抗高直链大米淀粉回生的物理修饰及其回生的检测和表征
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摘要
淀粉回生,即淀粉糊化后贮存过程中发生的自身结合,严重影响淀粉类食品的品质,是食品工业中一个尚未得到解决的技术难题。本文以易回生的高直链大米淀粉(HA-RS)为研究对象,在较易回生的贮存温度(4℃)和水分含量(66.7%)下,研究不同新型添加物和物性技术处理抑制淀粉回生的作用。同时,应用快速分子振动光谱技术(主要为红外和拉曼)研究了检测淀粉回生程度的可能性,应用分形理论研究了表征淀粉回生过程的分形生长。主要研究内容和结论如下:
研究了天然抗氧化提取物茶多酚(TPLs)、越橘提取物(BE)和葡萄籽提取物(GSE)对HA-RS回生的影响。采用差示扫描量热仪(DSC)、X-射线衍射仪(XRD)和扫描电镜(SEM)评价茶多酚对淀粉回生的抑制作用。结果表明,淀粉的糊化温度和焓值随着TPLs添加量的增加而明显降低,添加16%TPLs(基于淀粉重)淀粉样品的糊化To、Tp和Tc分别提前8.93、5.69和5.13℃,糊化焓值则降低2.27J/g;在4℃下贮存,淀粉的回生焓值和重结晶随着TPLs添加量的增加逐渐降低,添加16%TPLs(基于淀粉重)的糊化淀粉样品贮存15d后没有出现回生焓值以及重结晶。在SEM相同放大倍数下回生淀粉颗粒形貌显示,随着TPLs添加量的增加颗粒逐渐减小并呈网孔状。快速粘度仪(RVA)、1H-NMR分析以及傅立叶红外(FT-IR)测定证明,TPLs能与淀粉分子在糊化过程中能发生氢键相互作用,因此可推测正是由于该相互作用干扰了在贮存过程中糊化淀粉分子间的自身结合——即回生。越橘提取物(BE)和葡萄籽提取物(GSE)也具有抗HA-RS回生的能力,其中BE的效果好于GSE。与目前公认的抗淀粉回生效果最好的添加剂海藻糖相比,TPLs、BE(越橘提取物)、GSE(葡萄籽提取物)抑制HA-RS回生的能力均好于海藻糖,顺序为TPLs > BE > GSE >海藻糖。
研究了壳聚糖的降解方法及其产物抑制HA-RS回生能力。分别采用酶水解法和H2O2降解法制备壳低聚糖(COS)。FT-IR检测发现,酶法COS样品均含有活性氨基基团,而6mL/30min H2O2滴加量制备的COS样品中活性氨基基团被破坏。在4℃贮存7d条件下添加8%COS(基于淀粉重)糊化淀粉样品的抑制能力,5k < COSM < 10k明显,10k < COSM < 30k次之,COSM < 5k不显著。其中,5k < COSM < 10k的回生率降低14.5%,结晶率仅为空白样品的58.52%。这说明COS分子大小可影响淀粉回生的效果。6mL/30min H2O2降解法制备的COS均没有抑制HA-RS回生的能力,主要是由于COS结构发生变化,活性氨基被大量破坏。因此,COS的分子大小和活性氨基存在与否决定了其抗回生能力。
研究了“多次瞬时加热-搅拌”物性技术对高直链米粉(HA-RF)和HA-RS回生的影响。结果表明,通过20s的加热(100℃)-搅拌(500 r/min)的三次瞬时加热-搅拌处理能明显推迟HA-RF的回生。与空白样品(5.93J/g dry flour)相比,三次瞬时加热-搅拌处理的HA-RF显示最低的回生焓值(3.04J/g dry flour)和最小的重结晶(占空白52.32%)。SEM显示三次瞬时加热-搅拌处理可增加蜂窝状结构和减少结晶形貌。糊化后样品切片的SEM发现,三次瞬时加热-搅拌处理可增大网络结构的网孔,说明此种方法可使大量聚集水存在于网络结构。但这种物性技术对HA-RS的回生不产生影响。HA-RF和HA-RS处理后的不同结果说明,米粉的“多次瞬时加热-搅拌”物性技术可赋予米蛋白的抗回生功能。
研究了傅立叶红外(FT-IR)和激光共聚焦显微拉曼(Laser Confocal Micro-Raman,LCM-Raman)分子振动光谱快速检测HA-RS回生程度的可能性。结果表明,红外光谱中一些振动模式(包括3415cm-1附近的O-H伸缩振动,2927.46cm-1的CH2伸缩振动,1155.17cm-1处C-O和C-C伸缩振动,1081.89cm-1的C-O-H弯曲振动,1043.32cm-1、1020.18cm-1和998.96cm-1未确定的振动模式,609.41cm-1和578.55cm-1的吡喃环骨架振动模式)和拉曼光谱中一些振动模式(包括1122.37cm-1处的C-O伸缩和C-O-H弯曲振动,1082.83cm-1的C-O-H弯曲振动,1050.05cm-1的C-C伸缩振动,854.31cm-1的C-H变形振动,440.66cm-1处的吡喃环骨架振动)的相对强度均随淀粉回生程度的增加呈现降低的规律。这些振动模式的相对强度的倒数与DSC回生焓值的相关系数达到0.9以上的振动峰包括FT-IR的1081.89cm-1、1043.32cm-1、1020.18cm-1、609.41cm-1、578.55cm-1处吸收峰和LCM-Raman的854.31cm-1、1082.83cm-1、1050.05cm-1处光谱峰。因此,可以将这些振动模式作为定量检测淀粉回生程度的指标,其中LCM-Raman的灵敏性要高于FT-IR。
研究了基于计算机图像处理技术的分形理论用于表征淀粉回生过程的可能性。贮存5、10、15d的回生淀粉样品中心切面SEM显示,回生淀粉外观形貌具有自相似性,表现出分形特征。专业分形软件分析证明,不同回生程度的淀粉样品分形行为显著。贮存5、10、15d的回生淀粉样品分形维数分别为1.8267、1.7803、1.7333。通过分析计算,淀粉回生程度(回生焓值)与分形维数呈线性关系,线性方程为y = 0.0358x + 1.5934。利用团簇-团簇聚集模型(Cluster-Cluster aggregation,CCA模型)可模拟淀粉回生分形生长过程,模拟结果与实验结果的形态和分形维数均符合。此模型可用来表征淀粉回生的生长机制。
The retrogradation of starch is self-association during storage after gelatinization, it can markedly effect on quality of starchy foods. So it is an important and unresolved technology problem in food industry. To study retrogradation, gelatinized high amylose rice starch (HA-RS) with 66.7% w/w water content was stored at 4 oC different days as model. It was investigated that the effect of new additive and physical technology on preventing the retrogradation. Then rapid molecule vibration spectrum was used to monitor the retrogradation process and fractal theory was used to characterize fractal growth of this process. The major content and results were as following:
The effect of natural antioxidative extracts including tea polyphenols (TPLs), blueberry extract (BE) and grape seed extract (GSE) on the retrogradation of HA-RS were investigated. TPLs-fortified HA-RS exhibited retarding the retrogradation as assessed by differential scanning calorimetry (DSC), X-ray diffraction (XRD) and scanning electron microscopy (SEM). The temperature and enthalpy of starch gelatinization obviously decreased as the TPLs level increased. Gelatinization To, Tp and Tc of starch with 16%TPLs sample were respectively 8.93, 5.69 and 5.13 oC lower and enthalpy of gelatinization reduced by 2.27J/g. After storage at 4 oC, enthalpy and recrystallization of starch retrogradation gradually decreased with the increase of TPLs content. HA-RS with 16% TPLs had almost no retrogradation enthalpy and recrystallization until storage of 15 days. It was observed at same magnification by SEM that the size of retrograded starch granules decreased and appeared cancellous shape with adding TPLs increased. The overall results demonstrate that the marked inhibitory effect of TPLs on the retrogradation of HA-RS. Subsequently, it found that high active TPLs could interact with HA-RS to form hydrogen band through Rapid Visco Analyzer (RVA), 1H-NMR and FT-IR analyses, so this interaction might disturb self-association of starch retrogradation during storage. In addition, two other natural antioxidant extracts-blueberry extract (BE) and grape seed extract (GSE) also behaved the retarding the retrogradation of HA-RS, and BE has a better effect than GSE. The effect on preventing the retrogradation of TPLs, BE and GSE were all better than trehalose which is publicly known anti-retrogradation additive. The sequence of capability was TPLs > BE > GSE > trehalose.
It was studied that degradation methods of chitosan and the effect of product on preventing the HA-RS retrogradation. Chito-oligosaccharides (COS) were prepared using enzyme hydrolysis and hydrogen peroxide (H2O2) degradation, respectively. IR analysis proved that COS prepared by enzyme hydrolysis all had active amino-group. But this group of all the fractions of COS prepared by 6mL/30min H2O2 degradation was damaged. Subsequently, the effect of 8% COS (based on starch weight) on the retrogradation of HA-RS after storage for seven days at 4 oC was investigated. It found that the fraction of 5k < COSM < 10k had the best ability to prevent the retrogradation. Moreover, retrogradation ratio of HA-RS with this fraction was reduced by 14.5% as compared with the control and recrystallization ratio was only 58.52% of the control. Then, the inhibition capability of the fraction of 10k < COSM < 30k was followed. But the fraction of COSM < 5k has no this effect.
Therefore, it stated that the size of molecular weight of COS decided the effect on preventing the retrogradation. But all the fractions of COS prepared by 6mL/30min H2O2 degradation had no effect on retarding the retrogradation due to changes in the structure. Therefore, molecular size and existence of active amino-group of COS determined the capability of anti-retrogradation.
Multiple instant heating -stirring preventing the retrogradation of high amylose rice flour (HA-RF) and HA-RS were investigated. The results indicate that three instant heating (100 oC)-stirring (500 r/min) had a clear effect on retarding the retrogradation of HA-RF. Treated HA-RF by three instant heating-stirring exhibited the lowest retrogradation enthalpy (3.04J/g dry flour) as compared to the control (5.93J/g dry flour) in DSC analysis and had almost the least recrystallization (52.32% of the control) by XRD. The observation by SEM stated that treated HA-RF by this way granules had a more honeycomb-like structure and reduced the crystal morphologies. The gelatinized HA-RF by three instant heating-stirring treatments enlarged mesh of network structure by SEM section observation, it implied that network structure had more gathered water by this method treatment. But this physical technology had no effect on the retrogradation of isolated HA-RS. The different results of treated HA-RF and HA-RS by this physical technology might be due to rice protein having anti-retrogradation function after treatment.
It was investigated that Fourier transform infrared (FT-IR) and Laser Confocal Micro-Raman (LCM-Raman) spectroscopy for rapid monitoring the retrogradation process of HA-RS. The results indicate the relative intensity of some vibrational modes reduced regularly with increasing extent of the retrogradation including O-H (3415cm-1) stretching, C-H (2927.46cm-1) stretching, C-O and C-C (1155.17cm-1) stretching, C-O-H (1081.89cm-1) bending, undetermined (1043.32cm-1, 1020.18cm-1 and 998.96cm-1) modes, skeletal modes of pyranose ring (609.41cm-1 and 578.55cm-1) in FT-IR spectra and C-O stretching and C-O-H bending (1122.37cm-1), C-O-H (1082.83cm-1) bending, C-C (1050.05cm-1) stretching, C-H (854.31cm-1) deformation, skeletal modes of pyranose ring (440.66cm-1) in Raman spectra. By the correlation analysis between retrogradation enthalpy of DSC and the relative intensity reciprocal of above vibrational modes, these peaks of 1081.89cm-1, 1043.32cm-1, 1020.18cm-1, 609.41cm-1, 578.55cm-1of FT-IR and 854.31cm-1, 1082.83cm-1, 1050.05cm-1 of LCM-Raman had correlation coefficient above 0.9, so these vibrational modes listed above may be as index to quantify the retrogradation. Meantime, the sensitivity of LCM-Raman was higher than FT-IR.
It was studied that the probability of fractal theory to characterize process of starch retrogradation. SEM was adopted to observe section feature of retrograded HA-RS stored at 5, 10 and 15d. The surface topography of retrograded starch had self-similarity and manifested fractal characteristics. Based on professional fractal software analysis, fractal behavior of various retrograded starch was remarkable. The fractal dimensions of retrograded samples stored at 15d, 10d and 5d were respectively 1.8267, 1.7803, and 1.7333. The extent of retrogradation (retrogradation enthalpy) and the fractal dimension had a linear relationship, linear equation was y = 0.0358x + 1.5934. Cluster-Cluster aggregation model was used to simulate the fractal growth process of starch retrogradation. Simulation results resembled the feature and fractal dimension of the experimental results, so this model can be characterize the growth mechanism of starch retrogradation.
研究了天然抗氧化提取物茶多酚(TPLs)、越橘提取物(BE)和葡萄籽提取物(GSE)对HA-RS回生的影响。采用差示扫描量热仪(DSC)、X-射线衍射仪(XRD)和扫描电镜(SEM)评价茶多酚对淀粉回生的抑制作用。结果表明,淀粉的糊化温度和焓值随着TPLs添加量的增加而明显降低,添加16%TPLs(基于淀粉重)淀粉样品的糊化To、Tp和Tc分别提前8.93、5.69和5.13℃,糊化焓值则降低2.27J/g;在4℃下贮存,淀粉的回生焓值和重结晶随着TPLs添加量的增加逐渐降低,添加16%TPLs(基于淀粉重)的糊化淀粉样品贮存15d后没有出现回生焓值以及重结晶。在SEM相同放大倍数下回生淀粉颗粒形貌显示,随着TPLs添加量的增加颗粒逐渐减小并呈网孔状。快速粘度仪(RVA)、1H-NMR分析以及傅立叶红外(FT-IR)测定证明,TPLs能与淀粉分子在糊化过程中能发生氢键相互作用,因此可推测正是由于该相互作用干扰了在贮存过程中糊化淀粉分子间的自身结合——即回生。越橘提取物(BE)和葡萄籽提取物(GSE)也具有抗HA-RS回生的能力,其中BE的效果好于GSE。与目前公认的抗淀粉回生效果最好的添加剂海藻糖相比,TPLs、BE(越橘提取物)、GSE(葡萄籽提取物)抑制HA-RS回生的能力均好于海藻糖,顺序为TPLs > BE > GSE >海藻糖。
研究了壳聚糖的降解方法及其产物抑制HA-RS回生能力。分别采用酶水解法和H2O2降解法制备壳低聚糖(COS)。FT-IR检测发现,酶法COS样品均含有活性氨基基团,而6mL/30min H2O2滴加量制备的COS样品中活性氨基基团被破坏。在4℃贮存7d条件下添加8%COS(基于淀粉重)糊化淀粉样品的抑制能力,5k < COSM < 10k明显,10k < COSM < 30k次之,COSM < 5k不显著。其中,5k < COSM < 10k的回生率降低14.5%,结晶率仅为空白样品的58.52%。这说明COS分子大小可影响淀粉回生的效果。6mL/30min H2O2降解法制备的COS均没有抑制HA-RS回生的能力,主要是由于COS结构发生变化,活性氨基被大量破坏。因此,COS的分子大小和活性氨基存在与否决定了其抗回生能力。
研究了“多次瞬时加热-搅拌”物性技术对高直链米粉(HA-RF)和HA-RS回生的影响。结果表明,通过20s的加热(100℃)-搅拌(500 r/min)的三次瞬时加热-搅拌处理能明显推迟HA-RF的回生。与空白样品(5.93J/g dry flour)相比,三次瞬时加热-搅拌处理的HA-RF显示最低的回生焓值(3.04J/g dry flour)和最小的重结晶(占空白52.32%)。SEM显示三次瞬时加热-搅拌处理可增加蜂窝状结构和减少结晶形貌。糊化后样品切片的SEM发现,三次瞬时加热-搅拌处理可增大网络结构的网孔,说明此种方法可使大量聚集水存在于网络结构。但这种物性技术对HA-RS的回生不产生影响。HA-RF和HA-RS处理后的不同结果说明,米粉的“多次瞬时加热-搅拌”物性技术可赋予米蛋白的抗回生功能。
研究了傅立叶红外(FT-IR)和激光共聚焦显微拉曼(Laser Confocal Micro-Raman,LCM-Raman)分子振动光谱快速检测HA-RS回生程度的可能性。结果表明,红外光谱中一些振动模式(包括3415cm-1附近的O-H伸缩振动,2927.46cm-1的CH2伸缩振动,1155.17cm-1处C-O和C-C伸缩振动,1081.89cm-1的C-O-H弯曲振动,1043.32cm-1、1020.18cm-1和998.96cm-1未确定的振动模式,609.41cm-1和578.55cm-1的吡喃环骨架振动模式)和拉曼光谱中一些振动模式(包括1122.37cm-1处的C-O伸缩和C-O-H弯曲振动,1082.83cm-1的C-O-H弯曲振动,1050.05cm-1的C-C伸缩振动,854.31cm-1的C-H变形振动,440.66cm-1处的吡喃环骨架振动)的相对强度均随淀粉回生程度的增加呈现降低的规律。这些振动模式的相对强度的倒数与DSC回生焓值的相关系数达到0.9以上的振动峰包括FT-IR的1081.89cm-1、1043.32cm-1、1020.18cm-1、609.41cm-1、578.55cm-1处吸收峰和LCM-Raman的854.31cm-1、1082.83cm-1、1050.05cm-1处光谱峰。因此,可以将这些振动模式作为定量检测淀粉回生程度的指标,其中LCM-Raman的灵敏性要高于FT-IR。
研究了基于计算机图像处理技术的分形理论用于表征淀粉回生过程的可能性。贮存5、10、15d的回生淀粉样品中心切面SEM显示,回生淀粉外观形貌具有自相似性,表现出分形特征。专业分形软件分析证明,不同回生程度的淀粉样品分形行为显著。贮存5、10、15d的回生淀粉样品分形维数分别为1.8267、1.7803、1.7333。通过分析计算,淀粉回生程度(回生焓值)与分形维数呈线性关系,线性方程为y = 0.0358x + 1.5934。利用团簇-团簇聚集模型(Cluster-Cluster aggregation,CCA模型)可模拟淀粉回生分形生长过程,模拟结果与实验结果的形态和分形维数均符合。此模型可用来表征淀粉回生的生长机制。
The retrogradation of starch is self-association during storage after gelatinization, it can markedly effect on quality of starchy foods. So it is an important and unresolved technology problem in food industry. To study retrogradation, gelatinized high amylose rice starch (HA-RS) with 66.7% w/w water content was stored at 4 oC different days as model. It was investigated that the effect of new additive and physical technology on preventing the retrogradation. Then rapid molecule vibration spectrum was used to monitor the retrogradation process and fractal theory was used to characterize fractal growth of this process. The major content and results were as following:
The effect of natural antioxidative extracts including tea polyphenols (TPLs), blueberry extract (BE) and grape seed extract (GSE) on the retrogradation of HA-RS were investigated. TPLs-fortified HA-RS exhibited retarding the retrogradation as assessed by differential scanning calorimetry (DSC), X-ray diffraction (XRD) and scanning electron microscopy (SEM). The temperature and enthalpy of starch gelatinization obviously decreased as the TPLs level increased. Gelatinization To, Tp and Tc of starch with 16%TPLs sample were respectively 8.93, 5.69 and 5.13 oC lower and enthalpy of gelatinization reduced by 2.27J/g. After storage at 4 oC, enthalpy and recrystallization of starch retrogradation gradually decreased with the increase of TPLs content. HA-RS with 16% TPLs had almost no retrogradation enthalpy and recrystallization until storage of 15 days. It was observed at same magnification by SEM that the size of retrograded starch granules decreased and appeared cancellous shape with adding TPLs increased. The overall results demonstrate that the marked inhibitory effect of TPLs on the retrogradation of HA-RS. Subsequently, it found that high active TPLs could interact with HA-RS to form hydrogen band through Rapid Visco Analyzer (RVA), 1H-NMR and FT-IR analyses, so this interaction might disturb self-association of starch retrogradation during storage. In addition, two other natural antioxidant extracts-blueberry extract (BE) and grape seed extract (GSE) also behaved the retarding the retrogradation of HA-RS, and BE has a better effect than GSE. The effect on preventing the retrogradation of TPLs, BE and GSE were all better than trehalose which is publicly known anti-retrogradation additive. The sequence of capability was TPLs > BE > GSE > trehalose.
It was studied that degradation methods of chitosan and the effect of product on preventing the HA-RS retrogradation. Chito-oligosaccharides (COS) were prepared using enzyme hydrolysis and hydrogen peroxide (H2O2) degradation, respectively. IR analysis proved that COS prepared by enzyme hydrolysis all had active amino-group. But this group of all the fractions of COS prepared by 6mL/30min H2O2 degradation was damaged. Subsequently, the effect of 8% COS (based on starch weight) on the retrogradation of HA-RS after storage for seven days at 4 oC was investigated. It found that the fraction of 5k < COSM < 10k had the best ability to prevent the retrogradation. Moreover, retrogradation ratio of HA-RS with this fraction was reduced by 14.5% as compared with the control and recrystallization ratio was only 58.52% of the control. Then, the inhibition capability of the fraction of 10k < COSM < 30k was followed. But the fraction of COSM < 5k has no this effect.
Therefore, it stated that the size of molecular weight of COS decided the effect on preventing the retrogradation. But all the fractions of COS prepared by 6mL/30min H2O2 degradation had no effect on retarding the retrogradation due to changes in the structure. Therefore, molecular size and existence of active amino-group of COS determined the capability of anti-retrogradation.
Multiple instant heating -stirring preventing the retrogradation of high amylose rice flour (HA-RF) and HA-RS were investigated. The results indicate that three instant heating (100 oC)-stirring (500 r/min) had a clear effect on retarding the retrogradation of HA-RF. Treated HA-RF by three instant heating-stirring exhibited the lowest retrogradation enthalpy (3.04J/g dry flour) as compared to the control (5.93J/g dry flour) in DSC analysis and had almost the least recrystallization (52.32% of the control) by XRD. The observation by SEM stated that treated HA-RF by this way granules had a more honeycomb-like structure and reduced the crystal morphologies. The gelatinized HA-RF by three instant heating-stirring treatments enlarged mesh of network structure by SEM section observation, it implied that network structure had more gathered water by this method treatment. But this physical technology had no effect on the retrogradation of isolated HA-RS. The different results of treated HA-RF and HA-RS by this physical technology might be due to rice protein having anti-retrogradation function after treatment.
It was investigated that Fourier transform infrared (FT-IR) and Laser Confocal Micro-Raman (LCM-Raman) spectroscopy for rapid monitoring the retrogradation process of HA-RS. The results indicate the relative intensity of some vibrational modes reduced regularly with increasing extent of the retrogradation including O-H (3415cm-1) stretching, C-H (2927.46cm-1) stretching, C-O and C-C (1155.17cm-1) stretching, C-O-H (1081.89cm-1) bending, undetermined (1043.32cm-1, 1020.18cm-1 and 998.96cm-1) modes, skeletal modes of pyranose ring (609.41cm-1 and 578.55cm-1) in FT-IR spectra and C-O stretching and C-O-H bending (1122.37cm-1), C-O-H (1082.83cm-1) bending, C-C (1050.05cm-1) stretching, C-H (854.31cm-1) deformation, skeletal modes of pyranose ring (440.66cm-1) in Raman spectra. By the correlation analysis between retrogradation enthalpy of DSC and the relative intensity reciprocal of above vibrational modes, these peaks of 1081.89cm-1, 1043.32cm-1, 1020.18cm-1, 609.41cm-1, 578.55cm-1of FT-IR and 854.31cm-1, 1082.83cm-1, 1050.05cm-1 of LCM-Raman had correlation coefficient above 0.9, so these vibrational modes listed above may be as index to quantify the retrogradation. Meantime, the sensitivity of LCM-Raman was higher than FT-IR.
It was studied that the probability of fractal theory to characterize process of starch retrogradation. SEM was adopted to observe section feature of retrograded HA-RS stored at 5, 10 and 15d. The surface topography of retrograded starch had self-similarity and manifested fractal characteristics. Based on professional fractal software analysis, fractal behavior of various retrograded starch was remarkable. The fractal dimensions of retrograded samples stored at 15d, 10d and 5d were respectively 1.8267, 1.7803, and 1.7333. The extent of retrogradation (retrogradation enthalpy) and the fractal dimension had a linear relationship, linear equation was y = 0.0358x + 1.5934. Cluster-Cluster aggregation model was used to simulate the fractal growth process of starch retrogradation. Simulation results resembled the feature and fractal dimension of the experimental results, so this model can be characterize the growth mechanism of starch retrogradation.
引文
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