Waheed, Abdul (2011) Isolation and characterisation of potential anticancer compounds from medicinal plants. (PhD thesis), Kingston University.Full text not available from this archive.
The research work presented in this thesis deals with the anticancer activity of four medicinal plants: 'Caralluma tuberculata' (Asclepiadaceae), 'Fagonia indica' (Zygophyllaceae), 'Solanum surattense' (Solanaceae) and 'Arisaema utile' (Araceae) that originate from the North West and Himalayan regions of Pakistan. Through a bioactivity-guided fractionation approach, the crude and resultant organic fractions were tested on cultured breast cancer cells (MCF-7 and MDA MB-468) and colorectal carcinoma cells (Caco-2) in vitro. Five new compounds out of seven in total were isolated from potent fractions of the new medicinal plants using repeated flash column chromatography. Structural elucidation was carried out through a series of spectroscopic experiments (1-D and 2-D NMR, GC-MS, LC-MS). SIngle crystal X-ray structure was determined using X-ray crystallography for the crystalline compounds, which showed a defraction pattern. The apprent IC[sub]50 for compounds (1-6) were estimated from serial dilutions of eight concentrations (0.78-100 [mu]M) of each compound, tested against breast and colon cancer cell lines, using two cell viability assays (MTT and neutral red uptake assays) for 24 h and 48 h treatments. Two new steroidal glycosides, acylated pregnane (1) and acylated androstane (2) glycosides, isolated from the ethyl acetate fraction of 'Caralluma tuberculata' showed highly significant (P<0.001) percentage growth inhibition in Caco-2 cells (IC[sub]50) 1.56-6.25 [mu]M) and MCF-7 cells (IC[sub]50 6.25 - 25 [mu]M), however, oestrogen independent cancer cells (MDA MB-468) were less responsive with IC[sub]50 25 - 50 [mu]M. These steroidal glycosides induced apoptosis in cancer cells as measures of cytoxic activity (NRU, PARP clevage, DNA ladder) on MCF-7, MDA MB-468 and Caco-2 cells were inhibited by pre-treatment with the pan-caspase inhibitor (Z-VAD-FMK). Another pregnane glycoside (3), isolated for the first time from 'Fagonia indica', was found to be more potent in suppressing cell growth (IC 6.25-25 [mu]M), in oestrogen negative breast cancer cells (MDA MB-468,) as compared to oestrogen positive cancer cells (MCF-7). Although a cleaved PARP (89kDa) was detected by Western blotting, cytomorphological alterations and in cells pre-treated with a pan-caspase inhibitor (NRU assay), indicated that the necrosis mode of cell death is more likely. Moreover, three esters: hexadecanoic acid ethyl ester (4), phtalic acid 1-(1, 1-dimethyl-pentyl) ester 2-(2-ethyl-dec-5-enyl) ester (5) from chloroform fraction of 'Solanum surattense', and 5-Oxo-19-propyl-docosanoic acd methyl ester (6) from 'Arisaema utile', showed a highly significant )p<0.001) decrease in cell numbers for MDA MB-468 and Caco-2 cells with apparent IC 6.25-12.5 [mu]M in cell viability assays (MTT and NRU) after 48 h treatment, while MCF-7 cells were less responsive (IC[sub]50 25 [mu]M). Compunds 5 and 6 (first report from a natural source) did not restrict the growth inhibition in MCF-7 and Caco-2 cells, pre-treated with Z-VAD-FMK, which indicated less involvement of Capase-dependent apoptosis, while DAPI staining and Western blots (cleaved-PARP) showed characteristics of apoptosis that suggested the possibility of aponecrosis phenomenon of cell death. In preliminary screening (Western blot and DNA ladder assays), compounds (1-6) were not toxic to normal human cells (HUVEC and U937) and indicated some selctivly between malignant and normal cells.
|Item Type:||Thesis (PhD)|
|Physical Location:||This item is held in stock at Kingston University library.|
|Research Area:||Biological sciences
|Faculty, School or Research Centre:||Faculty of Science (until 2011) > School of Pharmacy and Chemistry|
|Depositing User:||Abdul Waheed|
|Date Deposited:||25 Jul 2012 08:36|
|Last Modified:||21 Sep 2012 14:16|
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