Supplementary Material

Figures List

Fig.S1. Chemical structure of ponatinib

Fig.S2.Chromatograms of ponatinib and warfalin in rat plasma

Fig.S3.Chromatograms of ponatinib and warfalin in rat urine

Fig.S4.Chromatograms of ponatinib and warfalin in rat feces

Fig.S5. Mean urinary/fecal recovery versus time profiles of ponatinib after single i.g. administration ponatinib 15 mg/kg ponatinib to rat(n=6)

Figure.S1. Chemical structure of ponatinib

Figure.S2. Typical chromatograms of ponatinib and warfalin in rat plasma: A, blank rat plasma sample; B, blank plasma sample spiked with Warfalin (50ng/mL in plasma); C, blank plasma sample spiked with ponatinib (500ng/mL in plasma) and warfalin (50ng/mL in plasma); and D, a rat plasma sample at 10h after intragastric administration of ponatinib.

Figure.S3. Typical chromatograms of ponatinib and warfalin in rat urine: A, blank rat urine sample; B, blank urine sample spiked with warfalin (50ng/mL in plasma); C, blank urine sample spiked with ponatinib (500ng/mL in urine) and warfalin (50ng/mL in urine); and D, a rat urine sample at 12h after intragastric administration of ponatinib.

Figure.S4. Typical chromatograms of ponatinib and warfalin in rat feces: A, blank rat feces sample; B, blank fece sample spiked with warfalin (50ng/mL in feces); C, blank feces sample spiked with ponatinib (200ng/mL in feces) and warfalin (50ng/mL in feces); and D, a rat feces sample at 12h after intragastric administration of ponatinib.

Fig. S5. Mean urinary/fecal recovery versus time profiles of ponatinib after single i.g. administration ponatinib 15 mg/kg ponatinib to rat (n=6), A, Mean urinary recovery versus time profiles of ponatinib; B, Mean fecal recovery versus time profiles of ponatinib.

Table. S1
Calibration curves of Ponatinib in rat plasma/urine/feces samples(n=5).
Biological matrix / Compound / Standard curves / r / Linear ranges ng/mL(ng/g) / LLOQ Precision(RE)
Plasma / Ponatinib / y =0.0187x +0.00546 / 0.9987 / 1-1000 / 0.0
Urine / Ponatinib / y =0.00605x +0.0265 / 0.9971 / 5-1000 / -4.6
Feces / Ponatinib / y =0.00183x +0.0236 / 0.9982 / 5-5000 / -2.9
Table.S2
Extraction recovery and matrix effect data for Ponatinib (n=5).
Plasma / Urine / Feces
Nominal concentration (ng/mL) / 2 / 50 / 1000 / 10 / 100 / 1000 / 10 / 100 / 1000
Recovery（%） / 65.73 / 59.32 / 61.12 / 85.32 / 75.78 / 80.23 / 108.71 / 93.66 / 97.08
Matrix effect（%） / 97.31 / 85.95 / 90.75 / 84.54 / 92.41 / 94.47 / 101.56 / 103.33 / 99.14
Table.S3
Intra-batch and inter batch precision and accuracy for the quantification of Ponatinib (n=5) in rat plasma.
Nominal concentration (ng/mL) / intra-batch / inter-batch
Precision (RSD%) / Accuracy (RE%) / Precision (RSD%) / Accuracy (RE%)
Plasma / 2 / 7.89 / 3.98 / 6.12 / 1.15
50 / 2.22 / 9.24 / 3.62 / 9.03
1000 / 3.82 / -4.55 / 2.59 / -5.95
Urine / 10 / - / - / 3.74 / 3.87
100 / - / - / 1.77 / 8.30
1000 / - / - / 6.94 / 2.51
Feces / 10 / - / - / 3.04 / -17.05
100 / - / - / 1.86 / 0.75
1000 / - / - / 5.39 / -1.42
Table.S4
Stability of Ponatinib in ratbio-samples under different storage conditions (n = 5).
Storage conditions / Nominal concentration (ng/mL) / Concentration determined (ng/mL) / Precision (RSD%) / Accuracy (RE%)
Plasma / Three freeze-thaw cycles / 2 / 2.28 / 4.24 / 13.85
50 / 56.27 / 3.15 / 12.53
1000 / 917.83 / 6.58 / -8.22
Short-term
(24 h at 20 ℃) / 2 / 2.08 / 11.28 / 4.04
50 / 53.88 / 2.08 / 7.77
1000 / 947.15 / 4.19 / -5.29
Long-term
(30 d at −80 ℃) / 2 / 2.08 / 2.96 / 3.99
50 / 56.85 / 0.27 / 13.70
1000 / 951.16 / 1.42 / -4.88
Autosampler
(24 h at 4 ℃) / 2 / 2.07 / 8.17 / 3.70
50 / 55.70 / 2.17 / 11.41
1000 / 1004.12 / 4.15 / 0.41
Urine / Three freeze-thaw cycles / 10 / 10.62 / 4.43 / 6.25
100 / 102.44 / 1.90 / 2.44
1000 / 1028.34 / 1.75 / 2.83
Short-term
(24 h at 20 ℃) / 10 / 10.31 / 2.40 / 3.06
100 / 104.06 / 2.14 / 4.06
1000 / 997.18 / 4.94 / -0.28
Long-term
(30 d at −80 ℃) / 10 / 10.75 / 3.89 / 7.45
100 / 102.97 / 2.26 / 2.97
1000 / 1000.36 / 5.14 / 0.04
Autosampler
(24 h at 4 ℃) / 10 / 11.02 / 6.45 / 10.21
100 / 103.43 / 3.71 / 3.43
1000 / 1032.14 / 1.47 / 3.21
Feces / Three freeze-thaw cycles / 10 / 9.89 / 14.97 / -1.15
100 / 108.07 / 1.02 / 8.07
800 / 783.36 / 2.11 / -2.08
Short-term
(24 h at 20 ℃) / 10 / 9.88 / 14.62 / -1.18
100 / 103.01 / 1.24 / 3.01
800 / 817.34 / 2.68 / 2.17
Long-term
(30 d at −80 ℃) / 10 / 10.09 / 9.88 / 0.94
100 / 104.74 / 2.59 / 4.74
800 / 821.52 / 3.54 / 2.69
Autosampler
(24 h at 4 ℃) / 10 / 10.30 / 8.75 / 2.96
100 / 100.59 / 1.89 / 0.59
800 / 834.23 / 3.25 / 4.28