counting (MSC) and other recently developed liquid radiochromatographic
techniques.
10.3 New Radiochromatography Techniques
10.3.1 HPLC-MSC
MSC was recently introduced as an off-line liquid radiochromatographic
detection technique for radioactive metabolite profiling (Boernsen et al., 2000;
Bruin et al., 2006; Kiffe et al., 2003; Wallace et al., 2004; Zhu et al., 2005b,
2005c). Compared to HPLC–LSC, HPLC–MSC not only increases analytical
throughput and sensitivity but also reduces radioactive waste and manual
operations involved in LSC vial handling. The HPLC–MSC has become the
method of choice for analysis of low level radioactive metabolites in some
metabolism laboratories (Boernsen et al., 2000; Bruin et al., 2006; Kiffe et al.,
2003; Wallace et al., 2004; Zhu et al., 2005b). In HPLC–MSC analysis (Fig.
10.3a), HPLC effluent is collected into 96-well microplates, and then
evaporated using a speed vacuum system. The radioactivity of the residue in
the 96-well plates is determined by counting up to 12 wells at a time with a
microplate scintillation counter. Two types of MSC instruments, TopCount
and MicroBeta counter (Nedderman et al., 2004; Zhu et al., 2005b), are
commercially available. The TopCount instrument uses deep-well LumaPlates,
in which yttrium silicate scintillators are deposited at the bottom of each well.
The MicroBeta counter uses 96-well Scintiplates^1 that consist of white frames
with clear wells embedded with solid scintillators. The radiodetection
sensitivity of TopCount is slightly better than MicroBeta counter, while
radiolabeled components can be readily recovered from Scintiplates but not
from LumaPlates. In HPLC–MSC analysis, counts per minutes (CPM) values
TABLE 10.1 Sensitivity of various liquid radiochromatographic techniques.
Radio-
detection
Background
(CPM)Counting
efficiency
(%)Counting
time
(min)Limit of
detectiona
(DPM)Limit of
quantificationb
(DPM)LC–LSC 25 90 10 10 31
LC–RFD 15 70 5–10 s 250–500 750–1500
LC–MSCc 270105 15
Stop-flow RFDd 15 70 1 25–50 75–150
LC–AMSe 0.0001
aLimit of detection (LOD) for LSC, RFD, MSC and stop-flow RFD was calculated based on
Equation 10.1. The parameters used for the calculation are listed Table 10.1.
bLimit of quantification (LOQ) was calculated based on an equation presented in reference (Zhu
et al., 2005b).
cTopCount instrument was used.
dStop by-fraction mode was utilized.
eSee references (Brown et al., 2005, 2006).
NEW RADIOCHROMATOGRAPHY TECHNIQUES 293