Article(id=1320166627065315695, tenantId=1146029695717560320, journalId=1260987677001138203, issueId=1320166220377215955, articleNumber=null, orderNo=null, doi=10.13386/j.issn1002-0306.2025060036, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=1749052800000, receivedDateStr=2025-06-05, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1792681851401, onlineDateStr=2026-10-22, pubDate=1780243200000, pubDateStr=2026-06-01, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1792681851401, onlineIssueDateStr=2026-10-22, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1792681851401, creator=13701087609, updateTime=1792681851401, updator=13701087609, issue=Issue{id=1320166220377215955, tenantId=1146029695717560320, journalId=1260987677001138203, year='2026', volume='47', issue='11', pageStart='1', pageEnd='514', issueExtLink='null', onlineDate='null', pubDate='1780243200000', pubDateStr='2026-06-01', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=null, createTime=1792681754440, creator='13701087609', updateTime=1792681754440, updator='13701087609', preIssue=null, nextIssue=null, articleTotal=null, ext=null, issueFiles=null, downloadFileDto=null}, startPage=149, endPage=155, ext={EN=ArticleExt(id=1320166628596236656, articleId=1320166627065315695, tenantId=1146029695717560320, journalId=1260987677001138203, language=EN, title=Effects of Ultrasound-assisted Treatment on the Structural and Functional Properties of Rice Protein-Ferulic Acid Complex, columnId=1261336277291548795, journalTitle=Science and Technology of Food Industry, columnName=Research and Investigation, runingTitle=null, highlight=null, articleAbstract=

This study investigated the preparation of rice protein (RP)-ferulic acid (FA) complex via ultrasound-assisted treatment, exploring both covalent and non-covalent interactions. The structure of these complexes was characterized using polyphenol grafting equivalents, sulfhydryl content analysis, UV spectroscopy, fluorescence spectroscopy, and Fourier transform infrared spectroscopy. Furthermore, the solubility, surface hydrophobicity, and antioxidant activity of the complexes were evaluated. The results demonstrated that the ultrasound-assisted covalent complex exhibited the highest polyphenol grafting equivalent, reaching 25.49 μg FA/mg. Complexation with FA reduced the fluorescence intensity of RP, whereas the ultrasound-assisted treatment of the RP-FA complex led to an enhancement in RP fluorescence. Compared to non-ultrasonic treatment, ultrasound processing decreased the relative content of α-helix by 5.46% and 3.41% for covalent and non-covalent complexes, respectively. In addition, protein solubility was enhanced by 25.85% and 8.13% for covalent and non-covalent complexes prepared by ultrasound processing, respectively. Both DPPH and ABTS+ free radical scavenging abilities were significantly improved, with increases of 9.75% and 3.72% for DPPH and 10.45% and 2.06% for ABTS+, respectively, for covalent and non-covalent complexes after ultrasound treatment. Ultrasonic-assisted treatment significantly promoted the complexation of RP and FA, altered the structure of the complexes, and enhanced their functional properties, providing technical references and theoretical support for the application of rice protein-phenol complexes in fields such as food antioxidants and functional ingredients.

, authors=Fang LUO1, 2, Shuyi LI1, *, Zhenzhou ZHU1, Yong SUI2, Jianbin SHI2, Tian XIONG2, Xin MEI2, *, authorsList=Fang LUO, Shuyi LI, Zhenzhou ZHU, Yong SUI, Jianbin SHI, Tian XIONG, Xin MEI, authorCompany=null, correspAuthors=Shuyi LI, Xin MEI, 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=1320166633495183746, articleId=1320166627065315695, tenantId=1146029695717560320, journalId=1260987677001138203, language=CN, title=超声辅助处理对大米蛋白-阿魏酸复合物结构和功能特性的影响, columnId=1261336277849391232, journalTitle=食品工业科技, columnName=研究与探讨, runingTitle=null, highlight=null, articleAbstract=

本文采用超声辅助制备大米蛋白(RP)与阿魏酸(FA)共价/非共价复合物,通过多酚接枝当量、巯基含量及紫外、荧光和傅里叶红外光谱分析表征复合物结构,并评价复合物的溶解度、表面疏水性和抗氧化活性。结果表明:超声辅助共价复合物多酚接枝当量最高,达到25.49 μg FA/mg,与FA复合降低了RP荧光强度,RP与FA的复合物经超声辅助处理后,其荧光强度增强。相比于非超声处理,超声辅助处理RP与FA共价与非共价复合物的α-螺旋相对含量分别降低5.46%和3.41%;蛋白溶解度分别提升25.85%和8.13%;DPPH自由基清除能力分别提高9.75%和3.72%;ABTS+自由基清除能力分别提高10.45%和2.06%。超声辅助可显著促进RP与FA的复合,改变复合物结构并提升其功能特性,为大米蛋白-多酚复合物在食品抗氧化剂、功能基料等领域的应用提供了技术参考和理论支撑。

, authors=罗芳1, 2, 李书艺1, *, 祝振洲1, 隋勇2, 施建斌2, 熊添2, 梅新2, *, authorsList=罗芳, 李书艺, 祝振洲, 隋勇, 施建斌, 熊添, 梅新, authorCompany=null, correspAuthors=李书艺, 梅新, authorNote=

罗芳(2000−),女,硕士研究生,研究方向:食品加工,E-mail:

, correspAuthorsNote=
李书艺(1987−),女,博士,副教授,研究方向:食品安全,E-mail:。
梅新(1978−),男,博士,研究员,研究方向:粮食加工及副产物利用,E-mail:。
, copyrightStatement=版权所有 © 2026《食品工业科技》编辑部, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=kGIc2sW0vqVjktvLyhYg/A==, magXml=pK5PHddr6OlZO8IrfuCFIw==, pdfUrl=null, pdf=Qt3ZO7pIkNIp8jdGcUlHmw==, pdfFileSize=1230046, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=HSMeoYq2KsVb6TPbkEfs2Q==, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=h5eNncx+Q/JSP/WDMVH9Tw==, mapNumber=null, fund=null)}, authors=[Author(id=1320166856367919111, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320166627065315695, orderNo=0, firstName=null, middleName=null, lastName=null, nameCn=null, orcid=null, stid=null, country=null, authorPic=null, dead=0, email=luofang1017@126.com, emailSecond=null, emailThird=null, correspondingAuthor=0, authorType=1, ext={EN=AuthorExt(id=1320166856485359626, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320166627065315695, authorId=1320166856367919111, language=EN, stringName=Fang LUO, firstName=Fang, middleName=null, lastName=LUO, prefix=null, suffix=null, authorComment=null, nameInitials=null, affiliation=null, department=null, xref=1, 2, address=1College of Modern Industry of Selenium Science and Engineering, Wuhan Polytechnic University, Wuhan 430023, China
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注:不同字母表示差异显著(P<0.05),图2、图6~图7同。

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注:不同大写字母表示样品的DPPH自由基清除能力差异显著(P<0.05),不同小写字母表示样品的ABTS+自由基清除能力差异显著(P<0.05)。

, figureFileSmall=WsRdjs+8T1LwRRazc8NF4w==, figureFileBig=2mZGjQleWFDIOoGaeoZGMA==, tableContent=null), ArticleFig(id=1320166861552078916, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320166627065315695, language=EN, label=Table 1, caption=

Relative content of secondary structures in RP-FA complexes

, figureFileSmall=null, figureFileBig=null, tableContent=
样品α-螺旋相对
含量(%)
β-折叠相对
含量(%)
β-转角相对
含量(%)
无规卷曲相对
含量(%)
RP20.99±0.15a29.10±0.46c28.60±0.91a21.31±0.37a
CRP-FA18.85±0.08bc35.94±0.73a25.69±0.30b19.17±0.49a
NRP-FA19.03±0.24b33.61±0.39b25.31±1.61b22.06±1.67a
CRP-FA-US17.82±0.58c35.46±0.49a25.84±0.08b21.95±0.57a
NRP-FA-US18.38±0.33bc35.42±0.13a25.73±0.17b20.01±1.11a
), ArticleFig(id=1320166863213023301, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320166627065315695, language=CN, label=表1, caption=

大米蛋白-阿魏酸复合物的二级结构相对含量

, figureFileSmall=null, figureFileBig=null, tableContent=
样品α-螺旋相对
含量(%)
β-折叠相对
含量(%)
β-转角相对
含量(%)
无规卷曲相对
含量(%)
RP20.99±0.15a29.10±0.46c28.60±0.91a21.31±0.37a
CRP-FA18.85±0.08bc35.94±0.73a25.69±0.30b19.17±0.49a
NRP-FA19.03±0.24b33.61±0.39b25.31±1.61b22.06±1.67a
CRP-FA-US17.82±0.58c35.46±0.49a25.84±0.08b21.95±0.57a
NRP-FA-US18.38±0.33bc35.42±0.13a25.73±0.17b20.01±1.11a
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超声辅助处理对大米蛋白-阿魏酸复合物结构和功能特性的影响
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罗芳 1, 2 , 李书艺 1, * , 祝振洲 1 , 隋勇 2 , 施建斌 2 , 熊添 2 , 梅新 2, *
食品工业科技 | 研究与探讨 2026,47(11): 149-155
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食品工业科技 |研究与探讨 2026 , 47 (11) : 149 -155
超声辅助处理对大米蛋白-阿魏酸复合物结构和功能特性的影响
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罗芳1, 2 , 李书艺1, * , 祝振洲1, 隋勇2, 施建斌2, 熊添2, 梅新2, *
作者信息
  • 1武汉轻工大学硒科学与工程现代产业学院,湖北武汉 430023
  • 2农业农村部农产品冷链物流技术重点实验室,特色农产品资源挖掘与高质利用湖北省重点实验室,湖北省农业科学院农产品加工与核农技术研究所,湖北武汉 430064
通讯作者:
李书艺(1987−),女,博士,副教授,研究方向:食品安全,E-mail:。
梅新(1978−),男,博士,研究员,研究方向:粮食加工及副产物利用,E-mail:。
作者简介:

罗芳(2000−),女,硕士研究生,研究方向:食品加工,E-mail:

Effects of Ultrasound-assisted Treatment on the Structural and Functional Properties of Rice Protein-Ferulic Acid Complex
Fang LUO1, 2 , Shuyi LI1, * , Zhenzhou ZHU1, Yong SUI2, Jianbin SHI2, Tian XIONG2, Xin MEI2, *
Affiliations
  • 1College of Modern Industry of Selenium Science and Engineering, Wuhan Polytechnic University, Wuhan 430023, China
  • 2Key Laboratory of Cold Chain Logistics Technology for Agro-product, Ministry of Agriculture and Rural Affairs, Hubei Key Laboratory of Characteristic Resources and Utilization, Institute of Agro-product Processing and Nuclear Agricultural Technology, Hubei Academy of Agricultural Sciences, Wuhan 430064, China
出版时间: 2026-06-01 doi: 10.13386/j.issn1002-0306.2025060036
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本文采用超声辅助制备大米蛋白(RP)与阿魏酸(FA)共价/非共价复合物,通过多酚接枝当量、巯基含量及紫外、荧光和傅里叶红外光谱分析表征复合物结构,并评价复合物的溶解度、表面疏水性和抗氧化活性。结果表明:超声辅助共价复合物多酚接枝当量最高,达到25.49 μg FA/mg,与FA复合降低了RP荧光强度,RP与FA的复合物经超声辅助处理后,其荧光强度增强。相比于非超声处理,超声辅助处理RP与FA共价与非共价复合物的α-螺旋相对含量分别降低5.46%和3.41%;蛋白溶解度分别提升25.85%和8.13%;DPPH自由基清除能力分别提高9.75%和3.72%;ABTS+自由基清除能力分别提高10.45%和2.06%。超声辅助可显著促进RP与FA的复合,改变复合物结构并提升其功能特性,为大米蛋白-多酚复合物在食品抗氧化剂、功能基料等领域的应用提供了技术参考和理论支撑。

大米蛋白  /  阿魏酸  /  超声处理  /  功能特性  /  结构

This study investigated the preparation of rice protein (RP)-ferulic acid (FA) complex via ultrasound-assisted treatment, exploring both covalent and non-covalent interactions. The structure of these complexes was characterized using polyphenol grafting equivalents, sulfhydryl content analysis, UV spectroscopy, fluorescence spectroscopy, and Fourier transform infrared spectroscopy. Furthermore, the solubility, surface hydrophobicity, and antioxidant activity of the complexes were evaluated. The results demonstrated that the ultrasound-assisted covalent complex exhibited the highest polyphenol grafting equivalent, reaching 25.49 μg FA/mg. Complexation with FA reduced the fluorescence intensity of RP, whereas the ultrasound-assisted treatment of the RP-FA complex led to an enhancement in RP fluorescence. Compared to non-ultrasonic treatment, ultrasound processing decreased the relative content of α-helix by 5.46% and 3.41% for covalent and non-covalent complexes, respectively. In addition, protein solubility was enhanced by 25.85% and 8.13% for covalent and non-covalent complexes prepared by ultrasound processing, respectively. Both DPPH and ABTS+ free radical scavenging abilities were significantly improved, with increases of 9.75% and 3.72% for DPPH and 10.45% and 2.06% for ABTS+, respectively, for covalent and non-covalent complexes after ultrasound treatment. Ultrasonic-assisted treatment significantly promoted the complexation of RP and FA, altered the structure of the complexes, and enhanced their functional properties, providing technical references and theoretical support for the application of rice protein-phenol complexes in fields such as food antioxidants and functional ingredients.

rice protein  /  ferulic acid  /  ultrasound treatment  /  functional properties  /  structure
罗芳, 李书艺, 祝振洲, 隋勇, 施建斌, 熊添, 梅新. 超声辅助处理对大米蛋白-阿魏酸复合物结构和功能特性的影响. 食品工业科技, 2026 , 47 (11) : 149 -155 . DOI: 10.13386/j.issn1002-0306.2025060036
Fang LUO, Shuyi LI, Zhenzhou ZHU, Yong SUI, Jianbin SHI, Tian XIONG, Xin MEI. Effects of Ultrasound-assisted Treatment on the Structural and Functional Properties of Rice Protein-Ferulic Acid Complex[J]. Science and Technology of Food Industry, 2026 , 47 (11) : 149 -155 . DOI: 10.13386/j.issn1002-0306.2025060036
大米蛋白(Rice Protein,RP)是一种优质且无麸质的植物蛋白,必需氨基酸种类齐全,具有较高的营养价值[1]。RP主要由谷蛋白、醇溶蛋白、白蛋白以及球蛋白等组成,其中谷蛋白的含量最高(约占总蛋白含量80%)。研究表明,RP在中性和酸性环境下的溶解度较低,且未经水解RP抗氧化活性较低[2],极大地限制RP在食品工业中的应用范围[3]。因此,改善RP的结构和功能特性,拓宽RP的应用范围具有重要的意义。
蛋白质和多酚主要通过共价和非共价两种方式结合进而诱导蛋白质构象发生改变,并对其溶解性、乳化性及稳定性等功能特性产生影响 [4]。共价相互作用是一种不可逆的相互作用,在碱性条件下,多酚可与蛋白质上的氨基酸残基共价结合,特别是赖氨酸、半胱氨酸和色氨酸残基[5]。非共价主要是蛋白质与小分子之间通过表面疏水力、氢键和范德华力结合[6] 。Jiang等[7]将绿原酸通过非共价相互作用与乳清蛋白及酪蛋白结合,复合物溶解性、起泡性和抗氧化活性明显高于乳清蛋白及酪蛋白;花生蛋白与多酚的共价结合可显著降低花生蛋白的过敏性,并增强其乳化、发泡特性以及溶解度和抗氧化活性[8];Chen等[9]发现大豆蛋白与阿魏酸非共价结合可以有效改善大豆蛋白的界面特性。蛋白质与多酚互作已成为改变蛋白质功能特性的重要手段之一。目前,蛋白-多酚复合物制备方法存在效率低和反应时间长等缺点,限制了这些功能组分的大规模商业化生产和应用[10]。超声辅助可以提高蛋白-多酚结合的反应效率,通过机械剪切、空化效应和热效应使蛋白质空间结构展开,增加蛋白质与多酚分子接触反应的机会,从而增强蛋白质分子与多酚的结合能力,提高蛋白质的抗氧化能力[11]。阿魏酸(Ferulic Acid,FA)具有调节脂质代谢、抗氧化、抗炎以及抑制细胞凋亡等多种生理功效[12]。研究表明,阿魏酸可以与卵清蛋白非共价结合,从而改善蛋白质的结构和发泡性能[13]。目前已有诸多有关多酚复合改性蛋白的研究,但通过超声辅助大米蛋白-多酚复合改善蛋白质功能特性鲜有报道。
基于此,本研究利用超声辅助处理制备大米蛋白-阿魏酸共价/非共价复合物(RP-FA),并采用光谱分析、溶解度和抗氧化活性测定,研究多酚复合对大米蛋白结构和功能特性的影响,为大米蛋白稳态利用及营养健康产品开发提供理论支撑。
大米蛋白 无锡金农生物科技有限公司;阿魏酸 纯度99%、5,5'-二硫代双(2-硝基苯甲酸)(DTNB) 纯度≥99%,上海麦克林生化科技股份有限公司;2,2-联氮-二(3-乙基-苯并噻唑-6-磺酸)二铵盐(ABTS) 色谱纯,上海源叶生物科技有限公司;1,1-二苯基-2-三硝基苯肼(DPPH)(色谱纯)、抗坏血酸(分析纯) 美国Sigma公司;福林酚试剂、碳酸钠、三羧甲基氨基甲烷(Tris)、氯化钠、氯化钾、磷酸氢二钠、磷酸二氢钾 分析纯,国药集团化学试剂有限公司;甘氨酸 纯度99%,德国Biofroxx有限公司。
722N可见分光光度计 上海仪电仪器有限公司;F4700荧光分光光度计 日立分析仪器有限公司;SCIENTZ-ⅡD超声细胞粉碎机 宁波新芝生物科技有限公司;Alpha Ⅱ傅里叶变换红外光谱仪 德国 Bruker 公司;SN-MS-6D多联磁力搅拌器 上海尚仪仪器设备有限公司;UV-2800A紫外可见分光光度计 尤尼柯上海仪器有限公司。
共价复合物制备参考Yan等[14]的方法并稍作修改,取2.0 g RP溶于100 mL去离子水中,常温下磁力搅拌2 h,4 ℃下过夜水合,取100 mL上清液并加入20 mg FA,充分混合后调节pH为9.0,此溶液一部分直接磁力搅拌,得CRP-FA溶液,另一部分超声(450 W,15 min)后磁力搅拌,得CRP-FA-US溶液。非共价复合物制备参考Shi等[15]的方法稍作修改,如共价复合物中方法制备RP与FA混合溶液,调节pH为5.5,一部分直接磁力搅拌,得NRP-FA溶液,一部分超声处理(450 W,15 min)后继续磁力搅拌,得NRP-FA-US溶液。将上述共价和非共价复合物于4 ℃下透析(透析袋截留分子量8000 Da)48 h ,截留物冷冻干燥备用。
复合物中多酚接枝当量测定参考He等[16]方法,取0.5 mL福林酚试剂与0.5 mL浓度为1 mg/mL的复合物溶液充分混合,后加入2 mL浓度为7.5%(w/v)的Na2CO3溶液,混合液于室温静置1 h后记录其760 nm下吸光值。以FA为标准品,制作FA浓度与吸光值的标准曲线,标准曲线为y=8.4778x+0.0456,R2=0.9951,通过标准曲线计算复合物的多酚接枝当量(以μg FA/mg表示)。
蛋白质及复合物中游离巯基测定参考Yan等[17]的方法,将RP、复合物分别溶解于Tris甘氨酸缓冲液中,配制成浓度为0.5 mg/mL的样品溶液。取50 μL DTNB 溶液与4 mL的样品溶液充分混合,并避光贮存1 h后记录其412 nm下的吸光值,蛋白质及复合物中巯基含量计算公式如下:
$ \mathrm{巯基含量}(\text{µ} \mathrm{mol}/\mathrm{g})=\frac{\text{75.53×}{\text{A}}_{\text{412}}\text{×D}}{\text{C}} $
式中:C为样品浓度,mg/mL;D为溶液稀释倍数;75.53为摩尔吸光系数。
制备0.25 mg/mL蛋白质及复合物溶液于220~300 nm范围内以1000 nm/min扫描速度和0.5 nm 的扫描间隔,进行紫外光谱分析。
采用傅里叶变换红外光谱仪(Fourier Transform Infrared Spectrometer,FTIR),蛋白质及复合物在波数4000~500 cm−1、分辨率4 cm−1下测定,时间分辨频谱500 ps,信噪比40000:1,扫描次数32次,采用PeakFit 4.12与Omnic软件分析数据作图[18]。
蛋白质及复合物溶液浓度为1.0 mg/mL,激发波长为280 nm,发射波长为300~500 nm,狭缝宽度为5 nm,扫描速率为6000 nm/min[19]。
表面疏水性分析参照Haskard等[20]描述的方法,取一定量蛋白质或复合物与PBS缓冲溶液(10 mmol/L,pH7.0)混合制备不同浓度样品溶液。取10 μL ANS溶液(8 mmol/L)分别与2 mL不同浓度样品溶液混合,避光放置10 min后检测其荧光强度,制作荧光强度与样品浓度曲线,其初始斜率即为H0。荧光激发和发射波长分别为390 nm和470 nm,狭缝均为5 nm。
取40 mg蛋白质或复合物与100 mL去离子水充分混合,后于6000 r/min下离心30 min,考马斯亮蓝法测定上清液中蛋白质含量[21],溶解度计算公式如下:
$ \text{溶解度}(\text{%})=\frac{{\text{m}}_{\text{1}}}{{\text{m}}_{\text{2}}}\text{×100} $
式中:m1为溶液中大米蛋白含量(mg);m2为大米蛋白总含量(mg)。
DPPH自由基清除活性测定参照Zhang等[22]方法,制备0.2 mmol/L DPPH乙醇溶液。将2 mL DPPH乙醇溶液(0.2 mmol/L)分别与2 mL蛋白质及复合物溶液(1 mg/mL)混合,黑暗中放置30 min后记录517 nm处吸光值。以抗坏血酸绘制标准曲线(y=40.656x−0.2104,R2=0.9984),样品DPPH自由基清除能力以g VC当量/100 g表示,样品清除活性计算公式如下:
$ \text{DPPH自由基清除}\mathrm{率}(\text{%}) =\left(1-\frac{{\mathrm{A}}_{\mathrm{i}}-{\mathrm{A}}_{\mathrm{j}}}{{\mathrm{A}}_{0}}\right)\times 100 $
式中:A0为空白样品的吸光度,Ai为样品的吸光度,Aj为空白试剂的吸光度。
ABTS+自由基清除活性测定参照Re等[23]的方法并略加修改,取2 mL蛋白质或复合物溶液(1 mg/mL)与2 mL ABTS溶液混合,室温避光反应6 min后记录其734 nm处吸光值。以抗坏血酸为对照,制作标准曲线(y=109.2x+18.58,R2=0.9914),计算ABTS+自由基清除能力,以g VC当量/100 g表示,样品清除活性计算公式如下:
$ \text{ABTS}^+\text{自由基清除}\mathrm{率}(\text{%})=\left(1-\frac{\mathrm{A}_{\mathrm{i}}-\mathrm{A}_{\mathrm{j}}}{\mathrm{A}_0}\right)\times100 $
式中:A0为空白样品的吸光度,Ai为样品的吸光度,Aj为空白试剂的吸光度。
所有实验均重复3次,结果采用平均值±标准差表示,用IBM SPASS Statistic 26将实验数据进行正态分布检验,对符合正态分布的数据进行单因素ANOVO检验,P<0.05具有统计学意义,采用 Origin 2019b 32-bit绘图。
如图1所示,RP共价结合多酚接枝当量显著高于非共价结合(P<0.05);相同结合方式下,超声辅助处理可明显提升复合物中多酚接枝当量,CRP-FA、NRP-FA、CRP-FA-US、NRP-FA-US的多酚接枝当量分别为22.81、18.36、25.49、20.45 μg FA/mg。超声辅助共价结合复合物多酚接枝当量较未超声处理的共价复合物提高了11.75%。研究表明,碱处理可破坏蛋白质空间结构,使其暴露出更多的多酚结合位点,从而提高蛋白质与多酚共价结合的接枝当量[24],此外,超声所引发空化和机械效应产生了大量羟基自由基,这些自由基与蛋白质中的氨基或巯基等位点的氢原子发生反应,从而使FA的苯环和蛋白质形成 C-N或C-S共价键,诱导RP和FA形成更稳定的共价复合物[25]。这与Zheng等[26]研究的超声辅助自由基修饰可以提高卵清蛋白-表没食子儿茶素没食子酸酯的接枝效率一致。
复合物的游离巯基测定结果如图2所示,相比RP,复合物中游离巯基含量均显著降低(P<0.05)。相比于非共价复合物,共价复合物中游离巯基含量下降幅度更大,这可能是因为FA中的苯环与蛋白质分子上的氨基或巯基相互作用最终形成C-N或C-S共价键,导致蛋白质中游离巯基含量下降[26]。相同结合方式下,超声辅助的复合物游离巯基含量高于非超声处理;超声处理下,非共价复合物游离巯基含量较高,达1.90 μmol SH/g,这可能是由于复合物形成过程中RP空间结构变化、聚集体解离以及超声处理导致游离巯基暴露量增多[27]。Chen等[28]的研究亦表明,超声辅助蛋白质的多酚修饰过程中,蛋白质空间结构发生变化,导致疏水基团暴露增加游离巯基含量。
紫外光谱可用于评估由酚类物质和蛋白质之间的相互作用引起的蛋白质氨基酸残基微环境的变化[29]。如图3所示,RP在265 nm处的特征吸收峰,与苯丙氨酸、酪氨酸和色氨酸残基有关。相比于RP,复合物的紫外吸收强度均有所提高,且最大吸收波长均出现明显蓝移。共价复合物的紫外吸收强度大于非共价复合物;相同结合方式下,超声辅助的复合物紫外吸收强度更高,这可能是由于FA的亲水性羟基与蛋白质疏水基团间的相互作用,以及超声处理过程中芳香族氨基酸的暴露,提升了微环境的亲水性[26]。
如图4所示,相比于RP,复合物的峰位置和峰强度发生明显变化。复合物在酰胺A带(3600~3100 cm−1)发生红移,尤其是CRP-FA和CRP-FA-US,其酰胺A带红移最为明显,达3271.75 cm−1和3272.61 cm−1,说明多酚接枝和超声处理的双重作用导致蛋白质结构重排和构象变化。Xue等[30]的研究发现,超声处理谷蛋白与多酚混合和偶联会促进酰胺A带的轻微红移,这与本研究结果一致,均证实了超声处理促进了蛋白质-多酚复合物酰胺A带红移。
蛋白质中肽主链的C-O拉伸振动可以提供有关不同二级结构的数据。FTIR光谱在酰胺Ⅰ带(1600~1700 cm−1)通过分峰拟合的二级结构相对含量如表1所示,相比于RP,复合物中α-螺旋和β-转角的相对含量均显著降低,而β-折叠的相对含量显著升高(P<0.05)。相比于非共价复合物,共价复合物的α-螺旋相对含量较低、β-折叠相对含量较高,说明共价复合物二级结构变化较非共价复合物更明显[31]。相同结合方式下,超声辅助处理复合物α-螺旋的相对含量低于非超声处理,CRP-FA-US和NRP-FA-US的α-螺旋的相对含量相较CRP-FA和NRP-FA分别下降了5.46%和3.41%,说明超声会进一步降低复合物α-螺旋的相对含量,这种变化可能是由羟基自由基的产生和超声波的空化效应引起的,这与Zheng等[32]的研究结果一致。
如图5所示,RP表现出最强的荧光强度,最大发射波长在351 nm处,相较于RP,复合物的荧光强度显著降低并发生蓝移(CRP-FA:347 nm、NRP-FA:348 nm、CRP-FA-US:347 nm、NRP-FA-US:348 nm)。共价复合物的荧光强度显著低于非共价复合物,且荧光发射峰蓝移更明显,这可能是由于蛋白质疏水基团参与共价结合,导致暴露于亲水环境中色氨酸残基数量增多,进而降低荧光强度[33]。相同结合方式下,超声处理复合物荧光强度高于非超声处理,这与超声处理产生的空化效应有关,在空化效应的影响下,蛋白质分子内部极性微环境发生改变,致使蛋白质的结构松散,色氨酸残基暴露,荧光强度升高[34]。
H0反映了极性水性环境中蛋白质分子表面疏水基团的数量,与蛋白质的表面功能特性密切相关[35]。如图6所示,相比于RP,复合物H0均显著降低(P<0.05),这可能是由于蛋白质的疏水性氨基酸残基与多酚极性或非极性组分主要通过疏水相互作用结合。其中共价复合物的H0显著低于非共价复合物(P<0.05),这可能是由于FA的酚羟基通过共价键结合,使其与RP的结合率更高,导致表面疏水基团暴露减少[36]。与未超声辅助处理的复合物相比,超声辅助处理的共价复合物的H0进一步降低,这可能归因于蛋白与多酚的结合增加[37]。
如图7所示,复合物溶解度均显著高于RP(P<0.05);共价复合物溶解度显著高于相同处理下非共价复合物;相比于非超声辅助处理复合物,超声辅助处理RP共价与非共价复合物的蛋白溶解度分别提升了25.85%和8.13%,CRP-FA-US溶解度最高,达70.30%。研究表明,超声产生的空化气泡和强剪切作用,导致蛋白质分子解离并诱导其结构展开[38],这种结构变化增强蛋白质与多酚相互作用,复合物中亲水性羟基数量增多,从而提高了复合物的水溶性[7];此外,酚羟基通过相互作用力诱导蛋白质结构发生变化,蛋白质内部亲水、疏水基团同时暴露出来,FA和疏水基团之间的相互作用会促进FA和RP的络合,进而增加了RP的溶解度[39]。
如图8所示,复合物抗氧化活性显著高于RP(P<0.05),相比于RP,CRP-FA、NRP-FA、CRP-FA-US和NRP-FA-US的DPPH自由基清除能力分别提升了71.20%、52.93%、87.88%和58.62%;ABTS+自由基清除能力分别提升了29.53%、29.30%、42.94%和31.97%。相比于非超声辅助处理复合物,超声辅助处理RP共价与非共价复合物的DPPH自由基清除能力分别提高了9.75%和3.72%;ABTS+自由基清除能力分别提高了10.45%和2.06%。超声处理可显著提升RP-FA的抗氧化活性,共价复合物抗氧化活性高于相同处理下非共价复合物,CRP-FA-US的DPPH和ABTS+自由基清除能力均最高,分别达1.81 g VC当量/100 g和0.55 g VC当量/100 g,这可能与超声促进RP和FA的共价结合以及FA结合当量高有关。Geng等[40]采用超声辅助制备大豆分离蛋白与表没食子儿茶素-3-没食子酸酯复合物,发现超声可提高复合物的ABTS+和DPPH自由基清除能力。
本研究通过超声辅助处理制备大米蛋白-阿魏酸复合物,并分析其结构与功能特性,结果表明,超声辅助可明显改变复合物结构,改善其功能特性。超声辅助共价结合复合物多酚接枝当量高达25.49 μg FA/mg,较未超声共价结合复合物(22.81 μg FA/mg)提高了11.75%。相同结合方式下,超声辅助处理复合物α-螺旋的相对含量低于非超声处理,CRP-FA-US和NRP-FA-US的α-螺旋的相对含量相较CRP-FA和NRP-FA分别下降了5.46%和3.41%。相比于RP,复合物的表面疏水性显著降低,溶解度显著升高(P<0.05),超声辅助共价复合物表面疏水性最低,溶解度最高(70.30%)。超声辅助共价结合的复合物抗氧化活性突出,DPPH和ABTS+自由基清除能力分别为1.81 g VC当量/100 g 和0.55 g VC当量/100 g,较RP分别提高87.88%和42.94%。总之,超声作为辅助手段,在赋予非共价/共价大米蛋白-阿魏酸复合物特征空间结构的同时,还提高复合物溶解度,增强其抗氧化活性,为大米蛋白-多酚复合物高效制备以及大米蛋白高效利用提供理论依据。

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2026年第47卷第11期
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doi: 10.13386/j.issn1002-0306.2025060036
  • 接收时间:2025-06-05
  • 首发时间:2026-10-22
  • 出版时间:2026-06-01
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  • 收稿日期:2025-06-05
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    1武汉轻工大学硒科学与工程现代产业学院,湖北武汉 430023
    2农业农村部农产品冷链物流技术重点实验室,特色农产品资源挖掘与高质利用湖北省重点实验室,湖北省农业科学院农产品加工与核农技术研究所,湖北武汉 430064

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李书艺(1987−),女,博士,副教授,研究方向:食品安全,E-mail:。
梅新(1978−),男,博士,研究员,研究方向:粮食加工及副产物利用,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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