Page 211 - Demo

SECTION I: MOLECULAR GENETIC ENGINEERING AND BIOCHEMICAL TECHNOLOGY 2152.3. Determination of total phenolic content and total flavonoid contentTotal phenolic content was determined by the Folin%u2013Ciocalteu method described by Kim et al. (2003) with some modifications [6]. Gallic acid was used as a standard and was prepared in a concentration range of 0-100 %u00b5g/mL. The supernatant of fermented MFP was obtained by centrifugation at 13000 rpm for 10 min. The reaction with Folin %u2013 Ciocalteau reagent was carried out with 100 %u00b5L of the standard or fermentation sample and 10 %u00b5L of Folin %u2013 Ciocalteau, mixed well. After 5 min of reaction, 100 %u00b5L of 7% Na2CO3 was added, mixed well, and left in the dark for 90 min before measuring OD at 750nm. The control sample was MFP without adding Lactobacillus bacteria. The experiment was repeated 3 times. The total phenolic content of the sample was determined in mg of gallic acid equivalents (GAE). The total flavonoid content was determined by the colorimetric method with Al(NO3)3 in an alkaline medium described by Zhao et al. (2018) with some modifications [7]. Quercetin standard was prepared in 5 concentrations: 0; 0.05; 0.1; 0.2; and 0.3 mg/mL with ethanol 80%. Each reaction of 100 %u00b5L of fermentation broth sample or quercetin standard was mixed with 20 %u00b5L of 10% Al(NO3)3, after 5 minutes, 20 %u00b5L of K-acetate 1M and 860 %u00b5L of ethanol 80% were added and left for 40 minutes. The absorbance was measured at 415 nm. The experiment was repeated 3 times. The total flavonoid content in the sample was determined as mg quercetin equivalents (QE).2.4. Evaluation of antioxidant activityThe antioxidant activity of fermented MFP was assessed based on the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging ability [8]. DPPH 0.1 mM was mixed with 80% ethanol. Ascorbic acid was used as a positive control with a concentration range of 0-0.5 mM in distilled water. The supernatant of fermented MFP