The client has developed the liposomal formulation of a hydrophilic small molecule drug using unsaturated phospholipids (>90% molar fraction) and ethanol injection method. After process optimization, we observed a stability issue with the formulation at 2-8 ° C storage. Containing unsaturated lipids as major components, and in the absence of any buffering media, the liposomal formulation was susceptible to peroxidative degradation. Not only that, formulation was susceptible to hydrolysis with >10% impurities measured within one month of normal storage. To resolve the stability issue, we optimized the use of different buffers and incorporation of antioxidants to make the formulation long-term stable at 2-8 °C.
The challenge was to select a buffer that could maintain the pH of the liposomes (intra-liposomal and external to the liposomes) near neutral and also could be used in ethanol injection method. Different buffers, including TRIS, HEPES, bicarbonate, and phosphate buffers were tested to measure the solubility of API in these buffers and the compatibility with ethanol injection method. Being a hydrophilic drug, the entrapment yield in the liposomes was <10 %. Those three buffers, TRIS – pH 7.0, HEPES – pH 7.0 and bicarbonate, were found suitable to reduce the pH of API solution and were compatible with ethanol injection method.
These were also tested for the formulation development at an equivalent osmolarity as the control saline formulation. In addition to buffers, the antioxidants, butylated hydroxy toluene (BHT, 0.002% in final formulation) and α-tocopherol (NMT 0.1% of total lipids), were added in the formulations to reduce lipid peroxidation of the lipids.
These candidate liposomal formulations prepared resulted in similar physicochemical characteristics as the control formulation in terms of size distribution (80-100 nm Z-ave), entrapment yield (~10%), final entrapment efficiency (>95%), total API, total lipids, residual ethanol and lipid to API ratio. The developed formulations were also studied at different storage conditions. We found that the formulations with TRIS or HEPES buffer and antioxidants were able to reduce the peroxidative lipid degradation after at least one-month storage at accelerated, long-term conditions as well as stressed storage conditions.
We were able to optimize buffers used in liposomal formulations of hydrophilic APIs to maintain the pH near neutral and to decrease peroxidative degradation. As such, the selection of the appropriate buffers and antioxidants during formulation development resulted in an improved purity profile of the lipids upon finished drug product storage.
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