Characterization and Effects of pH on the Conformation of Hemp Protein Isolate Based on Multi-Spectroscopic Technique
ZENG Jian-hua, MENG Yan, LIU Lin-lin, YANG Yang, LI Mei-ying, WANG Zi-yue, ZHU Xiu-qing*, SHI Yan-guo*
Key Laboratory of Grain Food and Comprehensive Processing of Heilongjiang Province, College of Food Engineering, Harbin University of Commerce, Harbin 150076, China
Abstract：Hemp seeds are the important resource of high-quality vegetable oil, while the numerous high-quality protein in the hemp seed meal has not been fully exploited. Hemp protein isolates (HPI) are the resource of the high-quality protein, containing 20 kinds of amino acids with a relative equilibrium of the content of 8 essential amino acid and a high content of Agr. However,the application of HPI in food processing was severely hindered by the poor protein solubility. At present, there was a few pieces of literature on the conformation of HPI, and the effect of pH change on its conformation during processing has not been reported. In this study, the conformation of hemp protein isolate (HPI) was characterized by Circular dichroism (CD) spectrum, Fourier transforms infrared (FTIR) spectroscopy, Fluorescence spectrum and Ultraviolet (UV) spectrum, which compared with soybean protein isolate (SPI). The effects of different pH conditions on the conformation of HPI were investigated. Conformation results showed that comparing with SPI, there was more α-helix content in HPI, determined by CD and FTIR, which was 39.1% and 33.1%, respectively, while SPI contained β-sheet. The exposure of aromatic amino acid residues from HPI was lower than SPI, revealed by intrinsic fluorescence and UV spectroscopy. Moreover, the content of disulfide bond of HPI (10.06 mmol·g-1) was more than SPI, as was as higher surface hydrophobicity (374), but the value of free sulfydryl/total free sulfydryl of HPI was significantly lower than that of SPI. These results suggested that the tertiary structure of HPI was more compact than SPI, leading to a smaller particle size of nature HPI. The structure characteristics of HPI were changed in strong acid and strong alkaline conditions, which induced the structure of HPI to unfold and increased the size. As a result, the distance between aromatic amino acid residues was increased, and the energy transfer efficiency of Phe to Tyr and Trp was decreased. Finally, the fluorescence intensity of Try residues was decreased, and the fluorescence of Phe and Tyr residues was partly showed. Comparatively speaking, under acidic conditions, the conformation stability of HPI was relatively poor, and dissociation-association reactions were prone to the protein subunits. However, the conformation of HPI was relatively stable and with more flexibility. Therefore, with the help of fluorescence spectrum and UV second derivative spectrum, changes in the microenvironment of amino acid residues of HPI could be predicted, so as to understand changes in the three-dimensional structure of protein molecules. In conclusion, further understanding of HPI conformation and clarifying the influence of pH changes on HPI conformation are conducive to providing a theoretical reference for specific modification of HPI to prepare high-quality HPI and active physiological polypeptides.
Key words：Hemp protein isolate; Conformation characterization; Spectrum technology; pH
曾剑华，孟 妍，刘琳琳，杨 杨，李美莹，王子玥，朱秀清，石彦国. 基于多光谱技术表征汉麻分离蛋白构象并分析pH对其构象的影响[J]. 光谱学与光谱分析, 2020, 40(12): 3748-3754.
ZENG Jian-hua, MENG Yan, LIU Lin-lin, YANG Yang, LI Mei-ying, WANG Zi-yue, ZHU Xiu-qing, SHI Yan-guo. Characterization and Effects of pH on the Conformation of Hemp Protein Isolate Based on Multi-Spectroscopic Technique. SPECTROSCOPY AND SPECTRAL ANALYSIS, 2020, 40(12): 3748-3754.
[1] Aluko R. Sustainable Protein Sources. Boston, Academic Press，2017. 121.
[2] Malomo S A, Aluko R E. Food Hydrocolloids, 2015, 43: 743.
[3] Teh S S, Bekhit A E A, Carne A, et al. Food Chem., 2016, 203: 199.
[4] Hadna ev M, Dapčevic′-Hadna ev T, Lazaridou A, et al. Food Hydrocolloids, 2018, 79: 526.
[5] Malomo S A, Onuh J O, Girgih A T, et al. Nutrients, 2015, 7(9): 7616.
[6] Malomo S A, He R, Aluko r E. J. Food Sci., 2014, 79(8): C1512.
[7] ZHANG Tao, LU Rong-rong, QIAN Ping, et al(张 涛, 卢蓉蓉, 钱 平, 等). Food and Fermentation Industries(食品与发酵工业)，2008, 34(8): 173.
[8] Tang C H, Wang X S, Yang X Q. Food Chem., 2009, 114(4): 1484.
[9] Beveridge T, Toma S, Nakai S. J. Food Sci., 1974, 39(1): 49.
[10] Maruyama N, Adachi M, Takahashi K, et al. Eur. J. Biochem., 2010, 268(12): 3595.
[11] Kundu A, Kundu S, Chattopadhyay K. Protein. Sci., 2017, 26(3): 536.
[12] Siano F, Sorrentino G, Riccardi M, et al. Eur. Food Res. Technol., 2018, 244(5): 795.
[13] Tang C H, Ten Z, Wang X S, et al. J. Agr. Food. Chem., 2006, 54(23): 8945.
[14] Abugoch L E, Romero N, Tapia C A, et al. J. Agr. Food Chem., 2008, 56(12): 4745.
[15] ZENG Jian-hua, YANG Yang, LIU Lin-lin, et al(曾剑华, 杨 杨, 刘琳琳, 等). Food Science(食品科学)，2019, 40(11): 303.
