Development and Validation of HPLC-UV Method for the Determination of Levothyroxine in Orodispersible Minitablets for Pediatric Application

Abstract

Background: Thyroid hormones play an important role in cognition and brain development. The determination of the content of levothyroxine, as well as related substances and dissolution test analysis, should be carried out by methods that are selective and highly sensitive due to the low concentration used in low dose orodispersible minitablets. Objectives: This study aims to develop and validate an analytical method by HPLC-UV for the quantification of levothyroxine and its related substances in pharmaceutical formulations; looking forward to being this method suitable for a future dissolution test analysis using tandem mass spectrometry detector. Methods: A Hypersil GOLD C18 (100 x 2.1 mm, 3µm) column was used with 25°C column temperature, 5 µL injection volume, 0.3 mL/min flow rate and detection at 225 nm. The mobile phase consisted of methanol: 0.05% formic acid (55:45). The developed method was validated for specificity, linearity, precision, accuracy and robustness. Results: The method is linear within the range of 2-20 µg mL-1 (R2=0.9982), which makes the method suitable for the evaluation of levothyroxine in pharmaceuticals formulations. LOQ was 0.17 µg/mL (0.85 ng on column) and LOD 0.05 µg/mL (0.25 ng on column) of LT4. Therefore, in terms of efficiency (1671), retention factor, k (6.79), Tailing factor, T (1.09) and resolution, Rs (5.11) the chromatographic method was found to be suitable according to USP 43. Conclusion: The HPLC UV method was found to be linear, specific, precise, accurate and robust, therefore it is suitable for the quality control of levothyroxine in pharmaceutical ODMTs.