Composição centesimal e de ácidos graxos do músculo Longíssimus de cordeiros confinados, alimentados com dietas contendo casca de mandioca / Centesimal and fatty acid composition of the longissimus muscle of confined lambs fed diets containing cassava peels

Objetivou-se avaliar os efeitos de inclusão da casca da mandioca (CM) sobre a composição centesimal e de ácidos graxos do músculo Longíssimus de cordeiros. Foram utilizados 32 cordeiros mestiços Santa Inês, machos não castrados, peso corporal médio de 21 ± 1,5kg. Os tratamentos foram distribuídos em delineamento inteiramente ao acaso aos animais, cujo os níveis de inclusão da CM foram (0, 10, 20, 30%) baseado na matéria seca da dieta. Utilizou-se como volumoso o feno da parte aérea de mandioca, e a relação volumoso:concentrado foi de 48:52. O experimento teve duração de 70 dias. Logo após, os cordeiros passaram por um jejum sólido de 16h e, depois, foram abatidos. A carcaça foi conduzida à câmara fria, permanecendo por 24h, a uma temperatura de 4°C. Na carcaça fria, foi retirada uma amostra do músculo Longíssimus, a qual foi congelada (4°C), até o início das análises. Os resultados de umidade, cinzas e proteína não foram influenciados pela CM, observando-se efeito linear decrescente para os teores de lipídeos. Com relação à composição de ácidos graxos, houve efeito linear para o C15:0 e efeito quadrático para C14:0, C18:0 e C22:0. Nos monoinsaturados, observou-se efeito linear para C16:1, C18:1-9c e C22:1-9c e efeito quadrático para C15:1, C17:1 e C20:1; já nos poli-insaturados, foi observado efeito linear para CLA, C20:3n-6, C20:4n-6, C20:5n-3 e C22:6n-3, e quadrático para C20:2 e C18:3n-6. Assim, conclui-se que a composição centesimal e de ácidos graxos apresenta variações em razão da inclusão da CM, porém não compromete a qualidade da carne.(AU)

This study sought to evaluate the effects of the inclusion of cassava peel on the centesimal composition and fatty acids of the Longissimus muscle of lambs. We used 32 uncastrated crossbred Santa Inês lambs, with average body weight of 21 ± 1.5 kg. Treatments were in a completely randomized design, with the inclusion of cassava peel (0, 10, 20, 30%) in the diet dry matter. Hay from the cassava shoot has been used as forage and the forage:concentrate ratio was 48:52. The experiment lasted 70 days and then the lambs underwent a fast for 16 hours, were slaughtered, and their carcasses remained at a temperature of 4°C for 24 hours. In cold carcass the Longissimus muscle were taken and frozen (4°C) until the analysis. The results for moisture, ashes and protein were not influenced by cassava peel, and a decreasing linear effect was observed for the lipids levels. In composition of unsaturated fatty acids, there was a linear effect for C15:0 and a quadratic effect for C14:0, C18:0 and C22:0. Regarding the monounsaturated fatty acids, a linear effect has been observed for C16:1, C18:1-9c and C22:1-9c, and a quadratic effect for C15:1, C17:1 and C20:1. With the polyunsaturated fatty acids, a linear effect was observed for CLA, C20:3n-6, C20:4n-6, C20:5n-3 and C22:6n-3, and a quadratic effect was seen for C20:2 and C18:3n-6. Thus, it is concluded that the centesimal and fatty acid composition varies depending on the inclusion of cassava peel, however, it does not compromise the quality of the meat.(AU)

Simultaneous detection of the C282Y, H63D and S65C mutations in the hemochromatosis gene using quenched-FRET real-time PCR.

Hereditary hemochromatosis (HH) is a common autosomal disorder of iron metabolism mainly affecting Caucasian populations. Three recurrent disease-associated mutations have been detected in the hemochromatosis gene (HFE): C282Y, H63D, and S65C. Although HH phenotype has been associated with all three mutations, C282Y is considered the most relevant mutation responsible for hemochromatosis. Clinical complications of HH include cirrhosis of the liver, congestive cardiac failure and cardiac arrhythmias, endocrine pancreatic disease, which can be prevented by early diagnosis and treatment. Therefore, a reliable genotyping method is required for presymptomatic diagnosis. We describe the simultaneous detection of the C282Y, H63D and S65C mutations in the hemochromatosis gene by real-time PCR followed by melting curve analysis using fluorescence resonance energy transfer (FRET) probes. The acceptor fluorophore may be replaced by a quencher, increasing multiplex possibilities. Real-time PCR results were compared to the results of sequencing and conventional PCR followed by restriction digestion and detection by agarose gel electrophoresis (PCR-RFLP). Genotypes from 80 individuals obtained both by the conventional PCR-RFLP method and quenched-FRET real-time PCR were in full agreement. Sequencing also confirmed the results obtained by the new method, which proved to be an accurate, rapid and cost-effective diagnostic assay. Our findings demonstrate the usefulness of real-time PCR for the simultaneous detection of mutations in the HFE gene, which allows a reduction of a significant amount of time in sample processing compared to the PCR-RFLP method, eliminates the use of toxic reagents, reduces the risk of contamination in the laboratory, and enables full process automation.

Simultaneous detection of the C282Y, H63D and S65C mutations in the hemochromatosis gene using quenched-FRET real-time PCR

Hereditary hemochromatosis (HH) is a common autosomal disorder of iron metabolism mainly affecting Caucasian populations. Three recurrent disease-associated mutations have been detected in the hemochromatosis gene (HFE): C282Y, H63D, and S65C. Although HH phenotype has been associated with all three mutations, C282Y is considered the most relevant mutation responsible for hemochromatosis. Clinical complications of HH include cirrhosis of the liver, congestive cardiac failure and cardiac arrhythmias, endocrine pancreatic disease, which can be prevented by early diagnosis and treatment. Therefore, a reliable genotyping method is required for presymptomatic diagnosis. We describe the simultaneous detection of the C282Y, H63D and S65C mutations in the hemochromatosis gene by real-time PCR followed by melting curve analysis using fluorescence resonance energy transfer (FRET) probes. The acceptor fluorophore may be replaced by a quencher, increasing multiplex possibilities. Real-time PCR results were compared to the results of sequencing and conventional PCR followed by restriction digestion and detection by agarose gel electrophoresis (PCR-RFLP). Genotypes from 80 individuals obtained both by the conventional PCR-RFLP method and quenched-FRET real-time PCR were in full agreement. Sequencing also confirmed the results obtained by the new method, which proved to be an accurate, rapid and cost-effective diagnostic assay. Our findings demonstrate the usefulness of real-time PCR for the simultaneous detection of mutations in the HFE gene, which allows a reduction of a significant amount of time in sample processing compared to the PCR-RFLP method, eliminates the use of toxic reagents, reduces the risk of contamination in the laboratory, and enables full process automation.