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dear members,
we are developing a method for a drug and its two metabolites having a
pka 8.9 (basic) on API 3000.Our problem is when we inject its aqueous
mixture, the response is stable. when we injected the extracted
(plasma) sample, we found the response is getting down in the
subsequent injections on continuous injectons from the same vial.we
have changed many columns and mobile phase combinations(pH 4.5 To 6.5)
but found no change in the decreasing trend. we are following SPE
method.when we injected the samples without column the aqueous samples
were stable but the same decreasing pattern found with the extracted
samples. our mobile phase is MEOH: 2mM ammonium acetate (80:20) and
reconstitution solution is MEOH:water (80:20).your suggestions will be
appreciated and if needed i ll provide further information on our
exercises.
thank you in advance
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how about sample (w/o plasma) stability/solubility in 80:20
MeOH:water? If the signal remains steady, then it could be something
from the SPE. You may want to double-check your SPE method.
Sherwin
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The following message was posted to: PharmPK
The decreasing pattern might be due to particulate build-up in the
ionization chamber leading to a decrease in the sensitivity of the
method. I would suggest trying the following:
1- To test this possibility, try few injections from the same vial
till the response drastically decreases. Then clean the ionization
chamber and re-inject from the same vial to see if you get the initial
response.
2- If this is the case, then may be using a deuterated form of your
compound in question as an internal standard could help, since its
response might degrade with injections proportionally to the analytes
decrease in response. 3- If this is not possible, then may be a fewer
number of injections per run, with ionization chamber clean-up in
between runs might help.
4- A lower buffer concentration along with a switch to amm formate
instead of acetate might help in increasing the number of runs before
the response significantly degrades.
Regards,
Maggie
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Dear Narendran,
Have you looked at the stability of sample in plasma? Knowing
structure or atleast chemical class will help to understand
instability issue.
Regards,
Vinayak
Vinayak Nadiger
Manager , Bioanalytical Chemistry
11 Biopolis Way, Helios #08-05
Singapore 138667
E Mail: vnadiger.-at-.combinatorx.com
Website:www.combinatorx.com
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The following message was posted to: PharmPK
Dear Narendran
Did you look at adsorption to the vial, may be silinized vials will
solve your problem. Another option is maybe try a different solvent
to recostitute the samplr (IPA, EtOH,..etc)
Good luck
Oliver
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