Hatoum et al. 1981 ). In addition, tetrahydrocannabivarin, a homologue of THC
with a propyl side chain, has been proven to antagonize mammalian cannabinoid
receptors (Thomas et al. 2005 ). In the 1990’s, endogenous cannabinoid receptor
ligands, such as anandamide and 2-arachidonoylglycerol, were identified in mam-
malian tissues (Devane et al. 1992 ; Mechoulam et al. 1995 ; Sugiura et al. 1995 );
thus, plant-derived cannabinoids are now often referred to as phytocannabinoids in
order to distinguish them from endocannabinoids.
Plant cannabinoids are classified into two types, neutral cannabinoids and
cannabinoid acids, based on whether they contain a carboxyl group or not. In live
Cannabisplants, cannabinoids are biosynthesized and accumulated as cannabinoid
acids, and non-enzymatically decarboxylized into their neutral forms during storage
and when smoked (Yamauchi et al. 1967 ; Kimura and Okamoto 1970 ). It was long
believed that tetrahydrocannabinolic acid (THCA) is biosynthesized by the iso-
merization of cannabidiolic acid (CBDA), whereas CBDA and cannabichromenic
acid (CBCA) were thought to be formed through the oxidative cyclization of
cannabigerolic acid (CBGA) (Mechoulam 1970 ). In order to confirm the steps in
the biosynthetic pathway, feeding experiments with radiolabeled precursors were
attempted, but no clear results were obtained due to the low incorporation rates of
radioactivity into cannabinoids (Shoyama et al. 1975 ; Kajima and Piraux 1982 ).
To overcome these challenges, starting in the middle of the 1990s, several
groups began to investigate the enzymes involved in cannabinoid biosynthesis, and
successfully elucidated the biosynthetic pathway of the major cannabinoids, as
illustrated in Fig.8.2. In the pathway, THCA, CBDA, and CBCA are biosynthe-
sized from the common precursor CBGA through the action of the unique oxi-
doreductases that include THCA synthase, CBDA synthase, and CBCA synthase,
respectively (Taura et al. 1995 , 1996 ; Morimoto et al. 1998 ). CBGA is synthesized
by the alkylation of olivetolic acid (OLA) with geranyl pyrophosphate through a
novel prenyltransferase known as geranyl pyrophosphate:olivatolate geranyltrans-
ferase (Fellermeier and Zenk 1998 ).
Of special interest, recent genomic and transcriptomic approaches conducted by
Page and co-workers demonstrated a novel mechanism for OLA biosynthesis: OLA
is formed by olivetolic acid cyclase (Gange et al. 2012 ), thefirst plant polyketide
cyclase, via the cyclization of a tetraketide-CoA intermediate produced by a
polyketide synthase (Taura et al.2009b). Novel omics-based studies have also
identified genes encoding geranyltransferase and the hexanoate-specific acyl-CoA
synthase (Page and Boubakir 2011 ; Stout et al. 2012 ). Thus, most structural genes
that encode biosynthetic enzymes are now available for molecular breeding in order
to control cannabinoid content inCannabisplants as well as for the biotechno-
logical production of cannabinoids in fermentation-friendly heterologous hosts such
as yeasts.
8 Cannabinoids: Biosynthesis and Biotechnological Applications 185