Poor water-solubility of many drug candidates represents high risk with regard to inadequate absorption and consequently poor and variable oral bioavailability of such drugs. A number of approaches have been developed to overcome poor oral bioavailability by enhancing solubility and dissolution rate of poorly soluble drugs. The aim of present work was to evaluate the influence of different solubility improvement technologies on stability of tacrolimus drug substance and its formulations. The effects of amorphization, solid dispersion preparation and nanosizing was studied. In addition, an influence of various environmental factors and effects of excipients on the stability behavior of tacrolimus was studied. Tacrolimus is a potent immunosuppressive drug. It is practically insoluble in water (1 – 2 μg/mL). Due to its complex structure based on 23-memberd macrolide lactone, tacrolimus exhibits different kinds of conformational heterogeneity in the solution and is subjected to various chemical transformations. In the present work, an advanced UPLC method for simultaneous separation of tacrolimus known impurities and its equilibrium compounds was applied for stability evaluation of tacrolimus. Stability indicating power of the method was confirmed by the appropriate separation of unknown tacrolimus degradation products formed in the forced degradation studies. Forced degradation study in solution proved tacrolimus instability under basic conditions, thermal stress, light and photolytic conditions and in the presence of radical initiator or metal ions. Tacrolimus was stable under neutral and acid conditions (pH 3-5). Solid-state degradation studies in addition to accelerated and long-term stability study confirmed that the amorphous form of tacrolimus is significantly less stable than crystalline tacrolimus monohydrate. Degradation studies conducted on amorphous drug substance revealed its sensitivity to light, elevated temperature, humidity and oxygen induced oxidation. Crystalline tacrolimus monohydrate was stable under conditions of accelerated and long-term stability, no degradation was observed at elevated temperature, high humidity or due to oxygen present in the air atmosphere. Crystalline tacrolimus was found to be sensitive to light. Solid-state excipient compatibility testing and stability evaluation of heterogeneous pharmaceutical systems containing amorphous solid dispersions revealed that amorphous tacrolimus is more susceptible to interactions with excipients than tacrolimus in crystalline form. Under the influence of environmental factors and in the presence of excipients, amorphous tacrolimus was subjected to thermal rearrangement, rearrangement due to presence of multivalent metal ions, epimerization, dehydration and other chemical transformations based on free radical catalyzed autoxidation. Significant degradation of amorphous tacrolimus was observed in the presence of magnesium stearate and stearic acid. On the other hand, hydroxypropyl methylcellulose, croscarmellose sodium and lactose were found to be compatible with amorphous tacrolimus. In the presence of magnesium stearate tacrolimus was decomposed toward a single degradation product. The impurity was isolated and its structure was elucidated. The impurity was found to be alpha-hydoxy acid, formed from the parent molecule through benzylic acid type rearrangement reaction in the presence of multivalent metallic ions under basic conditions. We have demonstrated that the stability of amorphous tacrolimus in heterogeneous systems can be improved by selection of suitable excipients, appropriate pH of the microenvironment and by applying dry storage conditions. Amorphous tacrolimus drug substance and formulations prepared as amorphous solid dispersions have also demonstrated an increased dissolution rate of tacrolimus. Stability evaluation of pharmaceutical system containing tacrolimus nanoparticles (nanocrystals) demonstrated enhanced chemical stability in comparison to amorphous tacrolimus and amorphous solid dispersions. During the milling process the crystalline form of tacrolimus monohydrate was retained. By wet ball milling approach a substantial reduction in particle size was achieved and consequently an increased dissolution rate of tacrolimus was observed.
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