Immobilization of microsomes into alginate beads is a convenient method for producing glucuronides from drugs.
The production of glucuronides from drugs by immobilized microsomal uridine diphosphate (UDP)-glucuronosyltransferase has been investigated. Of all the immobilization methods used (covalent binding, adsorption by ionic or hydrophobic interactions), only entrapment of microsomes into alginate beads in the presence of polyethyleneimine was effective in producing high glucuronidation rates, thus leading to the formation of large amounts of metabolites. The performance of the bioreactor was optimized with the drug 3'-azido-3'-deoxythymidine (AZT), active against the human immunodeficiency virus, as a model substrate of UDP-glucuronosyltransferase. Calcium (12 mM) could optimally improve the stability of microsomes entrapped in alginate beads. Upon immobilization, enzyme activation occurred, leading to a fivefold increase in specific activity. The determination of apparent Km and Vmax revealed that AZT was a better substrate for the immobilized enzyme than free microsomes. The AZT-glucuronide production obtained after 6 h was threefold higher than that observed with free microsomes. This bioreactor was also efficient in production of glucuronides from structurally different compounds such as bilirubin, 4-nitrophenol, clofibric acid, pirprofen, dextrorphan or morphine, the corresponding glucuronide of which possesses pharmacological or toxicological interest.
Centre du Médicament, U.R.A. CNRS no. 597, Nancy, France.
M
M
Haumont
J
J
Magdalou
J C
JC
Ziegler
R
R
Bidault
J P
JP
Siest
G
G
Siest
D000464:Alginates; D000818:Animals; D003902:Detergents; D004800:Enzymes, Immobilized / Q000737:chemistry*; D005782:Gels; D005965:Glucuronates / Q000737:chemistry*; D014453:Glucuronosyltransferase / Q000378:metabolism*; D007700:Kinetics; D008244:Lysophosphatidylcholines; D008297:Male; D008862:Microsomes, Liver / Q000737:chemistry / Q000201:enzymology*; D008863:Microspheres; D010316:Particle Size; D051381:Rats; D011919:Rats, Inbred Strains; D015215:Zidovudine / Q000737:chemistry
D000464:Alginates; D003902:Detergents; D004800:Enzymes, Immobilized; D005782:Gels; D005965:Glucuronates; D008244:Lysophosphatidylcholines; D015215:Zidovudine; D014453:Glucuronosyltransferase