Category: Formulation and Quality
Purpose: Griseofulvin is an antifungal antibiotic used in the treatment of dermatophyte and ringworm infections. It is insoluble in water (8.64 mg/L, at 25 °C, pH 6.5) and is classified as a BCS Class II drugs with limited oral absorption due to its poor water solubility. The aim of this study is to assess the beta-cyclodextrin derivatives capacity to enhance griseofulvin’s solubility by complex formation; the data was evaluated by phase solubility technique and DSC.
Methods: Two hydroxypropyl beta-cyclodextrins (Kleptose HPB = HPBCD, Kleptose HP =HPCD) and methylated beta-cyclodextrins (Kleptose Crysmeb) were kindly provided by Roquette, and griseofulvin (USP micronized #30137) was a gift from Roquette. Phase solubility and stability test: A phase solubility study was performed according to the method reported by Higuchi and Connors. An excess amount of API was added to 10 mL of aqueous solutions of increasing CDs concentrations (20, 40, 80, 160 and 200 mM) in deionized water. The vials are shaken at room temperature for 3 days. The supernatant was collected and filtered through a 0.45 μm membrane and analyzed by UPLC for solubility assessment. The filtered supernatant, 1 mL was placed into the stability chambers (25°C /60% RH and 40°C /60% RH) and will be analyzed after 6 and 12 months, respectively. UPLC method: The mobile phase was composed of a mixture of 20 mM aqueous sodium dihydrogen phosphate solution and acetonitrile (55:45, v/v). The final pH of the mobile phase was adjusted to 3.5 with phosphoric acid. The mobile phase was filtered through a 0.45 μm membrane filter and degassed before use. Griseofulvin was separated on a reversed phase XBridge C18 column (130Å, 3.5 µm, 4.6 mm X 100 mm). The flow rate was set at 1.0 mL/min and the injection volume was 30 μL. Griseofulvin fluorescence measurements were made at excitation wavelengths of 300 nm. Lyophilization method: 2 mL supernatant of each phase solubility sample was filtered and placed in amber vials and dried at -50°C for 24 hours by utilizing FreeZone 4.5 Liter Benchtop Freeze Dry System. DSC method: The freeze-dried powders, CDs and griseofulvin (4 – 6 mg) were loaded into Tzero pans and analyzed using a Discovery DSC 2500 (TA Instruments; New Castle, DE) by heating to 300°C at 10°C/min to confirm the formation of inclusion complex.
Results: Phase solubility
From the phase solubility diagram, we calculated stability constant (K1:1) and complexation efficiency (CE) of griseofulvin/CD complexes. The slope (m) of linear regression and the drug solubility (S0) in water were used to calculate stability constant (K1:1) and complexation efficiency (CE) by using Eq. (1) and (2), and the results are shown in Table 1:
Eq. (1): K1:1 = m/ (S0(1-m)); Eq. (2): CE = m/ (1-m)
Table 1 Generic name, substitution, M.W., m (slope), stability constant (K1:1) and complexation efficiency (CE) of each CD
Molecular weight (g/mol)
KLEPTOSE ® HPB (medium MS, approx. 0.65)
KLEPTOSE ® HP (high MS, approx. 0.85)