Distribution Study of 5-[5-(trifluoromethyl)-1,2-oxazole-3-yl]-furan-2-sulfonamide and Its Metabolites in Rats

INTRODUCTION. This article continues a series of publications on the pharmacokinetics of 5-[5-(trifluoromethyl)-1,2-oxazole-3-yl]-furan-2-sulfonamide (TFISA), a novel compound for the treatment of open-angle glaucoma. The distribution of TFISA and its metabolites in rat organs and tissues has not been previously studied in preclinical trials.

AIM. This study aimed to assess the tissue distribution and bioavailability of 5-[5-(trifluoromethyl)-1,2-oxazol-3-yl]-furan-2-sulfonamide and its metabolites in rat organs and tissues and to validate the analytical procedures developed for this purpose. MATERIALS AND METHODS. The study used 60 male Wistar rats. TFISA was administered by bilateral ocular instillation of 1% ophthalmic suspension at a dose of 40 μL (approximately 3.7 mg/kg). Tissue samples (liver, kidney, lung, brain, heart, spleen, skin, muscle tissue, and eyes) were collected 1, 2, 4, 8, 12, 24, 48, 72, 144, and 216 h after instillation (from 6 rats at each time point). The samples were immediately homogenised using methanol and were stabilised with ascorbic acid solutions. High-performance liquid chromatography with tandem mass spectrometric detection was used to quantify TFISA, N-hydroxy-5-[5-(trifluoromethyl)-1,2-oxazole-3-yl]furan-2-sulfonamide (M1), and N-acetyl-5-[5-(trifluoromethyl)-1,2-oxazole-3-yl]-furan-2-sulfonamide (M2) in homogenised organ and tissue samples.

RESULTS. This study involved full validation of the analytical procedures developed for the quantitative determination of TFISA and its metabolites, which was conducted separately for eye tissues and other biological samples. The tissue bioavailability (f_t) of TFISA decreased from 13.0 to 0.7 in the following order: eye tissues (administration and action site) > spleen > lungs ≥ heart ≥ liver > kidneys > brain > skin ≥ muscles. The f_t values for M1 decreased from 52.0 to 2.5 in the following order: spleen ≥ lungs ≥ heart ≥ liver > kidneys > brain > muscles ≥ eye tissues > skin. The f_t values for M2 were lower than those for TFISA and M1 and decreased from 6.4 to 0.3 in the following order: liver ≥ kidneys > heart > lungs > eye tissues > skin > spleen ≥ muscles > brain.

CONCLUSION. The validated bioassays have been successfully applied to study the distribution of TFISA and its metabolites in male rats. TFISA best penetrates into the eyes and well-vascularised organs. M1 is highly bioavailable in the spleen, heart, and lungs. M2 shows the highest bioavailability in the liver and kidneys of rats.

1. Khokhlov AL, Shetnev AA, Korsakov MK, Fedorov VN, Tyu shina AN, Volkhin NN, Vdovichenko VP. Pharmacological properties of sulfonamide derivatives — new inhibitors of carbonic anhydrase. Bulletin of Experimental Biology and Medicine. 2023;175(2):166–70 (In Russ.). https://doi.org/10.47056/0365-9615-2023-175-2-166-170

2. Yaichkov II, Khokhlov AL, Korsakov MK, Volkhin NN, Petukhov SS, Zaykova VE, Lasariants OE. Excretion study of 5-[5-(trifluoromethyl)-1,2-oxazole-3-yl]-furan-2-sulfonamide in rats. Regulatory Research and Medicine Evaluation. 2025;15(3) (In Russ.). https://doi.org/10.30895/1991-2919-2025-697

3. Chen M, Jin J, Ji X, Chang K, Li J, Zhao L. Pharmacokinetics, bioavailability and tissue distribution of chitobiose and chitotriose in rats. Bioresour Bioprocess. 2022;9(1):13. https://doi.org/10.1186/s40643-022-00500-y

4. Sheng Y-H, Siemiątkowska A, Kosicka-Noworzyń K, Brunetti L, Kagan L. A validated LC-MS/MS method for simultaneous quantitation of piperacillin, cefazolin, and cefoxitin in rat plasma and twelve tissues. J Pharm Biomed Anal. 2024;248:116259. https://doi.org/10.1016/j.jpba.2024.116259

5. Hu H, Xiao H, Bao H, Li M, Xue C, Li YT, et al. Tissue distribution comparison of six active ingredients from an Eucommiae cortex extract between normal and spontaneously hypertensive rats. Evid Based Complement Alternat Med. 2020;2020:2049059. https://doi.org/10.1155/2020/2049059

6. Körmöczi T, Szabó Í, Farkas E, Penke B, Janáky T, Ilisz I, Berkecz R. Heart-cutting two-dimensional liquid chromatography coupled to quadrupole-orbitrap high resolution mass spectrometry for determination of N,N-dimethyltryptamine in rat plasma and brain; Method development and application. J Pharm Biomed Anal. 2020;191:113615. https://doi.org/10.1016/j.jpba.2020.113615

7. Rosebooma IC, Thijssena B, Rosinga H, Alvesb F, Mondalc D, Teunissend MBM, et al. Development and validation of an HPLC-MS/MS method for the quantification of the anti-leishmanial drug miltefosine in human skin tissue. J Pharm Biomed Anal. 2021;207:114402. https://doi.org/10.1016/j.jpba.2021.114402

8. Eom HY, Jang S-I, Lee J-H. Development and validation of a bioanalytical method for 3'- and 6'-sialyllactose in minipig liver and kidney using liquid chromatography-tandem mass spectrometry and its application to analysis of tissue distribution. Molecules. 2020;25(23):5721. https://doi.org/10.3390/molecules25235721

9. Liang D, Wu Z, Liu Y, Li C, Li X, Yang B, Xie H, Sun H. HPLC-MS/MS-mediated analysis of the pharmacokinetics, bioavailability, and tissue distribution of schisandrol B in rats. Int J Anal Chem. 2021;2021:8862291. https://doi.org/10.1155/2021/8862291

10. Aladysheva ZhI, Belyaev VV, Beregovykh VV, Brkich GE, Greibo SV, Demina NB, et al. Industrial pharmacy. The way to create a product. Moscow: Russian Academy of Sciences; 2019 (In Russ.). EDN: KWJJUK

11. Popov NS, Gavrilenko DA, Balabanyan VYu, Petrova MB, Donskov SA, Atadzhanov IB, Shatokhina NA. Quantitative determination of monoamine neurotransmitters in rat brain homogenates using HPLC-MS/MS. Pharmacokinetics and Pharmacodynamics. 2022;(4):33–42 (In Russ.). https://doi.org/10.37489/2587-7836-2022-4-33-42

12. Barbosa da Silva Pereira HA, de Lima Leite A, Charone S, Vaz Madureira Lobo JG, Cestari TM, Peres-Buzalaf C, et al. Proteomic analysis of liver in rats chronically exposed to fluoride. PLoS One. 2013;8(9):e75343. https://doi.org/10.1371/journal.pone.0075343

13. Khokhlov AL, Yaichkov II, Korsakov MK, Kagramanyan IN, Volkhin NN, Petukhov SS, Zaikova VE. Development and validation of a method for the quantitative determination of monoamine neurotransmitters and their metabolites in rat brain tissue using HPLC-MS/MS. Acta Biomedica Scientifica. 2024;9(1):177–91 (In Russ.). https://doi.org/10.29413/ABS.2024-9.1.18

14. Plate AYA, Crankshaw DL, Gallaher DD. The effect of anesthesia by diethyl ether or isoflurane on activity of cytochrome P450 2E1 and P450 reductases in rat liver. Anesth Analg. 2005;101(4):1063–4. https://doi.org/10.1213/01.ane.0000166791.30963.ef

15. Koptyaeva KE, Muzhikyan AA, Guschin YaA, Belyaeva EV, Makarova MN, Makarov VG. Technique of dissection and extracting organs of laboratory animals. Message 1 (rats). Laboratory Animals for Science. 2018;(2):71–92 (In Russ.). https://doi.org/10.29296/2618723X-2018-02-08

16. Liang Y, Wang L, Zhang R, Pan J, Wu W, Huang Y, et al. Determination of the metabolites and metabolic pathways for three β-receptor agonists in rats based on LC-MS/MS. Animals. 2022;12(15):1885. https://doi.org/10.3390/ani12151885

17. Popovic MM, Schlenker MB, Thiruchelvam D, Redelmeier DA. Serious adverse events of oral and topical carbonic anhydrase inhibitors. JAMA Ophthalmol. 2022;140(3):235–42. https://doi.org/10.1001/jamaophthalmol.2021.5977

18. Kurysheva NI. Carbonic anhydrase inhibitors in the treatment of glaucoma. Review. Part II. Ophthalmology in Russia. 2020;17(4):676–82 (In Russ.). https://doi.org/10.18008/1816-5095-2020-4-676-682

19. Gu X-F, Mao B-Y, Xia M, Yang Y, Zhang J-L, Yang D-S, et al. Rapid, sensitive and selective HPLC–MS/MS method for the quantification of topically applied besifloxacin in rabbit plasma and ocular tissues: Application to a pharmacokinetic study. J Pharm Biomed Anal. 2016;117:37–46. https://doi.org/10.1016/j.jpba.2015.08.023

20. Kim E, Jang E, Jung W, Kim W, Lee J, Choi DH, et al. Establishment of an LC-MS/MS method for quantification of lifitegrast in rabbit plasma and ocular tissues and its application to pharmacokinetic study. J Chromatogr B Analyt Technol Biomed Life Sci. 2023;1229:123892. https://doi.org/10.1016/j.jchromb.2023.123892

21. Amo EM, Hammid A, Tausch M, Toropainen E, Sadegh A, Valtari A, et al. Ocular metabolism and distribution of drugs in the rabbit eye: Quantitative assessment after intracameral and intravitreal administrations. Int J Pharm. 2022;613:121361. https://doi.org/10.1016/j.ijpharm.2021.121361

22. Yaichkov II, Korsakov MK, Volkhin NN, Petukhov SS, Tyushina AN, Zaykova VE, Lasaraynz OE. Pharmacokinetic study of a new 4,5-dihydroisoxazole-5-carboxamide derivative in rats. Drug Development & Registration. 2024;13(4):238–50 (In Russ.). https://doi.org/10.33380/2305-2066-2024-13-4-1876

23. Al-Ghananeem AM, Crooks PA. Phase I and Phase II ocular metabolic activities and the role of metabolism in ophthalmic prodrug and codrug design and delivery. Molecules. 2007;12(3):373–88. https://doi.org/10.3390/12030373

24. Argikar UA, Dumouchel JL, Dunne CE, Saran C, Cirello AL, Mithat G. Ocular metabolism of levobunolol: Historic and emerging metabolic pathways. Drug Metab Dispos. 2016;44(8):1304–12. https://doi.org/10.1124/dmd.116.070458