Levothyroxine (T4)

For optimal results in HPLC analysis, it is recommended to measure absorbance at a wavelength that matches the absorption maximum of the compound(s) being analyzed. The UV spectrum shown can assist in selecting an appropriate wavelength for your analysis. Please note that certain mobile phases and buffers may block wavelengths below 230 nm, rendering absorbance measurement at these wavelengths ineffective. If detection below 230 nm is required, it is recommended to use acetonitrile and water as low UV-transparent mobile phases, with phosphoric acid and its salts, sulfuric acid, and TFA as buffers.
For some compounds, the UV-Vis Spectrum is affected by the pH of the mobile phase. The spectra presented here are measured with an acidic mobile phase that has a pH of 3 or lower.

Iodine is an important element in production of thyroid hormones, T3 and T4, which are essential regulators of organism’s metabolism. The glands’ concentration of Iodide, hormones T3 and T4, along with their precursors: 3,5-Diiodo-L-thyronine (T2), 3,5-Diiodo-L-tyrosine (DIT), and 3-Iodo-L-tyrosine (MIT) can now be studied simultaneously with this HPLC method.  In order to analyze the hydrophobic hormones along with iodide (a very polar inorganic ion) a special mixed-mode column, Primesep D, was used.  This column provides both strong hydrophobic and anion exchange properties.  The Primesep D silica-bonded ligand is comprised of a long alkyl chain and an embedded amino functional group.  The TFA was used as an ionic modifier to provide stable acidic pH and sufficient ion strength for the mobile phase.  The mobile phase composition is suitable for UV, MS, ELSD, and CAD detection.