Maria José Moreira Batatinha

In vitro ovicidal and larvicidal activity of Agave sisalana Perr. (sisal) on gastrointestinal nematodes of goats.

This study describes the in vitro anthelmintic activity of aqueous extracts (AE), ethyl acetate extracts (EE), flavonoid fractions (FF) and saponin fractions (SF) obtained from sisal waste (Agave sisalana) against gastrointestinal nematodes of goats. The activity of these extracts was evaluated by performing inhibition of egg hatch (EHA) and larval migration (LMI) assays. The EC(50) results of the EHA corresponded to 4.7, 0.1 and 0.05 mg/mL for EE, EA and FF, respectively. The SF fraction showed no ovicidal activity. The percent efficacies that were observed for the LMI were 50.3, 33.2 and 64.1% for the AE, EE and SF, respectively. The FF fraction did not show activity against the larvae. The analysis of the FF fraction indicates the presence of a homoisoflavonoid. This report suggests that the A. sisalana has activity in vitro against gastrointestinal nematodes of goats. This effect is likely related to the presence of homoisoflavonoid and saponin compounds, which have different actions for specific stages of nematode development.

In vivo anthelmintic activity of an aqueous extract from sisal waste (Agave sisalana Perr.) against gastrointestinal nematodes in goats.

The resistance of gastrointestinal nematodes (GINs) of small ruminants to anthelmintics has required the investigation of new alternatives. The aim of the present study was to evaluate the in vivo anthelmintic activity of an aqueous extract from sisal waste (Agave sisalana) (AESW) against GINs in goats and to observe the animals for toxic effects. Thirty animals that were naturally infected with GINs were distributed into three groups: group I, was treated with daily doses of AESW (1.7 g/kg) for eight days; Group II, the positive control, was treated with a single dose of levamisole phosphate (6.3mg/kg); and group III, the negative control, was left untreated. Faecal eggs counts (FECs), coprocultures and post-mortem worm counts were performed to assess the efficacy of the treatments. Clinical and laboratory analyses were performed to evaluate any toxic effects associated with the treatment. In the goats in groups I and II, a significant reduction (p<0.05) of the number of eggs and infective larvae (L(3)) was observed. The maximum reductions of the FECs were 50.3% and 93.6% for groups I and II, respectively, whereas the percent reductions of the total number of L(3) larvae were 80% (group I) and 85.6% (group II). There was no difference between groups I and III with respect to worm burden, and the percent reductions were 28.8% and 63.4% for Oesophagostomum columbianum and Trichostrongylus colubriformis, respectively. No reduction was detected for the Haemonchus contortus. The positive control group demonstrated a 74% reduction of the parasites that were recovered from the digestive tract. There were no changes in clinical and haematological parameters. The levels of serum urea and creatinine were higher in group I, but remained within the normal range. At necropsy, pale mucous membranes, abomasitis and enteritis were associated with parasitism. In addition, a histological analysis of the liver and kidney did not reveal any changes suggestive of toxicity. A chemical analysis of the AESW demonstrated the presence of saponins, which after acid-hydrolyses reaction, gave the sapogenins hecogenin and tigogenin. The AESW had a low efficacy for the parasitic stages and was moderately effective against eggs and free-living stages. Furthermore, the treatment was not toxic to the goats.

Action of sisal (Agave sisalana, Perrine) extract in the in vitro development of sheep and goat gastrointestinal nematodes.

Active compounds from Agave sisalana with antiparasitic action against gastrointestinal nematodes (GINs) could be an alternative to diversify the range of parasite management methods in the livestock sector. The objective of this study was to evaluate the in vitro action of A. sisalana extract on the development of sheep and goat GINs. The extract, obtained from shredded sisal leaves, was utilized at various concentrations in the egg hatch test (EHT), larval development test (LDT), larval feeding inhibition test (LFIT) and adult motility test (AMT). The LC(50) and LC(95) in the EHT were 6.90 and 24.79 mg/mL, in the LDT were 0.041 and 0.067 mg/mL and in the LFIT were 0.053 and 0.24 mg/mL, respectively, showing a dose-dependent relationship. The development and feeding inhibition on L(1) were both 100% at a dose of 0.12 mg/mL. In the AMT there was 100% inhibition at 75 mg/mL after 24h of exposure. The extract of A. sisalana therefore demonstrated significant action on L(1) at 0.12 mg/mL. So, if part of the A. sisalana extract passes through the animals gastrointestinal system, this material can have a significant effect on the parasites in the feces. This is an interesting approach because it can drastically reduce the pasture contamination as well as the infection of herds.

Genotoxicity and morphological changes induced by the alkaloid monocrotaline, extracted from Crotalaria retusa, in a model of glial cells

Plants of Crotalaria genus (Leguminosae) present large amounts of the pyrrolizidine alkaloid monocrotaline (MCT) and cause intoxication to animals and humans. Therefore, we investigated the MCT-induced cytotoxicity, morphological changes, and oxidative and genotoxic damages to glial cells, using the human glioblastoma cell line GL-15 as a model. The comet test showed that 24h exposure to 1-500microM MCT and 500microM dehydromonocrotaline (DHMC) caused significant increases in cell DNA damage index, which reached 42-64% and 53%, respectively. Cells exposed to 100-500microM MCT also featured a contracted cytoplasm presenting thin cellular processes and vimentin destabilisation. Conversely, exposure of GL-15 cells to low concentrations of MCT (1-10microM) clearly induced megalocytosis. Moreover, MCT also induced down regulation of MAPs, especially at the lower concentrations adopted (1-10microM). Apoptosis was also evidenced in cells treated with 100-500microM MCT, and a later cytotoxicity was only observed after 6 days of exposure to 500microM MCT. The data obtained provide support for heterogenic and multipotential effects of MCT on GL-15 cells, either interfering on cell growth and cytoskeletal protein expression, or inducing DNA damage and apoptosis and suggest that the response of glial cells to this alkaloid might be related to the neurological signs observed after Crotalaria intoxication.

Atividade anti-helmíntica do extrato aquoso das folhas de Zanthoxylum rhoifolium Lam. (Rutaceae)

The study evaluated the anthelmintic activity of aqueous extract of Zanthoxylum rhoifolium leaves in two experiments. In vitro test, cultures of goat fecal samples were treated with different concentrations of extract (134.5 to 335.0 mg.mL(-1)). In vivo test was composed of 20 sheep: G1: treated with 0.63 g.kg(-1), during four days; G2: same dose, for eight days; G3: ivermectin (200 microg.kg(-1)) and G4 untreated group. In vitro results showed a reduction of Haemonchus spp, Trichostrongylus spp. and Oesophagostomum spp. larvae greater than 95% in the concentrations between 335.0 and 193.7 mg.mL(-1). Faecal egg counting reduction was 51, 56 and 90% in G1, G2 and G3, respectively, while immature stages and adults ranged from 0 to 91% in G1 and from 26 to 94% in G2. Ivermectin effectiveness was 99% for L4 and L5 of H. contortus and 100% for other nematodes species. Clinical and biochemical parameters have remained in the normality and histophatologic analyses did not show alteration suggesting absence of toxicity. Although the great effectiveness of Z. rhoifolium leaves extract in vitro test, it displayed poor efficiency in vivo regarding gastrointestinal nematodes reduction.

Evaluation of anthelmintic activity of liquid waste of Agave sisalana (sisal) in goats

It was evaluated the anthelmintic activity of Agave sisalana (sisal) juice against gastrointestinal nematodes and its potential toxic effects in goats. In vitro tests showed more than 95% reduction in larval counts of the genus Haemonchus spp. at concentrations between 86.5 and 146.3 mg.mL(-1). In vivo the percent reduction of larvae of the fourth (L4) and fifth (L5) stages of Haemonchus, Oesophagostomum and Trichostrongylus was less than 95% in groups GI and GII, and between 80 and 90% in group GIII. A. sisalana juice at the concentrations tested in vitro was effective against gastrointestinal nematodes in goats; however, its anthelmintic efficacy was reduced when administered to animals.

Efeitos do suco de alho (Allium sativum Linn.) sobre nematódeos gastrintestinais de caprinos

The objective of this work was to evaluate the effects of the garlic juice on goat gastrointestinal nematodes. Twenty animals were alocated into two groups and treated with garlic juice. One group was treated with ivermectin and one control group was not treated. The percentage reductions in egg and larvae counts of the Strongyloidea were under 95% for all groups. The treatment of goats with garlic juice was not an effective control of gastrointestinal nematodes.

In vitro anthelmintic and cytotoxicity activities the Digitaria insularis (Poaceae)

This study aimed to evaluate the in vitro activity of D. insularis extracts and fractions against gastrointestinal nematodes of goats and its cytotoxicity on Vero cells. The egg hatch (EHT) and larval motility (LMT) tests were conducted to investigate the anthelmintic effects of the crude hydroethanolic (CH), ethyl acetate (EA), butanolic (BT) and residual hydroethanolic (RH) extracts. The elution of the active extract (EA) on column chromatography (SiO2) using organic solvents furnished six fractions (FR1 to FR6), which were also tested. Cytotoxicity was determined using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and Trypan Blue exclusion assays. All extracts, FR2 and FR3, inhibited egg hatching in a concentration-dependent manner. The EHT led to EC50 values (effective concentration 50%) of 0.64; 0.69; 0.77; 0.96; 0.27 and 0.65mg/mL for CH, EA, BT, RH, FR2 and FR3, respectively. However, the extracts exhibited low effect on the motility of L3. In the cytotoxicity evaluation (MTT assay), the IC50 (inhibitory concentration 50%) was 1.18 (EA), 1.65 (FR2) and 1.59mg/mL (FR3), which was relatively high (low toxicity) in comparison to the EC50 values in EHT, mainly for FR2. The chemical analyses of most active fractions (FR2) by Liquid Chromatography coupled to Mass Spectrometry (LC-MS) led the characterization of the flavones tricin and diosmetin. These results showed the high anthelmintic effect and low cytotoxicity of D. insularis and also that the flavones can be probably responsible for the nematocidal activity of this plant.

In vitro anthelmintic activity of the Zizyphus joazeiro bark against gastrointestinal nematodes of goats and its cytotoxicity on Vero cells.

This study examined the in vitro effect of the Zizyphus joazeiro bark against gastrointestinal nematodes of goats and its cytotoxicity on Vero cells. The ovicidal activity of the crude hydroethanolic extract (CE), its partitioned hexane (HE) and aqueous extract (AE) and saponins fraction (SF), including betulinic acid (BA), a biogenetic compound from this plant found in HE, were investigated using the inhibition of egg hatch assay (EHA). Thereafter, the extracts and the SF were evaluated through the larval motility assay (LMA) and larval migration inhibition assay (LMIA). The AE and SF promoted a complete inhibition of the egg hatch, and the effective concentration to inhibit 50% (EC50) values was 1.9 and 1.3mg/mL, respectively. The highest percentages of inhibition in EHA observed after treatments with CE, HE and BA corresponded to 79, 48 and 17%, respectively. The extracts and SF did not show larvicidal activity in LMA and LMIA. The AE and SF demonstrated cytotoxic effects in 3-4,5-dimethylthiazol-2-yl, 2,5diphenyltetrazolium bromide (MTT) and trypan blue tests; however, SF was more toxic (50% inhibitory concentration, IC50=0.20mg/mL). The chemical characterization of the SF was made through Proton Nuclear Magnetic Resonance ((1)H NMR) and Electrospray Ionization Mass Spectrometry (ESI-MS) analyses, which led to the identification of two saponins known as Joazeiroside B and Lotoside A. The results obtained from the research of this saponin content provide important information about the biological activity, especially the anthelmintic effect present in the plant investigated. That also suggests the types of bioactive compounds that may be responsible for this antiparasitic activity exhibited by the plant extracts.

In vitro acaricide and anticholinesterase activities of digitaria insularis (Poaceae) against Rhipicephalus (Boophilus) microplus

Medicinal plants have been proposed as an alternative for acaricide control, aiming to develop lower-cost and eco-friendly ectoparasiticide products. The aim of this study was to evaluate the in vitro activity of the extracts and fractions obtained from the leaves of Digitaria insularis on the reproductive efficacy of the bovine tick Rhipicephalus (Boophilus) microplus. Also, we investigated the possible relation with the anticholinesterase mechanism. The effect of the crude hydroethanolic (CH), hexanic (HE), ethyl acetate (EA), butanolic (BT) and residual hydroethanolic (RH) extracts, as well as four fractions of HE, were evaluated using adult immersion test. Only the HE and EA extracts (50 mg/mL) and fraction 2 (Fr2) (12.5 mg/mL) promoted reduction of the reproductive parameters (oviposition and hatching rate) greater than 90% and were not statistically different from the positive control. Higher reproductive activity was recorded in Fr2 with a lower effective concentration (EC50) value (6.65 mg/mL) than in HE (17.8 mg/mL) and EA (23.97 mg/mL). The anticholinesterase activity was assessed through spectrophotometry in microtiter assays, with enzymatic inhibition of 34.8, 43.2 and 57.9% of the HE, AE and Fr2, respectively. The chemical evaluation of the Fr2 was carried through Gas Chromatography coupled to Mass Spectrometry (GC-MS) and led to the characterization of nine compounds classified as fatty acids (3), esterified fatty acids with long-chain alcohol (4) and terpene (1). The effect of D. insularis extracts and fractions was focused on female reproductive parameters such as oviposition and hatching rates. The results obtained in this study suggest that D. insularis shows an in vitro acaricidal activity against R. (B.) microplus. Such action might be associated with the presence of secondary metabolites identified in the Fr2.