黃峻榕，王倩，楊麒，方晨璐，劉凱，陸鵬學(xué)

（1.陜西科技大學(xué)食品與生物工程學(xué)院，陜西西安710021；2.西安中糧工程研究設(shè)計(jì)院有限公司，陜西西安710082；3.陜西金中昌信農(nóng)業(yè)科技有限公司，陜西榆林719000）

淀粉分子量的測(cè)定及其與物化性質(zhì)關(guān)系的研究進(jìn)展

黃峻榕1，王倩1，楊麒1，方晨璐1，劉凱2，陸鵬學(xué)3

（1.陜西科技大學(xué)食品與生物工程學(xué)院，陜西西安710021；2.西安中糧工程研究設(shè)計(jì)院有限公司，陜西西安710082；3.陜西金中昌信農(nóng)業(yè)科技有限公司，陜西榆林719000）

綜述淀粉分子量的測(cè)定方法，包括凝膠滲透色譜、非對(duì)稱流場(chǎng)流分離和黏度法。對(duì)不同品種原淀粉的分子量分布，改變淀粉分子量的各類改性方法，以及分子量與物化性質(zhì)的關(guān)系進(jìn)行總結(jié)，并對(duì)今后淀粉分子量的研究方向進(jìn)行展望。

淀粉分子量；凝膠滲透色譜；非對(duì)稱流場(chǎng)流分離；黏度法；物化性質(zhì)

淀粉不僅是人類食物中最重要的能量物質(zhì)來源，也是一種常用工業(yè)原料。淀粉是由兩種結(jié)構(gòu)不同的高聚物組成的混合物，其中，直鏈淀粉的分子量在105g/mol～106g/mol之間，支鏈淀粉的分子量大約為108g/mol[1]。兩種淀粉分子的分子量都是不均一的，即它們是具有多分散性的物質(zhì)，因此實(shí)際測(cè)得的淀粉分子量是統(tǒng)計(jì)平均值。

淀粉分子量有4種表示方法：數(shù)均（Mn）、重均（Mw）、Z均（Mz）和黏均相對(duì)分子量（Mη）。Mn的測(cè)定方法有端基分析法、沸點(diǎn)升高和冰點(diǎn)降低法、蒸汽壓下降法和膜滲透法等；Mw和Mz的測(cè)定方法有光散射技術(shù)和超速離心技術(shù)等；Mη的測(cè)定方法為黏度法。淀粉分子量的多分散性可用多分散系數(shù)d（Mw/Mn或Mz/Mw）來表征，d＞1，d越大分布越寬。分子量是研究淀粉分子結(jié)構(gòu)的重要基礎(chǔ)參數(shù)之一，直接影響淀粉的物化性質(zhì)和用途。

1 不同測(cè)定方法及結(jié)果

1.1 淀粉分子量的測(cè)定方法

目前常用的淀粉分子的分離方法是凝膠滲透色譜、非對(duì)稱流場(chǎng)流分離和黏度法，三者的比較見表1。

表1 淀粉分子量測(cè)定方法的比較[1-4]Table 1 Comparison of determination methods of starch molecular weight

凝膠滲透色譜法（Gel permeation chromatography，GPC）又稱體積排阻色譜法（Size exclusion chromatograph，SEC），是基于分子的尺寸大?。ɑ蛄黧w力學(xué)體積）對(duì)淀粉分子進(jìn)行分離。非對(duì)稱流場(chǎng)流分離法（Asymmetrical flow field flow fractionation，A4F）是根據(jù)作用于樣品的外加場(chǎng)力與樣品的擴(kuò)散力相互作用，不同淀粉分子流速的差異對(duì)淀粉分子進(jìn)行分離。當(dāng)SEC與示差折光檢測(cè)器（Refractive index detectors，RI）串聯(lián)時(shí)，根據(jù)已知分子量標(biāo)準(zhǔn)品和待測(cè)樣品的結(jié)果，可間接獲得淀粉樣品的分子量。當(dāng)SEC或A4F與多角度激光光散射儀（Multi-angle laser light scattering，MALLS）串聯(lián)時(shí)，可以直接測(cè)定淀粉的Mw、Mn、Mz及平均回轉(zhuǎn)半徑（Rz）等數(shù)據(jù)。黏度法即用黏度計(jì)測(cè)量稀溶液的特征黏度值，再根據(jù)Mark-Houwink經(jīng)驗(yàn)式（公式1）確定分子量，該方法也需要已知分子量標(biāo)準(zhǔn)品。

式中：η為特征黏度值；Mη為黏均分子量；K為比例常數(shù)；α是與分子形狀有關(guān)的經(jīng)驗(yàn)參數(shù)。K和α值與溫度、聚合物、溶劑性質(zhì)有關(guān)，也和分子量大小有關(guān)。

無論采用哪種方法測(cè)定淀粉分子量，一個(gè)最重要的前提就是淀粉的完全溶解。90%二甲基亞砜（Dimethyl sulfoxide，DMSO）是目前最普遍使用和公認(rèn)的溶劑。在DMSO中添加低分子量電解質(zhì)（NaNO3或LiBr）可作為助溶劑改善淀粉的溶解效果。欒宏飛[5]研究了大米和玉米淀粉的有效溶解體系，結(jié)果表明，兩種淀粉在50 mmol/L NaNO3/DMSO或50 mmol/L LiBr/DMSO兩種配比的溶劑中溶解指數(shù)接近90%，NaNO3和LiBr對(duì)分散在DMSO中的淀粉都具有良好的增溶效果。Kapelko-Z˙eberska等[6]研究了馬鈴薯原淀粉和老化淀粉在兩種溶劑（DMSO和0.5 mol/L NaOH）中溶解，并利用檸檬酸處理后淀粉分子量的情況，結(jié)果表明，所有在DMSO中溶解的淀粉都比在NaOH中溶解的淀粉顯示較大的分子量；馬鈴薯原淀粉在兩種溶劑中測(cè)定的 Mw分別是 1.66×106g/mol和 1.23×106g/mol。

淀粉的溶解過程需要加熱，You等[7-8]是將普通和蠟質(zhì)大米淀粉分散在氫氧化鈉溶液中于50℃加熱10min后，再加少量蒸餾水并用鹽酸中和，最后置于微波爐中加熱使之完全溶解。而Bello-Pérez等[3，9]是將不同淀粉分散在95%DMSO中持續(xù)攪拌4 d，再加入乙醇沉淀淀粉，通過離心并干燥后，配置成淀粉水溶液置于微波爐中加熱使之完全溶解。

對(duì)于直鏈和支鏈淀粉分子的分離方法，Demiate等[10]、Ma等[11]和Du等[12]是將不同品種的豆類淀粉分散在90%DMSO中沸水浴加熱攪拌1 h，再置于室溫下攪拌24 h后，加入乙醇沉淀淀粉并離心，之后將淀粉顆粒重新溶解在沸水中加熱攪拌1 h，過膜后用高效體積排阻色譜（High-performance size exclusion chromatograph，HPSEC）系統(tǒng)分離。而Tran等[13]采取的方法是將菠蘿蜜種子淀粉乳于96℃水浴鍋中加熱攪拌1 h，過濾除去不溶成分后，用磷酸鹽緩沖液調(diào)整溶液pH為6.3，121℃高壓滅菌1 h，再于96℃下加熱攪拌2 h以分散淀粉分子，之后加入正丁醇，通過離心獲得直鏈淀粉正丁醇絡(luò)合物，殘留在上清液中的支鏈淀粉則通過加入乙醇沉淀再離心獲得。

1.2 淀粉分子量的測(cè)定結(jié)果

表2是幾種常見原淀粉分子量的測(cè)定結(jié)果。

不同品種豆類[10-12]（巴西豆、黑眼豆、鷹嘴豆、綠豆、扁豆、小紅豆、小利馬豆、花蕓豆、紅蕓豆、黑豆和菜豆）支鏈淀粉的Mw和Rz的測(cè)定結(jié)果表明，Mw值越大，對(duì)應(yīng)的Rz值也越大，報(bào)道中溶解淀粉樣品過程、所選擇的流動(dòng)相以及其他試驗(yàn)參數(shù)都基本相同，但出現(xiàn)結(jié)果的差異說明了Mw和Rz與淀粉品種有關(guān)。Bello-Pérez等[3]和You等[7-8]的測(cè)定結(jié)果也證實(shí)了上述的觀點(diǎn)。然而Agama-Acevedo等[14]研究了4種香蕉淀粉的結(jié)構(gòu)特性，結(jié)果卻顯示Mw最小的淀粉有較大的Rz值。這表明Mw和Rz值還受淀粉品種的基因型特征決定。

表2 不同淀粉分子量的測(cè)定結(jié)果Table 2 Determination of molecular weight of different starches

為使淀粉更適合生產(chǎn)應(yīng)用的要求，常利用物理、化學(xué)和生物（或酶）方法處理淀粉。表3、表4和表5分別是改變淀粉分子量的化學(xué)、物理和生物改性方法。

酯化反應(yīng)、乙?；磻?yīng)、酸處理和堿處理都會(huì)導(dǎo)致淀粉降解，且分子量隨著取代度或作用時(shí)間的增加而減小。酸堿處理造成分子量的降低程度比酯化和乙?；〈磻?yīng)大，前者為98.5%～99.9%，后者為37.1%～62.7%。而對(duì)脫支糯米和脫支蠟質(zhì)馬鈴薯淀粉進(jìn)行酯化反應(yīng)，結(jié)果分子量變大，這可能是因?yàn)槊撝У矸坻溤跉滏I與羥基、羥基與羧基之間發(fā)生交聯(lián)，和/或淀粉疏水部分之間的疏水性相互作用。

表3 改變淀粉分子量的化學(xué)改性方法Table 3 Chemical modification methods for changing starch molecular weight

表4 改變淀粉分子量的物理改性方法Table 4 Physical modification methods for changing starch molecular weight

表5 改變淀粉分子量的生物改性方法Table 5 Biological modification methods for changing starch molecular weight

續(xù)表5 改變淀粉分子量的生物改性方法Continue table 5 Biological modification methods for changing starch molecular weight

淀粉經(jīng)超聲波、超高壓、微波、脈沖電場(chǎng)、高速噴氣、線性偏振可見光（一種電磁輻射方式）、濕熱、退火和氧氣輝光等離子體（利用氣體電離產(chǎn)生的等離子體）處理后，分子量同樣隨著作用強(qiáng)度和作用時(shí)間的增加而降低，這是由于上述處理導(dǎo)致淀粉分子鏈的斷裂。超聲波和微波處理對(duì)分子量的降低程度最大（＞91.4%），高速噴氣和氧氣輝光等離子體處理對(duì)分子量的降低程度在64.3%～88.9%之間，超高壓、脈沖電場(chǎng)、線性偏振可見光、濕熱和退火處理的降低程度最?。?0%左右）。另外，氮?dú)夂秃廨x光等離子體處理卻得到相反的結(jié)果，表明這些處理方法誘導(dǎo)淀粉分子間發(fā)生聚合和/或交聯(lián)，導(dǎo)致分子量變大，且處理后造成分子量的增大程度為58.6%～70.2%。

酶作用會(huì)引起淀粉分子量減小，其中水解酶、脫支酶和普魯蘭酶對(duì)其降低作用最大，酶量和反應(yīng)時(shí)間越大，分子量越小，變化程度＞97.8%。糙米和小麥淀粉在預(yù)發(fā)芽和發(fā)芽過程中分子量的降低程度只有20%～36%左右，而大麥在發(fā)芽過程中（0～48 h）分子量逐漸增加，這表明發(fā)芽機(jī)制取決于谷物的植物來源。

2 淀粉分子量與物化性質(zhì)的關(guān)系

目前，對(duì)于淀粉分子量與物化性質(zhì)關(guān)系的研究已有較多報(bào)道。Zortéa-Guidolin等[53]研究表明巴西松子支鏈淀粉的Mw和Rz與糊化溫度和崩解值呈顯著負(fù)相關(guān)。Kowittaya等[54]的研究結(jié)果表明，大米支鏈淀粉的分子量與特征黏度值、糊化溫度、谷值黏度、最終黏度、回生值和溶解度呈顯著負(fù)相關(guān)，與峰值黏度、崩解值、膨脹能力和透光率呈正相關(guān)。而Park等[55]的研究表明大米支鏈淀粉的長(zhǎng)鏈分子量與直鏈淀粉含量和峰值溫度呈正相關(guān)，與峰值黏度呈負(fù)相關(guān)。以上報(bào)道中分子量與崩解值和峰值黏度的關(guān)系出現(xiàn)了不同的結(jié)果，可能與淀粉的品種有關(guān)。

對(duì)于凝膠質(zhì)構(gòu)和流變學(xué)特性，劉佳等[56]研究了小麥A、B型淀粉的凝膠質(zhì)構(gòu)特性與分子結(jié)構(gòu)的關(guān)系，結(jié)果發(fā)現(xiàn)A型淀粉更易形成有序結(jié)構(gòu)，使其凝膠硬度增強(qiáng)，B型淀粉更易形成網(wǎng)狀結(jié)構(gòu)，有利于凝膠的彈性及內(nèi)聚性增強(qiáng)。Sikora等[57]和Krystyjan等[58]根據(jù)不同濃度和糊化程度，分別研究普通和蠟質(zhì)馬鈴薯淀粉糊的流變學(xué)特性，得出普通馬鈴薯淀粉的分子量下降了近一半，且隨著糊化溫度升高顆粒的損傷程度增加，所有淀粉糊均屬于非牛頓流體，經(jīng)剪切變稀，流變性不穩(wěn)定；但對(duì)于蠟質(zhì)馬鈴薯淀粉，分子量的增加可能是由于已經(jīng)開始糊化時(shí)小分子淀粉葡聚糖聚集造成的。糊化溫度顯著影響糊狀物的流變學(xué)參數(shù)，使其具有觸變性，抗觸變性或混合觸變性/抗觸變性。

綜上所述，分子量是研究淀粉分子結(jié)構(gòu)的重要基礎(chǔ)參數(shù)之一，它直接影響淀粉的糊化溫度、特征黏度、玻璃化轉(zhuǎn)變溫度、凝膠質(zhì)構(gòu)特性和流變學(xué)特性等物化性質(zhì)，影響著淀粉的深加工及用途。

3 結(jié)語(yǔ)

凝膠滲透色譜、多角度激光光散射儀和示差折光檢測(cè)器串聯(lián)的方法是目前測(cè)定淀粉分子量最廣泛采用的方法，但由于測(cè)定過程中分析條件的不同，以及溶解、分離、檢測(cè)和數(shù)據(jù)處理等方面仍存在一些問題和困境（如淀粉樣品的溶解不完全，淀粉分子的降解或聚集，溶劑與分離和檢測(cè)儀器的不完全匹配，非淀粉成分的去除不完全等）使測(cè)定結(jié)果的準(zhǔn)確性和重現(xiàn)性不佳，進(jìn)而Mw和Rz的結(jié)果也可能受到用于淀粉分散，溶解、分離和數(shù)據(jù)擬合模型方法的影響，并影響到分子量與物化性質(zhì)相關(guān)性的研究。因此，精確測(cè)定淀粉分子質(zhì)量的方法有待進(jìn)一步研究與改進(jìn)。

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Research Progress on Determination of Molecular Weight and the Relationship with Physicochemical Properties of Starch

HUANG Jun-rong1，WANG Qian1，YANG Qi1，F(xiàn)ANG Chen-lu1，LIU Kai2，LU Peng-xue3
（1.School of Food and Biological Engineering，Shaanxi University of Science and Technology，Xi'an 710021，Shaanxi，China；2.Xi'an COFCO Engineering Research and Design Institute Co.Ltd.，Xi'an 710082，Shaanxi，China；3.Shaanxi Jinzhong Changxin Agricultural Science and Technology Co.Ltd.，Yulin 719000，Shaanxi，China）

The determination methods of starch molecular weight，including gel permeation chromatography，asymmetrical flow field flow fractionation and viscosity methods were reviewed.The molecular weight distribution of different native starches，the modification methods of starch molecular weight，and the relationship between molecular weight and physicochemical properties were summarized.Future research directions on molecular weight of starch were prospected.

starchmolecularweight；gelpermeationchromatography；asymmetricalflowfieldflowfractionation；viscosity method；physicochemical property

黃峻榕，王倩，楊麒，等.淀粉分子量的測(cè)定及其與物化性質(zhì)關(guān)系的研究進(jìn)展[J].食品研究與開發(fā)，2018，39（1）：182-188

HUANG Junrong，WANG Qian，YANG Qi，et al.Research Progress on Determination of Molecular Weight and the Relationship with Physicochemical Properties of Starch[J].Food Research and Development，2018，39（1）：182-188

10.3969/j.issn.1005-6521.2018.01.036

國(guó)家自然科學(xué)基金項(xiàng)目（31371786）；陜西省科技統(tǒng)籌創(chuàng)新工程計(jì)劃項(xiàng)目（2016KTCL02-23）

黃峻榕（1971—），女（漢），教授，博士，研究方向：淀粉結(jié)構(gòu)研究。

2017-10-16