Alternate Strategies to Obtain Mass Balance without the Use of Radiolabeled Compounds: Application of Quantitative Fluorine (19F) Nuclear Magnetic Resonance (NMR) Spectroscopy in Metabolism

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• 作者：Abdul Mutlib ; Robert Espina ; James Atherton ; Jianyao Wang ; Rasmy Talaat ; JoAnn Scatina ; Appavu Chandrasekaran
• 刊名：Chemical Research in Toxicology
• 出版年：2012
• 出版时间：March 19, 2012
• 年：2012
• 卷：25
• 期：3
• 页码：572-583
• 全文大小：382K
• 年卷期：v.25,no.3(March 19, 2012)
• ISSN：1520-5010

文摘

Nuclear magnetic resonance (NMR) spectroscopy is playing an increasingly important role in the quantitation of small and large molecules. Recently, we demonstrated that ¹H NMR could be used to quantitate drug metabolites isolated in submilligram quantities from biological sources. It was shown that these metabolites, once quantitated by NMR, were suitable to be used as reference standards in quantitative LC/MS-based assays, hence circumventing the need for radiolabeled material or synthetic standards to obtain plasma exposure estimates in humans and preclinical species. The quantitative capabilities of high-field NMR is further demonstrated in the current study by obtaining the mass balance of fluorinated compounds using ¹⁹F-NMR. Two fluorinated compounds which were radio-labeled with carbon-14 on metabolically stable positions were dosed in rats and urine and feces collected. The mass balance of the compounds was obtained initially by counting the radioactivity present in each sample. Subsequently, the same sets of samples were analyzed by ¹⁹F-NMR, and the concentrations determined by this method were compared with data obtained using radioactivity counting. It was shown that the two methods produced comparable values. To demonstrate the value of this analytical technique in drug discovery, a fluorinated compound was dosed intravenously in dogs and feces and urine collected. Initial profiling of samples showed that this compound was excreted mainly unchanged in feces, and hence, an estimate of mass balance was obtained using ¹⁹F-NMR. The data obtained by this method was confirmed by additional quantitative studies using mass spectrometry. Hence cross-validations of the quantitative ¹⁹F-NMR method by radioactivity counting and mass spectrometric analysis were demonstrated in this study. A strategy outlining the use of fluorinated compounds in conjunction with ¹⁹F-NMR to understand their routes of excretion or mass balance in animals is proposed. These studies demonstrate that quantitative ¹⁹F-NMR could be used as an alternate technique to obtain an estimate of the mass balance of fluorinated compounds, especially in early drug development where attrition of the compounds is high, and cost savings could be realized through the use of such a technique rather than employing radioactive compounds. The potential application of qNMR in conducting early human ADME studies with fluorinated compounds is also discussed.