Gi Prelab Final

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University of British Columbia *

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211

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Chemistry

Date

Dec 6, 2023

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pdf

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3

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Prelab 6 (UV-AO) Stephanie Ko J06 Nov 20, 2023 Jasmine Khan J05 1. Procedure for making FRAP and to produce a calibration curve a. Prepare FRAP reagent: i. Quantitatively weigh 0.0270 g of 20 mM FeCl ·6H O then QT into another 100 ml beaker, dissolve FeCl ·6H2O with 5 ml of DIH O. ii. Quantitatively weigh 0.0312 g of 10 mM TPTZ then QT into a 100 ml beaker, and dissolve TPTZ with 10 ml 0.040M HCl. iii. Use a volumetric pipette to QT 5 ml of TPTZ, 5 ml of FeCl ·6H O and 50 ml of buffer and combine into a 250 ml Erlenmeyer flask. b. Calibration Curve i. Quantitatively weigh 0.0348 g of FeSO ·7H O then QT into a 50 ml volumetric flask. Dilute up to the calibration line with DI water. Invert to mix well. c. Prepare a set of 6 Fe² standard solutions: i. Use 5 ml volumetric pipette to QT 5ml FRAP reagent into 6x 50 ml volumetric flask. ii. Use micropipette to pipette volumes of Fe² based on table below into each volumetric flask based on the table below. iii. Use Beer’s Law to calculate required [Fe² ] for absorbance wanted Flask number Vol of Fe² ( μL ) Vol of Buffer (mL_ Vol of FRAP (ml) 1 0 25 5 2 100 25 5 3 225 25 5 4 350 25 5 5 475 25 5 6 600 25 5 iv. Dilute flask to line with Buffer, invert to mix, AFTER WAITING 5-10 min v. Rinse cuvettes (length of 1 cm) with standard solutions and wipe with Kimwipe before running each of the 6 solutions with wavelength of 597 nm. vi. Starting with a blank which is distilled water. vii. After measuring absorbance against concentration of Fe² in each to determine the calibration curve 2. Procedure to prepare antioxidants (will be made for us) a. Clove extract i. Quantitatively weigh 1g of cloves then QT into 100 ml beaker ii. Add 20 ml DIH O and let it soak in it for 30 minutes iii. To decant into vial, funnel with glass wool to isolate out cloves b. Green tea i. Boil 180 ml of water in 250ml beaker on a hot plate (set at 6 to 7) ii. Turn off heat then take one tea bag and steep in boiled water for 2-3 min iii. Let it cool to room temperature
Prelab 6 (UV-AO) Stephanie Ko J06 Nov 20, 2023 Jasmine Khan J05 c. Cranberry juice i. Mix 6 ml of DIH O and micropipette 2 ml of concentrated Cranberry juice in a 100 ml beaker. Swirl and mix well with glass stir rod 3. Procedure to quantify percentage of Fe³ reduced to Fe² a. Antioxidant stock solutions and blank: Pipette volumes of FRAP solution and micropipette volumes of the antioxidant solutions following the table below into four 50 ml volumetric flasks. Antioxidant Volume of Antioxidant (µl) Volume of FRAP (ml) Clove Extract 100 5 Green tea 100 5 Cranberry juice 100 5 Blank 0 5 b. Dilute to mark with buffer solution and invert to mix. Repeat for the other two antioxidants c. Allow the reaction of FRAP and all the antioxidants to go to equilibrium letting the three solutions sit til the end of the lab. Measure absorbance of blank and all three solutions with a non scanning spectrophotometer at wavelength of 597 nm to find concentration of FRAP at equilibrium d. Place in a UV spectrophotometer for 1 min to observe changes in absorbance e. Plot absorbance vs concentration of Fe² in each to determine Beer’s Law to determine antioxidant strength. The concentration of Fe² produced is how much Fe³ was consumed. 4. Kinetics of FRAP-Ascorbic acid a. Procedure for determining rate constant k and the order of ascorbic acid a i. Ascorbic stock solution: Weigh out 0.0550 g of ascorbic acid and transfer into a 25 ml volumetric flask. Dilute to line with distilled water. ii. Prepare ascorbic acid standard solutions in 25 ml volumetric flasks following the table below. Dilute to line with DIH O. Solution # Volume of Stock solution (µl) Volume of volumetric flask (ml) 1 1000 25 2 2330 25 3 3670 25 4 5000 25
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