CN111088309A - 一种水解植物蛋白制备抗氧化肽的方法 - Google Patents
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Abstract
本发明公开了一种利用蛋白酶水解植物蛋白制备抗氧化肽的方法,主要以大豆分离蛋白粉作原料,以木瓜蛋白酶和菠萝蛋白酶作为催化剂,在10%(w/v)的蛋白浓度、原料质量5%(w/w)的酶浓度、适宜的温度及pH条件下,进行一锅水解反应。本发明使用的蛋白酶组合特异性高,肽产物的稳定性高,反应的可重复性高。水解得到的肽分子量适宜,肽收率高,且具有一定的抗氧化活性。而且,本发明得到的抗氧化肽氨基酸种类丰富,营养成分含量高,在食品及化妆品领域有着重要的应用。
Description
技术领域
本发明生物与医药技术领域,尤其涉及一种利用酶水解植物蛋白制备抗氧化肽的方法。
背景技术
人工合成抗氧化剂价格低、产量高等优点而广泛地应用于食品、护肤品、化妆品工业。但是,随着经济发展,人们对健康越来越重视,致使人们更加青睐安全、纯天然、无毒副作用的天然抗氧化剂,如茶多酚、抗坏血酸及蛋白肽等。食源性抗氧化肽的种类有很多,按照来源分类可分为植物源、动物源、乳源和蛋源等抗氧化肽。植物源主要有玉米、大豆、小麦谷、米糠、油菜籽、花生、豆鹰嘴、大米、米渣、花椒籽、核桃、黑豆、麦胚等。
大豆分离蛋白是以低温脱溶大豆粕为原料生产的一种全价蛋白。大豆分离蛋白中蛋白质含量在90%以上,氨基酸种类有近20种,能为人体提供丰富的营养。但由于大豆蛋白是大分子物质,其溶解度低,并且一些潜在的功能因子隐藏在分子内部,导致大豆分离蛋白功能性质较弱,严重制约其在应用。将大豆的蛋白质进行酶解制备具有抗氧化功能的多肽是常用的提高大豆分离蛋白功能特性的方法。
蛋白酶是一种重要的工业酶,它能选择性的降解蛋白类底物,在食品工业已经有广泛的应用;蛋白酶的来源有动物、植物、微生物。目前,人们已经利用蛋白酶将核桃蛋白、酪蛋白、大豆分离蛋白、黄粉虫蛋白、水产动物蛋白等水解之后制备得到了抗氧化肽。
采用酶水解蛋白制备抗氧化肽是主流的方法,但由于许多蛋白酶有特异性的酶切位点及蛋白质氨基酸序列的独特性,因此选用不同的蛋白及酶组合对于生成具有独特功能性的肽化合物具有重要意义。就现有的酶解方法而言,有些使用了无特异性或广泛特异性的蛋白酶,这样的生产重复性低并且会产生游离氨基酸,或是肽链会被水解的过小导致功能性降低,最后产物的收率也较低,总体而言还缺乏稳定、高效制备抗氧化肽的方法。
发明内容
为了克服现有技术的缺陷,本发明提供了一种酶解大豆蛋白制备抗氧化肽的方法,该方法收率高、产品稳定、肽具有抗氧化活性,同时还保留了大豆蛋白的营养成分,因此该肽可以被应用到食品、护肤品、化妆品中充当营养成分及功效成分。
本发明的具体技术方案如下:
一种水解大豆分离蛋白制备抗氧化肽的方法,其特征在于,包括如下步骤:
S1:用蒸馏水将豆粕粉配制成料液比为1:10豆粕溶液,搅拌均匀,55℃恒温水浴,调节pH值为8.5,碱提45min,然后浆液在20℃,6000r/min条件下离心20min,收集上清液;
S2:将上清液pH值调至4.5,并酸沉2h收集沉淀物,将沉淀物pH值调至7.0,冷冻干燥得大豆分离蛋白粉;
S3:将大豆分离蛋白粉溶解到去离子水中,搅拌均匀,其中大豆分离蛋白与去离子水的质量体积比为1:10;
S4:同时加入木瓜蛋白酶和菠萝蛋白酶,搅拌均匀得到反应液,其中木瓜蛋白酶的重量为大豆分离蛋白重量的5%,菠萝蛋白酶的重量为大豆分离蛋白重量的5%;
S5:调节反应液的pH至7.5,加热反应液至55℃,放入搅拌器中,在40rpm的条件下反应,反应时间为4h,反应结束后温度升高至90~95℃,灭活15min;
S6:过滤或离心去掉不可容物质,即得到所述的抗氧化肽。
进一步地,步骤S4中的离心方法为使用冷冻离心机6000rpm离心10min,过滤方法为使用0.45μm的滤膜进行抽滤。
本发明还涉及一种采用上述方法制备得到的抗氧化肽。
本发明还涉及一种采用上述方法制备得到的抗氧化肽在食品、护肤品、化妆品领域的应用。
本发明还涉及一种护肤品,其特征在于:含有采用上述方法制备得到的抗氧化肽。
本发明还涉及一种化妆品,其特征在于:含有采用上述方法制备得到的抗氧化肽。
本发明的方法有以下的优点:
1.本方法采用酶解蛋白制备抗氧化肽,反应过程温和,不会对于蛋白的结构造成影响,反应能耗低,反应过程中没有添加任何的化学成分,对于环境无污染同时对于人体无害。
2.本方法制备的产品均一、无刺激气味及颜色、具有功能性,对于人体而言更易接受,抗氧化肽不仅具有抗氧化活性,还保留有了原本大豆蛋白的一些优良功能性,具有营养的功效。
3.本方法工艺简单,在纯水体系里进行反应,使用的设备少、条件温和、成本低,适合在工业化大规模的条件下生产,原料利用率高,排放少,使用的是天然化合物,符合国家绿色、环保、低碳、低能耗、可持续的理念。
4.本方法的酶组合中的蛋白酶均加入到一个反应体系里,同时一锅的进行反应,提高了时空效率。
5.本方法制备的抗氧化肽大小较均一,批次间的重复性好;肽的得率高,反应液除去沉淀后均一、气味及颜色无刺激,具有一定的抗氧化活性。
6.本发明公开的抗氧化肽不仅具有抗氧化活性,还具有营养功效,在产品中可以当营养成分及功效成分,在食品、化妆品领域有着重要的应用。
具体实施方式
实施例1
豆粕粉碎过100目筛得豆粕粉,用蒸馏水将豆粕粉配制成料液比为1:10豆粕溶液,搅拌均匀后浆液转移到55℃恒温水浴锅,调节浆液pH值为8.5,碱提45min,然后浆液在20℃,6000r/min条件下离心20min,收集上清液;将上清液pH值调至4.5,并酸沉2h收集沉淀物,将沉淀物pH值调至7.0,冷冻干燥得大豆分离蛋白粉。
将100g大豆分离蛋白粉加入到1L的去离子水中,在室温的条件下手动搅拌使之混匀,制成相应的蛋白浆液。在浆液中分别加入5g木瓜蛋白酶和菠萝蛋白酶,手动混匀,反应pH用固体氢氧化钠调节至7.5,将以上的浆液放入到磁力搅拌器中,磁力搅拌器温度预先升到55℃后维持不变,在40rpm的条件下开始反应;反应4h后,反应温度升高至95℃并维持15min,灭活之后的反应液经过6000rpm离心6min后得到抗氧化肽。
对比实施例1
将100g燕麦蛋白粉加入到1L的去离子水中,在室温的条件下手动搅拌使之混匀,制成相应的蛋白浆液。在浆液中分别加入5g木瓜蛋白酶及菠萝蛋白酶,手动混匀,反应pH用固体氢氧化钠调节至7.5,将以上的浆液放入到磁力搅拌器中,磁力搅拌器温度预先升到55℃后维持不变,在40rpm的条件下开始反应;反应4h后,反应温度升高至95℃并维持15min,灭活之后的反应液经过6000rpm离心6min后得到抗氧化肽。
对比实施例2
将实施例1制备的大豆分离蛋白粉加入到1L的去离子水中,在室温的条件下手动搅拌使之混匀,制成相应的蛋白浆液。在浆液中加入5g中性蛋白酶,手动混匀,用固体氢氧化钠调节pH至7,将以上的浆液放入到磁力搅拌器中,磁力搅拌器温度预先升到55℃后维持不变(中性蛋白酶的最适温度为55℃,pH为7,它是非专一性的蛋白酶),在40rpm的条件下开始反应;反应4h后,反应温度升高至95℃并维持15min,灭活之后的反应液经过6000rpm离心6min后得到抗氧化肽。
对比实施例3
将实施例1制备的大豆分离蛋白粉加入到1L的去离子水中,在室温的条件下手动搅拌使之混匀,制成相应的蛋白浆液。在浆液中分别加入5g胃蛋白酶,手动混匀,反应pH用盐酸调节至1.5,将以上的浆液放入到磁力搅拌器中,磁力搅拌器温度预先升到40℃后维持不变(胃蛋白酶最适温度为37℃左右,最适pH范围为1~2),在40rpm的条件下开始反应;反应4h后,反应温度升高至90℃并维持15min,灭活之后的反应液经过滤纸过滤,后经抽滤过0.45微米的滤膜,得到抗氧化肽。
对比实施例4
将实施例1制备的大豆分离蛋白粉加入到1L的去离子水中,在室温的条件下手动搅拌使之混匀,制成相应的蛋白浆液。在浆液中分别加入5g碱性蛋白酶及木瓜蛋白酶,手动混匀,反应pH用固体氢氧化钠调节至8,将以上的浆液放入到磁力搅拌器中,磁力搅拌器温度预先升到55℃后维持不变,在40rpm的条件下开始反应;反应4h后,反应温度升高至95℃并维持15min,灭活之后的反应液经过6000rpm离心6min后得到抗氧化肽。
对比实施例5
将实施例1制备的大豆分离蛋白粉加入到1L的去离子水中,在室温的条件下手动搅拌使之混匀,制成相应的蛋白浆液。在浆液中分别加入5g碱性蛋白酶及胰蛋白酶,手动混匀,反应pH用固体氢氧化钠调节至8.5,将以上的浆液放入到磁力搅拌器中,磁力搅拌器温度预先升到45℃后维持不变,在40rpm的条件下开始反应;反应4h后,反应温度升高至95℃并维持15min,灭活之后的反应液经过6000rpm离心6min后得到抗氧化肽。
对比实施例6
将实施例1制备的大豆分离蛋白粉加入到1L的去离子水中,在室温的条件下手动搅拌使之混匀,制成相应的蛋白浆液。在浆液中分别加入5g胰蛋白酶及菠萝蛋白酶,手动混匀,反应pH用固体氢氧化钠调节至8,将以上的浆液放入到磁力搅拌器中,磁力搅拌器温度预先升到55℃,后维持不变,在40rpm的条件下开始反应;反应4h后,反应温度升高至95℃并维持15min,灭活之后的反应液经过6000rpm离心6min后得到抗氧化肽。
对比实施例7
将实施例1制备的大豆分离蛋白粉加入到1L的去离子水中,在室温的条件下手动搅拌使之混匀,制成相应的蛋白浆液。在浆液中分别加入5g碱性蛋白酶、中性蛋白酶及风味蛋白酶,手动混匀,反应pH用固体氢氧化钠调节至8,将以上的浆液放入到磁力搅拌器中,磁力搅拌器温度预先升到50℃后维持不变,在40rpm的条件下开始反应;反应4h后,反应温度升高至95℃并维持15min,灭活之后的反应液经过6000rpm离心6min后得到抗氧化肽。
实施例8:水解度检测
水解度采用pH-stat法测定,该方法只适用于pH大于6的反应(对比实施例3不适用)。记录反应前的pH,反应之后pH会降低,滴加1M的氢氧化钠至pH为初始值,记录消耗的氢氧化钠体积,消耗的氢氧化钠量与释放出来的肽键一一对应,根据公式F1计算水解度,其中B为氢氧化钠消耗量,N为氢氧化钠浓度,M为蛋白质质量,其中燕麦蛋白粉的含量经过测定为40%。
表1:α及htot值
大豆分离蛋白粉及燕麦蛋白在不同酶组合条件下的水解度结果如下表2所示。
表2:大豆分离蛋白粉及燕麦蛋白的水解度
实施例 | 水解度(DH%) |
实施例1 | 16.30 |
对比实施例1 | 12.80 |
对比实施例2 | 13.23 |
对比实施例4 | 15.97 |
对比实施例5 | 18.87 |
对比实施例6 | 21.90 |
对比实施例7 | 26.30 |
从结果上看,不同酶组合对于大豆分离蛋白粉的水解能力不同,并且根据水解度判断,本发明方法制备的抗氧化肽的平均长度在3-10个氨基酸之间,分子量在300~1200之间,这个范围在抗氧化肽范围之内。
实施例9:水解肽分子量分布检测
检测在不同酶组合条件下的水解抗氧化肽分子量分布。采用凝胶色谱法测定,色谱柱为TSKgeI G2000SWXL,标准品为谷胱甘肽(还原型)、谷胱甘肽(氧化型)以及维生素B12,流动相乙腈/水/三氟乙酸体积比为10:90:0.1,流速为0.5mL/min,在220nm波长下检测肽分布。
表3:大豆分离蛋白粉及燕麦蛋白的分子量分布
实施例 | <307 | 307-612 | 612-1355 | >1355 |
实施例1 | 0.20 | 45.80 | 53.89 | 1.63 |
对比实施例1 | 0.90 | 13.50 | 54.89 | 33.63 |
对比实施例2 | 1.20 | 18.79 | 53.69 | 28.60 |
对比实施例3 | 0.40 | 12.60 | 63.82 | 25.30 |
对比实施例4 | 5.60 | 51.00 | 37.56 | 8.30 |
对比实施例5 | 15.78 | 55.63 | 23.29 | 5.80 |
对比实施例6 | 19.67 | 59.98 | 18.89 | 1.37 |
对比实施例7 | 16.56 | 73.56 | 9.80 | 0.90 |
从结果上看,本发明方法制备的酶水解物的分子量主要分布在307~1355,与根据水解度所判断的肽链长短结果基本一致,与燕麦蛋白的酶水解物分子量相比,肽分子量分布更为集中。
实施例10:水解肽感官评价
水解肽感官评价:用肉眼及鼻判断实施例和对比实施例所制备的抗氧化肽的色泽与气味。
表4:大豆分离蛋白粉及燕麦蛋白水解液感官评价
实施例 | 色泽 | 气味 | 均一与沉淀情况 |
实施例1 | 黄色 | 豆奶香 | 均一无沉淀 |
对比实施例1 | 浅白色 | 燕麦蛋白粉气味 | 均一无沉淀 |
对比实施例2 | 浅橙黄色 | 豆奶香 | 有沉淀 |
对比实施例3 | 黄色 | 豆奶香 | 有沉淀 |
对比实施例4 | 橙黄色 | 豆奶香带有酶气味 | 均一无沉淀 |
对比实施例5 | 橙黄色 | 酶气味明显 | 均一无沉淀 |
对比实施例6 | 黄色 | 豆奶香 | 均一无沉淀 |
对比实施例7 | 红色 | 豆奶香带有酶气味 | 均一无沉淀 |
大豆分离蛋白粉具有浅黄颜色,燕麦蛋白粉呈白色,因此如果酶颜色不深,水解液颜色就接近蛋白本身颜色。碱性蛋白酶及中性蛋白酶都是红色酶,它们会将自己的颜色带入到水解液中。碱性蛋白酶及胰蛋白酶都有明显的气味,这样裂解液除了带有蛋白本身特别的气味外还会带有酶气味,特别是胰蛋白酶气味会掩盖蛋白气味。反应结束后各水解物都会被观察到有沉淀,这是因为其中都有酶存在,此外有一些水解度较低的水解物中含有一些未水解蛋白质,它们的尺寸较小,过滤之后放置一段时间仍会观察到水解液中有沉淀。
实施例10:水解肽的抗氧化活性
分别测定了实施例和的对比实施例所制备的抗氧化肽对于ABTS自由基、羟自由基及超氧自由基的清除率,结果以清除百分比表示,如下表5所示。
表5:大豆分离蛋白粉及燕麦蛋白水解液抗氧化活性
酶组合 | ABTS<sup>+</sup> | OH- | O<sub>2</sub><sup>-</sup> |
实施例1 | 91.01 | 63.32 | 98.70 |
对比实施例1 | 61.10 | 40.30 | 75.69 |
对比实施例2 | 40.59 | 16.36 | 40.33 |
对比实施例3 | 39.63 | 17.89 | 33.37 |
对比实施例4 | 24.60 | 34.70 | 66.67 |
对比实施例5 | 73.00 | 52.20 | 86.00 |
对比实施例6 | 77.00 | 59.42 | 84.00 |
对比实施例7 | 76.30 | 35.20 | 84.00 |
从上述结果上看,蛋白水解肽均有一定的自由基清除能力,其中对于羟自由基的清除能力稍差,大豆分离蛋白粉水解物的抗氧化能力明显强于燕麦蛋白水解物的抗氧化能力。而且,抗氧化活性与水解度有一定关系,当水解度过大时水解物的抗氧化活性反而会降低,本发明利用木瓜蛋白酶和菠萝蛋白酶制备的水解大豆分离蛋白粉得到的肽具有最高的抗氧化活性,对于ABTS+、OH-、O2 -自由基的清除率分别为91.0、63.32及98.7%。
上述实施例为本发明较佳的实施方式,但本发明的实施方式并不受上述实施例的限制,其他的任何未背离本发明的精神实质与原理下所作的改变、修饰、替代、组合、简化,均应为等效的置换方式,都包含在本发明的保护范围之内。
Claims (7)
1.一种水解大豆分离蛋白制备抗氧化肽的方法,其特征在于,包括如下步骤:
S1:用蒸馏水将豆粕粉配制料液比为1:10豆粕溶液,搅拌均匀,50~60℃恒温水浴,调节pH值为8.5~9,碱提45~60min,然后离心浆液,收集上清液;
S2:将上清液pH值调至4.0~4.5,并酸沉1~3h收集沉淀物,将沉淀物pH值调至7.0,冷冻干燥得大豆分离蛋白粉;
S3:将大豆分离蛋白粉溶解到去离子水中,搅拌均匀,其中大豆分离蛋白与去离子水的质量体积比为1:10;
S4:同时加入木瓜蛋白酶和菠萝蛋白酶,搅拌均匀得到反应液,其中木瓜蛋白酶的重量为大豆分离蛋白重量的5%,菠萝蛋白酶的重量为大豆分离蛋白重量的5%;
S5:调节反应液的pH至7.5,并加热至55℃,放入搅拌器中,在40rpm的条件下反应,反应时间为4h,反应结束后温度升高至90~95℃,灭活15min;
S6:过滤或离心去掉不可容物质,即得到所述的抗氧化肽。
2.根据权利要求1所述的方法,其特征在于:步骤S4中的离心方法为使用冷冻离心机6000rpm离心10min,过滤方法为使用0.45μm的滤膜进行抽滤。
3.一种采用根据权利要求1或2所述的方法得到的抗氧化肽。
4.根据权利要求1或2所述的方法得到的抗氧化肽在食品、护肤品、化妆品领域的应用。
5.一种食品,其特征在于:含有采用根据权利要求1或2所述的方法得到的抗氧化肽。
6.一种护肤品,其特征在于:含有采用根据权利要求1或2所述的方法得到的抗氧化肽。
7.一种化妆品,其特征在于:含有采用根据权利要求1或2所述的方法得到的抗氧化肽。
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Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2002069733A1 (en) * | 2001-03-05 | 2002-09-12 | Council Of Scientific And Industrial Research | Process for preparation of protein-hydrolysate from soy flour |
CN103719534A (zh) * | 2013-12-23 | 2014-04-16 | 江南大学 | 一种对大豆分离蛋白进行酶水解的方法 |
CN104286370A (zh) * | 2014-08-20 | 2015-01-21 | 华南理工大学 | 一种酶辅助亚临界水法制备功能性大豆蛋白的方法 |
US20170143001A1 (en) * | 2015-04-30 | 2017-05-25 | China National Research Institute Of Food And Fermentation Industries | Soybean oligopeptide with low allergenicity and little bitterness and preparation method and application thereof |
-
2019
- 2019-12-25 CN CN201911365369.1A patent/CN111088309A/zh active Pending
Patent Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2002069733A1 (en) * | 2001-03-05 | 2002-09-12 | Council Of Scientific And Industrial Research | Process for preparation of protein-hydrolysate from soy flour |
CN103719534A (zh) * | 2013-12-23 | 2014-04-16 | 江南大学 | 一种对大豆分离蛋白进行酶水解的方法 |
CN104286370A (zh) * | 2014-08-20 | 2015-01-21 | 华南理工大学 | 一种酶辅助亚临界水法制备功能性大豆蛋白的方法 |
US20170143001A1 (en) * | 2015-04-30 | 2017-05-25 | China National Research Institute Of Food And Fermentation Industries | Soybean oligopeptide with low allergenicity and little bitterness and preparation method and application thereof |
Non-Patent Citations (2)
Title |
---|
周利亘等: "大豆多肽复合酶解工艺条件研究", 《食品科技》 * |
陈丽花等: "大豆蛋白抗氧化活性肽的制备工艺研究", 《食品工业》 * |
Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN111670996A (zh) * | 2020-06-08 | 2020-09-18 | 颜如玉医药科技有限公司 | 一种大豆蛋白肽的制备方法 |
CN111670996B (zh) * | 2020-06-08 | 2021-01-29 | 颜如玉医药科技有限公司 | 一种大豆蛋白肽的制备方法 |
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