Initial inactivation rate

In this work, the influence of Ti02 concentration on E. coli inactivation was investigated in the range of 0.0025—2gL-1 at two different light intensities. In general, the time required for total inactivation increases as Ti02 concentration decreases. Figure 17.1a (107 CFU) shows that the initial inactivation rate (r0)... 

The inactivation is normally a first-order process, provided that the inhibitor is in large excess over the enzyme and is not depleted by spontaneous or enzyme-catalyzed side-reactions. The observed rate-constant for loss of activity in the presence of inhibitor at concentration [I] follows Michaelis-Menten kinetics and is given by kj(obs) = ki(max) [I]/(Ki + [1]), where Kj is the dissociation constant of an initially formed, non-covalent, enzyme-inhibitor complex which is converted into the covalent reaction product with the rate constant kj(max). For rapidly reacting inhibitors, it may not be possible to work at inhibitor concentrations near Kj. In this case, only the second-order rate-constant kj(max)/Kj can be obtained from the experiment. Evidence for a reaction of the inhibitor at the active site can be obtained from protection experiments with substrate [S] or a reversible, competitive inhibitor [I(rev)]. In the presence of these compounds, the inactivation rate Kj(obs) should be diminished by an increase of Kj by the factor (1 + [S]/K, ) or (1 + [I(rev)]/I (rev)). From the dependence of kj(obs) on the inhibitor concentration [I] in the presence of a protecting agent, it may sometimes be possible to determine Kj for inhibitors that react too rapidly in the accessible range of concentration. ... 

Two major difficulties must be considered in any assay for acid phosphatase. The enzyme is subject to surface inactivation (23, 24). Accordingly, reproducible initial hydrolytic rates are not always obtained, and the kinetic behavior should be checked in any new assay developed. Discrepancies between the amount of inorganic phosphate produced and phenol liberated from phenolic phosphates may be substantial if extensive phosphotransferase activity occurs because of phosphoryl acceptor action on the part of hydroxylic buffers or other constituents of the incubation mixture (25, 26). Fluorogenic assays have been developed with very high sensitivity (27). Reference will be made to particular assays in the discussion of the specific enzymes. 

The chloroplast thylakoids were first incubated in apH=5 medium and then rapidly changed to a basic medium with pH fixed at 8.2. The acid incubation period was restricted to 30 sec to minimize inactivation of the ATP synthase activity. AT values were maintained at 5,44 or 60 mV. At ApH=3.2 and each given AT, a linear increase in the ATP yield was observed up to a reaction time of 200 ms. The authors thus concluded that within this time span both ApH and AT remained practically constant. The slopes of the three curves, representing the rates of ATP synthesis at ApH=3.2, are 120,260 and 380 mmol ATP/mol Chi for AT equal to 5,44 and 60 mV, respectively. The rate of ATP synthesis gradually decreased after the initial linear portion and eventually approached zero, as ApH and AT decayed to approach threshold values completely. The three separate plots clearly show that the eventual ATP yield as well as the initial ATP rate increases with increasing amplitude of the diffusion potential AT. 

As can be seen in Figure 1, the reaction accelerated markedly with the increase in power output when it was below 80 W. Further increase in power output up to 100 W, however, resulted in little rise in conversion after reaction for 2 h and the ME content of the product was lower than 30% in spite of the higher initial reaction rate, suggesting partial inactivation of the enzyme. This has been proved by experiments showing that the enzyme retained only 81% of... 

Conventionally, reaction rates in enzyme kinetics refer always to initial reaction rates where the maximum catalytic potential of the enzyme is expressed and many factors affecting it (i.e. substrate depletion, accumulation of inhibitory products, enzyme inactivation, reverse reaction) are irrelevant (see section 1.3). The quantification of such effects on that maximum catalytic potential is the subject of sections 3.2, 3.3 and 3.4. 

Specific activity of biocatalyst molar concentration of substrate B (alternatively coefficient in Eq. 5.3) initial molar concentration of substrate B coefficient in Eq. 5.3 concentration of biocatalyst time of a cycle of reactor operation enzyme activity initial enzyme activity molar concentration of enzyme species Eij volumetric activity of enzyme species Ey enzyme volumetric activity initial enzyme volumetric activity bioreactor feed flow-rate total flow-rate to downstream operations initial feed flow-rate to bioreactor i number of half-lives of biocatalyst use film volumetric mass transfer coefficient for substrate Michaelis-Menten constant catalytic rate constant first-order inactivation rate constant transition rate constants... 

In these experiments initial reaction rates were measured by the decrease in absorbance at 265 nm and calculated as nanomoles per minute from = 6600. The final volume of 0.90 ml contained 0.01 M potassium citrate (pH 6.5), 0.016 / g of enzyme (Sigma Chemical Co., grade IV, from rabbit muscle), 0.005 M KCl, 60 /xM (I), and 50 /xM AMP. The order of addition was (a) buffer, (b) enzyme, (c) (I), followed 1 min later by AMP. In control assays carried out in conjunction with the inactivation experiments, the order of addition was (a) buffer, (b) (I), followed 1 min later by (c) the enzyme and (d) AMP. These control assays showed that hydrolyzed (I) was not inhibitory the observed rates varied between 1.11 and 1.13 nmoles/min. 

Here C is the flavor concentration in the stored spray-dried powder, Q is the initial flavor concentration, k o is the initial release rate constant, AG is the activation energy of flavor release, k is Boltzmann s constant and h is Planck s constant. Equation 6.9 was originally developed to express the inactivation kinetics of acovalently bound to a water-insoluble support in an aqueous system (Kawamura et id., 1981). Equation 6.9 was applied to express the oxidation kinetics of fish oil (EPA) in hnoleic add powder, based on the assumption that the free energy of activation AG follows a Gaussian distribution with the standard deviation a (Yoshii et al., 2003 Ishido et al., 2002, 2003). 

Cycling experiments are performed by first measuring the heat rates at some reference temperature at which the tissue is stable, then exposing the sample to a temperature near the high or low limit for viability for various time intervals. The samples are returned to the reference temperature to compare initial heat rate values with values after exposure to the temperature extreme. When inactivation occurs at the temperature extreme, heat rate values measured at the reference... 

Complex inactivation kinetics caused by enzyme-catalyzed decomposition of epoxide kinetic constants calculated from initial rates of inactivation. Approximate value calculated from half-life in the presence of 50 mAf inhibitor. 

Figure 8.9 Reaction progress curve in the presence of a mechanism-based inactivator when a second aliquot of enzyme is added to the reaction solution. The reaction is allowed to reach a plateau before a second, equal concentration aliquot of enzyme is added at the indicated time point. Note that the rate of inactivation for this second aliquot of enzyme is the same as that seen in the initial progress curve.

A second test for buildup of a free inactivator is to measure product formation in the presence of an excess of compound until the progress curve reaches a plateau, and to then add a second aliquot of enzyme. As described earlier in this section, the addition of a second aliquot of enzyme should result in renewed product formation, which will wane with time as the new molecules of enzyme are inactivated. The rate of inactivation of the second aliquot of enzyme (measured as kobs) should be the same as that of the first aliquot of enzyme in the experiment, if the compound is functioning as a true mechanism-based inactivator. If, instead, inactivation is due to buildup of an inhibitory species, then the second value of lcohs should be greater than the first value. This experiment can also be performed by preincubating the enzyme with compound and initiating the reaction with cognate substrate, as... 

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