development of an in vitro screening model for the biosynthesis of ...

FIG. 4.Nonlinear regression analysis of the degradation of the whole acyl glucuronide (top) and appearance of aglycone (bottom) during incubation at 37°C with
HSA, pH 7.4.
, tolmetin; f, zomepirac; Œ, ketoprofen; ‚, fenoprofen; F, suprofen; E, furosemide; , diclofenac; ✳, ibuprofen.
cannot be assessed in a screening process. Furthermore, HSA remains
the protein most extensively studied. It is widely distributed in the
plasma compartment and easily related to the immune system. The
knowledge gathered on covalent binding of acyl glucuronide and HSA
allowed the comparison and validation of the results achieved with
this model. A product showing a significant covalent binding to HSA
in our model will require specific attention during the development
process. The constant values calculated for the disappearance of acyl
glucuronides were lower than those previously published, especially
for zomepirac and tolmetin (Hasegawa et al., 1982; Ojingwa et al.,
1994). In fact, those reported values represented the global degradation
of 1-O-acyl glucuronide (hydrolysis isomerization), whereas
the values presented in this study only represent the degradation of
isomeric forms. In the conditions of analysis described here, the exact
IN VITRO ASSESSMENT OF ACYL GLUCURONIDE REACTIVITY
TABLE 2
Apparent first-order degradation of various acyl glucuronides and aglycone
appearance constants measured during incubation phases with 0.5 mM HSA, pH
7.4, at 37°C
Mean S.D. (n 3).
Compound Name Acyl Glucuronide Degradation Rate Aglycone Appearance Rate
Tolmetin 0.055 0.020 0.106 0.049
Zomepirac 0.025 0.008 0.103 0.027
Suprofen 0.046 0.021 0.035 0.018
Diclofenac 0.050 0.016 0.081 0.023
Fenoprofen 0.021 0.014 0.044 0.011
Ibuprofen 0.028 0.003 0.037 0.013
Ketoprofen 0.037 0.010 0.016 0.005
Furosemide 0.006 0.011 0.000 0.001
h 1
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