Measurement of specific activity

According to equation 5 the measured initial rate of reaction is proportional to the total concentration of enzyme. However, in practice, this is something that 

Let us consider the last of these to illustrate the sort of problems that can arise. If a concentration Cq of enzyme implies a concentration aco of a competitive inhibitor then the rate equation is not equation 5 but equation 15 (below), with the inhibitor concentration t written as aCo- Notice that this means that Cq appears not only as a factor of the munerator of the rate expression, but also as a term in the denominator, so the whole expression is no longer proportional to Cq. This makes the rate expression as a function of Cq have the same form that it has as a function of the substrate concentration a, so it approaches a limit of koKjc/Kj ct instead of increasing indefinitely as eo increases. Clearly this limit depends on all four of the quantities that it contains, but only one of them, a, is (in principle) under the control of the experimenter. If a cannot be made small enough to be negligible then use of the assay may need to be restricted to a low range of enzyme concentrations where the effect of the inhibitor is imperceptible. 

Once conditions have been determined that give a good proportionahty over a useful range, these can be used as the basis of an activity assay for the enzyme. As noted above, the rate is an intensive quantity, i.e. its magnitude does not vary with the total size of the system considered. The quantity required fix)m an assay, however, is the amount of enzyme, which is an extensive quantity because it does vary with the size of the system. It can be obtained by multiplying the rate obtained by the volume of the compartment in which the reaction takes place. The unit for measuring enzyme activity is mol s as the rate of conversion of a particular substrate under specified conditions. For example, a reaction rate of 5 (jlm s or 5 (ji,mol litre s , obtained in a 0.5 ml reaction volume would correspond to an enzyme activity of 5 x 0.5 x 10 = 0.0025 i.mol s .  

Notice that the definition of enzyme activity implies that all of the conditions, including the identity of the substrate, are specified. Thus a change of rate observed when a different substrate is used in the assay does not imply a change in enzyme activity (though if the change of substrate is adopted as part of the definition of the standard assay, it will imply a change in the definition of enz5nne activity). 

The SI unit of enzyme activity, 1 mol s , may also be called the katal, with symbol kat, but this term has not been widely adopted. It is a very large unit for most purposes, but its submultiples the nanokatal (nkat) and microkatal (jxkat) are of a convenient size for measuring enzyme activities in the laboratory. In the past it was usual to define a unit of enzyme activity measured in ixmol min (= 16.67 nkat), and this is still used, though it is not very satisfactory because the name is uninformative and fails to convey the incorporation of a factor of 10 and a non-SI unit (the minute) in the definition. 

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Measurement of specific activity. The half-life of a nuclide can be readily calculated if both the number of atoms and their rate of decay can be measured, i.e., if the activity A and the number of atoms of P can be measured, then X is known from A = XP. As instrumentation for both atom counting and decay counting has improved in recent decades, this approach has become the dominant method of assessing half-lives. Potential problems with this technique include the accurate and precise calibration of decay-counter efficiency and ensuring sufficient purity of the nuclide of interest. This technique provides the presently used half-lives for many nuclides, including those for the parents of the three decay chains, U, U (Jaffey et al. 1971), and Th. 

The challenge of FOPs demonstrating similar quality, safety, and efficacy to the innovator product relates to the poor predictability of physicochemical characteristics and biologic activity. For example, there are several different interferon-a and erythropoietin a and P products currently on the market. These variants are characterized by differences in sequence, glycosylation pattern, and in vitro measures of specific activity however, their clinical safety and efficacy profiles are considered similar [20],... 

Although the number of research papers reporting the use of NMR spectroscopy, especially by comparison to and is small the range of applications is extensive (and is described in more detail in the book by Evans et al.). For example, the measurement of specific activity, optical purity, stereochemistry, solution conformation, kinetic acidities, hydrogen isotope exchange, hydrogen bonding, fractionation factors, and radiolysis. 

Double (Multiple) IDA. Since the specific activity Ov is often unknown, double IDA can be used to determine m,.. Two equal aliquots containing the same, but unknown mass m, are diluted with different known carrier masses. i ,. and m, .. After. separation of portions m2, in 2 and measurement of specific activities at, a 2 one obtains from Equation (9) ... 

Quantitative measurements of optical activity are usually expressed in terms of the specific rotation, defined as... 

Normal measurements of optical activity are concerned with the ability of the optically active substance to rotate the plane of polarization of plane polarized light, its specific optical rotatory power ( ) being given by... 

Quantum yield of coelenterazine. The quantum yields of coelenterazine in the luminescence reaction catalyzed by luciferases A, B and C are all close to 0.30 at 24°C, which is one of the highest values among coelenterazine luciferases. The amount of luciferase L obtained was insufficient to measure reliable data of specific activity and quantum yield. 

Agarwal et al. 1978), the quantification of these specific enzymes may indicate that exposure to endosulfan has occurred. Blood tests, such as decay curves for aminopyrine in plasma, which are semiquantitative indices of liver enzyme induction, have been used successfully in the past to demonstrate enzyme induction in pesticide-exposed workers. Because numerous chemicals found at hazardous waste sites also induce these hepatic enzymes, these measurements are not specific for endosulfan exposure. However, measurements of enzyme activity, together with the detection of the parent compound or its metabolites in tissue or excreta, can be useful indicators of exposure. All of these potential biomarkers require further verification in epidemiological studies. Further studies with focus on the development of methods to separate and measure the estrogenicity of endosulfan in in vitro assays would be valuable since these assays are more sensitive and discriminative than other conventional biomarkers. Preliminary results have been presented by Sonnenschein et al. (1995). 

Lippi et. al (87) and Dirstine (88) circumvented titration by converting the liberated fatty acids into copper salts, which after extraction in chloroform are reacted with diethyldithio-carbamate to form a colored complex which is measured photometrically. While the end point appears to be more sensitive than the pH end point determination, the advantages are outweighed by the additional steps of solvent extraction, centrifugation and incomplete extraction when low concentrations of copper salts are present. Other substrates used for the measurement of lipase activity have been tributyrin ( ), phenyl laurate (90), p-nit ro-pheny1-stearate and 3-naphthyl laurate (91). It has been shown that these substrates are hydrolyzed by esterases and thus lack specificity for lipase. Studies on patients with pancreatitis indicate olive oil emulsion is definitely superior to water soluble esters as substrates for measuring serum lipase activity. 

Most measurements include the determination of ions in aqueous solution, but electrodes that employ selective membranes also allow the determination of molecules. The sensitivity is high for certain ions. When specificity causes a problem, more precise complexometric or titri-metric measurements must replace direct potentiometry. According to the Nernst equation, the measured potential difference is a measure of the activity (rather than concentration) of certain ions. Since the concentration is related to the activity through an appropriate activity coefficient, calibration of the electrode with known solution(s) should be carried out under conditions of reasonable agreement of ionic strengths. For quantitation, the standard addition method is used. 

The radioligand should also have a high specific activity so that very small quantities of bound ligand can be accurately measured. The specific activity, simply defined as the amount of radioactivity, expressed in becquerels (Bq) or curies (Ci) per mole of ligand, is dependent on the half-life of the isotope used and on the number of radioactive atoms incorporated into the ligand molecule. A radioisotope with a short half-life decays rapidly so that many disintegrations occur in unit time,... 

SS). Data for the metals of group VIII and for rhenium in group VIIA are given in Fig. 1, which is divided into three fields separating the metals of the first, second, and third transition series. The specific activity is defined as the activity per unit surface area of metal. Metal surface areas required for the determination of specific activities are derived from measurements... 

Measurement of exoenzymatic activities is potentially useful in detecting the effects of toxicants on heterotrophic biofilm communities. Sensitivity and direct relationship with organic matter use and, therefore, microbial growth make extracellular enzyme activities a relevant tool to assess the toxicity of specific compounds. Use of novel approaches that combine enzymatic and microscopic tools (e.g. ELF-phosphatase) may be extremely useful to detect anomalies at the sub-cellular scale. 

Other automated systems may be purchased for a specific purpose and are called dedicated instruments, e.g. glucose analyser. Others have fairly restricted applications, an example being the reaction rate analysers which are specifically designed for the kinetic measurements of enzyme activity. Some of the more recently developed instruments employ individual pre-prepared disposable test packs or strip devices which contain all the reagents for each particular assay in a dry form. 

Feature 14. Develop chemical facility-specific measures of security activities and achievements, and self-assess against these measures to understand and document program progress. 

All samples were dried for 72 hours at 80°C, and dry weights were calculated. Dried samples were milled to spectrometrically measure the specific activity of i Cs. The standard error of specific activity was in the range 10-20%. Statistical analysis used the software package MS Excel. 

In order to judge the efficiency of the vaccine system, the number of specifically activated CTL was measured. In case of a successful activation, the number of these cells should rise significantly in the timeframe of days after immunization. 

By active surface area we meant the kinetically active part of the total surface area. According to Helgeson et al. (1984), this area is restricted to etch pits. Alternative estimates of surface areas may be obtained from measurements of specific surface area s whenever solid particles have a narrow size range. The specific surface area for spherical particles is given by... 

EMSA assays can also be exploited to measure STAT nuclear localization, which is, similar to NFkB localization, a measure of STAT activity. Determination of JAK phosphorylation is carried out by immunoprecipitation of the JAK proteins from cell lysates, followed by SDS-PAGE electrophoresis, immunoblotting with antiphosphotyrosine antibody and JAK-specific antibody re-probing [99]. 

Since the early times of stereochemistry, the phenomena related to chirality ( dis-symetrie moleculaire, as originally stated by Pasteur) have been treated or referred to as enantiomericaUy pure compounds. For a long time the measurement of specific rotations has been the only tool to evaluate the enantiomer distribution of an enantioimpure sample hence the expressions optical purity and optical antipodes. The usefulness of chiral assistance (natural products, circularly polarized light, etc.) for the preparation of optically active compounds, by either resolution or asymmetric synthesis, has been recognized by Pasteur, Le Bel, and van t Hoff. The first chiral auxiliaries selected for asymmetric synthesis were alkaloids such as quinine or some terpenes. Natural products with several asymmetric centers are usually enantiopure or close to 100% ee. With the necessity to devise new routes to enantiopure compounds, many simple or complex auxiliaries have been prepared from natural products or from resolved materials. Often the authors tried to get the highest enantiomeric excess values possible for the chiral auxiliaries before using them for asymmetric reactions. When a chiral reagent or catalyst could not be prepared enantiomericaUy pure, the enantiomeric excess (ee) of the product was assumed to be a minimum value or was corrected by the ee of the chiral auxiliary. The experimental data measured by polarimetry or spectroscopic methods are conveniently expressed by enantiomeric excess and enantiomeric... 

Kabai, J. (1973) Determination of specific activation energies of metal oxides and metal oxide hydrates by measurement of the rate of dissolution. Acta Chem. Acad. Sci. Hung. 78 57-... 

Birchall, K., Gdlet, V.J., Harper, G. and Pickett, S.D. (2006) Training similarity measures for specific activities application to reduced graphs. Journal of Chemical Information and Modeling, 46 (2), 577-586. 

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