HPLC method for levothyroxine quantification in long-acting drug delivery systems. Validation and evaluation of bovine serum albumin as levothyroxine stabiliser

Deficiency of thyroid hormones (hypothyroidism) is treated with oral levothyroxine (LEVO). However, the effectiveness of oral administration is highly dependent on the co-administration of food and other drugs. This factor, in combination with the chronic nature of this condition, mean that there are concerns with patient compliance. Development of long acting formulations to treat hypothyroidism could potentially solve this problem. However, LEVO instability in solution could be problematic. In order to develop long acting LEVO delivery systems in vitro drug release experiments should be carried out. However, short term LEVO stability in aqueous solution will prevent this. BSA was used as a stabiliser for LEVO; extending the stability of the drug in aqueous solutions from a few hours to 2 weeks. In order to achieve this, the required concentration of the protein was 0.1 % w/v. Subsequently, an HPLC method capable of separating LEVO from the protein was developed and validated following ICH guidelines. The analysis was carried out using a reverse phase HPLC method on an Agilent 1220 Infinity II LC system. The column used to achieve separation was a Zorbax Eclipse plus C18 (95 Å pore size, 250 mm length x 4.6 mm internal diameter; 5 μm particle size). The mobile phase used was composed of acetonitrile and 0.1 % trifluoroacetic acid at a ratio of 50:50 % v/v. UV detection of LEVO sodium was carried out at 225 nm. The retention time for the drug was 6.6 min. The method showed a limit of detection of 0.03 μg/mL and a limit of quantification of 0.09 μg/mL. Finally, this method was used to evaluate the release from implants containing 20 % w/w of LEVO. These devices were prepared using a solvent casting method with poly(caprolactone) and LEVO. These devices showed an initial burst release over the first 3 days. Subsequently, they were capable of providing a linear release rate over the following 25 days.