Glucuronic acid kinetics

With UDP-glucuronic acid as the variable substrate and bilirubin at constant concentration, Michaelis-Menten kinetics were obeyed (A2, H2, HIO, P5, V2, W12). Rat liver microsomal preparations treated in different ways yielded apparent Km for UDP-glucuronic acid of 0.37-0.70 mM (A2, H2, HIO, V2), with albumin-bound bilirubin as the aglycon a higher value (1.66 mAf) was found in a carrier-free system (W12). These values probably do not represent Km for UDP-glucuronic acid at saturation with bilirubin (P5, V6). Under certain conditions of activation, cell extracts from livers of newborn and adult rats, wdicn tested with o-amino-... 

In determining enzyme activities, it is usually assumed that at a fixed set of so-called saturating substrate concentrations a sufficiently accurate value of F, ax is obtained. Bisubstrate kinetic analyses of UDP-glucu-ronyltransferase [assayed with bilirubin (P5) and p-nitrophenol (V6), respectively] indicate that a true measure of the amount of enzyme can be obtained only by suitable extrapolation procedures. This restriction applies in particular to bilirubin (A2, HIO, T8) and other aglycons (M15, V6) because of substrate inhibition. UDP-glucuronic acid was inhibitory at concentrations only about 10-fold higher than the apparent Km value (HIO) this was most pronounced at relatively short incubation times. Mg was noninhibitory at concentrations equal to 20 times the apparent Km values (F3, HIO). 

Investigations of the mechanism of decarboxylation of hexuronic acids have largely involved kinetic and tracer studies. When either D-xylo-5-hexulosonic acid or D-glucuronic acid is converted into 27 in acidified, tritiated water, the resulting 27 contains 18% and 15%, respectively, of the activity of the solvent as carbon-bound tritium.21 Further degradation studies showed that the isotope is situated on the furan ring at either position 3 or 4, or both these atoms correspond to C-3 or C-4 of the starting uronic acid. 

The simple self-crosslinking treatment also crosslinks GAG chains to collagen [30]. The reaction kinetics are outlined in Fig. 2. The mechanism probably involves condensation of amino groups of collagen with carboxylic groups of glucuronic acid residues on the repeat unit of chondroitin 6-sulfate. Dehydra-... 

Fig. 2. Kinetics of cross-linking of chondroitin 6-sulfate, a glycosaminoglycan (GAG), to collagen following exposure to 105 °C under 6.7 Pa (50 mtorr). The mechanism of cross-linking is most probably interchain amide condensation involving e-amino groups of lysyl residues on collagen chains with carboxylic groups on glucuronic acid residues in neighboring GAG chains (From [30] with permission).

A series of investigations of the metabolism of foreign organic compounds in the intact rabbit recently reported54-88 have given information about the kinetics of glucuronic acid conjugation in vivo. 

The kinetics of the conjugation of D-glucuronic acid in some cases seem to follow a first-order reaction. Bray4 has recently reviewed the work in his laboratory on this subject. 

Thus, as the first approach to kinetic studies of the action of /8-D-glu-cosiduronase toward synthetically prepared 1-O-acyl-a- and -/3-D-glu-copyranuronic acids, a reliable method that would permit monitoring of the rate of enzymic hydrolysis was needed. For this purpose, To-masic and Keglevic268 developed an analytical procedure, involving the colorimetric reaction of D-glucuronic acid with benzidine in the presence of acetic acid,269 that proved to be fully applicable to enzymic, kinetic studies performed with 1-esters of D-glucuronic acids and with D-glucosiduronic acids as the substrates. 

On the basis of kinetic studies [75-77], Smith suggested that the major factors governing the choice of two alternative metabohc routes are the first-and zero-order rate constants of the two reactions. The first-order rate constant in conjugation reactions may not be subject to serious species variations, but the zero-order rate constant depends not only on the mobihzation rate of glycine, glucuronic acid, etc., which is different in different species, but also on the compound metabohzed. The compound may affect the zero-order rate constant either by virtue of its tissue level, which depends on dose, or by its effect on the maximum velocity of the enzymatic reaction [23]. 

Valproic acid is eliminated by first-order kinetics and has an elimination half-life of 5-20 hours (average, 10.6 hours). Pediatric patients (3 months to 10 years) have a 50% higher clearance of the drug expressed by weight (i.e., mL/min/kg) over the age of 10 years, pharmacokinetic parameters of valproic acid approximate those in adults (Cloyd et al., 1993). Valproic acid is metabolized principally in the liver by (3 (over 40%) and CO oxidation (up to 15%-20%). Thirty through 50% of an administered dose is excreted as glucuron-ide conjugates (Cloyd et al., 1993). 

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