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Thermo Scienti c Poster Note
•
PN63784_E 03/13S
Analyte
Retention
time
Precursor
Ion
Product
ion
TL/CE
Product
ion
CE
Ion
Ratio
Bortezomib
3.11
367.1
226.0
192/-18
208.0
-28
60
Dasatinib
3.01
488.2
401.0
184/-29
231.9
-38
40
Erlotinib
3.12
394 .2
277.9
136/-21
336.0
-22
40
Imatinib
2.96
494.3
394.1
170/-25
222.0
-27
20
D8-Imatinib
2.96
502.3
394.1
170/-25
Lapatinib
3.28
581.1
349.9
185/-36
364.9
-38
75
Nilotinib
3.26
530.1
288.9
199/-29
261.0
-42
45
Sorafenib
3.59
465.1
251.9
176/-31
270.0
-21
75
Sunitinib
3.06
399.2
282.9
134/-28
326.0
-20
60
Vandetanib
2.99
475.1
83.1
142/-32
111.9
-64
15
Analyte
Slope
Inte
BORT
Mean
CV
0.000179
14.8
0.00
1
DASA
Mean
CV
0.000989
9.3
-0.0
1
ERLO
Mean
CV
0.00820
7.1
0.
4
IMAT
Mean
CV
0.0198
5.4
-0.0
1
LAPA
Mean
CV
0.000286
11.5
-0.0
NILO
Mean
CV
0.002519
3.88
-0.
9
SORA
Mean
CV
0.000657
10.8
-0.0
2
SUNI
Mean
CV
0.00514
6.9
0.0
1
VAND
Mean
CV
0.0000199
12.2
-0.0
1
ntional chemotherapy drugs
. Targeted therapies include
asatinib, Nilotinib, Sunitinib,
rlotinib) that present better
cancer drugs.
and sensitive method for the
Dasatinib, Nilotinib, Sunitinib,
ib) in plasma samples using
entific (Courtaboeuf, France)
ation was performed on an
nalytical column placed in a
phic system was coupled to
trometer (MS) from Thermo
lectrospray ionization (ESI)
(Thermo Fisher Scientific,
M ammonium formate buffer
acetonitrile with 0.1 % (v/v)
elivered using the following
95:5 (A:B) maintained for 0.5
in 5:95 (A:B) at 2 minutes,
ash using 100% of phase C
inutes to 95:5 (A:B) at 7.5
es for equilibration. The flow
was set at 50
°
C and the
ode, capillary temperature:
le 1; spray voltage: 3500 V;
and 25 (arbitrary units),
was 1.5 mTorr. Data are
ratio for each analyte are
red at 1 mg/ml by dissolving
of methanol. Stock solutions
rking solutions containing all
ation standards ranging from
50 to 3 500 ng/mL for the
ation of the 5 quality controls
Only QCs at 7, 75 and 150 ng/mL were used for BORT, DASA and SUNI while QCs
at 75, 150, 750 and 1 500 ng/mL were used for the other TKIs. A 0.5 mg/mL d8-
imatinib, internal standard (IS) stock solution was prepared by dissolving 1 mg of the
chemical in 2 ml of methanol. Plasma calibration samples and three plasma quality
control (QC) samples were prepared by adding the appropriate volume of each
working solution to blank plasma.
Table 1:
Retention time, precursor molecular ion/product ion for quantification,
precursor molecular ion/product ion for confirmation and detection parameters (tube
lens voltage (TL)/collision energy(CE)) for each analyte
Plasma sample extraction procedure
Aliquots of 50
µ
l of the plasma unknowns, blank, calibration standards and QCs were
placed in appropriate labeled 1.5 mL microcentrifuge tubes and mixed with 200
µ
l of
acetonitrile containing 20 ng/mL IS. After automatic vortexing for 10 minutes, each
sample was centrifuged at 6 000g at 4
°
C for 15 minutes. Hundred microliters of
supernatant were diluted two-fold using the mobile phases A and B in a 50/50 (v/v)
ratio. After capping and vortexing, the vials were transferred into the autosampler
tray that was maintained at +4
°
C. Twenty-five microliters aliquots of the extract were
injected into the HPLC system.
Results
Chromatograms
The proposed method enables the simultaneous quantification of commonly used
TKIs in 50
µ
L-plasma aliquots by liquid chromatography coupled with tandem MS.
Typical chromatographic profiles of the highest calibrator sample containing all are
shown in Fig. 2.
Internal standard, calibration curve and lower limit of
quantification
Imatinib-D8 was used as IS with a satisfactory chromatographic profile and a
negligible memory effect. Calibration curves over the entire ranges of concentrations
were best described by 1/x weighted linear regression of the peak-area ratio of each
TKI to IS
versus
the concentrations of the respective TKI in each standard sample.
This model was optimal for the 9 TKIs. Standar
biological plasmas (EDTA), were performed in pla
The assay proved to be linear and acceptable, a
>0.99 for each of the twenty standard curves exce
(Table 2).
A linearity test has been performed to compare th
deviations of the back-calculated values to each
low and the high standard curves. Then the acc
analyte. In all cases, slopes of these linear curve
1.019 and statistics showed slopes significantly
LLOQ was established at 2 ng/mL for BORT, DAS
others drugs in human plasma.
Fig. 1: Chromatogram of the highest calibrator sa
Table 2:
Data detailing the slopes, intercepts, coef
(n=20).