Study On Absorption Of Small Peptides In Broilers And Its Regulation

Six experiments were conducted to sdudy the mechanism of absorption of small peptide by broiler, and the effect of dietary nitrogen source, hormone and growth factor on transport of peptide in small intestine of broiler, the characteristic of protein feedstuff for releasing small molecular peptide in digestive tract.Exp. 1 was conducted to study the stability of some di-peptides when facing infusion solution, incubation temperature and time, acid, boiling. Gly-Gly, Gly-Leu and Met-Leu were treaed by resolving in PBS buffer with Ca²⁺ and Mg²⁺, heating at 39.5℃ for 30min, diluting with 0.6mol/L perchloric acid and boiling with 0.6mol/L perchlorate acid for f 5min. The results showed that three di-peptides were stability for these treatments and the availability retained was 97% above.Exp.2 was used to study the hydrolysis of Gly-Gly and Gly-L-Leu in small intestine of chick in vitro. Tewlve layers were divided into 4 groups, which include 4 replicates (birds). The birds were killed and their small intestine was divided into duodenum, upper jejunum, middle jejunum, lower jejunum and ileum. All segments of intestine were made into everted gut sacs. These sacs were suspended respectively in 4 incubation solutions, which were combined with various di-peptides and with or without peptidase inhibitor (Amastatin and Bestatin). The content of di-peptide in incubation solution was examined at series of sample time. The results showed that hydrolysis of di-peptides occured intensively. Gly-Gly was fully hydrolyzed in 5min. Gly-Leu was hydrolyzed mostly in 5 min, and entirely in 25min. Hydrolysis in the proximal of intestine was more intensively than the distal. Adding 10μmol/L Amastatin and Bestatin in the incubation solution could not inhibit hydrolysis of Gly-Gly, but inhibited of Gly-Leu. About 40% Gly-Leu was retained after 25min.Exp.3 was conducted to investigate the effect of dietary nitrogen source on intestinal absorption of broiler in vitro and content of small molecular peptides in serum. Broilers were divided into 3 groups and fed three kinds of isocaioric and iso-amino acid pure diet with different nitrogen source, amino acid, casein hydrolyte (oligopeptide), and casein from one to 21 day-aged, respectively. Nine birds per group were killed and their small intestine was cut into duodenum, jejunum and ileum. All segments of intestine were made into everted gut sacs. These sacs were incubated in various incubation solutions, which were blank solution, amino acid solution and Gly-Leu solution for 25min, respectively. The Cly-Leu in incubation solution, infusion solution and intestine tissue was examined. The results showed that dietary nitrogen source influenced intestinal absorption for Gly-Leu in vitro significantly. The birds fed with casein absorbed the most Gly-Leu, casein hydrolate the least. Significant difference was found among various intestinal segments. Absorption in duodenum was fewer than jejunum and ileum. The content of small molecular peptide was affected by dietary nitrogen source. Birds fed amino acid had more small molecular peptide than others.Exp.4 A method of semi-quantity RT-PCR was estabilished to evaluate gene expression of peptide transporter in small intestine of chick. The primers of β-actin and PepT1 were designed according to their gene sequence reported. The cycles and Mg²⁺ content was optimized by some experiments. Theresults showed that gene expression of PepTl varied with intestinal segments, decreased from the proximal to the distal, i.e. the level of the expression of PepTl in ileum was higher than in duodenum and jejunum significantly, no difference was found between duodenum and jejunum.Exp.5 was carried out to investigate the regulation role of hormone and growth factor on Peptl gene expression in broiler. Thirty broilers aged 25 days were divided into 6 groups. Five trial groups were stimulated by epidermal growth factor, thyroid hormone, glucagons, thiamazolum for 5d, respectively. The control was injected with distilled water. The abundance of Peptl mRNA in small intestine was examined by RT-PCR. The results showed that thyroid hormone and glucagon upregulated gene expression of PepTl on transcription level significantly. Thiamazolum and epidermal growth factor tended to downregulate it.Exp.6 was conducted to investigate the characteristic of releasing small molecular peptides for various protein feedstuffs in digestive tract of broiler. Thirty-five broilers aged 21 days were divided into 7 groups. Mixture of 70% feedstuff (soybean meal, fishmeal, rapeseed meal, peanut meal, corn gluten, blood meal and cottonseed meal) and 30% starch was fed by gavage, respectively. The results showed that percentage of small molecular peptides in soluable protein from digesta varied with digestive tract segments significantly. Proventriculus had the lowest percentage, and duodenum was the highest. No significant difference was found among upper and lower jejunum and upper ileum. Lower ileum had the lowest percentage among intestinal segments. Signifancant difference was found between different feedstuffs. The descendent sequence of percentage was cottonseed meal, peanut meal, blood meal, corn gluten, soybean meal, fishmeal, rapeseed meal.