M.G. Kulkarni

Antimicrobial properties and phenolic contents of medicinal plants used by the Venda people for conditions related to venereal diseases.

ETHNOPHARMACOLOGY RELEVANCE Many people are infected by venereal diseases and the human immunodeficiency virus (HIV) in rural areas. Sexual transmitted diseases are considered a disgrace in the community because of the stigmas attached to them. Indigenous people tend to use several medicinal plants to treat these infectious diseases rather than western medicines. AIM OF THE STUDY This study was aimed to evaluate the antibacterial, antifungal, antigonococcal, HIV-type 1 reverse transcriptase (RT) and to determine phenolic content of twelve medicinal plants used by the Venda people to treat venereal diseases. MATERIALS AND METHODS The dried plant materials of twelve medicinal plants were extracted with petroleum ether (PE), dichloromethane (DCM), 80% ethanol (EtOH) and water. The extracts were evaluated for their antimicrobial properties against two Gram-positive (Bacillus subtilis and Staphylococcus aureus), three Gram-negative (Neisseria gonorrhoeae, Escherichia coli and Klebsiella pneumoniae) bacteria and a fungus Candida albicans. The phenolic contents including total phenolics, flavonoids, gallotannins and condensed tannins of the methanolic extracts of the same plants were also determined. RESULTS DCM and EtOH extracts of Bolusanthus speciosus bark and stems exhibited good activity (<0.1mg/ml) against Escherichia coli and Staphylococcus aureus with MIC values ranging from 0.098 to 0.012 mg/ml. The DCM extracts of Bolusanthus speciosus bark showed the best activity against Candida albicans with MIC and MFC values of 0.012 mg/ml. Water and 50% methanol extracts of Acacia karroo bark, Bolusanthus speciosus stems and Ximenia caffra roots and leaves showed good RT inhibition percentages (>70%) at 1mg/ml. All tested extracts exhibited dose dependent IC(50) values ranging from (0.1 to 0.6 mg/ml). Almost all plant species investigated contained phenolic compounds, which were greater than 5mg/g with the exception of Adansonia digitata bark and Aloe chabaudii roots. The highest level of flavonoids (11.9 μg/g) were detected in Ximenia caffra leaves. Whereas the highest amount of gallotannins were detected in Ekebergia capensis bark (69 μg/g). Condensed tannins were higher in Ekebergia capensis bark and Ximenia caffra roots (0.47 and 0.48% respectively). Adansonia digitata bark and Aloe chabaudii roots exhibited low levels of phenolic compounds as well as antimicrobial activities. CONCLUSION The presented results offer supporting evidence for effective use of selected plant extracts. More in vitro confirmatory tests using other assays and/or in vivo tests are required. Toxicity studies are urgently needed to support the safe usage of these plants.

Pharmacological properties and protein binding capacity of phenolic extracts of some Venda medicinal plants used against cough and fever

ETHNOPHARMACOLOGY RELEVANCE Several ailments are caused by infectious bacteria and in other diseases; they act as co-infection which complicate human life by causing health hazards. In Venda (South Africa), many plants are used in traditional medicine to treat cough and fever. AIM OF THE STUDY This study was aimed at evaluating the antibacterial and antifungal properties, cyclooxygenases (COX), acetylcholinesterase (AChE) enzyme inhibitory effects and the phenolic composition as well as mutagenic properties of six medicinal plants used by the Venda people of Limpopo Province of South Africa against cough and fever. MATERIALS AND METHODS The petroleum ether (PE), dichloromethane (DCM), 80% ethanol (EtOH) and water extracts of six plants were tested against four infectious bacteria (Bacillus subtilis, Escherichia coli, Klebsiella pneumoniae and Staphylococcus aureus) and a fungus Candida albicans. The same extracts were evaluated for their ability to inhibit COX-1 and -2 enzymes. Methanolic and water extracts of the same plant were tested for acetylcholinesterase inhibitory effects. Total phenolics, flavonoids, gallotannins and condensed tannins were determined. The ability of the extracts to bind and precipitate proteins was also investigated. The extracts were investigated for genotoxicity with and without S9 (metabolic activation) against three Salmonella typhimurium tester strains TA98, TA100 and TA102. RESULTS The organic extracts of Rhus lancea leaves exhibited the best antibacterial activity with minimum inhibitory concentration (MIC) values ranging from 0.0061 to 0.049mg/ml. The best antifungal activity was observed from a DCM extract of Syzygium cordatum leaves with a MIC value of 0.195mg/ml. The methanolic and water extracts of the same plant exhibited high inhibitory effects towards AChE with IC(50) values of 0.22 and 0.26mg/ml, respectively. The highest levels of flavonoids and gallotannins were detected in Spirostachys africana bark; 11.57 and 48.88μg/g, respectively. The highest percentages (1.2%) of condensed tannins were detected in Uvaria caffra leaves. The high levels of phenolic compounds may have been responsible for high antimicrobial activities for extracts of S. africana bark and U. caffra leaves. S. cordatum leaves represented the highest affinity for protein binding with 93%. All the extracts were non-mutagenic towards the three tested strains with and without S9 metabolic activation. CONCLUSION The result obtained in this study goes a long way in validating the ethnobotanical usage of these medicinal plants in the treatment of cough and fever by the Venda people. However, more evidence obtainable from other assays not performed here are urgently required to confirm these results.

Cadmium induces hypodermal periderm formation in the roots of the monocotyledonous medicinal plant Merwilla plumbea

BACKGROUND AND AIMS Merwilla plumbea is an important African medicinal plant. As the plants grow in soils contaminated with metals from mining activities, the danger of human intoxication exists. An experiment with plants exposed to cadmium (Cd) was performed to investigate the response of M. plumbea to this heavy metal, its uptake and translocation to plant organs and reaction of root tissues. METHODS Plants grown from seeds were cultivated in controlled conditions. Hydroponic cultivation is not suitable for this species as roots do not tolerate aquatic conditions, and additional stress by Cd treatment results in total root growth inhibition and death. After cultivation in perlite the plants exposed to 1 and 5 mg Cd L(-1) in half-strength Hoaglands solution were compared with control plants. Growth parameters were evaluated, Cd content was determined by inductively coupled plasma mass spectroscopy (ICP-MS) and root structure was investigated using various staining procedures, including the fluorescent stain Fluorol yellow 088 to detect suberin deposition in cell walls. KEY RESULTS The plants exposed to Cd were significantly reduced in growth. Most of the Cd taken up by plants after 4 weeks cultivation was retained in roots, and only a small amount was translocated to bulbs and leaves. In reaction to higher Cd concentrations, roots developed a hypodermal periderm close to the root tip. Cells produced by cork cambium impregnate their cell walls by suberin. CONCLUSIONS It is suggested that the hypodermal periderm is developed in young root parts in reaction to Cd toxicity to protect the root from radial uptake of Cd ions. Secondary meristems are usually not present in monocotyledonous species. Another interpretation explaining formation of protective suberized layers as a result of periclinal divisions of the hypodermis is discussed. This process may represent an as yet unknown defence reaction of roots when exposed to elemental stress.

Variation in heavy metals and microelements in South African medicinal plants obtained from street markets

South African medicinal plants are traditionally harvested from a wide range of undisclosed locations by plant gatherers. Thus, there is a risk that plant material may be exposed to a variety of pollutants. The variation in five heavy metals (arsenic, cadmium, cobalt, nickel, and lead) and six essential elements (boron, copper, iron, manganese, molybdenum, and zinc) was determined in commonly used South African medicinal plants obtained from street markets. Elemental content was determined using inductively coupled plasma optical emission spectrophotometry. The reliability of the procedure was ensured by analysing a certified reference material. Medicinal plant samples contained arsenic and cadmium at levels exceeding the World Health Organization limits of 1 and 0.3 mg kg−1 respectively. Lead and nickel were detected in all samples. Elevated iron and manganese levels were recorded in certain plant species. Multiple metal contamination of parts of medicinal plants gives grounds for concern. This study emphasizes the unsafe consequences of the South African practice of collecting medicinal plants from undisclosed locations and making these readily available to the public.

Seedlings of medicinal plants treated with either a cytokinin antagonist (PI-55) or an inhibitor of cytokinin degradation (INCYDE) are protected against the negative effects of cadmium

Cytokinins are plant hormones that regulate diverse processes during plant growth and development. Targeted manipulation of their metabolism or perception has already shown benefits in agriculture. Recently, we described substances that can affect plants’ endogenous cytokinin status: a cytokinin antagonist, PI-55, and an inhibitor of cytokinin degradation, INCYDE (2-chloro-6-(3-methoxyphenyl)aminopurine). The effects of these substances on seedling growth in the medicinal plant Bulbine natalensis Baker (Asphodelaceae) and the metal tolerant Rumex crispus L. (Polygonaceae) under abiotic stress caused by cadmium (Cd) were tested. Cd is known for its negative effects on plant growth and its toxicity to humans. Treatment with either PI-55 or INCYDE had positive effects on seedling shoot and root growth and the fresh weight of treated seedlings grown in the presence of Cd. Even a single application of either compound at sub-micromolar concentrations was sufficient to reverse the inhibitory effects of Cd. Our results demonstrate that modulating cytokinin status with inhibitors of cytokinin perception and/or degradation may be useful in protecting plants against the adverse effects of high Cd levels.

Anti-inflammatory and mutagenic evaluation of medicinal plants used by Venda people against venereal and related diseases.

ETHNOPHARMACOLOGY RELEVANCE Inflammation is a major risk factor for various human diseases including venereal diseases, often resulting in treatment complications. Plants have been traditionally used for treatment of many different diseases and have been successfully proven to be an alternative source in treatment of infectious diseases. AIM OF THE STUDY This study was aimed at evaluating the anti-inflammatory activities and the mutagenic properties of 12 medicinal plants used by the Venda people against venereal and related diseases. MATERIALS AND METHODS The plants were evaluated for their anti-inflammatory activity against the cyclooxygenase (COX-1 and -2) enzymes and genotoxicity using the Ames test, with and without S9 (metabolic activation) against Salmonella typhimurium tester strain TA98. RESULTS DCM and PE extracts of Adansonia digitata bark, Bolusanthus speciosus bark, Pterocarpus angolensis bark and Pappea capensis leaves and EtOH and water extracts of Bolusanthus speciosus stem and Ekebergia capensis bark showed the best anti-inflammatory activity in both COX-1 and -2 assays at 250 μg/ml. These were further evaluated at three other concentrations (31.25, 62.5, and 125 μg/ml) to determine IC50 values. Water extracts of Ekebergia capensis bark showed the best IC50 value towards COX-1. The Ames test revealed that all plant extracts were non-mutagenic towards Salmonella typhimurium strain TA98 except for Elephantorrhiza burkei and Ekebergia capensis that showed weak mutagenicity. CONCLUSION The active plants may offer a new source of chemicals for the effective treatment of anti-inflammatory conditions related to venereal diseases.

Smoke Solutions and Temperature Influence the Germination and Seedling Growth of South African Mesic Grassland Species

Abstract Fire, natural or of anthropologic origin, is a recurrent phenomenon in South African mesic grassland. The species composition of these grasslands is sensitive to fire, particularly fire frequency. However, the mechanisms involved in influencing species composition are not fully understood. Currently there is a general suggestion that plant-derived smoke and smoke-isolated biologically active butenolide compound provide an important germination cue for a range of Poaceae species. Studies also show that these smoke solutions play a role in vegetative growth of many plants. We examined if this fire-response syndrome is related to the effect of plant-derived smoke-water (1∶500 v/v) and smoke-derived butenolide compound (10−8 M) on seed germination and seedling growth of six major constituent species of the grassland. In addition, the interaction of the smoke solutions with temperature was examined by incubating seeds at a range of temperatures. Treating seeds with smoke-water and butenolide, the germination rate and final germination percentage were greater in three of the six species. Themeda triandra Forssk. and Tristachya leucothrix Trin. ex Nees showed the greatest response, with final germination increased from 43% to 67% and 35% to 63%, respectively. With increasing temperature (> 30°C), Aristida junciformis Trin. & Rupr., Hyparrhenia hirta (L.) Staph, and Panicum maximum Jacq. responded positively to the test solutions. In nearly all the species tested, smoke-water–treated seeds produced significantly longer shoots or roots. However, the degree of response varied from species to species and across different temperatures. Findings from this study suggest that plant-derived smoke and its interaction with temperature may significantly influence the germination and seedling growth of the South African mesic grassland species, which can further alter the grassland composition.

Seed germination and phytochemical evaluation in seedlings of Aloe arborescens Mill.

Many Aloe species are exploited as natural products. Generally, the leaves are unsustainably picked from wild plants to meet the market demand. Basic scientific information on seed biology and the ways of increasing levels of secondary metabolites in seedlings is still lacking for Aloe species. This study investigated seed germination requirements and evaluated levels of secondary metabolites in seedlings of Aloe arborescens, an important species in traditional medicine. The highest percentage germination (78%) and the fastest germination rate (GR) (10% d− 1) with a mean germination time (MGT) of 9 days were achieved at 20°C under a 16-h photoperiod. At 25°C, maximum percentage germination (67%) (P < 0.05), higher GR (13% d− 1) and shorter MGT (6 days) were obtained under constant light. These results indicate that temperature and light play a significant role in germination of A. arborescens seeds. Increasing osmotic pressure on seeds decreased percentage germination, whereas buffering the solution to a range of pH values (4–10) did not significantly affect germination. Smoke–water (1:500 v/v), smoke-isolated karrikinolide (10− 8 and 10− 9 M) and potassium nitrate (10− 3 and 10− 4 M) significantly promoted germination compared with the control at 25°C (supra-optimal temperature) under a 16-h photoperiod. These treatments were also effective in increasing secondary metabolite levels (flavonoids and phenolics) in A. arborescens seedlings.

Effect of Nutrients and Smoke Solutions on Seed Germination and Seedling Growth of Tropical Soda Apple (Solanum viarum)

Abstract Solanum viarum, commonly known as tropical soda apple (TSA), is native to Brazil and Argentina but has become a harmful weed in many countries with tropical climates. This study was conducted to reassess the seed biology of TSA found in South Africa. Cold stratification (14 d), acid scarification (20% H2SO4 for 5 min), and sandpaper scarification (30 s) significantly improved percentage germination when compared to the control. The highest germination (99.5%) was achieved when seeds were germinated in 50% Hoaglands nutrient solution (HS). The lowest germination (66%) was recorded in the absence of phosphorus (P) under alternating light conditions. HS without nitrogen (N) completely inhibited seed germination of TSA under constant light conditions. These findings are useful in controlling TSA by amending the levels of N and P in soils. Seed germination of TSA was significantly enhanced by different concentrations of smoke-water and butenolide solution. Smoke-water dilution of 1∶500 v/v and butenolide concentration of 10−8M showed the highest seedling vigor indices (6,688 and 6,666, respectively) in comparison to the control (1,251) and gibberellic acid (GA3) concentrations (< 5,327). These findings suggest that germination of seeds or seedbanks of TSA might be successfully stimulated using smoke solutions. Subsequently, patches of seedlings emerging after treatment can be mechanically uprooted to reduce the infestation of TSA. However, justifying this with field trials is essential. Nomenclature: Tropical Soda Apple; Solanum viarum Dunal SOLVI.

Effect of smoke derivatives on in vitro pollen germination and pollen tube elongation of species from different plant families

Plant-derived smoke stimulates seed germination in numerous plant species. Smoke also has a positive stimulatory effect on pollen germination and pollen tube growth. The range of plant families affected my smoke still needs to be established since the initial study was restricted to only three species from the Amaryllidaceae. The effects of smoke-water (SW) and the smoke-derived compounds, karrikinolide (KAR1 ) and trimethylbutenolide (TMB) on pollen growth characteristics were evaluated in seven different plant families. Smoke-water (1:1000 and 1:2000 v:v) combined with either Brewbaker and Kwacks (BWK) medium or sucrose and boric acid (SB) medium significantly improved pollen germination and pollen tube growth in Aloe maculata All., Kniphofia uvaria Oken, Lachenalia aloides (L.f.) Engl. var. aloides and Tulbaghia simmleri P. Beauv. Karrikinolide (10(-6) and 10(-7) m) treatment significantly improved pollen tube growth in A. maculata, K. uvaria, L. aloides and Nematanthus crassifolius (Schott) Wiehle compared to the controls. BWK or SB medium containing TMB (10(-3) m) produced significantly longer pollen tubes in A. maculata, K. uvaria and N. crassifolius. These results indicate that plant-derived smoke and the smoke-isolated compounds may stimulate pollen growth in a wide range of plant species.