Assessment of aluminum ion concentration using chrome azurol S in [N-13]ammonia injection

Objectives: [¹³N]Ammonia is one of the most widely used PET tracers for myocardial blood flow measurement. Devarda’s alloy containing aluminum, copper and zinc is used for the reduction of [¹³N]nitrate/nitrite to [¹³N]ammonia. Since aluminum has neuronal and renal toxicity, the amount of aluminum should be carefully monitored before the administration into human. However, the procedure to measure the aluminum concentration described in USP is complicate and is not easy to perform. Therefore, we tried to develop a simple and practical method to measure the aluminum concentration in [¹³N]ammonia injection.

Methods: Each 0.1 mL aliquot of sample or aluminum standard solution (1, 2, 5, 10, 20, and 50 ppm) was added to a 10-mm diameter test tube containing 0.9 mL of pH 4.6 acetate buffer. Ten μL of 10 g/mL chrome azurol S solution was added to each solution, and then the mixture was shaken well at room temperature for 3 min. The colors of sample and standard solutions were observed and compared. To investigate the effect of pH, the reaction was performed at pH 3, 4, 5, 6, 7, and 9. For the quantitative analysis, the OD values of 0.2 mL of the reaction mixtures in a 96-well plate were measured by using a spectrometer at 570 nm. Finally, the colors of the reaction mixtures were compared visually at 0 and 4 h after the preparation.

Results: Six different concentration standard solutions including 10 ppm (the limit in the USP) were prepared. The color of sample reactant should be less intense than that of 10 ppm standard by visual inspection to pass the test. The color of the reactants changed by pH: from dark orange at low to yellow at high. Thus, pH 4.6 acetate buffer was used to adjust pH for the practical Q.C. of [¹³N]ammonia. The OD value increased quantitatively until 20 ppm of the aluminum ion concentration. However, the OD value started to be saturated at 50 ppm. The reaction mixtures of sample and the standard solutions didn’t show any differences between 0 and 4 h, suggesting that the aluminum test could be completed within short time and minimize the radiation exposure.

Conclusions: Aluminum ion concentration of [¹³N]ammonia injection could be straightforwardly quantified and evaluated using chrome azurol S method. Considering test time and reproducibility, this method might be applicable in many institutes under GMP. Acknowledgements: This research was partially supported by NRF-2017M2A2A7A01071134, NRF-2015M2C2A1047687, and HI15C3093.

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