Identification, characterization, and in silico ADMET prediction of nirmatrelvir and its degradation products using HPLC-PDA and LC-QTOF-MS/MS

IF 1.8 3区化学 Q4 BIOCHEMICAL RESEARCH METHODS Rapid Communications in Mass Spectrometry Pub Date : 2024-08-26 DOI:10.1002/rcm.9896

Matta Ashwin Kumar, Raja Sundararajan

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Abstract

Rationale

Nirmatrelvir is a protease inhibitor that is essential for virus replication. Nirmatrelvir is indicated for the management of mild to severe cases of COVID-19 in individuals who are 12 years of age or older. Forced degradation studies of nirmatrelvir were carried out on the drug substance in solid and solution forms, subjecting it to various stress conditions according to International Conference on Harmonisation (ICH) Q1A(R2) and Q1B guidelines. The analytical method was validated as per the ICH Q2(R1) guidelines.

Methods

The drug substance (nirmatrelvir) was subjected to hydrolysis (acidic, alkaline, and neutral), thermal, photolytic, and oxidative stress conditions. Five degradation products (DPs) of nirmatrelvir formed under hydrolytic (acidic and alkaline) and oxidative (2,2-azobisisobutyronitrile) stress conditions. These degradation products were identified and separated using reverse-phase HPLC on a phenomenex kinetex C8 column (250 mm × 4.6 mm × 5 μm) with gradient elution. The mobile phase consisted of 0.1% formic acid and acetonitrile, and detection was carried out at a wavelength of 210 nm.

Results and conclusions

Nirmatrelvir and its five DPs were efficiently separated using reverse phase–HPLC. These five DPs were identified and characterized using LC-electrospray ionization (ESI)-Q-TOF-coupled mass spectrometry analysis in the ESI-positive ionization mode. The formation mechanisms of the DPs and the most probable mass fragmentation pathways for both nirmatrelvir and its DPs were elucidated. The developed method demonstrated selectivity, accuracy, linearity, and reproducibility, making it appropriate for quality control of nirmatrelvir and future research studies. Additionally, the physicochemical and Absorption, Distribution, Metabolism, Excretion, and Toxicity (ADMET) properties of nirmatrelvir and its DPs were predicted using ADMET predictor software. The toxicity profile revealed that DP2 and DP3 have teratogenic effects while DP1 and DP3 caused phospholipidosis.

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使用 HPLC-PDA 和 LC-QTOF-MS/MS 对尼尔马特韦及其降解产物进行鉴定、表征和硅学 ADMET 预测。

理由：Nirmatrelvir 是一种蛋白酶抑制剂，对病毒复制至关重要。Nirmatrelvir 适用于治疗 12 岁或以上的 COVID-19 轻度至重度病例。根据国际协调会议（ICH）Q1A(R2)和Q1B指南，对固体和溶液形式的药物物质进行了强制降解研究，将其置于各种应力条件下。分析方法按照 ICH Q2(R1) 指南进行了验证：对药物（尼尔马特韦）进行水解（酸性、碱性和中性）、热解、光解和氧化应激试验。在水解（酸性和碱性）和氧化（2,2-偶氮二异丁腈）应力条件下，形成了五种尼尔马特韦降解产物（DPs）。采用反相高效液相色谱法对这些降解产物进行鉴定和分离，色谱柱为 phenomenex kinetex C8（250 mm × 4.6 mm × 5 μm），梯度洗脱。流动相为 0.1% 甲酸和乙腈，检测波长为 210 nm：采用反相高效液相色谱法高效分离了尼尔马特韦及其五种二磷酸甘油酯。采用液相色谱-电喷雾离子化（ESI）-Q-TOF耦合质谱在ESI正离子模式下对这五种DPs进行了鉴定和表征。该方法阐明了 nirmatrelvir 及其 DPs 的 DPs 形成机制和最可能的质量碎片途径。所开发的方法具有选择性、准确性、线性和可重复性，因此适用于尼尔马特韦的质量控制和未来的研究。此外，还利用 ADMET 预测软件预测了尼尔马特韦及其 DPs 的理化和吸收、分布、代谢、排泄和毒性（ADMET）特性。毒性曲线显示，DP2 和 DP3 有致畸作用，而 DP1 和 DP3 则会导致磷脂中毒。

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来源期刊

Rapid Communications in Mass Spectrometry 化学-分析化学

CiteScore

4.10

自引率

5.00%

发文量

219

审稿时长

2.6 months

期刊介绍： Rapid Communications in Mass Spectrometry is a journal whose aim is the rapid publication of original research results and ideas on all aspects of the science of gas-phase ions; it covers all the associated scientific disciplines. There is no formal limit on paper length ("rapid" is not synonymous with "brief"), but papers should be of a length that is commensurate with the importance and complexity of the results being reported. Contributions may be theoretical or practical in nature; they may deal with methods, techniques and applications, or with the interpretation of results; they may cover any area in science that depends directly on measurements made upon gaseous ions or that is associated with such measurements.