Extracts of the African potato have long been believed to have anti-cancer properties. The
aim of the current research was to isolate hypoxoside (HYP) from Hypoxis hemerocallidea
(African potato) and synthesize the dimethyl (DMH) and decaacetyl (DAH) derivatives and
to test their selective cytotoxicity on a model consisting of a normal (MCF10A) and
premalignant, invasive breast epithelial cells (MCF10A-NeoT).
Hypoxoside was extracted from the H. hemerocallidea corms using ethanol, purified using a
C-18 reverse phase column and the compound examined by nuclear magnetic resonance
(NMR) spectroscopy and high-resolution mass spectrometry and found to be of high purity.
This was also the case for the synthesized compounds. To assess possible selective effects
(cytotoxicity) of derivatized and underivatized hypoxoside, effects on the metabolism of
premalignant and normal cells were assessed using the 3-(4,5-dimethylthiazol-2-yl)-5-(3-
carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay. Effects on cell
number (total counts) and cell death [trypan blue and propidium iodide (PI) staining for dead
cells versus a lack of staining for live cells] were, thereafter, assessed. Imaging of live
adherent cells was also carried out using acridine orange (AO) and PI for live and dead cells
(respectively). Propidium iodide staining of detached cells was carried out for flow
cytometric determination of cell death (PI indicating early apoptotic or late apoptotic/necrotic
cells).
After treatment of normal (MCF10A) breast epithelial cells and premalignant cHa-rastransfected
(MCF10A-NeoT) derivative breast epithelial cells with HYP, DMH and the DAH
derivative, the MTS assay and the Duncan‟s multiple range, analysis of variance (ANOVA)
post hoc analysis of the MTS results revealed that only the 150 and 300 µM DAH derivative
had a statistically significant effect on the metabolic activity of the abnormal cell line relative
to the dimethyl sulfoxide (DMSO) and revealed no significant effect on the normal MCF-
10A cell line after treatment with any of the test compounds. Supravital PI staining of
adherent cells seemed to indicate a far higher rate of induction of cell death in abnormal cells
than evident in the MTS assay and the PI-based flow cytometry or the trypan blue exclusion
assays and need re-investigating, though result trends were similar.
Total cell counts, show that HYP and its derivatives appear to increase both cancer and
normal cell proliferation significantly, except in the case of DAH at 150 and 300 μM in the
MCF10A-NeoT, without affecting the MCF-10A cell line. The trypan blue method for
detection of cell death, together with total cell counts, the Duncan‟s analysis of MTS results
and a 24 hour exposure to test compounds, seems to constitute an optimal system for drug
screening and indicates the statistically significant selective toxicity of the DAH compound at
150 and 300 μM in the MCF10A-NeoT, suggesting that the DAH derivative at 150 and 300
µM would have significant, selective therapeutic potential on Ras-related malignancies. / Thesis (M.Sc.)-University of KwaZulu-Natal, Pietermaritzburg, 2013.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:ukzn/oai:http://researchspace.ukzn.ac.za:10413/9911 |
| Date | January 2013 |
| Creators | Xulu, Bongiwe Ziphelele. |
| Contributors | Elliott, Edith., Drewes, Siegfried E., Van Heerden, Fanie R. |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | English |
| Type | Thesis |
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