Category: Formulation and Quality
Purpose: The precipitation inhibitor ability of formulation components could determine supersaturating drug delivery systems accomplishment on improving poorly soluble drugs bioavailability. The role of glycyrrhizic acid (GA), a potential drug solubility enhancer, on preventing drug precipitation in supersaturated solutions has been poor explored. In this study, it was investigated the effect of GA as a precipitation inhibitor in buffer pH 6.5 and FaSSIF under supersaturated solutions of the poorly soluble drug griseofulvin. Also, it was studied the impact of the biorelevant medium on GA colloidal formation and how it affects the drug phase separation under supersaturation condition.
Methods: The effect o f GA on drug solubility in phosphate was investigated adding an excess of crystalline GSF to the medium (buffer pH 6.5 and FaSSIF) with and without 30 mmol L-1 of GA (approximately 1000 times drug solubility concentration), at 240 rpm, 37.0 °C, until reach the equilibrium (48 h). Supersaturation kinetic studies were conducted to evaluate the impact of GA on GSF precipitation behavior in both media, with and without 30 mmol L-1 of GA. The supersaturated solutions were generated by solvent-shift method, adding small aliquots of GSF stock solution (226.78 mmol L-1 dissolved in dimethylformamide (DMF)) to the medium, in order to achieve 4 different drug concentrations (0.283, 0.850, 1.701 and 3.402 mmol L-1). Samples were maintained at 37 °C, for 240 min in a shaker flask under agitation of 240 rpm. Aliquot samples were collected, filtered through a 0.45 μm membrane, diluted in mobile phase and quantified by HPLC methodology. Phase separation studies were conduct to investigate the maximum free drug concentration miscible in the different media. To produce supersaturated solutions, GSF stock solution (141.74 mmol L-1 dissolved in DMF) was titrated to 2 mL of medium in quartz cuvettes, flowed by sample’s homogenization (strong agitation was avoid to not support GSF fast recrystallization). The medium employed was phosphate buffer or FaSSIF, with and without 30 mmol L-1 of GA, maintained at 37.0 °C. After each titration, samples were studied through UV-Vis spectroscopy, fluorescence spectroscopy, dynamic light scattering (DLS) and polarized light microscopy (PLM). The impact of biorelevant medium on GA colloidal properties was also evaluated by conductivity, dynamic light scattering and pyrene fluorescence.
Results: GA presented remarkable drug solubilization in buffer pH6.5 (0.243 ± 0.011 mmol L-1), equivalent to 8-fold compare to the value found in buffer pH6.5 in its absence (0.030 ± 0.002 mmol L-1). However, in the biorelevant medium in the presence of GA, the drug solubility (0.028 ± 0.002 mmol L-1) was similar to the one found in buffer pH6.5, smaller than the one created in FaSSIF without its presence (0.063 ± 0.001 mmol L-1). It suggests that the drug solubilization ability of GA was strongly affected by the biorelevant medium, as well as its presence reduced drug micellar solubilization by bile salts. On precipitation studies (Figure 1), GA maintained high drug supersaturated solution concentration in buffer pH 6.5 ( > 0.38 mmol L-1) and FaSSIF ( > 0.60 mmol L-1) for 4 h. On GSF phase separation studies (Figure 2), the presence of GA in biorelevant medium increased the maximum free drug concentration to undergo this condition in FaSSIF (0.89 mmol L-1), compared to buffer pH 6.5 (0.49 mmol L-1), besides impacting on the precipitation type. While small crystalline particles were observed in buffer (with and without GA) and in FaSSIF without GA presence, amorphous aggregates were obtained under FaSSIF + GA supersaturated solutions (Figure 3). The differences between GA colloidal formation in buffer pH6.5 and in FaSSIF were investigated, since it might have impacted on drug solubilization, precipitation inhibition ability and phase separation phenomena of GSF supersaturated solutions. Different critical aggregation concentrations (0.00164 and 0.00497 mmol L-1) and different particle size number-weighted distribution (at around 1 and 4 nm) were observed for GA in buffer and FaSSIF, respectively. The pyrene I/III intensity ratios measured for the presence of GA in buffer and in FaSSIF were close to 0.8 and 0.5 at 30 mmol L-1, respectively. The intensity ratio values obtained in FaSSIF + GA provides a formation of a colloidal aggregates with polarity close to the organic solvent cyclohexane (0.7), with a substantial reduction in microenvironment polarity.
Conclusion: In conclusion, it was demonstrated the GA ability to act as an effective precipitation inhibitor under GSF supersaturated solutions in buffer pH 6.5 and FaSSIF. The FaSSIF + GA combination formed a mixed colloidal systems that exhibit smaller hydrodynamic diameter than FaSSIF and showed a microenvironment polarity similar to organics solvents in an aqueous medium. That colloidal phase enhance the system ability to resist recrystallization in solution. Despite of high recrystallization tendency of GSF under supersaturated solution, no crystallization was detected in FaSSIF + GA environment. The amorphous aggregates formatted could serve as a reservoir, re-dissolving as the free drug concentration decreases by the transport across biological membranes.
Maria Terezinha França– Florianopolis, Santa Catarina, Brazil
Tatyane Martins Marcos– Florianopolis, Santa Catarina, Brazil
Paulo F. A. Costa– Florianópolis, Santa Catarina, Brazil
Adriana Passarela Gerola– Florianópolis, Santa Catarina, Brazil
Hellen Stulzer– Professor, Universidade Federal De Santa Catarina, Florianopolis, Santa Catarina, Brazil