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

• Published in: Phytochemistry (Oxford) Vol. 87; pp. 51 - 59
• Main Authors: Happyana, Nizar, Agnolet, Sara, Muntendam, Remco, Van Dam, Annie, Schneider, Bernd, Kayser, Oliver
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.03.2013
• Subjects: biosynthesis; cannabidiol; Cannabinoid biosynthesis; Cannabinoids - analysis; Cannabinoids - chemistry; Cannabis - metabolism; Cannabis sativa; Capitate-sessile trichome; Capitate-stalked trichome; Chromatography, Liquid - methods; Cryogenic nuclear magnetic resonance (NMR); essential oils; flowering; Gland; Laser microdissection (LMD); Liquid chromatography–mass spectrometry (LCMS); Magnetic Resonance Spectroscopy - methods; Mass Spectrometry - methods; metabolites; nuclear magnetic resonance spectroscopy; scanning electron microscopy; Scanning electron microscopy (SEM); secretion; stems; trichomes; Cannabis sativa; Capitate-stalked trichome; Capitate-sessile trichome; Scanning electron microscopy (SEM); Cryogenic nuclear magnetic resonance (NMR); Laser microdissection (LMD); Cannabinoid biosynthesis; Gland; Liquid chromatography–mass spectrometry (LCMS)
• ISSN: 0031-9422; 1873-3700
• DOI: 10.1016/j.phytochem.2012.11.001

LCMS and cryogenic NMR analysis of cannabinoids in laser dissected glandular trichomes from Cannabis sativa L. [Display omitted] ► 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. 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 (8weeks) 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. Δ9-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, Δ9-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.