Biomedical Engineering Reference
In-Depth Information
Moreover, significant difference in extraction recovery has been observed for an
analyte and its deuterated internal standard. For example, the extraction recoveries
of haloperidol and d
4
-haloperidol were 72 % and 44 %, respectively [
19
] . The large
difference in extraction recovery could be due to differences in the aforementioned
physicochemical properties, such as p
K
a, or hydrogen-deuterium exchange.
Since no similar issue has been reported for other SIL internal standards, such
as
13
C and
15
N labeled internal standards, it is therefore necessary to distinguish
deuterated internal standards from other SIL internal standards. Nevertheless, one
should always have an open mind while using SIL internal standards in LC-MS
bioanalysis.
3.3
Internal Standard Response Variations During Incurred
Sample Analysis
3.3.1
Why Monitor Internal Standard Response Variations?
Bioanalytical chemists usually have contradictive expectations regarding internal
standard response variations. In one hand, internal standard response variations in
bioanalysis by LC-MS are somewhat expected due to the purpose of using internal
standards, i.e., correction of variations (otherwise there is no need to use an internal
standard), particularly if considering the many differences between spiked samples,
e.g., calibration standards (CS) and quality control (QC) samples prepared in a
pooled control blank plasma, and incurred samples (samples collected from dosed
subjects in a preclinical or clinical study, also termed as study samples), such as
absence of metabolites and formulation related components in spiked CS and QC
samples as well as difference in number of matrix lots and freeze-thaw cycles (Table 1).
On the other hand, too much variation in internal standard responses during bio-
analysis could trigger doubt in the integrity of the quantitative results obtained
because the same amount of an internal standard is added to all the samples. To
maintain the integrity of a study and to avoid economic losses, it is therefore critical
to monitor IS response variations during bioanalysis and to quickly identify the root
causes if variations are observed, especially for those causes that affect an analyte
and its internal standard differently.
3.3.2
How to Monitor Internal Standard Response Variations?
Though monitoring of internal standard response variations in incurred sample
analysis is necessary and has been recommended in the literatures (e.g., [
26
] ), no
agreement was reached on the inclusion of internal standard criteria or on the mag-
nitude of acceptable internal standard precision during the Third American
Association of Pharmaceutical Scientists (AAPS)/Food and Drug Administration
(FDA) Bioanalytical Workshop in 2007 [
9
]. It was only suggested that objective
