糖基化改性對花生蛋白膜性能的影響及其作用機理研究
發(fā)布時間：2018-08-07 14:07
【摘要】：花生蛋白膜是一種綠色環(huán)保的新型材料,但是由于花生蛋白質(zhì)基質(zhì)間缺乏柔韌性,同時又含有大量親水基團,導(dǎo)致蛋白膜具有脆而不耐水的缺點,限制了其在食品包裝中的應(yīng)用。針對以上問題,本研究對花生蛋白進行糖基化改性,并分析了改性對花生蛋白結(jié)構(gòu)的影響,再以改性蛋白為原料制備具有良好性能的花生蛋白膜,同時分析糖基化改性過程中蛋白膜性能的變化規(guī)律,通過蛋白結(jié)構(gòu)與蛋白膜性能之間的相關(guān)關(guān)系,明確糖基化改善花生蛋白膜性能的機理,最后將蛋白膜應(yīng)用在方便面調(diào)料包裝中,探討其在食品包裝應(yīng)用中的可行性。確定了花生蛋白糖基化改性的最佳工藝條件。對比了木糖、甘露糖、乳糖、半乳糖、葡萄糖和蔗糖對花生蛋白膜各種性能的影響,木糖在提高蛋白膜機械性能的同時能改善花生蛋白膜的耐水性,因此篩選出木糖對花生蛋白進行糖基化改性。結(jié)合單因素和正交試驗對改性條件進行優(yōu)化,得到花生蛋白-木糖糖基化的最適改性條件為:木糖濃度10%、p H 9.5、溫度91.5℃、時間95min。此條件下,利用改性蛋白制備的蛋白膜,其拉伸強度(TS)、斷裂延伸率(E)和溶解性(TSM)分別為10.37 MPa、96.47%和35.94%,同時呈現(xiàn)致密、緊湊的網(wǎng)絡(luò)結(jié)構(gòu),顯著優(yōu)于未改性花生蛋白膜。研究了花生蛋白結(jié)構(gòu)在不同木糖濃度、p H、溫度和時間的糖基化改性過程中的變化。SDS-PAGE結(jié)果表明,糖基化改性后,花生蛋白的條帶組成并無變化,但在糖基化改性的作用下,花生蛋白內(nèi)部發(fā)生交聯(lián),使花生球蛋白及伴花生球蛋白Ⅱ的條帶變淺,同時生成分子量較大的(130 k Da)聚合物。接枝度方面,隨著木糖濃度、p H、溫度和時間的增加,花生蛋白接枝度呈上升趨勢,在木糖濃度10%、p H 9、90℃反應(yīng)90 min的條件下,接枝度為10.49%。紅外光譜方面,改性蛋白光譜在1117 cm-l形成新的吸收峰,而糖基化使酰胺I和酰胺Ⅱ的吸收峰強度有所減弱。巰基含量方面,改性前花生蛋白中巰基含量為7.63μmol/g pro,隨著蛋白和木糖之間交聯(lián)程度的增加,蛋白結(jié)構(gòu)得以伸展,內(nèi)部巰基暴露使其含量增加,改性后巰基含量最高達9.72μmol/g pro。暴露在外的巰基在蛋白成膜過程中形成二硫鍵,從而使蛋白膜中巰基含量下降,最低降至1.65μmol/g pro。表面疏水指數(shù)方面,糖基化改性使蛋白表面疏水指數(shù)顯著增加,增加木糖濃度、p H和溫度,分別使表面疏水指數(shù)增加2.2-2.8倍、1.5-3.4倍和1.6-4.2倍�；瘜W作用力方面,改性前花生蛋白中的化學作用力以氫鍵為主,糖基化使蛋白中離子鍵的作用消失,部分氫鍵被破壞,同時由于內(nèi)部疏水基團暴露,使疏水相互作用得到加強。確定了利用木糖糖基化改性蛋白(PPI-X)制備蛋白膜的工藝。在上述最優(yōu)改性條件下對花生蛋白進行改性,并通過噴霧干燥獲得粉末狀改性蛋白,通過分析改性蛋白的溶解溫度及甘油添加量對蛋白膜性能的影響,最終確定蛋白膜制備工藝為PPI-X粉末溶解溫度20℃、甘油添加量為25%,此條件下蛋白膜TS、E和TSM分別為10.37 MPa、90.11%和35.94%。研究了糖基化改性過程對花生蛋白膜性能的影響。溶解性方面,木糖濃度的增加使蛋白膜溶解性顯著下降,木糖濃度達到10%時TSM比木糖含量為1%時下降17.4%;p H 9、溫度90℃時,蛋白膜溶解性最低,僅為33.64%。膨脹性方面,不同改性條件對花生蛋白膜膨脹性均有影響,其中提高p H使膜的膨脹率顯著增加,p H為11時蛋白膜膨脹率高達829.70,分別是p H 3、7和9時的7.7倍、3.5倍和2.0倍。表面疏水性方面,木糖濃度增加使花生蛋白膜的接觸角(CA)呈現(xiàn)先上升后下降的趨勢;改性p H和溫度增加,蛋白膜表面疏水性逐漸增加,p H為11時或溫度為90℃時,CA相對最高,分別為64.2°和68.91°;隨著改性時間延長,蛋白膜表面疏水性整體呈上升趨勢。微觀結(jié)構(gòu)形態(tài)方面,木糖濃度從1%增加至10%,蛋白膜從致密片狀結(jié)構(gòu)向網(wǎng)絡(luò)結(jié)構(gòu)轉(zhuǎn)變;增加p H有利于網(wǎng)絡(luò)結(jié)構(gòu)的形成,p H為9時,蛋白膜網(wǎng)絡(luò)結(jié)構(gòu)清晰可見,但p H高達11時分子間產(chǎn)生的過度交聯(lián),使蛋白網(wǎng)絡(luò)變得緊密;溫度低于50℃時,蛋白變性程度低,蛋白膜呈現(xiàn)致密的片層結(jié)構(gòu),隨著改性溫度上升,蛋白分子結(jié)構(gòu)伸展使蛋白膜呈現(xiàn)網(wǎng)絡(luò)結(jié)構(gòu)。其他性能方面,糖基化改性使蛋白膜顏色加深,同時能提高蛋白膜水蒸氣阻隔性能,但對膜的氧氣透過性和熱穩(wěn)定性影響不顯著。初步探討了糖基化改性改善花生蛋白膜性能的作用機理。對改性過程中花生蛋白結(jié)構(gòu)變化與性能改變進行相關(guān)性分析,發(fā)現(xiàn)接枝度、蛋白巰基含量、表面疏水指數(shù)與蛋白膜的強度和延伸性均呈極顯著的正相關(guān),與TSM則呈極顯著的負相關(guān)。上述結(jié)果表明,蛋白與木糖的接枝,使蛋白結(jié)構(gòu)得以伸展,大量的疏水基團以及巰基暴露,在蛋白成膜過程中分別形成疏水相互作用和二硫鍵,使蛋白膜能承受更大程度的形變,故而強度和延伸性均有所增強。此外,疏水基團的暴露,使蛋白親水性下降,疏水性增加,因此耐水性得到提高。研究了花生蛋白膜在方便面調(diào)料包裝中應(yīng)用的可行性。將花生蛋白膜包裝植物油,植物油保質(zhì)期可達180天以上,而無包裝對照組和塑料包裝的植物油保質(zhì)期較短,均為40天左右。將花生蛋白膜包裝調(diào)料粉,在貯藏60天后水分含量和菌落總數(shù)均符合標準要求。本研究通過糖基化改性顯著改善了花生蛋白膜的機械性能和耐水性,同時明確了糖基化改性改善蛋白膜性能的機理,為花生蛋白膜的實際應(yīng)用提供了理論依據(jù),有利于實現(xiàn)花生蛋白膜在食品包裝中的開發(fā)與利用。
[Abstract]:Peanut protein membrane is a new green material. However, due to the lack of flexibility between the peanut protein matrix and a large number of hydrophilic groups, the peanut protein membrane has the shortcoming of crisp and water resistant, which restricts its application in food packaging. The effect of modification on the structure of peanut protein was made, and the peanut protein membrane with good properties was prepared with modified protein as raw material, and the change law of the performance of the protein membrane during the process of glycosylation was analyzed. The mechanism of glycosylation to improve the performance of peanut protein membrane was determined by the relationship between the protein structure and the performance of the protein membrane. The feasibility of the application of white film in instant noodle dressing was discussed. The optimum technological conditions for the modification of peanut protein glycosylation were determined. The effects of xylose, mannose, lactose, galactose, glucose and sucrose on the performance of peanut protein membrane were compared. The xylose could improve the mechanical properties of the protein membrane while improving the mechanical properties of the protein membrane. The water resistance of peanut protein membrane was obtained. Therefore, the glycosylation of peanut protein was screened out by xylose. The modification conditions were optimized by single factor and orthogonal test. The optimum conditions for the modification of peanut protein xylose glycosylation were: the concentration of xylose concentration 10%, P H 9.5, temperature 91.5 C, time 95min., and the protein prepared by the modified protein The tensile strength (TS), fracture elongation (E) and solubility (TSM) of the membrane were 10.37 MPa, 96.47% and 35.94% respectively, showing a compact and compact network structure, which was significantly better than the unmodified peanut protein membrane. The changes of peanut protein structure in the glycosylated modification process of different xylose concentration, P H, temperature and time were studied. After glycosylation, the band composition of peanut protein has no change, but under the action of glycosylated modification, peanut protein and peanut globulin and peanut globulin II are shallower, and the molecular weight (130 K Da) polymer is produced. With the increase of the graft degree, the increase of the concentration of xylose, P H, temperature and time The graft degree of raw protein is on the rise. In the condition of the concentration of xylose 10%, P H 9,90 C reaction 90 min, the grafting degree is 10.49%. infrared spectrum, the modified protein spectrum forms a new absorption peak at 1117 cm-l, and the glycosylation makes the absorption peak intensity of amide I and amide II weaken. For 7.63 mol/g pro, with the increase of cross-linking degree between protein and xylose, the structure of protein is extended and the content of the internal sulfhydryl group is increased. The sulfhydryl group content of the modified sulfhydryl group is up to 9.72 mu mol/g and the sulfhydryl group is exposed to the protein film to form two sulfur bonds in the process of protein formation, thus the content of sulfhydryl group in the protein membrane is reduced, and the lowest to 1.65 mu mol/. G pro. surface hydrophobicity index, glycosylation modification makes the surface hydrophobicity index of protein increase significantly, increase the concentration of xylose, P H and temperature, make the surface hydrophobicity index increase 2.2-2.8 times, 1.5-3.4 times and 1.6-4.2 times respectively. In chemical force, the chemical force in the pre modified peanut protein is mainly hydrogen bond, and glycosylation makes the ionic bond in the protein. The effect was destroyed and some hydrogen bonds were destroyed, and the hydrophobic interaction was enhanced by the exposure of internal hydrophobic groups. The process of preparing protein membrane by xylose glycosylated protein (PPI-X) was determined. The peanut protein was modified under the above optimal modification conditions, and the powder modified protein was obtained by spray drying. The effect of the dissolved temperature of the modified protein and the amount of glycerol added on the performance of the protein membrane was determined. Finally, the preparation process of the protein membrane was PPI-X powder dissolution temperature 20, and glycerol 25%. The protein membrane TS, E and TSM were 10.37 MPa, 90.11% and 35.94%. respectively. The effect of glycosylated modification on the performance of peanut protein membrane was studied. When the concentration of xylose increased, the solubility of protein membrane decreased significantly. When the concentration of xylose reached 10%, TSM decreased by 17.4% when the content of xylose was 1%; P H 9, at the temperature 90, the solubility of the protein membrane was the lowest, only 33.64%. expansibility, and the different modification conditions had an effect on the expansibility of the peanut protein membrane, in which the increase of P H increased the expansion rate of the membrane significantly. When p H was 11, the swelling rate of protein membrane was up to 829.70, 7.7 times, 3.5 times and 2 times of P H 3,7 and 9, respectively. The increase of surface hydrophobicity made the contact angle (CA) of peanut protein film increase first and then decline; the modified P H and temperature increased, the surface hydrophobicity of protein membrane increased gradually, P H was 11 times or temperature 90 degrees, CA Relative highest, 64.2 and 68.91 degrees, respectively. With the prolongation of the modification time, the surface hydrophobicity of the protein film is increasing. In microstructure morphology, the concentration of xylose increases from 1% to 10%, the protein membrane is transformed from dense sheet structure to the network structure, and the increase of P H is beneficial to the formation of network structure. When p H is 9, the structure of the protein membrane network is clear. When p H is up to 11, the excessive cross-linking between molecules makes the protein network tight; when the temperature is lower than 50, the protein denaturation is low and the protein film presents a compact lamellar structure. With the increase of the temperature, the protein molecular structure extends to the network structure. The other performance, the glycosylation modification makes the protein film color deepened, At the same time, it can improve the water vapor barrier property of the protein membrane, but it has no significant influence on the oxygen permeability and thermal stability of the membrane. The mechanism of the effect of glycosylated modification on the performance of peanut protein membrane is preliminarily discussed. The correlation of the changes of the structure and performance of the peanut protein in the process of modification is analyzed, and the graft degree, the content of the protein sulfhydryl group and the surface of the protein are found. The hydrophobicity index has a very significant positive correlation with the strength and elongation of the protein membrane, which shows a very significant negative correlation with TSM. The results show that the graft of protein and xylose makes the protein structure extend, a large number of hydrophobic groups and sulfhydryl groups are exposed, and the hydrophobic interaction and two sulfur bonds are formed in the process of protein formation, and the protein membrane is made to make the protein membrane. It can bear a greater degree of deformation, so the strength and extensibility are enhanced. In addition, the hydrophobicity of the hydrophobic group decreases and the hydrophobicity increases, so the water resistance is increased. The feasibility of the application of peanut protein membrane in the instant noodle packing is studied. The vegetable oil is packaged with the peanut protein membrane, and the shelf life of the vegetable oil can reach 18. For more than 0 days, the shelf life of unpackaged control group and plastic package was shorter than 40 days. The water content of peanut protein membrane packing powder and the total number of colonies were in line with the standard requirements after 60 days of storage. The mechanical and water resistance of peanut egg white film were improved by glycosylation, and the sugar was clearly improved. The mechanism of improving the performance of the protein membrane by the modified modification provides a theoretical basis for the practical application of the peanut protein membrane, and is beneficial to the development and utilization of the peanut protein membrane in the food packaging.
【學位授予單位】：中國農(nóng)業(yè)科學院
【學位級別】：博士
【學位授予年份】：2015
【分類號】：TS206.4
本文編號：2170263
[Abstract]:Peanut protein membrane is a new green material. However, due to the lack of flexibility between the peanut protein matrix and a large number of hydrophilic groups, the peanut protein membrane has the shortcoming of crisp and water resistant, which restricts its application in food packaging. The effect of modification on the structure of peanut protein was made, and the peanut protein membrane with good properties was prepared with modified protein as raw material, and the change law of the performance of the protein membrane during the process of glycosylation was analyzed. The mechanism of glycosylation to improve the performance of peanut protein membrane was determined by the relationship between the protein structure and the performance of the protein membrane. The feasibility of the application of white film in instant noodle dressing was discussed. The optimum technological conditions for the modification of peanut protein glycosylation were determined. The effects of xylose, mannose, lactose, galactose, glucose and sucrose on the performance of peanut protein membrane were compared. The xylose could improve the mechanical properties of the protein membrane while improving the mechanical properties of the protein membrane. The water resistance of peanut protein membrane was obtained. Therefore, the glycosylation of peanut protein was screened out by xylose. The modification conditions were optimized by single factor and orthogonal test. The optimum conditions for the modification of peanut protein xylose glycosylation were: the concentration of xylose concentration 10%, P H 9.5, temperature 91.5 C, time 95min., and the protein prepared by the modified protein The tensile strength (TS), fracture elongation (E) and solubility (TSM) of the membrane were 10.37 MPa, 96.47% and 35.94% respectively, showing a compact and compact network structure, which was significantly better than the unmodified peanut protein membrane. The changes of peanut protein structure in the glycosylated modification process of different xylose concentration, P H, temperature and time were studied. After glycosylation, the band composition of peanut protein has no change, but under the action of glycosylated modification, peanut protein and peanut globulin and peanut globulin II are shallower, and the molecular weight (130 K Da) polymer is produced. With the increase of the graft degree, the increase of the concentration of xylose, P H, temperature and time The graft degree of raw protein is on the rise. In the condition of the concentration of xylose 10%, P H 9,90 C reaction 90 min, the grafting degree is 10.49%. infrared spectrum, the modified protein spectrum forms a new absorption peak at 1117 cm-l, and the glycosylation makes the absorption peak intensity of amide I and amide II weaken. For 7.63 mol/g pro, with the increase of cross-linking degree between protein and xylose, the structure of protein is extended and the content of the internal sulfhydryl group is increased. The sulfhydryl group content of the modified sulfhydryl group is up to 9.72 mu mol/g and the sulfhydryl group is exposed to the protein film to form two sulfur bonds in the process of protein formation, thus the content of sulfhydryl group in the protein membrane is reduced, and the lowest to 1.65 mu mol/. G pro. surface hydrophobicity index, glycosylation modification makes the surface hydrophobicity index of protein increase significantly, increase the concentration of xylose, P H and temperature, make the surface hydrophobicity index increase 2.2-2.8 times, 1.5-3.4 times and 1.6-4.2 times respectively. In chemical force, the chemical force in the pre modified peanut protein is mainly hydrogen bond, and glycosylation makes the ionic bond in the protein. The effect was destroyed and some hydrogen bonds were destroyed, and the hydrophobic interaction was enhanced by the exposure of internal hydrophobic groups. The process of preparing protein membrane by xylose glycosylated protein (PPI-X) was determined. The peanut protein was modified under the above optimal modification conditions, and the powder modified protein was obtained by spray drying. The effect of the dissolved temperature of the modified protein and the amount of glycerol added on the performance of the protein membrane was determined. Finally, the preparation process of the protein membrane was PPI-X powder dissolution temperature 20, and glycerol 25%. The protein membrane TS, E and TSM were 10.37 MPa, 90.11% and 35.94%. respectively. The effect of glycosylated modification on the performance of peanut protein membrane was studied. When the concentration of xylose increased, the solubility of protein membrane decreased significantly. When the concentration of xylose reached 10%, TSM decreased by 17.4% when the content of xylose was 1%; P H 9, at the temperature 90, the solubility of the protein membrane was the lowest, only 33.64%. expansibility, and the different modification conditions had an effect on the expansibility of the peanut protein membrane, in which the increase of P H increased the expansion rate of the membrane significantly. When p H was 11, the swelling rate of protein membrane was up to 829.70, 7.7 times, 3.5 times and 2 times of P H 3,7 and 9, respectively. The increase of surface hydrophobicity made the contact angle (CA) of peanut protein film increase first and then decline; the modified P H and temperature increased, the surface hydrophobicity of protein membrane increased gradually, P H was 11 times or temperature 90 degrees, CA Relative highest, 64.2 and 68.91 degrees, respectively. With the prolongation of the modification time, the surface hydrophobicity of the protein film is increasing. In microstructure morphology, the concentration of xylose increases from 1% to 10%, the protein membrane is transformed from dense sheet structure to the network structure, and the increase of P H is beneficial to the formation of network structure. When p H is 9, the structure of the protein membrane network is clear. When p H is up to 11, the excessive cross-linking between molecules makes the protein network tight; when the temperature is lower than 50, the protein denaturation is low and the protein film presents a compact lamellar structure. With the increase of the temperature, the protein molecular structure extends to the network structure. The other performance, the glycosylation modification makes the protein film color deepened, At the same time, it can improve the water vapor barrier property of the protein membrane, but it has no significant influence on the oxygen permeability and thermal stability of the membrane. The mechanism of the effect of glycosylated modification on the performance of peanut protein membrane is preliminarily discussed. The correlation of the changes of the structure and performance of the peanut protein in the process of modification is analyzed, and the graft degree, the content of the protein sulfhydryl group and the surface of the protein are found. The hydrophobicity index has a very significant positive correlation with the strength and elongation of the protein membrane, which shows a very significant negative correlation with TSM. The results show that the graft of protein and xylose makes the protein structure extend, a large number of hydrophobic groups and sulfhydryl groups are exposed, and the hydrophobic interaction and two sulfur bonds are formed in the process of protein formation, and the protein membrane is made to make the protein membrane. It can bear a greater degree of deformation, so the strength and extensibility are enhanced. In addition, the hydrophobicity of the hydrophobic group decreases and the hydrophobicity increases, so the water resistance is increased. The feasibility of the application of peanut protein membrane in the instant noodle packing is studied. The vegetable oil is packaged with the peanut protein membrane, and the shelf life of the vegetable oil can reach 18. For more than 0 days, the shelf life of unpackaged control group and plastic package was shorter than 40 days. The water content of peanut protein membrane packing powder and the total number of colonies were in line with the standard requirements after 60 days of storage. The mechanical and water resistance of peanut egg white film were improved by glycosylation, and the sugar was clearly improved. The mechanism of improving the performance of the protein membrane by the modified modification provides a theoretical basis for the practical application of the peanut protein membrane, and is beneficial to the development and utilization of the peanut protein membrane in the food packaging.
【學位授予單位】：中國農(nóng)業(yè)科學院
【學位級別】：博士
【學位授予年份】：2015
【分類號】：TS206.4
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