Inspire your students – tomorrow’s researchers
STAM Winnipeg October 25, 2013
STEM Electrophoresis with Food Dyes
Bio-Rad (Canada) Life Science Education
– Biotechnology Explorer Consultant
Robert Malyk BSc., MEd.
Senior Biology Teacher (retired)
Ridley College
St. Catharines, ON Canada
biobob4@hotmail.com
Combining
Science,
Engineering
and Math to
Develop a
Separation
Technology
Why Use
Bio-Rad?
• Real research!!
• 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
Separation
technology:
How do you
separate two
or more
compounds
from each
other?
Electricity can be
used to separate
molecules by
charge
Sieves can be
used to
separate
molecules by
size
Electrophoresis
separates
molecules by
CHARGE and SIZE
Electrophoresis means
“to carry with electricity”
Food dyes –
Think about
what you ate for
breakfast or
lunch…
Did you eat any cheddar cheese?
Was it orange?
A Starbucks Strawberry
Frapuccino?
Food dyes –
What flavor do
you think this
Skittle would
have?
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
Food dyes –
Canada Food
Inspection
Agency
There was NO
control or
regulations on
food dyes until
the early 1900s
Food dyes –
Canada Food
Inspection
Agency
Permitted
Natural Colours
in Canada and
Corresponding
European
Names
FD&C Dyes and
Natural dyes
FD&C
FD&C
FD&C
FD&C
FD&C
FD&C
FD&C
FD&C Dyes
Red 40 or FD&C Red 40 Lake
Yellow 6 or FD&C Yellow 6 Lake
Yellow 5 or FD&C Yellow 5 Lake
Blue 1 or FD&C Blue 1 Lake
Blue 2 or FD&C Blue 2 Lake
Red 3 or FD&C Red 3 Lake
Green 3 or FD&C Green 3 Lake
Natural Dyes
Carminic acid, carmine, or cochineal (from
ground up beetle abdomens)
Beetroot red or betanin (from beets)
Curcumin (from tumeric)
Caramel coloring (from sugar)
Annatto (seeds of achiote trees)
Lycopene (from tomatoes, watermelons,
papayas, and red carrots)
Food dyes in the
news
Food dyes have
an intrinsic SIZE
and CHARGE
and therefore
can be
separated using
Electrophoresis
So how do we
design an
electrophoresis
chamber to
separate food
dyes?
Dye Electrophoresis
Commercial versus built box comparisons
What are some of
the design factors
we want to think
about?
Dye extraction
from candies
Building and
running your
electrophoresis
system to
separate the
dyes
Building and
running your
electrophoresis
system to
separate the
dyes
Building and
running your
electrophoresis
system to
separate the
dyes
Building and
running your
electrophoresis
system to
separate the
dyes
Building and
running your
electrophoresis
system to
separate the
dyes
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
Other Activities
Chromatography
• Used to separate biomolecules based in their
physical characteristics may include:
•
Size
•
Charge
•
Hydrophobicity
•
interaction with other molecules
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)
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
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
What are some
of the design
factors we want
to think about?
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
Dye separation
from Kool Aid
Using Paper
Chromatography
4.
Place 1 ml of
your mobile
phase into a
medicine cup
5.
Place the strip
Kool-Aid side
down into the
mobile phase
Building and
Using A Column
to Separate Your
Kool-Aid
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
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
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
Compare
Results
• Which paper, mobile phase worked best for
paper chromatography?
• Which column, mobile phase worked best
for column chromatography?