ABSTRACT
Characterization studies were designed to evaluate the concentration and integrity of the L-thyroxine (T4) molecule (3,5,3',5'-tetraiodothyronine) in the free T4 stock solution (FT4SS) (code 99544). The determination of the concentration of T4 in FT4SS is critical to ensure that the free T4 calibrators and controls are manufactured with the least number of adjustments possible. The most significant conclusions drawn from these characterization studies are the following: (1) An accurate and sensitive HPLC method has been developed to measure the T4 concentration in FT4SS. The root cause of the failure of FT4SS to pass retest/ review is the presence of an unknown T4 degradation product with significantly higher molar extinction coefficient at 230 nm than T4 itself. The L-thyroxine concentration reference comparison spectrophotometric test with the current 43 to 58 ug/ml specification range (as per scp.99544, ed. 13A) is adequate to monitor the generation of the unknown T4 degradation product. The characterized T4 degradation product is not 3,5,3'-triiodo-thyronine (T3) and it is suspected that the identity of the degradation product is reverse T3 (3,3',5'-triiodothyronine). The use of sodium l-thyroxine pentahydrate (Na- T4-5H2O) as the equivalent of T4 (free base) is adequate provided that an excess of 15over the desired amount of T4 is weighed.