Biotransformation of artemisinin and asiaticoside: Malaysian tropical plant phytochemicals
Centella asiatica is one of the pioneering herbs that act actively in traditional remedy such as treating skin problems, including wound healing. Centella asiatica has been utilized as a traditional herbal medicine in Malaysia on different occasions and treatments including wound healing. Asiatico...
| Main Authors: | , , |
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| Format: | Monograph |
| Language: | English |
| Published: |
2016
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| Subjects: | |
| Online Access: | http://irep.iium.edu.my/50230/ http://irep.iium.edu.my/50230/1/EDW_B_14_END_OF_PROJECT_REPORT_EDW_B14-109-0994_edited.pdf |
| Summary: | Centella asiatica is one of the pioneering herbs that act actively in traditional remedy such as treating
skin problems, including wound healing. Centella asiatica has been utilized as a traditional herbal
medicine in Malaysia on different occasions and treatments including wound healing. Asiaticoside
(A.S) is a major bioactive constituent in Centella asiatica as it has been reported to have
antimicrobial effects and wound healing properties. Microbial transformation was extensively used to
create useful metabolites from various natural products and as an alternative to chemical synthesis
for the preparation of pharmacologically active compounds. This study aims to isolate A.S, the major
phytochemical in Centella asiatica that plays biological activities including wound healing, which will
presented for microbial transformation using Aspergillus niger fungi. Then, the wound healing activity
of A.S and the asiaticoside transformed product (T.P) will be investigated, and then the effect of
these products on TGFβ1 and TIMP1 gene expression will be examined to understand the mode of
action of A.S and T.P on wound healing. Extraction of the phytochemicals was achieved using
Soxhlet extraction with three types of organic solvents with different polarities, i.e. n-hexane,
dichloromethane, and methanol. Phytochemical screening of the crude extracts showed the
presence of several biological constituents and compounds, such as terpenoids and saponins, which
include the targeted phytochemical A.S. The results showed that the methanolic crude extract
contained the highest amount of A.S, This extract was fractionated using several techniques
including liquid-liquid separation, column chromatography and size exclusion purification to afford
A.S with 0.22 % recovery. Subsequently, A.S was identified by HPLC, FTIR, and NMR spectrometry.
A.S with a concentration of 0.1 mg/ml was introduced into the microbial culture of A. niger and
incubated for 8 days using a rotary shaker at 30 ºC. Then, extraction of the T.P was done by liquidliquid
extraction to afford 50 mg of dark brown residue analyzed by TLC, 1D and 2D FTIR and proton
NMR for metabolite profiling. In the third phase of the study, A.S and T.P were used in the cell
proliferation and wound healing experiments using human keratinocytes. A.S and T.P showed
promising results in vitro model. Results showed that A.S concentrations between 7.5 and 120 μg/ml
gave higher value of cell proliferation than negative control, prompting cell proliferation improvement.
The concentration of 240 μg/ml gave the lowest percentage of cell proliferation, though it did not
indicate to have a cytotoxic effect, however the IC50 of A.S was ~ 446.2 μg/ml. The result showed
that A.S has indeed low toxicity on human keratinocytes at the highest used concentration of 240
μg/ml. On the other hand, it was clear that T.P concentrations between 0.058 and 3.75 μg/ml
exhibited the higher viability values, whereas, the concentration at 7 μg/ml showed the lowest
viability value. Moreover it was detected that the concentrations from 15 to 240 μg/ml presented a
severe toxicity toward human keratinocytes, IC50 value of T.P was found ~ 7.423 μg/ml, which also
may confirm the change in the bioactivity properties of the original A.S. In wound healing assays
analysis it was shown that there were significant differences with comparison with the negative
control (P<0.05). Two-way ANOVA tests also showed that there was significant effect of the products
(drugs) as well as the exposure time with P values 0.0113 and 0.0001 correspondingly. Statistical
analysis showed that both A.S and T.P could increase the expression of TGFβ1 (P = 0.01 and 0.007)
respectively, and the expression of TIMP1 (P = 0.0002 and 0.0006) respectively, however T.P was
more effective than A.S (P = 0.009 for TGFβ1) and (0.001 for TIMP1). It was witnessed T.P showed
a faster migration and healing speed under macroscopic observation. In addition it was noted that as
well as T.P could considerably enhanced TIMP1 gene expression. It could be concluded that
biotransformation process could produce biological active products that can be used in wound
healing and pharmaceutical purposes. Since, this study was confirmed that T.P could enhance the
cell proliferation and wound healing of the human keratinocytes with very low concentration in comparison to the original substrate.
Meanwhile, artemisinin has remained an effective drug to treat malaria. Numerous approaches have
been adapted to increase the efficacy of artemisinin against antibiotic resistant malarial parasite.
Microbial biotransformation of artemisinin has been used recently to produce promising derivatives of
artemisinin on a large scale with low costs. During the last decade several biotransformation studies
on artemisinin, by using microorganisms, have been reported. This literature review focuses on the
most recent microbial transformation studies on artemisinin and its derivatives. |
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