By changing the secondary reagent (1-ethyl-3-(3-dimethylaminopropyl) carbodiimide instead of sodium cyanoborohydride), 4-APEBA is also applicable to the selective derivatization of carboxylic acids. Derivatization can be achieved under mild conditions (pH 5.7, 10 ☌). The new derivatization reagent, 4-(2-((4-bromophenethyl)dimethylammonio)ethoxy)benzenaminium dibromide (4-APEBA) contains a bromophenethyl group to incorporate an isotopic signature to the derivatives and to add additional fragmentation identifiers, collectively enhancing the abilities for detection and screening of unknown aldehydes. This original approach allowed us to evidence the occurrence of expected aldehydes according to their fatty acid precursors in the diet, and to characterize other aldehydes differentiating the different diets.īased on the template of a recently introduced derivatization reagent for aldehydes, 4-(2-(trimethylammonio)ethoxy)benzeneaminium dibromide (4-APC), a new derivatization agent was designed with additional features for the analysis and screening of biomarkers of lipid peroxidation. Derivatized aldehydes were first selectively detected on the basis of their isotopic pattern, then annotated and finally identified by tandem mass spectrometry. Following a metabolomics workflow, the developed methodology was applied to the characterization of aldehyde compounds formed by lipid peroxidation in rats fed two different diets differentially prone to lipoperoxidation. This original approach is based on the introduction of a characteristic isotopic labeling by selective derivatization of the carbonyl function using a brominated reagent. This work aimed at the development of an untargeted approach using high-performance liquid chromatography coupled with high-resolution mass spectrometry (HPLC–HRMS) for tracking aldehydes in both suspect screening and untargeted methods in fecal water, representing the aqueous environment of colon epithelial cells. Lipid peroxidation and subsequent formation of toxic aldehydes, such as 4-hydroxynonenal, is known to be involved in numerous pathophysiological processes, possibly including the development of colorectal cancer. Finally, plasma treated with radical inducer shows elevated aldehyde species compared to untreated plasma. An in vitro model shows how derivatized aldehydes in human and rat plasma are detected. The system is robust with low intra-/inter-day variation in retention time and peak area.
Detection limits of 0.5 nM can be reached for the detection of the derivatized aldehydes. Injection volumes up to 50 microl can be injected without overloading the WCX column. The WCXE column gives the advantages of direct injection of the sample after protein precipitation and centrifugation into the WCXE-LC-MS/MS system. Using a derivatization cocktail consisting of 4-(2-(trimethylammonio)ethoxy)benzenaminium dibromide (4-APC) and NaBH3CN in the screening and detection of known and unknown aldehyde biomarkers, one can take advantage of the specific fragmentation characteristics of this derivatization reagent in MS/MS. Aldehydes are important as biomarkers for oxidative stress. The system enables the selective purification and re-concentration of the in-vial derivatized aldehydes from plasma and urine samples. This paper focuses on the development and optimization of an on-line weak-cation exchange SPE (WCXE) coupled to gradient HPLC with tandem MS detection.
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