Abstract
Quantitative analysis by liquid chromatography-mass spectrometry (LC-MS) is frequently based on the application of isotopically labeled standards which usually have to be specially synthesized. Although the synthesis of deuterated isotopologues is relatively inexpensive, they are not considered as good internal standards due to the possible deuterium effect on the retention time during LC-MS analysis. We developed a method of deuterium labeling of denatonium benzoate (Bitrex) via H/D exchange of its α-carbon hydrogen atoms in CH2 group situated between carbonyl and quaternary ammonium groups. The proposed strategy is rapid, cost-efficient, and does not require derivatization reagents or further purification. The LC-MS analysis of isotopologues revealed that the introduced deuterons do not undergo back exchange under acidic conditions, and the co-elution of deuterated and non-deuterated forms is observed. The obtained deuterated standard was applied in the quantitative LC-MS analysis of Bitrex in commercially available household products.Electronic supplementary materialThe online version of this article (doi:10.1007/s00216-015-8815-y) contains supplementary material, which is available to authorized users.
Highlights
Denatonium benzoate, known as Bitrex, is a an extremely bitter compound used commonly to denature industrial alcohols and to make potential harmful household products extremely unpalatable [1]
It was found that quaternary ammonium (QA) group used as a fixed charge tag increases the ionization efficiency of peptides during ESI-MS analysis and significantly lowers the detection limit [6]
We present our studies on synthesis of the αC deuterated analogue of denatonium cation containing betaine derivative and its application in quantitative analysis of denatonium benzoate containing samples by liquid chromatographymass spectrometry (LC-MS)
Summary
Denatonium benzoate, known as Bitrex, is a an extremely bitter compound used commonly to denature industrial alcohols and to make potential harmful household products extremely unpalatable [1]. Several methods have been applied to determine denatonium benzoate in different samples, including highperformance liquid chromatography (HPLC) [2], UV-Vis spectrophotometry [3], capillary electrophoresis [4], and liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry (LC-APCI-MS) [5]. The proposed methods present several disadvantages; the investigation of the new analytical methodologies for the denatonium benzoate determination is required. Due to the low ionization efficiency of some compounds during the ESI-MS experiment, the reliable identification of trace amount of substance is often limited. It was found that quaternary ammonium (QA) group used as a fixed charge tag increases the ionization efficiency of peptides during ESI-MS analysis and significantly lowers the detection limit [6]. Many different stable isotope-labeled quantification reagents have been described [1] due to their complicated and expensive synthesis, the development of new isotopically labeled standards is still under investigation
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