Preclinical Pharmacokinetic Studies of a Novel Diuretic Inhibiting Urea Transporters
Abstract
:1. Introduction
2. Results and Discussion
2.1. LC–MS/MS Method Development and Optimization
2.2. LC–MS/MS Validation
2.2.1. Selectivity
2.2.2. Linearity
2.2.3. Accuracy and Precision
2.2.4. Recovery and Matrix Effect
2.2.5. Stability Evaluation
2.2.6. Dilution Integrity
2.2.7. Carry-Over
2.3. Pharmacokinetic and Pharmacodynamic Study
2.4. Tissue Distribution
2.5. Excretion Experiments
2.6. Plasma Protein Binding and Blood Plasma Ratio
3. Materials and Methods
3.1. Chemicals and Materials
3.2. LC–MS/MS Instrument Conditions
3.3. Sample Preparation
3.4. Method Validation
3.5. Pharmacokinetic Study in Rats
3.6. Tissue Distribution Study
3.7. Excretion Experiment
3.8. Plasma Protein Binding Study
3.9. Blood Plasma Ratio Study
3.10. Statistical Analyses
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
Sample Availability
References
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Time (min) | Mobile Phase A (%) | Mobile Phase B (%) |
---|---|---|
0.0 | 95 | 5 |
0.5 | 30 | 70 |
1.0 | 0 | 100 |
3.0 | 0 | 100 |
3.1 | 95 | 5 |
5.0 | 95 | 5 |
Matrix | y = a × x + b * | ||
---|---|---|---|
a | b | r | |
Plasma | 0.01201 | 0.00391 | 0.9983 |
Urine | 0.00965 | 0.00204 | 0.9997 |
Feces | 0.00967 | 0.00218 | 0.9996 |
Bile | 0.00708 | 0.00325 | 0.9989 |
Kidney | 0.01200 | 0.00390 | 0.9995 |
Heart | 0.01180 | 0.00314 | 0.9997 |
Lung | 0.01090 | 0.00400 | 0.9996 |
Spleen | 0.01060 | 0.00646 | 0.9997 |
Muscle | 0.01190 | 0.00332 | 0.9998 |
Skin | 0.01030 | 0.00640 | 0.9994 |
Bone | 0.00926 | 0.00694 | 0.9997 |
Brain | 0.01170 | 0.00395 | 0.9999 |
Fat | 0.01100 | 0.00259 | 0.9997 |
Liver | 0.00961 | 0.00599 | 0.9996 |
Testis | 0.01000 | 0.00878 | 0.9997 |
Matrix | Sample | Nominal Conc. (ng/mL) | Intraday | Interday | ||
---|---|---|---|---|---|---|
RE (%) | CV (%) | RE (%) | CV (%) | |||
Plasma | LLOQ | 1 | 1.2 | 6.0 | −4.6 | 8.4 |
LQC | 3 | −3.9 | 2.1 | −6.8 | 5.1 | |
MQC | 100 | 0.3 | 1.7 | −2.8 | 5.9 | |
HQC | 800 | −3.0 | 1.6 | −3.4 | 3.8 | |
Urine | LLOQ | 1 | −14.9 | 5.8 | −15.2 | 0.4 |
LQC | 3 | −11.4 | 8.0 | −10.6 | 2.1 | |
MQC | 100 | 3.3 | 1.7 | −0.7 | 6.3 | |
HQC | 800 | −3.0 | 4.3 | −4.7 | 3.0 | |
Feces | LLOQ | 1 | 7.0 | 2.3 | 9.1 | 2.4 |
LQC | 3 | 2.6 | 5.4 | 7.4 | 3.9 | |
MQC | 100 | 11.2 | 4.1 | 7.5 | 3.2 | |
HQC | 800 | 3.9 | 4.2 | 4.5 | 1.2 | |
Bile | LLOQ | 1 | 15.2 | 2.9 | 7.9 | 11.2 |
LQC | 3 | −12.1 | 4.2 | −9.7 | 3.5 | |
MQC | 100 | −7.8 | 2.8 | −8.0 | 0.6 | |
HQC | 800 | −11.2 | 3.5 | −11.6 | 0.8 |
Matrix | Quality Control | Nominal Conc. (ng/mL) | Recovery | Matrix Effect | ||||
---|---|---|---|---|---|---|---|---|
Mean (%) | SD (%) | CV (%) | Mean (%) | SD (%) | CV (%) | |||
Plasma | LQC | 3 | 97.6 | 6.0 | 6.1 | 105.4 | 2.8 | 2.5 |
MQC | 100 | 98.2 | 4.5 | 4.5 | 107.5 | 2.1 | 2.1 | |
HQC | 800 | 110.7 | 4.7 | 4.7 | 103.7 | 4.2 | 4.2 | |
Urine | LQC | 3 | 101.8 | 7.6 | 7.4 | 98.2 | 5.3 | 4.8 |
MQC | 100 | 94.4 | 8.4 | 8.9 | 99.3 | 2.6 | 2.4 | |
HQC | 800 | 100.9 | 3.6 | 3.5 | 98.0 | 0.9 | 0.8 | |
Feces | LQC | 3 | 98.5 | 8.0 | 7.5 | 102.1 | 8.0 | 6.8 |
MQC | 100 | 105.6 | 2.8 | 5.4 | 95.0 | 2.4 | 5.0 | |
HQC | 800 | 103.8 | 10.8 | 9.4 | 97.1 | 9.6 | 7.9 | |
Bile | LQC | 3 | 94.1 | 10.0 | 10.6 | 88.8 | 9.0 | 8.1 |
MQC | 100 | 88.7 | 3.1 | 3.5 | 82.8 | 9.6 | 8.6 | |
HQC | 800 | 92.5 | 10.3 | 11.1 | 83.5 | 5.5 | 5.0 |
Matrix | Quality Control | Nominal Conc. (ng/mL) | 25 °C for 4 h | 4 °C for 24 h | Three Freeze–Thaw Cycles | −80 °C for 30 days | ||||
---|---|---|---|---|---|---|---|---|---|---|
RE (%) | CV (%) | RE (%) | CV (%) | RE (%) | CV (%) | RE (%) | CV (%) | |||
Plasma | LQC | 3 | −4.9 | 2.0 | 2.8 | 5.6 | −3.6 | 1.4 | 1.1 | 5.6 |
MQC | 100 | −2.5 | 2.2 | 3.7 | 1.3 | −4.0 | 2.1 | 3.7 | 1.3 | |
HQC | 800 | −3.1 | 0.5 | −4.5 | 1.7 | −7.6 | 2.1 | −4.5 | 1.7 | |
Urine | LQC | 3 | 3.2 | 6.7 | −8.3 | 3.1 | 5.8 | 10.7 | 6.2 | 3.1 |
MQC | 100 | 4.3 | 1.7 | 2.5 | 1.7 | −5.2 | 4.2 | 2.5 | 1.7 | |
HQC | 800 | −2.3 | 4.4 | −2.3 | 3.9 | −1.5 | 3.5 | −2.3 | 3.9 | |
Feces | LQC | 3 | −5.2 | 4.9 | 3.9 | 7.4 | −5.2 | 2.9 | 1.2 | 7.4 |
MQC | 100 | 1.3 | 3.4 | 4.8 | 3.2 | −3.2 | 3.1 | 4.8 | 3.2 | |
HQC | 800 | −2.4 | 0.9 | −5.3 | 1.5 | −9.1 | 4.9 | −5.3 | 1.5 | |
Bile | LQC | 3 | −14.9 | 4.7 | −14.7 | 5.8 | −5.8 | 10.7 | −2.1 | 5.8 |
MQC | 100 | −5.1 | 9.0 | −3.9 | 2.2 | −5.2 | 4.2 | −3.9 | 2.2 | |
HQC | 800 | −7.8 | 3.7 | −7.1 | 3.5 | −12.2 | 3.5 | −7.1 | 3.5 |
Parameter | Unit | Single i.v. 1 mg/kg | Single p.o. 25 mg/kg | Single p.o. 50 mg/kg | Single p.o. 100 mg/kg | Multiple p.o. 100 mg/kg |
---|---|---|---|---|---|---|
t1/2 | h | 0.89 ± 0.52 | 2.86 ± 0.29 | 2.63 ± 0.20 | 3.04 ± 0.70 | 5.03 ± 0.82 |
tmax | h | - | 0.25 ± 0.00 | 0.50 ± 0.00 | 0.17 ± 0.04 | 2.00 ± 0.00 |
Cmax1 | ng/mL | 1711.0 ± 397.7 | 515.0 ± 106.7 | 693.2 ± 260.4 | 1324.0 ± 227.5 | 1711.7 ± 185.0 |
AUC0–t | μg/L·h | 500.9 ± 124.3 | 2517.9 ± 395.2 | 4729.6 ± 854.3 | 6921.7 ± 377.4 | 11,753.9 ± 1145.6 |
AUC0–∞ | μg/L·h | 502.7 ± 124.4 | 2526.5 ± 362.4 | 4743.3 ± 854.4 | 6957.4 ± 345.1 | 12,132.7 ± 1187.2 |
MRT | h | 0.6 ± 0.1 | 5.2 ± 0.4 | 5.4 ± 0.8 | 5.3 ± 0.3 | 6.5 ± 0.6 |
V/F 2 | L/kg | 2.6 ± 1.4 | 40.9 ± 7.4 | 40.1 ± 10.8 | 63.0 ± 17.6 | 59.8 ± 11.8 |
CL/F 3 | L/kg/h | 2.1 ± 0.6 | 9.9 ± 1.6 | 10.5 ± 2.2 | 14.4 ± 0.7 | 8.2 ± 0.8 |
F | % | - | 20.1 ± 0.8 | 18.9 ± 1.7 | 13.8 ± 0.8 | 23.5 ± 2.3 |
Species | Plasma Protein Binding (%) | ||
---|---|---|---|
50 ng/mL | 500 ng/mL | 2500 ng/mL | |
Human | 44.1 ± 2.5 | 38.8 ± 12.0 | 40.5 ± 2.5 |
Rat | 63.2 ± 3.3 | 61.4 ± 1.2 | 64.5 ± 2.5 |
Species | Blood Plasma Ratio | ||
---|---|---|---|
50 ng/mL | 500 ng/mL | 2500 ng/mL | |
Human | 1.39 ± 0.14 | 1.28 ± 0.05 | 1.22 ± 0.32 |
Rat | 0.84 ± 0.26 | 0.74 ± 0.06 | 0.79 ± 0.15 |
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Xu, Y.; Zhang, H.; Li, N.; Ma, W.; Wang, S.; Sun, J.; Yang, B. Preclinical Pharmacokinetic Studies of a Novel Diuretic Inhibiting Urea Transporters. Molecules 2022, 27, 2451. https://doi.org/10.3390/molecules27082451
Xu Y, Zhang H, Li N, Ma W, Wang S, Sun J, Yang B. Preclinical Pharmacokinetic Studies of a Novel Diuretic Inhibiting Urea Transporters. Molecules. 2022; 27(8):2451. https://doi.org/10.3390/molecules27082451
Chicago/Turabian StyleXu, Yue, Hang Zhang, Nannan Li, Wen Ma, Shuyuan Wang, Jianguo Sun, and Baoxue Yang. 2022. "Preclinical Pharmacokinetic Studies of a Novel Diuretic Inhibiting Urea Transporters" Molecules 27, no. 8: 2451. https://doi.org/10.3390/molecules27082451
APA StyleXu, Y., Zhang, H., Li, N., Ma, W., Wang, S., Sun, J., & Yang, B. (2022). Preclinical Pharmacokinetic Studies of a Novel Diuretic Inhibiting Urea Transporters. Molecules, 27(8), 2451. https://doi.org/10.3390/molecules27082451