1. STEM Electrophoresis with Food Dyes Bio-Rad Biotechnology Explorer™ IDEA and STEM Kits

2. Biotechnology Explorer™ | explorer.bio-rad.com 2 Instructors - Bio-Rad Curriculum and Training Specialists Sherri Andrews, Ph.D., Eastern US sherri_andrews@bio-rad.com Damon Tighe, Western US damon_tighe@bio-rad.com Leigh Brown, M.A., Central US leigh_brown@bio-rad.com

3. Biotechnology Explorer™ | explorer.bio-rad.com 3 Why Use Bio-Rad?  Guaranteed to work  Easy to prep  Cost Effective per student group  Easy, inexpensive first exposure to electrophoresis  Independent inquiry opportunities for what dyes are present in food items, what is the charge of the dyes, can they be separated based on charge/size?  Engineering inquiry with STEM kit ranging from optimizing metal composition and diameter for electrodes, gel matrix composition, buffer composition, etc.  Extensions including paper chromatography, spectroscopy, and researching food dyes commonly used

4. Biotechnology Explorer™ | explorer.bio-rad.com 4 Separation technology: How do you separate two or more compounds from each other? Materials to be SeparatedProperty Used to Separate Components Explanation Iron filings from other metalsMagnetismIron filings will be attracted to a magnet while other metals will not Salt from sandSolubilitySalt will dissolve in water and can be separated from the sand Caffeine from coffee beansSolubilityA solvent is used that selectively dissolves the caffeine (such as dichloromethane or ethyl acetate) but not most of the remaining coffee oils Ethanol from fermented grainBoiling pointEthanol will boil off at a lower temperature than the other components of the fermented grain Wheat from chaffDensityWheat kernels are more dense and will fall to the ground while chaff can be blow away by the wind Tea from tea leavesSizeFiltration can be used to separate the liquid extract from the large tea leaves

5. Biotechnology Explorer™ | explorer.bio-rad.com 5 Electricity can be used to separate molecules by charge Acetate ions Tris ions - + + - + - - - - + + + + - - - - + + + Apply an electrical charge CH 3 C O O-O- HO CH 2 C CH 2 OH NH 3 + CH 2 OH

6. Biotechnology Explorer™ | explorer.bio-rad.com 6 Sieves can be used to separate materials by their size

7. Biotechnology Explorer™ | explorer.bio-rad.com 7 Electrophoresis separates molecules by CHARGE and SIZE Molecular sieve Electricity Electrophoresis means “to carry with electricity”

8. Biotechnology Explorer™ | explorer.bio-rad.com 8 Food dyes – What have you recently eaten?… Did you eat any cheddar cheese? Was it orange? Starbucks Strawberry Frapuccino?

9. Biotechnology Explorer™ | explorer.bio-rad.com 9 Food dyes – Color tells our brains about foods. What flavor is this Skittle?

10. Biotechnology Explorer™ | explorer.bio-rad.com 10 Food dyes – We have been dying our foods for hundreds of years  In the early 1800s some cheese and cayenne pepper was colored with LEAD tetroxide  Pickles with COPPER sulfate  Green tea with COPPER carbonate

11. Biotechnology Explorer™ | explorer.bio-rad.com 11 Food dyes - Regulation There was NO control or regulations on food dyes until the early 1900s in the US Food and Drug Administration (1902) Due to public outcry over adulterated foods Upton Sinclair – The Jungle (1906) Elixir sulfanilamide Poisoning, 100 dead (1937) Food Drug and Cosmetics Act (1938) Synthetic Dyes – Seven approved for use Natural Dyes (derived from plants or animals) can also be used and are not regulated

12. Biotechnology Explorer™ | explorer.bio-rad.com 12 FD&C Dyes and Natural dyes FD&C Synthetic Dyes FD&C Red 40 or FD&C Red 40 Lake FD&C Yellow 6 or FD&C Yellow 6 Lake FD&C Yellow 5 or FD&C Yellow 5 Lake FD&C Blue 1 or FD&C Blue 1 Lake FD&C Blue 2 or FD&C Blue 2 Lake FD&C Red 3 or FD&C Red 3 Lake FD&C Green 3 or FD&C Green 3 Lake Natural Dyes Beetroot red or betanin (from beets) Curcumin (from tumeric) Caramel coloring (from sugar) Annatto (seeds of achiote trees) Carminic acid, carmine, or cochineal (from ground up beetle abdomens) Lycopene (from tomatoes, watermelons, papayas, and red carrots)

13. Biotechnology Explorer™ | explorer.bio-rad.com 13 Food dyes in the news

14. Biotechnology Explorer™ | explorer.bio-rad.com 14 Food dyes have an intrinsic SIZE and CHARGE and thus can be separated using Electrophoresis Food dyeMolecular weight Charge FD&C Blue 1792.86-2 FD&C Yellow 5534.37-3 FD&C Yellow 6452.37-2 FD&C Red 40496.43-2 Carminic Acid492.380 Beetroot red551.48+1

15. Biotechnology Explorer™ | explorer.bio-rad.com 15 So how do we design an electrophoresis chamber to separate food dyes? Dye Electrophoresis Commercial versus built box comparisons

16. Biotechnology Explorer™ | explorer.bio-rad.com 16 Combining Science, Engineering and Math to Develop a Separation Technology Engineering – Design parameter based; no single “correct answer” Science – Hypothesis driven; try to find specific cause/effect Conducts well, has low reactivity, inexpensive Understanding the chemistry of metals in salt solutions, physics of conductivity and currents Best electrode material Best gel matrix Conducts electricity, moldable, inexpensive, non-toxic, correct pore size Polymer chemistry, melting points, gelling properties Measurements Analysis

17. Biotechnology Explorer™ | explorer.bio-rad.com 17 What are some of the design factors we want to think about?

18. Biotechnology Explorer™ | explorer.bio-rad.com 18 Dye extraction from candies

19. Biotechnology Explorer™ | explorer.bio-rad.com 19 Building and running your electrophoresis system to separate the dyes

20. Biotechnology Explorer™ | explorer.bio-rad.com 20 Building and running your electrophoresis system to separate the dyes

21. Biotechnology Explorer™ | explorer.bio-rad.com 21 Building and running your electrophoresis system to separate the dyes

22. Biotechnology Explorer™ | explorer.bio-rad.com 22 Building and running your electrophoresis system to separate the dyes

23. Biotechnology Explorer™ | explorer.bio-rad.com 23 Building and running your electrophoresis system to separate the dyes

24. Biotechnology Explorer™ | explorer.bio-rad.com 24 Extensions  Paper chromatography  Spectroscopy  Food diary  Electrochemistry study –pH changes at the cathode and anode –Effect of material choice for electrode  Optimization of STEM box system –Electrode material choice and thickness –Different materials for gel thickness –Impact of TAE Concentration and Volume –Gel percentages Copper electrode Gelatin matrix