融合淀粉酶AmyP-Clo对大米生淀粉的高效降解

【目的】获得能高效降解大米生淀粉的α-淀粉酶。【方法】将来源Clostridium butyricum T-7的α-淀粉酶的淀粉结合结构域与能快速偏好性降解大米生淀粉的α-淀粉酶Amy P的催化结构域融合重组表达。【结果】融合蛋白Amy P-Clo保留了野生型Amy P催化优势的基础上,对大米生淀粉的比活为(373.9±8.4)U/mg,4 h内对5%大米生淀粉的最终降解率为(42.7±1.1)%,对大米生淀粉的最高结合率为(71.1±1.6)%,这些数据相比Amy P分别提高了3.1、2.8和1.3倍。【结论】融合蛋白Amy P-Clo能高效降解生大米淀粉,是一个具有优良应用价值的酶。     

Improvement in the In Vivo Digestibility of Rice Protein by Alkali Extraction Is Due to Structural Changes in Prolamin/Protein Body-I Particle

Rice prolamin, constituting type-I protein body (PB-I), is indigestible and causes deterioration of rice protein nutritional quality. In this study, the in vivo digestibility of rice protein isolates was investigated by tracing their intraluminal transit in the gastrointestinal (GI) tract of rats by western blotting and by observing the structures excreted in the feces by electron microscopy. Two types of rice protein isolates, produced by alkali extraction (AE-RP) and by starch degradation (SD-RP), were compared. The protein patterns in the isolates were similar, but their digestion in the GI-tract showed striking differences. In the AE-RP group, 13-kDa prolamin (13P) quickly disappeared in the lower GI tract and was not excreted in the feces. By contrast, in the SD-RP group, 13P accumulated massively and nearly intact PB-Is were excreted. These results indicate that the in vivo digestibility of prolamin can be improved by alkali extraction through structural changes to it.     

不同生态条件对云南和韩国粳稻品质及淀粉RVA谱特性的影响

以10个云南粳稻和6个韩国粳稻品种为材料,研究了不同生态条件对云南和韩国粳稻主要品质及RVA谱特性的影响.结果表明： 云南和韩国粳稻各指标的变异系数均以消减值最大.云南粳稻的粒长、垩白率、直链淀粉含量、蛋白质含量、碱消值、最终黏度、消减值和峰值时间的变异系数明显大于韩国粳稻,而其他品质指标则相反.随海拔升高,云南粳稻的粒长、粒宽、长宽比、精白度、直链淀粉含量、碱消值、最终黏度和回复值及韩国粳稻的粒长、粒宽、精白度、蛋白质含量、最高黏度、热浆黏度和崩解值明显降低,而云南粳稻的垩白率、蛋白质含量、热浆黏度和峰值时间及韩国粳稻的长宽比、直链淀粉含量、碱消值、消减值和回复值明显升高,但韩国粳稻的垩白率无明显变化.云南粳稻的最高黏度、崩解值和糊化温度及韩国粳稻的糊化温度随海拔升高呈先降后升趋势,而云南粳稻的消减值及韩国粳稻的最终黏度和峰值时间则表现相反趋势.     

一株产生淀粉酶杆菌Bacillus sp. GEL-09的筛选、鉴定及发酵条件优化

【背景】生淀粉酶可以水解生淀粉颗粒,在酒精发酵、白酒、黄酒和食醋的生料酿造工业中具有广阔的应用前景。【目的】从自然环境中筛选产生淀粉酶的菌,对其发酵条件及酶性能进行考察,为淀粉生料发酵过程提供优良菌种和酶资源。【方法】取木薯田土壤,经过稀释、热处理、富集培养以及木薯淀粉平板筛选培养基初筛,摇瓶复筛得到产高效降解生淀粉酶的菌株;经过菌落形态、细胞染色观察以及16S rRNA基因序列比对进行鉴定;对筛选菌株的发酵培养基和发酵条件进行优化,并对酶蛋白进行分离纯化和酶学性质分析。【结果】分离到一株具有较高生淀粉酶水解活力的菌株GEL-09,经鉴定为芽胞杆菌Bacillus sp.GEL-09;该菌在最优发酵条件下培养96 h,胞外酶活力达到430.6 U/m L,是优化前的2.8倍;酶学性质分析发现该酶为中温、中性酶,最适温度和p H为50°C和7.0;生淀粉降解能力对比发现,该酶的生淀粉降解能力值为62.3%,显著高于细菌α-淀粉酶、生麦芽糖淀粉酶和甘薯β-淀粉酶对生淀粉的降解能力。【结论】Bacillus sp.GEL-09在生淀粉酶生产方面具有良好的开发应用前景。     

Analysis of metabolites in Japanese alcoholic beverage sake made from the sake rice Koshitanrei

ABSTRACT

In sake brewing, the steamed rice is used in two ways, added to sake-mash (as kake-mai) and making koji. The rice is an important determinant for the quality of sake, as the metabolites in sake affect its taste/aroma. The sake rice Koshitanrei (KOS) was developed in Niigata Prefecture by genetically crossing two sake rice, Gohyakumangoku and Yamadanishiki. However, the metabolites in sake from KOS have not been analyzed. Here, to investigate the characteristic metabolites in sake from KOS, we performed two types of small-scale sake-fermentation tests changing only the rice used for kake-mai or total rice (both kake-mai and koji) by these three rice cultivars and examined the effect of KOS on sake metabolites by the metabolome analysis method using UPLC-QTOF-MS. We identified the peaks/metabolites, whose intensity in sake from KOS was higher/lower than those from the other cultivars. The brewing properties of KOS were partially characterized by this analysis.     

淀粉分支酶3存在于水稻胚乳的淀粉粒之中

对水稻胚乳淀粉颗粒结合的淀粉分支酶进行了研究.酶活性分析表明水稻胚乳中存在着与淀粉颗粒结合的淀粉分支酶.氨基酸测序分析结果表明结合于水稻胚乳淀粉粒的淀粉分支酶是分子量为84 kD的淀粉分支酶3(rice starch branching enzyme 3; RBE3).从开花后5 d到种子成熟,淀粉颗粒结合的RBE3蛋白都保持较为稳定的含量.Northern 分析表明水稻胚乳发育过程中RBE4最先表达而RBE3和RBE1的表达滞后.综合以上研究结果说明RBE3存在于水稻胚乳的淀粉之中是由于RBE3与淀粉葡聚糖链具有较高亲和性而难以和葡聚糖链解离,进而随着淀粉粒的增长而被其包裹.     

转反义Wx基因水稻颖果中与淀粉合成和积累相关酶活性的变化

将反义Wx基因转入水稻,导致Wx蛋白不同程度减少,颖果中的直链淀粉含量不同程度下降,总淀粉含量显著降低,直链淀粉与总淀粉的比值极显著降低。在水稻颖果发育过程中,ADPG-PPase、GBSS、SSS和SBE的活性在灌浆前期迅速升高,达最大值后很快下降,在灌浆中后期下降趋缓。Wx蛋白减少后的转基因水稻颖果中的GBSS活性明显下降,下降幅度与直链淀粉含量相一致,而且活性高峰期比其亲本有所提前。转基因水稻颖果中ADPG-PPase和SSS的活性在颖果发育的前中期,SBE则在中后期高于相应的亲本。     

不同气象条件下小麦冷源的品质变异

小麦冷源是一种冷温供体材料,有较强的降温作用,当受体小麦接受冷温供体的配子后,其后代冠温呈普遍下降态势,且部分受体小麦的温度型可由非冷型转化为后代的冷型,而冷型小麦具有代谢功能好且利于高产、稳产的特征,故小麦冷源不但对冷型小麦的问世有重要诱导作用,且能更好促进小麦的高产、稳产.将小麦冷源材料放置在不同的气象条件之下,以研究其籽粒品质的变异状况.结果表明,小麦冷源和非冷源相比,17项籽粒品质性状变异等级的加权平均值为1.9877,后者为2 8171,显示出小麦冷源的籽粒品质具有明显变异小、较稳定的特征,这和籽粒形成过程中蛋白质、淀粉及其组分的含量,相互间的比例变异小密切相关.由于小麦冷源具有籽粒品质变异小的特性,因而,这种特性就利于在其后代冷型小麦上显现,从而使冷型小麦的一些优良品质性状能在生产上反复重演,这不但进一步赋予了利于高产、稳产的冷型小麦以优质、稳质的性状,并能最终促进小麦生产的较大发展,其意义是十分重要的.     

水稻叶片淀粉的分离与理化性质分析

目的 分离高质量的叶片淀粉有助于研究叶片淀粉的理化特性及发育调控机制。利用已报道的叶片淀粉提取方法分离水稻叶片淀粉;步骤繁琐;操作要求高;且分离的水稻叶片淀粉纯度不高。因此;需要建立一种适合水稻叶片淀粉提取的方法。 方法 优化获得一种操作简单的水稻叶片淀粉分离和纯化方法;主要包括叶片磨粉、细胞裂解、淀粉释放、过筛、杂质去除、静置沉降、水洗淀粉、乙醇脱水和烘干保存9个步骤。该方法改良的核心在于利用静置沉降替代拟南芥叶片淀粉提取方法中的离心收集淀粉;通过3次悬浮沉降即可有效分离水稻叶片淀粉。 结果 利用该方法分离纯化3种水稻叶片的淀粉;经测定沉淀中淀粉纯度可以达到87%。与多角形的水稻胚乳淀粉粒形态相比;水稻叶片淀粉粒呈现扁平状和长的椭球形;且淀粉粒显著小于胚乳淀粉粒。热力学特性分析显示水稻叶片淀粉的糊化起始温度、峰值温度和终止温度显著高于胚乳淀粉;而热焓值低于胚乳淀粉。 结论 优化的叶片淀粉分离和提纯方法操作过程简单并且环保;获得的高纯度水稻叶片淀粉可以用于淀粉形态和理化特性的研究。     

氮磷水平与气象条件对春小麦籽粒蛋白质含量形成动态的影响

以3个品质类型春小麦品种的施肥和播期试验为基础,通过建立籽粒蛋白质含量形成动态的拟合方程,定量揭示籽粒蛋白质含量形成动态及氮磷肥与气象条件的影响效应。结果表明,灌浆期籽粒蛋白质含量随时间动态变化的普遍规律符合一元三次多项式曲线,即呈自开花始先降低后升高的单谷曲线变化。氮磷肥与气象条件的影响及基因型差异通过方程特征量而体现。氮水平增加,高蛋白品种蛋白质含量增加,动态曲线谷值和峰值均明显提高且出现时间分别提前和推后。磷水平增加,高蛋白强筋品种蛋白质含量降低,高蛋白中筋品种则增加;高蛋白品种谷值和峰值均提高且出现时间推迟。低蛋白品种蛋白质含量随氮磷肥变化不明显且幅度很小。在没有水分胁迫的情况下,光温互作是影响籽粒蛋白质含量动态形成的首要因子,其次为降水;而气温日较差则为最敏感因子。较高光温条件互作前提下,增加灌浆期温度日较差使高蛋白品种蛋白质含量提高,低蛋白品种则降低。高蛋白相比于低蛋白品种更易受氮磷水平和气象条件影响。     

The structure of starch can be manipulated by changing the expression levels of starch branching enzyme IIb in rice endosperm

When the starch branching enzyme IIb (BEIIb) gene was introduced into a BEIIb-defective mutant, the resulting transgenic rice plants showed a wide range of BEIIb activity and the fine structure of their amylopectins showed considerable variation despite having the two other BE isoforms, BEI and BEIIa, in their endosperm at the same levels as in the wild-type. The properties of the starch granules, such as their gelatinization behaviour, morphology and X-ray diffraction pattern, also changed dramatically depending on the level of BEIIb activity, even when this was either slightly lower or higher than that of the wild-type. The over-expression of BEIIb resulted in the accumulation of excessive branched, water-soluble polysaccharides instead of amylopectin. These results imply that the manipulation of BEIIb activity is an effective strategy for the generation of novel starches for use in foodstuffs and industrial applications.     

Production of Honbushi-like Flavor by the Fermentation of Nijiru

It takes about 2 months for the molding process of Katsuobushi (dried bonito) production. A model of a short-term system for the molding process was desirable for the efficient selection of useful fungi for Katsuobushi production by evaluating the flavor change in the model system.

A liquid culture system using Nijiru (waste fluid of Katsuobushi sterilization), which is obtained from the Kezuribushi (sliced Katsuobushi) manufacturing process and contains abundant phenolic compounds of smoke tar, was found to be suitable for the short-term evaluation of a 10 day culture.     

Effects of protein on crosslinking of normal maize, waxy maize, and potato starches

Channels of maize starch granules are lined with proteins and phospholipids. Therefore, when they are treated with reagents that react at or near the surfaces of channels, three types of crosslinks could be produced: protein–protein, protein–starch, starch–starch. To determine which of these may be occurring and the effect(s) of channel proteins (and their removal) on crosslinking, normal and waxy maize starches were treated with a proteinase (thermolysin, which is known to remove protein from channels) before and after crosslinking, and the properties of the products were compared to those of a control (crosslinking without proteinase treatment). After establishing that treatment of starch with thermolysin alone had no effect on the RVA trace, three reaction sequences were used: crosslinking alone (CL), proteinase treatment before crosslinking (Enz-CL), proteinase treatment after crosslinking (CL-Enz). Two crosslinking reagents were used: phosphoryl chloride (POCl₃), which is known to react at or near channel surfaces; STMP, which is believed to react throughout the granule matrix. Three concentrations of POCl₃ (based on the weight of starch) were used. For both normal maize starch (NMS) and waxy maize starch (WMS) reacted with POCl₃, the trends were generally the same, with apparent relative degrees of crosslinking indicated to be CL-Enz = CL > Enz-CL, but the effects were greater with NMS and there were differences when different concentrations of reagent were used. The basic trends were the same when potato starch was used in the same experiments. Crosslinking with STMP was done both in the presence and the absence of sodium sulfate (SS). Both with and without SS and with both NMS and WMS, the order of indicated crosslinking was generally the same as found after reaction with POCl₃, with the indicated swelling inhibition being greater when SS was present in the reaction mixture. Examination of the maize starches with a protein stain indicated that channel protein was removed by treatment with thermolysin when the proteinase treatment occurred before crosslinking with either POCl₃ or STMP, but only incompletely or not at all if the treatment with the proteinase occurred after crosslinking. Because the crosslinking reactions were less effective when the protein was removed, the results are tentatively interpreted as indicating that they involved protein molecules, although there may not be a direct relationship.