FT-NIRS技术应用于稻米直链淀粉含量分析研究

运用近红外光谱快速分析技术,使用偏最小二乘法建立了近红外光谱和水稻糙米直链淀粉含量的数学模型,并进行糙米直链淀粉含量预测.结果表明糙米近红外光谱与其直链淀粉含量具有良好的相关性,决定系数r2=0.8429,最大绝对误差4.82%,平均误差2.30%.该方法在不破坏样品的前提下快速分析水稻直链淀粉含量,可用于稻种资源的快速鉴定,对于水稻优质育种及其相关研究具有重要意义.     

基于NIRS法快速检测荞麦蛋白质与淀粉含量的研究

为建立近红外光谱技术测定荞麦蛋白质与淀粉含量的方法,本研究以217份荞麦样品为试验材料,采用最小二乘回归预测和交叉验证构建近红外预测模型。分析表明:前处理采用多元散射校正法(MSC),维数(Rank)分别为5和5,光谱区间6803.9~6094.2/cm所建立的荞麦蛋白质与淀粉含量模型的预测效果较好,其决定系数(R~2)分别为0.9481和0.9167,交叉验证均方根(RMSECV)分别为0.68和2.08,相对分析误差(RPD)分别为4.39和3.46,均大于3.0,外部验证相关系数均大于0.96。本试验所建立的蛋白质与淀粉含量近红外预测模型具有较高的准确度和稳健性,可用于荞麦品质的快速测定。     

籼稻品质分析的近红外光谱模型建立及其应用研究

为了满足籼稻品质快速分析的需求,本研究利用籼稻精米粉近红外光谱建立了直链淀粉含量、蛋白质含量、碱消值、垩白度的回归预测模型.结果表明,本研究提供的预测模型具有良好的测定效果,用偏最小二乘法(PLS)获得的籼稻精米粉直链淀粉含量、蛋白质含量、碱消值、垩白度的回归模型和交叉验证显示最优校正决定系数(R~2)和交叉检验均方误差(RMSECV)分别为0.9561、1.55,0.9510、0.258,0.9076、0.283,0.9014、4.14.说明所建的近红外光谱预测模型具有实用价值.     

近红外光谱测定小麦粉常规营养成分的模型优化

目的：应用近红外光谱（NIR）结合偏最小二乘法（PLS）建立小麦粉常规营养成分蛋白质、水分和脂肪的含量预测模型,并选择最佳模型。方法：收集117份小麦粉样品的近红外光谱,化学法测定蛋白质、水分和脂肪的含量,利用主成分分析（PCA）随机分组,81份样品用于构建模型、36份样品用作验证模型的预测能力。探讨波长范围和光谱预处理方法对所建模型预测能力的影响。结果：3个营养成分预测能力最好的模型分别是：对于蛋白质,预处理采用矢量归一化（SNV）,波长选取7 505.9~5 446.2 cm-1和4 605.4~4 242.8 cm-1,预测模型的RPD值是7.02;对于水分,无预处理,波长选择全谱12 800~3 960 cm-1,模型的RPD值是6.83;对于脂肪,无预处理,波长在9 000~4 000 cm-1,模型的RPD值是5.06。结论：近红外光谱法可以实现对小麦粉常规营养成分的快速预测,通过选择波长范围和光谱预处理方法可以显著提高模型的预测能力。     

豌豆品质性状近红外模型建立及区域差异分析

探讨了傅立叶变换近红外光谱技术(FT-NIRS)检测豌豆蛋白质、淀粉、脂肪和总多酚含量的可行性。用化学方法测定190份豌豆种质的蛋白质、淀粉、脂肪以及总多酚含量,采集其子粒与粉末的近红外光谱,采用偏最小二乘法(PLS)分别建立两种光谱与成份含量预测模型。豌豆粉末模型结果优于子粒模型,其中蛋白质和淀粉的粉末模型的预测残差(RPD)为5.88、5.82,相关系数r2达到0.99、0.99,具有很好的预测性能。对其中产地信息详细明确的150份豌豆种质的品质性状与产地进行两步聚类分析,明确得到3种类型,其特点分别为:类群1低蛋白质含量,类群2高总多酚含量,类群3高蛋白质、高淀粉和高脂肪含量。进一步分析了豌豆品质性状随播种期、经度、纬度、海拔高度的变化情况。结果表明,近红外光谱技术可对豌豆种质资源的部分品质性状进行快速筛选鉴定,聚类分析结论、地理坐标与播期对豌豆种质主要品质性状的影响规律,都可为收集高品质性状豌豆种质资源提供可靠依据。     

不同生态环境与播种期下小麦籽粒品质变异规律的研究

选用 6个不同品质类型小麦品种在 4个生态点进行分期播种试验, 系统分析了不同生态环境下小麦籽粒产量与品质的变异特征及其与主要气候生态因子间的关系。结果表明 :生态点、品种以及地点×品种互作对籽粒产量、千粒重、蛋白质、湿面筋和淀粉含量、沉降值与降落值的影响均达到显著水平 ;不同播种期处理对产量与淀粉含量的影响达到极显著水平, 而播种期×品种互作对千粒重、降落值、淀粉含量及沉降值的效应达到显著水平 ;地点×播种期×品种互作仅对产量、湿面筋、淀粉含量与沉降值有显著的影响。在 4个不同生态点中, 南京点的籽粒蛋白质与湿面筋含量最低, 但淀粉含量最高 ;徐州点的产量和千粒重最大 ;泰安点的蛋白质含量与湿面筋含量最高, 沉降值最小 ;保定点的产量、千粒重最小, 但沉降值最大。不同播种期处理下, 适播与晚播的籽粒蛋白质含量、湿面筋含量、淀粉含量、沉降值与降落值都显著高于早播, 而早播期的产量和千粒重最大。各小麦品种在不同地点与播种期下产量与品质性状的变异中以降落值的变异系数为最大, 淀粉的变异为最小。开花至成熟期的日均温与淀粉含量呈线性负相关, 与产量、蛋白质和湿面筋含量及降落值呈二次曲线相关关系 ;日温差与产量和千粒重呈二次曲线相关关系, 籽粒蛋白质和湿面筋含量、沉降值及降落值则随昼温差的增加线性递增 ;开花至成熟期降雨量与产量、千粒重都呈现先升后降的二次曲线相关关系, 而与籽粒蛋白质含量和降落值呈现线性负相关关系 ;籽粒蛋白质和湿面筋含量与降落值随开花至成熟期的累计日照时数呈现线性正相关关系。     

花后干旱和渍水对不同品质类型小麦籽粒品质形成的影响

 防雨池栽条件下研究了花后干旱和渍水胁迫对两个不同品质类型小麦（Triticum aestivum）品种籽粒产量和品质形成的影响。结果表明,花后渍水和干旱处理明显降低了小麦籽粒产量和蛋白质产量。在整个灌浆期内干旱处理明显提高了籽粒蛋白质和醇溶蛋白含量,而渍水处理降低了籽粒蛋白质及其组分的积累量。籽粒总淀粉和直链淀粉含量以渍水处理最高,而支链淀粉以对照最高。干旱处理提高了籽粒干、湿面筋含量、沉降值和降落值,而渍水处理降低了上述品质指标。试验表明干旱和渍水胁迫对小麦籽粒蛋白质与淀粉的含量和组分及面粉品质等均有不同程度的影响,从而改变了不同品质类型小麦的籽粒品质。     

水稻直链淀粉和蛋白质突变系筛选及其与主要农艺性状的关联分析

研究利用重离子辐照杂交籼稻9311创建农艺性状突变体库,使用化学方法筛选直链淀粉、蛋白质突变材料,并分析籽粒品质性状与农艺性状之间的相关性,为后续筛选直链淀粉、蛋白质突变体工作奠定基础。结果显示:在169份直链淀粉、蛋白质突变体中,直链淀粉含量变幅范围是7.64%~32.37%,其中高含量直链淀粉突变体材料有11份,低含量直链淀粉突变体材料有5份;蛋白质含量变幅范围是7.05%~13.79%,其中高蛋白突变体有34份,低蛋白突变体有3份。从169份突变体中筛选出直链淀粉、蛋白质含量都有梯度差异的突变体材料,筛选出材料的农艺性状分析结果表明:在直链淀粉、蛋白质突变体材料的农艺性状中,每穗实粒数、有效穗数、结实率和株高这四个农艺性状与籽粒品质性状间有关联,变异系数分别为37.53%、30.72%、24.70%、15.38%。相关性结果表明:直链淀粉含量和蛋白质含量呈负相关,直链淀粉含量与农艺性状呈正相关,蛋白质与农艺性状呈负相关,其中蛋白质含量和每穗实粒数、结实率呈极显著负相关,相关系数分别为-0.504、-0.592。     

灌溉方式对黄淮稻区优质粳米品质的影响

为探明不同水深处理对优质稻米的影响,研究了湿润、浅水、深水3种灌溉方式对优质粳稻(圣稻14、圣稻2572、圣稻18)籽粒外观、稻米直链淀粉、蛋白质含量及淀粉RVA谱的影响,分析了稻米品质与理化指标之间的关系.结果表明: 浅水灌溉下籽粒面积、周长、粒长、稻米蛋白质含量显著高于其他灌溉方式,垩白率和垩白度的表现相反;RVA谱的最高黏度和热浆黏度值以湿润灌溉最高,深水灌溉最低,最终黏度值以深水灌溉最低;不同品种RVA谱的崩解值、消减值、回复值及稻米直链淀粉含量对3种灌溉方式的响应存在明显的基因型差异.圣稻14以湿润灌溉的食味品质最好,圣稻18则以深水灌溉最好.相关分析表明,稻米RVA谱特征值与稻米外观指标及蛋白质含量之间存在显著的相关性.回归与通径分析表明,粒型是影响崩解值与消减值的主要因子,而稻米蛋白质与直链淀粉含量是影响回复值的主要因子.     

水稻籽粒直链淀粉含量的生态模型研究

在中国、日本和泰国不同生态环境下进行了多品种籼稻和粳稻的种植试验,通过分析水稻籽粒直链淀粉含量与纬度、海拔、温度、太阳辐射等气候生态因子的相互关系,确立了影响水稻籽粒直链淀粉积累的主要气候因子函数,并使用权重系数进一步修订各个气候生态因子对籽粒直链淀粉的作用,构建出基于生态效应(综合气候因子函数)的水稻籽粒直链淀粉含量预测模型.利用不同年份、不同生态点和不同品种类型的试验资料对所建模型进行了检验,籼稻和粳稻籽粒直链淀粉含量的预测误差(RMSE)平均分别为0.4%和0.5%;籼稻和粳稻种植区的预测误差(RMSE)平均为0.39%和0.50%,表明模型具有较好的预测性和实用性.     

Alpha-amylase activity of Rhyzopertha dominica (Coleoptera: Bostrichidae) reared on several wheat varieties and its inhibition with kernel extracts

Total progeny of Rhyzopertha dominica (F.) (Coleoptera: Bostrichidae) reared on 10 wheat, Triticum aestivum L., varieties was evaluated. Higher amylase activities were detected in populations with few individuals, whereas the opposite was observed in higher populations. As protein ingested increased, reproductive success increased. However, consumption of wheat protein was inversely correlated with amylase activity levels (r = -0.66). Amylase activity in homogenates of R. dominica populations showed variable inhibition by wheat extracts prepared from wheat varieties on which they were reared. Insect populations with lowest amylase activities were inhibited more by wheat extracts than those with higher amylase activity (r = -0.77). An electrophoretic analysis revealed four phenotypes showing combinations of three isoamylases (Rm 0.70, 0.79, and 0.90) in different populations of R. dominica. Some of the insect progeny that emerged from resistant wheat varieties contained the three isoamylases, whereas progeny that emerged from the most susceptible varieties showed reduced activity of isoamylases 0.70 or 0.90. These results suggest that the alpha-amylase activity levels and the composition of isoamylases in R. dominica populations are modulated by diet and that the alpha-amylase inhibitory activity of the wheat kernels influences these variations.     

Expression of a bi-functional and thermostable amylopullulanase in transgenic rice seeds leads to autohydrolysis and altered composition of starch

Overexpression of bacterial-derived starch metabolic enzymes in plant starch storage organs represents a valuable strategy for improving starch quality, bioprocessing and nutritional value. Transgenic rice seeds producing a thermostable and bifunctional starch hydrolase, amylopullulanase (APU) from Thermoanaerobacter ethanolicus 39E, were generated. Starch in these seeds could be hydrolyzed with optimal temperatures between 85 and 95 °C, which resulted in complete conversion of starch into soluble sugars and production of protein-enriched flour within a few hours. By expressing various levels of APU, rice seeds containing reduced amounts of amylose, which is an important factor affecting starch quality, were obtained without a significant impact on grain yield. Elevation in granule-bound pullulanase activity correlates with the reduction of amylose in developing APU-containing rice seeds. APU was found to be localized within amyloplasts and in cell walls, which could be the result of overexpression of APU with a signal peptide. This study establishes novel approaches to alter starch properties, accelerate bioprocessing of starch and production of protein-enriched flour from rice seeds, and could significantly impact the industrial and food uses of cereals.     

Biochemical analysis of starch degradation by Ruminobacter amylophilus 70.

Ruminobacter amylophilus is an obligate anaerobe that uses only alpha-linked glucose molecules (i.e., maltose, maltodextrins, and starch) as a source of energy, making it an excellent model for the study of bacterial starch degradation. Constitutive amylase, amylopectinase, and pullulanase activities were found in intracellular and extracellular fractions of R. amylophilus. However, extracellular activities apparently resulted from cell lysis. Both soluble and membrane-bound polysaccharidase activities were detected. Most of the soluble polysaccharidase activity partitioned with the periplasmic cell fraction. No alpha-glucosidase or maltase activity was detected in either the cellular or extracellular fraction. In addition, intact cells of R. amylophilus bound U-14C-starch. This binding could be saturated and was constitutive and sensitive to proteinase K, indicating protein or protein complex mediation. Competition experiments showed that these starch-binding sites had equally high affinities for starch and maltodextrins larger than maltotriose. The sites had a reduced affinity for maltose and virtually no affinities for glucose and nonstarch polysaccharides. These findings suggest that R. amylophilus binds starch molecules to the cell surface as an initial step in transporting the molecule through the outer membrane and into the periplasmic space. Extracellular polysaccharides do not appear to be involved in starch degradation.     

Selective Staining of Protein and Starch in Wheat Flour and its Products

The main constituents of wheat flour and many wheat flour products are wheat protein (gluten) and starch granules. The specific staining of the protein present was effected by 10 min in 0.1% aqueous ponceau 2R (C.I. No. 16150) acidified with 3—4 drops of 1 N H₂SO₄ per 50 ml of staining solution, followed by rinsing in 2 changes of distilled water, dehydrating, clearing and mounting in a resinous medium in the normal way. Staining of starch was as follows: sections or flour smears were brought to water, treated for 10 min in a protein-blocking reagent (Taninol ADR—Imperial Chemical Industries—used in 1% aqueous solution) rinsed, then stained for 3 mins in 0.5% aqueous chlorazol violet R (C.I. No. 32445) or for 10 min in either 0.5% aqueous chlorazol violet N (C.I. No. 22570), or chlorazol black E (C.I. No. 30235). Staining was followed by thorough rinsing, normal dehydration and clearing and mounting in a medium of R.I. about 1.49 to enhance visibility of unstained starch grains. The methods are applicable to flour smears, cryostat and wax sections.     

豌豆种子萌发时的含水量对子叶中蛋白酶和淀粉酶活性的影响

不同含水量的豌豆种子在饱和水蒸气中保持7d,在此过程中,当含水量低于萌动临界含水量时,限制了子叶中淀粉酶长寿mRNA的转译;含水量达到或超过萌动临界含水量时,子叶中淀粉酶长寿mRNA的转译得以进行,但转译程度因含水量不同而异,正常生长的轴器官是吸收子叶中蛋白质和淀粉水解产物的动力库,子叶中氨基酸含量的变化灵敏地调节着蛋白酶的活性,从而影响着蛋白质的动员程度,当环境中供水不足时,发现子叶中还原糖积累很快,非还原糖含量几乎没有变化,因此,认为对淀粉酶活性起灵敏调节作用的主要是还原糖。     

Histochemical studies on mobilization of storage components in cotyledons of germinatingPhaseolus mungo seeds

Phaseolus mungo seeds were allowed to germinate in the dark at 27 C, and time-sequence changes of mobilization of protein and starch reserves in cotyledons were observed by histochemical techniques. The distributions of amylase and protease activities in cotyledon sections were also examined during germination by use of the starch-polyacrylamide gel film and India ink-gelatin film methods, respectively. Amylolytic and proteolytic processes occurred more or less simultaneously during the germination. At the day 2 stage, low levels of hydrolytic enzyme activities were observed throughout cotyledon sections. At day 4, both amylase and protease activities appeared to increase in tissue areas farthest from vascular bundles, and the mobilization of starch and protein reserves also proceeded in these areas. At day 6, the reserves were found to remain only in the cells around vascular bundles. When cotyledons were detached from axis organs, allowed to imbibe water and incubated for 4 days at 27 C, the breakdown of reserves was markedly retarded and the patterns of enzyme localization in cotyledon sections appeared not as conspicuous as those in the sections from intact cotyledons. These histochemical results are discussed with reference to the previous results ofin vitro experiments.     

Particle size distribution of rice flour affecting the starch enzymatic hydrolysis and hydration properties

Rice flour is becoming very attractive as raw material, but there is lack of information about the influence of particle size on its functional properties and starch digestibility. This study evaluates the degree of dependence of the rice flour functional properties, mainly derived from starch behavior, with the particle size distribution. Hydration properties of flours and gels and starch enzymatic hydrolysis of individual fractions were assessed. Particle size heterogeneity on rice flour significantly affected functional properties and starch features, at room temperature and also after gelatinization; and the extent of that effect was grain type dependent. Particle size heterogeneity on rice flour induces different pattern in starch enzymatic hydrolysis, with the long grain having slower hydrolysis as indicated the rate constant (k). No correlation between starch digestibility and hydration properties or the protein content was observed. It seems that in intact granules interactions with other grain components must be taken into account. Overall, particle size fractionation of rice flour might be advisable for selecting specific physico-chemical properties.     

Ectopic expression of bacterial amylopullulanase enhances bioethanol production from maize grain

Key message

Heterologous expression of amylopullulanase in maize seeds leads to partial starch degradation into fermentable sugars, which enhances direct bioethanol production from maize grain.

Abstract

Utilization of maize in bioethanol industry in the United States reached ±13.3 billion gallons in 2012, most of which was derived from maize grain. Starch hydrolysis for bioethanol industry requires the addition of thermostable alpha amylase and amyloglucosidase (AMG) enzymes to break down the α-1,4 and α-1,6 glucosidic bonds of starch that limits the cost effectiveness of the process on an industrial scale due to its high cost. Transgenic plants expressing a thermostable starch-degrading enzyme can overcome this problem by omitting the addition of exogenous enzymes during the starch hydrolysis process. In this study, we generated transgenic maize plants expressing an amylopullulanase (APU) enzyme from the bacterium Thermoanaerobacter thermohydrosulfuricus. A truncated version of the dual functional APU (TrAPU) that possesses both alpha amylase and pullulanase activities was produced in maize endosperm tissue using a seed-specific promoter of 27-kD gamma zein. A number of analyses were performed at 85 °C, a temperature typically used for starch processing. Firstly, enzymatic assay and thin layer chromatography analysis showed direct starch hydrolysis into glucose. In addition, scanning electron microscopy illustrated porous and broken granules, suggesting starch autohydrolysis. Finally, bioethanol assay demonstrated that a 40.2 ± 2.63 % (14.7 ± 0.90 g ethanol per 100 g seed) maize starch to ethanol conversion was achieved from the TrAPU seeds. Conversion efficiency was improved to reach 90.5 % (33.1 ± 0.66 g ethanol per 100 g seed) when commercial amyloglucosidase was added after direct hydrolysis of TrAPU maize seeds. Our results provide evidence that enzymes for starch hydrolysis can be produced in maize seeds to enhance bioethanol production.     

Induction of digestive alpha-amylases in larvae of Zabrotes subfasciatus (Coleoptera: Bruchidae) in response to ingestion of common bean alpha-amylase inhibitor 1

Zabrotes subfasciatus larvae possess three alpha-amylase isoforms determined by in gel assays following SDS-PAGE. Two minor isoforms present lower electrophoretic mobility than the major form. When developed inside Vigna unguiculata (cowpea) seeds, fourth instar larvae have minor quantities of the slow-migrating isoforms, but when reared on seeds of Phaseolus vulgaris (common bean), the two slow-migrating forms are expressed in higher amounts, whilst the quantity of the major constitutive form is independent of the host bean. Larvae at the beginning of the fourth instar were fed on flour or cotyledons of cowpea and common bean and it was observed that the larvae fed on the common bean expressed the two slow-migrating forms in higher amounts when compared to the control larvae fed on cowpea. In order to investigate the possible correlation between the induction of alpha-amylases and the ingestion of the common bean alpha-amylase inhibitor 1 (alphaAI-1), this inhibitor was incorporated into artificial diet. It was observed that larvae fed on diet containing chronic doses of alphaAI-1 during their development, produced the two slow-migrating forms in higher amounts than control larvae, however, fourth-instar larvae fed on the same diet presented less amylase activity than control larvae. The data suggested that alphaAI-1 is involved in amylase induction and that it has inhibitory activity against the constitutive amylase, when starch granules are used as substrate.