CN110551790B - 一种乳源活性肽的制备方法及应用 - Google Patents
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Abstract
本发明公开了一种乳源活性肽的制备方法及应用,以大豆蛋白和酪蛋白为原料,通过酶解和微生物发酵相结合的方法获得乳源活性肽产品。大豆蛋白按一定质量比例添加蒸馏水,浸泡预处理,调整体系pH,添加酸性蛋白酶反应。酪蛋白按一定比例添加蒸馏水,调整体系pH,添加胰蛋白酶反应。然后大豆蛋白酶解液与酪蛋白酶解液混合,按照5%比例接种发酵,控制发酵pH在7.0左右,发酵温度为40~50℃,发酵时间为100~120小时,发酵完成后,低温干燥制成动物用乳源活性肽。乳源活性肽对动物消化道功能成熟与发育有重要的促进作用,本发明的动物用乳源活性肽,可显著提高畜禽采食量和生长性能,增强机体免疫能力和抗氧化能力。
Description
技术领域
本发明属于动物饲料领域,具体涉及一种乳源活性肽的制备方法及应用。
背景技术
动物肠道不仅是营养物质消化吸收的场所,也是重要的免疫器官,维护正常的肠道机能对动物生长发育至关重要。动物在营养素缺乏、感染、应激及病理等情况下,均可导致肠道结构和功能损伤、细菌和内毒素移位以及肠道黏膜屏障功能障碍等,严重影响动物健康和生产性能,给养殖生产带来重大的经济损失。
目前,大豆蛋白或酪蛋白均只采用发酵或酶解的单一方法制备活性肽,其中酶解法生产的小肽苦味较重,适口性差;微生物发酵法生产的小肽,得率不高,且平均分子质量较大,难以完全发挥潜在的生物学功能。通过酶解法和微生物发酵结合的方法生产活性肽,不仅可获得适口性好,产率高的小肽,而且还可获得更丰富的微生物代谢微量营养养分。
发明内容
本发明的目的是克服现有技术的不足,提供了一种乳源活性肽的制备方法和应用,本发明的方案具体如下:
乳源活性肽的生产方法的制备方法如下:
取大豆蛋白,按大豆蛋白:水质量比=1:2-5的比例添加蒸馏水,85~95℃浸泡处理3~4小时,冷却至40~50℃,用2mol/L HCl溶液调整体系pH至3.0-5.0,按每克原料的酶浓度为5000~6000U添加酸性蛋白酶,维持温度在40~50℃,反应10-15小时,制成大豆蛋白酶解液;
取酪蛋白,按酪蛋白:水质量比=1:5-10的比例添加蒸馏水,用2mol/L NaOH溶液调整体系pH至8.0-9.0,按每克大豆蛋白原料的酶浓度为1000~2000U添加胰蛋白酶,维持温度在40~50℃左右,反应2-5小时,制成酪蛋白酶解液;
然后大豆蛋白酶解液与酪蛋白酶解液以1:2-5体积混合备用;
按照麸皮4质量份,豆粕粉1质量份,硫酸铵4质量份,磷酸氢二钾0.3质量份,氯化钙0.5质量份,蒸馏水125质量份的比例配制发酵培养基,与混合酶解液等体积混合后115℃灭菌15min备用;
瑞士乳杆菌和黑曲霉分别使用MRS和查氏液体培养基进行扩大培养,37℃条件下静置培养24小时,按照5%-10%体积比例接种发酵,即菌液:(培养基+酶解物)=5-10:100;控制发酵pH在7.0,发酵温度为40~50℃,发酵时间为100~120小时;发酵完成后,低温干燥制成动物用乳源活性肽。
作为本发明的优选方案,所述大豆蛋白为粗蛋白质量百分比含量≥46%的低温豆粕制成的大豆蛋白粉;酪蛋白为牛乳源酪蛋白,粗蛋白质量百分含量≥90%,白色至淡黄色粉末或颗粒。
作为本发明的优选方案,混合酶解液发酵完成后,低温干燥的条件为真空度0.06-0.09MPa,β-环状糊精为干燥助剂,混合酶解液浓缩时间为40-60min,浓缩温度为40-55℃,干燥后的成品中肽分子量280~1000道尔顿的质量百分比含量为40-60%。
本发明公开了一种采用所述方法制备得到的乳源活性肽。所述的乳源活性肽可以在制备动物饲料或制备动物饲料添加剂中进行应用。作为本发明的优选方案,对于乳仔猪、肉禽,每吨饲料添加300~500克所述乳源活性肽,对于中大猪,每吨饲料添加800~1000克所述乳源活性肽。
本发明之乳源生物活性肽的生产制备,由于采用了酶解法和微生物发酵相结合的方法,不仅可生产适口性好、含量高的生物活性肽,还可获得儿茶酚胺类、GABA、5-羟色胺、褪黑激素及乙酰胆碱等神经信号物质,丰富了产品的生物活性成分,有效解决现有饲料工业中乳源活性肽来源有限,产业化程度低的问题。
本发明的乳源生物活性肽应用于动物生产,可使肥育猪平均日增重提高10%-15%,料重比降低8%-13%,瘦肉率提高5%-10%,背最长肌面积提高15%-30%,背膘厚降低10%-15%,脂肪率降低20%-30%;仔猪平均日增重提高10%-30%,料重比降低5%-10%,腹泻率下降30%-45%,空肠绒毛高度提高40%-50%,隐窝深度下降15%-20%,空肠sIgA含量提高10%-25%,十二指肠淀粉酶、脂肪酶和胰蛋白酶分别提高40%-55%,40%-50%和35%-40%;脾脏指数提高30%-40%,下颌淋巴指数提高35%-45%;血清IgG含量提高30%-35%,总抗氧化能力提高30%-35%,促炎性细胞因子TNF-α、IL-6和IL-22水平分别降低40%-50%、25%-35%和35%-45%。
大豆蛋白和酪蛋白酶解能产生丰富的氨基酸和生物活性肽,菌种发酵酶解液能产生儿茶酚胺类、GABA、5-羟色胺、褪黑激素及乙酰胆碱等神经信号物质。用本法制备的乳源活性肽产品具有广泛的生物活性,可促进消化道发育,增强机体抗氧化能力、免疫力及抗菌力,可增强矿物质吸收,缓解应激,调节动物行为,促进蛋白质合成和脂肪分解。试验证明,乳源活性肽能促进新生仔猪肠道生长,肠黏膜上皮细胞增殖和提高小肠黏膜麦芽糖酶和碱性磷酸酶活性,显著提高胃重、小肠总重,空肠绒毛高度与隐窝深度比值;可保护肠道屏障功能,降低血中内毒素含量、二胺氧化酶活性及肠黏膜通透性指标,减轻肠源性感染风险;可提高IGF-I及其受体基因的mRNA水平。乳源活性肽对肠道发育及维持肠道健康具有重要意义。
本发明的乳源生物活性肽,可显著提高畜禽采食量和生长性能,增强机体免疫能力和抗氧化能力。
具体实施方式
本发明结合以下实例作进一步的说明。
实施例1
以大豆蛋白为原料,按大豆蛋白:水=1:4的比例添加蒸馏水,85℃预处理3小时。冷却至45℃,用2mol/L HCl溶液调整体系PH至3.0左右,按每克原料的酶浓度为5000U添加酸性蛋白酶,维持温度在45℃左右,反应10小时。以酪蛋白为原料,按酪蛋白:水=1:10的比例添加蒸馏水,用2mol/L NaOH溶液调整体系PH至8.0左右,按每克原料的酶浓度为1500U添加胰蛋白酶,维持温度在45℃左右,反应2小时。1体积大豆蛋白酶解液与2体积酪蛋白酶解液混合备用。按照麸皮4g,豆粕粉1g,硫酸铵4g,磷酸氢二钾0.3g,氯化钙0.5g,蒸馏水125ml的比例配制发酵培养基,与酶解液等体积混合后115℃灭菌15min备用。瑞士乳杆菌和黑曲霉分别使用MRS和查氏液体培养基进行扩大培养,37℃条件下静置培养24小时,按照5%比例接种发酵,即菌液:(培养基+酶解物)=5:100。控制发酵PH在7.0左右,发酵温度为45℃,发酵时间为100小时,发酵完成发酵液泵入干燥仪内进行低温干燥,低温干燥的条件为β-环状糊精添加量4%,真空度0.06MPa,浓缩温度为45℃,浓缩时间为40min,干燥后制成的动物用乳源活性肽,水分含量为11%,分子量1000~280道尔顿的肽含量为49.3%。
实施例2
以大豆蛋白为原料,按大豆蛋白:水=1:2的比例添加蒸馏水,90℃预处理4小时。冷却至45℃,用2mol/L HCl溶液调整体系PH至4.0左右,按每克原料的酶浓度为5500U添加酸性蛋白酶,维持温度在45℃左右,反应12小时。以酪蛋白为原料,按酪蛋白:水=1:5的比例添加蒸馏水,用2mol/L NaOH溶液调整体系PH至9.0左右,按每克原料的酶浓度为2000U添加胰蛋白酶,维持温度在50℃左右,反应5小时。1体积大豆蛋白酶解液与5体积酪蛋白酶解液混合备用。按照麸皮4g,豆粕粉1g,硫酸铵4g,磷酸氢二钾0.3g,氯化钙0.5g,蒸馏水125ml的比例配制发酵培养基,与酶解液等体积混合后115℃灭菌15min备用。瑞士乳杆菌和黑曲霉分别使用MRS和查氏液体培养基进行扩大培养,37℃条件下静置培养24小时,按照10%比例接种发酵,即菌液:(培养基+酶解物)=5:100。控制发酵PH在7.0左右,发酵温度为48℃,发酵时间为120小时,发酵完成发酵液泵入干燥仪内进行低温干燥,低温干燥的条件为β-环状糊精添加量4%,真空度0.06MPa,浓缩温度为45℃,浓缩时间为40min,干燥后制成的动物用乳源活性肽,水分含量为11%,分子量1000~280道尔顿的肽含量为55.4%。
实施例3
25日龄仔猪断奶开始,对照组饲喂对照日粮(配方见表1)。试验组全价饲料在每吨对照组日粮基础上添加乳源活性肽400克,饲喂上述饲料至46日龄,测定仔猪生长性能。
表1试验饲粮组成和营养指标
试验结果表明,与对照组相比,试验组仔猪平均日增重提高31.67%(P<0.05),料重比降低8.52%(P<0.05),腹泻率下降13.67%(P<0.05)。十二指肠、空肠、回肠绒高度较对照组分别提高了6.25%(P<0.05),7.14%(P<0.05)和9.80%(P<0.05),隐窝深度较对照组降低了12.96%(P<0.05),8.37%(P<0.05)和7.54%(P>0.05)。十二指肠食糜淀粉酶、脂肪酶、胰蛋白酶分别较对照组提高了49.35%(P<0.05),40.00%(P<0.05)和38.40%(P<0.05)。脾脏指数提高了32.00%(P<0.05),下颌淋巴指数提高了37.50%(P<0.05)。血清IgG较对照组提高了89.83%(P<0.01),空肠sIgA含量提高13.57%(P<0.05)。总抗氧化能力较对照组提高32.71%(P<0.05),血清内毒素和D-乳酸含量分别降低12.57%(P<0.05)和8.97%(P<0.05),提高IGF-I及其受体基因的mRNA水平。
实施例4
21日龄健康AA肉鸡,对照组饲喂对照日粮(配方见表2),试验组全价饲料在每吨对照组日粮基础上添加乳源活性肽营养调节剂300克,饲喂上述饲料至35日龄,测定肉鸡生长性能。
表2试验饲粮组成和营养指标
试验结果表明,与对照组相比,试验组肉鸡日增重和采食量分别提高了6.77%(P<0.05)和7.02%(P<0.05),十二指肠绒毛长度提高14.33%(P<0.05),十二指肠绒毛长度/隐窝深度提高22.15%(P<0.05);回肠绒毛长度/隐窝深度提高10.32%(P<0.05),十二指肠胰蛋白酶、淀粉酶、脂肪酶和总蛋白水解酶活性分别提高了34.52%(P<0.05)、78.22%(P<0.05)、22.77%(P<0.05)和25.54%(P<0.05)。
Claims (5)
1.一种乳源活性肽的制备方法,其特征在于它的制备方法如下:
取大豆蛋白,按大豆蛋白:水质量比=1:2-5的比例添加蒸馏水,85~95℃浸泡处理3~4小时,冷却至40~50℃,用2mol/L HCl溶液调整体系pH至3.0-5.0,按每克原料的酶浓度为5000~6000U添加酸性蛋白酶,维持温度在40~50℃,反应10-15小时,制成大豆蛋白酶解液;
取酪蛋白,按酪蛋白:水质量比=1:5-10的比例添加蒸馏水,用2mol/L NaOH溶液调整体系pH至8.0-9.0,按每克原料的酶浓度为1000~2000U添加胰蛋白酶,维持温度在40~50℃,反应2-5小时,制成酪蛋白酶解液;
然后大豆蛋白酶解液与酪蛋白酶解液以1:2-5体积混合备用;
按照麸皮4质量份,豆粕粉1质量份,硫酸铵4质量份,磷酸氢二钾0.3质量份,氯化钙0.5质量份,蒸馏水125质量份的比例配制发酵培养基,与混合酶解液等体积混合后115℃灭菌15min备用;
瑞士乳杆菌和黑曲霉分别使用MRS和查氏液体培养基进行扩大培养,37℃条件下静置培养24小时,按照5%-10%体积比例接种发酵,即菌液:(培养基+酶解物)=5-10:100;控制发酵pH在7.0,发酵温度为40~50℃,发酵时间为100~120小时;发酵完成后,低温干燥制成动物用乳源活性肽;
对于乳仔猪、肉禽,每吨饲料添加300~500克所述乳源活性肽,对于中大猪,每吨饲料添加800~1000克所述乳源活性肽。
2.根据权利要求1所述的乳源活性肽制备方法,其特征在于,所述大豆蛋白为粗蛋白质量百分比含量≥46%的低温豆粕制成的大豆蛋白粉;酪蛋白为牛乳源酪蛋白,粗蛋白质量百分含量≥90%,白色至淡黄色粉末或颗粒。
3.根据权利要求1所述的乳源活性肽制备方法,其特征在于,混合酶解液发酵完成后,低温干燥的条件为真空度0.06-0.09MPa,混合酶解液浓缩时间为40-60min,浓缩温度为40-55℃,β-环状糊精为干燥助剂,干燥后的成品中肽分子量280~1000道尔顿的质量百分比含量为40-60%。
4.权利要求1-3任一项所述方法制备得到的乳源活性肽。
5.权利要求4所述的乳源活性肽在制备动物饲料或制备动物饲料添加剂中的应用。
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