Cytotoxicity, acute and sub-chronic toxicities of the leaves of Bauhinia thonningii (Schumach.) Milne-Redh. (Caesalpiniaceae).

BACKGROUND: Bauhinia thonningii is a plant traditionally used against many human diseases such as gastric ulcers, fever, inflammations, coughs, dysentery, diarrhea, and malaria. In the present investigation, the cytotoxicity of methanol extract of Bauhinia thonningii leaves (BTL), fractions and the isolated phytoconstituents was determined in a panel of 9 human cancer cell lines including drug sensitive and multidrug-resistant (MDR) phenotypes. The acute and sub-chronic oral toxicity of BTL was investigated as well. METHODS: Compounds were isolated using chromatographic techniques while their chemical structures were determined using spectroscopic methods. The resazurin reduction assay (RRA) was used to evaluate the cytotoxicity of samples, propidium iodide (PI) for apoptosis, 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide (JC-1) staining for mitochondrial membrane potential (MMP) analysis, 2´,7´-dichlorodihydrofluoresceine diacetate (H2DCFH-DA) staining for the quantification of reactive oxygen species (ROS), whereas Caspase Glo assays were combined by means of flow cytometry. Furthermore, the toxicological investigations were performed as recommended by the Organization for Economic Cooperation and Development (OECD). RESULTS: The botanicals as well as 6-C-methylquercetin-3,7-dimethyl ether (2), quercetin-3-O-L-rhamnopyranoside (5), quercetin-3-O-ß-glucopyranoside (6), 6,8-C-dimethylkaempferol 3,7-dimethyl ether (7), and 6,8-C-dimethylkaempferol-3-methyl ether (8) had promising cytotoxic effects in the 9 tested cancer cell lines. The IC50 values below 20 µg/mL (botanicals) or 10 µM (compounds) on at least 1/9 tested cancer cell lines were considered. The best cytotoxic effects with IC50 values below 5 µM were achieved with compounds 7 against CEM/ADR5000 leukemia cells (2.86 µM) and MDA-MB-231-pcDNA breast adenocarcinoma cells (1.93 µM) as well as 8 against CCRF-CEM leukemia cells (3.03 µM), CEM/ADR5000 cells (2.42 µM), MDA-MB-231-pcDNA (2.34 µM), and HCT116 p53-/- cells (3.41 µM). BTL and compound 8 induced apoptotic cell death in CCRF-CEM cells through caspase activation, alteration of MMP, and increased ROS production. BTL did not cause any adverse effects in rats after a single administration at 5000 mg/kg or a repeated dose of 250 mg/kg body weight (b. w.). CONCLUSION: Bauhinia thonningii and its constituents are sources of cytotoxic drugs that deserve more in-depth studies to develop novel antiproliferative phytomedicine to fight cancer including resistant phenotypes.

Antibacterial and antibiotic-potentiation activities of the hydro-ethanolic extract and protoberberine alkaloids from the stem bark of Enantia chlorantha against multidrug-resistant bacteria expressing active efflux pumps.

ETHNOPHARMACOLOGICAL RELEVANCE: Enantia chlorantha is traditionally used to treat various ailments including rickettsia fever, cough and wounds, typhoid fever, infective hepatitis, jaundice, and urinary tract infections. AIM OF THE STUDY: To isolate the antibacterial constituents of the hydro-ethanolic extract of the stem bark of E. chlorantha (ECB) and to evaluate the antibacterial and antibiotic-modifying activities of ECB and its constituents against the multidrug-resistant (MDR) phenotypes. MATERIALS AND METHODS: Chromatographic methods were used to isolate the constituents of ECB and Spectroscopic methods were used to elucidate the chemical structures of the isolated compounds. The antibacterial activity of samples was determined by the broth microdilution method while spectrophotometric methods were used to evaluate the effects of ECB and its most active constituent on bacterial growth. Their effects on bacterial proton-ATPase pumps was assessed through the acidification of the bacterial culture medium. RESULTS: Six protoberberine alkaloids were isolated and identified as columbamine (1), pseudocolumbamine (2), jathrorrhizine (3), palmitine (4), 4,13-dihydroxy-3,9,10-trimethoxyprotoberberine (5), and 13-hydroxy-2,3,9,10-tetramethoxyprotoberberine (6). The crude extract (ECB) inhibited the growth of all the tested MDR bacteria, with the minimal inhibitory concentration (MIC) values below 100 µg/mL obtained against Escherichia coli ATCC 10536, AG 102, Enterobacter aerogenes EA 27, Klebsiella pneumoniae ATCC 11296 and KP 55, Providencia stuartii NEA 16, and Staphylococcus aureus MRSA3 and MRSA6. Compound 1 had the best antibacterial effects with MIC values ranging from 16 to 64 µg/mL. The efflux pump inhibitor (EPI), phenylalanine-arginine-ß naphthylamide (PAßN) significantly improved the activity of compounds 1-6. Compounds 1-3 significantly potentiated the antibacterial activity of antibiotics such norfloxacin (NOR), ciprofloxacin (CIP), and doxycycline (DOX) against the tested MDR bacteria. CONCLUSION: The crude extract (ECB) and its isolated compounds 1-6 are potential antibacterial products from Enantia chlorantha. They could be explored more to develop the antibacterial agents that could be used alone or in combination with antibiotics to overcome MDR phenotypes.