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The following message was posted to: PharmPK
Dear all,
can you suggest regarding,
If drug is getting converted into metabolite, if we use mixed drug &
metabolite spiking solutions, as per area ratio, all the Calibration
Curve (CC) standards and Quality Control (QC) samples were found
within the acceptance criteria, STD BL and STD Zero were found clean
(not presence of drug & metabolite),
So what is the necessity to restrict the conversion of drug into
metabolite?
hoping for best reply
kanchan
veeda clinical research
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Hello,
I am not clear about your question.
If you ask that why do we need to restrict the conversion of drug into
metabolite during bioanalysis...depending on the matrix there could be
a possibility of ex vivo hydrolysis that can lead to underestimation
of the data.. In case of prodrugs, the same can lead to over estimation.
I am sorry if I had misunderstood your question
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Dear soni,
Even though the calibration curve is passing with
combined drug and metabolite spiking, you need to do some qualitative
experimentation, try spiking your parent compound and metabolite
seperately into matrix and prepare calibration curve for both
metabolite and parent compound. After extraction and reconstitution is
done try loading the metabolite spiked samples as calibration curve
standards, and parent drug spiked samples as unknown samples. In the
quantitation method try to quantify only metabolite, then you will
come to know what amount of your drug is getting converted into
metabolite. Is the metabolite calibration curve passing as such
without the addition of drug. Try doing this experiment and you will
end up with conclusion.
regards,
Vijay,
Aurigene discovery technologies ltd,
Bangalore.
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The following message was posted to: PharmPK
Dear Sir,
i got your answer but my doubt is why we are trying to not to convert
drug into metabolite, what will happen if some parts of drug is
converted into metabolite.
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Dear soni,
What I understand from your statement is that your analyte is
getting converted in to metabolite, but the % conversion is not
significant to effect the area ratio of the analytes and hence the
calibration curve and the QC samples are within acceptance limit. If
this is the case than in my opinion there is no harm in accepting the
results, but same should be mentioned in the pre-study method
validation report. As there can be concern about the stability of
analytes for long term in matrix (i.e. For overall study duration
Clinical + Analytical phase). Also in accordance to 3 AAPS/FDA
Bioanalytical workshop recommendations, you will have to generate a
incurred study samples stability data to prove and support your
statement.
With Due Regards,
Kuldeep Sharma
DMPK Lab.
Jubilant Biosys Ltd.
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The following message was posted to: PharmPK
Dear Soni,
It is very important to avoid the conversion to have the actual
concentration of metabolite and parent drug during study sample
analysis.
Linearity and QC's are passing may be because your calibration
standards and QC samples concentrations were same for both parent and
metabolite or proportional increase in concentration with respect to
metabolite in all level.
From regulatory perspective it is necessory to avoid the conversion
using appropriate stabilizer .
Specificity experiment should be performed separately for both parent
and metabolite.Parent specificity experiment should be performed in
presence of metabolite and metabolite specificity experiment should be
performed in presence of parent.
Also ensure the following during experimentation
Conversion in biological matrix?
Conversion in spiking stock?
Conversion in source?
Regards
Raghavendra Shetty
Manipal AcuNova LTD
Manipal
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The following message was posted to: PharmPK
You do need to halt conversion upon collection. Your preservative(s)
should
arrest metabolism of your product and its metabolite.
It must be able to do this through at least 3 (5-10) freeze thaw cycles,
including at least 30 minutes at room temperature and 12 hrs at the
storage
condition.
Analytes in samples must be stable at room temperature for some period
of
time (1hr-16 hrs). Processing over ice may be necessary or the
samples may
need to be heat denatured (if the analyte can tolerate this
treatment )56C
for 30 min.
The preservative should not interfere with any downstream processing or
analysis.
Were the calibrators and QC's prepared in the same matrix, treated the
same
way-as the samples. Often times the plasma used to prep QC and
calibrators
may be filtered and or heat denatured and may not reflect the true
activity
of the matrix towards your compound.
And we haven't yet addressed whole blood recovery/stability.
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