Influences of experimental parameters on the stability of a benzoporphyrin drug in water/ethanol mixtures: a statistical approach investigation

Abstract

The benzoporphyrin derivative, named B3B, is a promising new drug for application in photodynamic therapy. In aqueous organic solvent mixtures, B3B, can be used in topical application for superficial diseases, like for skin cancer. In this paper, the self-aggregation phenomena of porphyrin molecules in aqueous media was investigated using a chemometric tool to evaluate the effects of experimental variables on the stability of the B3B monomer in water/ethanol mixtures. A 23factorial design was employed which permitted determination of the magnitude of experimental parameters which influence aggregation of the drug at different pHs using the minimum number of experiments. At first, the pKavalues of the B3B in water/ethanol mixtures were determined by spectrophotometric measurements, resulting in pKa1= 3.3 and pKa2= 5.7, corresponding to the imino nitrogens and carboxylic groups, respectively. The experimental variables (B3B concentration, time and water/ethanol composition) were evaluated in three protonation states of the drug, at pH 7.0, 4.1 and 2.6. The two levels (low and high) were set up at: time 5 (-) and 360 minutes (+); [B3B] 4.4 × 10-6(-) and 26.5 × 10-6M (+); and water content in ethanol 40 (-) and 60% (+). The main parameter that determines the magnitude of the effects seems to be the charge component of B3B, which can be modulated by the pH. The results from the 23fractional design indicated that the main variable inducing B3B aggregation is the water percentage in ethanol at each pH selected. The magnitude of this effect was pronounced at pH 4.1 where the carboxylic and nitrogen free base forms of B3B are maintained. The factors of time and [B3B], despite being small, are not insignificant. Finally, the effect of water content was investigated in a wide range of water percentage in these three pHs.