Optimizing riboflavin delivery with co-crystal and in situ hydrogel formulations for management of keratoconus: A comprehensive investigation with in vitro, ex vivo and in vivo studies

AYTEKİN E., Kaya M. Z., POLAT H. K., ÇAKMAK H. B., ÇELEBİER M., PALASKA E., ...More

INTERNATIONAL JOURNAL OF PHARMACEUTICS, vol.674, 2025 (SCI-Expanded) identifier identifier

  • Publication Type: Article / Article
  • Volume: 674
  • Publication Date: 2025
  • Doi Number: 10.1016/j.ijpharm.2025.125435
  • Journal Name: INTERNATIONAL JOURNAL OF PHARMACEUTICS
  • Journal Indexes: Science Citation Index Expanded (SCI-EXPANDED), Scopus, Academic Search Premier, Aquatic Science & Fisheries Abstracts (ASFA), BIOSIS, Biotechnology Research Abstracts, CAB Abstracts, Chemical Abstracts Core, EMBASE, International Pharmaceutical Abstracts, Veterinary Science Database
  • Hacettepe University Affiliated: Yes

Abstract

Keratoconus is a progressive disease characterized by corneal thinning and conical deformation. Corneal crosslinking, a common treatment, strengthens collagen fibers using vitamin B2 (riboflavin) and UVA light. However, the surgical removal of the corneal epithelium (Epi-Off) required for riboflavin penetration causes complications and may affect treatment success. To address this, research has focused on delivering riboflavin without removing the epithelium (Epi-On). In this study, riboflavin-based hydrogel and co-crystal formulations were developed and evaluated through in vitro, ex vivo, and in vivo studies. Co-crystals were prepared using trehalose, dextrose, mannitol, and nicotinamide as agents, employing solvent evaporation and co-mixing methods. These formulations were characterized using DSC (Differential Scanning Calorimetry), XRD (X-Ray Diffraction), and FTIR (Fourier Transform Infrared Spectroscopy), identifying 1R1N (1 unit mol riboflavin and 1 unit mol nicotinamide co-crystals) and 1R1M (1 unit mol riboflavin and 1 unit mol mannitol co-crystals) groups as promising candidates. Thermosensitive hydrophilic gels containing riboflavin or riboflavin-5-phosphate sodium and Transcutol P (a permeation enhancer) were also developed, using Pluronic F127 as the polymer. The 18 % Pluronic F-127 gel formed at 31.4 +/- 0.2 degrees C. Drug release studies showed faster release from riboflavin-5-phosphate sodium formulations, while ex vivo retention studies revealed higher corneal retention for co-crystals. In vivo studies on rat corneas demonstrated superior drug concentrations for riboflavin formulations compared to riboflavin-5-phosphate sodium, with hydrophilic gels showing prolonged corneal contact time. The THJ-TP formulation (Riboflavin-5-phosphate sodium and permeation enhancer (Transcutol P) containing hydrophilic gel formulations), containing riboflavin-5-phosphate sodium and Transcutol P, emerged as the most promising candidate. This research represents a significant advancement towards a non-invasive riboflavin-based treatment for keratoconus.