Phenolic compounds HPLC-electrochemical detection

Knowledge of the identity of phenolic compounds in food facilitates the analysis and discussion of potential antioxidant effects. Thus studies of phenolic compounds as antioxidants in food should usually by accompanied by the identification and quantification of the phenols. Reversed-phase HPLC combined with UV-VIS or electrochemical detection is the most common method for quantification of individual flavonoids and phenolic acids in foods (Merken and Beecher, 2000 Mattila and Kumpulainen, 2002), whereas HPLC combined with mass spectrometry has been used for identification of phenolic compounds (Justesen et al, 1998). Normal-phase HPLC combined with mass spectrometry has been used to identify monomeric and dimeric proanthocyanidins (Lazarus et al, 1999). Flavonoids are usually quantified as aglycones by HPLC, and samples containing flavonoid glycosides are therefore hydrolysed before analysis (Nuutila et al, 2002). 

Sontag, G., Friedrich, O., Kainz, G., and Jorg, E. (1989). Determination of phenolic compounds by HPLC with electrochemical detection. Proc. EUR 5th Food Chem. Conference, Agric. Food Chem. Consum. 2, 703-707. 

The variety of detection modes available for HPLC analysis that provide additional information about the eluent as it exits the column greatly facilitates unknown characterization. The majority of analytical methods for phenolic compounds includes HPLC with spectrophotometric-based detection techniques (UV absorption, fluorescence, photo diode array—PDA) as well as HPLC with electrochemical detection. 

HPLC-based electrochemical detection (HPLC-ECD) is very sensitive for those compounds that can be oxidized or reduced at low voltage potentials. Spectrophotometric-based HPLC techniques (UV absorption, fluorescence) measure a physical property of the molecule. Electrochemical detection, however, measures a compound by actually changing it chemically. The electrochemical detector (ECD) is becoming increasingly important for the determination of very small amounts of phenolics, for it provides enhanced sensitivity and selectivity. It has been applied in the detection of phenolic compounds in beer (28-30), wine (31), beverages (32), and olive oils (33). This procedure involves the separation of sample constituents by liquid chromatography prior to their oxidation at a glassy carbon electrode in a thin-layer electrochemical cell. 

Fluorescence (FL) detection is another selective and sensitive technique. Vitamin E, a-, /3-, S-, and y-to-copherols have been analyzed by RP-HPLC using FL as well as UV detection. In addition, a newer technique, electrochemical detection, has been reported in the analysis of phenolic compounds of olive oil [7]. 

Electrochemical detection is very sensitive for the compounds that can be oxidized or reduced at low-voltage potentials. Therefore, it could also be applied in the HPLC analysis of phenolic acids that are present in natural samples at very low concentrations. With the recent advances in electrochemical detection, multi-electrode array detection is becoming a powerful tool for detecting phenolic acids and flavonoids in a wide range of samples. The multi-channel coulometric detection system may serve as a highly sensitive way for the overall characterization of antioxidants the coulometric efficiency of each element of the array allows a complete voltametiic resolution of analytes as a function of their reaction (redox) potential. Some peaks may be resolved by the detector, even if they are unresolved when they leave the HPLC column. ... 

The detection and identification of phenolic compounds, including phenolic acids, have also been simph-fied using mass spectrometry (MS) techniques on-hne, coupled to the HPLC equipment. The electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) interfaces dominate the analysis of phenohcs in herbs, fmits, vegetables, peels, seeds, and other plants. In some cases, HPLC, with different sensitivity detectors (UV, electrochemical, fluorescence), and HPLC-MS are simultaneously used for the identification and determination of phenolic acids in natural plants and related food products.In some papers, other spectroscopic instmmental techniques (IR, H NMR, and C NMR) have also been apphed for the identification of isolated phenolic compounds. 

Polarographic also can be utilized for the determination of many organic compounds present in the air, e.g. in the vicinity of factories, in the industrial atmosphere, etc. In [23] are listed compounds like formaldehyde, acrolein, acetaldehyde, furfurol, hexachlorbutadiene, and nitro-cyclohexane. The tested air is passed through a trap containing the supporting electrolyte. Application of electrochemical detection in HPLC to the measurement of toxic substances in air is the subject of a paper[24]. Derivatives of phenols and amines were chosen as examples. 

High-performance liquid chromatography (HPLC) is the most suitable technique to determine phenolic compounds in water using UV or diode-array detection (DAD) [23-29] or electrochemical detection (ECD) [30-33] but, although amperometric detection is more sensitive than UV detection, a preconcentration step is necessary in both cases to achieve the low levels allowed in real samples. LC nowadays is often the choice over GC, as it is more suitable for aqueous samples and as no derivatization step is needed for phenolic compoimds. The online coimection between SPE and the HPLC column is fairly straightforward this approach appears very suitable for the analysis of these compounds. However, the conventional ultraviolet (UV) detector is much less sensitive than most of... 

Whilst the application of electrochemical detection in HPLC is becoming established for certain classes of compounds, such as biogenic amines [2] and plant phenolic materials [3], its use for compounds that are less easily oxidized has not been widely examined. 

Air is drawn through a midget impinger or a bubbler containing 0.1 ANaOH solution. Phenol and ciesols are trapped as phenolates. The pH of the solution is adjusted <4 by H2S04. The compounds are determined by reverse-phase HPLC with UV detection at 274 nm. An electrochemical or fluorescence detector may also be used. The solution may be analyzed by colorimetric or GC-FID technique. 

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