Background:A decoction (hot-water extract) comprised of Nigella sativa (seeds), Hemidesmus indicus (roots), and Smilax glabra (rhizome) has been reported to prevent chemically-induced hepatocarcinogenic changes in rats and to exert significant cytotoxic effects on human hepatoma (HepG2) cells. However, the decoction used in previous studies to determine cytotoxicity was not standardized. Further, during preparation of pharmaceuticals for clinical use, it is more convenient to use an ethanolic extract. Therefore this study was carried out to (a) develop standardized aqueous and ethanolic extracts of the plant mixture (N. sativa, H. indicus, and S. glabra) used in the preparation of the original decoction, and (b) compare the cytotoxic effects of these two extracts by evaluating cytotoxicity to the human hepatoma (HepG2) cell line.Methods:Aqueous and ethanolic extracts have been standardized by evaluating organoleptic characters, physicochemical properties, qualitative and quantitative analysis of chemical constituents, and analysis of High Performance Liquid Chromatography (HPLC) and Thin Layer Chromatography (TLC) profiles. Cytotoxic potentials of the above standardized extracts were compared by evaluating their effects on the survival and overall cell activity of HepG2 cells by use of the 3-(4, 5-dimethylthiazol-2yl) -2, 5 – biphenyl tetrazolium bromide (MTT) and Sulphorhodamine B (SRB) assays.Results:Results from MTT and SRB assays demonstrated that both extracts exerted strong dose-dependent in vitro cytotoxicity to HepG2 cells. The standardized aqueous extract showed a marginally (though significantly, P<0.05) higher cyotoxic potential than the ethanolic extract. Thymoquinone, an already known cytotoxic compound isolated from N. sativa seeds was only observed in the standardized ethanolic extract. Thus, compounds other than thymoquinone appear to mediate the cytotoxicity of the standardized aqueous extract of this poly-herbal preparation.Conclusion:It may be concluded that results obtained in the present study could be used as a diagnostic tool for the correct identification of these aqueous or ethanolic extracts and would be useful for the preparation of a standardized pharmaceutical product that may be used in the future for clinical therapy of hepatocellular carcinoma.

The present study investigated the potential anticancer activity of the bark of Mangifera zeylanica, an endemic plant in Sri Lanka that has been traditionally used for cancer therapy. Cytotoxic and apoptotic effects were investigated in vitro using sulphorodamine assay, acridine orange and ethidium bromide staining, caspase-3 and −7 activity, DNA fragmentation and reverse transcription-quantitative polymerase chain reaction in estrogen receptor positive MCF-7 and triple-negative MDA-MB-231 breast cancer cell lines, SKOV-3 ovarian cancer cell line and MCF-10A normal mammary epithelial cells. Hexane extract demonstrated increased levels of cytotoxicity in cancer cells (IC50, 86.6–116.5 µg/ml) compared with normal cells (IC50, 217.2 µg/ml). Chloroform extract demonstrated increased cytotoxicity to normal cells (IC50, 92.9 µg/ml) compared with cancer cells (IC50, 280.1–506.5 µg/ml). Exposure to the hexane extract led to morphological changes characteristic of apoptosis and DNA fragmentation in the three cancer cell lines. Caspase-3 and −7 were significantly activated in MDA-MB-231 and SKOV-3 cells, indicating the occurrence of caspase-dependent apoptosis in these cells, and caspase-independent apoptosis in MCF-7 cells. Furthermore, upregulation of proapoptotic Bcl-2-associated X protein occurred in the three cancer cell lines, and antiapoptotic survivin was downregulated in MCF-7 and SKOV-3 cells; by contrast, tumor protein p53 was upregulated only in MCF-7 cells, suggesting p53-mediated apoptosis in MCF-7 cells and p53-independent apoptosis in the remaining cancerous cell lines. In addition, fraction M1 obtained from bioactivity-guided fractionation of the hexane extract demonstrated increased cytotoxicity in cancer cells (IC50, 15.4–38.7 µg/ml) compared with normal cells (IC50, 114.6 µg/ml), with the highest cytotoxicity observed in MDA-MB-231 triple-negative breast cancer cells. The hexane extract of M. zeylanica bark contained polyphenols and flavonoids, and caused free radical scavenging activity. Its gas chromatography-mass spectrometry profile revealed the presence of long-chain hydrocarbons, including β-sitosterol and β-amyrin. Fraction M1 contained seven unknown compounds and a small number of known non-cytotoxic compounds. Collectively, results obtained in the present study indicate that the hexane extract of M. zeylanica bark mediates cytotoxic activities through induction of apoptosis in three cancer cell lines; thus, the hexane extract may be used to isolate novel anti-cancer compounds.

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BackgroundA standardized poly-herbal decoction of Nigella sativa seeds, Hemidesmus indicus roots and Smilax glabra rhizomes used traditionally in Sri Lanka for cancer therapy has been demonstrated previously, to have anti-hepatocarcinogenic potential. Cytotoxicity, antioxidant activity, anti-inflammatory activity, and up regulation of p53 and p21 activities are considered to be some of the possible mechanisms through which the above decoction may mediate its anti-hepatocarcinogenic action. The main aim of the present study was to determine whether apoptosis is also a major mechanism by which the decoction mediates its anti-hepatocarcinogenic action.MethodsEvaluation of apoptosis in HepG2 cells was carried out by (a) microscopic observations of cell morphology, (b) DNA fragmentation analysis, (c) activities of caspase 3 and 9, as well as by (d) analysis of the expression of pro-apoptotic (Bax) and anti-apoptotic (Bcl-2) proteins associated with cell death.ResultsThe results demonstrated that in HepG2 cells, the decoction can induce (a) DNA fragmentation and (b) characteristic morphological changes associated with apoptosis (nuclear condensation, membrane blebbing, nuclear fragmentation and apoptotic bodies). The decoction could also, in a time and dose dependent manner, up regulate the expression of the pro-apoptotic gene Bax and down regulate expression of anti-apoptotic Bcl-2 gene (as evident from RT-PCR analysis, immunohistochemistry and western blotting). Further, the decoction significantly (p < .001) enhanced the activities of caspase-3 and caspase-9 in a time and dose dependent manner.ConclusionsOverall findings provide confirmatory evidence to demonstrate that the decoction may mediate its reported anti-hepatocarcinogenic effect, at least in part, through modulation of apoptosis.

Aerial parts of Trichosanthes cucumerina (Family: Cucurbitaceae) are used in traditional medical systems for treatment of diabetes and other diseases. The present study was designed to experimentally evaluate the antidiabetic potential of a hot water extract (HWE) of T. cucumerina (TC) aerial parts. In normoglycemic rats, HWE mediated (a) a dose dependent reduction in fasting blood glucose (FBS) levels (by 35% at 4h post-treatment with dose of 750 mg/kg) and (b) a significant (P<0.05) improvement of glucose tolerance. In STZ-induced diabetic rats, no immediate hypoglycemic effect was observed. However, with continuous administration, there was a gradual reduction in FBS (by 56.8% on day 14 and by 64.4% on day 28). In normoglycemic rats, on day 14 and day 28, the percentage reduction in FBS levels were 41% and 44% respectively. At the end of 28 days, in both normoglycemic and STZ-induced diabetic rats, there was a significant increase in the levels of liver glycogen and adipose tissue triglyceride levels, in comparison with the respective controls that did not receive HWE. However, HWE failed to inhibit intestinal glucose uptake. It may be concluded that T.C can exert significant antidiabetic activity, possibly through multiple effects involving pancreatic and extra pancreatic mechanisms.