CN1168770A - 蛋白水解产物和制备该产物的方法 - Google Patents
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Abstract
本发明提供了大豆球蛋白含量低的大豆蛋白水解产物以及生产该水解产物的方法,其中大豆蛋白之主要成分大豆球蛋白被选择性分解。通过使蛋白水解酶对大豆蛋白作用以选择性分解大豆蛋白中的大豆球蛋白而获得大豆球蛋白含量低的大豆蛋白水解产物,且生产大豆球蛋白含量低的大豆蛋白水解产物的方法包括在pH1.0至2.8,优选pH1.5至2.5时使蛋白水解酶对大豆蛋白作用。
Description
本发明涉及具有低含量特定组成蛋白的蛋白水解产物,特别是具有低含量大豆球蛋白的大豆蛋白水解产物,其中大豆蛋白中的主要成分大豆球蛋白被选择性地分解。本发明还涉及制备该水解产物的方法。发明背景
大豆含有大量高质蛋白质,长期以来一直被用作优良的蛋白来源。特别是由于其高蛋白含量和多种功能特性如乳化性,胶凝性,持水性质等优点,大豆蛋白分离产物被用作食品原料。
大豆蛋白由根据超离心沉降速率分类的2S、7S、11S和15S蛋白质等复杂的较高级结构的各种蛋白组成,并且这些蛋白质即使在物理性质方面也具有不同的特征。
例如,通过沉淀用水从脱脂大豆片中提取的豆奶而获得的大豆蛋白分离物基本上由7S球蛋白(主要是β-伴大豆球蛋白Conglycinin)和11S球蛋白(主要是大豆球蛋白)组成,并且各成份具有其固有的功能特性。但是,这些成份以其混合物的形式存在,因此各成份固有的功能特性在实际应用中不能被充分利用。
因此,为了利用其固有功能,已进行了许多偿试来分级分离各成份。例如对于实验性分级分离,Wolf等和Thanh等进行了研究和报导,方案已经在日本专利LOP公开号56843/1973,31843/1974,86149/1976,124457/1980,153562/1980,64755/1981,132844/1982和36345/1983中公开。但是这些现有方法只停留在实验阶段,不适于工业化分级分离。
在这种情况下,日本专利LOP公开187755/1986中提出在亚硫酸盐等存在时通过调节pH和温度的工业分离方法能够分级分离大豆蛋白成份,但在该方法中必须进行烦琐的pH和温度控制。
也进行了很多利用蛋白酶蛋白水解作用进行功能改进的研究。例如,日本专利公开号24262/1973,日本专利公开号1028/1980,日本专利LOP公开号232341/1987,日本专利公开号14941/1992等是有关这种改进的,但所有这些方法涉及如溶解性、非胶凝性等功能性改变,以在大豆蛋白热变性之前加速酶反应前的水解,而如对大豆蛋白中仅一种特定成份进行分解的这种功能性改变,未见有偿试。
对于天然形式的蛋白,包括大豆蛋白,用水解酶如蛋白酶将其分解常常是困难的(S.S.Nielsen等,《农业食品化学》(J.Agric.FoodChem),36,869(1988)),因此,在一般操作中,在蛋白酶解之前通过加热,醇等,使蛋白变性。
如上所述,大豆蛋白分离物是基本上由7S球蛋白(主要是β-伴大豆球蛋白)和11S球蛋白(主要是大豆球蛋白)组成的混合物,而且已知在相同条件下引起的各组分间的变性程度是不同的。例如,已知11S球蛋白在酸性pH下比7S球蛋白更容易变性(I.Koshiyama,《食品农业科学》(J.Sci、Fd Agrjc.,23,853(1972)),而且通过加热7S球蛋白在比11S球蛋白变性温度低(S.Damodaran,J.Agric,Food Chem.,36,262(1988))。
然而至今酶分解方法中不可能特异而专一地分解大豆蛋白中的一种特定成分,这可能是由于蛋白酶解前不可控制的蛋白变性处理,如过度加热,醇处理等。
如果能专一分解大豆蛋白中的特定成分,则可以从所需成分的混合物获得具有固有功能特性的大豆蛋白。
发明概述
在这种情况下,本发明的目的是提供有低含量特定成分蛋白的蛋白水解产物,特别是有低含量大豆球蛋白的大豆蛋白水解产物,其中大豆蛋白中的主要成分大豆球蛋白被选择性地分解,以及生产该大豆蛋白水解产物的方法。
作为深入研究的结果,本发明人发现这一事实,即作为大豆蛋白主要成分的大豆球蛋白和β一伴大豆球蛋白在特定酸性pH下具有不同程度的变性,并且他们发现通过使蛋白水解酶在该pH下作用能获得大豆球蛋白被选择性分解的大豆蛋白水解产物,从而实现本发明。
即本发明是一种生产低含量特定成分蛋白的蛋白水解产物的方法,其中使蛋白水解酶对含多种蛋白质的蛋白原料作用以选择性地分解特定成分蛋白。
所述蛋白水解酶对含有多种蛋白质的蛋白原料的作用在其中特定成分蛋白质被选择性地变性的条件下是有效的,且变性条件以pH调节和/或温度调节为基础。
在具有低含量特定成分蛋白质的蛋白水解产物中,特定蛋白质分解程度是60%或更多,优选80%或更多,非此特定成分蛋白质的主要成分蛋白质分解程度是40%或更少,优选20%或更少。
而且,本发明是一种生产具有低含量特定成分蛋白质的大豆蛋白水解产物的方法,其中使蛋白水解酶对大豆蛋白作用以选择性地分解大豆蛋白中的特定成分蛋白质。
作为大豆蛋白中特定成分蛋白质,提到的是大豆球蛋白。所述蛋白水解酶对大豆蛋白的作用在pH1.0至2.8,优选pH1.5至2.5时是有效的。
作为大豆蛋白中另一种特定成分蛋白质,提到的是β一伴大豆球蛋白。在这种情况下,所述蛋白水解酶对大豆蛋白的作用在高于50℃至低于90℃、优选55至85℃、更优选60至80℃的温度下是有效的。
而且,本发明是一种具有低含量β-伴大豆球蛋白的大豆蛋白水解产物,其中大豆球蛋白/β-伴大豆球蛋白之比是1.5或更大,优选2.5或更大,更优选3.0或更大,而三氯乙酸可溶蛋白与全部蛋白之比是5至20%(重量)。
附图简要说明
图1是SDS-电泳图。
发明的详细说明
下文详细说明本发明。
用于本发明的大豆蛋白包括大豆、脱皮大豆和以大豆蛋白为基础的全脂豆奶、脱脂豆奶、浓缩大豆蛋白、大豆蛋白分离物等,优选经不伴随蛋白质变性的加工处理的大豆蛋白加工产品,并且不限制原料大豆的品种和生产地区。一般情况下,优选原料是用正己烷作为提取剂进行低温提取处理的脱脂大豆片,而NSI(氮溶出率)为60或更大、优选80或更大的低度变性的脱脂大豆片是特别优选的。本发明中优选使用水提取这种低度变性的脱脂大豆片、脱脂豆奶、浓缩的大豆蛋白和大豆蛋白分离物。
用于本发明的蛋白水解酶必须是在pH1.0至2.8时有蛋白酶水解活性的酶制剂。这些可以是商购的来自植物、动物器官或微生物的酶制剂,并且它们的来源不受特别地限定,最优选使用胃蛋白酶。
为实施本发明,在生产大豆蛋白的过程中将蛋白水解酶加入到大豆蛋白中,其中在PH1.0至2.8,酶选择性地分解大豆球蛋白。在生产大豆蛋白分离物中,例如,用水提取低度变性的脱脂豆片,分离成水不溶部分(豆凝乳沉积物)和水可溶部分(豆奶),并且将水可溶部分等电沉淀进一步分离成水不溶部分(凝乳)和水可溶部分(乳清),并将酸沉淀的凝乳悬浮于水中,然后调节pH1.0-2.8并进行水解反应。然后将反应物中和,灭菌并干燥,得产品。或者,反应物可以在pH4.8左右即β-伴大豆球蛋白的等电点处进行酸沉淀,然后通过离心分离成上清液(主要是大豆球蛋白水解产物)和沉淀(主要是未分解的β-伴大豆球蛋白),这两者均可被中和,灭菌并干燥,得产品。
通常,在将含完整的大豆蛋白的水悬浮液调节至pH1.0-2.8后进行酶反应,然后向其中加入占所述水悬浮液的固体含量的0.001至0.5%,优选0.01至0.1%范围的蛋白水解酶。反应温度一般在20至50℃范围,优选30至40℃。反应一般进行5分钟至2小时,优选10至30分钟。通过将水悬浮液通过装载固定化酶的柱子也可以进行连续处理。
通过SDS-电泳并用考马斯蓝染色将酶解后大豆蛋白分离成它的成分。这样染色的各泳带密度能被用来评估大豆蛋白中各成分的变化。根据本发明,容易获得具有低含量大豆球蛋白的大豆蛋白水解产物,其中大豆球蛋白分解程度是60%或更多,优选80%或更多,β-伴大豆球蛋白的分解程度是40%或更少,优选20%或更少,换句话说,大豆球蛋白含量是大豆原料的40%或更少,优选20%或更少,而β-伴大豆球蛋白含量是大豆原料的60%或更多,优选80%或更多。
这种方法获得的具有低含量大豆球蛋白的大豆蛋白水解产物能有效地用作食品原料,以充分利用β-伴大豆球蛋白的功能。
实施例
下文中通过参照实施例来详细说明本发明,但并不意谓限制本发明范围。实施例1
向用正己烷为提取剂获得的100g低度变性的脱脂大豆片(氮溶出率:NSI>80)中加入10倍过量的水,室温和PH7时将悬浮液提取1小时,然后离心,得950g脱脂豆奶。用盐酸将950g脱脂豆奶调至pH4.5,然后离心移去乳清部分,这样得到100g酸沉淀凝乳。将100g酸沉淀凝乳悬浮于水中,然后用盐酸调至PH2.5。以所述悬浮液固体含量的0.05%的量向水悬浮液中加入胃蛋白酶(Sigma),且酶反应在37℃进行30分钟。用氢氧化钠中和酶反应物,且该溶液在140℃加热15秒并喷雾干燥得37g大豆蛋白(试验组)。作为对照组,将酸沉淀凝乳悬浮于水中,然后用氢氧化钠中和,在140℃加热15秒并喷雾干燥(对照组)。
用SDS-电泳分离试验组和对照组的每份10μg样品,并用光密度计检测用考马斯蓝染色的每条泳带的密度。表1给出试验组中与对照组的结果相比较表示的大豆球蛋白和β-伴大豆球蛋白减少的程度。结果表明几乎所有大豆蛋白中的大豆球蛋白被选择性地分解。实施例2
将用实施例1相同的方法制备的酸沉淀凝乳悬浮于水中,用盐酸将水悬浮液调至pH2.0,以悬浮液固体含量的0.05%的量加入胃蛋白酶(Sigma),酶反应在37℃进行30分钟。用氢氧化钠中和酶反应物,且溶液在140℃加热15秒并喷雾干燥制成大豆蛋白。实施例3
将用实施例1相同的方法制备的酸沉淀凝乳悬浮于水中,用盐酸将水悬浮液调至PH2.8,以悬浮液固体含量的0.05%的量加入胃蛋白酶(Sigma),酶反应在37℃进行30分钟。用氢氧化钠中和酶反应物,溶液在140℃加热15秒,并喷雾干燥制得大豆蛋白。对比实施例1
将用实施例1相同的方法制备的酸沉淀凝乳悬浮于水中,用盐酸将水悬浮液调至pH3.5,以悬浮液固体含量的0.05%的量加入胃蛋白酶(Sigma),酶反应在37℃进行30分钟。用氢氧化钠中和酶反应物,且溶液在140℃加热15秒并喷雾干燥制得大豆蛋白。对比实施例2
用实施例1相同的方法制备的脱脂大豆片奶在90℃加热30分钟,从中制备酸沉淀凝乳。凝乳悬浮于水中,用盐酸将水悬浮液调至pH2.5,并以悬浮液固体含量的0.05%的量加入胃蛋白酶(Sigma.),酶反应在37℃下进行30分钟。用氢氧化钠中和酶反应物,且溶液在140℃加热15秒,并喷雾干燥制得大豆蛋白。
用SDS-PAGE分离实施例2和3和对比实施例1和2的每份样品10μg,并用光密度计检测用考马斯蓝染色的各条泳带的密度。测定每份样品中大豆球蛋白和β-伴大豆球蛋白的减少程度,与实施例1中对照组中大豆球蛋白和β-伴大豆球蛋白含量(为100%)相比较。结果见表1。由对比实施例1和2的结果可以看出,在pH2.8或更大pH时很难发生大豆球蛋白和β-伴大豆球蛋白的分解,而如果在酶反应之前进行过量热变性,则发生大豆球蛋白和β-伴大豆球蛋白两者的分解,因而不可能得到选择性分解产物。
表1反应pH 大豆蛋白的减少(%) β-伴大豆蛋白的减少(%)pH2.5 96 4 实施例1pH2.0 98 15 实施例2pH2.8 65 2 实施例3pH3.5 8 2 对比实施例1pH2.5 96 94 对比实施例2(热变性后)
根据本发明,容易获得大豆球蛋白被选择性分解的大豆球蛋白含量低的大豆蛋白,所得大豆蛋白广泛用于各种食品工业,如肉加工、海产品加工、饮料等,因此对工业中的改进有很大贡献。
Claims (10)
1、一种生产特定成分蛋白含量降低的蛋白水解产物的方法,其中使蛋白水解酶作用于含有多蛋白的蛋白原料以选择性地分解特定成分蛋白质。
2、根据权利要求1的方法,其中使蛋白水解酶在选择性变性特定成分蛋白的条件下作用。
3、根据权利要求2的方法,其中通过pH调节和/或温度调节进行特定成分蛋白的选择性变性。
4、根据权利要求1-3的方法,其中特定成分蛋白的分解程度是60%或更多,优选80%或更多,且非特定成分蛋白的主要成分蛋白分解程度是40%或更少,优选20%或更少。
5、一种生产低含量特定蛋白的大豆蛋白水解产物的方法,其中使蛋白水解酶作用于大豆蛋白以选择性地分解大豆蛋白中的特定成分蛋白。
6、根据权利要求5的方法,其中特定成份蛋白是大豆球蛋白。
7、根据权利要求5和6的方法,其中使蛋白水解酶在pH1.0至2.8,优选pH1.5至2.5时作用。
8、根据权利要求5的方法,其中特定成分蛋白是β-伴大豆球蛋白。
9、根据权利要求5至8的方法,其中使蛋白水解酶在高于50℃至低于90℃、优选55至85℃、更优选60至80℃的温度作用。
10、一种β-伴大豆球蛋白含量低的蛋白水解产物,其中大豆球蛋白/β-伴大豆球蛋白之比是1.5或更大、优选2.5或更大、更优选3.0或更大,且三氯乙酸可溶性蛋白与总蛋白量之比是5至20%(重量)。
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