Flavonoid spectrophotometric methods

Direct ultraviolet spectrophotometric methods have been developed to measure naringin in grapefruit (19J and hesperidin in orange juice (20, 21j. While these methods are rapid, they are also nonspeciTTc for flavonoid bitterness. 

A spectrophotometric method for the detection of o-dihyrdoxyl groups in flavonoid compounds. Arch. Biochem. Biophys. 63, 376 (1956). 

Spectrophotometric methods are sensitive but not really suitable for estimating saponins in crude plant extracts since the reactions are not specific and colored products may form with accompanying compounds such as phytosterols and flavonoids. Similarly, hemolytic methods also suffer from a lack of specificity. Consequently, alternative ways of analyzing for saponins using different chromatographic techniques have been developed. 

Spectrophotometric methods especially colorimetric methods are nowadays intensively used for the quantification of different classes of polyphenols (total phenolic content, tannins content, flavonoids, and anthocyanins contents). 

Different spectrophotometric methods for the quantification of phenolic compounds in foods have been developed. Spectrophotometric methods are based on the formation of a compound or colored complex that is measured at a certain wavelength. To avoid interference, an effective extraction of flavonoids is necessary before spectrophotometric measurement. 

Quantification of individuals The foregoing quantitative methods provide rapid estimation of the total flavonoids or their subgroups however, they offer no information on specific flavonoids. To analyze the concentration of individual flavonoids, good separation procedures must first be developed, followed by quantification using various spectrophotometric or other types of detectors. Such a separation and... 

Several TLC methods have been widely used to quantitatively estimate the flavonoids for quality control purposes rather than to detect adulteration. The potential exists, however, for testing authenticity. Naringin is an important compound in grapefruit juice, since it is largely responsible for the bitter character of the juice. Fisher et al. (121) developed a TLC procedure for naringin estimation. This was later modified by Tatum and Berry (122). Swift (123) developed a TLC-spectrophotometric assay for the neutral methoxylated flavones in orange peel. The method was subsequently expanded to the determination of these compounds in orange juice (124). 

The estimation of flavonoids by measuring the UV absorbance is one of the most common and convenient methods. The spectra of a number of flavonoids was reported by Mabry et al. (1970). The availability and measuring capabilities of flavonoids UV spectra greatly facilitated their identification in complex mixtures. The spectrophotometric determination of flavonoids facilitated the quantitative and qualitative analysis of samples. In general, UV absorption can be used to determine the concentration of flavonoids in samples, e.g. citrus juices. 

The FRAP method (Benzie and Strain, 1996) is based on the measurement of the ability of the substance to reduce Fe to Fe ", which is determined spectrophotometrically through its colored complex with 2,4,6-tris(2-pyridyl)-s-triazine (TPTZ) at 595 nm. This assay was also modified to be used in 96-well micropiates for evaluation of the structure-antioxidant activity relationships of flavonoids (Firuzi et al., 2005). In this work, a good correlation was observed between the FRAP assay and electrochemical results, which confirms the reliability of the former method for the evaluation of the antioxidant activity. 

Method 3 - Physical separation of parts. Cranberries were physically separated by removing the peel and squeezing the juice from the remaining solid. Samples were prepared for HPLC analysis from the peels, solids, juice and the whole berry by extraction with 98 2 methanol/acetic acid. Solutions were passed through Whatman Puradisc 13 mm/0.45 pm filters prior to HPLC analysis. The samples were also analyzed spectrophotometrically for total flavonoid and anthocyanin content using the method of Lees and Francis (4). The data are shown in Table III. 

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