A new era of pain management could be on the horizon. How Terry Snutch turned snail venom into a multi-million dollar painkiller.

Thesis (M.Tech.: Homoeopathy)-Dept of Homoeopathy, Durban Institute of Technology, 2004 xiv, 154 leaves : ill. ; 30 cm / This study was conducted by administering Naja mossambica 30CH (a homoeopathic remedy derived from the venom of the Mozambican spitting cobra) to healthy individuals in order to elicit and document the resulting mental and physical symptomology. These symptoms were compared to the toxicology of Naja mossambica venom as well as a doctrine of signatures analysis of the snake in order to expand and clarify the remedy picture. Existing knowledge of the venom toxicology gives a clear indication of the organs and body systems that the substance has an affinity for. Many poisonous substances used homoeopathically rely heavily on inferences made from the toxicology of the substance, as much of the gross pathology in the symptom picture cannot safely be elucidated in a proving. The aim of this study was to determine the sphere of action of Naja mossambica by utilising symptoms obtained from the proving and from the toxicology of the venom. This was done in order to determine the remedy's usefulness in a homoeopathic clinical setting by expanding our understanding of the substance and thereby facilitating the treatment of disease based on the law of similars.

Predator harassment is an anti-predator behaviour that may increase a harasser’s risk of predation but decrease the potential for predation for other members of a group. The Cape ground squirrel (Xerus inauris) is a facultative cooperative breeder from southern Africa that harasses venomous snakes. The objective of my study was to examine predator harassment to determine: 1) whether predator harassment was part of alloparental care by comparing harassment behaviour among age and sex classes; 2) how individuals adjusted their behaviour against different snake species; 3) whether olfaction was used in discrimination of snakes; and 4) if individuals possessed venom resistance against venomous snakes. I found that females with juvenile offspring harass snakes longer and more intensely than other individuals, suggesting that predator harassment was a maternal behaviour. Squirrels increased harassment, inspection and vigilant behaviours with risk when exposed to both live snakes and snake odours suggesting they can use olfaction to discriminate snake predators. I also found no venom resistance in Cape ground squirrels concluding the cost of envenomation was significant.

Predator harassment is an anti-predator behaviour that may increase a harasser’s risk of predation but decrease the potential for predation for other members of a group. The Cape ground squirrel (Xerus inauris) is a facultative cooperative breeder from southern Africa that harasses venomous snakes. The objective of my study was to examine predator harassment to determine: 1) whether predator harassment was part of alloparental care by comparing harassment behaviour among age and sex classes; 2) how individuals adjusted their behaviour against different snake species; 3) whether olfaction was used in discrimination of snakes; and 4) if individuals possessed venom resistance against venomous snakes. I found that females with juvenile offspring harass snakes longer and more intensely than other individuals, suggesting that predator harassment was a maternal behaviour. Squirrels increased harassment, inspection and vigilant behaviours with risk when exposed to both live snakes and snake odours suggesting they can use olfaction to discriminate snake predators. I also found no venom resistance in Cape ground squirrels concluding the cost of envenomation was significant.

Orientador: Daniel Carvalho Pimenta / Resumo: O veneno de Crotalus durissus terrificus (Cdt), baseado no critério do perfil da cromatografia de exclusão molecular, pode ser descrito como sendo composto de quatro principais toxinas: Giroxina, Crotamina, Crotoxina e Convulxina. A crotoxina é uma neutoroxina, é um heterodímero não covalente, composta por duas subunidades: fosfolipase A2 (PLA2) e crotapotina (Ctp). A crotapotina constitui o componente da crotoxina que apresenta atividade anti-inflamatória além de ser chaperona de PLA2. Esta molécula é constituída por três cadeias peptídicas ligadas por pontes dissulfeto. Além disso, já foram descritas isoformas para a crotapotina. O objetivo deste estudo foi desenvolver um método cromatográfico para isolar a crotapotina, separar suas cadeias e analisá-las por LC/MS e LC-MS/MS para análise proteômica e/ou seqüenciamento de novo e avaliação da possível existência de um padrão de variação nas substituições de aminoácidos. O método C8-RP-HPLC foi capaz de separar os componentes do veneno de Cdt que puderam ser identificados por espectrometria de massas, de acordo com a sua massa molecular. A crotapotina foi isolada, reduzida e alquilada e submetida à outra separação cromatográfica RP-HPLC (C18) para isolamento das subunidades alquiladas. Conforme descrito na literatura, a cadeia α da crotapotina é composta por 40 resíduos de aminoácidos, enquanto que a cadeia β possui 35 resíduos e a cadeia γ 14, pelo que as subunidades podem ser identificadas pelas suas massas moleculares. As a... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The Crotalus durissus terrificus venom (Cdt), based on molecular size chromatography, can be described as being composed of four main toxins: Giroxin, Crotamine, Crotoxin and Convulxin. Crotoxin is a neutoroxin, a non-covalent heterodimer composed of two subunits: phospholipase A2 (PLA2) and crotapotin (Ctp). Crotapotin is the component of crotaxin that has anti-inflammatory activity besides being a PLA2 chaperone. This molecule consists of three peptide chains linked by disulfide bonds. In addition, isoforms for crotapotin have been described. The objective of this study was to develop a chromatographic methodology to isolate a crotapotin, separate its chains and analyze by LC/MS and LC-MS/MS for proteomic analysis and/or de novo sequencing and evaluation of the possibility of a variation pattern amino acid substitutions. The C8-RPHPLC method was able to separate the Cdt venom components that could be identified by mass spectrometry, according to their molecular mass. The crotapotin was isolated, reduced and alkylated and subjected to another RP-HPLC (C18) chromatographic separation for isolation of the alkylated subunits. As described in the literature, the α chain of crotapotin is composed of 40 amino acid residues, while the β chain has 35 residues and the γ chain, 14, and thus the subunits can be identified by their molecular masses. The analyzes of isolated chains of crotapotin showed the presence of more than one molecule per fraction, indicating the presence of isofor... (Complete abstract click electronic access below) / Mestre

Submitted by Patrícia da Rocha Mendes (prmbio@gmail.com) on 2018-08-15T16:34:30Z No. of bitstreams: 1 TESE FINAL 2.pdf: 1912543 bytes, checksum: 5c638621228b797d59f7811694025a8c (MD5) / Approved for entry into archive by Sulamita Selma C Colnago null (sulamita@btu.unesp.br) on 2018-08-15T19:06:08Z (GMT) No. of bitstreams: 1 mendes_pr_dr_bot.pdf: 1912543 bytes, checksum: 5c638621228b797d59f7811694025a8c (MD5) / Made available in DSpace on 2018-08-15T19:06:08Z (GMT). No. of bitstreams: 1 mendes_pr_dr_bot.pdf: 1912543 bytes, checksum: 5c638621228b797d59f7811694025a8c (MD5) Previous issue date: 2016-02-15 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Os acidentes ofídicos constituem um relevante problema de saúde pública, a qual deve receber cuidados rápidos e eficientes. Em relação ao ofidismo, no Brasil, somente há poucos anos foi definida uma política abrangente que enfocasse as múltiplas questões relacionadas aos acidentes humanos provocados por animais peçonhentos. Os venenos de serpentes apresentam uma mistura de compostos inorgânicos (íons metálicos) e orgânicos, sendo ricos em moléculas biologicamente ativas. Dentre essas moléculas, peptídeos, proteínas e compostos de baixa massa molecular são frequentemente estudados por apresentarem grande potencial terapêutico. Essas moléculas, se bem conhecidas, podem ser utilizadas na investigação de mecanismo moleculares e celulares. Além disso, estabelecem interessantes modelos moleculares para o desenvolvimento de estratégias biotecnológicas aplicáveis na criação de ferramentas experimentais e/ou agentes terapêuticos para a pesquisa básica e aplicada. Diante disso, a elucidação dos peptídeos hipotensores presentes no veneno crotálico da subespécie Crotalus durissus terrificus é o objetivo do presente trabalho. Este estudo poderá promover a identificação de outras moléculas hipotensoras podendo conter princípios ativos mais eficientes para o tratamento de quadros clínicos de hipertensão, doença esta que acomete milhares de pessoas no mundo todo. No presente trabalho, a porção peptídica do veneno da serpente C.d.t., apresentou rendimento médio de 12,8% em relação ao veneno bruto, sob condições de precipitação das proteínas totais na presença de ácido trifluoracético à 0,1% (v/v) e, quando submetida à estratégia de purificação por cromatografia líquida de fase reversa, 13 frações foram separadas. Por meio de ensaios de contratilidade de tecido isolado in vitro de músculo liso (artéria mesentérica), evidenciou-se que a fração I e a fração V apresentaram efeitos hipotensor e hipertensor, respectivamente. A fração I foi identificada com a sequência QHILIYVGVHD e massa molecular 1274,67 Da. Atualmente, esta molécula hipotensora identificada no presente trabalho foi submetida à síntese peptídica para posteriormente, avaliar-se este peptídeo hipotensor na sua forma sintética, seu efeito hipotensor por meio do ensaio de contratilidade de tecido isolado in vitro de músculo liso em relação ao fármaco Captopril. / The snakebites are an important public health problem, which must receive quick and efficient care. Regarding the ophidism, in Brazil, only a few years ago a comprehensive policy was defined that emphasized multiple issues related to human accidents caused by venomous animals. The snake venoms present a mixture of inorganic and organic compounds, being rich in biologically active molecules. Among these molecules, peptides, proteins and low molecular weight compounds are often studied by their ability therapeutic. Those molecules, when well known, may be used in the investigation of cellular and molecular mechanism. In addition, it provides interesting molecular models for the development of biotechnological strategies applicable in the creation of experimental tools and / or therapeutic agents for basic and applied research. Therefore, the elucidation of hypotensive peptides present in the venom of Crotalus durissus terrificus subspecies is the aim of this study, once this study may promote the identification of other hypotensive molecules may contain more effective active ingredients for the treatment of clinical conditions hypertension, this disease affecting millions of people in the world. In this study, the peptide portion of the venom of the snake Cdt, performed with an average yield of 12.8% compared to the venom under total protein precipitation conditions in the presence of trifluoro acetic acid to 0.1% (v / v) and, when subjected to purification strategy for reversed-phase liquid chromatography, 13 fractions were separated. Using isolated tissue contractility in vitro smooth muscle (mesenteric artery), was evidenced that the fraction I and fraction V showed hypotensive and hypertensive effects, respectively. Fraction I was identified with the QHILIYVGVHD sequence and molecular mass 1274.67 Da. Currently, this hypotensive molecule identified in this study was submitted to the peptide synthesis to subsequently evaluate this hypotensive peptide in its synthetic form, its hypotensive effect through tissue contractility assay isolated in vitro smooth muscle compared to Captopril medicine.

Les médicaments à base de substances naturelles telles que les plantes et les venins sont de plus en plus développés dans l’industrie pharmaceutique. Le contrôle qualité des matières premières ou teintures mères d’origine végétales ou animales, obligatoire dans un environnement BPF, est bien plus complexe que celui des petites molécules pharmaceutiques de synthèse. En effet, les substances responsables de l’activité thérapeutique recherchée sont souvent mal identifiées et on observe une variabilité qualitative et quantitative naturelle parfois importante, ce qui peut modifier de façon significative leur efficacité. Afin de s’assurer de l’homogénéité de chaque souche reçue, il est demandé, entre autre, de développer des méthodes de contrôle analytique permettant d’obtenir un « profil ou empreinte » analytique de la souche considérée, assimilable à une « carte d’identité » qui servira de référence pour le contrôle des lots utilisés en production. L’empreinte analytique (spectre, chromatogramme, électrophérogramme …) devra permettre une séparation la plus complète possible de l’ensemble des composés présents dans la souche considérée tout en tenant compte de sa « variabilité naturelle » afin d’assurer un contrôle qualitatif et semi-quantitatif.L’étude présentée a consisté à développer une approche analytique originale, combinant électrophorèse capillaire et chimiométrie, afin d’établir des profils électrophorétiques spécifiques («fingerprint») permettant d’effectuer le contrôle qualité de souches brutes de venins de serpents utilisés en thérapeutique. Les venins de serpent sont des mélanges complexes de peptides et de protéines, de plus en plus utilisés dans la conception de nouveaux médicaments. Comme toutes les substances naturelles, la composition des venins de serpents est sujette à une variabilité intra et inter individus relativement importante. De fait, la qualité de la matière première pour la production de médicament devient un challenge. Dans cette étude, l’électrophorèse capillaire s’est révélée comme une technique efficace pour la séparation des divers constituants composant les souches brutes de venins de serpent et l’obtention de leurs empreintes analytiques. L’étude a porté sur 4 venins de serpent, Lachesis muta, Bothrops lanceolatus, Vipera aspis aspis, Naja naja qui sont parmi les plus utilisés en thérapeutique. Une approche chimiométrique a alors été développée afin de tenir compte de variabilité qualitative et quantitative observée entre différents lots. Cette approche originale nous a permis d’extraire les pics d’intérêts (i.e. présents dans chaque lot d’un même venin) et d’établir une empreinte analytique de référence avec des seuils de conformité quantitatifs. / Although the therapeutic potential of natural substances is known for thousands of years in traditional medicine, it knows an increasing renewed interest in modern pharmaceutical industry as being an inexhaustible source of therapeutic active substances for various diseases.However, the quality control of these products, which is necessary to obtain reproducible biological activities and new commercial approvals, remains a great analytical challenge. Indeed, natural substances show a myriad of biological activities that can result from a specific or synergistic effects of many different components and this information remains still mostly unknown. Furthermore, because they are derived from living organisms and hence are affected by biotic and abiotic factors, it is extremely difficult to ensure a constant qualitative and quantitative composition for these natural substances among different batches. Consequently, there is a need to develop analytical strategies able to assess the “sameness” of natural productions that is analytical strategies able to assess required similarity between different batches by integrating acceptable qualitative and quantitative variations.To achieve this, the concept of analytical fingerprint has become more and more popular and is now starting to be accepted by regulatory authorities such as the FDA. This so-called “pattern-approach” aims at: (i) gaining effective and stable information about common features of a given strain (ii) evaluating similarity and difference with chemometric methods.If this concept has been quite largely studied for the quality assessment of herbal medicines, it is very still very little developed for animal active substances like snake venoms, which are of increasing interest in therapeutic research due to their rich composition in peptides and proteins. The routine quality control of venom raw substances is still often limited to a single gel electrophoresis which is clearly insufficient in terms of components resolution to achieve similarity-/dissimilarity between strains.MethodsIn this context, we have developed an original analytical fingerprint strategy that combines capillary electrophoresis and chemometrics. CE is a particularly well suited technique for peptides/proteins separation allowing to obtain complex specific fingerprints. Batches of different snake venoms have been analyzed with many replicates. All electropherograms have been processed using several chemometrics approaches (baseline correction, signals alignment, automatic recognition of common peaks …) to obtain a representative analytical trace that can be used for the quality assessment of different production lots.ResultsWe show that CE is a very well adapted method to produce highly specific and repeatable analytical profiles of different venom strains. Chemometric methods applied are very efficient to produce an average fingerprint for each strain that integrates features common to all batches and define acceptable variations in quantitative composition. Similarity/dissimilarity of different batches can then be assessed in an automatic manner.ConclusionAll presented results show the efficacy of CE combined with chemometrics to assess the quality of snake venom raw substances. It can be easily implemented in industrial quality control. This strategy can be implemented in future for other type of therapeutic substances of animal origin.

Acetylcholinesterase (AChE) belongs to a distinct family of serine hydrolases and is found in both synaptic and non-synaptic locations. At the synapses, it plays a major role in the hydrolysis of the neurotransmitter acetylcholine (ACh) while in non-synaptic tissue its function is unclear. Snake venom, particularly from the Elapidae family, is a common non-synaptic source of AChE. The venom of krait (Bungarus sindanus), an Elapidae snake, contained a high level of AChE activity. As there is no literature study about Bungarus sindanus venom AChE, it was selected as a main source of AChE activity. We characterized the venom AChE following comparative inhibitory studies with human, horse and rat cholinesterase using different ligands (tacrine, malathion, carbofuran, paraquat, antidepressants, TEMED). The enzyme of krait venom showed optimum activity at alkaline pH 8.5 with an optimal temperature of 45oC. We observed a significant reduction in substrate inhibition of krait venom AChE by using a high ionic strength buffer. With a low ionic strength buffer (10 mM PO4 pH 7.5) the enzyme was inhibited by 1.5 mM AcSCh, while with a high ionic strength buffer (62 mM PO4 pH 7.5) the enzyme was inhibited by 1mM AcSCh. Furthermore, we found that krait 5 venom acetylcholinesterase is thermally stable at 45oC. The enzyme lost only 5% of its activity after incubation at 45oC for 40 min. The Michaelis-Menten constant (Km) for the hydrolysis of acetylthiocholine iodide was found to be 0.052 mM. We noted that snake venom AChE was also inhibited by ZnCl2, CdCl2 and HgCl2 in a concentration dependent manner. In addition, this enzyme showed high sensitivity to tacrine, which is known to inhibit synaptic AChE. We observed that tacrine caused a mixed type of inhibition in krait venom as well as in human serum BChE. Snake venom AChE presents similar inhibitory behavior toward commonly used pesticides and herbicides as that of human serum BChE. The snake venom AChE exhibited a mixed type of inhibition for the pesticides malathion and carbofuran and the herbicide paraquat while human serum BChE presented a mixed inhibition for malathion and paraquat and an uncompetitive inhibition for carbofuran. The krait venom AChE was also affected by antidepressants such as paroxetine, imipramine, clomipramine and sertraline. Paroxetine and sertraline caused a mixed type of inhibition, while imipramine and clomipramine exhibited a competitive inhibition. Moreover, the well-known chemical N,N,N´,N´-tetramethylethylene diamine (TEMED) caused a mixed type of inhibition in snake (Bungarus) venom as well as in horse serum BChE. Futhermore, the inhibition of TEMED, was also confirmed from in vivo study in different structures of the brain, such as striatum, hippocampus, cortex, hypothalamus and cerebellum. Decrease in AChE activity was observed in all treated groups. The results suggest that TEMED exhibits toxic effect via inhibition of cholinesterase. Taken together the krait venom AChE showed similar behavior towards different ligands (tacrine, malathion, carbofuran, paraquat, antidepressants, TEMED) like other sources of cholinestease. Furthermore, krait venom contains large amount of acetylcholinesterase having highest catalytic activity and comparatively more stable than any other sources, making it more valuable for biochemical analysis. / A enzima acetilcolinesterase (AChE, E.C. 3.1.1.7) faz parte de uma família distinta de serina hidrolases, sendo uma importante enzima regulatória, encontrada principalmente no encéfalo, músculos, eritrócitos, neurônios colinérgicos e também em veneno de cobra. Nas sinapses, a sua principal função é a hidrólise no neutrotransmissor acetilcolina (ACh), enquanto que em tecidos não-sinápticos a sua função é desconhecida. O veneno de cobra, principalmente da família Elapidae, é uma fonte não sináptica abundante de AChE. Por exemplo, a espécie Bungarus sindanus, possui um alto conteúdo de AChE. No entanto, na literatura não existem muitos relatos sobre o estudo desta enzima neste veneno. Sendo assim, o veneno de cobra da espécie Bungarus sindanus, foi selecionado comno a principal fonte de AChE neste estudo. A partir dele fez-se estudos de caracterização, comparação com outras fontes de colinesterases (soro eqüino, encéfalo de ratos e sangue humano) e estudos de inibição da enzima por diversos agentes (tacrina, malation, carbofuran, paraquat e 2 antidepressivos). Foi observado que a enzima de veneno de cobra possui um pH ótimo alcalino de pH 8.5 e uma temperatura ótima de ensaio de 45oC. A inibição por substrato, característica cinética da AChE, foi reduzida significativamente ao se usar um tampão com alta força iônica (10 mM PO4 pH 7.5). A enzima possui estabilidade térmica a 45oC. A enzima perdeu somente 5% de sua atividade após 45 min de incubação à 45oC. A constante de Michaelis-Menten (Km) para a hidrólise de ACh foi de 0.052 mM com uma Vmax de 530 μmoles/min/mg de proteína. Também, foi observado que a AChE de veneno da cobra Bungarus sindanus é inibida por ZnCl2, CdCl2 e HgCl2. A enzima apresentou alta sensibilidade a tacrina, a aqual é sabidamente um inibidor da AChE sináptica. A inibição observada foi de tipo mista, tanto em veneno de cobra, tanto quanto em cholinesterase de soro humano (BChE). Também, a enzima apresentou padrão inibitório similar com a colinesterase de soro humano quando foram testados agentes pesticidas e herbicidas. A AChE de veneno de cobra exibiu uma inibição de tipo mista para os pesticidas malation e carbofuran e para o herbicida paraquat. A única diferença foi o padrão de inibição da BChE em relação ao carbofuran, que foi do tipo incompetitivo. A atividade da enzima AChE de veneno de cobra foi inibida, também, pelos antidepressivos paroxetina, imipramina, clomipramina e sertralina. A paroxetina e a sertralina causaram uma inibição de tipo mista, enquanto que a imipramina e a clomipramina exibiram um padrão de inibição competitivo. Também, o bem conhecido composto químico N,N,N´,N´-Tetramethylethylene diamine (TEMED), o qual é rotineiramente utilizado para iniciar processos de polimerização de géis de poliacrilamida causou uma inibição de tipo mista na Ache de veneno de cobra bem como em BChE de soro eqüino. Além disto, a inibição da AChE por TEMED foi observada em diferentes estruturas cerebrais de ratos, tais como estriato, hipocampo, cortex, hipotálamo e cerebelo. Estes resultados confirmam a hipótese de que o TEMED é colinotóxico. Analisando, em conjunto, os resultados podemos concluir que a AChE do veneno da cobra Bungarus Sindanus apresentou padrão cinético inibitório quase similar a outras colinesterases para os compostos tacrina, malation, carbofuran, paraquat, antidepressivos e TEMED. Além disto, o veneno desta cobra possui altas quantidades de AChE o que o torna uma fonte valiosa para estudos bioquímicos e cinéticos desta enzima.

Made available in DSpace on 2014-06-11T19:22:55Z (GMT). No. of bitstreams: 0 Previous issue date: 2006-06-09Bitstream added on 2014-06-13T19:47:18Z : No. of bitstreams: 1 aguiar_mb_me_sjrp.pdf: 1374388 bytes, checksum: 3f0881d7dfbe0cb771b8a1ffb4572604 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Os venenos de vespas socias são ricos em peptídeos biologicamente ativos que causam alguns males ao ser humano como: dores prolongadas, edema, eritema, reações alérgicas e sistêmicas. Possuem em sua composição vários tipos de aminas biogênicas, peptídeos e proteínas. Dentre eles, o que chama mais atenção na atividade farmacológica do veneno são os peptídeos policatiônicos. São diversas as atividades desses peptídeos como: neurotoxicidade, hemólise, liberação de histamina de mastócitos e antibatericida. Neste trabalho, foram estudados peptídeos catiônicos da família Polybine, sintetizados pelo Departamento de Biologia, Unesp, Rio Claro-SP. Os peptídeos Polybine I e II foram sintetizados na forma acetilada e não-acetilada para o estudo detalhado da sua estrutura. Com este objetivo, utilizamos as técnicas de Dicroísmo Circular (CD), para uma análise da estrutura secundária da amostra e espectroscopia de Ressonância Magnética Nuclear (RMN) para o estudo da estrutura tridimensional do peptídeo em solução. / Social wasp venoms are rich of biologically active peptides that may cause some malady to human such as prolonged pains, edema, erythema, allergies and systemic reactions. They have, in its composition, many kinds of biogenic amines, series of polycationics peptides and proteins. Among them, the most interesting thing in pharmacological activity are the polycationics peptides. These peptides show several activities like neurotoxicity, hemolytic activity, histamine releasing activity and antimicrobial activity. In this project, cationic peptides of the Polybine family synthesized by the Department of Biology, CEIS/IBRC, UNESP, Rio Claro, SP were studied. The cationic peptides Polybine I and II were synthesized in acetylad and non-acetylad forms to the detailed study of the structure. This way, CD spectroscopy were performed to analyze secondary structure of the sample and, to analyze treedimension structure, NMR spectroscopy were used.

Made available in DSpace on 2014-06-11T19:23:01Z (GMT). No. of bitstreams: 0 Previous issue date: 2003Bitstream added on 2014-06-13T18:09:09Z : No. of bitstreams: 1 ferreirajunior_rs_me_botfm.pdf: 1446696 bytes, checksum: dbfee5e5aa68ebc4a664a665cf902076 (MD5) / O ensaio imunoenzimático de ELISA foi utilizado para avaliar, acompanhar e comparar a resposta imune humoral de camundongos SWISS durante o processo de hiperimunização com veneno nativo e irradiado com Cobalto-60 (60Co) obtido das serpentes Crotalus durissus terrificus, Bothrops jararaca, Bothrops jararacussu e Bothrops moojeni. A potência e a capacidade de neutralização foram avaliadas por meio de desafios “in vitro”. A imunidade obtida ao final do processo de hiperimunização foi verificada mediante desafio “in vivo” e os efeitos colaterais foram avaliados. A imunização dos animais, com uma DL50 de cada veneno, ocorreu nos dias um, 15, 21, 30 e 45, quando foram colhidas amostras de sangue. No 60º dia ocorreram os desafios. Os resultados demonstraram que o teste de ELISA mostrou-se eficiente na avaliação, acompanhamento e comparação da resposta imune dos camundongos durante o processo de hiperimunização. Os títulos do soro produzido com veneno nativo foram semelhantes aos títulos do soro obtido com veneno irradiado. A capacidade imunogênica foi mantida após a irradiação com 60Co. O soro produzido a partir do veneno irradiado de Crotalus durissus terrificus apresentou potência e capacidade de neutralização maiores, quando comparado ao soro obtido a partir do veneno nativo.Os anticorpos antiveneno nativo e irradiado, mediante os desafios “in vitro”, mostraram-se igualmente eficazes na neutralização dos venenos de Bothrops jararaca, Bothrops jararacussu e Bothrops. Todos anticorpos foram capazes de neutralizar cinco DL50 dos respectivos venenos. As alterações clínicas foram mínimas durante o processo de hiperimunização com veneno irradiado e evidentes com veneno bruto. / ELISA was used to evaluate, follow, and compare the humoral immune response of Swiss mice during hyperimmunization with natural and Cobalt-60- irradiated (60Co) Crotalus durissus terrificus, Bothrops jararaca, Bothrops jararacussu, and Bothrops moojeni venoms. Potency and neutralization were evaluated by in vitro challenges. After hyperimmunization, immunity was observed by in vivo challenge and the side effects were assessed. The animals’ immunization with one LD50 of each venom was on days one, 15, 21, 30, and 45, when blood samples were collected; the challenges occurred on the 60th day. Results showed that ELISA was efficient in evaluating, following, and comparing mouse immune response during hyperimmunization. Serum titers produced with natural venom were similar to those with irradiated venom. Immunogenic capacity was maintained after 60Co irradiation. Serum produced from Crotalus durissus terrificus irradiated venom showed higher potency and neutralization capacity than that from natural venom. Antibodies from natural and irradiated venom by in vitro challenges were also efficient in neutralizing Bothrops jararaca, Bothrops jararacussu, and Bothrops moojeni venoms. All antibodies were capable of neutralizing five LD50 from these venoms. Clinical alterations were minimum during hyperimmunization with irradiated venom and evident with natural venom.