Abstract
ABSTRACTThe influence of extraction medium and sample preparation on the total antioxidant capacity (TAC) of pea plants leaves is investigated. Sodium acetate buffer (pH 5), potassium phosphate buffer (pH 7.5), methanol or 0.1% w/v trichloroacetic acid (TCA) were used for homogenization. TAC was measured in the crude supernatants (one-step procedure). The pellets were extracted with acetone and their TAC values were added to those from the previous assay (two-step procedure). The values for TAC of the acetone extract, the low-molecular and protein fractions derived from a partial fractionation of the probes are used in the proposed by us new three-step method. The results indicate that TAC of the probes processed with the two or three-step procedure is significantly higher than that of the crude supernatants (one-step procedure). On the basis of this study we recommend two-step protocol as a minimum and buffer with slightly acid pH or TCA for extraction when plant material is investigated. The highest results were obtained when the new three-step procedure for sample preparation and sodium acetate buffer as an extracting agent were applied. Moreover, the separation of the cellular antioxidants into three different fractions is especially useful for studying the effects of aging, processing and storage on the antioxidant capacity.
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