Bioautography Method Research Articles

In Vitro Assessment of Selected Medicinal Plants Used by the Bapedi Community in South Africa for Treatment of Bacterial Infections.

There is very little scientific information on indigenous plants used for medicinal purposes. Therefore, the aim of the study was to evaluate the antibacterial and antioxidant activities of the leaves of Commelina africana, Elephantorrhiza elephantina, Dombeya rotundifolia, and the whole plant excluding the roots of Schkuhria pinnata indigenous medicinal plants from the Limpopo Province, which may be used for the treatment in humans infected with bacterial pathogens. The screening of different phytoconstituents using standard methods revealed the presence of terpenoids, flavonoids, tannins, and saponins. The total phenolic, tannin, and flavonoid content were estimated using spectrophotometric methods. D rotundifolia had the highest amounts of phenol (259.00 ± 2.65 mg of gallic acid equivalent [GAE]/g), tannin (330.33 ± 15.63 mg of GAE/g), and flavonoid (19.90 ± 0.75 mg of quercetin equivalent [QE]/g) content, while S pinnata had the least. All the plants had antibacterial activity against Escherichia coli and Pseudomonas aeruginosa with the bioautography method. The microbroth dilution assay revealed antibacterial activity with minimum inhibitory concentration values raining from 0.02 to 2.50 mg/mL. D rotundifolia had the highest antioxidant activity in both the free radical scavenging and ferric reducing power methods. S pinnata and C africana extracts had high antibacterial activity, while D rotundifolia had high antioxidant activity, which made them potential plants for isolation of bioactive compounds and possible application in the pharmaceutical industry.

HPTLC – Bioautographic methods for selective detection of the antioxidant and a-amylase inhibitory activity in plant extracts

A high-performance thin-layer chromatography (HPTLC) method was developed for quantification of a-amylase inhibitory activity and stigmasterol content in ant plant extracts. An improved HPTLC method for the determination of total free radical scavenging activity in samples using DPPH is also reported. For quantification of a-amylase inhibitory activity, the developed HPTLC plate is dipped into an a-amylase solution, and the bioautogram is then incubated at 25?°C for 30?min under humid conditions. For visualization of enzyme inhibitory activity, the starch test with an iodine indicator solution is used. The blue zone observed comes from the starch-iodine complex formed from starch that was not hydrolyzed by the amylase due to enzyme inhibition by the compound(s) present in the sample. The area of the blue zones was used to compare and quantify relative a-amylase inhibitory activity in different extracts. Location of the blue zones (hRF) on the plate was used to detect compounds that are responsible for the a-amylase inhibitory activity. Relative a-amylase activity was not related to the antioxidant activity, but was highly correlated with the stigmasterol content in the sample extracts (R?=?0.95). Therefore, plant sterols present in the extracts might be responsible for a-amylase inhibitory activities in the extracts.•The developed method for quantification of a-amylase inhibitory activity provides an efficient and effective tool that can be used to screen, detect and quantify a-amylase inhibitory activity in plant extracts.•The proposed protocol is easy to run, involves minimal sample preparation, with multiple samples able to be analyzed in parallel on the same chromatographic plate, in a short time.•There were significant differences in a-amylase inhibitory activity, stigmasterol content, and total free radical scavenging activity between methanol, ethanol, dichloromethane, and ethyl acetate ant plant extracts.

<i>Sutherlandia frutescens</i> (Fabaceae) extracts used for treating tuberculosis do not have high activity against <i>Mycobacterium smegmatis</i>

Sutherlandia frutescens (L) R. Br. contains several essential, bioactive compounds with clinically proven pharmacological activities. Sutherlandia is prescribed for people with tuberculosis but it is still not known what compounds in this plant act against Mycobacterium tuberculosis and its mode of action. This study is aimed at determining if S. frutescens extracts contain antimycobacterial compounds. Aerial parts of S. frutescens were dried, ground and extracted with ethanol, dichloromethane: methanol 1:1 (v/v) and water. The chemical profiling was done using high-performance liquid chromatography-mass spectroscopy (HPLC-MS) and thin layer chromatography (TLC). TLC plates were developed in butanol:acetic acid:water (BAW) to the ratio of 21:6:3; chloroform:methanol:water:formic acid (CMWF1) [60:15:2:1] and (CMWF2) [21:9:1:0.3]. Qualitative antioxidant activity was done, using 2.2-diphenylpacryl-1-hydrazyl (DPPH). Antimycobacterial activity of the plant extracts was evaluated, using micro-dilution and bioautographic methods against Mycobacterium smegmatis. Low antimycobacterial activity against M. smegmatis was observed on the bioautograms. The ethanol extracts contained more compounds compared to water extracts on HPLC-MS chromatographic profiles. The average Minimum Inhibitory Concentration (MIC) values for all the extracts were 0.61 mg/mL units and the DCM:MeOH (1:1) extract had the lowest MIC value of 0.28 mg/mL. The results showed that the plant could be further explored for possible antimycobacterial agents. Low activity was observed, possibly due to low replication of bacilli and non-replicating organisms. The study provides preliminary scientific validation of the traditional medicinal use of this plant. Further studies are required to identify the bioactive compounds in the DCM:MeOH 1:1 extract which showed significant antimycobacterial activities. Research correlation: This article is the original version, of which an Afrikaans translation was made available to provide access to a larger readership, available here: https://doi.org/10.4102/satnt.v36i1.1494

High-performance thin-layer chromatographic methods in the evaluation of the antioxidant and anti-hyperglycemic activity of Myrmecodia platytyrea as a promising opportunity in diabetes treatment

The steady increase of diabetes is becoming a major burden on health care systems. As diabetic complications arise from oxidative stress, an antioxidant therapy along with anti-diabetic drugs is recommended. Myrmecodia or ant plant is highly valued as a traditional medicine in West Papua. It is used as an alternative treatment for diabetes, as the substances produced by ants can reduce blood sugar levels. The aim of this study was to develop and establish high-performance thin-layer chromatographic (HPTLC)-bioautographic methods to measure the antioxidant and hypoglycemic effects in different extracts from Myrmecodia platytyrea and to compare them with sterol content. Antioxidant activity in methanol, ethanol, dichloromethane (DCM) and ethyl acetate (EA) extracts were measured with a direct HPTLC-2,2-diphenyl-1-picrylhydrazyl free radical (DPPH) assay, while hypoglycemic effects were assessed using a newly developed α-amylase inhibitory activity assay. Stigmasterol is observed, after derivatization with anisaldehyde, as purple colored zones under visible light at hRF values of 0.66. The highest antioxidant activity was observed in the ethanol extract which is rich in polyphenols and flavonoids, while the DCM extract did not show antioxidant activity, but had significant α-amylase inhibitory activity. The highest α-amylase inhibitory activity was observed in the EA and DCM extracts and was related to their stigmasterol content.

Indole-3-Acetic Acid Production by Colletotrichum siamense, An Endophytic Fungus from Piper nigrum Leaves

Plant endophytic fungi have been recognized as an important and a good source of natural bioactive products with potential application in agriculture, medicine and food industry. Chemical and biological studies of fungal metabolites originating from endophytic fungi isolated in Sri Lanka are relatively less studied. Therefore we investigated the chemistry and bioactivity of endophytic fungi present in leaves of Piper nigrum L. (black pepper) of the family Piperaceae famed as the spices king due to its pungent quality. A fungal endophyte isolated from the leaves of P. nigrum was identified as Colletotrichum siamense through molecular biological means using internal transcribed spacer (ITS) region of rDNA gene. This is the first report of the isolation of C. siamense from Piperaceae. The fungus was fermented in potato dextrose media and the fungal media were extracted with EtOAc. Chromatographic separation of the EtOAc extract over silica gel, Sephadex LH-20 and preparative thin layer chromatography furnished indole-3-acetic acid (IAA) and uracil. IAA showed high antifungal activity against the common plant pathogenic fungus Cladosporium cladosporiodies by TLC bioautography method and antioxidant activity against DPPH radical by spectrophotometry method.

Inhibitory activity of bio-active compounds isolated from Anadara granosa in shrimp health management.

The crude extract isolated from the visceral mass of Anadara granosa, an intertidal bivalve mollusc was tested for inhibitory activity against pathogenic bacteria of the shrimp and fish viz. Vibrio harveyi and Staphylococcus aureus respectively by agar well diffusion and contact bioautography methods. Maximum inhibitory activity was shown against V. harveyi by methanol and chloroform (9:1) extract. Twelve fractions (1-12) could be separated from the crude extract through column chromatography. Five out of twelve fractions (7, 8, 9, 10, and 11) showed antibacterial activity and they were further run on column chromatography for purity. The fraction no. 9 showed highest antibacterial activity among the five and was subjected to NMR for the proton, C13 and H1-H1 correlation, IR and mass spectral analysis for structural elucidation. Structure of the compound isolated from fraction no: 9 was determined as 1-(((2Z, 4Z)-dodeca-2,4-dienoyl)oxy)-3-hydroxypropan-2-yl tetradecanoate.

Antibacterial and antibiofilm activity of Ammi majus seed against Gram-positive bacteria

Background and objective: Ammi majus belongs to family Apiaceae is an important plant used in a different country traditionally for treatment bacterial infection. This study aimed to investigate in vitro antibacterial and antibiofilm activity of Ammi majus seed extracts against six isolated Gram-positive bacteria. Methods: The antibacterial activity of seed extracts were screened by disk diffusion and agar overlay bioautography method and their activities were further determined by minimum inhibitory and minimum bactericidal concentration. Biofilm formation was evaluated by the microtiter plate crystal violet assay. The quantity of bound bacteria was determined by measuring absorbance at (OD630 nm) using microtiter plate reader. Results: Ethanol 80% and ethyl acetate extracts showed variable activity against all isolated bacteria while petroleum ether extract revealed resistance against studied bacteria. Streptococcus mitis showed more pronounced sensitivity against both extracts by disk diffusion (12 mm and 10 mm) and bioautography method with minimum inhibitory concentration 7.8125 mg/ml of ethanol 80% and 15.625 mg/ml for ethyl acetate extracts. Ethanol 80% and ethyl acetate extract significantly reduced biofilm formation as compared to control, showed antibiofilm activity at 0.4883-62.5 mg/ml and exhibited strongest antibiofim activity against Staphylococcus aureus. Conclusion: Ammi majus seed extracts revealed highest antibacterial activity against Streptococcus species and strongest antibiofilm activity on Staphylococcus species.

A TLC-Direct Bioautography Method for Detection of Antiurolithiatic Metabolites.

Hyperoxaluria is major urinary disorder troubling largest population throughout the world predominantly involving calcium oxalate (CaOx) crystals. Ancient Ayurvedic system of medicine in India claims better option in treatment of urolithiasis. A plant from "Pashanbheda" group is Phyllanthus niruri L., possessing antiurolithiatic activity, needed to be screened and validated. In the present study, a rapid, easy and efficient method forCaOx crystal inhibition in the agar gel system analogous to antimicrobial well diffusion assay is proposed. A novel thin-layer chromatography (TLC)-direct bioautography method was also proposed to detect the antilithiatic metabolites. It helps to localize the active metabolites in P. niruri, further the partial structure elucidation was characterized by High Resolution Liquid Chromatography by mass spectroscopy (LC-HRMS) analysis. The agar well diffusion method shows 50% inhibitory concentration (IC50) value at 228.55 and 493.529 mg/mL for tri-sodium citrate and P. niruri extract, respectively. The lowest concentration showing visible crystal inhibition (minimum inhibitory concentration, MIC) in both samples was found to be 20 mg/mL. In this study, a unique agar gel well diffusion and TLC-direct bioautography method successfully screened, detected and confirmed CaOx crystal inhibitory metabolites from P. niruri. The tuberonic acid was detected in bioactive fraction of P. niruri by LC-HRMS characterization.

DIVERSITY OF ANTIBACTERIAL COMPOUNDS FROM EUCHEUMA SERRA, HALIMEDA OPUNTIA, AND HYDROCLATHRUS CLATHRATUS

Fourteen compounds were isolated from acetone extracts of three species of seaweeds (Eucheuma serra, a red seaweed, Halimeda opuntia, a green seaweed, and Hydroclathrus clathratus, a brown seaweed) using bioautographic TLC methods and identified using GC-MS. From Eucheuma serra were isolated 8 compounds (3 fatty acids, 3 steroids, and 2 aldehyds). Only two compounds of fatty acid camefrom Halimeda opuntia, whereas Hydroclathrus clathratus produced 6 compounds (4 fatty acids, one compound each of steroid and ether). All isolated single compoundswere tested for their antibacterial activities by the agar diffusion method against the Gram-positive bacteria Staphylococcus aureus, Bacillus subtilis and Streptococcusfaecalis, and the Gram-negative bacteria Echerichia coli, Pseudomonas aeruginosa and Salmonella typhimurium. All 14 compounds showed activity against Gram-positivebacteria, especially Bacillus subtilis, and only 2 compounds showed activity against Gram-negative bacteria Escherichia coli. Nine compounds showed activity against Staphylococcus aureus, and 4 compounds showed activity against Streptococcusfaecalis. All compounds were not active against Pseudomonas aeruginosa andSalmonella typhimurium bacteria. This study indicated that there is indeed a diversityboth in kinds and in molecular structures of the antibacterial substances.

Antimicrobial, anti-inflammatory activity and cytotoxicity of Funtumia africana leaf extracts, fractions and the isolated methyl ursolate

Funtumia africana is used to treat and manage diverse ailments including fever, inflammation, malaria, cancer and urinary incontinence in South Africa. In this study, the antibacterial, antifungal, anti-inflammatory activities and cytotoxicity of the crude extracts, fractions and an isolated compound were determined. Serial microplate dilution and bioautography methods were used to determine the antimicrobial activities. The bacteria tested were ATCC reference strains of Escherichia coli, Enterococcus faecalis, Pseudomonas aeruginosa and Staphylococcus aureus. The fungal test organisms used were three clinical isolates (Aspergillus fumigatus, Cryptococcus neoformans and Candida albicans), C. albicans ATCC 10231 and three phytopathogenic fungi (Fusarium oxysporum, Penicillium janthinellum and Rhizoctonia solani). The anti-inflammatory activity was determined using cyclooxygenase (COX) enzymes and the MTT assay was used to determine cellular toxicity against Vero and human liver (C3A) cells. The crude extract had MICs as low as 80μg/ml against both bacteria and fungi. The chloroform fraction had the lowest MIC of 20μg/ml against P. aeruginosa. The hexane and chloroform fractions had MIC of 40μg/ml against C. albicans ATCC 10231. The crude extract, hexane and chloroform fractions had moderate activity against both COX-1 and COX-2. The chloroform fraction was more active than the crude extract (59.7%) with an inhibition of 68.2% against COX-1. Bioassay-guided fractionation using column chromatography led to the isolation of methyl ursolate (MU) with an MIC of 62.5μg/ml against F. oxysporum. It was relatively toxic against Vero cells with an IC50 of 10.4μg/ml. The antimicrobial and anti-inflammatory activities of the crude extract provide some support for the traditional use of the plant.

Antimicrobial Activity of Fractions of Ethyl Acetate Extract of Cladosporium oxysporum, An Endophytic Fungus Derived from Alyxia reinwardtii

The endophytic fungi Cladosporium oxysporum was isolated from medicinal plants Alyxia reinwardtii. The antimicrobial activity of fractions of ethyl acetate extract and also antioxidant and antimicrobial activities of fraction 7 of ethyl acetate extract of (AR-7) were determined in this study. Antimicrobial activity was determined by TLC bioautography and disc diffusion methods. Antioxidant activity was determined by TLC-DPPH autography assay. The fractions of ethyl acetate extract of C. oxysporum at dose of 100 mg/ml were active as antimicrobial against S. aureus, E. coli and C. albicans in disc diffusion method test. Fraction AR-7 was active as antioxidant in TLC-DPPH autography assay. Fraction AR-7 was active as antimicrobial against S. aureus, but was not active against another tested microoorganisms in TLC-bioautography

Olfactometry evaluation and antimicrobial analysis of essential oils from Baccharis dentata (Vell.) G.M. Barroso and Baccharis uncinella DC.

The species of genus Baccharis spp. (Compositae) are an important natural resource in the search for new bioactive compounds. This article presents an investigation of volatile compounds of two species of Baccharis that are native to southern Brazil (Baccharis dentata (Vell.) G.M. Barroso and Baccharis uncinella DC) collected in summer and autumn of 2011. The essential oils were extracted by steam distillation and analyzed by gas chromatography-mass spectrometry (GC-MS), olfactometry analysis (GC-O) and qualitative assessment of the antimicrobial potential using the indirect bioautography method against Staphylococcus aureus, Salmonella enterica subspecie enterica serovar Choleraesuis, Pseudomonas aeruginosa, Candida albicans, Aspergillus niger, Candida parapsilosis, Cryptococcus neoformans, Rhizopus stolonifer, Fusarium solani and Aspergillus oryzae. The results demonstrated that the harvest period has an influence on the composition of the essential oils, but little influence on extraction yield. It can also be observed that the joint analysis of GC-MS and GC-O resulted in complementary information about the essential oils studied because the intensities of volatile compounds detected by both methods are significantly different. Regarding antimicrobial assays, it was found that the essential oil of B. dentata was effective against S. aureus, A. niger, C. parapsilosis and R. stolonifer, whereas the essential oil of B. uncinella was effective against S. aureus, S. Choleraesuis, C. albicans, A. niger, C. parapsilosis, R. stolonifer and F. solani.

Bioassay-guided fractionation, GC–MS identification and in vitro evaluation of antioxidant and antimicrobial activities of bioactive compounds from Eucalyptus globulus stump wood methanolic extract

The biomass of Eucalyptus globulus, and particularly the stump wood, has received increased attention in recent years, including in a biorefinery perspective. A previous work evaluated the crude methanolic extract of E. globulus stump wood regarding its potential antioxidant and antimicrobial properties. That work concluded that E. globulus stump wood could be considered as a source of bioactive polyphenols. In the present study, the crude methanolic extract of E. globulus stump wood was thus fractionated in order to isolate the components responsible for such biological properties. This extract was subjected to bioassay-guided fractionation and the antioxidant and antimicrobial properties of the fractions were then evaluated. The antioxidant activity was assessed by the DPPH scavenging assay and the antimicrobial properties were studied by bioautography and resazurin microtiter methods. The fractions presented considerable capacity to scavenge the DPPH free radicals and to inhibit the growth of bacterial and yeast strains. The results obtained for the fractions were even better than the ones obtained with the crude extract. The GC–MS analysis revealed that the major compounds present in the extract were gallic acid, ellagic acid, citramalic acid and citric acid.

Abietane diterpenes from the cones of Abies numidica de Lannoy ex Carrière (Pinaceae) and in vitro evaluation of their antimicrobial properties

Eight known abietane diterpenes (1–8) were isolated for the first time from Abies numidica cones (Pinaceae). The structures of all compounds were established by means of 1D and 2D NMR, and UV spectral analyses. The hydromethanolic extract of A. numidica cones was tested for its antimicrobial activity against 17 gram-positive and gram-negative bacteria and against five yeasts by the use of liquid and solid medium and bioautography methods. The best antimicrobial activity was found against gram-positive bacteria (MIC ≤ 0.3 mg/mL) Bacillus subtilis, Enterococcus faecalis ATCC 1034, Staphylococcus aureus 8325.4, S. aureus CIP 53.154, Micrococcus luteus and Listeria innocua and against Candida yeasts. The determination of MIC’s of isolated products showed a high activity of compounds 4 and 6 against S. aureus, L. innocua (MIC = 62.5 μg/mL) and E. faecalis (MIC = 125 μg/mL).

Antimicrobial properties of biosynthesized silver nanoparticles studied by flow cytometry and related techniques.

This work reports the effect of silver bionanoparticles (Bio(AgNPs) synthesized by Actinobacteria CGG 11n on selected Gram (+) and Gram (-) bacteria. Flow cytometry, classical antibiogram method and fluorescent microscopy approach was used for evaluation of antimicrobial activity of Bio(AgNPs) and their combination with antibiotics. Furthermore, the performed research specified the capacity of flow cytometry method as an alternative to the standard ones and as a complementary method to electromigration techniques. The study showed antibacterial activity of both BioAgNPs and the combination of antibiotics/BioAgNPs against all the tested bacteria strains in comparison with a diffusion, dilution and bioautographic methods. The synergistic effect of antibiotics/BioAgNPs combination (e.g. kanamycin, ampicillin, neomycin and streptomycin) was found to be more notable against Pseudomonas aeruginosa representing a prototype of multi-drug resistant "superbugs" for which effective therapeutic options are very limited.

Antimicrobial Activities and Chemical Characterization of Propolis Collected from Three Different National Areas of Ethiopia

Background: Propolis has revealed versatile and valuable biological and pharmacological activities such as antibacterial, antimitotic and antiviral. It has other less obvious effects such as cytotoxic, antioxidant, anti-inflammatory and immunomodulation. The variation in the chemistry of propolis from different locations and plant sources has made it a source of new biologically active molecules, mainly antioxidative and antibacterial agents. Aim of the study: The purpose of the present study was primarily to establish and assess the variation (if any) in the antimicrobial activities of propolis samples from three different local areas of Ethiopia. The study also incorporates identification and characterization of the major antimicrobial component(s) of propolis that are responsible for the antimicrobial activity. Materials and methods: Pure cultures of two gram positive (S.aureus and methicillin resistant S.aureus (MRSA)), three gram negative (E. coli, P. aeruginosa, S. typhi) bacterial species and one fungal species (C. albicans) were obtained from the department of Microbiology and Bacteriology Laboratory center, Addis Ababa University. Three propolis samples were collected from South Wollo, Tembain and Holeta regions of Ethiopia during the summer season. 70% ethanolic extract preparation for each propolis sample was made by using maceration procedure. Agar plate disk diffusion using paper disks and Agar well diffusion methods were employed to study antimicrobial activity of ethanolic extract of propolis from Tembain (EEPT) and ethanolic extract of propolis from Holeta. Thin layer chromatography (TLC) phytochemical screening method using different TLC spray reagents (vanillin, aluminum chloride, diazotized sulfanilic acid and -anisaldehyde), TLC agar overlay bioautography method and column chromatography were employed to partially identify and characterize the antimicrobial active components of propolis from Holeta and Tembain regions. Conclusion: Ethanolic extract of propolis from Holeta and ethanolic extract of propolis from Tembain showed antibacterial activity against S. aureus and MRSA only without showing effect on other bacterial or the fungal strain. A variation in the chemical composition between EEPH and EEPT was noted. The antimicrobial activities of propolis could not be correlated with any single components of propolis therefore the bactericidal effects of propolis from this region may be as a result of the sum antimicrobial components.

Isolation, Antioxidant and Antifungal Activities of Two Newly Reported Compounds from Bergenia ciliata

Ethyl acetate fraction of Bergenia ciliata was subjected to various chromatographic techniques such as silica gel CC, size exclusion CC and reverse phase CC, which resulted in isolation of two new compounds from this specie. Modern spectroscopic techniques such as 1H-NMR, 13C-NMR (BB and DEPT), 2D NMR, ESI and ESI-HR-MS were utilized to elucidate the structures of isolated secondary metabolites. Both the compounds A, B showed a remarkable fungicidal activity against cladosporium cladosporioides in TLC bio-autography method. Compound B showed maximum activity (90.34±0.81%) against control drug.

THE STUDY OF ANTIFUNGAL ACTIVITY FROM INDIGENOUS PLANTS FROM INDONESIA: AN IN VITRO STUDY

Objective: This study aimed to look for new agents from extracted Indonesia herbs which have antifungal activity with better safety profile against Candida albicans, Microsporum gypseum, and Trichophyton mentagrophytes. Methods: Screening of eleven herbs was perform to determine the highest antifungal activity. Minimum Inhibitory Concentration (MIC) and Minimum Fungicidal Concentration (MFC) of selected extracts, rosemary (Rosmarinus officinalis) leaves, Java turmeric (Curcuma xanthorriza) rhizome, and ginger (Zingiber officinalis) rhizome; were determined by using microdilution and agar diffusion methods. Extracts were then combined to evaluate further activities. Selected extracts were fractionated by using liquid-liquid extraction, analyzed by TLC bioautography.Results: ethanolic extract of rosemary (Rosmarinus officinalis) leaves, Java turmeric (Curcuma xanthorriza) rhizome, and ginger (Zingiber officinalis) rhizome in combination showed the highest activity and synergistic interaction against Trichophyton mentagrophytes. There were several components from fractions were actively inhibiting corresponding fungi according to TLC bioautography method. Conclusion: ethanolic extracts of rosemary, Java turmeric, and ginger had the highest antifungal potency, both as extract and in combination comparable to ketoconazole as reference drug. These selected extracts are potential to be used as new antifungal agents.

Antibacterial, antifungal and antioxidant activity of Olea africana against pathogenic yeast and nosocomial pathogens.

BackgroundOlea africana leaves are used by Bapedi people to treat different ailments. The use of these leaves is not validated, therefore the aim of this study is to validate antimicrobial properties of this plant.MethodsThe ground leaves were extracted using solvents of varying polarity (hexane, chloroform, dichloromethane (DCM), ethyl acetate, acetone, ethanol, methanol, butanol and water). Thin layer chromatography (TLC) was used to analyse the chemical constituents of the extracts. The TLC plates were developed in three different solvent systems, namely, benzene/ethanol/ammonium solution (BEA), chloroform/ethyl acetate/formic acid (CEF) and ethyl acetate/methanol/water (EMW). The micro-dilution assay and bioautography method were used to evaluate the antibacterial activity of the extracts against Escherichia coli, Pseudomonas aeruginosa, Enterococcus faecalis and Staphylococcus aureus and the antifungal activity against Candida albicans and Cryptococcus neoformans.ResultsMethanol was the best extractant, yielding a larger amount of plant material whereas hexane yielded the least amount. In phytochemical analyses, more compounds were observed in BEA, followed by EMW and CEF. Qualitative 2, 2- diphenylpacryl-1-hydrazyl (DPPH) assay displayed that all the extracts had antioxidant activity. Antioxidant compounds could not be separated using BEA solvent system while with CEF and EMW enabled antioxidant compounds separation. The minimum inhibitory concentrations (MIC) values against test bacteria ranged between 0.16 and 2.50 mg/mL whereas against fungi, MIC ranged from 0.16 to 0.63 mg/mL. Bioautography results demonstrated that more than one compound was responsible for antimicrobial activity in the microdilution assay as the compounds were located at different Rf values.ConclusionsThe results indicate that leaf extracts of Olea africana contain compounds with antioxidant, antibacterial and antifungal activities. Therefore, further studies are required to isolate the active compounds and perform other tests such as cytotoxicity. Olea africana may be a potential source of antimicrobial compounds.

Detection of antibiotics, active against Bacillus subtilis, in grain and feed

Aim. Detection of antibiotic substances in samples of grain, extraction cake, and oilcake. Methods. The bioautography method using strains of Bacillus subtilis as test-microorganisms was used to study 102 samples of feed substrates (corn, maize gluten, barley, wheat, sorghum, chaff, dust middling, sunfl ower oilcake and extraction cake, soybean meal, feed yeast and grains). Results. From one to four antibiotic substances, inhibiting the growth of B. subtilis and characterized by a wide range of values of chromatographic mobility index, were detected in 95 % of samples of feed substrates. Average areas of the zones of absent growth of a test- microorganism, corresponding to 2.5 g of the sample, were in the range of 52–217 mm 2 . Conclusions. It was established that feeder grain and other feed substrates are highly contaminated with antibiotics which indicates the necessity of their identifi cation, search for contamination sources, study of prevalence and estimation of the possible impact on the indices of health, performance and reproduction of farm animals and poultry.