Article(id=1320881957412647357, tenantId=1146029695717560320, journalId=1260987677001138203, issueId=1320881182762447607, articleNumber=null, orderNo=null, doi=10.13386/j.issn1002-0306.2025080137, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=1755014400000, receivedDateStr=2025-08-13, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1792852399446, onlineDateStr=2026-10-24, pubDate=1786723200000, pubDateStr=2026-08-15, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1792852399446, onlineIssueDateStr=2026-10-24, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1792852399446, creator=13701087609, updateTime=1792852399446, updator=13701087609, issue=Issue{id=1320881182762447607, tenantId=1146029695717560320, journalId=1260987677001138203, year='2026', volume='47', issue='16', pageStart='1', pageEnd='537', issueExtLink='null', onlineDate='null', pubDate='1786723200000', pubDateStr='2026-08-15', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=null, createTime=1792852214756, creator='13701087609', updateTime=1792852214756, updator='13701087609', preIssue=null, nextIssue=null, articleTotal=null, ext=null, issueFiles=null, downloadFileDto=null}, startPage=148, endPage=154, ext={EN=ArticleExt(id=1320881957605585342, articleId=1320881957412647357, tenantId=1146029695717560320, journalId=1260987677001138203, language=EN, title=Enhancement of Rice Bran Protein Solubility via pH-Driven Modification, columnId=1261336277291548795, journalTitle=Science and Technology of Food Industry, columnName=Research and Investigation, runingTitle=null, highlight=null, articleAbstract=

To enhance the solubility of rice bran protein (RBP) and thereby improve its application performance in food processing, the pH-driven method was employed in this study, under which RBP was treated at pH conditions of 1.5, 2.5, 3.5, 10, 11, and 12, and structural and functional properties before and after modification were compared and analyzed. The results showed that the solubility of RBP was observed to be gradually increased with rising pH, with the optimal effect achieved at pH12, where a solubility of 70.57% was reached, representing a significant improvement of 63.85% compared to the untreated group (6.72%). The average particle size, absolute ζ-potential value, and surface hydrophobicity of RBP were found to be significantly altered as the pH increased; After treatment at pH12, the average particle size was reduced to the minimum (140.47 nm), the absolute ζ-potential value was increased to the maximum (14.8 mV), and the surface hydrophobicity was elevated to the highest value (2289), along with a marked enhancement in intrinsic fluorescence intensity and a red shift in the maximum emission wavelength. Following pH12 treatment, the relative content of β-sheets in RBP was increased by 10.43%, while the relative contents of α-helices and β-turns were reduced by 3.29% and 9.13%, respectively. These findings indicate that pH-driven method can induce depolymerization of RBP molecules and strengthen intermolecular electrostatic repulsion, thereby significantly improving its solubility. This study offers a new approach to promote the widespread application of RBP.

, authors=Fengzhu TIAN1, Dan LI1, 2, Feng ZUO1, 3, *, Hao HU4, authorsList=Fengzhu TIAN, Dan LI, Feng ZUO, Hao HU, authorCompany=null, correspAuthors=Feng ZUO, authorNote=null, correspAuthorsNote=null, copyrightStatement=Copyright © 2026 Science and Technology of Food Industry. All rights reserved., copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, fund=null), CN=ArticleExt(id=1320881959820177867, articleId=1320881957412647357, tenantId=1146029695717560320, journalId=1260987677001138203, language=CN, title=pH驱动法改善米糠蛋白溶解特性, columnId=1261336277849391232, journalTitle=食品工业科技, columnName=研究与探讨, runingTitle=null, highlight=null, articleAbstract=

为提高米糠蛋白(Rice bran protein,RBP)的溶解性,进而改善其在食品加工中的应用性能,本研究采用pH驱动法,在pH1.5、2.5、3.5、10、11及12条件下对RBP进行处理,并对比分析改性前后RBP的结构与功能特性变化。结果显示,RBP溶解度随pH升高呈逐渐提升趋势,其中pH12条件下处理效果最佳,溶解性达70.57%,较未处理组(6.72%)显著提高63.85%;RBP的平均粒径、ζ-电位绝对值及表面疏水性随pH升高发生显著变化,pH12处理后,其平均粒径降至最小(140.47 nm),ζ-电位绝对值增至最大(14.8 mV),表面疏水性达最高(2289),且内源荧光强度明显增强,最大发射波长出现红移;pH12处理后,RBP的β-折叠相对含量增加10.43%,α-螺旋与β-转角相对含量分别减少3.29%和9.13%。综上,pH驱动法可促使RBP分子发生解聚,并增强分子间静电斥力,从而显著提升其溶解性。本研究为推动RBP广泛应用提供了新方案。

, authors=田凤珠1, 李丹1, 2, 左锋1, 3, *, 胡昊4, authorsList=田凤珠, 李丹, 左锋, 胡昊, authorCompany=null, correspAuthors=左锋, authorNote=

田凤珠(2000−),女,硕士研究生,研究方向:农产品加工及贮藏工程,E-mail:

, correspAuthorsNote=
左锋(1979−),男,博士,教授,研究方向:植物蛋白及农副产品加工利用,E-mail:。
, copyrightStatement=版权所有 © 2026《食品工业科技》编辑部, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=zk3XhO+Ij81YRn1heF9DuA==, magXml=8L5+Vn/eu7p9qorLzDsMxg==, pdfUrl=null, pdf=cH8goxssHPVo6gwmD+HSFA==, pdfFileSize=1228961, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=3znorJJd6QtHXzljIxZotA==, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=0APKGD+IPHtJdzU/cTGzuQ==, mapNumber=null, fund=null)}, authors=[Author(id=1320881960625484249, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, orderNo=0, firstName=null, middleName=null, lastName=null, nameCn=null, orcid=null, stid=null, country=null, authorPic=null, dead=0, email=1456514212@qq.com, emailSecond=null, emailThird=null, correspondingAuthor=0, authorType=1, ext={EN=AuthorExt(id=1320881960730341851, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, authorId=1320881960625484249, language=EN, stringName=Fengzhu TIAN, firstName=Fengzhu, middleName=null, lastName=TIAN, prefix=null, suffix=null, authorComment=null, nameInitials=null, affiliation=null, department=null, xref=1, address=1College of Food Science, Heilongjiang Bayi Agricultural University, Daqing 163319, China, bio=null, bioImg=null, bioContent=null, aboutCorrespAuthor=null), CN=AuthorExt(id=1320881960818422236, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, authorId=1320881960625484249, language=CN, stringName=田凤珠, firstName=null, middleName=null, lastName=null, prefix=null, suffix=null, authorComment=null, nameInitials=null, affiliation=null, department=null, xref=1, address=1黑龙江八一农垦大学食品学院,黑龙江大庆 163319, bio={"content":"

田凤珠(2000−),女,硕士研究生,研究方向:农产品加工及贮藏工程,E-mail:

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田凤珠(2000−),女,硕士研究生,研究方向:农产品加工及贮藏工程,E-mail:

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Food Science, 2020, 41(18): 51−57., articleTitle=null, refAbstract=null)], funds=null, companyList=[AuthorCompany(id=1320881960067641804, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, xref=1, ext=[AuthorCompanyExt(id=1320881960076030413, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, companyId=1320881960067641804, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=1College of Food Science, Heilongjiang Bayi Agricultural University, Daqing 163319, China), AuthorCompanyExt(id=1320881960084419022, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, companyId=1320881960067641804, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=1黑龙江八一农垦大学食品学院,黑龙江大庆 163319)]), AuthorCompany(id=1320881960315105743, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, xref=2, ext=[AuthorCompanyExt(id=1320881960323494352, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, companyId=1320881960315105743, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=2National Engineering Technology Center of Cereals, Heilongjiang Bayi Agricultural University, Daqing 163319, China), AuthorCompanyExt(id=1320881960336077265, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, companyId=1320881960315105743, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=2黑龙江八一农垦大学国家杂粮工程技术中心,黑龙江大庆 163319)]), AuthorCompany(id=1320881960424157650, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, xref=3, ext=[AuthorCompanyExt(id=1320881960432546259, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, companyId=1320881960424157650, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=3Engineering Research Center of Processing and Utilization of Grain By-products and Utilization of Ministry of Education, Daqing 163319, China), AuthorCompanyExt(id=1320881960440934868, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, companyId=1320881960424157650, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=3粮食副产物加工与利用教育部工程研究中心,黑龙江大庆 163319)]), AuthorCompany(id=1320881960524820949, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, xref=4, ext=[AuthorCompanyExt(id=1320881960537403862, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, companyId=1320881960524820949, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=4Heilongjiang Beidahuang Rice Industry Group Co., Ltd., Harbin 150090, China), AuthorCompanyExt(id=1320881960545792471, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, companyId=1320881960524820949, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=4黑龙江省北大荒米业集团有限公司,黑龙江哈尔滨 150090)])], figs=[ArticleFig(id=1320881962546475510, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=EN, label=Fig.1, caption=Effects of pH-driven processing on RBP particle size distribution(A), average particle size, PDI (B), and Zeta potential (C), figureFileSmall=y0drwq0ZkJ3zC5CkoL2J4A==, figureFileBig=3znorJJd6QtHXzljIxZotA==, tableContent=null), ArticleFig(id=1320881962609390071, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=CN, label=图1, caption=pH驱动处理对RBP粒径分布(A)、平均粒径及PDI(B)和Zeta电位(C)的影响

注:不同字母表示差异显著(P<0.05),图2、图6同。

, figureFileSmall=y0drwq0ZkJ3zC5CkoL2J4A==, figureFileBig=3znorJJd6QtHXzljIxZotA==, tableContent=null), ArticleFig(id=1320881962785550840, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=EN, label=Fig.2, caption=Effect of pH-driven processing on surface hydrophobicity of RBP, figureFileSmall=NBP8FNcbqVkEhaldA3EdFQ==, figureFileBig=8wg+oueBu34ulsrms36dOA==, tableContent=null), ArticleFig(id=1320881962844271097, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=CN, label=图2, caption=pH驱动处理对RBP 表面疏水性的影响

注:不同字母的数值表示差异显著(P<0.05)。

, figureFileSmall=NBP8FNcbqVkEhaldA3EdFQ==, figureFileBig=8wg+oueBu34ulsrms36dOA==, tableContent=null), ArticleFig(id=1320881962907185658, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=EN, label=Fig.3, caption=Effect of pH-driven processing on intrinsic fluorescence of RBP, figureFileSmall=aRPD9y6hAoIlEYlPKXVEbw==, figureFileBig=I1Clu4CGpidGuytvTrnjNg==, tableContent=null), ArticleFig(id=1320881962965905915, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=CN, label=图3, caption=pH驱动处理对RBP内源荧光的影响, figureFileSmall=aRPD9y6hAoIlEYlPKXVEbw==, figureFileBig=I1Clu4CGpidGuytvTrnjNg==, tableContent=null), ArticleFig(id=1320881963033014780, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=EN, label=Fig.4, caption=Effect of pH-driven processing on fourier transform infrared spectroscopy of RBP, figureFileSmall=801zhHsRcyk7I35fM62L2A==, figureFileBig=cIEsr3g+oNB70q5PBhIBvA==, tableContent=null), ArticleFig(id=1320881963108512253, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=CN, label=图4, caption=pH驱动处理对RBP傅里叶红外光谱的影响, figureFileSmall=801zhHsRcyk7I35fM62L2A==, figureFileBig=cIEsr3g+oNB70q5PBhIBvA==, tableContent=null), ArticleFig(id=1320881963184009726, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=EN, label=Fig.5, caption=SDS-PAGE pattern of RBP treated by pH-driven, figureFileSmall=7T9ab04HtHHkRBG+TVr9IA==, figureFileBig=8/IOd4MOm+GdTEpvf9/q3g==, tableContent=null), ArticleFig(id=1320881963259507199, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=CN, label=图5, caption=pH驱动处理的米糠蛋白SDS-PAGE图

注:1~7为不同pH处理的样品:对照、pH10、pH11、pH12、pH1.5、pH2.5、pH3.5。

, figureFileSmall=7T9ab04HtHHkRBG+TVr9IA==, figureFileBig=8/IOd4MOm+GdTEpvf9/q3g==, tableContent=null), ArticleFig(id=1320881963322421760, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=EN, label=Fig.6, caption=Effect of pH-driven processing on solubility of RBP, figureFileSmall=+YPZ01gEVVpUesjw99l4JQ==, figureFileBig=jZg4IbsHb84AKtVzlS9HFA==, tableContent=null), ArticleFig(id=1320881963389530625, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=CN, label=图6, caption=pH驱动处理对RBP溶解度的影响, figureFileSmall=+YPZ01gEVVpUesjw99l4JQ==, figureFileBig=jZg4IbsHb84AKtVzlS9HFA==, tableContent=null), ArticleFig(id=1320881963473416706, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=EN, label=Table 1, caption=

Effect of pH-driven modification on the secondary structures of RBP

, figureFileSmall=null, figureFileBig=null, tableContent=
样品α-螺旋(%)β-折叠(%)β-转角(%)无规则卷曲(%)
对照11.44±0.06a17.12±0.08c13.46±0.12a20.55±0.03a
pH1.59.86±0.13b19.47±0.11c11.47±0.09d20.64±0.05a
pH2.58.87±0.1c19.08±0.07c11.75±0.11c20.62±0.05a
pH3.58.33±0.07d16.15±0.08c12.88±0.1b20.61±0.08a
pH101.16±0.08g16.64±0.13c9.15±0.07e20.63±0.02a
pH113.93±0.05f22.98±0.09b8.26±0.09f20.62±0.06a
pH128.15±0.06e27.55±0.1a4.33±0.11g20.65±0.06a
), ArticleFig(id=1320881963557302787, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320881957412647357, language=CN, label=表1, caption=

pH驱动处理对RBP二级结构的影响

, figureFileSmall=null, figureFileBig=null, tableContent=
样品α-螺旋(%)β-折叠(%)β-转角(%)无规则卷曲(%)
对照11.44±0.06a17.12±0.08c13.46±0.12a20.55±0.03a
pH1.59.86±0.13b19.47±0.11c11.47±0.09d20.64±0.05a
pH2.58.87±0.1c19.08±0.07c11.75±0.11c20.62±0.05a
pH3.58.33±0.07d16.15±0.08c12.88±0.1b20.61±0.08a
pH101.16±0.08g16.64±0.13c9.15±0.07e20.63±0.02a
pH113.93±0.05f22.98±0.09b8.26±0.09f20.62±0.06a
pH128.15±0.06e27.55±0.1a4.33±0.11g20.65±0.06a
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pH驱动法改善米糠蛋白溶解特性
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田凤珠 1 , 李丹 1, 2 , 左锋 1, 3, * , 胡昊 4
食品工业科技 | 研究与探讨 2026,47(16): 148-154
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食品工业科技 |研究与探讨 2026 , 47 (16) : 148 -154
pH驱动法改善米糠蛋白溶解特性
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田凤珠1 , 李丹1, 2, 左锋1, 3, * , 胡昊4
作者信息
  • 1黑龙江八一农垦大学食品学院,黑龙江大庆 163319
  • 2黑龙江八一农垦大学国家杂粮工程技术中心,黑龙江大庆 163319
  • 3粮食副产物加工与利用教育部工程研究中心,黑龙江大庆 163319
  • 4黑龙江省北大荒米业集团有限公司,黑龙江哈尔滨 150090
通讯作者:
左锋(1979−),男,博士,教授,研究方向:植物蛋白及农副产品加工利用,E-mail:。
作者简介:

田凤珠(2000−),女,硕士研究生,研究方向:农产品加工及贮藏工程,E-mail:

Enhancement of Rice Bran Protein Solubility via pH-Driven Modification
Fengzhu TIAN1 , Dan LI1, 2, Feng ZUO1, 3, * , Hao HU4
Affiliations
  • 1College of Food Science, Heilongjiang Bayi Agricultural University, Daqing 163319, China
  • 2National Engineering Technology Center of Cereals, Heilongjiang Bayi Agricultural University, Daqing 163319, China
  • 3Engineering Research Center of Processing and Utilization of Grain By-products and Utilization of Ministry of Education, Daqing 163319, China
  • 4Heilongjiang Beidahuang Rice Industry Group Co., Ltd., Harbin 150090, China
出版时间: 2026-08-15 doi: 10.13386/j.issn1002-0306.2025080137
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为提高米糠蛋白(Rice bran protein,RBP)的溶解性,进而改善其在食品加工中的应用性能,本研究采用pH驱动法,在pH1.5、2.5、3.5、10、11及12条件下对RBP进行处理,并对比分析改性前后RBP的结构与功能特性变化。结果显示,RBP溶解度随pH升高呈逐渐提升趋势,其中pH12条件下处理效果最佳,溶解性达70.57%,较未处理组(6.72%)显著提高63.85%;RBP的平均粒径、ζ-电位绝对值及表面疏水性随pH升高发生显著变化,pH12处理后,其平均粒径降至最小(140.47 nm),ζ-电位绝对值增至最大(14.8 mV),表面疏水性达最高(2289),且内源荧光强度明显增强,最大发射波长出现红移;pH12处理后,RBP的β-折叠相对含量增加10.43%,α-螺旋与β-转角相对含量分别减少3.29%和9.13%。综上,pH驱动法可促使RBP分子发生解聚,并增强分子间静电斥力,从而显著提升其溶解性。本研究为推动RBP广泛应用提供了新方案。

米糠蛋白  /  pH驱动法  /  溶解性  /  结构

To enhance the solubility of rice bran protein (RBP) and thereby improve its application performance in food processing, the pH-driven method was employed in this study, under which RBP was treated at pH conditions of 1.5, 2.5, 3.5, 10, 11, and 12, and structural and functional properties before and after modification were compared and analyzed. The results showed that the solubility of RBP was observed to be gradually increased with rising pH, with the optimal effect achieved at pH12, where a solubility of 70.57% was reached, representing a significant improvement of 63.85% compared to the untreated group (6.72%). The average particle size, absolute ζ-potential value, and surface hydrophobicity of RBP were found to be significantly altered as the pH increased; After treatment at pH12, the average particle size was reduced to the minimum (140.47 nm), the absolute ζ-potential value was increased to the maximum (14.8 mV), and the surface hydrophobicity was elevated to the highest value (2289), along with a marked enhancement in intrinsic fluorescence intensity and a red shift in the maximum emission wavelength. Following pH12 treatment, the relative content of β-sheets in RBP was increased by 10.43%, while the relative contents of α-helices and β-turns were reduced by 3.29% and 9.13%, respectively. These findings indicate that pH-driven method can induce depolymerization of RBP molecules and strengthen intermolecular electrostatic repulsion, thereby significantly improving its solubility. This study offers a new approach to promote the widespread application of RBP.

rice bran protein  /  pH-driven  /  solubility  /  structure
田凤珠, 李丹, 左锋, 胡昊. pH驱动法改善米糠蛋白溶解特性. 食品工业科技, 2026 , 47 (16) : 148 -154 . DOI: 10.13386/j.issn1002-0306.2025080137
Fengzhu TIAN, Dan LI, Feng ZUO, Hao HU. Enhancement of Rice Bran Protein Solubility via pH-Driven Modification[J]. Science and Technology of Food Industry, 2026 , 47 (16) : 148 -154 . DOI: 10.13386/j.issn1002-0306.2025080137
米糠是稻米加工的主要副产物,主要由清蛋白(34.39%)、球蛋白(30.74%)、谷蛋白(23.4%)和醇溶蛋白(5.54%)组成[1]。米糠富含蛋白质(约10%~20%)、脂肪(15%~20%)、膳食纤维(25%~40%)、碳水化合物(34%~54%),是一种营养价值较高的可再生资源[2−4]。然而,目前米糠的利用率较低,多用作动物饲料,造成资源浪费[5−6]。米糠蛋白(Rice bran protein,RBP)被称为“天赐营养源”[7],是一种优质植物蛋白,必需氨基酸组成接近FAO/WHO推荐标准[8],生物效价与牛奶、鸡蛋相当,其中赖氨酸含量高达5.8%[9]。RBP不含抗营养因子,过敏性低,适用于特殊人群及婴幼儿食品[10]。但由于其易聚集、溶解度低的特性,在食品工业中的应用受到限制[8]。
溶解性既是蛋白功能性质的基本指标[11],又影响着乳化性、起泡性和凝胶性等其他功能[12]。良好的蛋白溶解性是食品加工领域应用的前提和基础,这也有利于改进食品组织结构,以提高食品保存性[13]。因此溶解性是目前植物蛋白的研究热点。pH驱动法可有效诱导蛋白质亚基解离,并通过结构修饰提高其溶解性。pH驱动法是指在极端pH下,蛋白质分子侧链带同种电荷,静电斥力导致氢键和二硫键断裂,蛋白质从紧密结构变为松散状态从而改变蛋白质构象,恢复中性pH后,蛋白质重新折叠形成“熔融球”状态,疏水基团暴露但结构更松散,从而有效改善蛋白质功能特性的方法[14−15]。pH驱动法操作简便,无需使用有机试剂,具有成本低和效率高等优势[16−18]。该法能有效改善蛋白质的溶解特性[19]。Guo等[20]发现pH偏移处理可以破坏蛋白分子间氢键和疏水相互作用,从而提升溶解度。张唯唯等[21]将蛋白极端酸碱pH偏移处理后,尤其是pH为10时,其溶解性有所增强。
本研究主要利用pH驱动法对RBP进行简单、安全、高效的修饰,研究其结构和溶解性的变化,对解决RBP加工性能不佳、利用率较差等问题具有指导意义,也为今后RBP在食品行业中的广泛应用提供理论基础。
RBP(含量≥90%) 陕西帕尼尔生物科技有限公司;十二烷基硫酸钠(sodium dodecyl sulfate,SDS) 北京兰杰柯科技有限公司;1-苯胺基萘-8-磺酸(1-anilinonaphthalene-8-sulfonic acid,ANS) 美国Sigma-Aldrich公司;其他试剂均为分析纯。
ZEN3700电位测定仪 北京海鑫瑞科技有限公司;CF16RN高速冷冻离心机 日本日立公司;DYCZ-24K型电泳仪 北京六一生物科技有限公司;CHRISTALPha型冷冻干燥机 德国Marin Christ公司;FE28型pH计 梅特勒-托利多仪器(上海)公司;FST-RO-JK10超纯水机 上海富诗特环保科技有限公司;HJ-5搅拌器 常州市瑞华仪器制造有限公司;U-2910紫外分光光度计 上海美谱达仪器有限公司;FP-8000荧光光谱仪 德国Eppendorf公司;TY-80S脱色摇床 江苏金怡仪器科技有限公司。
分别配制6份浓度为0.01 g/mL的RBP,于磁力搅拌器室温搅拌1 h。其中3份分别用2 mol/L HCl溶液调节pH至1.5、2.5和3.5,另3份用2 mol/L NaOH溶液调节pH至10.0、11.0和12.0。所有样品在目标pH下保持2 h后,再用相应酸碱溶液将pH统一回调至7.0。将蛋白溶液装入截留分子量10000 Da的透析袋中,在4 ℃条件下持续透析48 h以去除盐离子。透析完成后,将所得蛋白质溶液进行冷冻干燥处理,冻干成粉末后贮藏于干燥器中室温保存备用。以未经酸碱处理的RBP溶液作为对照[22]。
参考邹赫楠[23]的研究方法并稍作修改,具体操作如下:取不同样品使用去离子水配制成0.1 mg/mL(pH7.0),磁力搅拌30 min后,于8000×g离心10 min,随后采用电位测定仪测定上清液的粒径、PDI及Zeta电位。
采用ANS荧光探针法测定RBP表面疏水性(H0),参照Yu等[24]研究方法并稍作调整。具体步骤如下:将不同样品用去离子水配制成1 mg/mL母液,梯度稀释至0.005~0.50 mg/mL系列浓度。取4 mL不同浓度蛋白溶液与50 μL、8 mmol/L ANS探针溶液于试管中混匀,在激发波长390 nm、发射波长470 nm、狭缝宽度5 nm条件下测定荧光强度。以蛋白浓度为横坐标、荧光强度为纵坐标绘制标准曲线,其线性回归方程的初始斜率即为RBP表面疏水性指数。
采用Zhou等[25]的方法并稍作改进,测定RBP的内源荧光光谱。具体步骤如下:将不同样品用去离子水稀释至0.1 mg/mL,室温下以8000×g离心10 min后取上清液,置于四面透光石英比色皿中。设置荧光分光光度仪参数:激发波长290 nm,发射波长扫描范围300~500 nm,扫描速率12000 nm/min,激发与发射狭缝宽度均为5 nm,随后进行荧光光谱测定。
参考Yang等[26]的研究方法并稍作改进,具体操作步骤如下:首先将不同冻干样品与溴化钾按1:100的质量比均匀混合,使用玛瑙研钵充分研磨至粉末状。随后将混合粉末压制成透明薄片,置于傅里叶变换红外光谱仪中进行蛋白质二级结构分析。仪器参数设置如下:扫描次数32次,分辨率4 cm−1,扫描波数范围400~4000 cm−1。所得光谱数据采用PeakFit 4.12软件进行基线校正和降噪处理。为进一步提高谱图分辨率,对数据进行二阶导数处理。最后通过分峰拟合分析确定各特征吸收峰,据此解析蛋白质二级结构的组成及其相对含量。
参考Ma等[27]的方法,采用不连续SDS-聚丙烯酰胺凝胶电泳(SDS-PAGE)进行实验。首先配制12%的分离胶和5%的浓缩胶,并以含0.05 mol/L Tris-0.384 mol/L甘氨酸和0.1%的SDS(pH8.3)的溶液作为电极缓冲液。将不同样品稀释至5 mg/mL后,分别与5× SDS上样缓冲液按4:1比例混合,并于95 ℃水浴加热5 min。取10 μL处理后的样品上样,初始电泳条件设置为10 mA,待样品进入分离胶后将电流调整为25 mA。电泳结束后,使用考马斯亮蓝染液染色1 h,经脱色液反复脱色4~5次后,最后采用凝胶成像系统进行结果分析。
参考Wang等[28]采用Lowry法对蛋白质溶解度进行测定。取不同样品使用去离子水配制成0.01 mg/mL,在室温下磁力搅拌30 min使其充分溶解,随后于8000×g离心10 min。取上清液进行适当稀释后,以牛血清白蛋白为标准蛋白,采用Lowry法测定蛋白质含量并绘制标准曲线。蛋白质溶解度计算如下。
$ \mathrm{溶解度}(\text{%})=\frac{\text{上清液中蛋白质含量}}{\text{样品中蛋白质含量}}\times 100 $
所有实验均重复3次,数据以均值±标准差表示。实验数据采用SPSS Statistics 26.0软件进行单因素方差分析,以最小显著差异法检验数据间的差异显著性(P<0.05,显著)。采用Origin 2021软件绘制图像,并使用PeakFit 4.12软件对色谱数据进行分峰拟合处理。
粒径可以间接反映蛋白质结构变化的指标,粒度分布可以充分反映样品溶液中粒子的大小和均匀性[29]。相比于未处理的RBP,采用pH驱动法处理的RBP粒径分布曲线明显向小尺寸方向移动,且分布范围变狭窄(图1(A)),蛋白质的平均粒径较对照组显著减小(P<0.05),碱性条件对粒径的减小效果明显优于酸性条件(图1(B))。其中pH调至12时平均粒径(137.1 nm)减小最为明显,这可能是极端pH条件下蛋白质分子间静电排斥力增强,诱导RBP中的大粒径聚集体发生解聚,形成更多小粒径颗粒,从而有效减弱蛋白质分子间的聚集作用。已有研究表明,较小粒径的蛋白质聚集体通常有利于提升体系的溶解特性[30]。
PDI可用于表征聚合物分子量分布的均匀性。经pH处理后,RBP的PDI值较对照组均减小,表明溶液分散性得到改善,蛋白质在溶液体系中的分布更加均匀。这种现象可能是由于酸碱处理过程中RBP产生较强的静电斥力,从而促使其均匀分布[31]。
蛋白质溶液的ζ-电位是表征蛋白质分子间相互作用的重要指标。在水溶液中,当RBP的亲水基团暴露于蛋白质表面,且带负电的氨基酸残基多于带正电的氨基酸残基时,溶液ζ-电位呈现负值。如图1(C)所示,不同pH处理会显著影响RBP的ζ-电位(P<0.05)。与对照组的ζ-电位绝对值(4.7 mV)相比,经pH处理后样品的ζ-电位绝对值均呈现增大趋势,其中碱性处理组的变化尤为显著。当pH调至12时ζ-电位绝对值最大(14.8 mV)。此外,当pH调至1.5的极端酸性条件下,ζ-电位绝对值出现下降现象,这可能是强酸性环境削弱了蛋白质与多肽间的静电相互作用所致。其余pH处理组的RBP ζ-电位绝对值均呈现上升趋势,这可能是pH处理诱导RBP空间结构发生改变,导致蛋白质部分解折叠,使原本埋藏在分子内部的带电基团(特别是带负电的氨基酸残基)暴露于分子表面[32]。这种构象变化不仅增加了蛋白质表面的负电荷量,同时增强了分子间的静电排斥作用,从而有效抑制了蛋白质的聚集行为。这一发现与pH驱动RBP形成更小粒径的结果具有良好的一致性。
表面疏水性(H0)是表征蛋白质分子表面疏水基团数量的重要指标,不仅反映蛋白质的构象特征,还与蛋白质的溶解性能密切相关[33]。采用ANS荧光探针法测定表面疏水性的结果表明(图2),pH驱动处理可显著提高RBP的表面疏水性(P<0.05),其中pH调至12时表面疏水性达到最大值(2289)。这一现象说明在pH驱动过程中,蛋白质分子发生部分展开导致其疏水基团(包括芳香族氨基酸)暴露,从而引起H0值升高[34],表明RBP的分子结构发生了改变。值得注意的是,碱性条件下处理的RBP其H0值显著高于酸性条件,这可能是因为强碱环境能够诱导更多蛋白质聚集体解聚,使蛋白质空间结构发生膨胀和伸展,形成熔融球状构象。在此状态下,蛋白质分子中的疏水肽段及氨基酸残基的疏水侧链得以暴露[35],蛋白质的两亲性平衡发生改变,疏水基团暴露的同时,碱性条件下蛋白质表面负电荷(-COO−)增加,静电斥力增强,反而提升溶解性。这与pH驱动RBP在碱性条件下溶解度优于酸性条件的实验结果一致。因此,RBP表面疏水性的变化可有效反映pH处理对其溶解性能的影响。张唯唯等[21]在研究白果蛋白时也发现,pH驱动法通过调控疏水/亲水平衡改善溶解性。这表明表面疏水性可作为表征pH处理对蛋白质溶解性能影响的有效指标。
蛋白质的内源荧光主要来源于其分子中带有芳香基团的氨基酸残基,该特性可用于表征蛋白质三级结构的变化[36]。如图3所示,对照组的最大荧光发射波长为342 nm。与对照组相比,经不同pH处理的蛋白质荧光光谱形状虽未发生明显改变,但其最大发射波长均出现红移现象,且荧光强度均有不同程度的增强。这种现象可能是pH处理破坏了蛋白质的三维构象,导致蛋白质结构展开,使原本包埋在疏水环境中的色氨酸(Trp)残基暴露于外部亲水环境所致。值得注意的是,碱性条件下荧光强度的增加幅度明显大于酸性条件,这可能是因为在碱性环境中蛋白质更易形成熔球态中间体,使得蛋白质表面疏水性氨基酸间的接触减少,疏水基团更多地暴露出来,导致荧光发射基团向疏水性更强的微环境转移。这与表面疏水性的测定结果(图2)一致,进一步证实了pH驱动法诱导蛋白质结构变化的有效性。
从图4中可以看出,蛋白分子中羟基的伸缩振动峰从3275 cm−1移至3286 cm−1,表明氢键网络发生重排,酰胺Ⅰ带特征峰从1625 cm−1移至1634 cm−1,该区域对蛋白质二级结构分析具有重要价值,因此采用Feakfit 4.2对FTIR光谱的酰胺I带进行二阶导数拟合。根据文献[37],酰胺I带中各特征峰对应的二级结构分别为:α-螺旋(1650~1660 cm−1)、β-折叠(1610~1640 cm−1)、β-转角(1660~1700 cm−1)和无规卷曲(1640~1650 cm−1)。通过分析各特征峰面积占比可定量评估不同二级结构的含量变化,进而反映蛋白质聚集状态和结构稳定性[38]。其中α-螺旋和β-折叠属于有序紧密结构,而无规卷曲和β-转角则属于无序松散结构[39]。由表1可见,对照组RBP的二级结构组成为:α-螺旋(11.44%±0.06%)、β-折叠(17.12%±0.08%)、β-转角(13.46%±0.12%)和无规卷曲(20.55%±0.03%)。经pH处理后,RBP中的α-螺旋含量降低,这与Tian等[40]报道的表面疏水性与α-螺旋含量呈负相关的研究结果一致,也验证了本研究的疏水性测定结果;β-折叠含量增加而β-转角含量降低,这可能是链间氢键断裂导致β-转角向β-折叠转化,同时疏水作用增强引发的分子聚集也会促进β-折叠形成;无规卷曲含量虽略有增加但未达显著水平。上述结果表明,pH处理通过改变蛋白质分子内相互作用力,使其二级结构发生由柔性向刚性的转变。
SDS-PAGE电泳分析可以观察到RBP在不同pH处理下分子质量的变化。如图5所示,由凝胶成像系统仪分析可知RBP主要亚基的相对分子质量分布为43.0~66.2 kDa、31.0 kDa、14.4~22.0 kDa、<14.4 kDa。与对照组(泳道1)相比,其他处理组的RBP相对分子质量分布在>31.0 kDa的条带颜色变浅;<14.4 kDa的条带更清晰,颜色加深,且碱性处理比酸性处理条带更清晰。这可能是在强酸强碱条件下,维持蛋白亚基结构的氢键遭到破坏,使得亚基解聚,在电泳中迁移速度加快,对应高分子质量的条带因亚基含量减少而颜色变浅甚至消失,低分子质量亚基的条带颜色变深甚至出现新条带,且碱处理比酸处理的变化更加明显,与粒径分析结果一致,因此碱处理可以比酸处理更好地促进RBP的亚基解离,增加其溶解度。已有研究发现[41],经pH偏移后蛋白质亚基会解聚为单体亚基。
蛋白质溶解度是反映其变性与聚集程度的关键指标,也是实现RBP加工利用的重要前提[42]。如图6所示,经pH处理的RBP溶解度较未处理组均显著提升(P<0.05),尤其是pH调至12时,溶解度由6.72%显著提高至70.57%。这主要是因为在极端pH条件下,蛋白质分子结构展开,内部疏水基团和极性基团暴露,从而增强了水化作用[43−44]。现有研究表明,RBP溶解度较低可能与其结合的植酸、纤维素等成分有关[45]。此外,碱性条件下RBP的溶解性优于酸性条件,其原因可能在于强碱环境能更有效地降解部分植酸等成分,削弱其与RBP的结合作用,进而提高RBP的溶解性。
本研究以米糠蛋白(RBP)为对象,系统考察了pH驱动法处理对其结构和溶解特性的影响规律。研究结果显示,随着pH升高,RBP的平均粒径显著减小(最低至140.47 nm),ζ-电位绝对值增大(最高达14.8 mV),表面疏水性呈上升趋势(最大值2289)。傅里叶变换红外光谱与荧光光谱分析表明,改性后RBP的二级和三级结构发生明显变化,具体表现为β-折叠相对含量增加,α-螺旋与β-转角相对含量降低,且内源荧光强度增强。在pH12条件下,RBP的溶解度达到70.57%,较未处理组(6.72%)显著提高63.85%。上述结果表明,pH驱动法可有效改善RBP的溶解性,为植物蛋白在功能性食品中的应用提供了一种可行的改性策略。然而,该技术在工业化放大过程中仍面临pH精确控制及营养品质变化等挑战,后续研究应聚焦于开发更温和的改性工艺,并系统评估产物的消化特性等营养指标,以推动该技术的实际应用。

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2026年第47卷第16期
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doi: 10.13386/j.issn1002-0306.2025080137
  • 接收时间:2025-08-13
  • 首发时间:2026-10-24
  • 出版时间:2026-08-15
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  • 收稿日期:2025-08-13
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    1黑龙江八一农垦大学食品学院,黑龙江大庆 163319
    2黑龙江八一农垦大学国家杂粮工程技术中心,黑龙江大庆 163319
    3粮食副产物加工与利用教育部工程研究中心,黑龙江大庆 163319
    4黑龙江省北大荒米业集团有限公司,黑龙江哈尔滨 150090

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左锋(1979−),男,博士,教授,研究方向:植物蛋白及农副产品加工利用,E-mail:。
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2种不同金属材料的力学参数
科
Family
属数
Number of
genus
种数
Number of
species
占总种数比例
Percentage of
total species (%)
属
Genus
种数
Number of
species
占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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