- Title
- Investigation of the biological and anticancer properties of bioactive compounds derived from Helicteres hirsuta Lour. and Catharanthus roseus (L.) G. Don
- Creator
- Pham, Hong Ngoc Thuy
- Relation
- University of Newcastle Research Higher Degree Thesis
- Resource Type
- thesis
- Date
- 2019
- Description
- Research Doctorate - Doctor of Philosophy (PhD)
- Description
- Selected plants, including Helicteres hirsuta Lour. (H. hirsuta) and Catharanthus roseus (L.) G. Don (C. roseus) have been used traditionally as medicinal plants for the treatment of various ailments in many countries. The root of H. hirsuta has been utilized for treating malaria and diabetes in Thailand, whereas Vietnamese people have used H. hirsuta leaf and root for pain relief, dysentery, measles, flu, antidote and poliakiuria. The hot water extract of the dried C. roseus plant has been taken orally for the treatment of cancer, heart disease and leishmaniasis in Peru. Also, the hot water extract of the dried C. roseus leaf has been used for the treatment of diabetes in Jamaica, Kenya and West India. Moreover, previous studies have found that these plants contain bioactive compounds with anticancer properties against various cancer cell lines. Some isolated lignans from H. hirsuta stem, including (±)-pinoresinol, (−)-boehmenan and (−)- boehmenan H, possessed cytotoxic activity against human lung carcinoma, hormone-dependent human prostate carcinoma and human breast carcinoma. Vinblastine and vincristine derived from C. roseus have been used in combination with other cancer chemotherapeutic drugs for treating advanced testicular cancer, breast and lung cancers. Consequently, H. hirsuta and C. roseus are important materials for the exploration of new phytochemicals for potential utilisation in the pharmaceutical industry. Cancer, a major cause of morbidity and mortality, affects populations in all countries. Cancer is also costly, being a significant financial burden on the people of low- and middle income countries. Of note, pancreatic cancer is the fourth leading cause of cancer death worldwide and has a low 5-year survival rate. Up to now, there have been limited studies conducted to investigate the anti-pancreatic cancer activity and potential of H. hirsuta and C. roseus. Therefore, these two medicinal plants were selected as the starting materials for extraction and isolation of known and un-identified bioactive compounds for in vitro assessment against a panel of cancer cell lines, in particular pancreatic cancer cells. Additionally, the derived bioactive compounds from these plants were investigated for their antioxidant and antimicrobial properties. The overall aim of this study was to extract and isolate bioactive compounds with antioxidant, antimicrobial, anticancer (particularly pancreatic cancer) activities from H. hirsuta and C. roseus. Specific objectives of the project included (1) determining the effect of different extraction solvents and drying methods on bioactive compound yield and antioxidant capacity of plant extracts; (2) optimisation of extraction conditions to maximise bioactive compound yield from dried plant materials; and (3) elucidation of the phytochemical profiles and assessment of the antioxidant, antimicrobial, and anticancer (particularly pancreatic cancer) properties in vitro of the derived extracts and fractions from H. hirsuta and C. roseus. This study revealed that H. hirsuta and C. roseus possessed high levels of bioactive compounds, including phenolics, flavonoids and saponins with strong antioxidant capacity. The yield of bioactive compounds obtained from these plants was significantly affected by the nature of the extraction solvent. Among the range of solvents tested, water was demonstrated to be suitable for phenolic extraction, whereas methanol revealed its dominant advantage in saponin extraction from both H. hirsuta and C. roseus. Various thermal drying conditions also had a remarkable effect on the retention of bioactive compounds and antioxidant property of the plant samples. The extraction process is a crucial step in liberating bioactive compounds from the plant matrix. It is necessary to determine the optimised extraction conditions as this directly impacts extraction efficiency, the stability and yield of the isolated bioactive compounds. Ultrasound-assisted extraction (UAE) was selected in this investigation to reduce extraction time, increase extraction yield and retain bioactive compounds when compared to conventional extraction. Response surface methodology (RSM) was employed to generate the optimal UAE conditions for enhanced total phenolic content, saponins and antioxidant capacity of H. hirsuta and C. roseus. The crude extracts from H. hirsuta and C. roseus contained a number of individual components with significant bioactivity. As such, the crude extracts were firstly separated using the liquid-liquid extraction technique to obtain extracts of increasing polarity, including n-butanol (saponin-enriched) and residual aqueous fractions. The phytochemical profiles of the crude extracts and their derived fractions, including saponin-enriched butanol and aqueous fractions, were elucidated by HPLC analysis. They were then subjected to antioxidant, antimicrobial and cytotoxic screening. The results highlighted that the saponin-enriched butanol fractions from either H. hirsuta leaf and stem or C. roseus stem and root exhibited strong antioxidant and antimicrobial capacity, as well as cytotoxic activity against a panel of cancer cell lines, including A2780 (ovarian), H460 (lung), A431 (skin), MIA PaCa-2 (pancreas), Du145 (prostate), HT29 (colon), MCF-7 (breast), BE2-C (neuroblastoma), SJ-G2, U87 and SMA (glioblastoma) at low doses. Of note, they possessed potent in vitro anti-proliferative activity towards MIAPaCa-2, BxPC-3 and CFPAC-1 pancreatic cancer cells. These findings indicated that n-butanol was effective in extracting antioxidants, antimicrobials and anticancer components from these plants. Thus, the saponin-enriched butanol fraction from H. hirsuta stem, one of the potent saponin-enriched butanol fractions, was subsequently fractionated using semi-preparative reversed-phase (RP) HPLC. The anti-proliferative data showed that the five xxvii sub-fractions derived from the saponin-enriched butanol fraction of H. hirsuta stem inhibited the growth of the pancreatic cancer cells via the induction of apoptosis, as well as promoting S phase cell cycle arrest. The identification of the five active sub-fractions were further determined using nanoflow RP Liquid Chromatography coupled directly to a Q-Exactive Plus Hybrid Quadrupole Orbitrap Mass Spectrometer. The LC/MS analysis revealed that the isolated fractions from H. hirsuta stem contained bioactive compounds, such as rosmarinic acid, caffeic acid, sagerinic acid, usnic acid, cucurbitacin S, 2-O-β-D-glucopyranosyl-cucurbitacin B (Arvenin I), 4-oxo-beta-isodamascol and absinthin, in which most have been previously demonstrated to exhibit diverse biological activities. In conclusion, these findings further confirm that H. hirsuta and C. roseus contain promising bioactive compounds with diverse biological activities, such as antioxidant, antimicrobial and anticancer (eg. pancreatic cancer) activities. It is expected that the outcomes from this research will provide a strong basis for further assessment in vivo and pre-clinical models, and perhaps then further investigation in clinical trials.
- Subject
- Helicteres hirsuta Lour.; Catharanthus roseus (L.) G. Don; biological property; anticancer property; bioactive compounds; thesis by publication
- Identifier
- http://hdl.handle.net/1959.13/1412999
- Identifier
- uon:36561
- Rights
- Copyright 2019 Hong Ngoc Thuy Pham
- Language
- eng
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