Deoxynivalenol (DON) is a mycotoxin that affects the intestinal morphology of animals, impairing nutrient intake and growth. On the other hand, dietary supplementation with functional oligosaccharides as chito-oligosaccharides (COS) has shown positive effects on the intestinal health of piglets. Therefore, the objective of the present study was to evaluate the effect of low doses of COS in preventing DON-induced intestinal histological changes, using a swine jejunal explant technique. The intestinal explants were incubated at 37 °C in culture medium for 4 h and exposed to the following treatments: (a) control (only culture medium), (b) DON (10 µM), (c) 25COS (0.025 mg·mL−1 of COS); (d) 50COS (0.05 mg·mL−1 of COS); (e) 25COS plus DON (25COS + DON); (f) 50COS plus DON (50COS + DON). Explants exposed to COS presented intestinal morphology similar to control samples. DON induced a significant decrease in the histological score as a consequence of moderate to severe histological changes (apical necrosis, villi atrophy, and fusion) and a significant decrease in morphometric parameters (villi height, crypt depth, villi height:crypt depth ratio, and goblet cells density). The intestinal morphology of samples exposed to COS + DON remained similar to DON treatment. In conclusion, low levels of COS did not counteract DON-induced intestinal lesions.
Antibiotics have been widely used in piglet diets to promote growth performance and reduce diarrhea incidence. However, the resistance of pathogens to antibiotics and the risk of residues of antibiotics in animal products induced a growing interest in the use of alternatives to in-feed antibiotics. Chito-oligosaccharide (COS), a natural alkaline polymer of glucosamine is currently being tested as a substitute for in-feed antibiotics. In weaned piglets, COS has positive effects on promoting growth, which may be related to its action on intestinal morphology, immune ability and beneficial microbiota. However, previous studies shown variable results with effective doses ranging from 30 mg/kg to 5 g/kg. Therefore, the goal of this study was to test the hypothesis that the use of COS can be an alternative to in-feed antibiotics by improve the intestinal morphology of piglets, using the jejunal explant model. The intestinal explants were exposed for 4 h to following treatments: control - only culture media and culture media with COS in doses of 0.025 mg/ml, 0.05 mg/ml, 0.1 mg/ml and 0.15 mg/ml. After the incubation period the explants were processed for histological and morphometrical analysis. The histological changes were evaluated using an adapted histological score based on the intensity and severity of lesions. Mild histological changes were observed in jejunal explants exposed to different treatments; however, no significant difference in the histological score, villi height, crypt depth or villus : crypt ratio were observed between the COS-groups and the control. In addition, goblet cells density in intestinal explants exposed to COS remained statistically similar to control group. Our results indicate that COS exposure in levels ranging from 0.025 to 0.15 mg/ml induced no effect on intestinal morphology of pig's explants. The research will provide guidance on the low dosage of COS supplementation on weaning pigs.
RESUMO:A presente pesquisa avaliou a ação mutagênica e antimutagênica de um biopolímero de glucose extraído da Agrobacterium radiobacter (Biopolímero de Agrobacterium radiobacter). O experimento foi realizado com camundongos Swiss machos divididos em oito grupos. O tratamento com o biopolímero foi realizado por gavage em dose única concomitante a uma dose de solução tampão fosfato nos grupos de avaliação da mutagenicidade, ou ao agente indutor de danos no DNA, ciclofosfamida, na concentração de 50 mg/kg (peso corpóreo -p.c.), nos grupos de avaliação da antimutagenicidade. Utilizou-se o teste de micronúcleo em sangue periférico e a coleta de sangue foi realizada 24 e 48 h após a aplicação das substâncias-teste. A análise estatística demonstrou que o biopolímero não possui atividade mutagênica e que é efetivo em prevenir danos no DNA. As porcentagens de redução de danos nos grupos de antimutagenicidade foram de 83,9%, 89,1% e 103,1% em 24 h e 101,24%, 98,14% e 120,64% em 48 h para as doses de 75, 150 e 300mg/kg (p.c.), respectivamente. A alta porcentagem de redução de danos associada à ausência de efeitos mutagênicos indica, além da atividade quimioprotetora, a possibilidade do biopolímero ser um alimento funcional candidato à utilização como co-adjuvante na quimioterapia para prevenir efeitos colaterais.Unitermos: Curdlana, biopolímero, Agrobacterium radiobacter, β-glucana, antimutagenicidade, camundongos Swiss.ABSTRACT: "Assessment of mutagenicity and antimutagenicity of a biopolymer extracted from the microorganism Agrobacterium radiobacter in mice." This study evaluated the mutagenic and ant mutagenic action of a biopolymer of glucose extracted from Agrobacterium radiobacter (Biopolymer of Agrobacterium radiobacter). The experiment was conducted with Swiss male mice divided into eight groups. Treatment with the biopolymer was performed in a single dose by gavage at a dose of concomitant phosphate buffer groups in the evaluation of mutagenicity, or the agent of inducing DNA damage, cyclophosphamide, the concentration of 50 mg/kg (body weight --b.w.), in groups of assessment ant mutagenic. We used the micronucleus test in peripheral blood. The blood sample was held 24 and 48 h after application of the test substances. Statistical analysis showed that the biopolymer has no mutagenic activity and it is effective in preventing damage to DNA. The percentages of damage reduction in groups of ant mutagenic were 83.9%, 89.1% and 103.1% in 24 h and 101.24%, 98.14% and 120.64% at doses of 48 to 75, 150 and 300 mg/kg (b.w.) respectively. The high percentage of damage reduction associated with the absence of mutagenic effects indicates the possibility of biopolymer chemoprotection action. It can also be considered a functional food candidate to be used as co-adjuvant chemotherapy to prevent side effects.
ResumoNo presente estudo, utilizando melaço de cana-de-açúcar como única fonte de carbono para Agrobacterium radiobacter k84 foi obtido, em processo fermentativo, um biopolímero composto por carboidratos (35%), proteínas (15%) e minerais (40%). O rendimento do processo foi de 10 g.L -1 do biopolímero que apresentou elevada solubilidade em água, pH neutro em solução aquosa e baixa atividade de água (0.52). As análises em Microscopia Eletrônica de Varredura revelaram microestrutura característica de um sólido amorfo, com partículas de formas irregulares e tamanhos variáveis. Na avaliação das propriedades tecnológicas, o biopolímero mostrou viscosidade à temperatura ambiente a partir da concentração 0.5% em solução aquosa, atividade geleificante em solução a 2%, atividade emulsificante (56.11±0.78%) e estabilizante (98.02±1.39%). Os resultados sugerem o biopolímero de Agrobacterium radiobacter k84 como um candidato promissor para uso industrial. AbstractIn this study, a biopolymer composed of carbohydrates (35%), protein (15%) and minerals (40%) was obtained through fermentation using sugar cane molasses as the sole carbon source for Agrobacterium radiobacter k84. The process yield was 10 gL -1 of biopolymer, which showed high solubility in water, neutral pH in aqueous solution and low water activity (0.52). The analysis in Scanning Electron Microscopy revealed microstructure characteristic of an amorphous solid, with particles of irregular shapes and sizes. In the evaluation of technological properties, the biopolymer showed formation of viscous solutions at room temperature from concentration of 0.5% in aqueous solution, gelling activity in solution at 2%, emulsifying (56.11±1.39%) and stabilizing activity (98.02±0.78%). The results suggest that the biopolymer from Agrobacterium radiobacter k84 is a promising candidate for industrial use.
Sugar cane molasses is a cheaper carbon source alternative than glucose traditionally used in fermentation processes. In the present study a biopolymer soluble from Agrobacterium radiobacter k84 (ARB) was obtained by fermentation using sugar cane molasses as a carbon source in a process with yield of 10.0 g.L -1. The ARB is composed by minerals (40%), carbohydrate (35%) and protein (15%). In vitro test of the cytotoxic effect of ARB at concentrations 2.5 mg/mL, 5.0 mg/mL and 10.0 mg/mL in LLC MK 2 (Rhesus Monkey Kidney) cells revealed a 50% cytotoxic concentration (CC 50 ) of 9.32 mg/mL. In a 30-day in vivo oral toxicity study, Swiss mice were administered ARB by gavage at 5 mg/mL, 15 mg/mL, 50 mg/mL and 150 mg/mL (approximately 25 mg/kg/day, 75 mg/kg/day, 250 mg/kg/day and 750 mg/kg/day). The results did not present any hematological or histopathological signs of adverse effects, leading us to define the no observed adverse effect level (NOAEL) as 150 mg/mL (approximately 750 mg/kg/day). Key words: Cane molasses, fermentation, cytotoxicity DOI: 10.5433/1679 Abstract O melaço de cana-de-açúcar é uma fonte de carbono alternativa de menor custo que a glicose tradicionalmente utilizada em processos fermentativos. No presente estudo, um biopolímero solúvel de Agrobacterium radiobacter k84 (ARB) foi obtido por fermentação utilizando melaço de cana de açúcar como fonte de carbono em um processo com rendimento de 10,0 g.L -1 . O ARB é composto de minerais (40%), carboidratos (35%) e proteínas (15%). O teste do efeito citotóxico do ARB in vitro nas concentrações de 2,5 mg/mL, 5,0 mg/mL e 10,0 mg/mL em células LLC MK 2 (Rim de Macaco Rhesus) revelou uma concentração citotóxica 50% (CC 50 ) de 9,32 mg/mL. Em estudo in vivo de toxicidade oral durante 30 dias, camundongos Swiss receberam por gavagem soluções de ARB nas concentrações de 5 mg/mL, 15 mg/mL, 50 mg/mL e 150 mg/mL (aproximadamente 25 mg/kg/dia, 75 mg/kg/dia, 250 mg/ kg/dia e 750 mg/kg/dia). Os resultados não apresentaram sinais hematológicos ou histopatológicos de efeitos adversos, levando a definir a dose sem efeito adverso observado (NOAEL) como 150 mg/mL (aproximadamente 750 mg/kg/dia). Palavras-chave: Melaço de cana, fermentação, citotoxicidade
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