The diversity of antifungal compounds of six South African Terminalia species (Combretaceae) determined by bioautography

A bioautography method was developed to determine the number of antifungal compounds in Terminalia species extracts. Acetone, hexane, dichloromethane and methanol leaf extracts of six Terminalia species (T. prunioides, T. brachystemma, T. sericea, T. gazensis, T. mollis and T.sambesiaca) were tested against five fungal animal pathogens (Candida albicans, Cryptococcus neoformans, Aspergillus fumigatus, Microsporum canis and Sporothrix schenkii). The Rf values and relative activities of separated compounds were determined. Hexane and dichloromethane extracts had at least three times more antifungal compounds than the other extracts indicating the nonpolar character of the antifungal compounds. From the Rf values, the non-polar character of the antifungal compounds was confirmed indicating that the antifungal activity is not due to tannins. M. canis had the highest number, up to ten, of antifungal compounds. All Terminalia species contained a compound (Rf =0.46 in benzene/ethanol/ammonium hydroxide (90/10/1) active against all tested pathogens. T. sericea and T. brachystemma were the most promising candidates for isolating antifungal compounds. The results demonstrate the value of bioautography in examining plant extracts with antifungal activity,selecting species for further study and dereplicating the isolation of compounds

Bioautography indicates the multiplicity of antifungal compounds from twenty-four southern African Combretum species (Combretaceae)

Dried ground leaves of 24 Combretum spp were extracted with hexane, dichloromethane, acetone and methanol and analysed by bioautography to determine the number of antifungal compounds againstfive animal fungal pathogens (Candida albicans, Cryptococcus neoformans, Aspergillus fumigatus, Microsporum canis and Sporothrix schenckii). There was some similarity in the chemical compositionof the non-polar components of extracts using extractants of varying polarity. Acetone extracted the most antifungal compounds from Combretum spp. Combretum spp. in the section Hypocrateropsis, C.celastroides ssp. celastroides and C. clelastroides ssp. orientale had 62 different antifungal zones of inhibition compared to the 7 to 8 of C. microphyllum and C. paniculatum in the Connivetaia section. C.collinum subspecies were not active against all the tested pathogens. C. neoformans was the most sensitive organism against all Combretum species, with 367 zones of inhibition using different TLCsolvent systems and extracts. A. fumigatus was the most resistant (192 zones of inhibition). The antifungal activity and number of active antifungal compounds were high enough to consider the use ofextracts for clinical application and to isolate antifungal compounds from the extracts. Based on the Rf values of the antifungal compounds determined using solvents of varying polarity, activity is not onlybe attributable to tannins found in Combretum extracts as was previously postulated

The phytochemical, antibacterial and antioxidant activity of five medicinal plants against the wound infecting bacteria

Leaf extracts of Senna italica, Ricinus communis, Lantana camara, Lippia javanica and Ziziphus  mucronata were screened for biological activity against bacteria which infect wounds. The leaves were extracted using different solvents of varying polarity (hexane, dichloromethane, acetone and methanol). Phytochemical analyses of the extracts were performed using thin layer chromatography (TLC). The extracts were loaded on TLC plates and developed in three solvent systems that is benzene/ethanol /ammonium solution (BEA), chloroform/ethyl acetate/formic acid (CEF) and ethyl acetate/methanol /water (EMW). Antibacterial activity of the plants was evaluated using micro-dilution and bioautography methods. The test organisms used were Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Enterococcus faecalis. Acetone extracts were chosen for antioxidant activity. Methanol was the best extractant, followed by acetone and dichloromethane (DCM). In the phytochemical analysis,  more compounds were observed on BEA, followed by EMW and CEF plates. Lantana camara had no  activity against any of the bacteria used. P. aeruginosa was the most resistant bacterium with only two plants active against it. E. faecalis and E. coli were sensitive to the extracts. More antibacterial  compounds were observed on BEA plates against all the test bacteria in bioautographic method. The Rf  values calculated from bioautography indicated that the selected plants have different active compounds. The most active compounds were from S. italica and Z. mucronata. BEA and EMW plates had good  antioxidant activity. No antioxidant activity was observed on the CEF plate. Most extracts were active against wound pathogens; their application on the wound area may prevent infection. Further studies are required to identify the active compounds in the plant extracts which showed significant anti-bacterial activities.Key words: Thin layer chromatography (TLC), plant extract, bacteria

Does seasonal variation influence the phytochemical and antibacterial properties of Carpobrotus edulis?

Carpobrotus edulis L. (family Aizoaceae) has been used by locals over the years to treat microbial infections. Extracts of varying polarities were prepared from the leaf debris and filtrate of a spring andan autumn harvest of C. edulis. Thin layer chromatography was used to analyze the phytocompounds of the extracts as well as to assay the plant for antioxidant compounds. The spring harvest showedequal distribution of the phytochemicals within the leaf debris and the filtrate, but there was a high prevalence of phytocompounds within the leaf debris extracts of the autumn sample. An antioxidantcompound was intensely pronounced in the autumn extracts of intermediate polarity and in the polar extract. The plant was evaluated for antibacterial activity against Escherichia coli, Enterococcusfaecalis, Pseudomonas aeruginosa and Staphylococcus aureus by using a two-fold serial microdilution method as well as bioautography. The spring extracts were more potent against all test organisms,having MIC values that were lower than the autumn extracts. When taking the total activity of each extract into account, the autumn extracts showed higher efficacy than the extracts from the springsample. The antibacterial activity observed in the extracts of both seasons somewhat validated the ethnomedicinal use of C. edulis

Characterisation of a chimeric Phanerochaete chrysosporium cellobiohydrolase expressed from Escherichia coli

The aim of this study was to purify and analyse a Phanerochaete chrysosporium cbhI.1 gene-product expressed as an inducible, secreted, heterologous protein from an Escerichia coli pGEXcbhI.1 clone. Using glutathione Sepharose 4B affinity chromatography, the expressed protein was purified from the supernatant of an induced E. coli transformed with pGEXcbhI.1 and ran as a single band on a Sodium dodecyl sulphate-polyacrylamide gel. The glutathione S-transferase (GST) fused CBHI.1 was approx-imately 80 kDa in size, approximately 2.2 kDa smaller than the theoretically predicted size. The purified protein exhibited time dependent hydrolytic reaction against carboxy-methyl-cellulose (CMC) and Avicel. On CMC the highest hydrolytic reaction occurred at 120 min. whereas for Avicel it was at 150 min. Optimum pH and temperature for activity of the protein against these cellulose substrates were pH 6 and 55oC, respectively, and the protein remained stable under these optimum conditions for 24 h. Key Words: Phanerochaete chrysosporium, cellobiohydrolase purification, heterologus expression. African Journal of Biotechnology Vol.3(7) 2004: 349-35

In vitro evaluation of the antifungal activity of Sclerocarya birrea extracts against pathogenic yeasts

The antifungal activity of Sclerocarya birrea which is used in South African traditional medicine for the treatment of skin diseases was evaluated against three yeasts; Candida parapsilosis, Cryptococcusalbidus and Rhodoturula mucilaginosa. Barks of S. birrea were extracted with hexane, dichloromethane (DCM), chloroform, ethyl acetate, acetone, methanol and ethanol and tested against these three yeasts.The antifungal assay was performed by the microdilution technique and bioautography. Thin layer chromatography was used to analyze the phytocompounds of the extracts as well as to assay the plantfor antioxidant compounds. More compounds with antioxidant activity were observed in polar separation system, ethyl  cetate:methanol:water (EMW). All test organisms were resistant against all non-polar extracts. Acetone, ethanol and methanol S. birrea extracts had average MIC values of 0.39, 0.22 and 0.27 mg/ml, respectively. C. albidus was the most sensitive organism with an average MIC value of 0.17 mg/ml. Average total activity was highest for methanol (387 ml/g) followed by ethanol (363 ml/g) and acetone (299 ml/g) bark extracts. Acetone and methanolic bark extracts were more active in EMW system at Rf values of 0.07, 0.32 and 0.70 against C. parapsilosis. The results showed that the plant could be further explored for possible antifungal agents and provides preliminary scientific validation of the traditional medicinal use of this plant

Antibacterial, Anti-HIV-1 Protease and Cytotoxic Activities of Aqueous Ethanolic Extracts from Combretum Adenogonium Steud. Ex A. Rich (Combretaceae).

\ud \ud Records have shown that Combretum adenogonium Steud. Ex A. Rich (Combretaceae) is used in traditional medicine systems of several tribes in Tanzania. This study focused on the investigation of antibacterial activity, anti-HIV-1 protease activity, toxicity properties and classes of phytochemicals in extracts from C. adenogonium Steud. Ex A. Rich (Combretaceae) to evaluate potential of these extracts for development as herbal remedies. Dried plant material were ground to fine powder and extracted using 80% aqueous ethanol to afford root, leaf and stem bark extracts. The extracts were assayed for anti-HIV-1 protease activities, antibacterial activities using microdilution methods and cytotoxicity using brine shrimps lethality assay. Screening for major phytochemical classes was carried out using standard chemical tests. All extracts exhibited antibacterial activity to at least one of the test bacteria with MIC-values ranging from 0.31-5.0 mg/ml. Two extracts, namely, root and stem bark exhibited anti-HIV-1 PR activity with IC50 values of 24.7 and 26.5 μg/ml, respectively. Stem bark and leaf extracts showed mild toxicity with LC50 values of 65.768 μg/ml and 76.965 μg/ml, respectively, whereas roots were relatively non-toxic (LC50 = 110.042 μg/ml). Phytochemical screening of the extracts indicated presence of flavonoids, terpenoids, alkaloids, tannins, glycosides and saponins. These results provide promising baseline information for the potential development of C. adenogonium extracts in treatment of bacterial and HIV/AIDS-related opportunistic infections

Characterization of n-Hexane sub-fraction of Bridelia micrantha (Berth) and its antimycobacterium activity

<p>Abstract</p> <p>Background</p> <p>Tuberculosis, caused by <it>Mycobacterium tuberculosis </it>(MTB), is the most notified disease in the world. Development of resistance to first line drugs by MTB is a public health concern. As a result, there is the search for new and novel sources of antimycobacterial drugs for example from medicinal plants. In this study we determined the <it>in vitro </it>antimycobacterial activity of <it>n</it>-Hexane sub-fraction from <it>Bridelia micrantha </it>(Berth) against MTB H₃₇Ra and a clinical isolate resistant to all five first-line antituberculosis drugs.</p> <p>Methods</p> <p>The antimycobacterial activity of the <it>n</it>-Hexane sub-fraction of ethyl acetate fractions from acetone extracts of <it>B. micrantha </it>barks was evaluated using the resazurin microplate assay against two MTB isolates. Bioassay-guided fractionation of the ethyl acetate fraction was performed using 100% <it>n</it>-Hexane and Chloroform/Methanol (99:1) as solvents in order of increasing polarity by column chromatography and Resazurin microtiter plate assay for susceptibility tests.</p> <p>Results</p> <p>The <it>n</it>-Hexane fraction showed 20% inhibition of MTB H₃₇Ra and almost 35% inhibition of an MTB isolate resistant to all first-line drugs at 10 μg/mL. GC/MS analysis of the fraction resulted in the identification of twenty-four constituents representing 60.5% of the fraction. Some of the 24 compounds detected included Benzene, 1.3-bis (3-phenoxyphenoxy (13.51%), 2-pinen-4-one (10.03%), N(b)-benzyl-14-(carboxymethyl) (6.35%) and the least detected compound was linalool (0.2%).</p> <p>Conclusions</p> <p>The results show that the <it>n-</it>Hexane fraction of <it>B. micrantha </it>has antimycobacterial activity.</p

Pentachlorophenol Induction of the Pseudomonas aeruginosa mexAB-oprM Efflux Operon: Involvement of Repressors NalC and MexR and the Antirepressor ArmR

Pentachlorophenol (PCP) induced expression of the NalC repressor-regulated PA3720-armR operon and the MexR repressor-controlled mexAB-oprM multidrug efflux operon of Pseudomonas aeruginosa. PCP's induction of PA3720-armR resulted from its direct modulation of NalC, the repressor's binding to PA3720-armR promoter-containing DNA as seen in electromobility shift assays (EMSAs) being obviated in the presence of this agent. The NalC binding site was localized to an inverted repeat (IR) sequence upstream of PA3720-armR and overlapping a promoter region whose transcription start site was mapped. While modulation of MexR by the ArmR anti-repressor explains the upregulation of mexAB-oprM in nalC mutants hyperexpressing PA3720-armR, the induction of mexAB-oprM expression by PCP is not wholly explainable by PCP induction of PA3720-armR and subsequent ArmR modulation of MexR, inasmuch as armR deletion mutants still showed PCP-inducible mexAB-oprM expression. PCP failed, however, to induce mexAB-oprM in a mexR deletion strain, indicating that MexR was required for this, although PCP did not modulate MexR binding to mexAB-oprM promoter-containing DNA in vitro. One possibility is that MexR responds to PCP-generated in vivo effector molecules in controlling mexAB-oprM expression in response to PCP. PCP is an unlikely effector and substrate for NalC and MexAB-OprM - its impact on NalC binding to the PA3720-armR promoter DNA occurred only at high µM levels - suggesting that it mimics an intended phenolic effector/substrate(s). In this regard, plants are an abundant source of phenolic antimicrobial compounds and, so, MexAB-OprM may function to protect P. aeruginosa from plant antimicrobials that it encounters in nature