role in cannabinoids production.


  • Cannabis sativa;
  • Laser microdissection (LMD);
  • Liquid chromatography–mass spectrometry (LCMS);
  • Cryogenic nuclear magnetic resonance (NMR);
  • Cannabinoid biosynthesis;
  • Capitate-stalked trichome;
  • Capitate-sessile trichome;
  • Gland;
  • Scanning electron microscopy (SEM)

Figures and tables from this article:

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Fig. 1. Structures of identified cannabinoids.
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Fig. 2. Trichomes of Cannabis sativa L.: (A) trichomes on the flower, (B) capitate-stalked trichome, (C) capitate-sessile trichome, (D) bulbous trichome, (E) trichomes on the bract, (F) trichomes on the stem, (G) trichomes on the adaxial surface of a floral leaf; a big capitate-sessile trichome is indicated with an arrow, (H) trichomes on the abaxial surface of a leaf; present abundant small capitate-sessile and bulbous trichomes.
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Fig. 3. Microdissection of trichomes of medicinal Cannabis sativa L. (A) intact capitate-stalked trichome before dissection, (B) capitate-stalked trichome after dissection of the head cells, (C) capitate-stalked trichome – complete dissection, (D) dissected head cells, (E) stem cells after dissection, (F) capitate-sessile trichome before dissection, (G) view after dissection of the capitate-sessile trichome, (H) dissected capitate-sessile trichomes.
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Fig. 4. Total ion chromatograms of ion pair multiple reaction monitoring (MRM) of identified cannabinoids and internal standards, LCMS analysis at week 8. ST: intact capitate-stalked trichomes, D: heads of capitate-stalked trichomes, B: stems of capitate-stalked trichomes, SE: intact capitate-sessile trichomes; 1: THCA, 2: CBDA, 3: CBGA, 4: THC, 5: CBD, 6: CBG, 8: CBC, 9: CBN.
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Fig. 5. 1H NMR spectra of dissected trichomes and reference compounds. (A) Capitate-stalked trichomes at week 8. (B) Capitate-sessile trichomes at week 8. (C) Reference THCA. (D) Reference CBDA. Signals in spectrum A are labeled with compound numbers 12, and 7 (CBCA).
Table 1. Density and distribution of trichomes on flowers, leaves, and stems of Cannabis sativa L. during flowering period. T: trichomes, NT: number of trichomes, D: density of trichomes, B: bract, Fd: adaxial surface of a floral leaf, Fb: abaxial surface of a floral leaf, Ld: adaxial surface of a leaf, Lb: abaxial surface of a leaf, S: stem, A: average, NF: not found, NC: not counted.
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Table 2. Multiple reaction monitoring (MRM) transitions of the components measured by electrospray ionization; positive voltages in positive ionization, negative voltages in negative ionization.
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Table 3. Concentrations of cannabinoids in dissected trichomes samples based on LCMS analysis. N: number of dissected trichomes, ST: intact capitate-stalked trichomes, D: heads of capitate-stalked trichomes, B: stems of capitate-stalked trichomes, SE: intact capitate-sessile trichomes; number in the sample name represents the collection week; ND for CBD, CBG, CBN, OA: <3 ng/mL in measured sample, ND for CBC: <30 ng/mL in measured sample.
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Corresponding author contact information
Corresponding author. Tel.: +49 231 7557 487; fax: +49 231 7557 489.

Copyright © 2012 Elsevier Ltd. All rights reserved.

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