2014 Cell 9 Presentation

Rebecca!
Kirby!
Ashley!
Ian! David!
Boshra!
Thomas!
Lauren!
Jeannelle!
Howdy!
My name is
Archibald and we
have an analysis that
we need help with. I
heard that this
Chem 315 class is
really smart. Can
you help us?
I heard that some
VeggieTales might
have some special
powers called
FLAVONOIDS Can
you guys teach us
about them and tell
us which of us has a
lot of them?
OkSo What are Flavonoids?
Over 6000 different compounds
Secondary plant metabolites
Latin word flavus means yellow
Albert Szent-Gyorgyi in 1938
First named vitamin P
Classes of flavonoids:
Quercetin
Flavones
etc.

Why Quantify
Flavonoids?
Reduce risk of cancer
Reduce cardiovascular diseases
Posses anti-oxidants
Free radical scavenging activity
Horticultural approaches toward
health promotion
Optimizing growth and storage
conditions
Cooking advices to increase
flavonoid intake
(SHHHHH. Dont tell them)
YOU HAVE TO EAT
THE VEGGIETALES
TO GET THE
FLAVONOIDS.
But there is ONE LITTLE Problem:
Objective of Analysis
So why
should I
volunteer for
this study?
Sounds Awesome!
I VOLUNTEER AS TRIBUTE!
Liquid
Nitrogen is
COLD!!!!
Strategy
Extraction from plant tissues
Plant material is freeze-dried
Flavonoids are extracted with
acid/enzyme hydrolysis
Reverse-phase high
performance liquid
chromatography
Used because there is higher
resolving power than
traditional chromatography
Due to having smaller
particles

Fundamental Principles
Retention time, Partition coefficient,
Gradient Elution, Isocratic Elution,
Chromatogram
More Fundamentals

Hydrophobic Interactions:
Hydrophobic regions
(Need to know structure of
molecule)
Advantages:
Takes advantage of
hydrophobicity characteristics
of protein or molecule
Large volume of sample can
be loaded
High Ionic strength sample
can be loaded
sample eluted with low salt

Fundamentals of Elution
Gradient Elution:
Salt concentration in mobile phase changes
Isocratic Elution:
Salt concentration in mobile phase remains
constant

Fundamentals of the
Chromatogram

Reverse Phase High Performance
Liquid Chromatography
non-polar stationary phase and an aqueous, polar
mobile phase
Retention time is longer for non-polar molecules
Flavonoids have varying levels of polarity, so this
allows separation and identification

https://www.youtube.com/watch?v=mPO7Tv2mIJU
What is the Measurable
Quantity?
Retention Time
To identify specific
flavonoid glycoside
Compare to reference
standards
Peak Area
Determine the amount
of quercetin
Detective
Larry is going
to help!
Lets Measure It!
Standards are used
Mass spectroscopy is used
to confirm identification

Calculate Results/ Interpret Data
Finding the
Area
Find the Percent
Composition
Matching the
Peaks
Take Mass
Spectrometry to
verify the
fractions identity.
So what did we learn about
Bob?
He contains 5 different
flavonoids! (4-26 mg/100 g)
Chalconaringenin (35-
71%)
Quercetin 3-rutinoside
Another form of
quercetin
kaempferol 3-rutinoside
A form of phloretin


Accuracy and Precision
Variation in results could be due to:
Light regimes under which the plants were
grown
UV-B irradiation can induce accumulation of
flavonoids
Varietal differences in crops
Conditions it was kept under during
transport from grower to supermarket
Data presented is in line with previous
studies that utilized similar tissues
Accurate measurement of flavonoids in
foods is relatively cheap and not especially
time consuming and thereby offers method
for product quality assurance
Bob does
have lots of
Flavonoid
Special
Powers!!!

Thank you Chem 315 for
teaching us the concepts of
chromatography and the
special powers of Flavonoids!
The Flavonoids of Tomatoes, Rune
Slimestad, Torgils Fossen, and Michel J.
Verheul, Journal of Agricultural and Food
Chemistry 2008 56 (7), 2436-2441
Quantitative Analysis of the Flavonoid
Content of Commercial Tomatoes,
Onions, Lettuce, and Celery, Alan Crozier,
Michael E. J. Lean, Morag S. McDonald,
and Christine Black, J. Agric. Food Chem,
1997, 45, 590-595
Sources