Comparative toxicological evaluation of carvacrol, acetylcarvacrol anda fipronil-based pesticide in human blood cells.

Plant-derived chemicals are promising substances to control arthropod pests, although synthetic ones are still the most frequently used. Thus, comparative toxicological studies are needed to determine if natural substances are safe alternatives to replace the use of synthetic chemicals. This study aimed to compare the toxicity of carvacrol (natural origin), acetylcarvacrol (semi-synthetic) and a fipronil-based pesticide (synthetic). We assessed the effects of these chemicals on hemolytic activity, erythrocytes morphology and leucocyte viability using whole blood from human subjects. Additionally, DNA damage was evaluated through comet and DNA fragmentation assays. Fipronil and carvacrol caused hemolysis at concentrations ranging from 0.5 to 2.0%, whereas acetylcarvacrol did not cause hemolysis at 0.5 and 0.75%. Fipronil and carvacrol caused severe alterations in erythrocytes' morphology at 2%, such as ghost erythrocytes, elliptocyte-like shape and rouleau-like shape, presenting only 3.3 and 8.3% normal cells, respectively, at this concentration. However, 73.3% erythrocytes incubated with 2% acetylcarvacrol exhibited normal morphology. Fipronil considerably reduced leucocytes viability, decreasing it to 78% at 2%. Carvacrol and acetylcarvacrol showed no differences in leucocyte viability for 0.5 to 1.0%, but a decrease was observed for 2% carvacrol. The comet assay showed similar DNA damage for fipronil and carvacrol, but it was significantly lower for 1 and 2% acetylcarvacrol. Incubation with genomic DNA showed that only fipronil caused fragmentation of this molecule. Thus, we conclude that carvacrol and fipronil can present similar toxicity at higher concentrations. However, acetylation of carvacrol significantly reduced its toxicity to human blood cells compared with the other chemicals.

Achyrocline satureioides (Lam) DC extracts acting as enzyme modulators: digestion, inflammation, and hemostasis / Extratos de Achyrocline satureioides (Lam) DC atuando como moduladores enzimáticos: digestão, inflamação e hemostasia

Achyrocline satureioides is popularly known for its richness in phenolic compounds and medicinal properties (anti-inflammatory, analgesic, and hepatoprotective). The present study aimed at broadening the knowledge about the pharmacological potential exerted by the aqueous and ethanolic extracts of A. satureioides. These extracts were characterized by HPLC and tested for their modulatory action on phospholipases A2 and proteases of snake venoms. In addition, they were tested on the activities of digestive enzymes. Snake venoms were used as tools since they have enzymes with high functional and structural homology to human enzymes. The results demonstrate that the extracts of A. satureioides act as enzymatic inhibitors or potentiators, interfering in processes related to the hemostasis, such as coagulation and thrombus dissolution. In addition, the anti-genotoxic activity and inhibitions exerted on digestive enzymes suggests their potential use in the prevention and/or treatment of several pathologies. New studies could provide information on how the compounds present in the extracts and the different enzymes interact.

A Achyrocline satureioides é popularmente conhecida por sua riqueza em compostos fenólicos e por suas propriedades medicinais (anti-inflamatória, analgésica e hepatoprotetora). No presente estudo, com o objetivo de ampliar o conhecimento sobre o potencial farmacológico exercido por esses extratos, os extratos aquoso e etanólico de A. satureioides foram caracterizados por HPLC e testados quanto à sua ação modulatória sobre as fosfolipases A2 e proteases de peçonhas de serpentes. Além disso, também foram testados em atividades de enzimas digestivas. As peçonhas de serpentes foram usadas como ferramentas por apresentarem enzimas com alta homologia funcional e estrutural às humanas. Os resultados demonstram que os extratos de A. satureioides atuam como inibidores ou potencializadores enzimáticos, interferindo em processos relacionados à hemostasia, como coagulação e dissolução do trombo. Além do mais, destacam seu potencial antigenotóxico e as inibições exercidas sobre as enzimas digestivas direcionando seu potencial de uso na prevenção e/ou tratamento de diversas patologias. Novos estudos poderão fornecer informações sobre os mecanismos de interação entre os compostos presentes nos extratos e as diferentes enzimas.

Thrombolysis, clotting, and genotoxic activities modulated by essential oils extracted from Lippia alba

Abstract The composition and pharmacological properties of Lippia alba (Mill.) (L. alba) (Verbenaceae) flower and leaf essential oils (EO) were determined in this study. The major constituents in the flower EO were geranial (49.83%) and neral (32.75%), and in the leaf EO were geranial (38.06%), neral (31.02%), and limonene (18.03%). Flower EO inhibited thrombolysis induced by Bothrops moojeni (B. moojeni) and Lachesis muta muta (L. muta muta) venoms (0.05-1.2 µL mL-1). When tested against L. muta muta venom, the protective effect was smaller in both EO. The EOs prolonged the clotting time induced by L. muta muta venom and a procoagulant effect was observed on B. moojeni. In the comet assay, the flower EO presented anti-genotoxic action (damage frequency of only 11.6 - 34.9%) against the L. muta muta venom. The positive control (Doxorubicin) and the venom alone presented a damage frequency of 80.3% and 70.7%, respectively. The flower EO protected DNA from damage induced by L. muta muta venom. L. alba leaf and flower EOs presented anti-genotoxic action

In vivo and in vitro prospection of the anti-ophidic properties exercised by the extracts of Jacaranda decurrens L

The research and development of alternative treatments for snakebites (e.g., medicinal plants) is necessary due to the high costs of the existing ones. The effects of the aqueous extracts from Jacaranda decurrens leaves, roots, and xylopodium were analyzed upon the venom-induced (Bothrops spp. and Crotalus spp.) systemic and local toxicity. The extracts were able to partially inhibit the phospholipase activity of the venoms from Bothrops jararacussu and Crotalus durissus terrificus. The myotoxic, edema-inducing, coagulant, and hemorrhagic activities were also inhibited. The SDS-PAGE showed that the venom proteins were intact after their incubation with the extracts. This suggests that the possible mechanism of inhibition is not related to the degradation of the protein but rather to their binding to specific sites of the enzymes. The extracts significantly prolonged the survival time of animals in the lethality assay performed with Crotalus durissus terrificus venom and its toxin (crotoxin). The anti-ophidic activity of medicinal plants may aid in the management of snakebites in distant locations by reducing the victim’s local effects and time to heal.

Mitochondriotropic action and DNA protection: Interactions between phenolic acids and enzymes.

The protective action of caffeic (CA) and syringic (SA) acids on the genotoxicity exercised by snake venoms was investigated in this study. Molecular interactions between phenolic acids and the enzyme succinate dehydrogenase were also explored. In the electrophoresis assay, SA did not inhibit the genotoxicity induced by the venom. However, CA partially inhibited DNA degradation. In the comet assay, SA and CA exerted an inhibitory effect on the venom-induced fragmentation. Succinate dehydrogenase presented, in computational analyzes, favorable energies to the molecular bond to both the malonic acid and the phenolic compounds evaluated. In the enzymatic activity assays, SA inhibited succinate dehydrogenase and interfered in the interaction of malonic acid. Meanwhile, CA potentiated the inhibition exerted by the malonic acid. The results suggest transient interactions between toxins present in venoms and phenolic acids, mainly by hydrogen interactions, which corroborate with the data from previous works.

Enzymes modulation by dried grape pomace from the manufacture of wines and juices

The processing of grapes for the manufacture of juices and wines, generates large quantities of by-products rich in metabolites with antioxidant, antimicrobial, anti-inflammatory and cicatrizing activities. The high homology between human enzymes and snake venoms makes the latter valuable laboratory tools for the study of pathophysiological processes. Proteases and phospholipases A2 act in processes related to hemostasis and inflammatory response. Thus, in this work, dried pomace obtained from grape (Isabel, Niagara, Bordô, BRS Violeta and Blend cultivars) processing were evaluated on phospholipase, proteolytic, hemolytic and thrombolytic activities induced by snakes venoms and the content of phenolic compounds and minerals was evaluated. The dried pomace exerted inhibitory and potentiating actions in all analyzed activities. The enzymatic modulators present in the evaluated dried pomace have potential for therapeutic use, although their broad characterization is still necessary, in order to define adequate amounts and formulations to obtain efficacy and safety in their use.

Inhibition by essential oils of Mentha viridis and Mentha pulegium (Lamiaceae) in proteolysis, fibrinogenolysis and coagulation caused by venomous snakes

Snake venoms are widely used as laboratory tools for studies of physiological, pharmacological and toxicological mechanisms. Venoms used here are rich sources of several classes of proteases that act on factors of the coagulation cascade, fibrinogenolysis and fibrinolysis, altering the hemostatic processes, and phospholipases A2 which are involved mainly in inflammatory and clotting processes. Natural products such as essential oils are made up of active ingredients with wide application in the food, pharmaceutical and cosmetic industries. Thus, this study evaluates the essential oils from Mentha viridis and Mentha pulegium on coagulation, fibrinogenolysis and degradation of azocasein, induced by Bothrops sp and Lachesis muta muta venoms. These oils were achieved by hydrodistillation and presented, respectively, as the main constituents linalool (40.70 %), carvone (13.52 %) and α-terpinene (8.56 %); pulegone (50.01 %), menthol (31.90 %) and menthone (16.56 %). The essential oils were previously incubated with Bothrops alternatus venom, for two different times, then plasma was added and time was recorded. The M. viridis oil presented greater anticlotting potential. Meanwhile, the M. pulegium oil presented anticlotting or proclotting activity dependent on the dose tested. The incubation time also influenced the effect of the oils on the coagulation time. At azocaseinolytic assay, the oil from M. pulegium reduced the activity for all evaluated venoms. The highest inhibition was of 39.99 %; on activity induced by B. jararacussu, M. viridis reduced the activity in 57.72 %. On B. moojeni, the major inhibition observed was of 74.67 %. The fibrinogenolysis induced by B. moojeni venom was totally inhibited by both oils in the evaluated proportions. The results show the presence in oils of protease inhibitors, considering serine and metalloproteases (acting on clotting factors or with hemorrhagic activity), as well as phospholipase A2 (enzymes involved in inflammation and clotting processes) inhibitors of wide application in medical and biotechnology areas.

Los venenos de serpientes son ampliamente utilizados como herramientas de laboratorio para estudios de mecanismos fisiológicos, farmacéuticos y toxicológicos. Los venenos son recursos ricos en diferentes clases de proteasas que actúan sobre los factores de la cascada de coagulación, fibrinogenólisis y fibrinólisis, alterando los procesos homeostáticos y las fosfolipasas A2, los cuales están involucrados en procesos inflamatorios y de coagulación. Los productos naturales como los aceites esenciales se componen de ingredientes activos de amplia aplicación en las industrias alimentaria, farmacéutica y cosmética. El presente estudio evalúa los aceites esenciales de Mentha viridis y Mentha pulegium sobre la coagulación, fibrinogenólisis y degradación de azocaseína, inducida por los venenos de Bothrops sp. y Lachesis muta muta. Los aceites esenciales fueron obtenidos por hidrodestilación y sus componentes principales fueron identificados respectivamente como el linalool (40.70 %), carvona (13.52 %) y α-terpineno (8.56 %); pulegone (50.01 %), mentol (31.9 %) y mentona (15.56 %). Los aceites esenciales fueron previamente incubados con veneno de Bothrops alternatus, durante dos tiempos diferentes, luego se agregó el plasma y fue registrado el tiempo. El aceite de M. viridis presentó el más grande potencial anticoagulante. Mientras tanto, el aceite de M. pulegium presentó actividad anticoagulante o procoagulante dependiendo de la dosis evaluada. El tiempo de incubación también influyó en el efecto de los aceites sobre el tiempo de coagulación. En el ensayo azocaseínolitico, el aceite de M. pulegium redujo la actividad para todos los venenos evaluados. La inhibición más alta inducida para el veneno de B. jararacussu fue de 39.99 %, para M. viridis la actividad fue reducida en 57.72 %, mientras que para B. moojeni, la mayor inhibición observada fue de 74.67 %. La fibrinogenólisis inducida por el veneno de B. moojeni fue totalmente inhibida por ambos aceites en las proporciones evaluadas. Los resultados muestran la presencia de inhibidores de proteasas en los aceites, como las serinas y metaloproteasas (actuando sobre los factores de coagulación o sobre la actividad hemorrágica), como también fosfolipasas A2 (enzimas involucradas en los procesos de inflamación y coagulación), los cuales son inhibidores de amplia aplicación en áreas médicas y biotecnológicas.

Exploring the structural and functional aspects of the phospholipase A2 from Naja spp.

Naja spp. venom is a natural source of active compounds with therapeutic application potential. Phospholipase A2 (PLA2) is abundant in the venom of Naja spp. and can perform neurotoxicity, cytotoxicity, cardiotoxicity, and hematological disorders. The PLA2s from Naja spp. venoms are Asp 49 isoenzymes with the exception of PLA2 Cys 49 from Naja sagittifera. When looking at the functional aspects, the neurotoxicity occurs by PLA2 called ß-toxins that have affinity for phosphatidylcholine in nerve endings and synaptosomes membranes, and by α-toxins that block the nicotinic acetylcholine receptors in the neuromuscular junctions. In addition, these neurotoxins may inhibit K+ and Ca++ channels or even interfere with the Na+/K+/ATPase enzyme. The disturbance in the membrane fluidity also results in inhibition of the release of acetylcholine. The PLA2 can act as anticoagulants or procoagulant. The cytotoxicity exerted by PLA2s result from changes in the cardiomyocyte membranes, triggering cardiac failure and hemolysis. The antibacterial activity, however, is the result of alterations that decrease the stability of the lipid bilayer. Thus, the understanding of the structural and functional aspects of PLA2s can contribute to studies on the toxic and therapeutic mechanisms involved in the envenomation by Naja spp. and in the treatment of pathologies.

Snake Venom Disintegrins: An Overview of their Interaction with Integrins.

Disintegrins are non-enzymatic proteins that interfere on cell-cell interactions and signal transduction, contributing to the toxicity of snake venoms and play an essential role in envenomations. Most of their pharmacological and toxic effects are the result of the interaction of these molecules with cell surface ligands, which has been widely described and studied. These proteins may act on platelets, leading to hemorrhage, and may also induce apoptosis and cytotoxicity, which highlights a high pharmacological potential for the development of thrombolytic and antitumor agents. Additionally, these molecules interfere with the functions of integrins by altering various cellular processes such as migration, adhesion and proliferation. This review gathers information on functional characteristics of disintegrins isolated from snake venoms, emphasizing a comprehensive view of the possibility of direct use of these molecules in the development of new drugs, or even indirectly as structural models.

Mechanism of firmness loss in guava cv. Pedro Sato during ripening at room temperature

Abstract Guava (Psidium guajava L.) is fruit with very short shelf life due to associated with the loss of firmness in the pulp and information on the activity of enzymes that degrade pectic substances, as well as the amount of pectin, is very contradictory and not clearly defined. This work showed that the firmness of the fruit decreased sharply on the first four day­s of ripening. The identified phenolic compounds had their content increased with fruit maturation. The addition of the enzymes cellulase and hemicellulase in the pectin extraction in fruits of Psidium guajava revealed higher contents of this polyssacaride that the ones reported in literature, highlighting pectin as the responsible for the firmness of these fruits at more adequate contents. β-D-glucosidase was identified as one of the responsible enzymes for the maturation of P. guajava fruits. Thus, studies about possible inhibitory effects of this enzyme in P. guajava fruits may reveal an important tool to reduce pectin release and early maturation of these fruits.

Anticancer Properties of Essential Oils: An Overview.

BACKGROUND: Essential oils are complex mixtures of low molecular weight compounds extracted from plants. Their main constituents are terpenes and phenylpropanoids, which are responsible for their biological and pharmaceutical properties, such as insecticidal, parasiticidal, antimicrobial, antioxidant, anti-inflammatory, analgesic, antinociceptive, anticarcinogenic, and antitumor properties. Cancer is a complex genetic disease considered as a serious public health problem worldwide, accounting for more than 8 million deaths annually. OBJECTIVE: The activities of prevention and treatment of different types of cancer and the medicinal potential of essential oils are addressed in this review. CONCLUSION: Several studies have demonstrated anti-carcinogenic and antitumor activity for many essential oils obtained from various plant species. They may be used as a substitution to or in addition to conventional anti-cancer therapy. Although many studies report possible mechanisms of action for essential oils compounds, more studies are necessary in order to apply them safely and appropriately in cancer therapy.