Transcript BIOCHEMISTRY (The study of chemicals in living organisms)

Topic Outline
3. Proteins
a. Definition
b. Classification
c. Formation of peptide bond
d. Test for proteins
e. Principle source and dietary importance of proteins
4. Fats
a. Definition
b. Classification
c. Test for fats
d. Principle source and dietary importance of fats
3. PROTEIN
(meaning 'foremost or first')
• (a) Definition: Proteins are complex
organic compounds always containing
the elements carbon, hydrogen,
oxygen and nitrogen and in some
cases sulphur.
Basic
units/building blocks are amino acids.
• 170 aa known, 20 occur commonly.
• Proteins as diverse as:
– enzymes, hormones , mushroom poisons,
antigens, antibodies , nutritive milk proteins
(globular proteins)
– feathers, spider webs, lens protein of the eye,
turtle shell, nail, silk thread (fibrous proteins).
Amino Acids
General Formula
R = residual group
R
H
N
H
AMINO GP
(BASIC)
aC
H
O
C
OH
CARBOXYL GP
(ACIDIC)
(b) Classification of proteins
Criteria used in classification:
1) Structure
• Fibrous (eg. Keratin, collagen, silk)
• Globular (eg. Enzymes, plasma proteins
some hormones)
• Intermediate (eg. Fibrinogen)
(NB: Differences between fibrous and globular
proteins.)
(b) Classification of proteins
Criteria used in classification:
2) Composition
Simple, Conjugated
3. Functions of Proteins
1) Nutrition -- enzymes
2) Respiration & Transport -- Haemoglobin, myoglobin
3) Growth -- hormones
4) Excretion -- urease
5) Support & movement -- muscles
7) Sensitivity & coordination -- hormones
8) Reproduction -- hormones
d. Formation of peptide bonds
O
R1
H
N
C
H
C
N
H
OH
amino end
C
H
C
H
OH
carboxyl end
H2O
H
N
H
O
R2
H
R1
O
C
C
H
O
R2
N
C
H
H
C
OH
Peptide bond
(covalent bond)
Structure = dipeptide
Process = Condensation
(d)Test for Proteins
Biuret’s test
1) Add to the test soln an equal amount of 10%
NaOH. (test works in an alkaline medium)
2) Add 0.5% copper sulphate solution dropwise,
shaking after each drop.
3) A purple/violet colour indicates the presence
of protein
Care is needed since addition of excess CuSO4
results in the disappearance of the purple
colouration (complex formation)
Basis of Test:
O
C
N
• a test for the peptide linkage
H
• N atoms in the peptide chain forms a
purple complex with Cu2+
• works in alkaline medium
• purple complex is the Biuret cpd
which is derived from urea which also
contains the -CONH- gp giving a +ve
result
Biuret Test for Protein
The peptide bond of a protein
is sufficiently reactive to change
the Biuret reagent from blue to
purple.
This test is based on the
interaction between the copper
ions in the Biuret reagent and
the amino groups in the
Original blue
peptide bonds.
colour
Positive test
with biuret's
reagent
Tube 1: Water (control)
Tube 2: Albumin (protein)
Tube 3: Starch
1
2
3
e.
Principle source and dietary
importance of proteins
• http://www.nutristart.com/protein.html - detailed
descriptions of the dietary source and importance of
proteins
• http://www.nutramed.com/nutrition/proteins.htm
• http://www.americanheart.org/presenter.jhtml?identifi
er=11234 - an account of why high-protein diet is not
recommended
Importance of proteins
• Protein is important for everyone, regardless of age
or activity level. Protein is best known for its ability to
build and maintain lean body mass. It also maintains
strong hair, skin, and teeth. But it doesn’t stop there –
protein is also vitally important in maintaining blood,
organs, tendons, and in the production and smooth
functioning of hormones, enzymes, immune cells and
brain neurotransmitters. In short, every tissue and
cell in our body relies on protein for survival.
Source of proteins
• Conventional protein sources include meat,
fish, poultry, eggs, milk and cheese. Beans,
legumes and soybeans (i.e. soymilk and tofu)
are fairly high in protein.
Essential vs. Non-essential AA
• Essentials - those that cannot be
synthesized by body  source: from food
e.g. Leucine, Lysine
• Non-essentials - can be synthesized by the
human body (usually using “backbone” from
other aa or sugars )
e.g. glutamine, proline
How much proteins do I need?
• In order to maintain healthy muscles and to
encourage fat loss, we need to eat high
quality protein 4 to 6 times per day. The
servings should be evenly divided throughout
the day, and roughly 2 to 3 hours apart. For
women, servings of protein should be
between 15 to 25 grams, and men 30 to 40
grams. (The only exception is immediately
after a work out when our bodies can absorb
25 per cent of our total daily protein intake).
(I) What are lipids?
• Diverse group defined by their solubility
rather than by their structures.
• Therefore, they differ widely in size,
structure and functions.
• They are not soluble in water (inorganic
solvent) but soluble in organic solvents (like
benzene, ether, chloroform).
• Basically, lipids are esters of fatty acids
and alcohols.
• They contain C, H and O.
• Examples are:
triglycerides,
phospholipids,
waxes and steroids.
(II) Components of lipids
(1) What are fatty acids?
Contain
the acidic group (carboxyl group – COOH)
General
formula:
R-COOH
(R= H, CH3, C2H5, etc)
R = CnH2n+1
R
is referred to as the
‘hydrocarbon’ tail hydrophobic
(therefore insoluble in
water)
C13H27COOH
(2) Alcohols
• Organic compounds containing the hydroxyl group
(-OH) as the functional group.
• General formula: R-OH where R is the alkyl group
or a hydrogen atom
• E.g. Ethanol - C2H5 OH
Glycerol -
(b) Classification of lipids
• Based on different number and composition
of fatty acids and alcohol and
• Whether they contain other elements like
sulphur, phosphorus, etc.
Triglycerides
• Most common lipids
• Called FATS if solid at 20oC
• Called OILS if liquid at 20oC
c. Formation of lipids
• Organic compounds formed when an acid and an
alcohol undergo condensation reaction / or
dehydration (where water is removed).
Alcohol + Acid --------------- ester + water
Esterification
R-OH + HOOC-R1 --------- R-OOC-R1 + H-OH
•An ester bond (-COO-) is formed
•Hydrolysis is the opposite of condensation, in which
water is added to esters to regenerate the acid and
alcohol. Ester ---------------------- Alcohol + Acid
Acid/ Alkali/ enzyme
Triglycerides
• Structure : 1 glycerol + 3 fatty acids
O
||
H-O-C-R1
O
||
H-O-C-R2
O
||
H-O-C-R3
R1
R2
R3
3 H2O
(d) Test for lipids
(1) Ethanol emulsion test
• Method:
– Add 2cm3 of sample to ethanol.
– Shake vigorously.
– Add equal volume of cold water.
• Result: cloudy white emulsion observed
• Theory:
– Ethanol emulsifies fats.
– When water is added, an emulsion of tiny lipid
droplets reflect light to give an opalescent
appearance.
Test for lipids
Ethanol emulsion test
The one on
the left
forms an
emulsion.
The oil is
dispersed
within the
water.
The tube on the
right contains oil
and water.The
oil can be seen
floating on the
water in the
tube on the
right.
Ethanol emulsion test
In this test, the ethanol will first dissolve the
lipids. Vigorous shaking is required to facilitate
this process.
Cold water will cause the formation of an
emulsion, of tiny ethanol-lipid droplets
suspended in water. These reflect light and give
a white opalescent appearance.
(2) Grease Spot test
• Method:
– Put a small drop of sample n a piece of
paper.
– Soak in water for 1-2 mins.
– Dry over bunsen burner.
– Hold paper up against light.
– Result: If fats are present, spot remains
translucent when the surrounding is dry.
Note: this test is less reliable than the Ethanol Emulsion Test
Grease spot test
(Left negative, right positive)
Sudan III Test
Fat globules are stained red and are less dense than water
(red- stained layer of oil separates from colourless water
below)
SUDAN III test
(Left negative, right positive)
Under the microscope
Below left: oil and water X 40 - Note the large fat
droplet/ globule on the upper, right half of the
photograph.
Below right: oil, water and detergent (emulsifier) X 40 The large oil droplets have been broken up into smaller
droplets after shaking.
e. Principle source and dietary
importance of fats
• http://www.findarticles.com/p/articles/mi_m0K
FY/is_4_20/ai_98464597 - good and bad
fats?
• http://www.healthgoods.com/Education/Nutriti
on_Information/Nutrition_and_Health/nutrition
_guide_men.htm - overall recommended
nutrition standards
Principal source of fats
 Fats and oils used in cooking, such as
peanut oil, olive oil, corn oil etc.
 Salad dressings and spreads, such as
butter, margarine, mayonnaise, peanut
butter etc.
• Skin of meat
Importance of trigyclerides
• Most efficient energy stores
compared to carbohydrates – due to
higher proportion of H compared to O
• Good storage material – compact and
insoluble
• Oxidation provides water
• Heat insulator and keeps marine
animals buoyant
Lipids and diet
• Non-essential FA  CAN be synthesized
by body from carbohydrates
• Essential FA  CANNOT be synthesized
by body; obtained from diet
– Actual function not known -> important for
proper structure and functioning of cells
– Shortage leads to retarded growth, kidney
failure, reproductive deficiency
– E.g. linoleic acid and linolenic acid (unsaturated
FA)
– Especially abundant in vegetable and seed oils