M. P. Mokgotho

Isolation and Chemical Structural Characterisation of a Compound with Antioxidant Activity from the Roots of Senna italica

Senna italica, a member of the Fabaceae family (subfamily Caesalpiniaceae), is widely used in South African traditional medicine to treat a number of disease conditions. Aqueous extracts of the plant are mainly used to treat sexually transmitted infections and intestinal complications. The roots of S. italica were ground to a fine powder and sequentially extracted with n-hexane, dichloromethane, acetone, and methanol using serial exhaustive extraction (SEE) method. Thin layer chromatography was used to analyse the phytochemical composition of the extracts and DPPH radical scavenging method to detect the presence of antioxidant compounds. The bioassay guided fractionation of the acetone fraction afforded an antioxidant compound with free radical scavenging activity. The isolated compound was subsequently identified as 3,4′,5-trihydroxystilbene (resveratrol). This study represents the first report of the stilbene resveratrol in S. italica.

Antibacterial and Antimetastatic Potential of Diospyros lycioides Extract on Cervical Cancer Cells and Associated Pathogens

Cervical cancer is among the most prevalent forms of cancer in women worldwide. Diospyros lycioides was extracted using hexane, ethyl acetate, acetone, and methanol and finger print profiles were determined. The leaf material was tested for the presence of flavonoids, tannins, saponins, terpenoids, and cardiac glycosides using standard chemical methods and the presence of flavonoids and phenolics using thin layer chromatography. The total phenolic content was determined using Folin-Ciocalteu procedure. The four extracts were tested for antibacterial activity using bioautography against Staphylococcus aureus, Enterococcus faecalis, Pseudomonas aeruginosa, and Escherichia coli. The acetone extract with the highest number of antibacterial and antioxidant compounds was assessed for its cytotoxicity on BUD-8 cells using the real-time xCELLigence system and its potential effects on metastatic cervical cancer (HeLa) cell migration and invasion were assessed using wound healing migration and invasion assays. The leaf extract tested positive for flavonoids, tannins, and terpenoids while the four different extracts tested in the antimicrobial assay contained constituents active against one or more of the organisms tested, except E. coli. The cytotoxicity of the acetone extract in real-time was concentration-dependent with potent ability to suppress the migration and invasion of HeLa cells. The finding demonstrates the acetone extract to contain constituents with antibacterial and antimetastatic effects on cervical cancer cells.

Free Radical Scavenging Activity: Antiproliferative and Proteomics Analyses of the Differential Expression of Apoptotic Proteins in MCF-7 Cells Treated with Acetone Leaf Extract of Diospyros lycioides (Ebenaceae).

Breast cancer is the most common cancer in South Africa. The acetone leaf extract of Diospyros lycioides was evaluated qualitatively and quantitatively for its antioxidant potential using DPPH assay and nitric oxide radical scavenging effect, while the viability of MCF-7 cells was evaluated using the MTT. MCF-7 treated cells were stained with Hoechst 335258 dye and annexin-V-FITC to be evaluated for apoptotic effect of the extract, while mRNA expression levels of apoptotic genes were assessed by quantitative real-time PCR and deferential protein expression levels using 2D gel electrophoresis and mass spectrometry. Results revealed presence of antioxidant constituents in the extract. Extract was shown to be cytotoxic in a concentration- and time-dependent manner. Cytotoxicity was demonstrated to be due to apoptosis, with 70% of the extract-treated cells being annexin-V-positive/PI negative at 48 hours. The extract was also shown to upregulate the expression of p53 gene with concomitant downregulation of the Bcl-2 antiapoptotic gene while differentially expressed proteins were identified as enolase, pyruvate kinase, and glyceraldehyde-3-phosphate. The extract in this study was shown to induce apoptosis at an early stage which makes it an ideal source that can be explored for compounds that may be used in the treatment and management of cancer.

Potential Hypoglycaemic and Antiobesity Effects of Senna italica Leaf Acetone Extract

Background Type II diabetes is on the rise while obesity is one of the strongest risk factors of type II diabetes. The search for a drug for type II that can equally mitigate obesity related complication is desired. Methods The acetone leaf extract of Senna italica was evaluated for its cytotoxic, antiglycation, and lipolytic effect, glucose uptake, and GLUT4 translocation and expression using published methods, while that for adipogenesis and protein expression levels of obesity related adipokines was assessed using adipogenesis assay and mouse adipokine proteome profiler kit, respectively. The possible mechanism of glucose uptake was assessed through the inhibition of PI3K pathway. Results The extract had no adverse effect on 3T3-L1 cell viability (CC50 > 1000 μg/ml). High antiglycation effect was attained at 10 mg/ml, while at 25–200 μg/ml it showed no significant increase in adipogenesis and lipolysis. The extract at 100 μg/ml was shown to decrease the expression levels of various adipokines and minimal glucose uptake at 50–100 μg/ml with a nonsignificant antagonistic effect when used in combination with insulin. GLUT4 translocation and expression were attained at 50–100 μg/ml with an increase in GLUT4 expression when in combination with insulin. Conclusion The acetone leaf extract of S. italica stimulates glucose uptake through the PI3K-dependent pathway and can serve as a source of therapeutic agent for the downregulation of obesity-associated adipokines in obesity and antiglycation agents.

Antioxidant and Anti-Proliferative Capacity of a Dichloromethane Extract of Dicerocaryum senecioides Leaves

Leseilane J. Mampuru1,*, Pirwana K. Chokoe1, Maphuti C. Madiga1, Annette Theron2, Ronald Anderson2 and Matlou P. Mokgotho1 1Department of Biochemistry, Microbiology and Biotechnology, Faculty of Science and Agriculture, University of Limpopo, Sovenga, 2Medical Research Council Unit for Inflammation and Immunity, Department of Immunology, Faculty of Health Sciences, University of Pretoria and Tshwane Academic Division of the NHLS, Pretoria, South Africa

Dichloromethane extract of Dicerocaryum senecioides leaves exhibits remarkable anti-inflammatory activity in human T-lymphocytes

The leaves of Dicerocaryum senecioides are used in South Africa as a traditional remedy for many ailments, including inflammatory disorders. The purpose of this study was therefore to evaluate the anti-inflammatory potential of a dichloromethane extract of D. senecioides leaves. Methanol extracts of the leaves were sub-fractionated with dichloromethane and the anti-inflammatory potential of this fraction investigated according to its effects on the mitogen-induced proliferative responses and cytokine profiles of isolated human blood mononuclear leucocytes (MNL). The cells were pre-treated with the extract (25–100 µg mL−1) followed by addition of the mitogen, phytohaemagglutinin (PHA, 5 µg mL−1 final), and measurement of lymphocyte activation and proliferation, using flow cytometric detection of up-regulation of expression of CD25 and incorporation of radiolabelled thymidine into newly synthesised DNA, respectively. Cytokine production by unstimulated and PHA-activated cells was measured using multiplex suspension bead array technology. Treatment of cells with the Dicerocaryum extract resulted in dose-related inhibition of PHA-activated lymphocyte proliferation and expression of CD25, as well as decreased production of Th1 (IFN-γ, TNF-α) and Th2 (IL-10) cytokines. These observations not only underscore the anti-inflammatory potential of components of Dicerocaryum leaves, but also provide a basis for future definitive studies.

Antioxidant, Antiglycation, and Hypoglycaemic Effect of Seriphium plumosum Crude Plant Extracts

Diabetes is a severely debilitating metabolic disorder characterised by chronic hyperglycaemia. Traditional medicinal plants provide an important avenue for the development of novel antidiabetic agents. The antidiabetic potential of the methanol, acetone, and hexane extracts of S. plumosum was assessed using different parameters. These included secondary metabolite quantification, hypoglycaemic, cytotoxic effects, and GLUT4 translocation augmentation on C2C12 cells. The methanol extract contained the highest amount of total phenolic and flavonoid compounds and showed enhanced antioxidant activity. The methanol extracts had the best DPPH scavenging (EC50 = 0.72 mg/ml) and ferric reducing powers (EC50 = 2.31 mg/ml). The hexane extract resulted in the highest glucose uptake activity of 35, 77% with respect to all other treatments after a 6-hour exposure period. Immunocytochemistry technique further revealed that the increased glucose utilisation may be due to increased membrane fused GLUT4 molecules in C2C12 cells. The hexane extract was also shown to upregulate the phosphorylation of p70 S6 kinase and Akt1/2. The study highlights a probable insulin-mimetic activity of the hexane extract via the augmentation of Akt1/2 phosphorylation which is involved in the GLUT4 translocation pathway. Furthermore, the study represents the first report on the cytotoxic effect, GLUT4 translocation, and glucose uptake potential of S. plumosum.

Semi-purified extracts of Commelina benghalensis (Commelinaceae) induce apoptosis and cell cycle arrest in Jurkat-T cells

BackgroundCommelina benghalensis (CB) is a small plant whose fleshy stems are used in South Africa to treat skin conditions (e.g., cancerous skin outgrowths). This study was aimed at evaluating the effect of sub-fractions of acetone extracts of CB stems on growth-associated molecular events of apoptosis and cell division cycle of Jurkat-T (JT) cells.MethodsAcetone extract of CB stems were subfractioned into n-hexane (F1) and dichloromethane (F2) fractions. After treatment of JT cells with these subfractions, cell proliferation, viability and apoptosis were determined using a haemocytometer, the trypan blue dye exclusion assay, and Hoechst 33258 staining, respectively. Cell division cycle distribution profiles were analysed using an Epics Alba Flow Cytometer and the expression of cell division cycle regulatory genes was analysed using RT-PCR, while immunoreactive proteins were detected on western blots.ResultsThe F1 and F2 fractions inhibited the proliferation and viability of JT cells in a concentration- and time-dependent manner, with IC50 values of 32.5 μg/mℓ and 56 μg/mℓ, respectively. The observed cytotoxicity was established to be a consequence of apoptosis. as verified using Hoechst staining method. Both fractions induced a G1/S interphase arrest of the cell division cycle of JT cells.RT-PCR analyses showed an up-regulatory effect by the F1 fraction in the expression of cyclin B1, cdc2 and bax, with a down-regulatory effect in the expression levels of bcl-2. Fraction F1 also increased the protein expression levels of p53 and its downstream regulators, p21 and Cdc2. However, protein Bax and p21 and p53 transcripts were undetectable under the same experimental conditions. On the other hand, fraction F2 increased the mRNA expression levels of bax, bcl-2, cyclin B1 and cdc2. Concomitantly, fraction F2 showed an up-regulation in the protein expression levels of Cdc2, Bcl-2, Cyclin B1 and p21. Despite the up-regulation in protein expression levels by fraction F2, there was no observable expression levels of the p53 protein and p21 and p53 mRNAs under similar experimental conditions.ConclusionThese findings suggest that the F1 and F2 fractions of CB may provide a valuable lead for the development of novel and effective anti-neoplastic drug(s).