Experiment: Extraction of Vitamins A & C Objective

NMR experiment involving vitamins to study intermolecular forces & solubility
Experiment: Extraction of Vitamins A & C
Objective: The objective of this experiment is separate vitamins A and C through an
extraction and to confirm the separation using NMR.
Retinol (Vitamin A)
Ascorbic acid (Vitamin C)
D2O (deuterated water)
CDCl3 (deuterated chloroform)
Thin-tipped spatulas (2)
Microcentrifuge tubes (2)
Pasteur pipets & bulb
CH 113 (M.Hall) Ch11
Experiment: Extraction of Vitamins A & C
Data & Observations
Part I. Extraction of the Vitamins
1. Label a microcentrifuge tube clearly on the top and side of the tube as “Bottom Layer.” Also
write your initials and the date.
2. Use a thin-tipped spatula to transfer a small amount of ascorbic acid to the microcentrifuge
tube. The solid is very staticky, so it clings to the spatula and tube. Simply dip the spatula
into the bottle, and tap whatever sticks to the spatula into the tube.
1. To the same microcentrifuge tube, add a small amount of retinol using a clean spatula.
Retinol is very sticky and thick (like earwax), so just scoop some out of the container and
scrape it along the inside of the microcentrifuge tube.
2. Use a Pasteur pipet to transfer 0.5-0.7 mL of CDCl3 to the same tube. You can use the
graduations on the tube to guide the volume added. Close the tube firmly and shake it to
dissolve the solids as much as possible.
3. Use a fresh Pasteur pipet to add 0.5-0.7 mL D2O to the same tube. Mix the contents well,
then let the tube sit until the layers separate.
4. Using a Pasteur pipet, decant off the top layer and transfer it to a new microcentrifuge tube
labeled with “Top Layer,” your initials, and the date.
CH 113 (M.Hall) Ch11
Experiment: Extraction of Vitamins A & C
Data & Observations, cont’d
Part II. NMR Spectra of the Separated Layers
1. Using a Pasteur pipet, transfer the top layer to a labeled NMR tube.
2. Using a fresh Pasteur pipet, transfer the bottom layer to another labeled NMR tube.
3. Obtain a 1H NMR spectrum of each sample (using the 300 MHz NMR spectrometer and ICONNMR).
1. Compare your spectra to the reference spectra on the following pages. Which solvent extracted
the ascorbic acid (Vitamin C)? Which solvent extracted the retinol (Vitamin A)?
CH 113 (M.Hall) Ch11
Experiment: Extraction of Vitamins A & C
Summarize all your results including the identity of the top and bottom layers and the vitamin that
was extracted in each layer.
Describe the results of the experiment and account for the presence of two layers in the extraction as
well as the separation of the vitamins into the two solvents. Mention specific intermolecular forces
to explain why each vitamin dissolved better in one solvent than in the other. Relate this information
to the lifetime of vitamins in the body.
CH 113 (M.Hall) Ch11
Experiment: Extraction of Vitamins A & C
full spectrum
Reference 1H NMR Spectrum
Pure Ascorbic Acid (Vit.C) in D2O
CH 113 (M.Hall) Ch11
zoom in
CH 113 (M.Hall) Ch11
Experiment: Extraction of Vitamins A & C
Reference 1H NMR Spectrum
Pure Retinol (Vit.A) in CDCl3
full spectrum
CH 113 (M.Hall) Ch11
zoom ins
CH 113 (M.Hall) Ch11

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