Stereoselective Hydrolysis and Penetration of Propranolol Prodrugs: In Vitro Evaluation Using Hairless Mouse Skin

Abstract

Stereoselective hydrolysis of two ester prodrugs of propranolol, isovaleryl propranolol (IV-PL) and cyclopropanoyl propranolol (CP-PL), was studied in Tris-HCI buffer (pH 7.4) containing 0.15M KCI, skin and liver homogenates, 5% plasma in Tris-HCI buffer, skin cytosol and microsomes, and liver cytosol and microsomes. The hydrolysis rate constants of (R)-isomers of the prodrugs were 1.1–30.3 times greater than those of the respective (S)-isomers in tissue preparations. Skin showed considerable metabolic activity and very high stereoselectivity (R/Sratio: 7.3–30.3). The hydrolyzing capacities of buffer and different tissue preparations per milligram of protein content were in the following increasing order: buffer < skin homogenate < plasma < liver homogenate. The studies with microsomes and cytosol indicated that the esterases, which are responsible for the hydrolysis of prodrugs, were mainly present in the cytosolic and microsomal fractions of skin and liver, respectively. There was a good correlation between the octanol–buffer partition coefficients of propranolol and its prodrugs and the skin partition coefficient. In vitro stereoselective penetration of propranolol and the prodrugs through full-thickness hairless mouse skin was evaluated with flow-through diffusion cells. Although the concentration of propranolol was 14–22 times greater than those of the prodrugs in the donor chamber, the steady-state flux of propranolol isomers [10.72 and 10.64 μg/cm2.h for (R)- and (S)-isomers, respectively] were similar to those of CP-PL [10.80 and 10.78 μg/cm2.h for (R)- and (S)-isomers, respectively] and even lower than those of IV-PL [14.51 and 14.33 μg/cm2.h for (R)- and (S)-isomers, respectively]. Moreover, the permeability coefficients of IV-PL [2.82 × 10−3and 2.78 × 10−3cm/h for (R)- and (S)-isomers, respectively] and CP-PL (1.29 × 10−3cm/h for each isomer) were 14–30-fold greater than those of propranolol isomers (0.09 × 10−3cm/h for each isomer). The diffusion coefficients of all the compounds were similar, but their solvent membrane distribution coefficients differed greatly and proved that the higher permeability coefficients of the prodrugs were due to the higher affinity of the prodrugs for skin. Neither propranolol nor the prodrugs showed stereoselective penetration. However, highly stereoselective hydrolysis occurred during penetration of the prodrugs, and theR/Sratios of the cumulative amount of delivered propranolol in 12h were 11 and 13 for IV-PL and CP-PL, respectively. A skin irritation test was performed in Japanese white male rabbits and no irritation was observed. In conclusion, the hairless mouse skin possesses highly stereoselective esterase activity, and IV-PL and CP-PL might be promising prodrugs for transdermal delivery of higher amounts of drug from a much lower initial concentration compared with propranolol.