DISCUSSION
In order to better understand the phytotoxic potential of C.
cardunculus and increase the production of secondary metabolites, the
effect of light stress (shading) in field conditions was studied on the
STL profile of cultivated cardoon leaf extracts. The influence of light
stress on the production of plant STLs has never been studied. Given the
importance of these chemical compounds for their numerous biological
activities(Chaturvedi 2011), the knowledge of the factors affecting
their biosynthesis and production represents an important aspect that
should be considered for their future application.
In this study, the yield extract was greater in the April harvest than
January, with the stressed treatment showing the highest yield. These
yields are similar to those reported by Ramos et al. (Ramos et
al. 2013) and Scavo et al. (Scavo et al. 2019f). The yields of
extracts were closely correlated to STL concentrations. In fact,
extracts belonging to April harvest generally showed higher
concentrations of total and single STLs than January, probably due to
the higher levels of global radiation and longer photoperiod. A similar
trend was reported by Scavo et al. (Scavo et al. 2019f) for STLs
on the three C. cardunculus taxa and by Pandino et al. (Pandino,
Lombardo, Lo Monaco & Mauromicale 2013) on globe artichoke leaf
polyphenols. The leaves of many Asteraceae members, including C.
cardunculus , are known for the high cocentrations of STLs, mainly
represented by cynaropicrin, which is responsible of its bitter
taste(Seaman 1982). Here, in accordance with Ramos et al. (Ramoset al. 2013), cynaropicrin was the most abundant STL detected.
However, most of previous studies are focused on the globe
artichoke(Rouphael et al. 2016), while cultivated cardoon has
been little investigated.
While the influence of light quality (e.g. ionizing or ultraviolet
radiation, intensity of visible light, red and far-red light) on the
allelopathic expression of plants has been reported in previous
researches (del Moral 1972; Kato-Noguchi 1999; Li et al. 2009),
only few works investigated the effect of daylength. Zucker et al.
(Zucker, Nitsch & Nitsch 1965) stated that chlorogenic acid
concentration in Nicotiana spp. species is affected by
photoperiodic alteration. Rice (Rice 1984) indicated that long days
generally increase the production of inhibitory compounds in donor
plants. Similarly, Taylor (Taylor 1965) found that the concentration of
different phenolics and cinnamic acid derivatives increase in the leaves
of Xanthium strumarium L. with increases in daylength. Here we
found an opposite trend to those reported in literature. Indeed, the
amount and composition of STLs in cultivated cardoon leaf extracts
varied in relation to both light stress and harvest time, with a more
pronounced effect of the latter factor, suggesting how the
meteorological conditions (e.g. global radiation, photoperiod, air
temperature and total rainfall) affect their biosynthesis. However,
except for aguerin B, the shading significantly increased
the concentration of total and
single STLs in the April harvest, compared to control, and this increase
was more marked if combined with harvest time. It is known that once a
stress factor is recognised at cellular level by a plant, a signal
transduction begins leading to gene expression and metabolic responses
with the aim of helping the plant survive or minimizing the
effectiveness of the stress agent (Reigosa et al. 1999).
Eljounaidi et al. (Eljounaidi et al. 2014) elucidated the genes
and the enzymes involved in the biosynthetic pathway of STLs in C.
cardunculus . However, the alteration of gene expression in this species
is still unknown, but appears reasonable how this plant, as commonly
happens in other ones, prevents the oxidative damages induced by ROS
(reactive oxygen species such as hydrogen peroxide, superoxide, etc.)
through the synthesis of secondary metabolites, STLs and polyphenols in
this case. However, in nature plants interact with the complex of
abiotic and biotic influences of the environment, hence an allelopathic
phenomenon depends on the type and intensity of stress, on the genotype
and development stage of the plant, as well as on the seasonal changes
and, therefore, may vary through time (Scavo, Abbate & Mauromicale
2019a). The influence of genotype and harvest time have been previously
studied (Scavo et al. 2019f), while light stress is the first
abiotic factor evaluated in C. cardunculus .
With the aim to find a correlation between the STL composition and
phytotoxicity, cultivated cardoon leaf extracts were bioassayed on the
wheat coleoptile elongation, as well as on germination, root and shoot
length of A. retroflexus and P. oleracea . In agreement
with previous results with the same bioassay and target species bothC. cardunculus (Scavo et al. 2019e) and other donor plants
(da Silva et al. 2017), seed germination percentage was the least
affected parameter. Contrariwise, wheat coleoptile elongation and root
length of both weeds were significantly inhibited. The phytotoxic
activity increase at increasing concentrations, as widely reported in
literature (Ambika 2013), and in correspondence of the April harvest, as
found also by Scavo et al. (Scavo et al. 2019f) and Pandino et
al. (Pandino et al. 2013) for polyphenols. These results are
corroborated by STL profile. During April, in fact, the plant
accumulated more STLs in the leaves, mainly cynaropicrin. Probably, the
combination of high temperatures, low rainfall, high global radiation
and long photoperiod stimulates the production of C. cardunculussecondary metabolites (caffeoylquinic acids, flavones and STLs) and
increases its inhibitory potential. Nevertheless, the imposition of
plant shading in April was found to stimulate the production of STLs in
the leaves and to enhance, albeit to a lesser extent, the allelopathic
activity.
In conclusion, here we demonstrated, for the first time, that plant
shading in field conditions enhances the production of STLs in the
leaves of cultivated cardoon, especially in the April harvest. This
increase was accompanied by a higher phytotoxicity on wheat coleoptile
elongation, germination percentage, root and shoot length of A.
retroflexus and P. oleracea . The inhibitory activity of
cultivated cardoon extracts caused by shading was weak if taken alone,
becoming stronger when combined with the harvest time due to the optimal
combination of climatic conditions, development stage of the plant and
intensity of light stress. The induction of light stress by plant
shading could be a useful tool to stimulate the biosynthesis of STLs for
industrial applications and to increase the phytotoxic potential of
cultivated cardoon as a sustainable weed control method within an
integrated weed management system for the future production of a
bioherbicide.