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Can someone please shed some light on how to inhibit glucuronidation
reaction without addition of organic solvents. I read some literature
that the addition of UDP inhibits the reaction. Not sure whether it
is UDP or UDPGA that is added.
Question:
If UDP or UPDGA inhibits formation of glucuronidation reaction then
what is the mechanism involved?
Is it UDP or UDPGA that inhibits the reaction?
Listed below is the reference that recommends using 10 mM UDP to
inhibit glucuronidation.
Reference: A simple in vitro model to study the stability of
acylglucuronides
Leung et al.. Journal of Pharm and Tox. Methods, 55, (2007) 91-95
Thanks a bunch
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The following message was posted to: PharmPK
Dear Dr Gallagher,
It would appear that not all UGT isoforms are equally sensitive to
the presence of solvents. Let me suggest the following reading:
Uchaipichat et al
HUMAN UDP-GLUCURONOSYLTRANSFERASES: ISOFORM SELECTIVITY AND KINETICS
OF 4-METHYLUMBELLIFERONE AND 1-NAPHTHOL GLUCURONIDATION, EFFECTS OF
ORGANIC SOLVENTS, AND INHIBITION BY DICLOFENAC AND PROBENECID
Drug Metab Dispos 32:413-423, 2004
Hope this helps.
Best regards,
Henri MERDJAN
Head of Drug Metabolism and Pharmacokinetics
NOVEXEL S.A., Parc Biocitech, 102 Route de Noisy
F-93230 Romainville, France
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Dear S Gallagher,
I have come across the following inhibitors of UGT-activity:
-Non specific inhibitors:
--
-probenecid (ref. Markowitz et al. Clin Pharmacol Ther 2002; 71(1):
30-8),
-hepatic UGTs are inhibited by: borneol or 7,7,7- triphenylheptyl-
UDP (ref. Sallustio et al. Curr Drug Metab 2000,1,163-180).
More or less specific inhibitors for:
--
-UGT1A1- inhibited by bilirubin (also UGT1A4), oestradiol (but also
UGT1A9),
-UGT1A3 - inhibited by imipramine (but also UGT1A4),
-UGT1A4 - specifically inhibited by hecogenin,
-UGT1A6 - by 4-nitrophenol (but also UGT1A9),
-UGT1A8 - by emodin (but also UGT1A10),
-UGT2B4 - by hydroxyeholic acid,
-UGT2B7 - by morphine.
References:
[Watanabe,Drug Metab Dispos. 2002; 30(12):1462-9],
[Ghosal,Drug Metab. Dispos. 2004; 32(2):267-71. ],
[Uchaipichat,Drug Metab. Dispos. 2006; 34(3):449-56.],
[Kuehl,Drug Metab. Dispos. 2005; 33(7):1027-35],
Hope any of this helps,
Best regards,
Jurij Trontelj
Faculty of pharmacy,
University in Ljubljana
Slovenia
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Dear Sue,
I am looking at the REPLY to your question and I think folks are
misunderstanding your question. If I understood correctly you want to
use the inhibitor to stop the reaction to go further so you can carry
out glucuronidation (most likely acylglucuronidation) stability as
they are responsible to cause potential toxicity. Mainly I believe
your goal is to synthesize the metabolite and make sure the reaction
is stopped before carrying out stability assay.
In my opinion, the literature reference cited adds 10 mM final
concentration of UDP (uridine- 5-diphosphate) to inhibit the
reaction. UDP will compete with enzyme and 10 mM concentration is
high enough to exhaust enzyme activity and therefore the reaction is
stopped.
Best regards,
Anila
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All,
Anila, Yes, you are correct in understanding my question. I would
like to understand the mechanism involved. Interestingly, the
literature does not mention about UDP manufacturer or full name.
I would like to know if anyone else has different theory than Anila.
Please reply.
Thank you
Sue
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The following message was posted to: PharmPK
Hi Sue,
It looks like the UDP competes with UDPGA for binding to the enzyme.
As enzymes are in theory just catalysts, they can be blocked by any
structurally similar substance. Generally they have a high affinity
for substrates and cofactors and a low affinity for the products; in
this case, UDP has a higher affinity than UMP and competes
effectively with UDPGA but is inactive as a cofactor, so no
glucuronidation can take place. The following ref describes how UDP
competes with UDPGA but not their substrate (entacapone) on UGT1A9:
DMD 33:1017-1026, 2005 KINETIC CHARACTERIZATION OF THE 1A SUBFAMILY
OF RECOMBINANT HUMAN UDP-GLUCURONOSYLTRANSFERASES
Leena Luukkanen, et al.
Thanks for bringing this up. It is an interesting technique.
Regards.
Ted
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