Enzymes Present

When substitutions are made at carbon 6 of jS-glucosides, the effects are quite different than those arising from substitutions at the ring carbons (2, 3, 4, 5). In the accompanying formula, this type of substitution is represented by variations in the nature of the atoms or groups represented byX  

The ease of hydrolysis of a series of such 6-substituted jS-glucosides (34) is compared in Table VII. 

Effect of Pubification on the Rates of Hydrolysis OF A Series of /J-Glucosidbs 

Substituent group (group X) Phenyl glucoside (E,E,) Vanillin 6-X-jS-glucoside Volume of group (Biltz) 

Changes of Ring Structure and Configuration. When the structure of a hydrolyzable pyranoside is changed to that of a furanoside, the latter is unaffected by the enzyme. Thus, the a- and jS-D-glucofuranosides are unaffected 36) by almond emulsin as well as by yeast a-glucosidase (yeast maltase). Similarly, yeast invertase catalyzes the cleavage of sucrose and other jS-fructofuranosides, but not that of the known fructopyranosides 37). 

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Phosphatase Test. The phosphatase [9001-78-9] test is a chemical method for measuring the efficiency of pasteurization. AH raw milk contains phosphatase and the thermal resistance of this enzyme is greater than that of pathogens over the range of time and temperature of heat treatments recognized for proper pasteurization. Phosphatase tests are based on the principle that alkaline phosphatase is able, under proper conditions of temperature and pH, to Hberate phenol [108-95-2] from a disodium phenyl phosphate substrate. The amount of Hberated phenol, which is proportional to the amount of enzyme present, is determined by the reaction of Hberated phenol with 2,6-dichloroquinone chloroimide and colorimetric measurement of the indophenol blue formed. Under-pasteurization as well as contamination of a properly pasteurized product with raw milk can be detected by this test. 

Enzyme Assays. An enzyme assay determines the amount of enzyme present in sample. However, enzymes are usually not measured on a stoichiometric basis. Enzyme activity is usually determined from a rate assay and expressed in activity units. As mentioned above, a change in temperature, pH, and/or substrate concentration affects the reaction velocity. These parameters must therefore be carefully controlled in order to achieve reproducible results. 

Enzymes are proteins of high molecular weight and possess exceptionally high catalytic properties. These are important to plant and animal life processes. An enzyme, E, is a protein or protein-like substance with catalytic properties. A substrate, S, is the substance that is chemically transformed at an accelerated rate because of the action of the enzyme on it. Most enzymes are normally named in terms of the reactions they catalyze. In practice, a suffice -ase is added to the substrate on which die enzyme acts. Eor example, die enzyme dial catalyzes die decomposition of urea is urease, the enzyme dial acts on uric acid is uricase, and die enzyme present in die micro-organism dial converts glucose to gluconolactone is glucose oxidase. The diree major types of enzyme reaction are ... 

FIGURE 14.7 Substrate saturation curve for au euzyme-catalyzed reaction. The amount of enzyme is constant, and the velocity of the reaction is determined at various substrate concentrations. The reaction rate, v, as a function of [S] is described by a rectangular hyperbola. At very high [S], v= Fnax- That is, the velocity is limited only by conditions (temperature, pH, ionic strength) and by the amount of enzyme present becomes independent of [S]. Such a condition is termed zero-order kinetics. Under zero-order conditions, velocity is directly dependent on [enzyme]. The H9O molecule provides a rough guide to scale. The substrate is bound at the active site of the enzyme. 

Studies of chemical reactions in solution and in enzymes present an enormous challenge because of the enormous size and complexity of these systems. MM force fields have made a tremendous impact in certain areas, but they cannot... 

The minute quantities of enzymes present in cells complicate determination of their presence and concentration. However, the abifity to rapidly transform thousands of molecules of a specific substrate into products imbues each enzyme with the abifity to reveal its presence. Assays of the catalytic activity of enzymes are fre-quendy used in research and cfinical laboratories. Under appropriate conditions (see Chapter 8), the rate of the catalytic reaction being monitored is proportionate to the amount of enzyme present, which allows its concentration to be inferred. 

The physicochemical properties of the reactants in an eiKyme-catalyzed reaction dictate the options for the assay of enzyme activity. Spectrophotometric assays exploit the abihty of a substrate or product to absorb hght. The reduced coenzymes NADH and NADPH, written as NAD(P)H, absorb hght at a wavelength of 340 run, whereas their oxidized forms NAD(P) do not (Figure 7—9). When NAD(P)+ is reduced, the absorbance at 340 run therefore increases in proportion to—and at a rate determined by—the quantity of NAD(P)H produced. Conversely, for a dehydrogenase that catalyzes the oxidation of NAD(P)H, a decrease in absorbance at 340 run will be observed. In each case, the rate of change in optical density at 340 nm will be proportionate to the quantity of enzyme present. 

Of the thousands of different enzymes present in the human body, those that fulfill functions indispensable to cell vitality are present throughout the body tissues. Other enzymes or isozymes are expressed only in specific cell types, during certain periods of development, or in response to specific physiologic or pathophysiologic changes. Analysis of the presence and distribution of enzymes and isozymes— whose expression is normally tissue-, time-, or circumstance-specific—often aids diagnosis. 

Results of these investigations indicated that there were enzymes present in the littleneck clam that facilitated decarbamoylation of either the carbamate or sulfocar-bamoyl toxins to their decarbamoyl form see Figure 5). These enzymes may be unique to this particular species of shellflsh since, although the decarbamoyl toxins have been found in small quantities in other species, no shellflsh species examined to date contains the predominance of the decarbamoyl toxins found in the littleneck clams. 

There are important methodologic considerations which apply to the use of cultured amniotic fluid cells for the detection of biochemical disorders. The first is that the enzymes which can be sampled are those which are usually present in fibroblasts or fibroblast-like cells. Therefore, conditions such as phenylketonuria and glycogen storage disease type I, which are associated with deficiencies of enzymes present only in liver and kidney, are not amenable to this approach. The same also pertains to enzyme deficiencies affecting other specific tissues. 

Subsequently, a clear Juice is obtained by ultrafiltration. A serious problem in this process is the fouling of the ultrafiltration membrane, causing a reduced flux rate. For apple processing, the material responsible for this effect has been isolated and extensively characterized [2-4]. It appeared to consist mainly of ramified pectic hairy regions (MHR), which were not degraded by the pectolytic enzymes present in the technical pectinase preparation. 

Table 1 shows some biochemical properties of the pectolytic enzymes present in pool 1. The pectin lyase/pectate lyase activities (pool I) and polygalacturonase activity (pool II) were not significantly affected by NH4+, Na+ and K+ (0,25 - 2,5mM), while Al +, p-mercaptoethanol, Hg2+, EDTA, Ba + and Zn+2 (2,5mM) inhibited 30-100% these activities. On the other hand, Ca2+, Mg + and Mn + at 2,5mM concentration activated 20-100% pectin/pectate lyases but Ca " " and Cu " " (2,5mM) inhibited polygalacturonase activity about 42 - 70%. 

The activities of pectic enzymes present in cultivation medium (98 mg of protein extracted from 2.5 1 of pectin medium) were poor, not leading to the clarification of cultivation medium indicating the cleavage of pectate chains, with values 0.024 pmol/min.mg for polygalacturonase, 0.004 pmol/min.mg for exopolygalacturonase, 0.034 pmol/min.mg for pectinesterase and 0.005 pmol/min.mg for pectin lyase. The production of individual pectic enzymes was dependent on the C-source used in the cultivation medium (Tab. 1). 

The pectinase was supported on y-alumina and the three enzymes present in the pectinase sample were found still active after the immobilisation. The supported biocatalyst was used in several reaction cycles to perform consecutive depectinisations of a cloudy apple juice with a negligible loss of biocatalytic activity. 

There are also enzymes present that participate in the formation of many of the several hundred volatile compounds found in tea aroma. The important enzyme systems responsible for the biosynthesis for the methylxanthines have already been mentioned. 

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