臺灣博碩士論文加值系統

本研究旨在探討利用蛋白水解酵素AP2®處理大豆粕(soybean meal, SBM)應用於離乳仔豬飼糧之效果。於SBM中添加商業化蛋白酵素AP2® (活性為12,000U/ml)並與水以1:1.5比例混合均勻後置於37℃下培養72小時，於50℃下烘乾12小時(此產品簡稱為protease-treated soybean meal, PSBM)，再經粉碎處理。利用凝膠電泳(SDS-PAGE)分析PSBM蛋白肽，可發現經酵素處理後可去除SBM中抗原蛋白質β-conglycinin並降低glycinin的含量，pH由6.5降至4.78，乳酸含量從0 mmole/g提高至62 mmole/g。以in vitro法測定蛋白質消化率由77 ﹪提升到90 ﹪，in vitro總胺基酸消化率由70.9 ﹪提高到75.29 ﹪。經過酵素處理後可提高SBM可溶性蛋白質含量，其中90%水溶性蛋白質降至10KDa以下。
為進一步探討PSBM在離乳仔豬飼糧之應用性，採用28日齡之三品種(Duroc♂與Landrace x Yorkshire雜交母豬♀)雜交仔豬48頭進行試驗，所有仔豬依體重及性別平均分配至含SBM、PSBM與脫脂乳粉(Skim milk, SM)之飼糧中，各處理組四欄，每欄四頭，試驗期28天，分別於試驗第14天與第28天每組各屠宰4頭。結果顯示，SBM、PSBM與SM處理組間下痢指數評分有顯著性差異(P<0.05)分別為1.2、2.0與2.7。顯示PSBM具改善仔豬下痢之程度。於動物試驗開始後第二週與第四週進行木糖吸收試驗評估仔豬腸道吸收能力，顯示灌食後一小時PSBM與SM處理組血中木糖含量顯著高於SBM處理組，PSBM與SM處理組仔豬血液中之IgG抗體含量也顯著(P<0.05)較SBM處理組低。檢測消化道性狀中發現，PSBM可降低胃與小腸前段pH值，並降低腸道中coliform菌數，PSBM與SM處理組仔豬小腸中pepsin、trypsin與chymotrypsin活性較SBM處理組高。餵飼PSBM與SM仔豬有較高之腸道絨毛高度與較低之腺窩深度。綜合以上結果得知，SBM經蛋白水解酵素AP2處理後可降低大豆中之抗原蛋白含量，提高in vitro蛋白質與胺基酸消化率，有改善仔豬下痢情形，降低腸道中coliform菌數與大豆抗原對腸道之傷害，提高SBM於離乳仔豬飼糧之應用性。

Abstract
The purpose of this study was using protease-treated to degrade soybean antigenic-protein and to improve SBM application in weaned pigs. The protease AP2 (12000U/g) was added to SBM and incubated at 37℃ for 72 hours in a tank. After incubation, the SBM was dried at 50℃ for 12 hours and ground(protease-treated soybean meal, PSBM).
Using SDS polacrylamide gel electrophoresis analysis the soybean protein. After protease-treatment can breakdown β-conglycinin and decrease the amount of glycinin. The pH decrease from 6.5 to 4.7, lactic acid concentration from 0 mmole/g to 62 mmole/g. The protein digestion by in vitro can increased from 77% to 90% and in vitro total amino acid digestion also increased from 70.9% to 75.29%. The soluble protein of PSBM was higher than SBM, and the soluble protein molecular weight was below 10kDa.
Thirty-six crossbred [Landrace♂x (Yorkshire x Duroc) ♀] weaning 28-day-old were used in vivo experiment. According to body weight and sex the piglets were allotted into three dietary treatments that were diets containing SBM or PSBM or skim milk (SM). Each treatment contained four pens, each pen contained four piglets and experiment period was for 28 days. Piglets were slaughtered at 14 and 28 days.
The result showed that PSBM could decrease the diarrhoea severity. At 14 and 28 days using xylose absorption test to assess intestinal absorptive ability. PSBM and SM treatments had significant xylose absorptive ability (P<0.05) than SBM treatment. The IgG concentration of plasma in pigs fed PSBM and SM had lower than SBM treatment. Piglets fed PSBM can decrease stomach and intestinal 25% location pH on 28 test days, had lower coliform counts than SBM and SM treatments. Piglets fed SBM had less active of pepsine in stomach mucosa, and less active of trypsin and chymotrypsin in intestinal mucosa (p<0.05). In the intestinal mucosa histology, piglets fed PSBM and SM had greater height of villi (P<0.05) and less crypt depth (P<0.05) than SBM treatments.
It is concluded that SBM, treated with protease can decrease the amount of soy antigenic-protein. Protein and amino acid digestion was increased by in vitro and the molecular weight of soluble protein was below 10kDa. Improve the diarrhoea severity in piglet, decrease coliform counts and soy antigenic-protein damage in intestinal. Improve SBM application in the feed of weaned pigs.

壹、中文摘要 1
貳、前言 3
參、文獻檢討 4
一、大豆之特性 4
二、大豆之加工產品 4
(一) 大豆粕（soybean meal, SBM） 4
(二) 脫殼大豆粕 (dehulled soybean meal,DSBM) 4
(三) 濃縮大豆蛋白 (soy protein concentrate, SPC) 5
三、大豆之抗營養因子(anti-nutritional factors, ANFs) 6
(一) 胰蛋白酶抑制因子(trypsin inhibitor, TI) 6
(二) 寡糖(oligosaccharides) 8
(三) 非澱粉多醣類(non starch polysaccharides, NSP) 9
(四) 大豆抗原蛋白(soya antigenic- protein) 9
四、不同加工處理改善大豆之應用性 11
(一) 擠壓處理 11
(二) 加熱處理 11
(三) 溶劑處理 12
(四) 醱酵處理 13
(五) 酵素處理 14
肆、材料與方法 16
一、SBM之酵素處理 16
二、利用兩段式in vitro 模擬消化試驗測量SBM與PSBM 之利用率 16
(一) in vitro 模擬消化試驗後之蛋白質與胺基酸消化率 16
(1) 蛋白質消化率 16
(2) 胺基酸消化率 17
(二) in vitro 模擬消化試驗後水溶性蛋白質之含量 18
(三) in vitro 模擬消化試驗後可溶性蛋白質分子量分布 18
(四) SBM處理前後蛋白肽分佈 18
(五) 乳酸含量之測定 19
(六) 胰蛋白酶抑制因子(trypsin inhibitor)之測定 20
三、以動物試驗探討SBM與PSBM之應用效果 21
(一) 試驗動物與飼養管理 21
(二) 試驗處理與日糧 21
(三) 樣品採集與處理 23
(1) 屠宰與採樣 23
(2) 粗酵素液製備 23
(四) 分析項目與方法 23
(1) 下痢指數 23
(2) 木糖吸收率試驗 23
(3) 血清中IgG抗體濃度測定 25
(4) 胃蛋白酶(pepsin)之測定 27
(5) 胰蛋白酶(trypsin)測定方法 28
(6) 胰凝乳蛋白酶(chymotrypsin)之測定 29
(7) 腸道菌相之分布 29
(8) 腸道組織切片之觀察 30
(9) 掃描式電子顯微鏡組織觀察(Scanning electron microscopy ,SEM) 31
四、統計分析 33
伍、結果與討論 34
一、利用兩段式in vitro 模擬消化試驗探討SBM與PSBM 之利用率 34
（一）SBM與PSBM成分與蛋白肽分佈 34
（二）in vitro 模擬消化試驗後之蛋白質與胺基酸消化率 39
（三）in vitro 模擬消化試驗後水溶性蛋白質之含量與分佈 39
二、以離乳豬試驗探討SBM與PSBM之應用效果 43
（一）下痢程度 43
（二）木糖(D+-xylose)吸收能力 45
（三）血清中IgG抗體濃度 46
（四）盲腸之菌相 48
（五）消化道之性狀 50
(1) 消化道pH 50
(2) 消化道重量與長度 52
(3) 蛋白水解酵素活性 54
（六）腸道絨毛之型態 56
陸、結論 62
柒、參考文獻 63
捌、英文摘要 76

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