Aplicação de nanopartículas de quitosana com potencial de adjuvante na produção de soro contra o venneno do escorpião Tityus serrulatus

In Brazil, several species of scorpions are known to cause accidents which can lead to death, which are mainly belonging to the genus Tityus. The scorpion Tityus serrulatus is the main responsible for more severe cases. Anti-scorpion serums are routinely produced by various institutions, despite their effectiveness, quality and action depends on how quickly treatment is started. Studies have been developed in the search for appropriate technologies to encapsulate and release recombinant or natives proteins capable of inducing antibody production. In this context, chitosan copolymer which can be obtained from the partial deacetylation of chitin or in some microorganisms and it is biocompatible and biodegradable has been widely used for this purpose. This study aimed to search for a system release from chitosan nanoparticles for peptide / protein of the venom of the scorpion T. serrulatus, able to provide a new model of immunization in animals, in order to obtain a potential novel polyclonal serum, anti-venom T. serrulatus. The chitosan nanoparticles were prepared by ionic gelation with polyanion tripolyphosphate (TPP). After standardizing the concentrations of TPP and chitosan was evaluated the efficiency of incorporation of bovine serum albumin (BSA) and scorpion venom, showed particle size compatible with the intended purpose. The particles showed adequate size around 200nm. The crosslinking was confirmed by absorption spectroscopy in the infrared. After verified the high encapsulation efficiency (EE) for acid bicinconínico method (BCA) protein assay and the particle size distribution, the success of the technique was proven and the potential for in vivo application of nanoparticles. The experimental animals were vaccinated and the antibodies measured by ELISA

Eletrofiação e caracterização de membranas biopoliméricas a base de quitosana extraídas dos exoesqueletos de crustáceos

Chitin and chitosan are nontoxic, biodegradable and biocompatible polymers produced by renewable natural sources with applications in diverse areas such as: agriculture, textile, pharmaceutical, cosmetics and biomaterials, such as gels, films and other polymeric membranes. Both have attracted greater interest of scientists and researchers as functional polymeric materials. In this context, the objective of this study was to take advantage of the waste of shrimp (Litopenaeus vannamei and Aristeus antennatus) and crabs (Ucides cordatus) from fairs, beach huts and restaurant in Natal/RN for the extraction of chitin and chitosan for the production of membranes by electrospinning process. The extraction was made through demineralization, deproteinization, deodorization and deacetylation. Morphological analyzes (SEM and XRD), Thermal analysis (TG and DTG), Spectroscopy in the Region of the Infrared with Transformed of Fourier (FTIR) analysis Calorimetry Differential Scanning (DSC) and mechanical tests for traction were performed. In (XRD) the semicrystalline structure of chitosan can be verified while the chitin had higher crystallinity. In the thermal analysis showed a dehydration process followed by decomposition, with similar behavior of carbonized material. Chitosan showed temperature of maximum degradation lower than chitin. In the analysis by Differential Scanning Calorimetry (DSC) the curves were coherent to the thermal events of the chitosan membranes. The results obtained with (DD) for chitosan extracted from Litopenaeus vannamei and Aristeus antennatus shrimp were (80.36 and 71.00%) and Ucides cordatus crabs was 74.65%. It can be observed that, with 70:30 solutions (v/v) (TFA/DCM), 60 and 90% CH3COOH, occurred better facilitate the formation of membranes, while 100:00 (v/v) (TFA/DCM) had formation of agglomerates. In relation to the monofilaments diameters of the chitosan membranes, it was noted that the capillary-collector distance of 10 cm and tensions of 25 and 30 kV contributed to the reduction of the diameters of membranes. It was found that the Young s modulus decreases with increasing concentration of chitosan in the membranes. 90% CH3COOH contributed to the increase in the deformation resulting in more flexible material. The membranes with 5% chitosan 70:30 (v/v) (TFA/DCM) had higher tensile strength

Estudos das interações de quitosana/CTAB/C12E8

Surfactant-polymer interactions are widely used when required rheological properties for specific applications, such as the production of fluids for oil exploration. Studies of the interactions of chitosan with cationic surfactants has attracted attention by being able to cause changes in rheological parameters of the systems making room for new applications. The commercial chitosan represents an interesting alternative to these systems, since it is obtained from partial deacetylation of chitin: the residues sites acetylated can then be used for the polymer-surfactant interactions. Alkyl ethoxylated surfactants can be used in this system, since these non-ionic surfactants can interact with hydrophobic sites of chitosan, modifying the rheology of solutions or emulsions resultants, which depends on the relaxation phenomenon occurring in these systems. In this work, first, inverse emulsions were prepared from chitosan solution as the dispersed phase and cyclohexane as the continuous phase were, using CTAB as a surfactant. The rheological analysis of these emulsions showed pronounced pseudoplastic behavior. This behavior was attributed to interaction of "loops" of chitosan chains. Creep tests were also performed and gave further support to these discussions. Subsequently, in order to obtain more information about the interaction of chitosan with non-ionic surfactants, solutions of chitosan were mixed with C12E8 and and carried out rheological analysis and dynamic light scattering. The systems showed marked pseudoplastic behavior, which became less evident when the concentration of surfactant was increased. Arrhenius and KWW equations were used to obtain parameters of the apparent activation energy and relaxation rate distribution, respectively, to which were connected to the content of surfactant and temperature used in this work

Inclusões de quitosana no subcutâneo de rato: avaliação clínica, histológica e morfométrica

FUNDAMENTOS: A quitosana é polímero derivado da quitina, com vários tipos de aplicação na área médica. OBJETIVO: Avaliar a biocompatibilidade de membranas de quitosana no subcutâneo de ratos. MÉTODOS: Foram utilizados 20 ratos Wistar machos, nos quais foram implantadas membranas de quitosana, na região mediana dorsal. Os animais foram sacrificados: sete, 15, 30 e 60 dias após a cirurgia, tendo sido avaliados clinicamente durante o período experimental e com fotodocumentação no momento do sacrifício. Após o sacrifício, as membranas e tecidos adjacentes foram removidos e preparados para exame histológico e morfométrico. RESULTADOS: Nenhum animal apresentou efeitos adversos que pudessem ser atribuídos à implantação das membranas. O exame histológico mostrou que as inclusões são lisas e homogêneas e não são colonizadas por células do hospedeiro, sendo circundadas por pseudocápsula composta por fibroblastos e células inflamatórias. A morfometria da pseudocápsula revelou espessura semelhante durante todo o período experimental (P>0,05). CONCLUSÃO: A quitosana pode ser opção para uso como implante não integrado. Novos estudos devem ser realizados para comprovar a biocompatibilidade a longo prazo.

Histological evaluation of chitosan-based biomaterials used for the correction of critical size defects in rat's calvaria

Chitosan, a biopolymer obtained from chitin, and its derivates, such as chitosan hydrochloride, has been reported as wound healing accelerators and as possible bone substitutes for tissue engineering, and therefore these Substances could be relevant in dentistry and periodontology. The purpose of this investigation was to make a histological evaluation of chitosan and chitosan hydrochloride biomaterials (gels) used in the correction of critical size bone defects made in rat's calvaria. Bone defects of 8 mm in diameter were surgically created in the calviria of 50 Holtzman (Rattus norvegicus) rats and filled with blood clot (control), low molecular weight chitosan, high molecular weight chitosan, low molecular weight chitosan hydrochloride, and high molecular weight chitosan hydrochloride, numbering 10 animals, divided into two experimental periods (15 and 60 days), for each biomaterial. The histological evaluation was made based on the morphology of the new-formed tissues in defect's region, and the results indicated that there was no statistical difference between the groups when the new bone formation in the entire defect's area were compared (p > 0.05) and, except in the control groups, assorted degrees of inflammation Could be Seen. In Conclusion, chitosan and chitosan hydrochloride biomaterials used in this study were not able to promote new bone formation in critical size defects made in rat's calvaria. (C) 2009 Wiley Periodicals, Inc. J Biomed Mater Res 93A: 107-114, 2016

Evaluation of chitosan gel as antibiotic and photosensitizer delivery

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Different Molecular Weight Chitosan-Based Membranes for Tissue Regeneration

Natural polymers, such as chitosan, obtained from chitin, are been widely studied for use in the tissue regeneration field. This study established a protocol to attain membranes made from this biopolymer, consisting of high or low molecular weight chitosan. The biocompatibility of these membranes was histologically evaluated, comparing them to collagen membrane surgically implanted in rat subcutaneous tissue. Fifteen Holtzmann rats were divided in three experimental groups: High and Low Molecular Weight Chitosan membranes (HMWC and LMWC) and Collagen membranes (C-control group); each of them with three experimental periods: 7, 15 and 30 days. As a result, after the seven days evaluation, the membranes were present and associated with a variable degree of inflammation, and after the 15 and 30 days evaluations, the membranes were absent in all groups. It is concluded that the chitosan-based membranes were successfully attained and presented comparable resorption times to collagen membranes.

Enzymatic activity of hypopharyngeal gland extractsfrom workers of Apis mellifera (Hymenoptera, Apidae, Apinae)

This research presents a comparative study of enzymatic activity of the hypopharyngeal gland extracts from workers of Apis mellifera in three physiologic stages: newly emerged, nurse and forager workers, with the objective of contributing to the comprehension of the gland function. In order to determinate the enzymes present in the extracts, the Api Zym kit (Bio Merieux) was used to test the activity of 19 different enzymes. The enzymes found in larger amounts only in the hypopharyngeal glands from certain individuals were the following: in newly emerged workers, the N-acetyl-double down arrow-glucosaminidase that may be digesting the chitin of some food ingested by the bee; in forager workers, the acid phosphatase that is likely acting in authophagic processes, the a-glucosidase, in the processing of nectar into honey, and the double down arrow-glucosidases, in the pollen digestion.

Venomous mollusks: the risks of human accidents by conus snails (gastropoda: conidae) in Brazil

Os moluscos do gênero Conus apresentam um aparato venenoso composto de uma rádula quitinosa ligada a glândulas de peçonha, causando envenenamentos humanos graves e mesmo óbitos pela ação neurotóxica indutora do bloqueio de vários receptores e paralisia muscular. Não há casos descritos de envenenamento no país, mas determinadas populações correm risco de acidentes.

cDNA cloning and 1.75 angstrom crystal structure determination of PPL2, an endochitinase and N-acetylglucosamine-binding hemagglutinin from Parkia platycephala seeds

Parkia platycephala lectin 2 was purified from Parkia platycephala (Leguminosae, Mimosoideae) seeds by affinity chromatography and RP-HPLC. Equilibrium sedimentation and MS showed that Parkia platycephala lectin 2 is a nonglycosylated monomeric protein of molecular mass 29 407 +/- 15 Da, which contains six cysteine residues engaged in the formation of three intramolecular disulfide bonds. Parkia platycephala lectin 2 agglutinated rabbit erythrocytes, and this activity was specifically inhibited by N-acetylglucosamine. In addition, Parkia platycephala lectin 2 hydrolyzed beta(1-4) glycosidic bonds linking 2-acetoamido-2-deoxy-beta-D-glucopyranose units in chitin. The full-lengthamino acid sequence of Parkia platycephala lectin 2, determined by N-terminal sequencing and cDNA cloning, and its three-dimensional structure, established by X-ray crystallography at 1.75 angstrom resolution, showed that Parkia platycephala lectin 2 is homologous to endochitinases of the glycosyl hydrolase family 18, which share the (beta alpha)(8) barrel topology harboring the catalytic residues Asp125, Glu127, and Tyr182.

Effect of the insect growth regulator diflubenzuron on the predator Podisus nigrispinus (Heteroptera: Pentatomidae)

Podisus nigrispinus (Dallas) (Heteroptera: Pentatomidae) is a common natural predator of defoliating caterpillars in agricultural and forest systems. Insecticides acting as growth regulators of insect pests can indirectly affect their predators through consumption of contaminated prey. We examined the reproductive performance of P. nigrispinus fed on caterpillars of Anticarsia gemmatalis Hubner (Lepidoptera: Noctuidae) reared on soybean leaves exposed to the chitin synthesis inhibitor, diflubenzuron. Caterpillars of A. gemmatalis were fed for 12 h with treated soybean leaves and offered to adults of the predator P. nigrispinus over five consecutive days. The fertility of P. nigrispinus was reduced when feeding on diflubenzuron treated caterpillars, especially at the beginning of the reproductive period, but recovered 3 weeks later. The effects of diflubenzuron ingestion on the life table parameters of P. nigrispinus included an increase in the period taken to double the population size, and reductions in the intrinsic rate of population increase, generation duration, and net reproductive rate. Diflubenzuron therefore had an indirect negative effect on the reproduction and the population dynamics of the non-target predator P. nigrispinus. Clearly, its use in integrated pest management requires further evaluation.

Enzymatic activity of hypopharyngeal gland extracts from workers of Apis mellifera (Hymenoptera, Apidae, Apinae)

This research presents a comparative study of enzymatic activity of the hypopharyngeal gland extracts from workers of Apis mellifera in three physiologic stages: newly emerged, nurse and forager workers, with the objective of contributing to the comprehension of the gland function. In order to determinate the enzymes present in the extracts, the Api Zym kit (Bio Mérieux) was used to test the activity of 19 different enzymes. The enzymes found in larger amounts only in the hypopharyngeal glands from certain individuals were the following: in newly emerged workers, the N-acetyl-down double arrow sign-glucosaminidase that may be digesting the chitin of some food ingested by the bee; in forager workers, the acid phosphatase that is likely acting in authophagic processes, the a-glucosidase, in the processing of nectar into honey, and the down double arrow sign-glucosidases, in the pollen digestion.

cDNA cloning and 1.75 Å crystal structure determination of PPL2, an endochitinase and N-acetylglucosamine-binding hemagglutinin from Parkia platycephala seeds

Parkia platycephala lectin 2 was purified from Parkia platycephala (Leguminosae, Mimosoideae) seeds by affinity chromatography and RP-HPLC. Equilibrium sedimentation and MS showed that Parkia platycephala lectin 2 is a nonglycosylated monomeric protein of molecular mass 29 407 ± 15 Da, which contains six cysteine residues engaged in the formation of three intramolecular disulfide bonds. Parkia platycephala lectin 2 agglutinated rabbit erythrocytes, and this activity was specifically inhibited by N-acetylglucosamine. In addition, Parkia platycephala lectin 2 hydrolyzed β(1-4) glycosidic bonds linking 2-acetoamido-2-deoxy-β-d-glucopyranose units in chitin. The full-length amino acid sequence of Parkia platycephala lectin 2, determined by N-terminal sequencing and cDNA cloning, and its three-dimensional structure, established by X-ray crystallography at 1.75 Å resolution, showed that Parkia platycephala lectin 2 is homologous to endochitinases of the glycosyl hydrolase family 18, which share the (βα) 8 barrel topology harboring the catalytic residues Asp125, Glu127, and Tyr182. © 2006 The Authors.

Susceptibility/resistance of Anticarsia gemmatalis larvae to its nucleopolyhedrovirus (AgMNPV): Structural study of the peritrophic membrane

This investigation compares the peritrophic membrane (PM) morphology along the midgut of susceptible (SL) and resistant (RL) Anticarsia gemmatalis larvae to the AgMNPV. The PM increased the thickness from the anterior to the posterior midgut region in both insects strain; however, the intensity of FITC-WGA reaction of the PM in the RL were greater than in SL. The PM in RL was ultrastructurally constituted by several layers of fibrous/vesicular materials in comparison with the few ones in SL. Our results showed that the structure of PM in the RL could be one of the resistance barriers to AgMNPV. © 2007.

Toxicidade aguda e risco ambiental do diflubenzuron para Daphnia magna, Poecilia reticulata e Lemna minor na ausência e presença de sedimento

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)