扁杏仁水解蛋白的抗氧化活性、理化性质及组成分析
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摘要
扁杏仁为大扁杏的种子，具有很高的营养价值。我国大扁杏在三北地区的栽培面积很大，扁杏仁资源丰富，而对于扁杏仁蛋白的研究尚处于初步阶段。对扁杏仁蛋白的深入研究可提高扁杏仁蛋白的利用率，并且为开发功能性扁杏仁蛋白产品提供依据。
本文以扁杏仁蛋白为对象，采用碱性蛋白酶、中性蛋白酶、木瓜蛋白酶、胰蛋白酶分别对扁杏仁蛋白进行酶解，碱性蛋白酶对扁杏仁蛋白酶解效果较好，当底物浓度为2%，E/S为10000u/g时，水解度可达19.3%。胰蛋白酶酶解扁杏仁蛋白最适底物浓度为1%，最适E/S为10000u/g。对扁杏仁水解蛋白体外抗氧化实验表明，水解度在14%～16%时，扁杏仁蛋白酶解物表现出较好的抗氧化活性。SDS-PAGE实验表明，通过蛋白酶的水解，扁杏仁蛋白分子量有一部分被水解为分子量小于10KDa的短肽。
为了进一步研究扁杏仁水解蛋白的活性组分，采用Sephadex G-100、G-25凝胶层析分离技术，对碱性蛋白酶酶解得到的扁杏仁水解蛋白进行分离，得出APH-1、APH-2、APH-21、APH-22、APH-23五个组分。并用HPLC方法对扁杏仁水解蛋白及分离组分进行检测，发现APH-2组分中小分子肽类达到90%以上。
为拓展水解蛋白的潜在应用领域，对扁杏仁水解蛋白与分离蛋白物理性质进行比较研究发现：水解蛋白的溶解性、吸水吸油性均优于分离蛋白。分离蛋白的起泡性优于水解蛋白，但水解蛋白的泡沫稳定性较好。NaCl浓度接近0.2mol/L时水解蛋白和分离蛋白的乳化能力最强，分别达到37.04m/g和47.44m/g。水解蛋白和分离蛋白的乳化能力在蔗糖浓度达到0.4g/L时最强，分别为17.58m/g和18.34m/g，分离蛋白的乳化稳定性稍高于水解蛋白。
鉴于喷雾干燥工艺在实际生产中对扁杏仁水解蛋白的干燥效果及抗氧化活性的影响，设计了响应面实验对其进行了探讨，得出结论：进料速度在270～320mL/h，脱水比例在0.5～0.6之间，进风温度200℃左右，可以获得较好的综合效果。从而为扁杏仁水解蛋白的实际生产提供理论依据。
As seeds of apricot, the sweet almonds have very high nutritional value. Theapricot is widely planted in Northeast of China, North China and Northwest ofChina. Although the harvest resources of the almond are rich, researching on theprotein of them is still in a primary stage. Study of almond protein will improvetheir availability and provide the basis theory for the development of variousalmond products.
This dissertation studied the almond protein by using alkaline protease,neutral protease, papain and trypsin to decompose the protein. The results showedthat alkaline protease was better under the conditions of concentration of2%and10000u/g of E/S and the degree of hydrolysis could reach to19.3%. For trypsin,the most optimized concentration was1%and the optimazed E/S was10000u/g.The experiment proved that under such conditons, the degree of hydrolysis isbetween14%～16%and showed the best antioxidant activity. SDS-PAGE showedthat part of the almond protein was decomposed to shorter peptide whichmolecular weight was less than10KDa.
In order to further researching the component of almond protein hydrolysate,the chromatography of Sephadex G-100and Sephadex G-25were adopted toseparate the almond protein hydrolysate which was modifed by alkali protease.Five constituents such as APH-1、APH-2、APH-21、APH-22、APH-23wereobtained.And the method of HPLC was used to analyze almond proteinhydrolysate and related components, which showed that low molecular weightpeptide consisted over90%in the constituents of APH-2.
To expand the potential application area of the protein hydrolysate, thephysical property between almond protein hydrolysate and protein isolate wasstudied comparely, which proved that the protein hydrolysate was better than protein isolate in dissolubility, hydroscopicity and oil absorption. In0minutes, theprotein isolate was better than protein hydrolysate in foamability, while was worsethan protein hydrolysate in foam stability. When the concentration of NaClapproached to0.2mol/L, the emulsification ability of protein hydrolysate andprotein isolate were the best that attained to37.04m/g and47.44m/g separately.When the concentration of sugar appoached to0.4g/L, the emulsification ability ofprotein hydrolysate and protein isolate were the best and reached to17.58m/g and18.34m/g. The emulsion stability of protein isolate was better than proteinhydrolysate.
In practical production, since spray drying technology had influces on thedrying effect and antioxidant activity of almond protein hydrolysate, a responsesurface experiment was designed to study the information. It concluded that thebest comprehensive effect could be reached when feed speed was270～320mL/h,dehydration ratio was between0.5～0.6and the inlet temperature was about200℃.The study will provide theoretical basis for the practical production of almondprotein hydrolysate.
本文以扁杏仁蛋白为对象，采用碱性蛋白酶、中性蛋白酶、木瓜蛋白酶、胰蛋白酶分别对扁杏仁蛋白进行酶解，碱性蛋白酶对扁杏仁蛋白酶解效果较好，当底物浓度为2%，E/S为10000u/g时，水解度可达19.3%。胰蛋白酶酶解扁杏仁蛋白最适底物浓度为1%，最适E/S为10000u/g。对扁杏仁水解蛋白体外抗氧化实验表明，水解度在14%～16%时，扁杏仁蛋白酶解物表现出较好的抗氧化活性。SDS-PAGE实验表明，通过蛋白酶的水解，扁杏仁蛋白分子量有一部分被水解为分子量小于10KDa的短肽。
为了进一步研究扁杏仁水解蛋白的活性组分，采用Sephadex G-100、G-25凝胶层析分离技术，对碱性蛋白酶酶解得到的扁杏仁水解蛋白进行分离，得出APH-1、APH-2、APH-21、APH-22、APH-23五个组分。并用HPLC方法对扁杏仁水解蛋白及分离组分进行检测，发现APH-2组分中小分子肽类达到90%以上。
为拓展水解蛋白的潜在应用领域，对扁杏仁水解蛋白与分离蛋白物理性质进行比较研究发现：水解蛋白的溶解性、吸水吸油性均优于分离蛋白。分离蛋白的起泡性优于水解蛋白，但水解蛋白的泡沫稳定性较好。NaCl浓度接近0.2mol/L时水解蛋白和分离蛋白的乳化能力最强，分别达到37.04m/g和47.44m/g。水解蛋白和分离蛋白的乳化能力在蔗糖浓度达到0.4g/L时最强，分别为17.58m/g和18.34m/g，分离蛋白的乳化稳定性稍高于水解蛋白。
鉴于喷雾干燥工艺在实际生产中对扁杏仁水解蛋白的干燥效果及抗氧化活性的影响，设计了响应面实验对其进行了探讨，得出结论：进料速度在270～320mL/h，脱水比例在0.5～0.6之间，进风温度200℃左右，可以获得较好的综合效果。从而为扁杏仁水解蛋白的实际生产提供理论依据。
As seeds of apricot, the sweet almonds have very high nutritional value. Theapricot is widely planted in Northeast of China, North China and Northwest ofChina. Although the harvest resources of the almond are rich, researching on theprotein of them is still in a primary stage. Study of almond protein will improvetheir availability and provide the basis theory for the development of variousalmond products.
This dissertation studied the almond protein by using alkaline protease,neutral protease, papain and trypsin to decompose the protein. The results showedthat alkaline protease was better under the conditions of concentration of2%and10000u/g of E/S and the degree of hydrolysis could reach to19.3%. For trypsin,the most optimized concentration was1%and the optimazed E/S was10000u/g.The experiment proved that under such conditons, the degree of hydrolysis isbetween14%～16%and showed the best antioxidant activity. SDS-PAGE showedthat part of the almond protein was decomposed to shorter peptide whichmolecular weight was less than10KDa.
In order to further researching the component of almond protein hydrolysate,the chromatography of Sephadex G-100and Sephadex G-25were adopted toseparate the almond protein hydrolysate which was modifed by alkali protease.Five constituents such as APH-1、APH-2、APH-21、APH-22、APH-23wereobtained.And the method of HPLC was used to analyze almond proteinhydrolysate and related components, which showed that low molecular weightpeptide consisted over90%in the constituents of APH-2.
To expand the potential application area of the protein hydrolysate, thephysical property between almond protein hydrolysate and protein isolate wasstudied comparely, which proved that the protein hydrolysate was better than protein isolate in dissolubility, hydroscopicity and oil absorption. In0minutes, theprotein isolate was better than protein hydrolysate in foamability, while was worsethan protein hydrolysate in foam stability. When the concentration of NaClapproached to0.2mol/L, the emulsification ability of protein hydrolysate andprotein isolate were the best that attained to37.04m/g and47.44m/g separately.When the concentration of sugar appoached to0.4g/L, the emulsification ability ofprotein hydrolysate and protein isolate were the best and reached to17.58m/g and18.34m/g. The emulsion stability of protein isolate was better than proteinhydrolysate.
In practical production, since spray drying technology had influces on thedrying effect and antioxidant activity of almond protein hydrolysate, a responsesurface experiment was designed to study the information. It concluded that thebest comprehensive effect could be reached when feed speed was270～320mL/h,dehydration ratio was between0.5～0.6and the inlet temperature was about200℃.The study will provide theoretical basis for the practical production of almondprotein hydrolysate.
引文
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