Ion-pairing HPLC methods to determine EDTA and DTPA in small molecule and biological pharmaceutical formulations
Authors: George Wang, Frank P.Tomasella
Abstract:
We have developed and validated ion-pairing high-performance liquid chromatography-ultraviolet (HPLC-UV) methods for the quantification of ethylenediaminetetraacetic acid (EDTA) in Abilify® oral solution and diethylenetriaminepentaacetic acid (DTPA) in Yervoy® intravenous formulation. As EDTA and DTPA do not possess chromophores, we incorporated transition metal ions (Cu2+, Fe3+) during sample preparation to generate stable metallocomplexes, enhancing UV detection. This approach, combined with ion-pairing chromatography, allowed us to achieve the desired sensitivity and selectivity for method development. Copper facilitated the determination of EDTA, while iron was utilized for DTPA analysis. We employed a gradient mobile phase to separate the components of the formulation from the analyte in the case of EDTA, and the active drug substance, ipilimumab, was eluted separately in the DTPA method. Additionally, we optimized the concentration of the ion-pairing reagent to enhance retention of aminopolycarboxylic acids (APCAs), including EDTA and DTPA, and to improve method specificity. Our method development was based on the analytes' chromatographic properties, sample matrix characteristics, and the intended purpose of the method. We present validation data for both methods, and successfully applied them to investigate the fate of the chelates.
Keywords
EDTA; DTPA; Ion-pairing HPLC; Aripiprazole; Ipilimumab
Citation: George Wang, Frank P.Tomasella Ion-pairing Hplc Methods To Determine Edta And Dtpa In Small Molecule And Biological Pharmaceutical Formulations doi:10.1016/j.jpha.2016.01.002
Received: 6 October 2015, Revised: 15 January 2016, Accepted: 20 January 2016, Available online: 21 January 2016
Copyright: © 2016 Xi'an Jiaotong University. Production and hosting by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/)
Conclusion
In conclusion, two fit for purpose, ion-pairing reversed-phase HPLC methods were developed to determine EDTA in Abilify® oral solution and DTPA in Yervoy® intravenous solution drug products. TBA was added into the mobile phase to form ion pairs with metallocomplexes of EDTA and DTPA to enhance retention on reversed-phase HPLC columns. The analytical method utilized gradient and isocratic mobile phases in the determination of an APCA in a small molecule and a biological formulation, respectively. The use of metallocomplexes with ion-pairing chromatography provides the ability to achieve the desired sensitivity and selectivity in the development of the method.
Specifically, the sample preparation involving metallocomplex formation allows UV detection with high sensitivity. Copper (II) was utilized for the determination of EDTA and iron (III) was utilized for the determination of DTPA. For the determination of EDTA, a gradient mobile phase separated the components of the formulation from the analyte. In the case of DTPA, the active drug substance, ipilimumab, was eluted in the void. To the best of our knowledge, there is no report on analytical HPLC method for APCAs by direct injections of monoclonal drug formulations. In addition, the optimization of the concentration of the ion-pairing reagent was discussed as a means of enhancing the retention of the APCAs and the specificity of the method. The analytical method development was designed based on the chromatographic properties of the analytes, the nature of the sample matrix and the intended purpose of the method.
Acknowledgments
The authors wish to acknowledge the following that provided the needed supplies or stimulating conversation: Jonathan D. Basch, Jacob Bongers, Mi Jin, David K. Lloyd and Rao V. Mantri, Bristol-Myers Squibb Company New Brunswick, NJ, USA.
