Serum kinetic properties

The application of quality control procedures to ensure that satisfactory analytical performance of enzyme assays is maintained on a day-to-day basis is complicated by the tendency of enzyme preparations to undergo denaturation with loss of activity. This maltes it difficult to distinguish between poor analytical performance and denaturation as possible causes of a low result obtained for a control sample introduced into a batch of analyses. Assured stability within a defined usable time span is therefore the prime requirement for enzyme control materials, as it is for enzyme calibrators. However, specifications for the two types of materials can differ in other respects. Because the function of a calibrator is to provide a stated activity under defined assay conditions, it is not necessary for it to show sensitivity to changes in the assay system identical to those of the samples under test therefore within certain Umits, enzymes from various sources can be considered in the search for stability. However, it is the function of a control to reveal small variations in reaction conditions, so it must mimic the samples being analyzed. The preparation of enzymes from human sources is not by itself a guarantee of an effective control. For example, human placental ALP is very stable, but it differs significantly in kinetic properties from the liver and bone enzymes that contribute most of the ALP activity of human serum samples it is therefore not an ideal enzyme for use in control material for the determination of ALP. 

The physiological levels of cocaine can reach a maximum of about 0.3 micromolar (M) in serum immediately after cocaine administration (Jeffcoat et al. 1989). These levels are much less than the Km of the liver benzoyl esterase (0.7 mM). Hence, the enzyme will obey first-order kinetics, where activity equals k at/KM, called the catalytic efficiency, times the cocaine concentration. The catal5Tic efficiency of the liver benzoyl esterase, 11 min mM", is similar to human serum cholinesterase, 7 min mM ", another enzyme that catalyzes the hydrolysis of the benzoyl group of cocaine. The content and kinetic properties of serum cholinesterase and liver benzoyl esterase need to be evaluated to determine which enz5mie has the greater capacity for hydrolysis of cocaine in human. 

A comparative study of the properties of human and rat 3-D-2-acetamido-2-deoxyhexosidases has been reported. There are four j3-D-2-acetamido-2-deoxy-hexosidases isoenzymes in rat serum one is thermostable and the remaining three are thermolabile. Human serum contains three isoenzymes two are thermostable and one is thermolabile. In general, the /3-D-2-acetamido-2-deoxy-hexosidases isoenzymes in rat serum are more thermolabile than those in human serum. In contrast, the kinetic properties of the j8-D-2-acetamido-2-deoxyhexo-sidase isoenzymes in serum from the two species are similar. Rat liver and human placenta contain two 3-D-2-acetamido-2-deoxyhexosidase isoenzymes, one of which is thermolabile and the other thermostable. These tissue isoenzymes from the two species have similar physical and kinetic properties. 

A neutral a-D-glucosidase from porcine serum preferentially hydrolysed malto-oligosaccharides. The substrate specificity and kinetic properties, etc., of a jS-l,4-D-glucanase (mol. wt. 5.1 x 10 pH optimum 5.5—6.0, temperature optimum 45 °C) isolated from the intestinal juices of a snail Helixpomatia) have been determined. The purified enzyme, which degrades poly- and oligosaccharides (d.p. >3) containing /8-(l -> 4)- and j8-(l 3)-linked D-glucosyl residues, was inhibited by Hjedta and appeared to be activated by Ca + ions. 

It appears that, generally, sialic acid is not a part of an antigenic determinant (atropinesterase and orosomucoid) nor a functioning part of a catalytic or binding site ( i-antitrypsin, atropinesterase, cemloplasmin, and serum cholinesterase). However, sialic acid does seem to play a role in transport ( i-antitrypsin). This effect is discussed more fully later in this chapter. Johnson et al. (1970) found that removal of sialic acid residues from the copper-containing protein cemloplasmin had no effect on its kinetic properties as an oxidase. They concluded that the terminal sialic acid residues do not influence the stmcture and function of the active sites in cemloplasmin. 

These prodrugs underwent spontaneous hydrolysis in aqueous solution. The mechanism of reaction (Fig. 11.10) was postulated to involve nucleophilic hydration of the C=N bond to yield an intermediate carbinolamine. The latter breaks down with loss of the dialkylamine to form an Af4-formyl intermediate, which, in turn, hydrolyzes to the active agent (11.68, Fig. 11.10). The hydrolysis of these prodrugs followed pseudo-first-order kinetics with tm values at pH 7.4 and 37° that were mostly influenced by the steric properties of the dialkylamino group. Thus, tm values were ca. 4, 9 - 10, 14 -15, and 48 - 52 h for R = Me, Et, Pr, and i-Pr, respectively. Interestingly, the rates of hydrolysis were decreased in human serum, indicating protective... 

The combination of the health-relevant time-window and the toxicokinetic properties of the agent of interest determine the optimal exposure assessment strategy. Dioxin, a contaminant of chlorophenoxy herbicides and fungicides, has a relatively long biological half-life, estimated at about seven years and is measurable in serum. Serum measurements of dioxin are therefore relatively stable, and simple first-order kinetics have been used to back-estimate serum dioxin levels on the basis of an occupational history. Such exposure data have been used quite successfully in epidemiological analyses of cohorts of pesticide producers (Hooiveld et al, 1998). 

Alkan, M. et al.. Surface properties of bovine serum albumin-adsorbed oxides Adsorption, adsorption kinetics and electrokinetic properties, Micropor. Mesopor. Mater., 96, 331, 2006. 

The inhibitory properties of PED were also shown in the pregnant-mare-serum-gonadotropin-stimulated rat ovarian microsomes (PMSGSROM). Comparison of the inhibitory kinetic parameters of PED in the human placental and PMSGSROM preparations showed close resemblance time-dependent Ki of 5 nM and 14.5 nM and t i2 of inactivation of 11.8 min and 16.2 min, respectively. The Fmax of human placental microsomes was, however, 10 times higher than that of PMSGSROM (51.8 9.4 vs 4.2 0.9 pmol/min/mg protein, respectively, with testosterone as substrate). The affinities of aromatase from the two sources were also similar Johnston and coworkers showed that baboon placental aromatase, but not the rhesus placental enzyme, was similar in activity to the human placental preparation. 

In contrast to the naturally occurring Gd compounds, pharmacokinetic and pharmacological properties of the element s DOTA and DTPA chelates that are used in MRl have been studied more intensively. From a kinetic point of view, these highly hydrophilic compounds behave in an identical fashion. As expected from hydrophilic substances, the volume of distribution is small being 0.17 0.01 liters/kg in humans [13] while the plasma elimination half-life is around 20 and 90 min in rats and humans, respectively [4]. In mice 89% of the administered Gd-DOTA and Gd-DTPA doses was recovered in the urine within 1 hr [14]. In correlation with the reduced GFR in patients with chronic renal failure (median creatinine clearance 25.5 mL/min) the serum half-life of Gd-DTPA was prolonged and the renal clearance decreased. Recovery of the Gd-DTPA after administration of a 0.1 mmol/kg dose was 92 12%, whereas extrarenal elimination in these subjects accounted for less than 0.4% [15], indicating that glomerular filtration is the predominant route of elimination of the chelates. 

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