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Degradants of Tenofovir Disoproxil Fumarate Under Forced Yet Mild Thermal Stress: Isolation, Comprehensive Structural Elucidation, and Mechanism

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Abstract

In addition to understanding the mechanism of action for a specific drug candidate, information regarding degradation pathways/products under various stress conditions is essential to know about their short- and long-term effects on health and environment. In line with that, tenofovir disoproxil fumarate (TDF, a co-crystal form of the prodrug tenofovir with fumaric acid), particularly used as an antiretroviral drug for treatment of HIV and hepatitis-B among others, is subjected to primarily thermal and other ICH-prescribed forced degradation conditions and their various degradation products are identified. Upon thermal degradation at 60°C for 8 h, five different degradants (namely DP-1 to DP-5) are isolated, and their structures are unambiguously confirmed using advanced analytical and spectroscopic techniques including ultra-performance liquid chromatography–mass spectrometry (UPLC-MS), high-resolution mass spectrometry (HRMS), state-of-the-art 1- and 2-dimensional nuclear magnetic resonance (1D and 2D NMR), and Fourier-transform infrared spectroscopic (FT-IR) techniques. Among fully characterized five degradants, two new degradants (DP-2 and DP-4) are identified which can potentially impact the stability of TDF via different pathways. Plausible mechanisms leading to all five thermal degradation products are also proposed including the generation of carcinogenic formaldehyde for some cases. The present systematic structural study especially combining MS and advanced NMR investigations unequivocally confirms the structures of the degradants and opens opportunities for connecting the various degradation pathways especially for the TDF-related pharmaceutical candidates.

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Data Availability

All the data for this work are represented either in the manuscript or in the supplementary information of the manuscript (https://doi.org/10.1208/s12249-023-02598-5).

Abbreviations

TDF:

Tenofovir disoproxil fumarate

ICH:

International Council for Harmonization

UPLC-MS:

Ultra-performance liquid chromatography-mass spectrometry

FTIR:

Fourier transform infrared spectroscopy

HRMS:

High-resolution mass spectrometry

NMR:

Nuclear magnetic resonance spectroscopy

SQD:

Single quadrupole detector

DAD:

Diode array detector

ESI:

Electron spray ionization

gDQF-COSY:

Gradient double quantum filtered correlated spectroscopy

g-HSQC:

Gradient heteronuclear single quantum coherence spectroscopy

g-HMBC:

Gradient heteronuclear multiple bond coherence spectroscopy

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Acknowledgements

Heartful thanks to Aragen Life Sciences Pvt. Ltd. management for support and providing the laboratory facility to perform this research.

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Authors and Affiliations

  1. Analytical Discovery Chemistry, Aragen Life Sciences Pvt. Ltd., IDA Nacharam, Hyderabad, Telangana, 500076, India

    Mahesh Ranga, Arun Kumar Modini, Raju Doddipalla & Muralidharan Kaliyaperumal

  2. Department of Chemistry, School of Applied Science and Humanities, Vignan’s Foundation for Science, Technology and Research (Deemed to be University), Vadlamudi, Guntur, 522 213, Andhra Pradesh, India

    Mahesh Ranga, Arun Kumar Modini, Anil Kumar Nallajarla & Anandarup Goswami

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  1. Mahesh Ranga
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  2. Arun Kumar Modini
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Contributions

MR and AG conceived the idea. MR performed the studies. AG, AKN and MR analyzed the results. AKM supported for purification of Thermal degradation products, RD helped in the literature survey and MK helped with the HRMS analysis of degradation products. All authors contributed in compiling the data and writing the manuscript.

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Correspondence to Anandarup Goswami.

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Ranga, M., Modini, A.K., Nallajarla, A.K. et al. Degradants of Tenofovir Disoproxil Fumarate Under Forced Yet Mild Thermal Stress: Isolation, Comprehensive Structural Elucidation, and Mechanism. AAPS PharmSciTech 24, 139 (2023). https://doi.org/10.1208/s12249-023-02598-5

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