Evaluation of the Methods for Determination of the Free Radical Scavenging Activity by DPPH

G. MARINOVA and V. BATCHVAROV
Institute of Cryobiology and Food Technologies, BG - 1407 Sofia, Bulgaria

Abstract

MARINOVA, G. and V. BATCHVAROV, 2011. Evaluation of the methods for determination of the free radical scavenging activity by DPPH. Bulg. J. Agric. Sci., 17: 11-24

The survey of the methods for determination of free radical scavenging activity by DPPH has been done.The differences between methods conditions and their evaluation are presented. It was determined the effect of methods conditions by ruggedness testing of methods. It was specified that the accuracy of the method for determination of free radical scavenging activity is effected by the solvent used (ethanol or methanol) and the sample/reagent DPPH volume ratio. The coefficient of variation of the method with ethanol is twice lower that the respective one determined with using of methanol. The calibration curves with ascorbic acid (Vitamin C) and α-Tocopherol (Vitamin E) and solvent ethanol and methanol were plotted. They are characterized with very high regression coefficients. Based on the analysis and evaluation of the methods, the results of ruggedness testing of methods, coefficient of variations of determination with solvent ethanol and methanol and recommendations of some authors it was proposed modification of the method for determination of free radical scavenging activity of beer and beverages with DPPH. The modification of the method includes: 0.06 mM solution of DPPH in ethanol, reaction mixture 1.5 ml diluted sample and 1.5 ml DPPH solution, 30 minutes time of reaction in dark, measurement of absorbance at 517 nm, presentation of the results as equivalent of Vitamin C antioxidant activity. It was investigated the effect of malt and hops on the antioxidant activity of wort and beer. It was established that the main free radical scavenging activity of beer is attributed by the malt used. The hopping increases additionally the values of the parameter. During the different stages of the brewing process the free radical scavenging activity is changed. The differences between the free radical scavenging activity of laboratory and production beers indicated the very important role of raw materials and technology used. The free radical scavenging activity of beers determined by ethanol is higher (an average 8,2 - 38.9 % for the used beer samples) than the values obtained by solvent methanol.

Key words: method DPPH, modification of method, ethanol, methanol, malt, beer

Abreviations: DPPH - 2,2-Diphenyl-1-picrylhydrazyl, BHA - Butylated xydroxy anisole, BHT - Butylated hydroxy toluene, EVCAA - equivalent vitamin C antioxidant activity, EVEAA - equivalent vitamin E antioxidant activity, FRSA - free radical scavenging activity, EBC - European Brewery Convention

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