Evaluation of seeds ethanolic extracts of Triplaris gardneriana Wedd. using in vitro and in vivo toxicological methods.

Triplaris gardneriana Wedd. is a tree used in folk medicine to treat venereal diseases and inflammation as well as a source of biological compounds with antioxidant capacity. In order to assess the safety of these bioactive compounds, the present study aimed to determine the toxicity of an ethanolic extract of T. gardneriana, (EETg). Toxicological tests included hemolytic activity, toxicity toward the brine shrimp Artemia, cytotoxicity against breast cancer cells (MCF7) and acute oral toxicity in rodents. In addition, toxicogenomics techniques were used to determine genome expression in MCF7 cells exposed to EETg. The results showed that the extract exhibits approximately 60% of hemolytic activity at the highest tested concentration (64 µg/ml) and toxicity against nauplii of Artemia sp. (LC50 of 67.85 µg/ml). Further, EETg appears to be cytotoxic to MCF7 (cell viability reduced to 40% at 250 µg/ml after 24 hr). Genomic data demonstrated differential expression of 14 genes. Data analysis indicated possible altered pathways (e.g., xenobiotic metabolism), possible adverse health risks (e.g., hepatotoxicity), and drugs with similar gene expression profile (e.g., antimicrobials). The investigation provides important information on potentially adverse aspects of EETg, which need to be considered prior to the therapeutic utilization of this plant.Abbreviations: EETg: ethanolic extract of T. gardneriana seeds; MCF7: michigan cancer foundation-7 which refers to a human breast cell line (adenocarcinoma); NGS: next-generation sequencing; edgeR: empirical analysis of digital gene expression data in R; Consensus: consensus path database; FDR: false discovery rate; NCBI: national center for biotechnology information; KEGG: kyoto encyclopedia of genes and genomes; Ingenuity: ingenuity pathway analysis software; CMAP: connectivity map; OECD: organization for economic co-operation and development; HL-60: human promyelocytic leukemia cells; PC3: prostate cancer cells.

Proteomics analysis of zebrafish larvae exposed to 3,4-dichloroaniline using the fish embryo acute toxicity test.

The zebrafish (Danio rerio) is a small teleost fish that is becoming increasingly popular in laboratories worldwide and several attributes have also placed the zebrafish under the spotlight of (eco)toxicological studies. Since the 1990s, international organizations such as ISO and OECD have published guidelines for the use of zebrafish in ecotoxicological assessment of environmental toxicants such as the Fish Embryo Acute Toxicity (FET) test, OECD n° 236 guideline. This protocol uses 3,4-dichloroaniline (DCA), an aniline pesticide whose toxicity to fish species at early life stages is well known, as a positive control. Despite its use, little is known about its molecular mechanisms, especially in the context of the FET test. Therefore, this study aimed to investigate such changes in zebrafish larvae exposed to DCA (4 mg/L) for 96 hours using gel-free proteomics. Twenty-four proteins detected in both groups were identified as significantly affected by DCA exposure, and, when considering group-specific entities, 48 proteins were exclusive to DCA (group-specific proteins) while 248 were only detected in the control group. Proteins modulated by DCA treatment were found to be involved in metabolic processes, especially lipids and hormone metabolism (eg, Apoa1 and Apoa1b and vitelogenins), as well as proteins important for developmental processes and organogenesis (eg, Myhc4, Acta2, Sncb, and Marcksb). The results presented here may therefore provide a better understanding of the relationships between molecular changes and phenotype in zebrafish larvae treated with DCA, the reference compound of the FET test.

A Protein Isolate from Moringa oleifera Leaves Has Hypoglycemic and Antioxidant Effects in Alloxan-Induced Diabetic Mice.

Moringa oleifera has been used in traditional medicine to treat diabetes. However, few studies have been conducted to relate its antidiabetic properties to proteins. In this study, a leaf protein isolate was obtained from M. oleifera leaves, named Mo-LPI, and the hypoglycemic and antioxidant effects on alloxan-induced diabetic mice were assessed. Mo-LPI was obtained by aqueous extraction, ammonium sulphate precipitation and dialysis. The electrophoresis profile and proteolytic hydrolysis confirmed its protein nature. Mo-LPI showed hemagglutinating activity, cross-reaction with anti-insulin antibodies and precipitation after zinc addition. Single-dose intraperitoneal (i.p.) administration of Mo-LPI (500 mg/kg·bw) reduced the blood glucose level (reductions of 34.3%, 60.9% and 66.4% after 1, 3 and 5 h, respectively). The effect of Mo-LPI was also evidenced in the repeated dose test with a 56.2% reduction in the blood glucose level on the 7th day after i.p. administration. Mo-LPI did not stimulate insulin secretion in diabetic mice. Mo-LPI was also effective in reducing the oxidative stress in diabetic mice by a decrease in malondialdehyde level and increase in catalase activity. Mo-LPI (2500 mg/kg·bw) did not cause acute toxicity to mice. Mo-LPI is a promising alternative or complementary agent to treat diabetes.

AOP Report: Glutathione Conjugation Leading to Reproductive Dysfunction via Oxidative Stress.

We propose an adverse outcome pathway (AOP) for reproductive dysfunction via oxidative stress (OS). The AOP was developed based on Organisation for Economic Co-operation and Development (OECD) Guidance Document 184 and on the specific considerations of the OECD users' handbook supplement to the guidance document for developing and assessing AOPs (no. 233). According to the qualitative and quantitative experimental data evaluation, glutathione (GSH) conjugation is the first upstream key event (KE) of this AOP to reproductive dysfunction triggering OS. This event causes depletion of GSH basal levels (KE2 ). Consequently, this drop of free GSH induces an increase of reactive oxygen species (KE3 ) generated by the natural cellular metabolic processes (cellular respiration) of the organism. Increased levels of these reactive species, in turn, induce an increase of lipid peroxidation (KE4 ). This KE consequently leads to a rise in the amount of toxic substances, such as malondialdehyde and hydroxynonenal, which are associated with decreased quality and competence of gamete cell division, consequently impairing fertility (KE5 and adverse outcome). The overall assessment of the general biological plausibility, the empirical support, and the essentiality of KE relationships was considered as high for this AOP. We conclude that GSH conjugation is able to lead to reproductive disorder in fishes and mammals, via OS, but that the amount of stressor needed to trigger the AOP differs between stressors. Environ Toxicol Chem 2023;42:2519-2528. © 2023 SETAC.

A brief report on some health aspects of rats fed with crescent levels of recombinant chagasin, a potential plant defense protein.

Chagasin may be considered a potential plant-incorporated protectant (PIP) protein due to its deleterious effects on insect pests. However, extensive safety studies with PIP's are necessary before introducing them into the target plant. Thus, a short-term feeding trial in rats with high doses of r-chagasin was conducted to provide evidences about its safety. Three test diets containing casein + r-chagasin (0.25, 0.5 and 1% of total protein) were offered to rats (10 days). The test diets did not show adverse effects upon the development, organ weight, hematological parameters and serum protein profiles of rats, providing preliminary information on the safety of r-chagasin.

Neutralizing Effect of Synthetic Peptides toward SARS-CoV-2.

The outbreak caused by SARS-CoV-2 has taken many lives worldwide. Although vaccination has started, the development of drugs to either alleviate or abolish symptoms of COVID-19 is still necessary. Here, four synthetic peptides were assayed regarding their ability to protect Vero E6 cells from SARS-CoV-2 infection and their toxicity to human cells and zebrafish embryos. All peptides had some ability to protect cells from infection by SARS-CoV-2 with the D614G mutation. Molecular docking predicted the ability of all peptides to interact with and induce conformational alterations in the spike protein containing the D614G mutation. PepKAA was the most effective peptide, by having the highest docking score regarding the spike protein and reducing the SARS-CoV-2 plaque number by 50% (EC50) at a concentration of 0.15 mg mL-1. Additionally, all peptides had no toxicity to three lines of human cells as well as to zebrafish larvae and embryos. Thus, these peptides have potential activity against SARS-CoV-2, making them promising to develop new drugs to inhibit cell infection by SARS-CoV-2.

Study of the antiproliferative potential of seed extracts from Northeastern Brazilian plants.

This study assessed the antiproliferative and cytotoxic potential against tumor lines of ethanolic seed extracts of 21 plant species belonging to different families from Northeastern Brazil. In addition, some underlying mechanisms involved in this cytotoxicity were also investigated. Among the 21 extracts tested, the MTT assay after 72 h of incubation demonstrated that only the ethanolic extract obtained from Myracrodruon urundeuva seeds (EEMUS), which has steroids, alkaloids and phenols, showed in vitro cytotoxic activity against human cancer cells, being 2-fold more active on leukemia HL-60 line [IC(50) value of 12.5 (9.5-16.7) µg/mL] than on glioblastoma SF-295 [IC(50) of 25.1 (17.3-36.3) µg/mL] and Sarcoma 180 cells [IC(50) of 38.1 (33.5-43.4) µg/mL]. After 72h exposure, flow cytometric and morphological analyses of HL-60-treated cells showed that EEMUS caused decrease in cell number, volume and viability as well as internucleosomal DNA fragmentation in a dose-dependent way, suggesting that the EEMUS triggers apoptotic pathways of cell death.

Water extracts of Brazilian leguminous seeds as rich sources of larvicidal compounds against Aedes aegypti L.

This study assessed the toxicity of seed water extracts of 15 leguminous species upon Aedes aegypti larvae. A partial chemical and biochemical characterization of water extracts, as well as the assessment of their acute toxicity in mice, were performed. The extracts of Amburana cearensis, Anadenanthera macrocarpa, Dioclea megacarpa, Enterolobium contortisiliquum and Piptadenia moniliformis caused 100% of mortalit y after 1 to 3 h of exposure. They showed LC(50) and LC(90) values ranging from 0.43 ± 0.01 to 9.06 ± 0.12 mg/mL and from 0.71 ± 0.02 to 13.03 ± 0.15 mg/mL, respectively. Among the secondary metabolite constituents, the seed water extracts showed tannins, phenols, flavones, favonols, xanthones, saponins and alkaloids. The extracts also showed high soluble proteins content (0.98 to 7.71 mg/mL), lectin (32 to 256 HU/mL) and trypsin inhibitory activity (3.64 = 0.43 to 26.19 = 0.05 gIT/kg of flour) The electrophoretic profiles showed a great diversity of protein bands, many of which already described as insecticide proteins. The extracts showed low toxicity to mice (LD(50) > 0.15 = 0.01 g/kg body weight), but despite these promising results, further studies are necessary to understand the toxicity of these extracts and their constituents from primary and secondary metabolism upon Ae. aegypti.

Identification, characterization, and antifungal activity of cysteine peptidases from Calotropis procera latex.

Cysteine peptidases (EC 3.4.22) are the most abundant enzymes in latex fluids. However, their physiological functions are still poorly understood, mainly related to defense against phytopathogens. The present study reports the cDNA cloning and sequencing of five undescribed cysteine peptidases from Calotropis procera (Aiton) Dryand (Apocynaceae) as well as some in silico analyses. Of these, three cysteine peptidases (CpCP1, CpCP2, and CpCP3) were purified. Their enzymatic kinetics were determined and they were assayed for their efficacy in inhibiting the hyphal growth of phytopathogenic fungi. The mechanism of action was investigated by fluorescence and atomic force microscopy as well as by induction of reactive oxygen species (ROS). The deduced amino acid sequences showed similar biochemical characteristics and high sequence homology with several other papain-like cysteine peptidases. Three-dimensional models showed two typical cysteine peptidase domains (L and R domains), forming a "V-shaped" active site containing the catalytic triad (Cys, His, and Asn). Proteolysis of CpCP1 was higher at pH 7.0, whereas for CpCP2 and CpCP3 it was higher at 7.5. All peptidases exhibited optimum activity at 35 °C and followed Michaelis-Menten kinetics. However, the major difference among them was that CpCP1 exhibited highest Vmax, Km, Kcat and catalytic efficiency. All peptidases were deleterious to the two fungi tested, with IC50 of around 50 µg/mL. The peptidases promoted membrane permeabilization, morphological changes with leakage of cellular content, and induction of ROS in F. oxysporum spores. These results corroborate the hypothesis that latex cysteine peptidases play a role in defense against fungi.

Biotechnological potential of a cysteine protease (CpCP3) from Calotropis procera latex for cheesemaking.

This article reports the characterization and evaluation of the biotechnological potential of a cysteine protease purified from Calotropis procera (CpCP3). This enzyme was highly stable to different metal ions and was able to hydrolyze κ-casein similarly to bovine chymosin. Atomic force microscopy showed that the process of casein micelle aggregation induced by CpCP3 was similar to that caused by chymosin. The cheeses made using CpCP3 showed higher moisture content than those made with chymosin, but protein, fat, and ash were similar. The sensory analysis showed that cheeses made with CpCP3 had high acceptance index (>80%). In silico analysis predicted the presence of only two short allergenic peptides on the surface of CpCP3, which was highly susceptible to digestive enzymes and did not alter zebrafish embryos' morphology and development. Moreover, recombinant CpCP3 was expressed in Escherichia coli. All results support the biotechnological potential of CpCP3 as an alternative enzyme to chymosin.

Larvicidal activity of the water extract of Moringa oleifera seeds against Aedes aegypti and its toxicity upon laboratory animals.

In this work, biological effects of the water extract of Moringa oleifera seeds (WEMOS) were assessed on eggs and 3rd instar larvae of Aedes aegypti and on its toxicity upon laboratory animals (Daphnia magna, mice and rats). Crude WEMOS showed a LC50 value of 1260microg/mL, causing 99.2 +/- 2.9% larvae mortality within 24 h at 5200microg/mL, though this larvicidal activity has been lost completely at 80 masculineC/10 min. WEMOS did not demonstrate capacity to prevent egg hatching. After extensive dialyses of the crude WEMOS into watersoluble dialyzable (DF) and nondyalizable (NDF) fractions, only DF maintained its efficacy to kill larvae. Acute toxicity evaluations on daphnids (EC50 of 188.7microg/mL) and mice (LD50 of 446.5 mg/kg body weight) pointed out to low toxicity. Despite the thymus hypertrophy, WEMOS revealed to be harmless in orally and subacutelytreated rats. In conclusion, WEMOS has thermostable bioactive compounds against Ae. aegypti larvae with apparent molecular mass lower than 12 kDa and moderately toxic potential.

Insecticidal action of sodium anacardate from Brazilian cashew nut shell liquid against Aedes aegypti.

Aedes aegypti is the major vector of 1 of the most concerning arboviruses of the world, the dengue fever. The only effective way of reducing the incidence of dengue fever is to control the vector mosquito, mainly by application of insecticides to its breeding places. This study was aimed at assessing the insecticidal activity of sodium anacardate, isolated from Brazilian cashew nut shell liquid (CNSL), against the eggs, 3rd instars or pupae of Ae. aegypti. In addition, the acute toxicity of sodium anacardate to mice was also investigated. Sodium anacardate showed toxicity against Ae. aegypti eggs (median effective concentration [EC50] = 162.93 +/- 29.93 microg/ml), larvae (median lethal concentration [LC50] = 55.47 +/- 3.0 microg/ml) and pupae (LC50 = 369.78 - 52.30 microg/ml). On the other hand, even at high dose (0.3 g/kg body weight), this compound did not cause any adverse effects on mice, suggesting that this compound is safe to mammals. Therefore, sodium anacardate may be a viable low-cost alternative to help combat Ae. aegypti.

Food safety knowledge on the Bt mutant protein Cry8Ka5 employed in the development of coleopteran-resistant transgenic cotton plants.

Insecticidal Cry proteins from Bacillus thuringiensis (Bt) have been exploited in the development of genetically modified (GM) crops for pest control. However, several pests are still difficult to control such as the coleopteran boll weevil Anthonomus grandis. By applying in vitro molecular evolution to the cry8Ka1 gene sequence, variants were generated with improved activity against A. grandis. Among them, Cry8Ka5 mutant protein showed coleoptericidal activity 3-fold higher (LC50 2.83 µg/mL) than that of the original protein (Cry8Ka1). Cry8Ka5 has been used in breeding programs in order to obtain coleopteran-resistant cotton plants. Nevertheless, there is some concern in relation to the food safety of transgenic crops, especially to the heterologously expressed proteins. In this context, our research group has performed risk assessment studies on Cry8Ka5, using the tests recommended by Codex as well as tests that we proposed as alternative and/or complementary approaches. Our results on the risk analysis of Cry8Ka5 taken together with those of other Cry proteins, point out that there is a high degree of certainty on their food safety. It is reasonable to emphasize that most safety studies on Cry proteins have essentially used the Codex approach. However, other methodologies would potentially provide additional information such as studies on the effects of Cry proteins and derived peptides on the indigenous gastrointestinal microbiota and on intestinal epithelial cells of humans. Additionally, emerging technologies such as toxicogenomics potentially will offer sensitive alternatives for some current approaches or methods.

Increased Levels of Antinutritional and/or Defense Proteins Reduced the Protein Quality of a Disease-Resistant Soybean Cultivar.

The biochemical and nutritional attributes of two soybean (Glycine max (L.) Merr.) cultivars, one susceptible (Seridó) and the other resistant (Seridó-RCH) to stem canker, were examined to assess whether the resistance to pathogens was related to levels of antinutritional and/or defense proteins in the plant and subsequently affected the nutritional quality. Lectin, urease, trypsin inhibitor, peroxidase and chitinase activities were higher in the resistant cultivar. Growing rats were fed with isocaloric and isoproteic diets prepared with defatted raw soybean meals. Those on the Seridó-RCH diet showed the worst performance in terms of protein quality indicators. Based on regression analysis, lectin, trypsin inhibitor, peroxidase and chitinase appear to be involved in the resistance trait but also in the poorer nutritional quality of Seridó-RCH. Thus, the development of cultivars for disease resistance may lead to higher concentrations of antinutritional compounds, affecting the quality of soybean seeds. Further research that includes the assessment of more cultivars/genotypes is needed.

Food safety assessment of an antifungal protein from Moringa oleifera seeds in an agricultural biotechnology perspective.

Mo-CBP3 is an antifungal protein produced by Moringa oleifera which has been investigated as potential candidate for developing transgenic crops. Before the use of novel proteins, food safety tests must be conducted. This work represents an early food safety assessment of Mo-CBP3, using the two-tiered approach proposed by ILSI. The history of safe use, mode of action and results for amino acid sequence homology using the full-length and short contiguous amino acids sequences indicate low risk associated to this protein. Mo-CBP3 isoforms presented a reasonable number of alignments (>35% identity) with allergens in a window of 80 amino acids. This protein was resistant to pepsin degradation up to 2 h, but it was susceptible to digestion using pancreatin. Many positive attributes were presented for Mo-CBP3. However, this protein showed high sequence homology with allergens and resistance to pepsin digestion that indicates that further hypothesis-based testing on its potential allergenicity must be done. Additionally, animal toxicity evaluations (e.g. acute and repeated dose oral exposure assays) must be performed to meet the mandatory requirements of several regulatory agencies. Finally, the approach adopted here exemplified the importance of performing an early risk assessment of candidate proteins for use in plant transformation programs.

Insecticidal activity against Aedes aegypti of m-pentadecadienyl-phenol isolated from Myracrodruon urundeuva seeds.

BACKGROUND: Myracrodruon urundeuva Fr. Allemao is a common tree in the Caatinga that has been widely used for various medical purposes. Previous studies showed that the ethanol seed extract of M. urundeuva has potent activity against the larval stage of the dengue vector Aedes aegypti. Given this potential insecticidal activity, bioguided separation steps were performed in order to isolate the active compound(s). RESULTS: The isolation process resulted in only one active chemical compound, identified by infrared spectroscopy and mass spectrometry as m-pentadecadienyl-phenol. This compound presented potent larvicidal and pupicidal activity (LC50 10.16 and 99.06 µg mL(-1) respectively) and great egg hatching inhibitory activity (IC50 49.79 µg mL(-1)). The mode of action was investigated through observations of behavioural and morphological changes performed in third-instar larvae treated with m-pentadecadienyl-phenol solution after 1, 6, 12, 16 and 20 h of exposure. Some changes were observed as flooding of the tracheal system, alterations in siphonal valves and anal gills and lethargy, probably caused by the strong anticholinesterasic activity reported previously. CONCLUSION: The compound isolated from M. urundeuva seeds, m-pentadecadienyl-phenol, showed potent activity against immature stages of dengue vector, Ae. aegypti, being considered the main larvicidal principle.

Combination of chemical analyses and animal feeding trials as reliable procedures to assess the safety of heat processed soybean seeds.

This study assessed whether chemical analyses are sufficient to guarantee the safety of heat processing of soybeans (SB) for human/animal consumption. The effects of extrusion and dry-toasting were analyzed upon seed composition and performance of broiler chicks. None of these induced appreciable changes in protein content and amino acid composition. Conversely, toasting reduced all antinutritional proteins by over 85%. Despite that, the animals fed on toasted SB demonstrated a low performance (feed efficiency 57.8 g/100 g). Extrusion gave place to higher contents of antinutrients, particularly of trypsin inhibitors (27.53 g/kg flour), but animal performance was significantly (p < 0.05) better (feed efficiency 63.2 g/100 g). Upon the basis of chemical analyses, dry-toasting represents the treatment of choice. However, considering the results of the feeding trials, extrusion appears to be the safest method. In conclusion, in order to evaluate the reliability of any processing method intended to improve nutritional value, the combination of chemical and animal studies is necessary.

Study of the antiproliferative potential of seed extracts from Northeastern Brazilian plants

This study assessed the antiproliferative and cytotoxic potential against tumor lines of ethanolic seed extracts of 21 plant species belonging to different families from Northeastern Brazil. In addition, some underlying mechanisms involved in this cytotoxicity were also investigated. Among the 21 extracts tested, the MTT assay after 72 h of incubation demonstrated that only the ethanolic extract obtained from Myracrodruon urundeuva seeds (EEMUS), which has steroids, alkaloids and phenols, showed in vitro cytotoxic activity against human cancer cells, being 2-fold more active on leukemia HL-60 line [IC50 value of 12.5 (9.5-16.7) μg/mL] than on glioblastoma SF-295 [IC50 of 25.1 (17.3-36.3) μg/mL] and Sarcoma 180 cells [IC50 of 38.1 (33.5-43.4) μg/mL]. After 72h exposure, flow cytometric and morphological analyses of HL-60-treated cells showed that EEMUS caused decrease in cell number, volume and viability as well as internucleosomal DNA fragmentation in a dose-dependent way, suggesting that the EEMUS triggers apoptotic pathways of cell death.

Este estudo avaliou o potencial antiproliferativo e citotóxico contra linhagens de células tumorais de extratos etanólicos de sementes de vinte e uma espécies vegetais pertencentes a diferentes famílias do Nordeste brasileiro. Além disso, alguns mecanismos subjacentes envolvidos nesta citotoxidade também foram investigados. Dentre os 21 extratos testados pelo ensaio do MTT após 72 h de incubação, apenas o extrato etanólico obtido a partir de sementes de Myracrodruon urundeuva (EEMUS), o qual apresentou traços de esteróides, alcalóides e fenóis em sua composição, demonstrou atividade citotóxica in vitro contra células tumorais humanas, sendo 2 vezes mais ativo sobre a linhagem leucêmica HL-60 [IC50 valor de 12,5 (9,5-16,7) μg/mL] do que sobre células de glioblastoma SF-295 [IC50 de 25,1 (17,3-36,3) μg/mL] e de sarcoma 180 [IC50 de 38,1 (33,5-43,4) μg/mL]. Após 72 h de exposição, as análises morfológicas e por citometria de fluxo de células HL-60 tratadas com EEMUS mostraram diminuição no número de células, seu volume e viabilidade, assim como fragmentação internucleosomal do DNA de forma dose-dependente, sugerindo que a ação antiproliferativa de EEMUS pode ser ativada por vias apoptóticas.

Water extracts of Brazilian leguminous seeds as rich sources of larvicidal compounds against Aedes aegypti L.

This study assessed the toxicity of seed water extracts of 15 leguminous species upon Aedes aegypti larvae. A partial chemical and biochemical characterization of water extracts, as well as the assessment of their acute toxicity in mice, were performed. The extracts of Amburana cearensis, Anadenanthera macrocarpa, Dioclea megacarpa, Enterolobium contortisiliquum and Piptadenia moniliformis caused 100 percent of mortalit y after 1 to 3 h of exposure. They showed LC50 and LC90 values ranging from 0.43 ± 0.01 to 9.06 ± 0.12 mg/mL and from 0.71 ± 0.02 to 13.03 ± 0.15 mg/mL, respectively. Among the secondary metabolite constituents, the seed water extracts showed tannins, phenols, flavones, favonols, xanthones, saponins and alkaloids. The extracts also showed high soluble proteins content (0.98 to 7.71 mg/mL), lectin (32 to 256 HU/mL) and trypsin inhibitory activity (3.64 = 0.43 to 26.19 = 0.05 gIT/kg of flour) The electrophoretic profiles showed a great diversity of protein bands, many of which already described as insecticide proteins. The extracts showed low toxicity to mice (LD50 > 0.15 = 0.01 g/kg body weight), but despite these promising results, further studies are necessary to understand the toxicity of these extracts and their constituentsfrom primary and secondary metabolism upon Ae. aegypti.

Este trabalho objetivou avaliar a toxicidade dos extratos aquosos de sementes de 15 espécies de leguminosas contra larvas de Aedes aegypti. Foi realizada uma caracterização química e bioquímica parcial dos extratos aquosos e a avaliação da toxicidade aguda em camundongos. Os extratos de Amburana cearensis, Anadenanthera macrocarpa, Dioclea megacarpa, Enterolobium contortisiliquum e Piptadenia moniliformis causaram 100 por cento de mortalidade depois de 1 a 3 h de exposição e mostraram valores de CL50 e CL90 entre 0,43 = 0,01 e 9,06 ± 0,12 e entre 0,71 = 0,02 e 13,03 = 0,15 mg/mL, respectivamente. Dentre os constituintes do metabolismo secundário, os extratos das sementes apresentaram taninos, fenóis, flavonas, flavonóis, xantonas, saponinas e alcalóides. Os extratos apresentaram alto teor de proteínas solúveis (0,98 to 7,71 mg/mL), lectina (32 to 256 UH/mL) e inibidor de tripsina (3,64 ± 0,43 to 26,19 = 0,05 gIT/kg de farinha). O perfil eletroforéticomostrou uma grande diversidade de proteínas, muitas dasquais já descritas como inseticidas. Os extratos mostraram baixa toxicidade ao camundongo (DL50 > 0,15 = 0,01 g/kg peso corporal), porém apesar desses resultados promissores, estudos posteriores são necessários para compreender a toxicidade desses extratos e de seus constituintes do metabolismo primário e secundário sobre Ae. aegypti.

Larvicidal activity of the water extract of Moringa oleifera seeds against Aedes aegypti and its toxicity upon laboratory animals

In this work, biological effects of the water extract of Moringa oleifera seeds (WEMOS) were assessed on eggs and 3rd instar larvae of Aedes aegypti and on its toxicity upon laboratory animals (Daphnia magna, mice and rats). Crude WEMOS showed a LC50 value of 1260µg/mL, causing 99.2 ± 2.9 percent larvae mortality within 24 h at 5200µg/mL, though this larvicidal activity has been lost completely at 80ºC/10 min. WEMOS did not demonstrate capacity to prevent egg hatching. After extensive dialyses of the crude WEMOS into watersoluble dialyzable (DF) and nondyalizable (NDF) fractions, only DF maintained its efficacy to kill larvae. Acute toxicity evaluations on daphnids (EC50 of 188.7µg/mL) and mice (LD50 of 446.5 mg/kg body weight) pointed out to low toxicity. Despite the thymus hypertrophy, WEMOS revealed to be harmless in orally and subacutelytreated rats. In conclusion, WEMOS has thermostable bioactive compounds against Ae. aegypti larvae with apparent molecular mass lower than 12 kDa and moderately toxic potential.

Neste trabalho, o extrato aquoso das sementes de Moringaoleifera (EASMO) foi avaliado quanto aos seus efeitos biológicos sobre ovos e larvas de Aedes aegypti no 3ºestágio de desenvolvimento e sua toxicidade sobre animais de laboratório(Daphnia magna, camundongos e ratos). O EASMO bruto revelou uma CL50 de 1.260 µg/mL, causando 99, 2 ± 2, 9 por cento de mortalidade em 24 h na concentração de 5.200 µg/mL, embora o mesmo não tenha sido capaz de impedir a eclosão dos ovos. A atividade larvicida extinguiu-se após aquecimento do extrato a 80ºC/10 min. Diálises sucessivas do EASMO bruto resultaram em duas frações solúveis em água (Fração dializável, FD; Fração nãodializável, FND), dentre as quais apenas a FD mostrou ação larvicida. Testes de toxicidade aguda realizadosem dáfnias (CE50 de 188, 7 µg/mL) e camundongos (DL50 de446,5 mg/kg de peso corpóreo) evidenciaram baixa toxicidade. Apesar da hipertrofia tímica, o EASMO mostrou ser atóxicoapós tratamento subagudo via oral em ratos. Conclui-se, portanto, que o EASMO apresenta substâncias com capacida de larvicida contra Ae. aegypti, as quais possuem massa molecular aparente menor que 12 kDa e potencial tóxico moderado.