In-depth immunorecognition and neutralization analyses of Micrurus mipartitus and M. dumerilii venoms and toxins by a commercial antivenom.

In Colombia, the Micrurus genus comprises 30 species, including M. mipartitus and M. dumerilii, which are of major clinical relevance due to their wide geographical distribution and the number of snakebites inflicted by them. These neurotoxic envenomations are characterized by neuromuscular paralysis attributed to venom components such as three-finger toxins (3FTx) and phospholipases (PLA2). Additionally, there is limited information available on the neutralizing coverage of commercially available antivenoms, underscoring the need to perform studies to assess the cross-neutralizing ability of these life-saving products. Therefore, we present an in-depth immunorecognition analysis by the anticoral-INS antivenom from Colombia on the M. mipartitus and M. dumerilii venoms. The antivenom cross-recognized the whole venoms and their components with different intensities. For instance, the antivenom showed better recognition on PLA2s than on 3FTxs in both venoms. Moreover, at doses tested, the antivenom totally neutralized the lethal effect of M. dumerilii venom; however, it did not neutralize this effect induced by M. mipartitus venom and its main toxic components from the southwestern region of the department of Antioquia. Furthermore, the anticoral-INS antivenom displayed better cross-immunorecognition of PLA2-predominant Micrurus venoms than of 3FTx-predominant Micrurus venoms. This highlights the need to include venoms from both types of venom patterns in the immunization mixture to produce antivenoms against coral snakes. Finally, our results suggest the need for further research to optimize the composition of immunizing mixtures for antivenom production and improve their efficacy against coral snake envenomation in Colombia and the Americas.

Knowledge about Snake Venoms and Toxins from Colombia: A Systematic Review.

Colombia encompasses three mountain ranges that divide the country into five natural regions: Andes, Pacific, Caribbean, Amazon, and Orinoquia. These regions offer an impressive range of climates, altitudes, and landscapes, which lead to a high snake biodiversity. Of the almost 300 snake species reported in Colombia, nearly 50 are categorized as venomous. This high diversity of species contrasts with the small number of studies to characterize their venom compositions and natural history in the different ecoregions. This work reviews the available information about the venom composition, isolated toxins, and potential applications of snake species found in Colombia. Data compilation was conducted according to the PRISMA guidelines, and the systematic literature search was carried out in Pubmed/MEDLINE. Venom proteomes from nine Viperidae and three Elapidae species have been described using quantitative analytical strategies. In addition, venoms of three Colubridae species have been studied. Bioactivities reported for some of the venoms or isolated components-such as antibacterial, cytotoxicity on tumoral cell lines, and antiplasmodial properties-may be of interest to develop potential applications. Overall, this review indicates that, despite recent progress in the characterization of venoms from several Colombian snakes, it is necessary to perform further studies on the many species whose venoms remain essentially unexplored, especially those of the poorly known genus Micrurus.

Drugs as Possible Triggers of Takotsubo Cardiomyopathy- Update 2022: Systematic Review.

BACKGROUND: A list of drugs that can induce takotsubo cardiomyopathy (TCM) was published in 2011 and 2016. The aim of the present review was to update this list. METHODS: Similar to the 2011 and 2016 reviews, from April 2015 to May 2022 case reports of druginduced TCM were identified by a comprehensive search in Medline/PubMed database. The search terms were: takotsubo cardiomyopathy, tako-tsubo cardiomyopathy, stress cardiomyopathy, transientleft- ventricular ballooning syndrome, apical ballooning syndrome, ampulla cardiomyopathy OR broken heart syndrome; together with "iatrogenic", "induced by" OR "drug-induced". Registers published in English or Spanish, in humans, and with full texts were retrieved. Articles that recognized any drug associated with the development of TCM were selected. RESULTS: Overall, 184 manuscripts were identified by the search. A total of 39 articles were included after an exhaustive revision. Eighteen drugs as possible triggers of TCM were identified in the current update. Of them, 3 (16.7%) have been previously identified, and 15 (83.3%) are different from the previous reports. Thus, the list of drugs as possible triggers of TCM updated in 2022 includes 72 drugs. CONCLUSION: There are new case reports that link drugs with the development of TCM. The current list is principally made up of drugs that generate sympathetic overstimulation. However, some of the listed drugs do not have a clear link with sympathetic activation.

Antibodies against a single fraction of Micrurus dumerilii venom neutralize the lethal effect of whole venom.

The coralsnake Micrurus dumerilii (Elapidae) is reported to cause envenomings of medical importance. Previous studies characterized the protein composition of its venom, with phospholipase A2 (PLA2) proteins the most abundant. However, it is unknown which venom components are responsible for its lethal toxicity. Fractionation of M. dumerilii venom from Colombia was carried out using RP-HPLC and each fraction was screened for lethal effect in mice at a dose of 20 µg by intraperitoneal route. Results showed that only one fraction, F9, was lethal. This fraction displayed PLA2 activity, induced indirect hemolysis in vitro, as well as edema and myotoxicity in vivo. SDS-PAGE of unreduced F9 evidenced two bands of 8 and 15 kDa, respectively, consistent with the detection of proteins with masses of 13,217.77 Da, 7144.06 Da, and 7665.55 Da. Tryptic digestion of F9 followed by nESI-MS/MS revealed peptide sequences matching proteins of the three-finger toxin (3FTx) and PLA2 families. Immunization of a rabbit with F9 proteins elicited antibody titers up to 1:10,000 by ELISA. After serum fractionation with caprylic acid, the obtained IgG was able to neutralize the lethal effect of the complete venom of M. dumerilii using a challenge of 2 ×LD50 at the IgG/venom ratio of 50:1 (w/w). In conclusion, present results show that the lethal effect of M. dumerilii venom in mice is mainly driven by one fraction which contains 3FTx and PLA2 proteins. The antibodies produced against this fraction cross-recognized other PLA2s and neutralized the lethal effect of whole M. dumerilii venom, pointing out to the potential usefulness of F9 as a relevant antigen for improving current coral snake antivenoms.

Inhibitory Effects of Varespladib, CP471474, and Their Potential Synergistic Activity on Bothrops asper and Crotalus durissus cumanensis Venoms.

Snakebite is a neglected tropical disease that causes extensive mortality and morbidity in rural communities. Antivenim sera are the currently approved therapy for snake bites; however, they have some therapeutic limitations that have been extensively documented. Recently, small molecule toxin inhibitors have received significant attention as potential alternatives or co-adjuvant to immunoglobulin-based snakebite therapies. Thus, in this study, we evaluated the inhibitory effects of the phospholipase A2 inhibitor varespladib and the metalloproteinase inhibitor CP471474 and their synergistic effects on the lethal, edema-forming, hemorrhagic, and myotoxic activities of Bothrops asper and Crotalus durissus cumanensis venoms from Colombia. Except for the preincubation assay of the lethal activity with B. asper venom, the mixture showed the best inhibitory activity. Nevertheless, the mix did not display statistically significant differences to varespladib and CP471474 used separately in all assays. In preincubation assays, varespladib showed the best inhibitory activity against the lethal effect induced by B. asper venom. However, in independent injection assays, the mix of the compounds partially inhibited the lethal activity of both venoms (50%). In addition, in the assays to test the inhibition of edema-forming activity, the mixture exhibited the best inhibitory activity, followed by Varespladib, but without statistically significant differences (p > 0.05). The combination also decreased the myotoxic activity of evaluated venoms. In these assays, the mix showed statistical differences regarding CP471474 (p < 0.05). The mixture also abolished the hemorrhagic activity of B. asper venom in preincubation assays, with no statistical differences to CP471474. Finally, the mixture showed inhibition in studies with independent administration in a time-dependent manner. To propose a mode of action of varespladib and CP471474, molecular docking was performed. PLA2s and SVMPs from tested venoms were used as targets. In all cases, our molecular modeling results suggested that inhibitors may occupy the substrate-binding cleft of the enzymes, which was supported by specific interaction with amino acids from the active site, such as His48 for PLA2s and Glu143 for the metalloproteinase. In addition, varespladib and CP471474 also showed interaction with residues from the hydrophobic channel in PLA2s and substrate binding subsites in the SVMP. Our results suggest a synergistic action of the mixed inhibitors and show the potential of varespladib, CP471474, and their mixture to generate new treatments for snakebite envenoming with application in the field or as antivenom co-adjuvants.

Tako-tsubo cardiomyopathy in clinical toxinology: A systematic review.

Tako-tsubo cardiomyopathy (TTC) is a transient left ventricular dysfunction, normally triggered by emotional or physical stress, although it is also associated with to use of drugs, drug abuse, or some intoxications. In addition, TTC has been reported in some case reports derived from the exposure of patients to animal venoms, toxins or poisons, or bacterial infections. However, to date, a systematic assessment of TTC in clinical toxinology is lacking. Therefore the aim of this study was to collect and integrate the available information about TTC in clinical toxinology. After our search strategy, 19 articles were retrieved, resulting in 20 case reports. Most cases occurred in women (75.0%). The venomous species that trigger TTC are bee/wasp, including probable Africanized honey bee and Vespa orientalis (15.0%), scorpions (Tytius serrulatus and Androctonus australis, 15.0%), a spider (Latrodectus tredecimguttatus, 5.0%), snakes (Gloydius blomhofii and Naja nivea, 10.0%), Clostridium sp (C. tetani, C. botulinum and C. difficile, 45.0%) and jellyfish (Pelagia noctiluca and Carukia barnesi, 10.0%). Among the affected people there were two deaths. In all case reports authors diagnosed TTC by using the combination of some of the following strategies: clinical findings, echocardiography, magnetic cardiac resonance, electrocardiogram changes and/or the increased plasma levels of cardiac damage biomarkers. In most cases images were available. We hypothesized the possible mode of action of venoms, toxins or poisons to induce TTC, however other mechanisms may exist, but they have not been described yet. Therefore, further studies are needed. In some cases, venoms, toxins, or poisons might cause catecholamine discharge either directly or indirectly, therefore, this was suggested as the trigger of TTC. Finally, the appearance of TTC should be considered in clinical toxinology.

Immunorecognition and Neutralization of Crotalus durissus cumanensis Venom by a Commercial Antivenom Produced in Colombia.

In Colombia, on average 2.9% of the nearly 5600 snakebite events that occur annually involve the rattlesnake Crotalus durissus cumanensis. The envenomation by this snake is mainly characterized by neurotoxicity and the main toxin is crotoxin (~64.7% of the total venom). The Instituto Nacional de Salud (INS) produces a polyvalent antivenom aimed at the treatment of bothropic, crotalid, and lachesic envenomations; nonetheless, its immune reactivity profile and neutralizing capacity over biological activities of the C. d. cumanensis venom has been poorly evaluated. In this sense, the study aims: (1) to describe an in-depth exploration of its immunoreactivity through second-generation antivenomics and HPLC fraction-specific ELISA immunoprofiles; and (2) to evaluate the neutralization pattern of the rattlesnake venom in vitro and in vivo biological activities. The results obtained showed a variable recognition of crotoxin subunits, in addition to a molecular mass-dependent immunoreactivity pattern in which the disintegrins were not recognized, and snake venom metalloproteinases and L-amino acid oxidases were the most recognized. Additionally, a high neutralization of proteolytic and coagulant activities was observed, but not over the PLA2 activity. Further, the median effective dose against C. d. cumanensis venom lethality was 962 µL of antivenom per mg of venom. In conclusion, (1) the antivenom recognition over the crotoxin and the disintegrins of the C. d. cumanensis should be improved, thus aiming upcoming efforts for the exploration of new techniques and approaches in antivenom production in Colombia, and (2) the neutralization activity of the antivenom seems to follow the molecular mass-dependent recognition pattern, although other explanations should be explored.

Snake venomics, experimental toxic activities and clinical characteristics of human envenomation by Bothrocophias myersi (Serpentes: Viperidae) from Colombia.

Venoms of the viperid genus Bothrocophias, restricted to Colombia and Ecuador, are poorly known. Only a proteomic analysis of B. campbelli venom has been described. In this work we present a proteomic study of B. myersi venom, its biological activities, and describe the clinical characteristics of a patient bitten by this species. B. myersi venom mainly consists of phospholipases A2 (54.0%) and metalloproteinases (21.5%), among proteins of twelve different families. This venom exhibited proteolytic, phospholipase A2, myotoxic, edema-forming, and lethal activities. Enzymatic activities did not show statistically significant differences in comparison to Bothrops asper venom, but B. myersi venom displayed weaker hemorrhagic and coagulant activities. Polyvalent Viperidae antivenoms produced in Costa Rica and Colombia cross-recognized B. myersi venom by ELISA, however only the latter neutralized its lethal activity in mice when tested at a ratio of 3 mg venom/mL antivenom, suggesting it should be useful to treat envenomings inflicted by this species. A patient bitten by B. myersi developed edema and myotoxicity, evidenced by an increased creatine kinase activity in plasma. A good correlation was found between experimental biological activities of Bothrocophias myersi venom and the clinical features of an envenoming provoked by this species. SIGNIFICANCE: The proteomic characterization, toxicity, immunorecognition and neutralization of Bothrocophias myersi venom have been determined for the first time. The distribution of this pit viper is restricted to Colombia and Ecuador, and its venom contains a high proportion of phospholipases A2 and metalloproteinases. The polyvalent antivenom produced in Colombia neutralized the lethal activity of this venom in vivo, and therefore should be effective in the treatment of envenomings by this snake.

Sulfur Compounds as Inhibitors of Enzymatic Activity of a Snake Venom Phospholipase A2: Benzyl 4-nitrobenzenecarbodithioate as a Case of Study.

Snakebite is a neglected disease with a high impact in tropical and subtropical countries. Therapy based on antivenom has limited efficacy in local tissue damage caused by venoms. Phospholipases A2 (PLA2) are enzymes that abundantly occur in snake venoms and induce several systemic and local effects. Furthermore, sulfur compounds such as thioesters have an inhibitory capacity against a snake venom PLA2. Hence, the objective of this work was to obtain a carbodithioate from a thioester with known activity against PLA2 and test its ability to inhibit the same enzyme. Benzyl 4-nitrobenzenecarbodithioate (I) was synthesized, purified, and characterized using as precursor 4-nitrothiobenzoic acid S-benzyl ester (II). Compound I showed inhibition of the enzymatic activity a PLA2 isolated from the venom of the Colombian rattlesnake Crotalus durissus cumanensis with an IC50 of 55.58 µM. This result is comparable with the reported inhibition obtained for II. Computational calculations were performed to support the study, and molecular docking results suggested that compounds I and II interact with the active site residues of the enzyme, impeding the normal catalysis cycle and attachment of the substrate to the active site of the PLA2.

Toxinology in Colombia: contributions of ophidism/scorpionism program and other research groups / Toxinología en Colombia: contribuciones del programa de ofidismo/escorpionismo y de otros grupos de investigación

Background: Toxinology is a sub-field of toxicology dedicated to studying toxins produced by animals, plants and, microorganisms. In Colombia, during the last thirty years, this area has been mainly investigated by Ophidism/Scorpionism Program of Universidad de Antioquia. However, some other research groups have also contributed to our knowledge of venoms and toxins, as well as their related effects and treatments. Objective: to highlight the most significant findings in toxinology made by the Ophidism/Scorpionism Program and other research groups in Colombia. Methods: 119 papers dealing with the history of ophidiology and toxinology in Colombia were collected and analyzed. Results: some useful terms are described to understand toxinology and its scope. Also, a brief history of ophidiology is presented, spanning from the discovery of America until present-day findings. Finally, an overall description of several results related to toxin isolation, characterization, antivenoms, clinical trials, description of new species, proteomic and transcriptomic, among others. The nineteens were characterized by the study of snakebites, their clinic manifestations, and the use of antivenoms. In addition, the ethnopharmacological studies of medicinal plants used in snakebite treatments began to be explored. The 2000s included the newly ethnopharmacology, toxin isolation, clinical trials, inhibitor studies, scorpion venom characterization, and scorpion stings features. Finally, from 2010 until today, proteomic and transcriptomic gave the most important findings. Conclusions: Toxinology works in Colombia have contributed to our knowledge about endemic species, clinical manifestations of snakebite and scorpion stings, and the development of new therapeutic agents. However, we invite Colciencias and other funding agencies to assign more resources to support a higher number of researchers in this field, since snakebite is considered a neglected tropical disease by the World Health Organization, which needs more attention from governments and scholars. Finally, the venoms of some species and their possible mode of action are still unknown to us. Besides, given the complexity of venoms, we are not yet aware of the potential use of toxins in current biomedicine. Thus, studies in toxinology must continue.

Antecedentes: La Toxinología es el campo de la Toxicología que estudia las toxinas producidas por animales, plantas y microorganismos. En Colombia, durante los últimos treinta años, los estudios realizados en esta área han sido desarrollados principalmente por el Programa de Ofidismo/Escorpionismo de la Universidad de Antioquia. Sin embargo, otros grupos de investigación también han contribuido en el conocimiento de venenos, toxinas, efectos y tratamientos. Objetivo: Destacar los hallazgos más relevantes en toxinología realizados por el Programa de Ofidismo Escorpionismo y otros grupos de investigación en Colombia. Métodos: Se recopilaron 119 artículos referentes a la historia de la ofidiología y la toxinología en Colombia. Resultados: Se describieron algunos términos útiles para el entendimiento de la toxinología y sus alcances. Se construyó una breve historia de la ofidiología que inicia con el descubrimiento de América y finaliza con hallazgos recientes. Se realizó una amplia descripción de varios resultados relacionados con el aislamiento y caracterización de toxinas, antivenenos, ensayos clínicos, descripciones de nuevas especies, proteómica y transcriptómica, entre otras. Así, la década de los noventa se caracterizó por el estudio de las mordeduras de serpientes, sus manifestaciones clínicas, el uso de antivenenos y la exploración de la etnofarmacología asociada a las mordeduras de serpiente. La década del 2000 incluyó nuevamente etnofarmacología, el aislamiento de toxinas, ensayos clínicos, estudios sobre inhibidores de toxinas, caracterización de venenos y picaduras de escorpión. Finalmente, desde 2010 hasta hoy, la proteómica y transcriptómica aportaron los hallazgos más importantes. Conclusiones: Los estudios de Toxinología en Colombia han contribuido al conocimiento de especies endémicas, manifestaciones clínicas de mordeduras de serpientes y picaduras escorpiones, y el desarrollo de nuevos agentes terapéuticos. No obstante, se invita a Colciencias y a otras agencias de financiamiento a apoyar la investigación en este campo, ya que es considerada una enfermedad tropical desatendida por la Organización Mundial de la Salud y necesita mayor atención por parte del gobierno e instituciones académicas. Además, dada la complejidad de los venenos, se desconoce el uso potencial de las toxinas en la biomedicina actual. Así, se deben continuar realizando estudios en toxinología.

Potential of Matrix Metalloproteinase Inhibitors for the Treatment of Local Tissue Damage Induced by a Type P-I Snake Venom Metalloproteinase.

Snake bite envenoming is a public health problem that was recently included in the list of neglected tropical diseases of the World Health Organization. In the search of new therapies for the treatment of local tissue damage induced by snake venom metalloproteinases (SVMPs), we tested the inhibitory activity of peptidomimetic compounds designed as inhibitors of matrix metalloproteinases on the activities of the SVMP Batx-I, from Bothrops atrox venom. The evaluated compounds show great potential for the inhibition of Batx-I proteolytic, hemorrhagic and edema-forming activities, especially the compound CP471474, a peptidomimetic including a hydroxamate zinc binding group. Molecular dynamics simulations suggest that binding of this compound to the enzyme is mediated by the electrostatic interaction between the hydroxamate group and the zinc cofactor, as well as contacts, mainly hydrophobic, between the side chain of the compound and amino acids located in the substrate binding subsites S1 and S1 ' . These results show that CP471474 constitutes a promising compound for the development of co-adjuvants to neutralize local tissue damage induced by snake venom metalloproteinases.

Inhibition of a Snake Venom Metalloproteinase by the Flavonoid Myricetin.

Most of the snakebite envenomations in Central and South America are caused by species belonging to Bothrops genus. Their venom is composed mainly by zinc-dependent metalloproteinases, responsible of the hemorrhage characteristic of these envenomations. The aim of this study was to determine the inhibitory ability of ten flavonoids on the in-vitro proteolytic activity of Bothrops atrox venom and on the hemorrhagic, edema-forming and myonecrotic activities of Batx-I, the most abundant metalloproteinase isolated from this venom. Myricetin was the most active compound, exhibiting an IC 50 value of 150 µ M and 1021 µ M for the inhibition of proteolytic and hemorrhagic activity, respectively. Independent injection experiments, with a concentration of 1600 µ M of myricetin administered locally, immediately after toxin injection, demonstrated a reduction of 28 ± 6 % in the hemorrhagic lesion. Additionally, myricetin at concentrations 800, 1200 and 1600 µ M promoted a reduction in plasma creatine kinase activity induced by Batx-I of 21 ± 2 % , 60 ± 5 % and 63 ± 2 % , respectively. Molecular dynamics simulations coupled with the adaptive biasing method suggest that myricetin can bind to the metalloproteinase active site via formation of hydrogen bonds between the hydroxyl groups 3', 4' and 5' of the benzyl moiety and amino acid Glu143 of the metalloproteinase. The hydroxyl substitution pattern of myricetin appears to be essential for its inhibitory activity. Based on this evidence, myricetin constitutes a candidate for the development of inhibitors to reduce local tissue damage in snakebite envenomations.

Interactions between Triterpenes and a P-I Type Snake Venom Metalloproteinase: Molecular Simulations and Experiments.

Small molecule inhibitors of snake venom metalloproteinases (SVMPs) could provide a means to rapidly halt the progression of local tissue damage following viperid snake envenomations. In this study, we examine the ability of candidate compounds based on a pentacyclic triterpene skeleton to inhibit SVMPs. We leverage molecular dynamics simulations to estimate the free energies of the candidate compounds for binding to BaP1, a P-I type SVMP, and compare these results with experimental assays of proteolytic activity inhibition in a homologous enzyme (Batx-I). Both simulation and experiment suggest that betulinic acid is the most active candidate, with the simulations predicting a standard binding free energy of Δ G ∘ = - 11.0 ± 1.4 kcal/mol. The simulations also reveal the atomic interactions that underlie binding between the triterpenic acids and BaP1, most notably the electrostatic interaction between carboxylate groups of the compounds and the zinc cofactor of BaP1. Together, our simulations and experiments suggest that occlusion of the S1 ' subsite is essential for inhibition of proteolytic activity. While all active compounds make hydrophobic contacts in the S1 ' site, ß -boswellic acid, with its distinct carboxylate position, does not occlude the S1 ' site in simulation and exhibits negligible activity in experiment.

How the Triton X-100 modulates the activity/stability of the Thermomyces lanuginose lipase: Insights from experimental and molecular docking approaches.

The lipase and Triton X-100 mixture is common for stabilization, immobilization and application processes of these kinds of enzymes. The objective of this article was to study the structural behavior and catalytic performance of Thermomyces lanuginose lipase in the presence of Triton X-100 at 25 °C and different pHs. The structural changes were followed by circular dichroism, correlating them with the catalytic performance, which is reported as the initial lipase activity in the hydrolysis of p­nitro phenyl butyrate at zero time and residual activity after 48 h of incubation in the absence or presence of surfactant, at the selected pHs. Computational simulations allowed to explain the correlations between the physicochemical changes and the formation of surfactant protein complex, leading to the elucidation of the main interactions that drive activity and stability of this lipase in presence of the Triton X-100 surfactant. Main results showed the Triton X-100-enzyme complex modulates the site active geometry, favoring a better substrate-enzyme adjustment, which influences the activity and stability at evaluated pHs. This study contributes to understand the effect of some additives commonly used to improve the biocatalytic performance on several applications for different industrial fields.

Betulinic, oleanolic and ursolic acids inhibit the enzymatic and biological effects induced by a P-I snake venom metalloproteinase.

Betulinic acid (BA), Oleanolic acid (OA) and Ursolic acid (UA), are pentacyclic triterpenoids with widespread occurrence throughout the plant kingdom, these compounds are widely recognized by their pharmacological and biological properties, such as, anti-tumoral, anti-inflammatory, anti-microbial and hepatoprotective activity. In this work we determined the inhibitory ability of these compounds on the enzymatic, hemorrhagic, myotoxic and edema-inducing activities of Batx-I, a P-I metalloproteinase isolated from Bothrops atrox venom. BA, UA and OA inhibited the proteolytic activity of Batx-I on gelatin with IC50 values of 115.3, 223.0 and 357.3 µM, respectively. Additionally, these compounds showed inhibition of the hemorrhagic activity of Batx-I in skin with IC50 345.7, 643.5 and 1077.0 µM for BA, UA and OA in preincubation experiments. In studies with independent-injection, in which Batx-I was injected and then, at the same site, a concentration of 600 µM of each compound were administered at either 0, 5 or 10 min, BA showed a significant reduction of hemorrhage at 0 and 5 min. In addition, these compounds inhibited myotoxicity and edema-forming activity of Batx-I at 600 µM concentration. Molecular docking studies suggested that these compounds could occupy part of the substrate binding cleft of the enzyme affecting its catalytic cycle. In this manner, triterpenic acids are candidates for the development of inhibitors for the prevention of local tissue damage in snakebite envenomation.

Acute oral toxicity from a fraction rich in phenolic compounds from the leaf extract of Swietenia macrophylla king in a murine model / Determinación de la toxicidad aguda oral de una fracción rica en compuestos fenólicos del extracto de hojas Swietenia macrophylla king en un modelo murino

Background: The mahogany tree (Swietenia macrophylla King) is widely used in traditional medicine, especially its seeds, which are used in Malaysia, Indonesia and some countries of South America. Recently an active fraction of the leaf ethanolic extract (labelled "Sm13-16,23"), showed promising results against some toxins of South American pit vipers. Objectives: In this study the aim was to evaluate the acute oral toxicity of fraction Sm13-16,23 from the leaf ethanolic extract, using Swiss Webster mice. Methods: Fraction Sm13-16,23 was administered orally in a single dose to 2000 mg/kg and 300 mg/kg, clinical follow-up for 14 days was performed and then euthanasia, necropsy and histopathology of organs were performed. Results: Overall, there were not deaths recorded during the study period. Further not signs of toxicity in doses of 300mg/kg were observed, but at doses of 2000 mg/kg, histopathological lesions in the liver, as karyomegaly and binucleation were observed. Conclusions: Fraction Sm13-16, 23 does not produce toxicity lesion at doses of 300mg/kg, indicating that the acute oral toxicity risk is low.

Antecedentes: El árbol de la caoba (Swietenia macrophylla King) es utilizado ampliamente en la medicina tradicional, especialmente sus semillas, que son usadas en Malasia, Indonesia y algunos países de Suramérica. Recientemente una fracción activa del extracto etanólico de las hojas (llamada "Sm13-16,23"), demostró resultados promisorios frente algunas toxinas de las víboras suramericanas. Objetivos: En este estudio se evaluó la toxicidad aguda oral de la fracción Sm13-16,23 del extracto etanólico de las hojas de S. macrophylla utilizando ratones Swiss Webster. Métodos: La fracción Sm13-16,23 se administró vía oral como dosis única a 2000 mg/kg y 300 mg/kg, se realizó seguimiento clínico de los animales durante 14 días y finalmente se les aplicó la eutanasia. Se practicó la necropsia de cada animal y se tomaron algunos órganos para su evaluación histopatológica. Resultados: No se registraron muertes durante el tiempo de estudio, ni se evidenciaron signos de toxicidad a dosis de 300mg/kg, sin embargo, a la dosis de 2000 mg/kg, se encontraron lesiones histopatológicas en el hígado; como cariomegalia y binucleación de los hepatocitos. Conclusión: La fracción Sm13-16,23 no produjo lesiones de toxicidad a dosis de 300mg/ kg, lo que indicia que el riesgo de toxicidad aguda oral es bajo.

Inhibitory effect of pinostrobin from Renealmia alpinia, on the enzymatic and biological activities of a PLA2.

Pinostrobin is a flavanone isolated from Renealmia alpinia, a plant used in folk medicine to treat snakebites. We tested the inhibitory ability of pinostrobin on the enzymatic, anticoagulant, myotoxic and edema-inducing activities of a PLA2 isolated from Crotalus durissus cumanensis venom. The compound displayed IC50 values of 1.76mM and 1.85mM (95% Confidence intervals: 1.34-2.18 and 1.21-2.45) on the PLA2 enzymatic activity, when either aggregated or monodispersed substrates were used, respectively. When mice were injected with PLA2 preincubated with 0.4, 2.0 and 4.0mM of pinostrobin, myotoxic activity induced by the PLA2 was inhibited up to 87%. Nevertheless, these values decreased up to 56% when the pinostrobin was injected into muscle after PLA2. Pinostrobin inhibited edema-forming and anticoagulant activities of the PLA2. In order to have insights on the mode of action of pinostrobin, intrinsic fluorescence and ultraviolet studies were performed. Results suggest that pinostrobin interacts directly with the PLA2. These findings were supported by molecular docking results, which suggested that pinostrobin forms hydrogen bonds with residues His48 and Asp49 of PLA2, besides, a π-π stacking interactions with those of residues Phe5 and Trp31, and rings C of flavanone and Tyr52 of the toxin.

The need for an integrated approach in confronting snakebite envenoming in Latin America: the relevance of endogenous scientific and technological research / Hacia un abordaje integral del envenenamiento ofídico en américa latina: la importancia de la generación de conocimiento científico-tecnológico endógeno

Snakebite envenomings constitute a serious public health problem on a global level, especially in Africa, Asia and Latin America [1, 2]. In our Latin American region, it is estimated that at least 70,000 snakebite cases occur every year, although the actual number is likely to be higher [3]. This pathology is one of the so-called 'neglected tropical diseases', i.e. a group of diseases that affect primarily poor people in poor settings and, therefore, does not receive the necessary attention from research agencies, pharmaceutical companies, and health authorities. Consequently, it has been described as a 'disease of poverty' [3, 4]. In our continent it largely affects impoverished populations living in rural areas, including indigenous groups [3]

Characterization of the most promising fraction of Swietenia macrophylla active against myotoxic phospholipases A2: identification of catechin as one of the active compounds / Caracterización de la fracción más promisoria de Swietenia macrophylla activa contra fosfolipasas A2 miotoxicas: identificación de catequina como uno de los compuestos activos

Background: The pharmacological effects produced by snakebite accidents involve the actions of several enzymes, of which those of the phospholipases A2 (PLA2) exhibit a wide variety of effects such as edema and myotoxicity. Some plant extracts have been antagonists of crude snake venoms and toxins. Based on promising bioactivity, Swietenia macrophylla King was selected for further studies. Objective: The purpose of this study was to identify the PLA2 inhibitors present in the crude extract of S. macrophylla that could be promising leads in neutralizing the local effects of ophidian accidents. Methods: Bioassay-guided fractionation of the ethanolic extract of the leaves of S. macrophylla lead to the detection of (+)-catechin, characterized through gas chromatography coupled with mass spectrometry (GC-MS), and confirmed by HPLC. The PLA2 inhibitory activity was measured with the Dole method and a spectrophotometric assay with 4-Nitro-3 octanoyloxy-benzoic acid (4N3OBA). Cytotoxicity was done on C2C12 murine myoblast. Results: Fraction F5 and (+)-Catechin inhibited the PLA2 activity of B. asper venom, in a dosedependent way. In addition, (+) Catechin showed an inhibition level of 83.1 ± 3.1 % of the enzymati activity of one PLA2 purified from the venom of Crotalus durissus cumanensis using 4N3OBA as substrate. Also the ethanolic extract and fraction F5 showed inhibition of the cytotoxicity induced by the Bothrops atrox venom and their Lys 49 PLA2 (80 and 100% respectively). Molecular docking results suggested that OH from 4´ and 5' carbons of (+)-catechin could form hydrogen bonds with carboxylate moiety of residue Asp49, while OH from 5 could form a hydrogen bond with Asn 6. Additional Van der Waals interactions were also proposed. Conclusion: Swietenia macrophylla exhibited strong inhibitory activity against PLA2s enzymes. Catechin, one of the components in the active fraction F5, is proposed as being partially responsible for the bioactivity.

Antecedentes: Los efectos farmacológicos producidos en el accidente ofídico implican la acción de varias enzimas, como las fosfolipasas A2 (PLA2), que exhiben una amplia variedad de efectos como edema y miotoxicidad. Algunos extractos de plantas han demostrado ser antagonistas de los venenos crudos y sus toxinas. En base a una bioactividad promisoria previa, Swietenia macrophylla King fue seleccionada para estudios posteriores. Objetivo: El propósito de este estudio fue identificar metabolitos inhibidores de PLA2 presentes en el extracto crudo de S. macrophylla que podrían ser prometedores en la neutralización de los efectos locales del accidente ofídico. Métodos: Un fraccionamiento biodirigido del extracto etanólico de hojas de S. macrophylla llevó a la detección de la (+)-catequina mediante cromatografía de gases acoplada a espectrometría de masas (GC-MS), y se confirmó mediante HPLC. La actividad inhibidora de PLA2 se determinó por el método de Dole y un ensayo espectrofotométrico con Ácido 4-Nitro-3-octanoiloxibenzoico (4N3OBA). La citotoxicidad se determinó en mioblastos C2C12 murinos. Resultados: La fracción F5 de S. macrophylla, mostró la mayor inhibición del veneno de Bothrops asper y PLA2s aisladas de B. atrox y Crotalus durissus cumanensis e inhibición completa de la citotoxicidad. La (+)-catequina fue el metabolito más abundante en F5, mostró una inhibición de PLA2 de 89,8%, 81,1% y 74,3%, a diferentes relaciones de veneno: catequina. Mediante estudios de docking molecular se demostró la unión de la (+)-catequina al sitio activo de la PLA2. Conclusión: Swietenia macrophylla exhibió una fuerte actividad inhibitoria sobre las enzimas PLA2s. La (+)-catequina, uno de los metabolitos presentes en la fracción activa F5, se propone como uno de los compuestos responsables de la bioactividad.

Colombian biodiversity, an opportunity for the strengthening of the pharmaceutical and cosmetic industries / La biodiversidad colombiana, una oportunidad para el fortalecimiento de las industrias farmacéutica y cosmética

Colombia, a country privileged by nature, where diversity of climates are created by large altitude gradients which generate a variety of ecosystems and beautiful badlands, punas, mountain forests, dry valleys, high deserts and salt flats; landscapes that have diversity of plants and animals and we are cataloged as one of the countries as the greatest diversity in the world, preceded by Brazil and followed by Indonesia, China and Mexico [1]. We have 28.000 plants and lichens; it is an invaluable heritage that can give to our country competitive advantages in the discovery and development of active ingredients for the pharmaceutical and cosmetic industry by sustainable use of our biodiversity.