FIGURE 11 Principal component analysis of secondary metabolites
of A. trifoliata in different habitats
4 | DISCUSSION
Environmental stress can not only inhibit plant growth, but also
stimulate the synthesis and accumulation of plant secondary metabolites
(Zhang et al., 2020). However, the biggest characteristic of rocky
desertification habitat is that it does not keep water, resulting in a
large amount of water loss. Thus causing certain drought stress to
plants. The biomass of various organs of A. trifoliatasignificantly decreased in the rocky desertification habitat, which
indicates that the desertification habitat has caused a certain abiotic
stress on A. trifoliata , and the metabolic physiological
processes of its plants will inevitably change.
4.1 | Effects of rocky desertification on the
contents of basic secondary metabolites in A. trifoliata
The secondary metabolites such as phenols and flavonoids are closely
related to plant stress defense (Sanchez et al., 2011). In this study,
we found that rocky desertification could promote the increase of total
phenol content in roots, stem and leaf of A. trifoliata , which
was the same as that of total phenol secretion of Desi ajwain
(Trachyspermum ammi L.) under drought stress (Azhar et al.,
2011). The possible reason is that the increase of total phenol content
can help to reduce the water potential in cells, prevent the loss of
water in cells; At the same time, phenol can also remove reactive oxygen
species produced under drought (Nakabayashi et al., 2015). In this
study, it was found that the content of polyphenols in leaves changed
the most, which may be due to the fact that leaves are the largest organ
of plants exposed to the outside world, and they are most seriously
exposed to oxidative stress. Therefore, they secrete a large number of
polyphenols, remove reactive oxygen species and free radicals through
hydrogen atom transfer mechanism, and are used for the synthesis of
antioxidants to resist oxidative damage (Ping et al., 2017).
Tannins and flavonoids belong to the phenolic compounds of plant
secondary metabolites, which are good free radical scavengers and lipid
peroxidation inhibitors (Dicko et al., 2005). The results showed that
the content of tannin in A. trifoliata increased rapidly in the
early stage and decreased slowly in the later stage. The possible reason
is that water shortage in rocky desertification habitat can induce the
increase of secondary metabolism related enzyme activities in A.
trifoliata , so a large number of secondary metabolites are activated.
Finally, it is transported to various organelles to repair the damage of
membrane system and improve the tolerance of plants to drought stress,
which is a self-protection mechanism for plants to cope with stress
(Castellarin et al., 2007). The tannin content would slow down and
decrease in the later rocky desertification habitat. The possible reason
is that with the aggravation of stress, the plant organ damage is
serious, the metabolism speed in the body slows down, resulting in the
reduction of carbon sources required for the synthesis of phenols.
Therefore, the tannin content in the plant does not increase or even
decrease. Flavonoids usually have special structures such as hydroxyl,
double carbon bond, glycosylation, acylation, methylation, etc., which
can effectively resist oxidative damage induced by stress. At the same
time, they can also complete antioxidant function by preventing the
generation of reactive oxygen species and scavenging reactive oxygen
species (Agati et al., 2010; Singanusong et al., 2015). In this study,
we found that rocky desertification habitat can improve the content of
flavonoids in the root and stem of A. trifoliata , which is the
same as Zahir et al., (2014) study on water stress in Silybum marianum.
The possible reason is the water deficiency caused by rocky
desertification, which affects the up regulation of genes related to
flavonoid metabolism pathway, such as MYB transcription factor,
flavone-3-hydroxylase (F3H) and Flavonol Synthase (FLS) (Ying et al.,
2018; Fan et al., 2018), thus increasing the content of flavonoids.
However, there is no obvious rule for the change of flavonoids content
in A. trifoliata leaves in rocky desertification habitat in the
two bases. The possible reason is that the flavonoids content in leaves
is less affected by rocky desertification habitat, so there is no
obvious change rule.
4.2 | Effects of rocky desertification on
enzyme activities of phenylpropanoid pathway in A. trifoliata
The changes of biological and abiotic factors do not directly affect the
synthesis of plant secondary metabolites, but affect the synthesis of
secondary metabolites by affecting the expression of key genes and key
enzyme activities in secondary metabolism (Havaux and Kloppstech 2001).
PAL, C4H and 4CL are key enzymes in phenylalanine metabolism pathway,
and their activities are affected by environmental factors such as
light, temperature and water (Dupont and Aksnes 2010). This study found
that A. trifoliata in rocky desertification habitat can improve
the activities of pal, C4H and 4CL in roots, 4CL in stems and PAL, 4CL
in leaves. Obviously, different enzymes have different responses to
rocky desertification habitat, and the changes in different tissues and
organs are also different. In rocky desertification habitat, the three
enzymes in roots were significantly increased, which may be due to the
root as the main organ of plant drought resistance, its flavonoids
content is also the most, so its response changes are also the most
obvious, especially pal and C4H changes are the most obvious. It can be
seen that the appropriate degree of drought stress can promote the
accumulation of flavonoids by stimulating the expression and activity of
key enzymes involved in flavonoid biosynthesis. This is consistent with
the research results of Cheng et al., (2018) that moderate drought
stress can promote baicalin accumulation by stimulating the key enzyme
activity and expression of Baicalin biosynthesis.
4.3 | Effects of rocky desertification on
oleanolic acid and α-hederagenin contents in A. trifoliata
Oleanolic acid and α-hederagenin are the two most important medicinal
components of A. trifoliata . They are triterpenoids in
terpenoids. They can not only participate in interspecific competition
as interspecific sensing compounds (Arimura et al., 2000), but also be
used as raw materials for spices, flavorings and cosmetics (Martin et
al., 2003), pesticides and industrial raw materials (George et al.,
2015), As well as anti-tumor, anti-inflammatory, antibacterial,
antiviral, antimalarial, promoting percutaneous absorption, prevention
and treatment of cardiovascular disease, hypoglycemic and other
biological activities (Chen et al., 2021; Sun et al., 2019). However,
their ecological functions, biosynthetic pathways and environmental
factors are not very clear. In this study, we found that rocky
desertification could increase the contents of oleanolic acid and
α-hederagenins in the roots, stems and leaves of A. trifoliata .
The possible reason is the water stress caused by rocky desertification
habitat, which induces the expression of BPW and BPY genes and reduces
the expression of BPX genes, resulting in the accumulation of
triterpenoid saponins (Yin et al., 2015). This is the same as the result
of Chen et al., (2011) which found that moderate drought would increase
oleanolic acid in Prunella vulgaris L., and the possible reason
is that oleanolic acid has antioxidant effect, which can help A.
trifoliata resist the oxidative damage caused by rocky desertification
habitat (Zhao et al., 2013). However, the role of α-hederagenins under
drought stress is still unclear, so it needs to be studied. At the same
time, this study found that rocky desertification habitat did not affect
the distribution of oleanolic acid content in all organs of A.
trifoliata , which was still root > stem >
leaf. However, in Huaxi base, the peak period of oleanolic acid content
in roots was delayed to a certain extent, but not in Guanling base. The
possible reason is that A. trifoliata adapts to rocky
desertification habitat stress for a long time in Huaxi base, which
causes the peak period of oleanolic acid content to shift. However, the
content peak and distribution of α-hederagenins in various organs did
not change.
Although the biomass of roots, stems and leaves of A. trifoliatawill decrease in the rocky desertification habitat, the content of
medicinal ingredients has increased significantly. This not only brings
new economic benefits to A. trifoliata for restoring rocky
desertification habitats, but also further improves the quality ofA. trifoliata medicinal ingredients.
5 | CONCLUSION
The response of secondary metabolites of A. trifoliata to rocky
desertification was studied. The results showed that the rocky
desertification habitat reduced the biomass of A. trifoliata , but
significantly increased the tannin content in the leaves, the flavonoid
content in the roots, and the total phenol content in the roots, stems
and leaves, and reduced the flavonoid content in the leaves; The
activities of PAL, C4H and 4CL in roots, 4CL in stems and PAL, 4CL in
leaves of A. trifoliata in rocky desertification habitat were
increased. The content of oleanolic acid and α-hederagenin in the root,
stem and leaf of A. trifoliata was increased in rocky
desertification habitat. In addition, the content of oleanolic acid in
the root of A. trifoliata was the highest from July to August,
which was suitable for picking medicine. The content of α-hederagenin in
the stem was the highest in July, which was suitable for picking
medicine. The principal component analysis showed that the main response
index of secondary metabolites of A. trifoliata in rocky
desertification habitat was total phenol. The results of this study are
of great signifificance to cultivators and researchers of medicinal
plants.