Transcript Slide 1

Making Sense of Science
Megan Tichy, Ph.D.
Lecturer, Texas A&M University
Highlights

Toxic grains
 Scientific and non-scientific names

Quantification
 Parts per million

Gluten
 Chemical structure

Processing
 Distillation
Scientific names
More specific
• Classification based on physical, behavioral or genetic diversity
• Subject to revisions
Human
Pea
Wheat
Domain
Eukarya
Eukarya
Eukarya
Kingdom
Animalia
Plantae
Plantae
Phylum
Chordata
Magnoliophyta
Magnoliophyta
Class
Mammalia
Magnoliopsida
Lilliopsida
Order
Primates
Fabales
Cyperales
Family
Hominidae
Fabaceae
Poaceae
Genus
Homo
Pisum
Triticum
Species
H. sapiens
P. sativum
T. aestivum
Mnemonic device: Kids Prefer Cheese Over Fried Green Spinach
The grass family
KINGDOM – Plantae – plants
PHYLUM – Magnoliophyta – flowering plants
CLASS – Liliopsida – monocotyledons
ORDER – Cyperales – grasses/sedges
TRIBE
SUBFAMILY
Festucoideae
FAMILY
Oryzeae - rice
Aveneae - oats
Triticeae - wheat, rye, barley
Poaceae
Paniceae - millet
Panicoideae
Andropogoneae - sorghum
Tripsaceae - corn
Donald D. Kasarda, Ph.D., Research Chemist (Retired), USDA
http://wheat.pw.usda.gov/ggpages/index.shtml
Toxic grains
Modern wheat varieties
Wheat
Triticum
aestivum
Durum
Barley
Hordeum
vulgare
Rye
Secale
cereale
Hard Red
Winter
Soft Red
Winter
Hard Red
Spring
Hard White
Wheat
Soft White
Wheat
Images: Home-Grown Cereals Authority (HGCA); http://www.hgca.com
Wheat products
Atta
Bulgur
Couscous
Farina
Seitan
Semolina
whole wheat, stone-grinded
whole wheat, precooked, dried, cracked, and sifted
moistened semolina, rolled and shaped,
coated with finely ground wheat four
endosperm milled to fine granular consistency, sifted
washed dough; water removes starch
inner endosperm of wheat not ground into flour
Atta (India)
Images: Wikipedia commons
Farina
Seitan (Taiwan)
Ancient forms of wheat
Genetics:
Diploid = 14 chromosomes
Tetraploid = 28 chromosomes
Hexaploid = 42 chromosomes
More chromosomes = easier
to breed varieties suitable for
non-ideal soils/climates
Einkorn
Emmer (farro)
Spelt (dinkel)
Triticum monococcum
Triticum dicoccum
Triticum spelta
Diploid
Tetraploid
Hexaploid
Kamut
Triticum turanicum
Tetraploid
Wheat proteins
Protein
Soluble in
Albumin
WATER - neutral pH
Cause of most
wheat allergies
Globulin
WATER - salt solutions
Prolamin
ETHANOL - 70-90% alcohol
Glutelin
NONE OF THE ABOVE
dilute acids/bases, detergents,
or reducing agents
Gluten
Gluten proteins
Prolamine = proline + glutamine
GLUTEN
GLIADINS



GLUTENINS

HMW
LMW
All coded
by different
genes (DNA)
“It is highly probable that glutenin proteins are toxic. Attempts to breed
wheat with satisfactory baking properties tolerated by coeliac patients will
be very difficult.” Eur J Gastroenterol Hepatol. 2006 May;18(5):483-91.
Grain proteins
Grain
Prolamin fraction
% total protein
Wheat
Gliadin
69
Rye
Secalin
30-50
Barley
Hordein
46-52
Oats
Avenin
16
Millet
Panicin
40
Corn
Zien
55
Rice
Orzenin
5
Sorghum
Kafirin
52
toxic
safe
still questioned
contaminated? pure?
Anatomy of a kernel
Manufacturers remove the germ and bran to make refined bread flour
Endosperm
Storage tissue
starch, gluten
Germ
Embryo
vitamin E, magnesium,
riboflavin, thiamin,
phosphorus, niacin, iron and
zinc. Some fat and protein.
Bran
Rich source of magnesium
and iron. ¼ cup contains 6
grams fiber.
Whole-grain: endosperm,
bran, and embryo (all three
parts of the kernel!) left intact
Wheat flour: gluten and starch
flour
H2O
dough
starch (sugars)
75%
dough
yeast
gluten (protein)
12% (varies)
C6H12O6 + 6 O2 → 6 CO2 + 6 H2O + energy
bread
glucose + oxygen → carbon dioxide + water
Image: http://www.exploratorium.edu/cooking/bread/activity-gluten.html
How much gluten?
• Flour contains 10-15% protein
• The protein is 80% gluten
Q: If 1 slice of bread uses 40 g flour
how much gluten is in it?
A: solve for protein 40 g X 0.13 = 5.2 g protein
solve for gluten 5.2 g X 0.8 = 4 g gluten
Q: How many mg gluten is in 1/8 teaspoon flour?
A: Conversion for wheat flour 1 tsp = 2.5 g
1/8 tsp all purpose flour = 0.31 g flour
0.31 g X 0.13 X 0.8 = 0.033 g gluten
1/8 tsp  33 mg gluten
Measuring spoon image: http://www.tubularspices.com/accessories/spoons.asp
How much is too much?
“Ingestion of contaminating gluten should be kept lower than 50
mg/day in the treatment of CD” – C. Catassi et al. AmJ Clin Nutr
2007;85:160–6. Some people are far more sensitive*
Normal gluten intake by healthy individuals = 13 g gluten per day
Compare – 50 mg (0.05 g) vs. 13 g
Normal intake is 260 times as much as the suggested threshold
Codex Alimentarius (WHO/FAO), has two limits for “gluten-free”:
0.02% gluten content - rendered gluten-free
0.002% gluten content - naturally gluten-free food
A person could ingest 3-6 mg gluten per day by consuming
6-8 ounces of “naturally gluten free” grain food
0.002% = 20 parts per million or ppm
*Reference: A milligram gluten a day keeps the mucosal recovery away: a case report,
F. Biagi, J. Campanella and S. Martucci et al., Nutr Rev 62 (2004), pp. 360–363
A part per what?
Examples of a part per twelve
• One egg out of a dozen
• One cupcake out of a dozen
• One pane from this stained-glass window
A part per hundred (%)
per cent means one part per hundred
X 100 =
A part per million (ppm)
One bushel of wheat (60 lbs) contains 1 million individual kernels
A part per million (ppm)
Use % for larger fractions
1 part per 100
= 0.01
Use ppm for smaller fractions
1 part per 1,000,000
= 0.000001
= 1%
= 0.0001%
Calculating ppm:
• Start with the fraction (part/whole)
• Rather than 2 decimal places to the right → %
• Move 6 decimal places to the right → ppm
20 ppm = 20 minutes in:
A) 4 days
B) 10 weeks
C) 2 years
Quiz answer
20 minutes
4 days
1 hour
1 day
60 minutes
24 hours
=
20
5760
= 0.003472
= 3,472 ppm
20 minutes 1 hour
2 years
1 day
60 minutes 24 hours
1 year
365 days
=
20
1051200
= 0.00001902
= 19 ppm
Perspective: How much is 20 ppm?
20 ppm
20 seconds in 11.5 days
20 inches in 15.8 miles
20 ounces in 62,500 pounds
20¢ in $10,000
Tiny amounts are biologically significant!
Most fish need  4 ppm dissolved oxygen to survive;
Bass and trout seek out waters with 9-12 ppm oxygen
Using ppm to calculate quantity gluten
ppm
gluten

2g
20 g
200 g
Tsp non-dairy
GF creamer
10 GF
crackers
4 slices GF
bread
20
0.04 mg
0.4 mg
4 mg
50
0.1 mg
1 mg
10 mg
100
0.2 mg
2 mg
20 mg
200
0.4 mg
4 mg
40 mg
Reminder: The
ingestion of
contaminating
gluten should be
kept lower than 50
mg/day in the
treatment of CD
Starch vs. Protein
Starches
Proteins
Contain carbon, oxygen,
hydrogen
Contain carbon, oxygen,
hydrogen, nitrogen, often
sulfur
Broken down to only one
material, glucose
Broken down to 22 naturally
occurring amino acids
OH
OH
HO
HO
glucose
O
H
N
OH
OH
O
HO
SH
O
O
O
OH
O
HO
N
H
OH
H
N
N
H
O
O
N
H
OH
O
HN
aminoHO
acid
HN
O
OH
OH
O
HO
O
OH
O
O
O
SH
Essential amino acids (from diet)
HN
HO
O
O
O
HN
O
S
HN
N
H 2N
H 2N
OH
NH 2
H2 N
NH2
OH
Arginine
Arg
R
Histidine
His
H
Isoleucine
Ile
I
OH
Methionine
Met
M
OH
O
O
H2 N
Lysine
Lys
K
H2N
OH
H 2N
Phenylalanine
Phe
F
H 2N
OH
OH
H2 N
OH
Threonine
Thr
T
Leucine
Leu
L
All amino acids have a
1-letter abbreviation
Mnemonic device:
I Have Received Much Kudos
For Learning These Very Well
O
O
O
O
N
H
H 2N
Valine
Val
V
OH
Tryptophan
Trp
W
H 2N
OH
Protein primary structure
Proteins are polymers of amino acids with directionality
C (carboxy)-terminus → N (amino)-terminus
“Beads on a string”
Stability: Proteins vary widely in their stability to heat
Proline is known to provide stability to proteins
Gliadin primary structure (sequence)
MKTFLILALLAIVATTATTAVRVPVPQLQPQNPSQQQPQ
EQVPLVQQQQFLGQQQPFPPQQPYPQPQPFPSQQPYLQL
QPFLQPQLPYSQPQPFRPQQPYPQPQPQYSQPQQPISQQQ
QQQQQQQQQQQQQQQQIIQQILQQQLIPCMDVVLQQHN
IVHGKSQVLQQSTYQLLQELCCQHLWQIPEQSQCQAIHN
VVHAIILHQQQKQQQQPSSQVSFQQPLQQYPLGQGSFRP
SQQNPQAQGSVQPQQLPQFEEIRNLARK
Q: Why does gliadin survive in the presence of enzymes and heat?
Reference: L. Shan, O. Molberg and I. Parrot et al., Structural basis for gluten intolerance in celiac
sprue, Science 297 (2002), pp. 2275–2279.
Protein folding
Secondary structure: Regularly repeating local structures
stabilized by hydrogen bonds. Ex: alpha helix, beta sheet
Tertiary structure: Overall shape or “fold”; the spatial relationship
of the secondary structures relative to one another
Quaternary structure: Results from interaction of more than one
protein molecule (subunit) which functions as part of the larger
assembly or protein complex.
Protein function requires folding
5 common protein functions:
amino acids
•Structure: keratin
•Transport: hemoglobin
•Storage: myoglobin
•Hormones: insulin
•Catalysis: lactase
binding site
Just a few “beads” from the string ≠ gluten
MSG ≠ gluten
NH2
H2N
OH
O
O
L-glutamine
NH2
HO
OH
O
O
L-glutamic acid
NH2
O
+
Na
OH
O
The average person consumes
• 10 - 20 g bound glutamate/day
• 0.5 - 1.5 g MSG/day
O
monosodium glutamate
(both L and D)
L versus D
• In higher mammals, all proteins
contain “L” amino acids.
• During hydrolysis “L” → “D”
• The “D” form is tasteless
Other small molecules ≠ gluten
Not all symptoms are related to celiac disease
O
The structure of caffeine
N
200-300 milligrams daily appears to be safe
N
N
Starbucks’ Tall Coffee = 260 milligrams
N
O
OO
The structure of calcium oxalate
O
Oxalate binds with calcium in your urinary tract
Calcium oxalate can form crystals which eventually grow
into kidney stones
Ca 2+
O-
Hydrolyzed wheat starch
Hydrolysis = hydro (water) + lysis (cleavage/breakage)
A chemical reaction in water that
breaks down wheat starch
North America – starch products usually derived from corn
Any of the following can be made from wheat starch:
• Maltodextrin, glucose syrup, dextrose
• Poly-ols, sugar alcohols (sorbitol, xylitol, mannitol etc…)
• Lactic acid, citric acid, acetic acid (vinegar), ascorbic acid
How do we test for gluten?
ELISA – Enzyme Linked Immunosorbent Assay
ELISA detection limit between 1-2 ppm, with <0.5 ppm reported*
*Sousa et al. Am J Clin Nut, Vol. 87, No. 2, 405-414, February 2008
Alcoholic fermentation
Safe liquors
Wine/brandy
Rum
Whiskey
Vodka
Gin
-
natural sugars present in fruits
cane sugar or molasses
amylase-treated grain
amylase-treated grain or potatoes
grain infused with juniper berry
A vessel allows carbon dioxide to escape,
but prevents outside air from coming in
Without oxygen present:
C6H12O6 → 2 C2H5OH + 2 CO2
glucose → ethanol + carbon dioxide
Distillation: separation based on volatility
Volatility - how readily substances become gases
Stronger intermolecular forces decrease volatility
It takes more energy to break these molecules apart
Volatile
H2O water
acetone
O
C
ethanol
HO
CH 3
O
H3N
CH C
CH 2
Non-volatile
O
CH 3
H3C
H3C
NaCl
H2
C
CH 2
C
H2
C
C
O
CH3
ethyl acetate
O
Also not volatile:
large peptides (proteins)
starches, any polymer
O
NH 2
glutamine
Not solely based on size
of the molecule
Volatility ≠ size
O
Increasing boiling point
Acetone
Molecular Weight: 58.08 g/mole
Boiling Point: 56.5 C
Ethanol
Molecular Weight: 32.04 g/mole
Boiling Point: 64.7 C
Ethyl acetate
Molecular Weight: 88 g/mole
Boiling Point: 77 C
Butanol
Molecular Weight: 74.12 g/mole
Boiling point: 117.7 C
C
3 C’s
1O
CH 3
H3C
HO
H3C
H3C
H2
C
H2
C
H2
C
2 C’s
1O
CH 3
O
CH3
4 C’s
1O
OH
4 C’s
1O
C
O
C
H2
H2
C
Simple distillation procedure
Ethanol gas floats up
circulates through
condensing tubes
Volatile portion
enters gas phase
Tubes are cooled
gas → liquid
Liquid heated
in boiling chamber
Peptides/amino acids
do not evaporate
The separated liquid (distillate)
drips out into a new chamber
Conclusions
There are MANY other names for wheat
 Spelt is NOT an alternative for wheat!
A ppm is not a quantity
 20 ppm can be many different quantities, depends on how
much you ate!
Gluten is a highly stable protein fraction
 Several different fragments of gliadin and glutenin are toxic
 Small molecules do not mimic gluten, but can have negative
health effects of their own
Distillation is separation based on volatility
 Distilled products are gluten-free as long as gluten was not
added post-distillation
THANK YOU!
Brazos Valley Gluten Intolerance Group
Denise Fries, Taylor Jensen
Julie Hoyle (secretary), Jen
Sara Boswell
Kim Melissari (treasurer), Lauren
Monica Michalka
GIG mentors
Cynthia Kupper, RD
Betty Barfield
Family
Shane (husband), Nathan (son)
Karen Jones & Family
McLean family – Rochester, NY
Santiago family – Bryan, TX
Texas A&M University
Dr. Ganesa Gopalakrishnan