Activity Of Acetone Leaf Extracts Research Articles

In vitro antibacterial potential of acetone leaf extract of Otostegia integrifolia Benth against human selected bacterial pathogens

Otostegia integrifolia Benth (O. integrifolia) is the endogenous medicinal plant of Ethiopia mostly used for the treatment of Stomach ache, tonsillitis, hypertension, malaria, ascariasis, and lung diseases. The current study was focused on phytochemical analysis and evaluation of the antibacterial activity of O. integrifolia Benth leave extracts against selected human bacterial pathogen by the agar well diffusion and microtube broth dilution method. Plant leaves were extracted with less polar to high polar solvents such as petroleum ether, chloroform, acetone, and ethanol. Phytochemical investigation was carried out for the identification of secondary metabolites responsible for antibacterial activity. In vitro antibacterial potential of O.integrifolia Benth leaf organic extracts against human pathogenic gram-negative (E. coli, K. pneumonia, V. cholera) and positive bacteria (B.subtilis, E. faecalis, S.aureus) were assessed by agar well diffusion, and bacterial inhibitory concentration of effective plant extracts was determined by 96 well plate broth dilution assay. Among all the tested organic leaf extracts, the acetone leaf extract of O.integrifolia Benth exhibited a promising broad spectrum of in vitro antibacterial activity against all tested multiple drug-resistant bacteria pathogens with significant MIC values of K.pneumoniae (2.144µg/ml), V. cholera (2.025µg), B.substilis (2.604 µg), and S.aureus (3.028 µg), respectively. The significant antibacterial activity of acetone leaf extracts of O. integrifolia Benth was due to the existence of flavonoids and phenolic compounds. The current studies demonstrated that the broad-spectrum antibacterial activity of acetone leaf extracts of O.integrifolia Benth might be helpful for the isolation of novel potent antibacterial agents against infectious bacterial pathogens without any side effects.

Ultrastructure changes induced by the phloroglucinol derivative agrimol G isolated from Leucosidea sericea in Haemonchus contortus

Plant extracts used for the treatment of helminth infections in sheep are an alternative to chemical anthelmintic drugs. Previous studies have reported the anthelmintic activity of acetone leaf extracts of Leucosidea sericea. For this study, we evaluate the ultrastructure changes induced by the acetone leaf extract of L. sericea and the component agrimol G (AG) that was isolated for the first time on adult haemonchus parasites. Adult haemonchus parasites harvested from sheep were incubated with the plant extract and AG for 3 h and evaluated by both scanning and transmission electron microscopy in comparison and in combination with albendazole or ivermectin. In all cases the method of evaluation shows ultrastructural changes, with albendazole inducing mitochondrial damage and ivermectin inducing muscle degeneration, both as previously described. Incubation with the plant extract and AG resulted in the formation of numerous non-membrane bound multi-vesicular like bodies and evenly spread disruptions/erosion in the epicuticle. Combining AG with ivermectin or albendazole resulted in an absence of effect of AG. Based on the structural changes induced by AG, together with the absence of an effect in combination with ivermectin and albendazole would suggest a disrupted microtubular network. The latter does however require biochemical confirmation.

Antibacterial and antibiofilm activity of acetone leaf extracts of nine under-investigated south African Eugenia and Syzygium (Myrtaceae) species and their selectivity indices

BackgroundAntimicrobial resistance (AMR) remains an important global health issue but the gap between AMR and development of new antimicrobials is increasing. Plant extracts may have good activity per se or may be sources of effective antimicrobial compounds which can act against planktonic and/or biofilms of pathogens. We determined the antimicrobial efficacy and cytotoxicity of some under-investigated plants from the Myrtaceae family endemic to South Africa. The ability of the plant extracts to inhibit or destroy pre-formed bacterial biofilms was also determined.MethodsBased on previous preliminary in vitro screening and on chemotaxonomy, nine species from the Myrtaceae family were selected. The antimicrobial activity of the crude acetone leaf extracts was determined against six common nosocomial pathogens, namely: Gram-positive bacteria (Bacillus cereus, Enterococcus faecalis, Staphylococcus aureus), Gram-negative bacteria (Escherichia coli, Pseudomonas aeruginosa, Salmonella Typhimurium) using a two-fold serial microdilution assay with p-iodonitrotetrazolium violet as growth indicator. The number of antimicrobial compounds present in extracts was determined by bioautography. Cytotoxicity of extracts was determined against Vero kidney cells using a colorimetric tetrazolium-based assay. The total antibacterial activity (TAA) in ml/g and selectivity index (LC50/MIC) of the plant extracts were calculated. A modified crystal violet assay was used to determine the antibiofilm activity of the extracts.ResultsSyzygium legatii, Syzygium masukuense, and Syzygium species A had the best activities against Gram-negative and Gram-positive bacteria (MIC) values ranging from 0.04–0.08 mg/ml. Eugenia erythrophylla had the best MIC (0.02 mg/ml) against Bacillus cereus. Many extracts had relatively low cytotoxicity (LC50 > 20 μg/ml) leading to reasonable selectivity indices. Three leaf extracts (Syzygium masukuense, Syzygium species A, and Eugenia natalitia) were moderately cytotoxic (20 μg/ml < LC50 < 100 μg/ml). The plant extracts had a good capacity to reduce biofilm formation and good to poor potential to destroy pre-formed biofilms.ConclusionsThe plant species examined in this study had varying degrees of antibacterial activity against bacterial planktonic and biofilm forms with some having good activity against both forms. Several of these selected species may be potential candidates for further investigation to isolate antimicrobial compounds and to determine the mechanism of activity.

Temperature stress does not affect antimicrobial activity of some South African medicinal plants

Some traditional healers believe that cultivated medicinal plants are not as active as plants collected from nature that may be growing under more environmental stress. This perception has implications for the conservation of popular medicinal plants. We previously found that water stress on plants had a limited effect on antimicrobial activity of its extracts. We now investigated the antimicrobial activity of leaf extracts of seedlings of Leonotis dysophylla Benth. from one batch and clones of Bulbine frutescens (L.) Willd. and Tulbaghia violacea Harv. plant species grown under temperature regimes of 15 and 30 °C. The aim was to determine if temperature stress would influence the antimicrobial activity of acetone leaf extracts of these widely used plant species against four bacteria and three fungi. The minimum inhibitory concentration (MIC) and total activity was determined to evaluate the antimicrobial activity of plant extracts. All leaf extracts across both temperature regimes had intermediate antimicrobial activity with MIC values varying from 0.42 to 2.50 mg/ml. With an exception of one plant extract against one fungus, there were no statistically significant differences in the MIC values of plant extracts against bacteria and fungi between the different temperatures. Total activity values that also take into account the quantity extracted were significantly higher particularly for B. frutescens and T. violacea grown under a temperature of 30 °C. High temperature might have led to a larger percentage of the plant material being soluble in acetone with little change in the composition of antibacterial compounds. The temperature regimes applied in this study may be optimal for cultivating these plant species, but 30 °C may be preferable to 15 °C for B. frutescense. These results as well as results found earlier on the effect of water stress questions the perception that cultivation weakens medicinal properties. Cultivating medicinal plants may lead to herbal medicines with more consistent quality and biological activities.

Influence of annual rainfall on antibacterial activity of acetone leaf extracts of selected medicinal trees

With the increasing demand for medicinal plants there is a danger that some populations may be destroyed by over collection. Cultivation of medicinal plants may address this problem, but many traditional healers believe that cultivated plants are not as active as plants growing in nature. There is uncertainty around the extent to which environmental stress affects the biological activity of plant extracts. We determined the effects of different annual rainfalls on antibacterial activity of tree leaf extracts of Terminalia sericea, Combretum collinum and Sclerocarya birrea growing under annual rainfalls of ≥870mm, 650mm and ≤480mm in the Lowveld, South Africa. The minimum inhibitory concentration of acetone leaf extracts was determined by a serial microplate dilution technique using tetrazolium violet as growth indicator on four nosocomial bacteria Enterococcus faecalis, Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa. MICs of leaf extracts of different trees varied between 60 and 1460μg/ml. Variations were for T. sericea (60–820μg/ml), for C. collinum (70–820μg/ml) and for S. birrea (70–1460μg/ml). Total activity values of the same extracts ranged between 48ml/g and 539ml/g. Antibacterial activity and total activity of plant extracts against test bacteria generally significantly (P≤0.05) increased under lower rainfall. In some cases there were no clear correlations. There were varying sensitivities of different bacteria of leaf extracts of plants grown under different annual rainfalls. This variation suggests that possible water stress caused by different rainfalls did not exclusively affect antibacterial activity of plant extracts. It is possible that other factors such as genetic, edaphic, microclimate, herbivory and pathogens could also have had an effect. More experiments using clonal plant material under well controlled environmental conditions are required to provide a more definitive answer.

Using plant extracts to control ticks and tick-borne diseases: A review

Plant extracts used for the treatment of helminth infections in animals are an alternative to chemical anthelmintics. Previous studies have reported the anthelmintic activity of acetone leaf extracts of Leucosidea sericea. However the mechanisms of action remain unknown. The aim of this study was to determine through scanning and transmission electron microscopy the effect of the acetone leaf extract of L. sericea and two phloroglucinol derivatives on the ultrastructure of Haemonchus contortus. A positive drug control treatment using albendazole and ivermectin was included. The main changes identified were the alteration and destruction of the cuticle, changes to the hypodermis, vacuoles within the cytoplasm, cytoplasmic degradation, cellular swelling and in a few cases, some abnormality within the mitochondria. The control for albendazole and ivermectin had the expected destruction of internal architecture and paralysis respectively. The lesions seen were probably due to bioactive plant metabolites and the route of entry. The possible mode of action is direct as it depends on contact between the bioactive plant metabolite and Haemonchus contortus.

Establishing the effects of phloroglucinol derivatives isolated from Leucosidea sericea on the ultrastructure of Haemonchus contortus, using electron microscopy

Plant extracts used for the treatment of helminth infections in animals are an alternative to chemical anthelmintics. Previous studies have reported the anthelmintic activity of acetone leaf extracts of Leucosidea sericea. However the mechanisms of action remain unknown. The aim of this study was to determine through scanning and transmission electron microscopy the effect of the acetone leaf extract of L. sericea and two phloroglucinol derivatives on the ultrastructure of Haemonchus contortus. A positive drug control treatment using albendazole and ivermectin was included. The main changes identified were the alteration and destruction of the cuticle, changes to the hypodermis, vacuoles within the cytoplasm, cytoplasmic degradation, cellular swelling and in a few cases, some abnormality within the mitochondria. The control for albendazole and ivermectin had the expected destruction of internal architecture and paralysis respectively. The lesions seen were probably due to bioactive plant metabolites and the route of entry. The possible mode of action is direct as it depends on contact between the bioactive plant metabolite and Haemonchus contortus.

In vitro antibacterial, antioxidant and cytotoxic activity of acetone leaf extracts of nine under-investigated Fabaceae tree species leads to potentially useful extracts in animal health and productivity.

BackgroundThe Fabaceae family is the second largest family of medicinal plants, containing more than 490 species which are being used as traditional medicine. The aim of this study was to determine the antioxidant and antibacterial activity as well as the cytotoxicity of acetone leaf extracts of nine tree species from the Fabaceae family that have not been investigated well previously for possible use in animal health and production.MethodsThe antibacterial activity was determined by a serial microdilution method against three Gram-positive and three Gram-negative bacteria. Antioxidant activity was determined using free-radical scavenging assays. The safety of the extracts was ascertained using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay on Vero African green monkey kidney cells.ResultsSix of the nine acetone extracts had significant antibacterial activity against at least one of the six bacterial species with (MIC 20–80 μg/mL). The Crotalaria capensis extract had the highest activity against Salmonella typhimurium, followed by Indigofera cylindrica with MICs of 20 μg/mL and 40 μg/mL respectively. The Dalbergia nitidula extract had free radical scavenging capacity (IC50 of 9.31 ± 2.14 μg/mL) close to that of the positive control Trolox in the DPPH assay. The Xylia torreana extract also had high activity (IC50 of 14.56 ± 3.96 μg/mL) in the ABTS assay. There was a good correlation between antioxidant activity and total phenolic content (R2 values > 0.8). The extracts had weak or no toxicity to Vero cells, compared to the positive control doxorubicin with the LC50 varying from 10.70 ± 3.47 to 131.98 ± 24.87 μg/mL at the concentrations tested.ConclusionExtracts of D. nitidula, X. torreana, C. capensis and I. cylindrica had a low cytotoxicity and high antimicrobial and/or antioxidant activity. These species are therefore promising candidates for the development of useful antimicrobial/antioxidant preparations with a low cytotoxicity that may be useful in promoting animal health and productivity.

Acetone leaf extracts of Breonadia salicina (Rubiaceae) and ursolic acid protect oranges against infection by Penicillium species

The activity of acetone leaf extracts of Breonadia salicina and the main antifungal compound isolated from the extract, ursolic acid, was determined against three important plant fungal pathogens (Penicillium expansum, P. janthinellum and P. digitatum) to evaluate their potential use in combating post-harvest infections of oranges. In an in vitro assay, acetone extracts had good antifungal activity against P. janthinellum with an MIC (minimum inhibitory concentration) of 0.08mg/ml. P. digitatum and P. expansum were more resistant both with MICs of 1.25mg/ml. We evaluated the potential use of an acetone extract and ursolic acid against these fungal pathogens in artificially infected oranges. A crude leaf extract at a concentration of 1mg/ml gave the same level of protection as 1mg/ml ursolic acid indicating synergistic activities within the crude extract. The acetone extract had an MIC of 0.16mg/ml compared to the MIC of 0.08mg/ml of amphotericin B against P. digitatum. Cytotoxicity of the crude extract and ursolic acid was determined using a tetrazolium-based colorimetric assay (3-(4,5-dimethylthiazol)-2,5-diphenyl tetrazolium bromide (MTT)) against Vero monkey kidney cells. The acetone extract had sufficient antifungal activity in vitro against these organisms to consider its use in the citrus industry after it has been tested under production and natural infection conditions and if it does not affect the fruit quality. The extracts were however more toxic to the kidney cells than to the fungi. The results show the potential use of plant extracts to combat plant fungal infections if extracts with lower cellular toxicity can be found or if the toxicity of the extract can be decreased without changing the antifungal activity.

Some southern African plant species used to treat helminth infections in ethnoveterinary medicine have excellent antifungal activities

Diseases caused by microorganisms and parasites remain a major challenge globally and particularly in sub-Saharan Africa to man and livestock. Resistance to available antimicrobials and the high cost or unavailability of antimicrobials complicates matters. Many rural people use plants to treat these infections. Because some anthelmintics e.g. benzimidazoles also have good antifungal activity we examined the antifungal activity of extracts of 13 plant species used in southern Africa to treat gastrointestinal helminth infections in livestock and in man. Antifungal activity of acetone leaf extracts was determined by serial microdilution with tetrazolium violet as growth indicator against Aspergillus fumigatus, Cryptococcus neoformans and Candida albicans. These pathogens play an important role in opportunistic infections of immune compromised patients. Cytotoxicity was determined by MTT cellular assay. Therapeutic indices were calculated and selectivity for different pathogens determined. We proposed a method to calculate the relation between microbicidal and microbistatic activities. Total activities for different plant species were calculated. On the whole, all 13 extracts had good antifungal activities with MIC values as low as 0.02 mg/mL for extracts of Clausena anisata against Aspergillus fumigatus a nd 0.04 mg/mL for extracts of Zanthoxylum capense, Clerodendrum glabrum, and Milletia grandis, against A. fumigatus. Clausena anisata extracts had the lowest cytotoxicity (LC50) of 0.17 mg/mL, a reasonable therapeutic index (2.65) against A. fumigatus. It also had selective activity against A. fumigatus, an overall fungicidal activity of 98% and a total activity of 3395 mL/g against A. fumigatus. This means that 1 g of acetone leaf extract can be diluted to 3.4 litres and it would still inhibit the growth. Clerodendrum glabrum, Zanthoxylum capense and Milletia grandis extracts also yielded promising results. Some plant extracts used for treatment of parasitic infections also have good antifungal activity. Because it is much easier to isolate antifungal compounds by bioassay guided fractionation, this approach may facilitate the isolation of anthelmintic compounds from active plant extracts. The viability of this approach can be tested by isolating the antifungal compounds and then determining its anthelmintic activity. Some of these plant extracts may also be useful in combating fungal infections.

Efficacy and toxicity of thirteen plant leaf acetone extracts used in ethnoveterinary medicine in South Africa on egg hatching and larval development of Haemonchus contortus

BackgroundHelminthiasis is a major limitation to the livestock industry in Africa. Haemonchus contortus is the singular most important helminth responsible for major economic losses in small ruminants. The high cost of anthelmintics to small farmers, resistance to available anthelmintics and residue problems in meat and milk consumed by humans further complicates matters. The use of plants and plant extracts as a possible source of new anthelmintics has received more interest in the last decade. Our aim was not to confirm the traditional use, but rather to determine activity of extracts.Based on our past experience acetone was used as extractant. Because it is cheaper and more reproducible to evaluate the activity of plant extracts, than doing animal studies, the activity of acetone leaf extracts of thirteen plant species used traditionally in ethnoveterinary medicine in South Africa were determined using the egg hatch assay and the larval development test. Cytotoxicity of these extracts was also evaluated using the MTT cellular assay.ResultsExtracts of three plant species i.e. Heteromorpha trifoliata, Maesa lanceolata and Leucosidea sericea had EC50 values of 0.62 mg/ml, 0.72 mg/ml and 1.08 mg/ml respectively for the egg hatch assay. Clausena anisata; (1.08 mg/ml) and Clerodendrum glabrum; (1.48 mg/ml) extracts were also active. In the larval development assay the H. trifoliata extract was the most effective with an EC50 of 0.64 mg/ml followed by L. sericea (1.27 mg/ml). The activities in the larval development test were generally lower in most plant species than the egg hatch assay. Based on the cytotoxicity results C. anisata was the least toxic with an LC50 of 0.17 mg/ml, while Cyathea dregei was the most toxic plant with an LC50 of 0.003 mg/ml. The C. anisata extract had the best selectivity index with a value of 0.10 and 0.08 for the two assays, followed by H. trifoliata and L. sericea with values of 0.07, 0.07 and 0.05, 0.04. The C. dregei extract had the worst selectivity index with a value of 0.00019 for both assays.ConclusionThe result of this study indicates which species should be further investigated in depth for isolation of compounds.

Some southern African plant species used to treat helminth infections in ethnoveterinary medicine have excellent antifungal activities

BackgroundDiseases caused by microorganisms and parasites remain a major challenge globally and particularly in sub-Saharan Africa to man and livestock. Resistance to available antimicrobials and the high cost or unavailability of antimicrobials complicates matters. Many rural people use plants to treat these infections. Because some anthelmintics e.g. benzimidazoles also have good antifungal activity we examined the antifungal activity of extracts of 13 plant species used in southern Africa to treat gastrointestinal helminth infections in livestock and in man.MethodsAntifungal activity of acetone leaf extracts was determined by serial microdilution with tetrazolium violet as growth indicator against Aspergillus fumigatus, Cryptococcus neoformans and Candida albicans. These pathogens play an important role in opportunistic infections of immune compromised patients. Cytotoxicity was determined by MTT cellular assay. Therapeutic indices were calculated and selectivity for different pathogens determined. We proposed a method to calculate the relation between microbicidal and microbistatic activities. Total activities for different plant species were calculated.ResultsOn the whole, all 13 extracts had good antifungal activities with MIC values as low as 0.02 mg/mL for extracts of Clausena anisata against Aspergillus fumigatus and 0.04 mg/mL for extracts of Zanthoxylum capense, Clerodendrum glabrum, and Milletia grandis, against A. fumigatus. Clausena anisata extracts had the lowest cytotoxicity (LC50) of 0.17 mg/mL, a reasonable therapeutic index (2.65) against A. fumigatus. It also had selective activity against A. fumigatus, an overall fungicidal activity of 98% and a total activity of 3395 mL/g against A. fumigatus. This means that 1 g of acetone leaf extract can be diluted to 3.4 litres and it would still inhibit the growth. Clerodendrum glabrum, Zanthoxylum capense and Milletia grandis extracts also yielded promising results.ConclusionsSome plant extracts used for treatment of parasitic infections also have good antifungal activity. Because it is much easier to isolate antifungal compounds by bioassay guided fractionation, this approach may facilitate the isolation of anthelmintic compounds from active plant extracts. The viability of this approach can be tested by isolating the antifungal compounds and then determining its anthelmintic activity. Some of these plant extracts may also be useful in combating fungal infections.

Five &lt;i&gt;Ochna&lt;/i&gt; species have high antibacterial activity and more than ten antibacterial compounds

New measures to control infections in humans and other animals are continuously being sought because of the increasing resistance of bacteria to antibiotics. In a wide tree screening survey of the antimicrobial activity of extracts of tree leaves (www.up.ac.za/phyto), Ochna pulchra, a small tree found widely in southern Africa, had good antibacterial activity. We therefore investigated the antibacterial activity of acetone leaf extracts of some other available Ochna spp. Antibacterial activity and the number of antibacterial compounds in acetone leaf extracts of Ochna natalitia, Ochna pretoriensis, O. pulchra, Ochna gamostigmata and Ochna serullata were determined with a tetrazolium violet serial microplate dilution assay and bioautography against Staphylococcus aureus, Escherichia coli, Enterococcus faecalis and Pseudomonas aeruginosa, bacteria commonly associated with nosocomial infections. The percentage yields of the extracts varied from 2.5% to 8%. The minimum inhibitory concentrations of the five species ranged from 40 µg/mL to 1250 µg/mL. E. coli was sensitive to all the extracts. The O. pretoriensis extract was the most active with minimum inhibitory concentrations of 0.065 mg/mL and 0.039 mg/mL against E. coli and E. faecalis, respectively. The O. pretoriensis extract also had the highest total activities of 923 mL/g and 1538 mL/g, indicating that the acetone extract from 1 g of dried plant material could be diluted to 923 mL or 1538 mL and would still kill these bacteria. Based on the bioautography results, the two most active species, O. pretoriensis and O. pulchra, contained at least 10 antibacterial compounds with similar R f values. Some of these antibacterial compounds were polar and others were non-polar. Variation in the chemical composition of the species may have some taxonomic value. The order of activity of the species to the bacteria were O. pretoriensis > O. pulchra > O. gamostigmata > O. serullata > O. natalitia. Even before toxicity and bioavailability issues are considered, some Ochna spp. leaf extracts have the potential to be used in treating skin infections.

With a different approach it is possible to develop commercially useful antimicrobial products from plants

Many compounds from plants play a major role as pharmaceutical products in several therapeutic applications in human and animal health. Despite thousands of publications in the field there has been a remarkable absence of therapeutic products from plants to combat microbial infections. Data will be presented to show that the following factors play a role in this absence: the extractant used, the bioassay used, the unexpectedly low activity of isolated antimicrobial compounds, the presence of synergism in crude extracts and focussing on plants traditionally used. The antibacterial activity of acetone leaf extracts of more than 700 South African tree species on four important nosocomial pathogens Staphylococcus aureus, Enterococcus faecalis, Escherichia coli and Pseudomonas aeruginosa were determined. A high proportion of these extracts had antibacterial activities with MICs lower than 0.08mg/ml. Unfortunately in many cases the extracts are toxic to mammalian cells. Many of these species have a good potential to be used as crude extracts to treat topical infections in humans. Examples will be demonstrated where these extracts have been as effective as commercial products in controlling microbial infections in animals and plants. The limitations associated with commercializing plant extracts as antimicrobials such as quality control, availability of material and potentising the extracts to yield patentable products will also be discussed. It appears that if the focus is on using extracts rather than isolated pure compounds there is a considerable opportunity to use the compounds present in plants to combat microbial infections.