DPPH-Assay

Sanchez-Morcno (2002) considered that this assay is an easy and accurate method for determining antioxidant capacity in fruit and vegetable samples. The DPPH assay has been used to determine the antioxidant activity of polyphenols (Sanchez-Moreno and others 1998 Bao and others 2004) flavonols (Jimenez and others 1998 1999 Choi and others 2002) anthocyanin-based natural colorants from berries (Espin and others... 

However, the DPPH assay is not suitable for measuring the antioxidant capacity of plasma because protein is precipitated in the presence of the ethanol/methanol solvent used. 

Fig. 7.3 Radical scavenging activity as measured in the DPPH assay versus (a) total ginsenoside content (% w/w) and (b) total phenolics (pg quercetin per mg extract) of eight ginseng root samples (n = 3). The log IC50 of each sample was divided by the log IC50 of the standard ascorbic acid to...

Glavind and Holmer proposed a method of determination of antioxidants by TLC using the DPPH radical. They sprinkled a plate with separated substances with methanol solutionofthe radical and observed discoloration where substances able to quench radicals were present [57]. The TLC-DPPH assay allows a researcher to access the analysed substances and to assess the biological activity of individual compounds. Another advantage of the method is the possibili-... 

Muller L, Frohlich K, Bohm V. Comparative antioxidant activities of carotenoids measured by ferric reducing antioxidant power (FRAP), ABTS bleaching assay (aTEAC), DPPH assay and peroxyl radical scavenging assay. Food Chemistry. 2011 129 139-148... 

Popular EPR-based assays of antioxidant activity include the DPPH assay, in which the ability of compounds to quench (by H-atom transfer) the 1,1-diphenyl-2-picrylhydrazyl radical is used to rank their antioxidant activity. This method has been applied widely to the analysis of dietary antioxidants and extracts from medicinal plants.213 219 Extensive use has also been made of assays based on the competition between spin traps and test compounds for reaction with enzymatically-generated 02 and, in the presence of a metal catalyst, the OH rad-... 

A19. Atsumi, T., Iwakura, I., Kashiwagi, Y., Fujisawa, S., and Ueha, T., Free radical scavenging activity in the nonenzymatic fraction of human saliva A simple DPPH assay showing the effect of physical exercise. Antioxid. Redox Signal. 1, 537-546 (1999). 

DPPH (l,l-diphenyl-2-picryhydrazyl) is a purple-colored stable free radical that is reduced to the yellow-colored diphenylpicrylhydrazine by free radicals. The DPPH assay measures one electron, such as hydrogen atom donating activity and hence provides a measure of free radical scavenging activity. This assay is suitable for the initial screening of multiple samples, such as plant extracts. Reaction mixtures containing test samples dissolved in DMSO and DPPH in absolute ethanol are incubated at 37°C for 30 min in a 96-well plate and absorbance measured at 515 nm. ... 

A Chinese isolate of Penicillium terrestre derived from sediments yielded a series of new gentisyl alcohol polymers, including the trimeric terrestriol A (89) and the dimeric terrestrols B-H (90-96)/° All compounds showed moderate cytotoxicity toward four different cancer cell lines as well as moderate radical scavenging activity in the DPPH assay. Furthermore, 95 displayed moderate inhibitory activity against protein tyrosine kinases Src and KDR. 

Chemical investigation of the endophytic fungus Chaetomium globosum, which was isolated from the inner tissue of the Chinese marine red alga Polysiphonia urceolata, resulted in the isolation of chaetopyranin (107), a new benzaldehyde secondary metabolite. Compound 107 displayed moderate radical scavenging activity in the DPPH assay, and also exhibited moderate to weak cytotoxicity toward several tumor cell lines. 

Several studies described the interaction of flavonoids with the 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical [76, 79, 107-108] (Fig. (3)). DPPH is a stable free radical and is frequently used in ESR studies [109]. The DPPH assay provides information on the reactivity of flavonoids with a stable free radical [79]. Because of its odd electron, DPPH gives a strong absorption band in ethanol at 517 nm. As this... 

The antioxidant activity using DPPH ranged from 21 mg to 361 mg VGE per g dry weight. The overall antioxidant capacity of plant parts in VCEAC which was evaluated by DPPH assay decreased in the following order T. macroptera root bark > A. leiocarpus leaves >T. macroptera trunk bark > C. populnea root bark > Z. mucronata leaves = A. leiocarpus trunk bark... 

For lignin fractions soluble in methanol it has been confirmed that the OHphen group plays a crucial role in radical scavenger activity of lignin in the DPPH assay. 

Antioxidant molecules themselves are not free radicals and are not EPR-active. However, these compounds do react rapidly with reactive oxygen species, ROS, and other free radicals to render them harmless. The EPR-based DPPH assay described below can be used to provide quantitative information concerning the relative effectiveness of various dietary substances and their antioxidant capacities. 

Figure 6 Flow chart for the DPPH assay procedure using the e-scan with LC autosampler.

Figure 7 Antioxidant capacity measurements obtained by the DPPH method. EPR signal intensities were determined at 60 s after mixing of the DPPH assay solution with each of the samples listed at the bottom of the automatically generated chart. From left to right, the test solutions were distilled water (control), ascorbic acid at 5, 2.5, 1.25, 0.625, and 0.3125 mM, distilled water, white wine, red wine.

Electron Paramagnetic Resonance (EPR, ESR) spectroscopy is a unique tool to answer specific questions evolving radical processes in both research and development (R D) as well as quality control (QC). Key QC examples comprise the food irradiation control for customer care according to European Union (EU) norms and the flavour stability and shelf life assessment on fresh beer. For the first time, an automated DPPH-assay to measure the effectiveness of antioxidants by CW-EPR is reported. 

Note 5 On applying the DPPH assay, the determined ICj value for arbutin ( 768 mM) represents a 176 times lower antioxidant capacity than the one determined for quercetin (4.36 mM). Although the arbutin ICj(, value correlates well with those published by other investigators applying similar DPPH-assay conditions [23], it should be pointed out that there is an experimental conflict The DPPH assay, which uses alcohols (methanol) as the l,l-diphenyl-2-picrylhydrazyl-radical dissolvent, is preferably used to investigate the antioxidant capacity of lipophilic (alcohol... 

Antioxidant activity is also a measure for substance ability to prevent free radical concentration increment, oxidative stress, and risk for development-related diseases. And for the purpose of measuring antioxidant activity, many assays are applied such as 2,2-diphenyl-l-picrylhydrazyl (DPPH) assay, 2,2 -azino-bis(3-ethylbenzthiazoline-6)-sulfonic acid (ABTS) assay, p-carotene bleaching test, ferric and cupric reducing power, and linoleic acid assay (Akrout et ah, 2011 Chabir et ah, 2011 Jia et al, 2010 Serrano et al, 2011). 

Thymol, p-cymene, and y-terpinene were major components of Thymus proximus Serg. and Thymus marschallianus Will, essential oils, growing wild in Xinjiang. In a dose-dependent manner, both essential oils showed results between 12,000 and 20,000 g/mL in reducing power assays, while the results of the positive controls BHT and thymol were better than from the oil. Results from the DPPH assay were also concentration dependent. T. proximus essential oil exhibited an... 

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