Free Radical Scavenging Assay

Abstract Recently, we have investigated Aconitum cochleare Woroschin and obtained three new alkaloids cochleareine, acoleareine from the aerial parts of the plant and cochleareinine from the roots. Cochlearenine exhibited antioxidant activity against DPPH free radical scavenging assay. The cardio active effect of has also have been studied on isolated heart preparations. 

In Xanthine oxidase inhibition assay, the SFE extract show almost no bioactivity as the case of the LSE. In the oxygen free radical scavenger assay, the SFE extracts of Schizandrae fructus and Moutan Cortex Radicis were more active than the case of the LSE. 

In an analysis of the different components of the extracts of the roots and leaves of E. purpurea, E. angustifolia, and E. pallida, all possessed antioxidant properties in a free-radical scavenging assay and in a lipid peroxidation assay (26). 

Antioxidant activity of petroleum ether, chloroform, ethyl acetate and methanol extracts of Usnea pictoides lichen was determined by DPPH free radical scavenging assay and ferric reducing assay (Pavithra et al. 2013). The scavenging potential of methanol extract was higher than other extracts, and also, in ferric reducing assay, methanol extract showed stronger reducing power than other extracts. 

The aqueous and ethyl acetate flower extracts rich in luteolin and luteolin-7-gluco-side exhibited antioxidant activity in the 2,2-iphenyl-l-picrylhydrazyl (DPPH) free radical scavenging assay as well as in the phosphatidylcholine liposome assay. Lower concentrations, however, had a pro-oxidant effect. 

Alcoholic extracts of the roots and leaves of E. angustifolia, E. pallida, and E. purpurea exhibited comparable in vitro antioxidant activities in ABTS free radical scavenging assay, and the root extracts were also active in the lipid peroxidation inhibition assay.Methanolic extracts of freeze-dried roots of the three species act as in vitro antioxidants through the scavenging of DPPH and ABTS radicals, as well as by chelation of the copper ion (Cu Copper-catalyzed oxidation of human LDL was further demonstrated in vitro by different preparations from E. purpurea root containing alkamides, caffeoyl derivatives, and polysaccharides, as well as by pure caffeoyl derivatives. The antioxidant effect was synergistic and dose dependent. ... 

Choi CW, Kim SC, Hwang SS, Choi BK, Ahn HJ, Lee MY, Park SH and Kim SK. 2002. Antioxidant activity and free radical scavenging capacity between Korean medicinal plants and flavonoids by assay-guided comparison. Plant Sci 163(6) 1161-1168. 

Free radical scavenging activity was assessed using the DPPH (1, 1-di-phenyl-2-picrylhydrazyl) assay as described by Harbilas et al. [22] with incubation time increased to 65 min. Briefly, 250 tiL of 100 timol/L DPPH dissolved in methanol was added to 40 p.L of extract (tested at 5 concentrations) in a microplate well. A standard curve of ascorbic acid (positive control) was included as a reference and all data were blanked against a treatment with only methanol. Absorbance was read with a microplate reader at 517 nm. The inhibitory concentration for 50% scavenging (IC50) of each extract was calculated and compared to the IC50 of the ascorbic acid standard curve. 

To this end, the authors perform a second comet assay. Only limited results, however, are shared PM2 5 from only one city (Baton Rouge) was tested using only one cell line (K562 myeloid leukemia cells), thereby following a broad-to-narrow approach. The results are presented in excerpt 4E. As shown in Eigure 3 (excerpt 4E), cells were exposed to (A) a blank extract, (B) a PM2 5 extract, (C, D, E) a PM2 5 extract mixed with one of three different free radical scavengers, (E) a positive control, and (G, El) a PM2 5 extract mixed with one of two different metal-ion chelators. The free radical scavengers remove free radicals the... 

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. ... 

Most stilbenoids possess antioxidant activities because they possess polyphenol functions in the molecules. Some of their beneficial effects, hepatoprotective action, cardiovascular protection, for instance, are in close relation to their antioxidant activities. Several models have been employed in the assay such as lipid peroxidation system, human low-density lipoprotein model, xanthine oxidase system and l,l-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging model, which is the most commonly used protocol. 

Bagchi et al. (1997) studied the oxygen free-radical scavenger ability of grape seed proanthocyanidins using a chemiluminescence assay and cytochrome c reduction. The results revealed that on a weight basis (100 mg/L), the grape seed proantho-cyanidin extract was a better inhibitor of both superoxide anion and hydroxyl radical than vitamin C and vitamin E succinate. 

The free-radical-scavenging activity of compounds was evaluated in the TEAC and CL assays. The first measures the relative abiUty of antioxidant substances to scavenge the radical cation 2,2 -azinobis(3-ethylbenzothiozoline-6-sulfonate) (ABTS" ) as compared to a standard amount of the synthetic antioxidant Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid). The CL assay measures the inhibition of iodophenol-enhanced chemiluminescence by a horseradish peroxidase/perborate/luminol system. Trolox was used as the reference antioxidant. The results showed that xanthones exhibited free-radical-scavenging activity at potency levels comparable to those of reference antioxidant compounds quercetin and rutin, while the benzophenone and phloroglucinol type compoimds had more moderate activities [87]. 

The free radical-scavenging capacity of oleuropein and hydroxy-tyrosol was tested, using some radical generators. As a radical initiator, AAPH 2,2 -azobis (2-amidinopropane)-hydrochloride) was used for peroxyl radicals generation, because it is a common free radical found in the body [59] and has often been used in several antioxidant activity assays [60]. It appears to be slightly less reactive than OH-radical [61]. 

Recently, an indirect method to measure the free radical scavenging activity of extracts and, thus, their antioxidant activity has been developed using HPLC coupled to a coulometric array detector system [61]. This was developed to characterise the overall antioxidant activity status of fruit and vegetables. A significant positive linear correlation was demonstrated between the total antioxidant activity determined by using the oxygen radical absorbance capacity assay and that measured using the electrochemical data obtained from the coulometric array detectors. 

---