葡萄糖氧化酶的固定化及其性能的研究

本文对葡萄糖氧化酶的固定化进行了研究,考查了载体、缓冲溶液pH值,以及酶担载量对固定化酶性能的影响,确定了固定化过程的工艺条件,并对固定化葡萄糖氧化酶的自由态葡萄氧化酶的性能进行了比较。研究结果表明,以TiCl_4处理的5A分子筛作载体所制备的固定化葡萄糖氧化酶具有良好的化学与物理性能,在主要指标上优于文献报道的结果。     

The Metabolic Fate of Hydrogenated Glucose Syrups

The physiological response to the ingestion of glucose and glucose syrups is well documented in the literature. More recently, hydrogenated glucose syrups have become available but little is known of their physiological effect in man although sorbitol (hydrogenated glucose) has been used for many years in diabetic foods. Some metabolic effects of maltitol (hydrogenated maltose), which is a major component in hydrogenated syrups, have been described but the results obtained are equivocal. Our present work concerns physiological studies of hydrogenated glucose syrups and maltitol in animals and human subjects. In rats it has been shown that maltitol is utilized in some quantity by the gut microflora as shown by differences between conventional and germ-free animals. Intravenously injected maltitol is excreted almost quantitatively in the urine producing no significant rise in blood glucose. This contrasts sharply with intravenously injected maltose which induces a significant blood glucose rise and of which less than 10% is excreted in the urine. In human subjects hydrogenated glucose syrup may be tolerated at levels between 80 and 120g per day without diarrhoea. Our studies indicate that although after an oral dose maltitol and hydrogenated glucose syrup induce lowered blood glucose and serum insulin peak values, compared with glucose, total influx of the three carbohydrates shows no difference. This may have importance regarding the use of maltitol and hydrogenated glucose syrup in diabetic foods. Long term feeding trials with human subjects, using maltitol and hydrogenated glucose syrup as the sole dietary carbohydrates, show that some adaption to the carbohydrates occurs during the trial as judged by elevated blood glucose and serum insulin peak values at the end of the trial compared with initial values.     

Variation of Microwave Dielectric Properties in the Glucose Biosensor System

Glucose biosensor is generally based on the reaction between glucose and glucose oxidase, which produces gluconic acid and hydrogen peroxide. The gluconic acid is a conductive material, while hydrogen peroxide has polar molecules. This article examines the changes of dielectric properties due to the conductive loss below 2 GHz and dipole orientation of above that frequency of this reaction. The difference between the dielectric loss of glucose solution and the dielectric loss of glucose-enzyme reaction can be related to the glucose concentration in the samples, such as orange juice, black grape juice, sugarcane juice, and sapodilla juice. A good sensitivity to these differences due to the effect of ionic conductivity and dipole orientation was found at 1 and 16.44 GHz, respectively. The minimum detection limit of glucose concentration in the proposed technique was about 0.01 M (0.20 g/100 ml) with an optimum reaction ratio of about 1:1 between the enzyme solution and the glucose solution. This technique could benefit the future development of microwave biosensor by which both ionic conductivity and dipole effects can be considered simultaneously.     

Production of High Fructose Syrup by Glucose Isomerase Immobilized on Phenol-Formaldehyde Resin

An immobilized glucose isomerase which is active and stable in concentrated glucose solution has been developed. The enzyme has been immobilized to Duolite A7 resin simply by adsorption. Industrial application of this immobilized enzyme for the continuous production of high fructose syrup is quite promising because the immobilized enzyme having enough mechanical strength against the pressure of column flow, is active over broad ranges of both temperature and pH and is less dependent on Mg and Coions. Long-run continuous experiments in column system have been carried out, isomerizing 45% of glucose with flow rate of SV 1 at 60°C. A gradual decrease of activity was observed after 25 d operation and the half-life was calculated to be 36 d. One liter of the immobilized enzyme in column isomerized 288 kg of glucose in a month. The values of kinetic parameters of the immobilized enzyme in column operation varied with flow rates and approached to those of the native enzyme when they were assayed at the flow rate approaching to SV 0.     

固定化葡萄糖异构酶的研究 Ⅰ 游动放线菌(Actinoplanes missouriensis)-Ac-81-2葡萄糖异构酶的活力测定

测定游动放线菌胞内的葡萄糖异构酶时,无论在新鲜菌体、干菌体或用戊二醛直接交联的固定化菌体中,找不到酶量与反应生成物间的直线范围。但是在新鲜菌体中培养液量的倒数与反应速度的倒数呈直线关系。在干菌体或固定化菌体中则是所测酶量的对数与反应生成物的对数呈直线关系;酶量的对数与酶活力(单位/克)的对数呈直线关系。据此,对测定反应生成物果糖的半胱氨酸一咔唑反应作了修改和补充。     

A comparison of two glucose syrup drinks with D-glucose monohydrate solutions as test-loads in the Glucose Tolerance Test

A solution of d-glucose monohydrate is a commonly used test-load in the Glucose Tolerance Test, but it is also permissible to use a solution of glucose syrup (WHO, 1965). However, since 1965, a greater variety of glucose syrups has become available and efforts have been made to define the Glucose Tolerance Test more rigidly. Thus, a considerable section of this paper is devoted, by way of introduction, to clarifying and standardising various details of this test.     

湿热条件下大豆分离蛋白与葡萄糖、麦芽糖的美拉德反应

研究湿热条件下反应温度、反应时间及混合质量比对大豆分离蛋白（soybean protein isolate,SPI）与葡萄糖、麦芽糖之间美拉德反应的影响,并对所得美拉德反应产物的溶解性、乳化性、溶液pH值、电势、热性能、结构和分子质量的变化进行表征。结果表明,SPI与2?种还原糖发生美拉德反应的最适条件为按质量比4∶1混合后在80?℃条件下反应6?h。美拉德反应明显改善了SPI的溶解度,并且麦芽糖比葡萄糖对SPI的改性效果更佳;美拉德反应降低了SPI的乳化性,并对其ζ电势有一定影响,随着美拉德反应程度的增加,SPI的负电荷明显减少;美拉德反应会使SPI溶液的pH值降低;热稳定性分析表明美拉德反应降低了SPI的热稳定性;傅里叶变换红外光谱分析表明美拉德反应在SPI中引入了新的化学键;荧光分析表明美拉德反应增强了SPI的荧光强度;十二烷基硫酸钠-聚丙烯酰胺凝胶电泳测定表明美拉德反应导致了大分子物质的形成。     

Oxidized Glucose Syrup Production,Parameters and Food Applications

This contribution deals with the production, properties and food applications of a novel glucose syrup. For the production of this syrup, two methods are compared and from the obtained results the most suitable preparation process is recommended. In this connection some properties like optical rotation, dextrose equivalents (DE), molecular weight and mineral matter content were determined. Food applications like mineral-complexing power and rat feeding were investigated. The produced syrup was examined along with conventional types of syrup for its mineral-complexing properties, and it has been found that oxidized glucose syrup may have an important mineral-complexing power. The short-term rat feeding trials have shown no external abnormalities in the animals offered experimental diets. This suggests that oxidized glucose syrup deserves further study as a possible food component.     

Glucose Isomerase and Its Behaviour under Hydrogenation Conditions

A survey is given of glucose isomerase, its sources, its mechanism of isomerization and its data and properties in three different immobilized forms. In addition, the effect of a number of parameters on the activity of immobilized glucose isomerase has been investigated, e. g. hydrogen pressure, Mg(II) and Ca(II), transition metal ions, borate and sugar alcohols. Immobilized glucose isomerase remains sufficiently active under hydrogenation conditions to maintain D -glucose and D -fructose in equilibrium. D -Glucitol, in contrast to D -mannitol, has some inhibiting effect on the enzyme action. The D -glucose/D -fructose equilibrium constant is independent of the total sugar concentration (between 0.2 – 2.2-M).     

亚甲基蓝为介体的丝网印刷双酶修饰电极测定葡萄糖

以亚甲基蓝作为电子媒介,通过壳聚糖固定葡萄糖氧化酶和辣根过氧化酶在丝网印刷电极上,制备成葡萄糖生物传感器。循环伏安法和恒电位法用于研究修饰电极的电化学特性,在优化的实验条件下,获得传感器对葡萄糖响应的线性范围为8．0×10-5～3．5×10-3mol/L。此方法测定葡萄糖抗干扰能力强、重现性好、准确度高,电极制作简单,使用方便。     

静电场对葡萄糖溶液和蔗糖溶液冰晶生长的影响

为了探讨静电场对糖溶液冷冻过程冰晶生长的影响,本文采用自行设计的静电场辅助冷冻实验台,以不同浓度(5、50、100、150 g/L)的葡萄糖溶液和蔗糖溶液作为研究对象,在样品冷冻过程中施加不同强度(分别为0、4、6、8、10 k V外加电压)的静电场。以过冷温度和相变时间为考察参数,分析静电场对糖溶液冰核产生和冰晶成长的影响。结果表明:不同强度的静电场对糖溶液冰晶生长的影响不同,且其影响与溶液浓度有关。对于5 g/L、50 g/L葡萄糖溶液和5 g/L蔗糖溶液,较高强度(6 k V和8 k V)静电场有促进冰核产生的影响;对于150 g/L葡萄糖溶液和蔗糖溶液,较高强度(6 k V)静电场可能对冰核产生有促进的影响。静电场对葡萄糖溶液和蔗糖溶液的冰晶成长都有抑制水分子扩散运动的不利影响。     

The Emi2 Protein of Saccharomyces cerevisiae is a Hexokinase Expressed under Glucose Limitation

Hexokinases catalyze glucose phosphorylation at the first step in glycolysis in eukaryotes. In the budding yeast Saccharomyces cerevisiae , three enzymes for glucose phosphorylation have long been known: Hxk1, Hxk2, and Glk1. In this study, we focus on Emi2, a previously uncharacterized hexokinase-like protein of S. cerevisiae . Our data show that the recombinant Emi2 protein (rEmi2), expressed in Escherichia coli , possesses glucose-phosphorylating activity in the presence of ATP and Mg ²⁺ . It was also found that rEmi2 phosphorylates not only glucose but also fructose, mannose and glucosamine in vitro . In addition, we examined changes in the level of endogenous Emi2 protein in S. cerevisiae in the presence or absence of glucose and a non-fermentable carbon source. We found that the expression of Emi2 protein is tightly suppressed during proliferation in high glucose, while it is strongly upregulated in response to glucose limitation and the presence of a non-fermentable carbon source. Our data suggest that the expression of the endogenous Emi2 protein in S. cerevisiae is regulated under the control of Hxk2 in response to glucose availability in the environment.     

Temperature Dependence of Stability and Activity of an Immobilized Glucose Isomerase in a Packed Bed

A mathematical model describing the kinetics of the conversion of glucose to fructose by an immobilized glucose isomerase has been developed. The model is a straight-forward one and based on known kinetic behaviour of glucose isomerase and on limitations posed by diffusional resistance when a spherical particle containing the enzyme is considered. The model has been used to derive a relationship between initial enzyme activity and temperature. It can be demonstrated that the relationship between activity and temperature shows deviations from the familiar Arrhenius plot. The same holds for the activity half life time in a packed bed column. Experimental material confirming the theoretical considerations is presented. The experimental results have been applied to estimate some important parameters of the material used. An Arrhenius-type relationship is shown to hold for the estimated enzyme activity as well as for the kinetics of the deactivation process. The parameters obtained can be used in temperature optimization studies for the packed bed conversion process. The influences of initial activity, particle size and temperature on the production of fructose are discussed.     

Continuous Isomerization of Glucose to Fructose on a Commercial Basis

Immobilized glucose isomerase has been used to make more than 500,000 metric tons of high fructose corn syrups in the United States. These syrups have found wide application as high quality sweeteners to replace sucrose. A commercial process is described for the continuous enzymatic conversion of glucose to fructose. The use of immobilized glucose isomerase in a multiple bed reactor system is an economical method for making fructose. A comparison between immobilized glucose isomerase in a batch versus a continuous process shows the continuous process has the following advantages:

• 1 Lower enzyme usage and cost for converting glucose to fructose.
• 2 Lower capital equipment and labor costs.
• 3 Better and easier process control.
• 4 Lower refining costs and improved product quality.

Both the continuous isomerization process and the immobilization technique described are developments of Clinton Corn Processing Company, a Division of Standard Brands, Inc.     

The Production and Properties of Glucose Syrups. II. Physiological Properties of Glucose Syrups

The study indicates that when gum arabic, fat and protein are taken in conjunction with glucose syrup, changes in blood glucose profiles occur compared with glucose syrup alone. With gum arabic the effect is greatest in the 21% D.E. syrup, 30 min post ingestion (causing an elevation of blood glucose) and thereafter the effect is confined to the higher D. E. syrups. Added protein produces an elevated blood glucose peak and fat a decreased peak. Smaller loadings (10 g instead of 50 g) of different D.E. syrups showed no significant differences in blood glucose profiles.     

壳聚糖凝胶固定葡萄糖氧化酶制备酶电极的工艺

以壳聚糖作为固定GOD载体 ,研究了壳聚糖溶液的粘度 ,交联剂戊二醛的浓度、用量以及铂丝在酶膜母液中浸涂时间等对葡萄糖传感性能的影响。在优化固定化条件的基础上 ,建立了通过调整固定化条件来适应载体溶液粘度变化的工艺 ,制备出性能基本一致的酶电极     

Glucose Oxidase/Cqtalase Improves Preservation of Shrimp (Heterocarpus reedi)

The preservative effect of glucose oxidase (GOX) and catalase (CAT) in a 4% glucose aqueous solution were evaluated on shrimp (Heterocarpus reedi) stored at 0–2°C. On-board and immediately after catch, shrimp were washed with tap water at 10°C, and dipped or kept in the enzyme solution. Microbial (total psychrotrophs and Pseudomonas spp. counts), chemical (total volatile bases, ammonia and pH) and sensory tests (whole-raw and cooked-peeled samples) showed that holding shrimp in the GOX/CAT/glucose solution retarded microbial spoilage and preserved quality more effectively than dipping shrimp in the same solution. The importance of treating shrimp as early as possible was demonstrated by comparing shrimp treated 4 and 82 hr after catch.     

Immobilization of Glucose Isomerase on Radiation-Modified Gelatine Gel

Whole-cell glucose isomerase immobilization on radiation-modified gelatine gel was demonstrated. The enzyme was stabilized within the cells by heat-treatment at 70°C. The heat-treated cells were mixed with radiation-modified gelatine gel and cross-linked by glutaraldehyde. Immobilized enzyme exhibited 70–75% activity as compared to the free cells. Several properties of immobilized glucose isomerase were studied under various reaction conditions. A continuous isomerization was performed using a column packed with cell granules. It has been found, that the half-life of the immobilized glucose isomerase amounted to 650 h at 60°C.     

葡萄糖的化学发光检测法及对发酵过程糖耗曲线的测定

本文首次将葡萄糖和Ｃｕ~（２＋）的氧化反应与Ｃｕ~（２＋）催化下鲁米诺和Ｈ_２Ｏ_２的化学发光反应偶联，建立了葡萄糖的化学发光（ＣｈｅｍｉｌｕｍｉｎｅｓｃｅｎｃｅＣＬ）检测法，并成功地测定了酒精酵母发酵过程中的糖耗曲线。该方法操作简便，线性范围宽（１０~（－６）～１０~（－３）ｍｏｌ／Ｌ）。Ｆ－检验结果表明，ＣＬ法的测量精度明显高于传统的２，５－二硝基水扬酸比色法（ＤＮＳ法）。ＣＬ法的变异系数和最低检测限分别为２．６％和２．３×１０~（－６）ｍｏｌ／１（０．２ｍｌ，８３ｎｇ），均大大小于ＤＮＳ法的变异系数（５．０％）和最低检测限（１．１１×１０~（－５）ｍｏｌ／Ｌ，３ｍｌ，６μｇ）。     

高效液相色谱法测定烟草中的果糖、葡萄糖和蔗糖

以72%(体积比)乙腈/28%水预混合溶液作流动相,High-Performance Carbohydrate卡套柱作色谱柱,等梯度洗脱,采用带示差折光检测器的高效液相色谱仪测定了烟草中的果糖、葡萄糖和蔗糖含量。结果表明:方法的日内精密度和日间精密度的范围分别为0.46%~1.54%和0.96%~4.74%,回收率97.0%~102.3%,果糖、葡萄糖和蔗糖的检出限(LOD)分别为95,107和105μg/mL(S/N=3∶1),可以满足烟草中果糖、葡萄糖和蔗糖含量的日常检测要求。