Step 1. Graph the calibration curve relating absorbance at 520 nm to gallic acid concentration. Fit a line function to the measured data points applying regression analysis with r2 > 0.80 (refer Calculations in Experiment 1).
Step 2. Calculate concentration of free gallic acid (Cfree) in the tannin extract as under:
Cfree (mM) = A520 x e where, A520 is absorbance at 520 nm and e is extinction coefficient obtained from linear regression.
Step 3. Calculate concentration of total gallic acid in free and gallotannin forms in 0.2 ml of the sample (Cs):
However, original volume of tannin extract was 3.34 mL. Hence, total concentration (CT) should be:
Step 4. Calculate concentration of gallic acid in gallotannin forms (Cg form), subtracting free gallic acid found in tannin extract:
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