Determination of Dissociation Constants of ^99mTechnetium Radiopharmaceuticals by Capillary Electrophoresis

Capillary electrophoresis was applied to investigate pK_a values of ^99mTc radiotracers used in nuclear medicine. Therefore, the protonation equilibria of the carboxyl groups of ^99mTc- mercaptoacetylglycylglycylglycine (^99mTc-MAG₃) and ^99mTc-ethylenecysteine dimer (^99mTc-EC) were studied by pH-dependent determination of electrophoretical velocities. ^99methylenecysteine dimer diethyl ester (^99mTc-ECD) was used as a non-protonable standard. The capillary electrophoresis system was equipped with a radioactivity detector. Measurements were performed using a pressure-driven capillary zone electrophoresis which allowed runs even in the low pH range. For the determination of pK_a values, the electrophoretical velocities of the analytes were referred to the electrophoretical velocities of tetraphenyle arsonium chloride as a positively charged marker. Calculation of pK_a values was accomplished by non-linear curve fitting of both structure-based equilibria equations and sigmoidal decay functions to the experimental data. ^99mTc-MAG₃ was shown to have a carboxyl group pK_a value of 4.22. the value for the carboxyl groups of ^99mTc-EC is 2.90 (determined by structure-based equilibria equations), which represents a common value for both carboxyl groups. By the use of sigmoidal functions, similar values were elucidated. As expected, ^99mTc-ECD shows no protonation step.