Direct RP‐HPLC determination of underivatized amino acids with online dual UV absorbance, fluorescence, and multiple electrochemical detection

Abstract

Original PaperDirect RP-HPLC determination of underivatized amino acids with online dual UV absorbance, fluorescence, and multiple electrochemical detectionThe combined use of a dual-UV detector, a fluorimetric one and of a multiple electrochemical (EC) detector equipped with a dual electrode, consisting of a conventional size 3 mm diameter glassy carbon electrode (GCE) and of a pair of 30 lm thick carbon microfibers, is proposed for the determination of 15 amino acids, two dipeptides and creatinine. This online coupling of the above detection modes could partially replace amino acid analysis by derivatization methods, since it solves problems concerning the direct detection of selected underivatized amino acids. Additionally, it was proved that the use of multiple-detection allows positive peak identification in a single chromatographic run, yields more information for free amino acids and solves in some cases the problem of chromatographic resolution. In order to optimize the detection conditions of the underivatized amino acids and related compounds by different detectors, their detection characteristics were determined by adequate preliminary experiments. The electro-oxidation characteristics of the underivatized compounds of interest were determined by hydrodynamic voltammetry using a flow cell with a macrodisc GCE and by ex-situ voltammetry using both a GCE of conventional size and a carbon fiber disk microelectrode. Important practical advantages of microfiber and microdisk electrodes with respect to macroelectrodes were demonstrated.Keywords: Dual electrode / Microdisc electrode / Multiple UV-fluorescence-electrochemical detection / Reversed phase liquid chromatography (RPLC) / Underivatized amino acids and related compounds /

A targeted metabolomic protocol for short-chain fatty acids and branched-chain amino acids

“…For example, previous studies employed various techniques, such as direct infusion into capillary electrophoresis (CE) (Arellano et al 2000; Barbas et al 1998; Poinsot et al 2012), high performance liquid chromatography (HPLC) (Agrafiotou et al 2009; Albert et al 1997; Kotani et al 2009; Stein et al 1992), or gas chromatography (GC) (Bachmann et al 1979; Zhao et al 2006). However, these methods have disadvantages of low sensitivity for SCFAs or BCAAs determination in complex biological matrix.…”

The Modern Art of Identification of Natural Substances in Whole Plants

“…Their detection is ensured by the electrical current resulting from oxidation or reduction reactions in the detector cell. Such a detection method allows the quantification of sugars, (Corradini et al 2012) and amino acids (Marioli and Sereno 1996;Agrafiotou et al 2009). The use of ECD is obviously not reserved to nonabsorbing compounds and it may possibly also be applied to the detection of catechins (Maoela et al 2009) resveratrol (Kolouchova-Hanzlikova et al 2004) and many other antioxidants in medicinal plants.…”

Study on urinary metabolic profile of phenylketonuria by micellar electrokinetic capillary chromatography with dual electrochemical detection—Potential clinical application in fast diagnosis of phenylketonuria

“…Therefore, combining the universal C 4 D with the high selective and sensitive AD, the dual electrochemical detection (C 4 D/AD) could detect more kinds of compounds just in one run regardless of their electroactivity, avoiding repeated measurements, thus greatly reducing the assay cost. A variety of dual detection schemes have been proposed for chromatographic methods such as HPLC [26] and CE [27], while, to the best of our knowledge, CE coupled with C 4 D and AD (CE-C 4 D/AD) has not been employed for the determination of marker compounds of PKU disease [28]. In our previous work, CE with dual electrochemical detection method was applied for the determination of PKU marker compounds in the spiked urine samples of healthy volunteers [29].…”