Tasha Magloire
Instrumental Chemistry
Real World Project: HPLC and Raman Analysis of Ascorbic Acid in Fruit Juices
Introduction: Ascorbic acid (also known as Vitamin C) is a water soluble vitamin that is needed for the growth and repair of tissues in the body. Vitamin C protects against scurvy, heart disease, cancer, the common cold, and high blood pressure. Vitamin C can also be used as an antioxidant and dietary supplement. It’s naturally present in some fruits and vegetables and In some commercial juices.
Purpose: To evaluate and analyze the Vitamin C content in four commercial fruit juices using Raman scattering and High Performance Liquid Chromatography (HPLC), and construct a calibration plot relating peak area for the ascorbic
…show more content…
We retested two spiked samples of apple juice and orange juice on the HPLC, adding only .05 g of Ascorbic acid to obtain clean peaks that fit under the calibration curve
HPLC Method
1) Make sure there is enough degassed methanol and water.
2) Turn on computer and check pump lines A and B for air bubbles
3) Turn on detector (SPD-10 AV)
4) On desktop, select HPLC, select Shimadzu icon, select the Methods icon and choose TSW.met
5) Set parameters by clicking the yellow box icon, click Parameters and set as following:
a. Flow rate A: .500 mL/min
b. Flow rate B: .500 mL/min
c. Flow rate C: 1.000 mL/min
d. Max pressure:6000 psi
e. Min pressure: 1
…show more content…
Flow rate A: .010 mL/min
b. flow rate B: .010 mL/min
c. Flow rate C: 1.000 mL/min
11) Click download and turn detector off, leave computer and pumps on
FT-Raman Method
1) Fill the blue dewar with liquid nitrogen and wait 20 minutes then top off
2) make sure the FTIR is on
3) Turn on the laser power supply switch and then turn the key to the on position
4) Make sure the lever between the Raman and FTIR is up, the lever between the Raman and microscope is down, and the internal lever in the Raman should be up, meaning bypass closed
5) Double click on the Varian Resolutions Pro icon to open software
6) From the current scan menu select Raman scan
7) Optics parameters should be:
a. IR source: off
b. Beam: right
c. Detector: Raman Ge
d. Beamsplitter: quartz near IR
e. Accessory: FT-Raman
f. ATR Crystal: none
g. Optical filter: holographic notch
h. Aperture: open
8) Select the laser tab and click turn on diode
9) Press the shutter switch on the front of the Raman (to open) and turn the power to its highest,