Tolfenamic acid (TA) is a fenamate nonsteroidal anti-inflammatory drug with expanding therapeutic potential, but its clinical use is limited by systemic side effects and the lack of topical formulations. Developing a stable topical gel may enable localized delivery of TA while minimizing systemic exposure. In the present study, simple hydroalcoholic gel formulations of TA with carboxymethylcellulose sodium (CMC) were prepared, with or without propylene glycol (PG), at pH 6.5. Before the formulation step, the interactions and compatibility of the TA/CMC (1:1) physical mixtures were studied using Fourier transform infrared spectroscopy and high-performance liquid chromatography. The formulated gels were stored at room temperature (27 ± 2°C) for six months and evaluated for their physical and chemical stability. The results indicated that gels prepared without PG showed greater moisture evaporation, resulting in higher viscosity, spreadability, and swelling index, with lower weight loss. No syneresis was observed in either gel formulation under different conditions, except under acidic conditions, where syneresis was observed. Microscopic evaluation of the formulations revealed polymorphic changes of the drug in the gels during storage. Both gels demonstrated optimal chemical stability, with negligible degradation throughout the study. Release kinetic studies indicated that TA followed the Higuchi model, with almost 100% drug release within 2.5 h.

1. ABrayfield A. Martindale: The complete drug reference, 38th ed. London, UK: Pharmaceutical Press. 2014. doi: 10.1007/s11096-014-0007-x

2. British Pharmacopoeia. Monograph on tolfenamic acid. London, UK: Her Majesty’s Stationery Office. 2026.

3. O’Neil MJ. Monograph on tolfenamic acid. In: The Merck Index, 15th ed., White House Station, NJ, USA: Merck & Co., Inc. 2013.

4. 4. British National Formulary (BNF) 85. London, UK: Pharmaceutical Press. 2025.

5. Christelle K, Norhayati MN, Jaafar SH, et al. Interventions to prevent or treat heavy menstrual bleeding or pain associated with intrauterine-device use. Cochrane Database Syst Rev. 2022;8:CD006034. doi: 10.1002/14651858.CD006034.pub3

6. Basha R, Baker CH, Sankpal UT, et al. Therapeutic applications of NSAIDs in cancer: Special emphasis on tolfenamic acid. Front Biosci. 2011;3:797–805. doi: 10.2741/s188

7. Feldman D, Leahy E, Lee SH, Chemopreventive properties of tolfenamic acid: A mechanistic review. Curr Med Chem. 2018;25:1598–1608. doi: 10.2174/0929867324666170414155107

8. Hu X, Li J, Zhang H, et al. Discovery of dual inhibitors of topoisomerase I and cyclooxygenase-2 for colon cancer therapy. Eur J Med Chem. 2022;240:114560. doi: 10.1016/j.ejmech.2022.114560

9. Hurtado M, Prokai L, Sankpal UT, et al. Next generation sequencing and functional pathway analysis to understand the mechanism of action of copper-tolfenamic acid against pancreatic cancer cells. Process Biochem. 2020;89:155–164. doi: 10.1016/j.procbio.2019.10.022

10. Kang SU, Shin YS, Hwang HS, et al. Tolfenamic acid induces apoptosis and growth inhibition in head and neck cancer: Involvement of NAG-1 expression. PLoS ONE. 2012;7:34988. doi: 10.1371/journal.pone.0034988

11. Li J, Hu X, Zhang H, et al. N-2-(Phenylamino) benzamide derivatives as dual inhibitors of COX-2 and Topo I deter gastrointestinal cancers via targeting inflammation and tumor progression. J Med Chem. 2022;65:10481–10505. doi: 10.1021/acs.jmedchem.2c00635

12. Matsunaga T, Horinouchi M, Saito H, et al. Availability of aldo-keto reductase 1C3 and ATP-binding cassette B1 as therapeutic targets for alleviating paclitaxel resistance in breast cancer MCF7 cells. J Biochem. 2023;173:167–175. doi: 10.1093/jb/mvac098

13. Poradowski D, Obmińska-Mrukowicz B, Effect of selected nonsteroidal anti-inflammatory drugs on the viability of canine osteosarcoma cells of the D-17 line, In vitro studies. J Vet Res. 2019;63:399–403. doi: 10.2478/jvetres-2019-0051

14. Sankpal UT, Lee CM, Connelly SF, et al. Cellular and organismal toxicity of the anticancer small molecule, tolfenamic acid: A pre-clinical evaluation. Cell Physiol Biochem. 2013;32:675–686. doi: 10.1159/000354471

15. Siraj S, Kurri A, Patel K, et al. Risk factors for esophageal cancer, with an emphasis on the role of specificity protein transcription factors in prognosis and therapy. Crit Rev Oncog. 2020;25:355–363. doi: 10.1615/CritRevOncog.2020036449

16. Xing L, Yang CX, Zhao D, et al. A carrier-free antiinflammatory platinum (II) self-delivered nanoprodrug for enhanced breast cancer therapy. J Control Release. 2021;331:460–471. doi: 10.1016/j.jconrel.2021.01.037

17. Leso A, Bihaqi SW, Masoud A, et al. Loss in efficacy measures of tolfenamic acid in a tau knock-out model: relevance to Alzheimer’s disease. Exp Biol Med. 2019;244:1062–1069. doi: 10.1177/1535370219871249

18. Zhu ZY, Liu YD, Gong Y, et al. Mitochondrial aldehyde dehydrogenase (ALDH2) rescues cardiac contractile dysfunction in an APP/PS1 murine model of Alzheimer’s disease via inhibition of ACSL4-dependent ferroptosis. Acta Pharmacol Sin. 2022;43:39–49. doi: 10.1038/s41401-021-00635-2

19. Andrews PC, Ferrero RL, Junk PC, et al. Bismuth (III) complexes derived from non-steroidal anti-inflammatory drugs and their activity against Helicobacter pylori. Dalton Trans. 2010;39(11):2861–2868. doi: 10.1039/c000164c

20. Yang F, Liu J, Gu Y, et al. Antimicrobial activity of auranofin, cannabidivarin, and tolfenamic acid against multidrugresistant Neisseria gonorrhoeae. Microbiol Spectr. 2022;10(6):0395222. 10.1128/spectrum.03952-22

21. Bishop YM. The Veterinary Formulary, 6th ed., British Veterinary Association, Pharmaceutical Press, London, UK. 2005.

22. Abdelwahab NS, Ali NW, Zaki MM, et al. Validated chromatographic methods for simultaneous determination of tolfenamic acid and its major impurities. J Chromatogr Sci. 2015;53:484–491. doi: 10.1093/chromsci/bmu071

23. Ahmad I, Sheraz MA, Ahmed S, et al. Photostability and interaction of ascorbic acid in cream formulations. AAPS PharmSciTech. 2011;12(3):917–923. doi: 10.1208/s12249-011-9659-1

24. Sheraz MA, Khan MF, Ahmed S, et al. Factors affecting formulation characteristics and stability of ascorbic acid in water-in-oil creams. Int J Cosmet Sci. 2014;36(5):494–504. doi: 10.1111/ics.12152

25. Kazi SH, Sheraz MA, Musharraf SG, et al. Analysis of tolfenamic acid using a simple, rapid, and stability-indicating validated HPLC method. Antiinflamm Antiallergy Agents Med Chem. 2024;23(1):52–70. doi: 10.2174/1871523022666230608094152

26. Mutimer MN, Riffkin C, Hill JA, Glickman ME, et al. Modern ointment base technology II. Comparative evaluation of bases. J Am Pharm Assoc. 1956;45(4):212–218. doi: 10.1002/jps.3030450406

27. Arora R, Aggarwal G, Harikumar SL, et al. Nanoemulsionbased hydrogel for enhanced transdermal delivery of ketoprofen. Adv Pharm. 2014;2014:1-12. doi: 10.1155/2014/468456

28. Khullar R, Kumar D, Seth N, et al. Formulation and evaluation of mefenamic acid emulgel for topical delivery. Saudi Pharm J. 2012;20(1):63–67. doi: 10.1016/j.jsps.2011.08.001

29. Khurana S, Jain NK, Bedi PMS. Nanoemulsion-based gel for transdermal delivery of meloxicam: physico-chemical, mechanistic investigation. Life Sci. 2013;92(6-7):383–392.. doi: 10.1016/j.jsps.2011.08.001

30. Ravindra J, Pratibha P, Patil P. Formulation and evaluation of semisolid preparation (ointment, gel & cream) of thiocolchicoside. J Pharm Biomed Sci. 2011;8:1–6.

31. Bao Y, Ma J, Li N. Synthesis and swelling behaviors of sodium carboxymethyl cellulose-g-poly(AA-co-AM-co-AMPS)/MMT superabsorbent hydrogel. Carbohydr Polym. 2011;84(1):76–82. doi: 10.1016/j.carbpol.2010.10.061

32. Curcio M, Spizzirri U, Iemma F, et al. Grafted thermoresponsive gelatin microspheres as delivery systems in triggered drug release. Eur J Pharm Biopharm. 2010;76(1):48–55. doi: 10.1016/j.ejpb.2010.05.008

33. Nesseem DI, Eid SF, El-Houseny SS. Development of novel transdermal self-adhesive films for tenoxicam, an antiinflammatory drug. Life Sci. 2011;89(13-14):430–438. doi: 10.1016/j.lfs.2011.06.026

34. Oprea A, Ciolacu D, Neamtu A, et al. Cellulose/chondroitin sulfate hydrogels: synthesis, drug loading/release properties and biocompatibility. Cellul Chem Technol. 2010;44(9):369–378.

35. Rokhade A, Agnihotri SA, Patil SA, et al. Semiinterpenetrating polymer network microspheres of gelatin and sodium carboxymethyl cellulose for controlled release of ketorolac tromethamine. Carbohydr Polym. 2006;65(3):243–252. doi: 10.1016/j.carbpol.2006.01.013

36. Mishra RK, Datt M, Banthia AK, Synthesis and characterization of pectin/PVP hydrogel membranes for drug delivery system. Carbohydr Polym. 2008;9(2):395–403. doi: 10.1016/j.carbpol.2006.01.013

37. Jana S, Das A, Nayak AK, et al. Aceclofenac-loaded unsaturated esterified alginate/gellan gum microspheres: in vitro and in vivo assessment. Int J Biol Macromol. 2013;57:129–137. doi: 10.1016/j.ijbiomac.2013.03.015

38. Franklin DS, Guhanathan S, Synthesis and characterization of citric acid-based pH-sensitive biopolymeric hydrogels. Polym Bull. 2014;71(1):93–110. doi: 10.1007/s00289-013-1047-4

39. Allen L, Ansel’s Pharmaceutical Dosage Forms and Drug Delivery Systems. Lippincott Williams & Wilkins, Philadelphia, USA. 2014.

40. Gan T, Guan Y, Zhang Y. Thermogelable PNIPAM microgel dispersion as 3D cell scaffold: effect of syneresis. J Mater Chem. 2010;20(28):5937–5944. doi: 10.1039/c0jm00338g

41. Helal DA, El-Rhman DA, Abdel-Halim SA, et al. Formulation and evaluation of fluconazole topical gel. Int J Pharm Sci. 2012;4(5):176–183.

42. Higuchi T. Rate of release of medicaments from ointment bases containing drugs in suspension. J Pharm Sci. 1961;50(10):874–875. doi: 10.1002/jps.2600501018

43. Higuchi T. Mechanism of sustained-action medication, Theoretical analysis of rate of release of solid drugs dispersed in solid matrices. J Pharm Sci. 1963;52(12):1145–1149. doi: 10.1002/jps.2600521210

44. Shoaib MH, Al Sabah Siddiqi S, Yousuf RI, et al. Development and evaluation of hydrophilic colloid matrix of famotidine tablets. AAPS PharmSciTech. 2010;11(2):708–718. doi: 10.1208/s12249-010-9427-7

45. Gilpin RK, Zhou W. Infrared studies of the polymorphic states of the fenamates. J Pharm Biomed Anal. 2005;37(3):509–515. doi: 10.1016/j.jpba.2004.11.009

46. Ahmed S, Sheraz MA, Rehman I. Studies on tolfenamic acidchitosan intermolecular interactions: effect of pH, polymer concentration and molecular weight. AAPS PharmSciTech. 2013;14(2):870–879. doi: 10.1208/s12249-013-9974-9

47. Jabeen S, Dines TJ, Leharne SA, et al. Raman and IR spectroscopic studies of fenamates–conformational differences in polymorphs of flufenamic acid, mefenamic acid and tolfenamic acid. Spectrochim. Acta A Mol Biomol Spectrosc. 2012;96:972–985. doi: 10.1016/j.saa.2012.07.129

48. Mattei A, Li T. Polymorph formation and nucleation mechanism of tolfenamic acid in solution: an investigation of pre-nucleation solute association. Pharm Res. 2012;29(2):460–470. doi: 10.1007/s11095-011-0574-7

49. Moffat AC, Osselton MD, Widdop B. Clarke’s Analysis of Drugs and Poisons, 4th ed. London, UK: Pharmaceutical Press. 2011.

50. Sheraz MA, Ahmed S, Rehman IU. Effect of pH, polymer concentration and molecular weight on the physical state properties of tolfenamic acid. Pharm Dev Technol. 2015;20(3):352–360. doi: 10.3109/10837450.2013.871027

51. Ahmed S, Sheraz MA, Ahmad I. Tolfenamic Acid. In: Brittain HG (ed.) Profiles of Drug Substances, Excipients and Related Methodology. Cambridge, MA, USA: Academic Press. 2018;43:255–319. doi: 10.1016/bs.podrm.2018.01.001

52. Ahmed S, Sheraz MA, Yorucu C, et al. Quantitative determination of tolfenamic acid and its pharmaceutical formulation using FTIR and UV spectrometry. Cent Eur J Chem. 2013;11(9):1533–1541. doi: 10.2478/s11532-013-0284-6

53. Andleeb S, Ahmed S, Sheraz MA, et al. Development and validation of a spectrofluorimetric method for the analysis of tolfenamic acid in pure and tablet dosage form. Luminescence. 2020;35(7):1017–1027. doi: 10.1002/bio.3810

54. Sheskey PJ, Cook WG, Cable CG, Handbook of Pharmaceutical Excipients, 8th ed. London, UK: Pharmaceutical Press. 2017.

55. Rozou S, Antoniadou-Vyza E. An improved HPLC method overcoming Beer’s law deviations arising from supramolecular interactions in tolfenamic acid and cyclodextrin complexes. J Pharm Biomed Anal. 1998;18(4-5):899–905. doi: 10.1016/S0731-7085(98)00227-1

56. Bergstrom CAS, Wassvik CM, Johansson K, et al. Poorly soluble marketed drugs display solvation limited solubility. J Med Chem. 2007;50:5858–5862. doi: 10.1021/jm0706416

57. Rozou S, Michaleas S, Antoniadou-Vyza E. Supramolecular interactions between tolfenamic acid and various cyclodextrins: effects of complexation on physicochemical and spectroscopic data. Pharm Pharmacol Commun. 1999;5(2):79–84. doi: 10.1211/146080899128734497

58. Banker GS, Anderson NR. Tablets. In: Lachman L, Lieberman HA, Kanig JL (eds.) The Theory and Practice of Industrial Pharmacy. Philadelphia, PA, USA: Lea & Febiger. 1986;293–345.

59. Nechipadappu SK, Trivedi DR. Structural and physicochemical characterization of pyridine derivative salts of anti-inflammatory drugs. J Mol Struc. 2017;1141:64–74. doi: 10.1016/j.molstruc.2017.03.086

60. Andersen KV, Larsen S, Alhede B, et al. Characterization of two polymorphic forms of tolfenamic acid, N-(2-methyl-3-chlorophenyl)anthranilic acid: their crystal structures and relative stabilities. J Chem Soc Perkin Trans. 1989;2(10):1443–1447. doi: 10.1039/p29890001443

61. Fabian L, Hamill N, Eccles KS, Moynihan HA, Maguire AR, McCausland L, et al. Cocrystals of fenamic acids with nicotinamide. Cryst Growth Des. 2011;11(8):3522-3528. doi: 10.1021/cg200429j

62. Lopez-Mejias V, Kampf JW, Matzger AJ. Polymer-induced heteronucleation of tolfenamic acid: structural investigation of a pentamorph. J Am Chem Soc. 2009;131(13):4554–4555. doi: 10.1021/ja806289a

63. Sacchi P, Reutzel-Edens SM, Cruz-Cabeza AJ. The unexpected discovery of the ninth polymorph of tolfenamic acid. Cryst Eng Comm. 2021;23(20):3636–3647. doi: 10.1039/D1CE00343G

64. Surov AO, Szterner P, Zielenkiewicz W, et al. Thermodynamic and structural study of tolfenamic acid polymorphs. J Pharm Biomed Anal. 2009;50(5):831–840. doi: 10.1016/j.jpba.2009.06.045

65. Surov AO, Terekhova IV, Bauer-Brandl A, et al. Thermodynamic and structural aspects of some fenamate molecular crystals. Cryst Growth Des. 2009;9(7):3265–3272. doi: 10.1021/cg900002q

66. Wittering KE, Agnew LR, Klapwijk AR, et al. Crystallisation and physicochemical property characterisation of conformationally-locked co-crystals of fenamic acid derivatives. Cryst Eng Comm. 2015;17(19):3610–3618. doi: 10.1039/C5CE00297D

67. Lin TJ. Bubble formation in hydroalcoholic gels. J Soc Cosmet Chem. 1969;20:795–805. 68. Sowmya J, Gowda DV, Srivastava A. Topical gels: A recent approach for novel drug delivery. Int J Health Sci Res. 2015;5(10):302–312.

69. Batheja P, Sheihet L, Kohn J, et al. Topical drug delivery by a polymeric nanosphere gel: Formulation optimization and in vitro and in vivo skin distribution studies. J Control Release. 2011;149(2):159–167. doi: 10.1016/j.jconrel.2010.10.005

70. Contreras MD, Sanchez R, Application of a factorial design to the study of the flow behavior, spreadability and transparency of a Carbopol ETD 2020 gel: Part II. Int J Pharm. 2002;234(1-2):149–157. doi: 10.1016/S0378-5173(01)00954-1

71. Hoffman AS. Hydrogels for biomedical applications. Adv Drug Deliv Rev. 2002;54(1):3–12. doi: 10.1016/S0169-409X(01)00239-3

72. Davidovich-Pinhas M, Bianco-Peled H. A quantitative analysis of alginate swelling. Carbohydr Polym. 2010;79(4):1020–1027. doi: 10.1016/j.carbpol.2009.10.036

73. Birch DJS, Geddes CD. Cluster dynamics, growth and syneresis during silica hydrogel polymerisation. Chem Phys Lett. 2000;320(3-4):229–236. doi: 10.1016/S0009-2614(00)00241-4

74. Walters KA. Drug delivery: topical and transdermal routes. In: Swarbrick J (ed.) Encyclopedia of Pharmaceutical Technology, 3rd ed. New York, NY, USA: Informa Healthcare Inc. 2007;2:1311–1325.

75. Food and Drug Administration (FDA). Guidance for Industry – Nonsterile Semisolid Dosage Forms, Scale-Up and Postapproval Changes: Chemistry, Manufacturing and Controls; In Vitro Release Testing and In Vivo Bioequivalence Documentation, US Department of Health and Human Services, Food and Drug Administration, Center for Drug Evaluation and Research (CDER). 1997.