Anthocyanins as Acid/Base Indicators: Applications and Activities for High School Students

1. Anthocyanins as Acid/Base Indicators: Applications and Activities for High School Students Brett Hale Heath Dawley Southeast Missouri State University BioQUEST 2008 Fruits_and_vegetables2. (Picture). URL: http://www.firodiyaheartfoundation.org/images/fruits_and_vegetables2.jpg

2. *Wanted to use anthocyanins as acid/base indicators *Were familiar with the famous red cabbage acid/base indicator. *Decided to try out other anthocyanins as acid/base indicators, having students explore the uses of chemicals derived from plants. *Designed 4 activities to help students explore this area Using plants in high school chemistry

3. *For later activities reference solutions at the unit pHs from 1-14 are needed. *Though the teacher can pre-make these, it is also a good activity to have students do. *They will take 1.00 M HCl and 1.00M NaOH, and add it to distilled water, while exploring the math relationships of dilution. *Goal here is to see that pH is a logarithmic pattern. Taking 5 mL of each acid or base and diluting it to 50 mL and repeating the pattern gives you what you need. *Students are required to show all their work, or figure out the pattern and explain it. Pre-activity: Chemistry and Math pH=-log [H3O+]

4. *Students will be given leaves from plants with a high anthocyanin concentration. *Students will perform paper chromatography to separate out the plant pigments in the leaves. *Students will roll a coin over the leaf to place the pigments on the filter paper. *Suggested solvent is a petroleum ether/acetone mixture. *Students will calculate Rf values for the pigments and compare to some standard Rf values. *Using this information the students will identify the plant pigments. Activity One- Plant Pigment Chromatography Chorophyll a and b, Xanthophylls, Carotenes, Anthocyanins

5. Using the chromatographs produced in part one, students will test which pigments can be used as acid/base indicators • The pH range samples generated in the pre-activity are used • The students dip the paper from each pigment type into the standard solutions • They will discover that anthocyanins change color in very acidic and basic environments Activity one cont.- Which act as acid/Base indicators? Which of the pigments will change color?

6. *Many high school students have used pH paper, but now they will construct their own scale based on anthocyanins from plants. *Each student group will be given an anthocyanin rich plant. Examples include red cabbage, grapes, blueberries, tomatoes, strawberries, plums, or raspberries. The plants should be different for each group. *Each will then place some of their plant material in a beaker and boil it to extract the anthocyanins. Decanting the liquid will give them their own acid/base indicators (Withers, 2001). *The students will then take a little of each of the pH reference solutions and put them into 14 test tubes, adding a small amount of their natural indicator. Activity Two: Construction of a ph scale A single group of plant pigments provides a lot of different colors.

7. *Students need a reason to use their new color scale, so we will use it to find a pH range of an unknown acid or base. *Placing a small amount of their natural indicator into a sample of the unknown, students will compare their color to their color scale to get an estimated pH range. *Students will then collaborate all of their estimates to get the most precise estimate possible. *Students must record all their observations and reasoning. Activity two cont. finding the ph of an unknown Plants + Acid or Base -> Color Change

8. *Now students will test the accuracy of their pH measurement using a handheld pH meter. *Students will also graph real data as they titrate their unknown. The main data they will be collecting will be pH and volume of titrant added. *Using Excel, students will construct a graph of pH vs. mL of titrant, and analyze the graph to determine the unknown’s concentration. If their anthocyanin indcator changes color they will indicate that, too. Activity Three: Quantitative Analysis Time to use some math!

9. *Students will then be required to come up with a hypothesis for the existence of yellow tomatoes based on the data they have learned. Activity Three Continued At halfway to the equivalence point pKa=pH

10. *Students could look at spectroscopy of the anthocyanins to measure absorbance at different wavelengths of light, and use a database to help identify specific anthocyanins (See Kalb and Howard). *While fresh fruit was used in our trials, it would be a great variation to try frozen and canned, to see if these processes had any effect on the anthocyanins. Extensions There’s lots more to learn!

11. Questions?? ???

12. Blueberry_Cluster. (Picture). URL:http://www.oregonberry.com/Blueberries.htm Accessed: June 19, 2008. Fruits_and_vegetables2. (Picture). URL: http://www.firodiyaheartfoundation.org/images/fruits_and_vegetables2.jpg Kalb, Dee A., and G. W. Howard. "Analysis of Plant Pigments Using Paper Chromatography and Visible and/or UV Spectroscopy." Purdue University Instrument Van Project. Purdue University. <http://www.chem.purdue.edu/teacher/table_of_contents/UVVUS/UVVIS.Plant%20Pig ments_CH.pdf>. Accessed: June 19, 2008. LYC11. (Yellow tomato picture). URL: http://www.magicgardenseeds.com/images/product/LYC11/LYC11.jpg. Accessed June 19, 2008. References Learning requires you to get help from others.

13. Marsden, Steve. "Separation of Plant Pigments by Paper Chromatography." Chemtopics.Com. 19 Aug. 2007. <http://www.chemtopics.com/unit06/pchrom.pdf>. Accessed: June 19, 2008. Massengale. Chromatography of plant pigments. URL: http://www.biologyjunction.com/chromatography_plant_pigments.htm NASA. Chromatography of leaf pigments: Lesson 2 of 2. URL: http://media.nasaexplores.com/lessons/01-037/9-12_2. Accessed: June 18, 2008. Plant Pigment Chromotography. Milanareaschools.Org. <http://milanareaschools.org/~wbenya/Biology/documents/chromatography%20lab.pdf >. Accessed June 19, 2008. Red Acre Cabbage. (Picture).URL: www.seedfest.co.uk/seeds/cabbage/cabbage.html Accessed: June 19, 2008. References So does teaching!

14. Red Cabbage Acid Base Indicator. 27 Feb. 2002. University of Minnesota. <http://www.chem.umn.edu/services/lecturedemo/info/Cabbage_Indicator .html>. Accessed: June 19, 2008. Red Cabbage pH paper. URL: http://www.ipse.psu.edu/activities/paper/red_cabbage.pdf Accessed: June 19, 2008. Reiss, Carol. Experiments in Plant Physiology. Englewood Cliffs, NJ: Prentice Hall, 1994. Senese, Fred. Water to Wine. General Chemistry Online. 31 July 2007. Frostburg. <http://antoine.frostburg.edu/chem/senese /101/features/water2wine .shtml>. Accessed: June 19, 2008. References In fact, teaching is learning!

15. Strawberry. (Picture). URL: http://www.arabidopsisthaliana.com/strawberry/strawberry.jpg Accessed: June 19, 2008. Tomato_pd. (Picture). URL: http://collectingtokens.files.wordpress.com/2007/08/tomato_pd.jpg. Accessed: June 19, 2008 Withers, G. (2001). Natural indicators: How do they work?GIPSE. URL: http://www.cmu.edu/gipse/materials/pdf-2001/natural_ph_indicators.pdf. Accessed: June 19, 2008. References Thank you!