Feeding deterrence,larvicidal and haemolymph protein profiles of an Indian traditional medicinal plant Alangium salviifolium (L.F.) Wangerin on cluster caterpillar,Spodoptera litura (Fabricius) (Lepidoptera: Noctuidae)

Different organic solvent crude extracts from the leaves of Alangium salviifolium (L.F.) Wangerin, were tested for their feeding deterrence, larvicidal activity and protein concentrations on the fourth instar larvae of Spodoptera litura L. under laboratory conditions. Treatments were given through two different host plant leaves such as brinjal and castor. The bioassay was carried out at different concentrations viz., 0.625, 1.25, 2.5 and 5%. The maximum antifeedant activity in brinjal (52.0%) and castor (29.97%) leaf discs was recorded in ethyl acetate extract as compared with hexane and chloroform extracts at 5% concentration. The result of larvicidal activity revealed that the maximum larval mortality was registered in brinjal leaves (65.81%) and castor with (57.48%) in ethyl acetate extract as compared to hexane and chloroform extracts at 5% concentration. The ethyl acetate extract of A. salviifolium contained alkaloids, diterpenoids, and saponins. The treatment also reduced haemolymph protein concentration after 48 h.     

Biological activity of Xanthium strumarium seed extracts on different cancer cell lines and Aedes caspius,Culex pipiens (Diptera: Culicidae)

Effects of methanol extracts of Xanthium strumarium on different cancer cell lines and on the mortality rates of Aedes caspius, Culex pipiens (Diptera: Culicidae) were investigated. Among the cell lines tested, the Jurkat cell line was the most sensitive to the methanol extract and ethyl acetate fraction, with reported LC₅₀ values of 50.18 and 48.73 μg/ml respectively. Conversely, methanol extracts were not that toxic to the A549 cell line though the toxicity increased on further purification. The percentage of growth inhibition was dose dependent for the methanol extract and ethyl acetate fraction. The ethyl acetate fraction showed higher toxicity to all cell lines tested when compared to the methanol extract. The results showed that methanol extracts of plant seeds caused 100% mortality of mosquito larvae at a concentration of 1000 μg/ml after 24 h of treatment. The LC₅₀ and LC₉₀ values of X. strumarium were found to be 531.07 and 905.95 μg/ml against Ae. caspius and 502.32 and 867.63 μg/ml against Cx. Pipiens, respectively. From the investigations, it was concluded that the crude extract of X. strumarium showed a weak potential for controlling the larval instars of Ae. caspius and Cx. pipiens. However, on further purification the extract lost the larvicidal activity. The ethyl acetate fraction showed higher toxicity to all cell lines tested when compared to the methanol extract. The ethyl acetate fraction investigated in this study appears to have a weak larvicidal activity but a promising cytotoxic activity. Future studies will include purification and investigation in further detail of the action of X. strumarium on Cancer Cell Lines and mosquitoes.     

Insect growth regulator and insecticidal activity of beta-dihydroagarofurans from Maytenus spp. (Celastraceae)

From the aerial parts of Maytenus disticha, we have isolated 9beta-benzoyloxy-1alpha,2alpha,6beta,8alpha,15penta-acetoxy-dihydro-beta-agarofuran (1) and from seeds of Maytenus boaria 9beta-furoyloxy-1alpha,6beta,8alpha-triacetoxy-dihydro-beta-agarofuran (2). These compounds and their MeOH and hexane/ethyl acetate (1:1 v/v) extracts were evaluated for their effects on the fall armyworm (Spodoptera frugiperda). Toosendanin, a commercial insecticide derived from Melia azedarach was used as a positive control. When tested for activity using neonate larvae in a nochoice artificial diet bioassays, the agarofurans 1, 2 and toosendanin as well as the MeOH and hexane/EtOAc extracts caused significant growth inhibitory effects with GC50 of 7.55; 3.84; 1.75; 14.0 and 7.3 ppm at 7 days, respectively. Compounds 1 and 2 caused 100% larval mortality at 25 and 15 ppm, respectively. MeOH and hexane/EtOAc extracts caused 100% larval mortality at 25.0 ppm, respectively, they also increased the development time of surviving larvae and a significant delay for the time of pupation and adult emergence. These compounds showed comparable potency of activity with toosendanin. Acute toxicity against adults of S. frugiperda was also found, for hexane/EtOAc extract and 2 had the most potent activity with LD50 value of 4.7 and 1.9 ppm, respectively. MeOH extract, hexane/EtOAc extract, 1 and 2 caused acetylcholinesterase inhibition with 78.0, 89.2, 79.3 and 100% inhibition at 15.0 ppm, respectively. Therefore, the furoyloxy agarofuran may be responsible for the insecticidal activity of these plants.     

Evaluation of free radical scavenging activity of Butea monosperma Lam

In the present study, ethyl acetate, butanol and aqueous fractions derived from total methanol extract of Butea monosperma flowers were evaluated for radical scavenging activities using different in vitro models like reducing power assay, scavenging of 2,2 diphenyl-1-picrylhydrazyl (DPPH) radical, nitric oxide radical, superoxide anion radical, hydroxyl radical and inhibition of erythrocyte hemolysis using 2, 2' azo-bis (amidinopropane) dihydrochloride (AAPH). Methanol extract along with its ethyl acetate and butanol fractions showed potent free radical scavenging activity, whereas aqueous fraction was found to be devoid of any radical scavenging properties. The observed activity could be due to the higher phenolic content in the extracts (16.1, 25.29, and 17.74% w/w in methanol extract, ethyl acetate and butanol fractions respectively). HPTLC fingerprint profile of the ethyl acetate and butanol fractions were developed which would serve as reference standard for quality control of the extracts.     

Anti-genotoxic and free-radical scavenging activities of extracts from (Tunisian) Myrtus communis

The effect of extracts from leaves of Myrtus communis on the SOS reponse induced by Aflatoxin B1 (AFB1) and Nifuroxazide was investigated in a bacterial assay system, i.e. the SOS chromotest with Escherichia coli PQ37. Aqueous extract, the total flavonoids oligomer fraction (TOF), hexane, chloroform, ethyl acetate and methanol extracts and essential oil obtained from M. communis significantly decreased the SOS response induced by AFB1 (10 microg/assay) and Nifuroxazide (20 microg/assay). Ethyl acetate and methanol extracts showed the strongest inhibition of the induction of the SOS response by the indirectly genotoxic AFB1. The methanol and aqueous extracts exhibited the highest level of protection towards the SOS-induced response by the directly genotoxic Nifuroxazide. In addition to anti-genotoxic activity, the aqueous extract, the TOF, and the ethyl acetate and methanol extracts showed an important free-radical scavenging activity towards the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical. These results suggest the future utilization of these extracts as additives in chemoprevention studies.     

Bioefficacy of Ceasalpinea bonduc (L.) Roxb. against Spodoptera litura Fab. (Lepidoptera: Noctuidae)

Bioefficacy of hexane, chloroform and ethyl acetate extracts of the seeds of Ceasalpinea bonduc (L.) Roxb. was studied against third instar larvae of Spodoptera litura at 0.25, 0.5, 1, 2.5 and 5.0% concentrations. Significant antifeedant, larvicidal and pupicidal activities and least LC₅₀ values were observed in chloroform extract. The chloroform extract was subjected to fractionation using silica gel column chromatography. Six fractions were obtained; among these, the third fraction showed high antifeedant, larvicidal and pupicidal activities at 1000 ppm concentration. Abnormalities in adults were also observed. All the activities were concentration dependent. C. bonduc could be useful in integrated pest management programme.     

Insect Feeding Deterrents in Endophyte-Infected Tall Fescue

The presence of an endophytic fungus, Acremonium coenophialum, in tall fescue (Festuca arundinacea) deterred aphid feeding by Rhopalosiphum padi and Schizaphis graminum. Both species of aphid were unable to survive when confined to endophyte-infected tall fescue plants. Feeding deterrents and toxic factors to R. padi and Oncopeltus fasciatus, large milkweed bug, were primarily associated with a methanol extract obtained when endophyte-infected tall fescue seed was serially extracted with hexane, ethyl acetate, and methanol. The concentrations of pyrrolizidine alkaloids were determined to be 30 to 100 times greater in the methanol extract than in the hexane and ethyl acetate extracts.     

Hepatoprotective and antineoplastic properties of Protaetia brevitarsis larvae

Protaetia brevitarsis larvae have been used as a traditional medicine to treat various liver diseases. Freeze‐dried powder and solvent partitioned fractions of Protaetia brevitarsis larvae, in this study, were examined for hepatoprotective and antitumor activities in vivo and in vitro. Oral administration of the powder reduced signs of acute and chronic liver injuries in diethylnitrosamine‐induced hepatotoxic mouse model. Various cancer cell types, but not normal primary hepatocytes, were killed by the hexane, ethyl acetate and butanol fractions, which were partitioned from larval extracts. Apoptosis and autophagy were primarily induced in cancer cells by hexane and ethyl acetate fractions, respectively, and can be the cytotoxic mechanisms of the fractions. Further analyses of the insect fractions could lead to identification of novel bioactive substances that can be exploited as therapeutics for various human liver diseases.     

Inhibition of the growth and development of mosquito larvae of Culex pipiens L. (Diptera: Culicidae) treated with extract from flower of Matricaria chamomilla (Asteraceae)

Mosquitoes can transfer many adverse diseases to human and animals therefore, there is a need to fight their spread. Among promising larvicidal sources is the use of plant extracts, which will play an important role in the future. This study was conducted to assess the larvicidal activity of Ferula hermonis Boiss,Achillea millefolium, Salvia officnalis, Psidium guaja and Matricaria chamomilla extract against Culex pipiens. The plant materials were extracted with hexane, dichloromethane, ethyl acetate, and methanol using a Soxhlet extractor. The extracts were evaluated for larvicidal activity against Cx. pipiens the mortality was monitored after 24 and 48 h of exposure. None of the extracts tested showed larvicidal acitivties except M. chamomilla. The ethyl acetate extract showed the most promising larvicidal activity with LC₅₀ values of 287.1 and 209.4 ppm after 24 and 48 h of exposure, respectively. Treatment of the eggs with different concentrations of the active extract decreased the hatchability of the eggs dose dependently from 95 to 86.49%. Similarly, the pupal duration increased in treated groups. The larval period lasted for 12 d, whereas that of the control group lasted for 10 d. Furthermore, the pupal period lasted 3 d (control 2 d) in treated groups. The data also revealed a significant decrease in the growth index in treated groups (0.00–7.53) than that of the control (8.5). The GC–MS analysis revealed the presence of 1.6.10‐dodecatriene, 7,11‐dimethyl‐3‐methylene (89.68%), 1,6‐cyclodecadiene, 1‐methyl‐5‐methylene‐8‐(1‐methylethyl)‐, [S‐(E,E)] (6.34%), 2H‐pyran ?3‐ol (4.04%), and 2H‐1‐benzopyran‐2‐one (0.079%).     

飞机草提取物对红脉穗螟的产卵忌避及杀卵活性

为探讨飞机草提取物对红脉穗螟Tirathaba rufivena Walker的防控潜力,采用室内生物测定法,研究了飞机草提取物对红脉穗螟的产卵忌避作用和杀卵活性。结果表明,飞机草乙酸乙酯提取物对红脉穗螟的忌避效果最好,选择性和非选择性忌避率分别为41.85%和46.84%;不同提取物处理红脉穗螟的卵后,均造成卵孵化率降低和初孵幼虫的死亡,其中乙酸乙酯提取物对红脉穗螟卵的影响最大,孵化率和1龄幼虫死亡率分别达到65.89%和40.37%。对飞机草乙酸乙酯提取物进行分级萃取测试各萃取物的活性发现,正己烷萃取物对红脉穗螟的产卵忌避作用和杀卵活性最强。田间试验表明,采用正己烷萃取物稀释50倍和100倍浓度进行田间喷雾,药后15 d对红脉穗螟的种群控制率可达到50%以上。     

Reduced physiological and reproductive fitness induced by Nerium oleander leaf extracts in the cotton bollworm,Helicoverpa armigera (Lepidoptera: Noctuidae)

Helicoverpa armigera is one of the most devastating, cosmopolitan, polyphagous and multivoltine pests of agricultural crops. The growing side-effects of synthetic insecticides used to control agricultural pests, have amplified demand for the relatively safe and biodegradable compounds which offer benefits over synthetic chemical insecticides. Hence, the current study investigates the effects of Nerium oleander methanol leaf extracts on the physiological and reproductive fitness of H. armigera. Addition of the range of extract concentration; 0.5%–10.0%; in the larval diet induced an appreciable and dose-dependent effect on the developmental parameters of H. armigera. The extract, though resulted in moderate larval and pupal mortality, caused significant growth regulatory effects. A prolonged larval duration, formation of larval-pupal intermediates and significant reduction in the average larval weight gain by 26.5% - 63.4%; and pupal weight by 23.9% - 63.4% was recorded. The 74.6% to 54.9% reduction in the oviposition duration in the emerged adults, and diminished fecundity and egg hatch in the corresponding range of 0.78–0.63 and 0.89–0.62 demonstrated negative impact of the extract on the adult reproductive fitness. The adverse impact of N. oleander methanol leaf extracts on the growth and development of H. armigera leading to the reproductive disadvantage in adults can be beneficial in the field management; though its use can only be ascertained after identification of bioactive constituents and elucidating their mode of action.     

Anticariogenic and phytochemical evaluation of Eucalyptus globules Labill.

In the present study, in vitro anticariogenic potential of ethyl acetate, hexane and methanol and aqueous extracts of plant leaves of Eucalyptus globules Labill. were evaluated by using four cariogenic bacteria, Lactobacillus acidophilus, Lactobacillus casei, Staphylococcus aureus and Streptococcus mutans. Agar well diffusion method and minimum inhibitory concentration (MIC) were used for this purpose. The ethyl acetate extracted fraction of plant leaves showed good inhibitory effects against all selected bacteria. In Eucalyptus globules, hexane and ethyl acetate extracts found highly effective against, Lactobacillus acidophilus with MIC value of 0.031 and 0.062 mg/mL, respectively. Qualitative phytochemical investigation of above extracts showed the presence of alkaloids, phenolic compounds, steroids, cardiac glycosides and terpenes. Based on the MIC value and bioautography, ethyl acetate of plant leaf was selected for further study. Further investigation on the structure elucidation of the bioactive compound using IR, GC-MS and NMR techniques revealed the presence of alpha-farnesene, a sesquiterpene. Eucalyptus globules plant leaf extracts have great potential as anticariogenic agents that may be useful in the treatment of oral disease.     

Biological effects of active fraction isolated from Hydnocarpus pentandra (Bunch. –Ham.) Oken seeds against Helicoverpa armigera (Hub.) (Lepidoptera: Noctuidae)

Hexane, chloroform, ethyl acetate and methanol extracts of Hydnocarpus pentandra (Flacourtiaceae) seeds were tested for antifeedant, larvicidal, pupal mortality and adult deformations activities against Helicoverpa armigera. Crude extracts were screened at 0.5, 1.0, 1.5 and 2.0% concentrations. Bioassay-guided fractionation method was followed to isolate the active fraction from the crude extract. Active fraction was analysed by FT-IR, ¹H NMR, ¹³C NMR and GC-MS. Hexane extract presented the highest antifeedant (87.89%), pupal mortality (41.67%) and adult malformation activities at 2% concentration. Seven different fractions were isolated from hexane extract, among which fraction-2 showed the highest antifeedant (81.43%) activity and recorded the lowest LC₅₀ of 792.07 ppm. The fraction-2 contained two cyclopentenyl carboxylic acids, such as hydnocarpic acid (1) and chaulmoogric acid (2) in the ratio of 2:1. These compounds were major constituents in the active fraction of hexane extract of H. pentandra seeds. Fraction-2 can be used for agricultural pest management.     

Chemical compositions of leaf extracts from Conocarpus erectus L. (Combretaceae) and their bioactivities against Tribolium castaneum Herbst (Coleoptera: Tenebrionidae)

Botanical insecticides have long been considered as alternatives to synthetic chemical insecticides in IPM programs. Effects of aqueous and ethanolic extracts obtained from buttonwood, Conocarpus erectus L. (Combretaceae), leaves on a major kind of stored product pests, Tribolium castaneum Herbst (Coleoptera: Tenebrionidae), were evaluated under laboratory conditions. For this purpose, LC₅₀, repellency, antifeedant properties, and some biological effects (including body weight, immature developmental time and survival) of the insect were determined. The aqueous and ethanolic extracts were highly toxic to the larvae and adults. Calculated LC₅₀ values ranged between 2.6 and 193.4 (g/kg). Both extracts had repellent and antifeedant properties against the adults. The extracts adversely affected the larval and pupal weights, developmental time, and survival. Aqueous extracts were more effective only for the LC₅₀ values and only in females. All other measured parameters do not differ between the two extracts. The bioactive properties might be related to high amounts of alkaloid, phenol and tannin. Aqueous extract of the plant leaves may be a useful alternative for chemical insecticides.     

Phytotoxic activity of Cleome arabica L. and its principal discovered active compounds

The aim of this study was to assess the phytotoxic potential of Cleome arabica L, as well as to isolate the main bioactive compounds. Phytotoxicity was evaluated on germination and seedling growth of Lactuca sativa, Raphanus sativus, Peganum harmala and Silybum marianum, through testing aqueous and organic extracts of different C. arabica organs (roots, shoots, siliquae and seeds). Results showed that siliquae methanol extract caused the greatest negative effect on lettuce germination and growth. For the bioactive subfractions (petroleum ether, ethyl acetate and methanol–water), the ethyl acetate induced highly significant reduction, showing 100% inhibition of lettuce growth at 6 g/L. The bioactive ethyl acetate subfraction was chromatographed and subjected to NMR techniques. Based on bio-guided chromatographic fractionation, five bioactive allelochemical compounds were isolated from ethyl acetate extract of siliquae of C. arabica. The most inhibitory compound on lettuce seedling growth was elucidated as 11-α-acetylbrachy-carpone-22(23)-ene.     

Pest-managing activities of plant extracts and anthraquinones from Cassia nigricans from Burkina Faso

Insecticidal activity of eight plants collected from Burkina Faso was studied using mosquito (Ochlerotatus triseriatus), Helicoverpa zea and Heliothis virescens larvae and adult white fly (Bemisia tabaci). The n-hexane, ethyl acetate and methanol extracts of Pseudocedrela kotschyi, Strophantus hispidus, Securidaca longepedunculata, Sapium grahamii, Swartzia madagascariensis, Cassia nigricans, Jatropha curcas and Datura innoxia were used in this study. Extracts were tested at 250 microg/mL concentration. All three extracts of C. nigricans, J. curcas (skin and seeds) and D. innoxia exhibited 100% mortality on fourth instar mosquito (O. triseriatus) larvae. In addition, the n-hexane and ethyl acetate extracts of S. hispidus, S. longepedunculata, S. grahamii showed 100% mortality. The ethyl acetate extract of S. madagascariensis was the most active on adult white fly and exhibited 80% mortality. Extracts of all other plants exhibited 30-50% mortality on B. tabaci. In the antifeedant assays against H. zea and H. virescens, the MeOH extracts of C. nigricans, S. madagascarensis and S. hispidus were more effective against H. zea as indicated by 74% larval weight reduction as compared to the control. Since C. nigricans is commonly used in West Africa to protect grain storage from insects, we have characterized the insecticidal components present in its extract. Bioassay directed isolation of C. nigricans leaf extract yielded anthraquinones emodin, citreorosein, and emodic acid and a flavonoid, luteolin. Emodin, the most abundant and active anthraquinone in C. nigricans showed approximately 85% mortality on mosquito larvae Anopheles gambiaea and adult B. tabaci at 50 and 25 microg/mL, respectively, in 24 h. These results suggest that the extract of C. nigricans has the potential to be used as an organic approach to manage some of the agricultural pests.     

Growth regulatory and growth inhibitory effects of Thevetia neriifolia stem extracts on Helicoverpa armigera (Lepidoptera: Noctuidae)

Abstract

Helicoverpa armigera, a serious global destructive pest of agricultural crops, is on continuous rise despite several control measures undertaken. The detrimental effects of these measures have created a dire need to explore alternate eco-safe strategies. The present study investigates the growth-regulatory and growth-arrest potential of hexane and methanol extracts of Thevetia neriifolia stems against H. armigera. Investigations revealed that larval feeding and rearing on the extract-containing diet did not result in appreciable larval mortality but delayed the larval growth and development. Both the extracts demonstrated dose-dependent effects exhibiting negative correlation between the weights gained by developmental stages and the extract concentrations. Feeding with extracts also resulted in formation of few larval–pupal and pupal–adult intermediates and significantly reduced percent adult emergence of H. armigera. Current study, however, revealed higher growth inhibitory potential of methanol extracts as compared with hexane extracts. The study attempts to provide an eco-friendly approach for H. armigera management.     

Antiulcerogenic activity of crude extract, fractions and populnoic acid isolated from Austroplenckia populnea (Celastraceae)

Many plant crude extracts and their isolated compounds are the most attractive sources of new drugs and show promising results for the treatment of gastric ulcers. Austroplenckia populnea is commonly known as "marmelinho-do campo, mangabeira-brava, mangabarana and vime" and it has been used in folk medicine as anti-dysenteric and anti-rheumatic. Powdered bark wood (3.25 kg) was macerated with aqueous ethanol (96%) and the extract was concentrated under reduced pressure to yield 406 g of crude hydralcoholic extract. The hydralcoholic extract was suspended in aqueous methanol and partitioned with hexane, chloroform and ethyl acetate (EtOAc) in sequence, yielding 8.0 g, 9.5 g and 98.17 g of crude extracts, respectively. Chromatography of the hexane extract over a silica gel column led to the isolation of the triterpene populnoic acid. The oral administration of hydralcoholic, hexane, chloroform and EtOAc extracts (200 mg/kg) decreased the ulcer lesion index (ULI) by 83.15%, 46.87%, 32.2%, 68.12%, respectively. Oral administration of populnoic acid (100 mg/kg) diminished the ULI by 55.29%. All the obtained results were significant in comparison with the negative control, with exception of the chloroform extract.     

Antimicrobial activity and phytochemical screening of Ocimum americanum L extracts against pathogenic microorganisms

The aim of the present study is to assess the antimicrobial activities of various leaf extracts of Ocimum americanum were tested against pathogenic microorganisms. Preparation of different extracts viz., aqueous, acetone, ethyl acetate and methanol through soxhlet extraction method. Various extracts were investigated against MTCC strains of Bacillus cereus, Clostridium penfrigens, Klebsilla pnemoniae, Salmonella paratyphi, Candida albicans and Aspergillus niger by agar well diffusion and disc diffusion methods. Minimum inhibitory concentration (MIC), Minimum Bactericidal/Fungicindal Concentration (MBC/MFC) were determined through micro dilution method. Elucidation of phytochemicals and functional groups were observed by HPLC and FT-IR respectively. Ethyl acetate leaf extract of O.americanum showed significant antimicrobial activity against the all tested pathogens in agar well diffusion method in which B.cereus (17 mm) was observed high zone of inhibition. Whereas lowest inhibition was observed in aqueous extract against C.pentrigens (7 mm). The ranges of MIC values from 0.78 μg/ml to 50 μg/ml and MBC/MFC 1.56 μg/ml to 50 μg/ml were observed. Phytochemicals such as alkaloids, steroids, saponins, flavonoids, tannins, terepenes, phenolic compounds cardiac glycosides were detected. Saponinns, flavonoids, tannins, phenolic compounds were observed in only ethyl acetate leaf extracts. Functional group of the leaf extracts was exhibited by FTIR and HPLC analysis of the ethyl acetate leaf extract was elutated at six peaks. Based on the results we concluded that ethyl acetate leaf extract of O.americanum has proved to be potentially effective than the other extracts. Therefore, ethyl acetate leaf extract of O.americanum could act as antimicrobial agent and further studies are recommended for isolation of compounds and toxicological studies.     

Cytotoxic,hypoglycemic activity and phytochemical analysis of Rubus imperialis (Rosaceae)

Rubus imperialis, Artemia salina, 3-O-methylellagic-4'-O-alpha-rhamnose Acid Screening of different extracts, fractions and compounds from Rubus imperialis Chum. Schl. (Rosaceae) has been conduced using the brine shrimp microwell cytotoxicity assay. Three parts of the plant (methanolic extract from leaves, roots and stems), three fractions from roots (hexane, ethyl acetate and butanol) and three isolated compounds (niga-ichigoside F1, 23-hydroxytormentic acid, ellagic acid derivative) were tested. The most promising material (LC50 <1000 microg/ml) were the methanolic extract and ethyl acetate fraction from roots. However, there was little correlation observed in the degree of toxicities observed between the isolated compounds. On the other hand, the cytotoxicity and in vivo assays confirmed the hypoglycemic activity of methanolic extract and validated the Brazilian popular use of R. imperialis as an antidiabetic agent.