Determination of Chlorophyll in Olive Oil Using the Vernier Spectrometer

David A. Reierson
Roosevelt High School, Minneapolis, MN
ISD Special District #1
Retrieved from Vernier Spectrometer Lab #5
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Summary

In this activity, students will investigate how much chlorophyll is in olive oil using a Varnier Spectrometer. Students will measure and analyze the visible light absorbance spectra of three standard olive oils obtained from any supermarket: extra virgin, regular, and light. Students will then measure and determine the absorbance spectrum of an "unknown" olive oil sample. Last, students will determine and identify the "unknown" olive oil sample as one of the three standard types, extra virgin, regular, or light.

Learning Goals

From this activity, students will understand the absorbance spectrum, in the visible light range, of chlorophyll creates three yellow absorbance peaks at 413, 454, and 482 nanometers, and two blue absorbance peaks at 631 and 669 nanometers. Students will examine combinations of these wavelengths which are green to the human eye. They will determine that different sources of chlorophyll in olive oil have different ratios of these absorbance peaks, which create various shades of green. Students will realize that in the world of chemistry, specifically, chlorophyll, that all greens are not the same. Lastly, students will learn how to use the Vernier Spectrometer (V-SPEC) to measure the absorbance of the olive oil samples over the visible-near infrared (NIR) light spectrum between 380 and 950 nanometers. Vocabulary words that students will learn through this activity are , spectrometer, absorbance, absorbance spectrum, light spectrum, chlorophyll, cuvette, Beral pipet, and nanometer.

Context for Use

This activity could include students in grades 9-12. We are an urban public school. There are 20-35 students per class on any given day and class session. Students are divided up into groups of four. Special equipment includes the Vernier Spectrometer, computers, cuvettes, Beral pipets, three different types of olive oil, distilled water, and isopropyl alcohol. I would suggest using olive oil or other green leaved plants to extract olive oil samples. This lab requires an extensive pre-lab component and demonstration by the teacher. You should expect a few days to get students acclimated to the Varnier spectrometer, lecture, and completing the lab and analysis. It would be encouraged to complete a demo and show students what the expected results graph will look like and discuss absorbance and concentration before the lab begins. Students should be able to differentiate the different types of olive oil absorbance Vs wavelength on the final graphs. The final activity is for each group to identify an "unknown" determined by the teacher. The "unknown" should be either one of the three types of olive oils they have already analyzed. An extension would be to soak another type of green leaved plant in the isopropyl alcohol to analyze the chlorophyll absorbance and wavelength.

Description and Teaching Materials

The lesson will be introduced during the properties unit in chemistry and physical science. Before beginning this lab, I would highly suggest that students become familiar with both Lab Quest and the spectrometer hardware and software. This would require a couple of days of exploratory and simple canned lessons.
To get started, show students during a demonstration how to complete the following tasks: setting up the spectrometer, using Logger Pro 3.4.6 program on the computer or Lab Quest, calibrating the spectrometer, conducting the spectrum analysis of the olive oil samples, saving the experimental files on the computer, and saving and printing the absorbance Vs wavelength graphs.
The procedure used to conduct the lab is as follows: (Vernier Software and Technology, Spectroscopy with Vanier, 2006, P. 6-2)
SETUP:
1. Obtain and wear goggles.
2. Use a USB cable to connect a Vernier Spectrometer to a computer.
3. Start the Logger Pro 3.4.6 program on your computer.
4. Obtain small volumes of the three standard olive oils to be tested.
5. To set up the spectrometer, open the Experiment menu and select Connect Interface → Spectrometer → Scan for Spectrometers.

6. CALIBRATE THE SPECTROMETER:

a. Prepare a blank by filling an empty cuvette ¾ full of distilled water.
b. Open the Experiment menu and select Calibrate → (Spectrometer). The following
message appears in the Calibrate dialog box: "Waiting.. . seconds for the device to warm up." After 60 seconds, the message changes to: "Warm up complete."
c. Place the blank in the cuvette holder of the spectrometer. Align the cuvette so that the
clear sides are facing the light source of the spectrometer. Click "Finish Calibration", and then click "OK."

7. Conduct a full spectrum analysis of an olive oil sample:

a. Empty the blank cuvette and rinse it twice with small amounts of extra virgin olive oil.
Fill the cuvette ¾ full with the olive oil and place it in the spectrometer.
b. Click "collect." A full spectrum graph of the olive oil will be displayed. Review the graph to identify the peak absorbance values. Click "stop" to complete the analysis.
c. To save your data, select "Store Latest Run" from the Experiment menu.
8. Repeat Step 7 with the remaining olive oil standard samples.
9. Obtain an unknown sample of olive oil. Repeat Step 7 with the unknown.
10. Save and print a copy of each graph for your lab report.
11. Select "Exit" from the File menu to close down Logger Pro 3.4.6
12. At the end of the experiment, clean up by rinsing and cleaning the cuvettes and other oil-bearing containers with isopropyl alcohol.

Teaching Notes and Tips

When locating olive oil in the supermarket, if the olive oil grade is not labeled, grab any three samples that appear, to your eyesight, different greenish tints. Chlorophyll concentration determines the greenish tint. Light olive oil should be the sample with the lightest green tint.
The cuvette should be about 3/4 full to get good absorbance measurements and will give plenty of room to seal the cuvette with a plastic cap. Try not spilling the sample in the spectrometer, it is difficult to get it cleaned up, and time consuming.
Because this lab is qualitative in nature, students can use different cuvettes and get good results.
A good extension is to have your speedy students complete a similar lab by extracting chlorophyll from fresh green leaves and comparing the absorbance to their olive oil samples. A simple method is to tear and or grind a few leaves and soak them in ethanol or isopropanol for at least thirty minutes. Another interesting fact, according to a Vernier Software & Technology resource, is that the isopropanol and ethanol used for cleaning might show up on the absorbance Vs wavelength graphs around the 900 nm wavelength.
As a good way to become familiar with this experiment, plan to keep a set of sample data, graphs, as well as an answer key to the analysis section questions. Additionally, set up the Lab Quest and Logger Pro 3.4.6 and become familiar with the setup and experiment. The results vary a lot, because of many different factors. Have fun with this lab and remember, it is more about the process than the results, plus it is qualitative, which means no math calculations. A simple explanation about concentration and absorbance will go along ways during the lecture if you take the time to explain the relationship to your students.
Having enough materials is another problem. Make sure that you know the quantity of materials you will need ahead of time. This might be a great time to invite parent volunteers to work in your class. They can oversee material distribution and sit in with small groups.

Assessment

Data Analysis
1. Describe the graph of each of the olive oil samples. Discuss the differences between each grade of olive oil.

2. Identify the unknown olive oil as extra virgin, regular, or light. Explain your results.

3. The introduction of this experiment identified six visible wavelengths of peak absorbance for chlorophyll. Describe each of the olive oil samples in your lab including the unknown, in the terms of these expected peaks. Were your peaks similar to the introduction peak values given in the introduction?

Standards

9-12. II.A.1-Structure of Matter
9-12. I.B.1-Scientific Inquiry
9-12. I.B.1. analyzing data

References and Resources