1. What Colors Your World? Biotechnology Activities with Food Dyes

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 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

4. Biotechnology Explorer™ | explorer.bio-rad.com 4 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 Inexpensive separates well Understanding the chemistry of mobile phase- Salt? Polar? Non-polar? Best separation material? Measurements Analysis

5. Biotechnology Explorer™ | explorer.bio-rad.com 5 Chromatography Used to separate biomolecules based in their physical characteristics may include: size charge hydrophobicity interaction with other molecules

6. Biotechnology Explorer™ | explorer.bio-rad.com 6 The System  All forms of chromatography have a –Stationary phase (remains stationary and is a solid or a liquid supported by a solid) –Mobile phase (a liquid or a gas that travels through the stationary phase and carries the parts of the mixture)

7. Biotechnology Explorer™ | explorer.bio-rad.com 7 Types of Chromatography  Paper –Paper serves as the stationary phase. –The paper strip is placed in a solvent (mobile) phase which carries the mixture through the paper. –Parts of the mixture will travel at different rates separating the parts  Column –The stationary phase is packed into a column –The column is equilibrated (saturated with solvent) –Sample is added to the column –Buffers are added to separate the mixture –Samples are collected in intervals called fractions

8. Biotechnology Explorer™ | explorer.bio-rad.com 8 Be an Engineer – Design your own!  Select materials that you will use to separate your Kool Aid into the food dyes that make it the color it appears  You will do this by –Paper chromatography –Column chromatography

9. Biotechnology Explorer™ | explorer.bio-rad.com 9 What are some of the design factors we want to think about? TypeMobile Phase Stationary Phase Column Type PaperWaterPaper type Column1X PBSHIC MediaSyringe AlcoholCottonCommercial Column

10. Biotechnology Explorer™ | explorer.bio-rad.com 10 Dye separation from Kool Aid Using Paper Chromatography 1.Choose your paper type (stationary phase) 2.Cut it into.75 x 4 inch strips 3.Place 50ul of Kool-Aid at one end using a DPTP

11. Biotechnology Explorer™ | explorer.bio-rad.com 11 4.Place 1 ml of your mobile phase into a medicine cup 5.Place the strip Kool-Aid side down into the mobile phase

12. Biotechnology Explorer™ | explorer.bio-rad.com 12 1.Choose your Column Type (HIC, Empty Column, Syringe) 2.If empty column or syringe add stationary phase by stuffing it with a cotton ball Building and Using A Column to Separate Your Kool-Aid

13. Biotechnology Explorer™ | explorer.bio-rad.com 13 3.Choose your mobile phase (water, alcohol, or 1xPBS) 4.Place column in collection tube 5.Add 2 ml of mobile phase to column 6.Let it flow until it no longer drips Building and Using A Column to Separate Your Kool-Aid

14. Biotechnology Explorer™ | explorer.bio-rad.com 14 7.Move column to next collection tube 8.Using a DPTP add 1ml of Kool-Aid to the top of the column 9.Let it flow until it no longer drips (this is fraction 1) 10.Move to the next column 11.Continue steps 5 and 6 until the eluate (fluid coming off the column) is clear Building and Using A Column to Separate Your Kool-Aid

15. Biotechnology Explorer™ | explorer.bio-rad.com 15 Compare Results Which paper, mobile phase worked best for paper chromatography? Which column, mobile phase worked best for column chromatography?