Brazilian freshwater stingrays, Potamotrygon gr. orbigyni, are relatively common in the middle-western regions of Brazil, where they are considered an important public health threat. In order to identify some of their naturally occurring toxin peptides available in very low amounts, we combine analytical protocols such as reversed-phase high-performance liquid chromatography (RP-HPLC), followed by a biological microcirculatory screening and mass spectrometry analysis. Using this approach, one bioactive peptide was identified and characterized, and two analogues were synthesized. The natural peptide named Porflan has the primary structure ESIVRPPPVEAKVEETPE (MW 2006.09 Da) and has no similarity with any bioactive peptide or protein found in public data banks. Bioassay protocols characterized peptides as presenting potent activity in a microcirculatory environment. The primary sequences and bioassay results, including interactions with the membrane phospholipids, suggest that these toxins are a new class of fish toxins, directly involved in the inflammatory processes of a stingray sting.

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In order to investigate the relationship between the primary structure of Orpotrin, a vasoactive peptide previously isolated from the freshwater stingray Potamotrygon gr. orbignyi, and its microcirculatory effects, three Orpotrin analogs were synthesized. The analogs have a truncated N-terminal with a His residue deletion and two substituted amino acid residues, where one Nle is substituted for one internal Lys residue and the third analog has a substitution of a Pro for an Ala (Orp-desH(1) , Orp-Nle and Orp-Pro/Ala, respectively). Only Orp-desH(1) could induce a lower vasoconstriction effect compared with the natural Orpotrin, indicating that besides the N-terminal, the positive charge of Lys and the Pro residues located at the center of the amino acid chain is crucial for this vasoconstriction effect. Importantly, the suggestions made with bioactive peptides were based on the molecular modeling and dynamics of peptides, the presence of key amino acids and shared activity in microcirculation, characterized by intravital microscopy. Moreover, this study has demonstrated that even subtle changes in the primary structure of Orpotrin alter the biological effects of this native peptide significantly, which could be of interest for biotechnological applications.

As espécies de arraias de água doce (família Potamotrygonidae) têm sua história evolutiva intrinsecamente conectada à história geológica das bacias hidrográficas neotropicais. Os representantes desse grupo acumulam uma série de adaptações fisiológicas que possibilitam a manutenção de suas populações em condições ambientais variáveis, sendo a maioria dessas ainda desconhecidas. Assim, este trabalho teve por objetivo analisar as respostas fisiológicas das arraias Potamotrygon rex, frente às variações do ambiente, ocorridas durante o ciclo hidrológico (períodos chuvoso/estiagem) na região do Alto/Médio rio Tocantins. O ambiente foi verificado na tomada de amostras da água durante sete semanas consecutivas, no período chuvoso e de estiagem, na mesma região onde foram coletadas as arraias. Os espécimes foram coletados na área de transição do reservatório do Lajeado, no rio Tocantins, dos quais foram registrados os dados biométricos e coletadas amostras de sangue para verificação dos indicadores fisiológicos (hematócrito, hemoglobina e glicose plasmática). Foi calculado o fator de condição relativo das arraias (Kn), e a relação ente os dados biométricos e as variáveis fisiológicas foi avaliada pela correlação de Pearson. Como resultado, verificou-se as alterações ambientais que ocorrem sazonalmente, durante o ciclo hidrológico (períodos chuvoso e estiagem). No período chuvoso (de outubro a abril), a temperatura da água (°C), o oxigênio dissolvido na água (mg/L) e a turbidez apresentaram valores mais elevados do que no período de estiagem (de maio a setembro), com exceção da condutividade elétrica. Para os espécimes coletados na estiagem verificou-se o aumento do peso, do Kn, do hematócrito, hemoglobina e glicose plasmática. Contudo, não houve correlação significativa entre os dados biométricos e fisiológicos. Dessa forma, as mudanças ocorridas na fisiologia de Potamotrygon rex sugerem plasticidade da espécie para suportar as variações sazonais da região.

Freshwater stingrays are benthic animals that almost always behave passively and preferentially inhabit shallow water. Their tail barb is rigid and serrated, and is covered by glandular epithelium. Injury from the barb is followed by local symptoms such as bleeding, intense throbbing pain, and edema in the affected member. Nausea, salivation, convulsion, and tachycardia are reported systemic changes. The Potamotrygon sp. C is stingray species most often encountered in rivers of the state of Tocantins, Brazil.ObjectiveTo evaluate the physiopathological changes induced by poisoning caused by Potamotrygon sp. C.MethodsMucus was removed from the epithelial tissue at the barb and from the disc of the animal (far from the barb), and an extract was prepared from the glandular epithelium that covers the barb and analized. Different concentrations of this material (25, 50, 100 μg/ml) were used to evaluate the edematogenic, nociceptive, hemorrhagic, and necrotic effects in male and female Swiss mice.ResultsThe nociceptive and edematogenic activity was dose‐dependent, with the most intense response observed for the mucus collected from the disc. No hemorrhagy was observed, and necrosis was limited to small dots on the tissue evaluated.ConclusionIt was evident that the greatest changes caused by poisoning with Potamotrygon sp. C are related to contact of the barb‐injured tissue with the mucus that covers the animal.