BIOACTIVITY OF FACTOR-2 (6,7,4'-trihydroxiisoflavone) FROM TEMPE FOR CELLS OXIDATIVE STRESS IN BACTERIA

• Main Author: PEBRINA (NIM: 20512025), RELITA
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/23878
• Institution: Institut Teknologi Bandung

Tempe is a traditional Indonesian food containing isoflavone compounds which available in the form of isoflavone glucosides that are, genistin, daidzin and glisitin, as well as in the form of isoflavone aglycons, genistein (5,7,4'-trihydroxiisoflavone), daidzein (7,4'-dihidroksiisoflavone) and glisitein (6-methoxy-7,4'-dihydroxiisoflavone), and factor-2 (6,7,4'-trihydroxiisoflavone). Factor-2 in tempe thought to have stronger activity than the other isoflavones. Several studies have been conducted to determine the activity of some of the isoflavones are genistein, glisitein, and daidzein. It gives its own interest for researchers to conduct research on the benefits of factor-2 in tempe. Therefore, aims of this research to study the role of factor-2 in tempe for cells oxidative stress in bacteria. <br /> <br /> <br /> Isolation of factor-2 was done by macerating, the samples (tempe and soybean) in methanol (p.a), followed by sequencial liquid extraction using hexane and ethyl acetate respectively. The separation and identification of factor-2 was performed using TLC by comparing the soybean extract and tempe extract. The result of identification with Thin Layer Chromatography (TLC) showed 5 stains of isoflavone extract, hereinafter referred to fractions 1, 2, 3, 4, dan 5. Fraction 2 of tempe extract was absent compared to those of soybean extract, it was suspected as a factor-2 whose Rf value was 0.62. Further identification of fraction 2 with UV-Vis spectrophotometer showed the optimum wavelength of 261 nm was observed. The functional group identification of fractions 2 using IR spectrophotometer showed three important functional groups in fraction 2 which are OH group, C = O, and C-O-C group. The groups are characteristic for such group for isoflavone. <br /> <br /> <br /> Cell's oxidative stress test was conducted using crude extract from isoflavone, daidzein, and fractions 2 suspected as a factor-2 against gram-positive bacteria (Bacillus cereus and Staphylococcus aureus) and gram-negative bacteria (Pseudomonas aeruginosa, and Escherechia coli) using the disc diffusion test. 0.3% H2O2 was chosen as positive controle to induce cell’ oxidative stress, GSH 50 mM was chosen as a positive controle to suppress the cell's oxidative stress by its activity as antioxidant. Cell was treated with 20 &#956;L of 0.3% H2O2 compared with those of mixture of 20 &#956;L of 0.3% H2O2 and each sample extracts (daidzein, farction 2, and GSH). Diameter of clear zone observed around the paper disc were examined after incubation for 16-24 hours at 37oC. <br /> <br /> <br /> The percentage reduction in the diameter of clear zone of the crude extract against B.cereus, S. aureus, E. coli, and P.aeruginosa are 43.50%, 40.50%, 31.30%, and 44.16% respectively. While those of daidzein are 13.08%, 11.50%, 11.82% and 10.83% respectively. The test by fraction 2 showed the results of 20.51%, 16.67%, 11.82%, and 20.63% respectively. <br /> <br /> <br /> Quantitative analysis of cell's oxidative stress was conducted using DPPH methode. As a result showed that IC50 of GSH, daidzein, and fraction 2 were 0.85 ppm; 22.45 ppm; and 21.84 ppm respectively.