Biomedical Engineering Reference
In-Depth Information
Substantial matrix effects can be specific for individual patients' samples, e.g.
due to co-administered drugs or other xenobiotics. It is in general tried to detect
such samples by assessing the peak areas of the internal standard compound over a
whole series: Outliers with respect to the peak area of the internal standard may
indicate unusually pronounced matrix effects in an individual sample. In such cases
it is speculative if the evident matrix effects affect analyte and internal standard to
an identical degree, and it is up to the valuation of the analyst if quantitative results
of such samples may be reported or not. Re-analysis after dilution can be useful in
case of evident matrix effects—given sufficiently high analyte concentration in the
respective sample.
Such decisions are always more or less arbitrary since in many routinely used
LC-MS/MS instruments the “normal” variation of IS peak-areas between subse-
quent samples in a series can be substantial. Moreover, it is always uncertain if
matrix effects in an individual sample might impact the target analyte but not at all
the internal standard compound. This consideration underscores the importance of
the appropriateness of internal standard compounds for reliable LC-MS/MS results.
We believe, that matrix related modulation of ionization will remain an important
issue in clinical LC-MS/MS and has to be approached with a multiple strategy span-
ning the whole life time of an assay:
•
Matrix effects have to be detected by systematic experiments during method
development.
Matrix effects have to be minimized by adequate sample preparation and
•
chromatography.
Matrix effects have to be compensated by use of most appropriate internal stan-
•
dard materials.
3.2
In-source Transformation
API techniques as electrospray ionization (ESI) and atmospheric pressure chemical
ionization (APCI) are generally looked upon as “soft.” Nevertheless, if weak bonds
are present, molecules can be disintegrated during the process of ionization before
the ion enters the mass spectrometer. In bioanalysis, such “in source transforma-
tion” is particularly prominent if conjugated metabolites, i.e. if glucuronides or sul-
phates of target analytes are present in a sample [
53
] . Consequently, analytical
inaccuracy can be easily introduced whenever a target analyte co-elutes together
with its conjugate metabolites. (e.g. mycophenolic acid (MPA) with MPA-
glucuronide) [
34
]. In such a case, analyte molecules may be generated from the
conjugate metabolites within the ion source by fragmentation. By principle, this
effect cannot be compensated by the subsequent SRM based mass spectrometric
analysis of the once generated ions. Hence, in general the risk of in-source transfor-
mation related inaccuracy is particularly given whenever the selectivity of the cho-
sen chromatographic process is too low to separate a target analyte from its more
