中国科学院过程工程研究所机构知识库

火麻(Cannabis sativa L)又称大麻，桑科大麻属植物，其栽培和利用已有上千年历史。火麻仁为火麻的种子仁，收录于2015年版《中国药典》，属于药食同源物质。四氢大麻酚含量低于0.3%的火麻称之为工业大麻，随着大麻二酚(CBD)开发持续火爆，工业大麻种植面积持续攀升，火麻籽产量剧增，迫切需要进行髙值化综合利用。蛋白质是火麻仁的主要成分，其提取分离通常采用碱提酸沉技术，产品颜色重、蛋白含量低限制了火麻仁蛋白的深度利用。针对上述问题，本文研究改进盐溶盐析法提取蛋白质，在理化性质和功能特性等方面与碱提酸沉制备的蛋白质进行了比较，并探索了超声改性火麻仁蛋白，以期为火麻仁蛋白深度开发利用提供依据。取得的主要研究结果如下：1.火麻仁蛋白制备研究。火麻仁蛋白通过两种方法制备，分别是碱提酸沉法和经过实验室改进的盐溶盐析法。采用正交试验优化这两种工艺，结果表明：碱提酸沉最佳工艺为提取温度50℃、碱液pH值9.5、料液比1:20，蛋白提取率83.43%，蛋白含量85.9%，外观颜色浅黄色。盐溶盐析最佳工艺为提取温度50℃、氯化钠质量分数5%、料液比1:20, 蛋白提取率69.09%，蛋白含量94.8%，外观颜色乳白色。2.火麻仁蛋白性能表征。构建了高效液相色谱检测氨基酸组成的方法，用于探究火麻仁蛋白中17种氨基酸的组成。火麻仁蛋白氨基酸组成丰富，包含人体所需的8种必需氨基酸，且含量高于30%；同时发现精氨酸含量高，其中HPI-SE精氨酸含量可达15.09%。SDS-PAGE表明，两种方法制备的火麻仁蛋白主要成分均为麻仁球蛋白，它由六个分子量为55 kDa的亚基组成，每个亚基由一个分子量34 kDa的酸性基团和一个分子量为18或19 kDa的碱性基团通过二硫键连接组成。不同pH条件下，两种火麻仁蛋白的溶解性曲线均呈U型，但是在相同pH条件下，二者的溶解性存在较大差异。溶液pH值小于5时，HPI-SE溶解性更高，当溶液pH值大于9.0时，HPI-AE溶解性更高。火麻仁蛋白的起泡性、持水力同蛋白质的溶解性呈现正相关；当pH值远离等电点时，同种电荷的静电斥力提高了蛋白质的溶解性，改善了蛋白质的起泡性、持水力等功能特性。3.火麻仁蛋白超声改性研究。以盐溶盐析法制备的火麻仁蛋白为原料，在超声功率400 w时，超声时间的延长可显著影响火麻仁蛋白的溶解性、起泡性。超声处理未对蛋白质的氨基酸组成、肽链分子量分布产生影响，但使火麻仁蛋白的表面结构变得疏松，覆盖的球形颗粒粒径减小，部分活性位点暴露出来，进而强化了蛋白质-水的相互作用，改善了火麻仁蛋白的溶解性、起泡性。;Hemp (Cannabis sativa L) has strong adaptability and wide distribution. Its cultivation and utilization have thousand year history. Hemp seed is included in Chinese Pharmacopoeia (2015 Edition), which is medicinal and edible substance. Hemp with tetrahydrocannabinol (THC) content less than 0.3% is called industrial hemp. With the popularity of research on cannabidiol (CBD), the planting area of hemp continues to rise, and the output of hemp seed increases dramatically, so it is urgent to carry out high-value comprehensive utilization. Protein is the main component of hemp seed, and the traditional technology of alkali extraction/acid precipitation is still used in the extraction of hemp seed protein. The heavy color, low protein content of the product limit the utilization of hemp seed protein. In view of the above problems, this paper studied the method of salt extraction/salt precipitation to extracted protein, and compared it with the protein obtained by alkali extraction/acid precipitation in the aspects of and functional properties, and explored the ultrasonic modification of hemp seed protein. The main research results are as follows:1. The preparation of hemp seed protein isolate. Hemp seed protein isolates (HPIs) from defatted hemp seed meal (HPM) were prepared using two methods, one was the traditional extraction method of alkaline extraction/acid precipitation (HPI-AE), the other was a laboratory improved extraction method of salt extraction/salt precipitation (HPI-SE). The orthogonal experiment was used to optimize the extraction process of hemp seed protein. The results showed that the best extraction process of alkali extraction/acid precipitation was extraction temperature 50 ℃, pH value 9.5, and solid-to-solvent ratio 1:20 (g/mL). And the best extraction process of salt extraction/salt precipitation was extraction temperature 50 ℃, mass fraction of sodium chloride 5%, and solid-to-solvent ratio 1:20 (g/mL). The protein content of HPI-SE 94.8% was higher than that of HPI-AE 85.9%, but the protein extraction rate of HPI-SE 69.09% was lower than that of HPI-AE 83.43%, the appearance color of HPI-SE was white, while that of HPI-AE was light yellow.2. The characterization of hemp seed protein. A HPLC method was constructed to study the composition of 17 amino acids in hemp seed protein. It was found that the content of essential amino acids in HPIs was higher than 30%, the content of arginine in HPIs was high, and the content of arginine in HPI-SE was as high as 15.09%. SDS-PAGE showed that the main component of HPIs was hemp seed globulin, which was composed of six subunits with molecular weight of 55 kDa, each of which was composed of an acid subunit with molecular weight of 34 kDa and an alkaline subunit with molecular weight of 18 or 19 kDa connected by a disulfide bond. Under different pH conditions, the solubility curves of HPIs were U-shaped, but under the same pH conditions, the solubility of HPIs were different: The solubility of HPI-SE was higher than HPI-AE when the pH value was less than 5; the solubility of HPI-AE was higher than HPI-SE when the pH value was greater than 9.0. The foaming and water holding capacity of HPIs were positively related to the solubility of protein. When the pH value was far away from the isoelectric point, the electrostatic repulsion of the same charge improved the solubility, foaming capacity and water holding capacity.3. The ultrasonic modification of hemp seed protein. The effect of ultrasonic treatment on the functional properties of HPI-SE and its mechanism were studied. It was found that the solubility and foaming capacity of HPI-SE were significantly affected by the prolongation of ultrasound time when the ultrasonic power was 400 w. The results showed that the ultrasonic treatment had no effect on the amino acid composition and the molecular weight distribution of peptide chain; the surface structure of hemp seed protein became loose, the size of covered spherical particles decreased, and the content of sulfhydryl group increased. The results showed that the three-dimensional structure of hemp seed protein became loose and some active sites were exposed, which enhanced the interaction between protein and water, thus improving the solubility and foaming capacity of hemp seed protein.