Phenols quantification

High-performance liquid chromatographic separation with electrochemical detection may provide the best sensitivity for phenol quantification in biological samples. The use of gas chromatography with a flame ionization detector may be a more versatile method, if other non-ionic pollutants must be quantified. The advantages and disadvantages of different methods available for the quantification of phenol and metabolites in biological and environmental samples have been discussed by Tesarova and Packova(1983). 

A search of federal research programs has indicated that no additional research is in progress for improving the method of phenol quantification in biological or environmental samples. 

HPLC-DAD is still the main technique used for phenolics quantification, a parameter not usually determined by other techniques such as LC-MS, given its poor sensibility. 

When two compounds with very close, or even superimposable, retention times and identical UV spectra are found in a chromatogram, more sophisticated detectors that can yield much more structural information are required. Liquid chromatography-diode array detection (LC-DAD) is still the main technique used for phenolics quantification, a parameter not usually determined by other techniques, such as liquid chromatography-mass spectrometry (LC-MS), given its poor sensibility. 

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). 

The use of HPLC for quantification of phenols is often limited to a single class of phenolics and then often only to low-molecular weight compounds that are available as standards. It is, therefore, often necessary to use colorimetric assays such as the Folin-Ciocalteau assay which rely on the reducing ability of phenols to quantify the amount of total phenolics in a sample (Waterman and Mole, 1994 Singleton et al, 1999 Schofield et al, 2001). The degree of condensation of polyphenols can be quantified by colorimetric assays such as the acid-butanol assay and the vanillin assay (Waterman and Mole, 1994 Schofield et al, 2001). 

Amiot, M. Aubert, S., Gonnet, M., and Tacchini, M. (1989). Phenolic composition of honeys Preliminary study on identification and group quantification. Apidologie 20, 115-125. 

Plasticiser/oil in rubber is usually determined by solvent extraction (ISO 1407) and FTIR identification [57] TGA can usually provide good quantifications of plasticiser contents. Antidegradants in rubber compounds may be determined by HS-GC-MS for volatile species (e.g. BHT, IPPD), but usually solvent extraction is required, followed by GC-MS, HPLC, UV or DP-MS analysis. Since cross-linked rubbers are insoluble, more complex extraction procedures must be carried out. The determination of antioxidants in rubbers by means of HPLC and TLC has been reviewed [58], The TLC technique for antidegradants in rubbers is described in ASTM D 3156 and ISO 4645.2 (1984). Direct probe EIMS was also used to analyse antioxidants (hindered phenols and aromatic amines) in rubber extracts [59]. ISO 11089 (1997) deals with the determination of /V-phenyl-/9-naphthylamine and poly-2,2,4-trimethyl-1,2-dihydroquinoline (TMDQ) as well as other generic types of antiozonants such as IV-alkyl-AL-phenyl-p-phenylenediamines (e.g. IPPD and 6PPD) and A-aryl-AL-aryl-p-phenylenediamines (e.g. DPPD), by means of HPLC. 

A method suitable for quantification of the functional class of bis(ethanol)amine antistatics, which lack UV chromophores, consists of reaction with methyl orange [53]. Atmer 163 (alkyl-diethanol amine) has been determined as a yellow complex at 415 nm after interaction with a bromophenol/cresole mixture [64]. Hilton [65] coupled extracted phenolic antioxidants with diazotised p-nitroaniline in strongly acidic medium and carried out identification on the basis of the visible absorption spectrum in alkaline solution. The antioxidant Nonox Cl in... 

Sidwell and Zondervan [10] used LC-MS with APCI detection for the identification and quantification of extractable antioxidants from food-contact plastic materials. Identification is based on the presence of the molecular ion (M + FI)+, (M—H) , other key ions or on further ion breakdown (MSn) transitions. The following antioxidant/stabiliser types were examined hindered phenols,... 

Gao L and Mazza G. 1995. Characterization, quantification, and distribution of anthocyanins and colorless phenolic in sweet cherries. J Agric Food Chem 43 343—346. 

Rivera-PastarnaD, YahiaEM and Gonzalez-Aguilar G. 2009. Identification and quantification of carotenoids and phenolic compounds in papaya using mass spectroscopy. In preparation. 

In order to preserve, as much as possible, the phenolic content in fruit and vegetable samples, the literature proposed the application of cold temperatures, even reaching to freezing, when lyophilization is the objective. These procedures also could inactivate the enzymes. The freeze-drying is largely the main preservation technique used in the studies related to the identification and quantification of the phenolic compounds of fruit... 

A number of spectrophotometric methods for the quantification of phenolic compounds in plant materials have been developed. Based on different principles, these assays are used to determine various structural groups present in phenolic compounds. Spectrophotometric methods may quantify all extractable phenolics as a group (Marshall and others 2008), or they may determine a specific phenolic substance such as sinapine (Ismail and Eskin 1979) or a given class of phenolics such as phenolic acids (Brune and others 1989). 

The Folin-Denis assay is used as a procedure for the quantification of total phenolics in plant materials, food, and beverages. Reduction of phosphomolybdic-phosphotungstic acid (Folin-Denis reagent) to a blue-colored complex in an alkaline solution occurs in the presence of phenolic compounds (Folin and Denis 1912). 

De la Torre-Carbot K, Jauregui O, Gimeno E, Castellote AI, Lamuela-Raventos RM and Lopez-Sabater M. 2005. Characterization and quantification of phenolic compounds in olive oils by solid-phase extraction, HPLC-DAD, and HPLC-MS/MS. J Agric Food Chem 53(11 ) 4331 —4340. 

These experts collectively have knowledge of phenol s physical and chemical properties, toxicokinetics, key health end points, mechanisms of action, human and animal exposure, and quantification of risk to humans. All reviewers were selected in conformity with the conditions for peer review specified in Section 104(I)(13) of the Comprehensive Environmental Response, Compensation, and Liability Act, as amended. 

Capillary columns may provide the best method for the separation of phenols prior to their quantification (Eichelberger et al. 1983 Shafer et al. 1981 Sithole et al. 1986). Of the various methods available for detection, the two commonly used methods that are most sensitive are mass spectrometry and flame ionization detection. Although electron capture detectors provide good sensitivities for higher chlorine-substituted phenols, they are poor for phenol itself (Sithole et al. 1986). The best method for the quantification of phenol may be mass spectrometric detection in the selected ion mode, but the loss of qualitative information may be significant (Eichelberger et al. 1983). 

The changes in metabolite concentrations in human blood, urine, or other appropriate biological media over time may be useful in estimating phenol s rate of metabolism in humans. In some instances, the quantification of metabolites may be useful in correlating the exposure doses to the human body burden. Studies that correlate phenol exposure with levels of metabolites in human biological matrices are not available for this compound, although analytical methods for the quantification of the metabolites are available. 

Methods used for the detection of PAs in cmde or partially purified extracts can also be adapted for post-column analysis after fractionation (see below). Direct quantitative analysis of PAs in crude grape phenolic extracts is often impossible due to the complex sample matrix. Thus, fractionation or purification is often necessary before analysis. The Folin-Ciocalteu and Pmssian Blue assays are widely used for the quantification of total polyphenols in plants [27,28]. These methods are not specific for PAs due to the reaction of other phenolic compounds with these reagents. 

Stobiecki, M. and Makkar, H.P.S., Recent advances in analytical methods for identification and quantification of phenolic compounds, EAAP Publ, 110, 11-28, 2004. 

Whilst the updating aspect of the chapters is seen as the prime contribution of this book, an effort also has been made to include a summary of previous knowledge in the field to enable the reader to place new advances in this context. Chapters 1 and 2 review the application of contemporary isolation, quantification, and spectroscopic techniques in flavonoid analysis, while Chapter 3 is devoted to molecular biology and biotechnology of flavonoid biosynthesis. Individual chapters address the flavonoids in food (Chapter 4) and wine (Chapter 5), and the impact of flavonoids and other phenolics on human health (Chapter 6 and, in part, Chapter 16). Chapter 8 reviews newly discovered flavonoid functions in plants, while Chapter 9 is the first review of flavonoid-protein interactions. Chapters 10 to 17 discuss the chemistry and distribution of the various flavonoid classes including new structures reported during 1993 to 2004. A complete listing of all known flavonoids within the various flavonoid classes are found in these later chapters and the Appendix, and to date a total of above 8150 different flavonoids has been reported. 

A similar study by Verrelli and coworkers, based on PCA of potentio-metric data, allowed the differentiation of Verdicchio wines according to the winemaker and the vintage. Furthermore, MLR and PLS models, validated by means of cross-validation, permitted the quantification of a number of oenological parameters, such as SO2, L-malic acid, and the total phenols content (Paolesse et al., 2008 Verrelli et ah, 2007). 

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