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Dear Dr Sivakumar.
The problem in your analysis showing lower extraction efficiency by
LCMS under SRM and higher extraction efficiency by HPLC could be
because of degradation of your molecule in plasma. If your molecule
undergoes simple change in its structure say reduction (ie) addition of
two more hydrogen to your molecule then there wouldn't be much of a
change in its polarity and hence will co-elute at the same retention
time in your HPLC run while your LCMS run is run under SRM it detects
only the molecule in its undegraded state and hence indicating reduced
Aurigene discovery Technologies
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It is difficult to say whether a degradation product will coelute with
your analyte. If your LC conditions are "strong" then it is more
probable for coelution to occur. Another, and I think more likely
reason for lower extraction recovery with LCMS, is matrix associated
ion suppression. This might be a significant problem if you are dealing
with human plasma samples. Human plasma can cause different ion
suppression effects, depending on it's source.
Here are some good references on the subject and how to deal with it
Matuszewski, BK; Constanzer, ML; Chavez-Eng, CM. Strategies for the
assessment of matrix effect in quantitative bioanalytical methods based
on HPLC-MS/MS. Analytical Chemistry. 2003; 75, 3019.
Muller, C; Schafer, P; Stortzel, M; Vogt, S; Weinmann, W. Ion
suppression effects in liquid chromatography-electrospray-ionisation
transport-region collision induced dissociation mass spectrometry with
different serum extraction methods for systematic toxicological
analysis with mass spectra libraries. J Chromatogr B Analyt Technol
Biomed Life Sci. 2002; 773, 47.
Another comment. It's always helpful to indicate the detector used in
a Liquid Chromatographic (LC) method. The term "HPLC" only refers to
the separation and says nothing about the detector used.
Howard Hendrickson, PhD
Research Assistant Professor
University of Arkansas for Medical Sciences
Department of Pharmacology & Toxicology
4301 West Markham St #611
Little Rock AR 72205
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