Download Structure of Enzymes

МИНИСТЕРСТВО ОБРАЗОВАНИЯ И НАУКИ РЕСПУБЛИКИ КАЗАХСТАН
ГОСУДАРСТВЕННЫЙ УНИВЕРСИТЕТ имени ШАКАРИМА города СЕМЕЙ
Документ СМК 3 уровня
УММ
УММ
042-18-25.1.55/03-2016
Учебно –методические
Редакция №1
материалы "Basis of bioот 8 сентября 2016 г.
chemistry "
УЧЕБНО-МЕТОДИЧЕСКИЙ КОМПЛЕКС
ДИСЦИПЛИНЫ
"Basis of biochemistry "
Для специальности
5BB080100 – Agronomy
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5B120200 – Veterinary Sanitation
УЧЕБНО-МЕТОДИЧЕСКИЕ МАТЕРИАЛЫ
Семей
2016
СОДЕРЖАНИЕ
1 Глоссарий
2 Лекции
3 Практические и лабораторные задания
4 Самостоятельная работа студента
5 Лист регистрации
3
5
154
194
218
1. The conceptual apparatus
"Rule of 10%" (rule of the pyramid energy R. Lindemann): from one
trophic level ecological pyramid moves to another higher its level (the "ladder"
producer - consumer), on average, about 10% received the previous uro¬ven energy .
Abiotic factors - factors of inanimate nature (cosmic, geophysical, climatic,
spatial, temporal, etc.) that have a direct or indirect impact on living organisms.
Act of tolerance (V.Shelford): environmental factors, with specific conditions pessimal (unfavorable as a minimum, and excess) value that limits the ability
of the species in these conditions, in spite of and in spite of the optimal combination of certain other conditions.
Agrocenoses - community of organisms cultured and accompanying them in
agriculture.
Amensalizm - type of interspecies relationships, in which in a joint environment, one kind of organism suppresses susche¬stvovanie another species without experiencing resistance.
Anthropogenic factors - factors that have arisen as a result of human activity.
Autotrophs - organisms can synthesize or¬ganicheskoe agent of carbon dioxide, water and salts mine¬ralnyh. energy sources are used for the biosynthesis of
light (in photoautotrophs) or oxidation of a number of inorganic substances (in
chemoautotrophs).
Bio-accumulation - the accumulation of substances (man-made pollutants)
in the body increasing trophic levels.
Biogen - a nutrient; nutrients, nutrients essential chemical elements that
make up the substance of living organisms, carbon, hydrogen, oxygen, nitrogen,
sulfur, phosphorus.
Boreal zone - the zone of temperate forests.
Chemosynthesis - synthesis of organic substances in chemoautotrophic bacteria using as power sources of certain inorganic oxidizing substances.
Co-evolution - in parallel, the joint conjugate evolution of mankind and nature.
Consuments - heterotrophic organisms (mostly animals) who consume organic matter other plant organisms (herbivores - herbivores) and animals (carnivores - zoophages).
Cryostasis - temporary total suspension of the body's vital functions associated with the onset of unfavorable conditions or with extreme phase of individual
development.
Depopulation - a reduction in population, population.
Desertification (aridity) - the process of depletion of vegetation associated
with a persistent reduction in moisture areas, its transformation in the arid zone,
topically, followed by the previous member of the chain.
Detritophages - organisms that feed on detritus (saprophagous).
Detritus - dead organic matter, isolation and decay organisms products.
Disadaptation - violations of vital activity caused by the incompleteness of
acclimation, the inability to fully adapt to changing environmental conditions.
Dissimilation - the disintegration of complex organic substances in the body,
accompanied by the release of energy, which is used in the processes of life.
Ecological culture - system of scientific knowledge about the human interaction of society and nature; environmental value orientations, rules and regulations; moral and aesthetic attitude towards nature; skills for the study of nature and
its protection.
Edafon - a set of soil animal population of the Earth's thermal radiation air.
Education - a relatively meaningful and purposeful nurturing person in accordance with the specific objectives of groups and organizations, in which it is
carried out.
Ektotermy - organisms, the body temperature is a little different from the
temperature of the environment and follow its changes: lower organisms, plants,
cold-blooded animals.
Emergence - the emergence of completely new properties of the interaction
of two or more objects or phenomena, properties that are not simply the sum of the
original.
Endotherm - warm-blooded animals birds and mammals, are capable of using the internal mechanisms of thermoregulation to maintain a relatively constant
body temperature, to a certain extent independent of the ambient temperature. "
Environmental - trudnosti crisis, environmental problems due to anthropogenic human activities.
Environmental education - the formation of the human conscious perception of the environment, the conviction of the need for respect for nature, rational
use of its wealth of natural resources.
Environmental education - tselenapravlennaya specially organized, systematic educational activities aimed at the development of environmental education
and upbringing of children, on the formation of environmental awareness and skills
for the study of nature and its protection.
Environmental education and training of students - pedagogical process,
which ultimately should provide insight into the importance of proper behavior in
the natural environment, the ability to anticipate and assess the impact of its activities, the realization that the man part of nature /
Environmental upbringing - purposeful human development, including the
formation of its ecological culture, the perception of not only the public, but also of
environmental norms and values;
Eurybionts (evrieki) - organisms that exist in a wide range of changes in environmental conditions: temperature (evritermy), humidity (evrigidridnye organisms), food choices (euryphages), etc.
Eutrophication - the excessive enrichment of water with nutrients.
Gene flow - the process of undirected random changes in gene frequency in
a population.
Heterotrophic organisms - organisms that feed on organic matter ready.
Hibernation - a significant reduction in the level of life upon the occurrence
of adverse external conditions (for example, hibernating animals).
Homeostasis - the ability of an organism or organisms of the system to
maintain stable dynamic equilibrium in a changing environment.
Humid Zone - area or natural-climatic zone with high
Law of constancy of the amount of living matter of the biosphere (Vernadsky): The number of living matter (biomass of all organisms) for the biosphere of
the geological eras.
Noogenesis (noospherogenesis) - the process of formation of the noosphere.
optimality law: any system with the highest efficiency in the functioning of
some specific spatio-temporal limits to her.
Phenotype - a set of genetically determined characteristics and properties
of the organism.
Photoperiodism - change the state of biological systems due to the natural
rhythm of light exposure, the change of day and night, seasonal changes in the
length of daylight.
Phytocoenosis - multispecies plant community.
Phytophagy - herbivorous animals.
Phytoplankton - a set of micro-algae, small plant organisms that live in the
water column
Rule D.Allena: increase protruding body parts of one species or closely related species of warm-blooded animals (limbs, tail, ears) when moving from north
to south.
Rule K.Bergmana: warm-blooded animals, subject to geographical variation, the body size of individuals statistically (on average) more than in populations
living in colder parts of its range.
Security Environment - the degree of protection of the territorial complex
ecosystems, the human potential of the eco-logical lesions derived from the magnitude of environmental risk.
Technosphere - "technical envelope" - artificially transformed space of the
planet, being under the influence of human industrial activity products.
The capacity of the ecosystem - the maximum size of the population of one
species, this ecosystem which is capable of supporting in certain environmental
conditions for a long time.
The law of irreversibility of evolution (L. Dollo): evolution is irreversible;
organism (population, species) can not return to their previous state, already implemented in a number of his ancestors.
The noosphere - the letters "thinking envelope", the scope of reason; according to Vernadsky - a qualitatively new, higher stage of development of the biosphere under the control of a reasonable human activity.
The ontogenesis - the individual development of the organism; multicellular
egg from fertilization to aging and death.
Valence Environment - (tolerance limits) the characteristic type of ability,
populations exist in different
Valeology - science for the preservation and strengthening of health, healthy
lifestyles.
Zoophages - carnivorous organisms that feed on other animals or their species (cannibalism).
Environmental education - process of mastering by students the system of
scientific knowledge about the natural environment as the reality of human life,
about the impact of industrial activity on the environment of society, as well as the
knowledge and skills of environmental activities.
Environmental awareness -environmental knowledge (information, conclusions and generalizations) about the natural environment and interacting with her
man, ecological thinking, feeling and will.
Environmental science is a generic of the relation of organisms in the environment (Haeckel), the science of organization and functioning supraorganismal
systems at various levels: the populations of species, biocenoses (communities),
ecosystems and the biosphere.
2.Lectures
Module 1.Introduction.
Biochemistry subject
The main principles of chemical logic of a live condition. The concept about
macro- and microelements.
I. Basic Chemical Concepts
A. Atoms
1. Def.- the smallest unit of an element that can combine chemically
with other elements
Structure
a. Proton (+) charged
b. Neutron (not charged)
c. Electron (-) charged
1. Electrons exist in distinct orbital clouds
2. s, p, and d orbitals
3. Orbitals combine to form energy levels: K, L, M, N, etc
d. Protons and neutrons are the same mass and make up the nucleus
2. Identification
a. Atomic number: number of protons
b. Atomic mass number: number of protons + neutrons
c. Atoms are organized into groups in the periodic table
3. Isotopes
a. Two atoms with the same atomic number but different atomic
mass numbers
b. Differ only in the number of neutrons
c. Some are radioactive (radioisotopes)
B. Compounds
4. Def: a combination of two or more elements which are joined chemically
5. Chemical bonding
a. Ionic: when an atom will either give or take an electron from
another atom
1. Cation: positive ion
2. Anion: negative ion
3. Electrostatic forces hold the atoms together
b. Covalent: when atoms share electrons
1. Forms single or multiple bonds
2. Sharing of electrons hold the atoms together
c. Hydrogen bonds: weak links between the hydrogen (+) end of
one polar molecule and the negative end of another polar molecule
C. Acids and Bases
6. Acid: a substance which releases a H+ ion
7. Base: a substance which releases an OH- ion
8. pH scale
a. A method of determining how acidic or basic a solution is
b. Negative logarithmic scale: 0 (acidic) to 14 (basic) (alkaline)
c. pH 7.0 is neutral (water)
9. Buffers: a substance which limits the change of pH
D. Basic chemical reactions
10.Synthesis: two or more atoms or molecules are combined
11.Decomposition: molecules are broken down into simpler forms
12.Reduction
a. The addition of electrons to a molecule
b. Often accompanied by a gain of a hydrogen nucleus (proton)
13.Oxidation
a. The removal of electrons from a molecule
b. Often accompanied by a loss of a proton
c. Oxidized atoms are more reactive than reduced atoms
II. Basic Biochemistry Concepts
A. Building Materials of Life
1. Inorganic compounds
2. Organic compounds
a. All contain some form of carbon
b. Biosynthesis: the manufacture of things by a living organism
3. Carbohydrates
a. Structure
1. Contain only C, H, and O
2. Ratio of O:H is 1:2 (same as water H2O)
b. Reactions involving carbohydrates
1. Dehydration synthesis: joining two molecules by removing water
2. Hydrolysis: splitting two molecules by adding water
c. Types
1. Monosaccharides (simple sugars)
a. 5-carbon: ribose
b. 6-carbon: C6H12O6 (Glucose, Galactose, Fructose)
2. Disaccharides
a. Two monosaccharides joined together (dehydration synthesis)
b. Sucrose (table sugar): Glucose + Fructose
c. Maltose (malt sugar): Glucose + Glucose
d. Lactose (milk sugar): Glucose + Galactose
3. Polysaccharides
a. Starch: straight chain of glucose (food storage in plants)
b. Glycogen: branched chain of glucose (food storage in animals)
c. Cellulose: Zig-zag chain of glucose (non-digestible
roughage)
4. Lipids
a. Fats (triglycerides)
1. 3 fatty acid molecules + 1 glycerol joined by dehydration synthesis
2. Saturated: no double bonds between carbons
3. Unsaturated: at least one double bond
b. Phospholipids
1. 2 fatty acids + 1 glycerol + 1 phosphate
2. Hydrophobic end (fat): water fearing (non-polar)
3. Hydrophilic end (phosphate): water loving (polar)
4. Used extensively in cell membranes
c. Sterols: multi-ringed compounds
1. Cholesterol
a. HDL: High density lipoprotein ("good" cholesterol)
b. LDL: Low density lipoprotein ("bad" cholesterol)
2. Hormones: i.e. prostaglandins, cortisone, etc
5. Proteins
a. Structure: composed of 20 basic amino acids
b. Protein synthesis
1. Two amino acids are brought together and dehydration synthesis between the amino acids forms a peptide bond
2. Protein = polypeptide chain
3. The order of the amino acids is critical to the function of a protein
c. Enzymes: large proteins which catalyze reactions
1. Structure
a. Active site: attachment site for substrates
b. Substrate: molecule which reacts with the enzyme and is
changed
c. Coenzyme: non-protein which helps to complete the active site (vitamins)
2. Enzyme action
a. Enzyme & substrate bind at the active site
b. Reaction proceeds (lytic- splitting apart, synthetic - putting together)
c. Enzyme and product(s) separate
6. Nucleic acids
a. Consist of long chains of repeating subunits (nucleotides)
b. Nucleotide structure
1. 5-carbon sugar (ribose)
2. Phosphate group (PO4)
3. Organic nitrogen-containing base
c. DNA: Deoxyribonucleic acid
1. Used to store biological information
2. DNA base pairs
a. Guanine - Cytosine (G - C)
b. Adenine - Thymine (A - T)
3. Double-stranded helix shape formed by hydrogen bonds
d. RNA: Ribonucleic acid
1. Used as working blueprints for protein synthesis
2. RNA base pairs
a. Guanine - Cytosine (G - C)
b. Adenine - Uracil (A - U)
3. Single strand
III. Energy and its Changes
A. Kinetic energy: energy of motion
B. Potential energy: energy of position (stored energy)
C. Kinetic and potential energy are interconvertable
D. Energy in chemical reactions
1. Exothermic: reactions which release energy (heat)
2. Endothermic: reactions which require energy
3. Activation energy: energy needed to start a chemical reaction
Module 2.Aminoacids.
Amino acids: classification, structure, stereochemistry, physical and chemical
properties and classification amino acids forming proteins.

Properties of the 20 amino acids that occur in peptides and proteins are crucial to the structure and function of proteins.
stereochemistry
o relative hydrophobicity or polarity
o hydrogen bonding properties
o ionization properties
o other chemical properties
 Condensation of 2 amino acids forms the peptide bond, the amide linkage
holding amino acid residues in peptide and protein polymers.
 Properties of the peptide bond have major consequences in terms of the 3dimensional structures of proteins
There's an excellent website on amino acids being developed here in the Department of Biochemistry and Molecular Biophysics; parts of it are still under construction, but there are links to various very useful parts of it here in these notes,
and indeed parts of it may be used in class.
BASICS
o


Proteins are polymers of -amino acids:
 There are 20 different amino acids found in proteins
and they differ by the nature of the R group.
Both the -amino group (amino group substituent on the C) and the carboxyl group (carboxyl substituent on the C) are ionizable.
o
-COOH group: a weak acid, can DONATE its proton, with a pKa
of about 2-3. What's the conjugate base form of the carboxyl group?
Which form is charged, and is it a positive or a negative charge?
o
-NH2 group: a weak base (there's an unshared pair of electrons on
the N; the neutral amino group can ACCEPT a proton). What's the
conjugate acid form of the amino group? Which form is charged, and
is it a positive or a negative charge?
o pKas of -amino and -carboxyl groups are different for different
amino acids, and also are altered if they're the terminal groups on a
chain of amino acids, i.e., a peptide or protein.
Predominant form in H2O is the zwitterion:
.
Stereochemistry of the amino acids

-carbon is asymmetric (has four different substituents) except for one amino acid, for which the R group is a hydrogen atom.





amino acids occur as enantiomers (nonsuperimposable complete mirror images)
L-amino acids are the naturally occurring enantiomers found in all proteins
There are naturally occurring D-amino acids, but not in proteins (found in
some bacterial cell wall peptide structures, in some peptide antibiotics,
etc.) (D_L)
Perspective formulas show stereochemistry; projection formulas CAN be
written "correctly", with convention that horizontal bonds project out of paper and vertical bonds behind plane of paper, but often biochemists use projection formulas casually (inaccurately), knowing that if it's in a protein, it's
always an L-amino acid.
Absolute configurations of D-glyceraldehyde as the reference compound
for -amino acids. D- and L- apply only to the absolute configuration
around the chiral carbon; 2 of the 20 amino acids (threonine and isoleucine) have a second chiral center, requiring the RS system to describe their
structures accurately, but we aren't going to worry about using the RS system here.
Which of the amino acids does NOT have a chiral center, so has no D/L isomers?
Amino Acid Abbreviations
amino acid (or residue
3-letter
1-letter
Mnemonic for 1-letter
in protein)
abbreviation
abbreviation
abbreviation
Glycine
Gly
G
Glycine
Alanine
Ala
A
Alanine
Valine
Val
V
Valine
Leucine
Leu
L
Leucine
Isoleucine
Proline
Methionine
Phenylalanine
Ile
Pro
Met
Phe
I
P
M
F
Isoleucine
Proline
Methionine
Fenylalanine
tWyptophan (or tWo
Tryptophan
Trp
W
rings)
Tyrosine
Tyr
Y
tYrosine
Serine
Ser
S
Serine
Threonine
Thr
T
Threonine
Cysteine
Cys
C
Cysteine
Aspartic Acid
Asp**
D
asparDic acid
Glutamic Acid
Glu*
E
gluEtamic acid
Asparagine
Asn**
N
asparagiNe
Glutamine
Gln*
Q
Q-tamine
Histidine
His
H
Histidine
Lysine
Lys
K
(before L)
Arginine
Arg
R
aRginine
* Glx = either acid or amide (when it isn't known which it is)
**Asx = either acid or amide (when it isn't known which it is)
Properties of Amino Acid Side Chains
Side chains ("R groups") provide proteins with unique structural and functional
properties.
Additional C atoms in R groups (after the C) designated by successive Greek
letters:
as shown in the structure of the amino acid LYSINE
(Nelson & Cox: Lehninger Principles of Biochemistry, 3rd ed., p. 116):
Side chain classes
 The side chains of the amino acids play an essential role in determining the
properties of proteins.
There is a wide diversity in the chemical properties of amino acid side
chains, but they can be grouped into classes, sometimes with overlapping
"membership" (e.g., tyrosine is both aromatic and hydroxyl-containing).
Other classifications are also possible (for example, the 5 classes in textbook, Fig. 5-5, discussed below). You are expected to know all 20 amino
acid structures and their R group properties, including ionization properties
(see table below with "generic" pKa values for groups in peptides and proteins and links to titration curves, and the PDF of proton dissociation reactions).


Side Chain Class
Amino Acids
Aliphatic
glycine, alanine, valine, leucine, isoleucine
Cyclic
proline
Aromatic
phenylalanine, tyrosine,
tryptophan
Hydroxyl-Containing
serine, threonine, tyrosine
Sulfur-Containing
cysteine, methionine
Basic
histidine, lysine, arginine
Acidic and Their Amides
aspartic acid, glutamic acid,
asparagine, glutamine
Nonpolar, aliphatic R groups
o Gly: quite water-soluble (as is Pro)
o Ala, Val , Leu and Ile: increasing hydrophobicity with increasing
number of C atoms in hydrocarbon chain
o Pro: cyclic (--> unusual properties)
 shares many properties with the aliphatic group
 rigidity of ring plays critical role in protein structure (more
about that later)
o Met: methyl thioether (S-containing)
 quite hydrophobic
 Met's terminal methyl group important in metabolism
Aromatic R groups
o Phe: phenyl group (linked to -CH2, so Phe = alanine with a phenyl
substituent on the methylene C)
 VERY hydrophobic.
o Trp: indole functional group on C
 electronegative atom in ring system
 not as hydrophobic as Phe
 hydrogen bonding capability (donor? acceptor? how many hydrogen bonds?)
o Tyr: phenylalanine with aromatic OH group (phenolic OH) = phydroxyphenylalanine
 ionizable (pKa around 10; loss of proton gives phenolate anion)



hydrogen bonding capability (donor? acceptor? how many hydrogen bonds?)
Tyr R group is the least hydrophobic of the 3 aromatic amino
acid side chains.
Polar, uncharged R groups
o Ser and Thr: aliphatic OH groups, not ionizable in pH range 1-13
 pKa values so high that under any biologically reasonable pH
conditions they're polar but not ionizable.
 hydrogen bonding capability (donor? acceptor? how many hydrogen bonds?)
o Asn and Gln: amide functional groups
 VERY polar, but NOT ionizable
 hydrogen bonding capability (donor? acceptor? how many hydrogen bonds?)
o Cys: thiol (also called a sulfhydryl group) -- not very polar,
and IS ionizable
 sulfur atom makes protonated -SH group more hydrophobic
than an aliphatic OH group
 thiol DOES lose its proton in physiologically relevant pH range
(pKa about 8.5)
 generates -S (thiolate anion is quite hydrophilic due to the
charge).
o

Positively charged R groups (sometimes called "basic" R groups)
o Arg: guanidino group
 VERY high pKa (~12+), so a very weak acid (stronger base)
 carries + charge all across physiological pH range
 resonance forms of guanidino group stabilize protonated form
(charge is delocalized)
 hydrogen bonding capability (donor? acceptor? how many hydrogen bonds?)
o Lys: -amino group (a primary amine)
 pKa about 10
 protonated form (predominates at physiological pH) carries +
charge
 hydrogen bonding capability (donor? acceptor? how many hydrogen bonds?)
o His: imidazole functional group (has 2 N atoms in 5-membered unsaturated ring)
 pKa about 6-6.5
 protonated form carries + charge, but at pH 7 predominant form
is neutral (despite textbook's categorization as "positively
charged")
 very important player in catalytic activity of many enzymes
 hydrogen bonding capability, and also proton donor/acceptor

Negatively charged R groups (sometimes called "acidic" R groups)
o Asp and Glu: side chain carboxyl groups
 pKa values around 4
 predominant form at physiological pH = carboxylate anion

hydrogen bonding capability (donor? acceptor? how many hydrogen bonds?)
Relative hydrophobicity/hydrophilicity of amino acid R groups
 Table 12.2 : Polarity scale for amino acid residues based on free energy
changes for moving a residue from a hydrophobic environment (dielectric
constant = 2) into H2O.
 Similar trends for relative hydrophobicities in text Table 5-1 (diff. numerical
scale, and not arranged in order of relative polarity)
 Depending on how transfer experiments are done, different absolute numbers can be obtained, but the general trends of relative polarity are clear
o Phe, Met, Ile, Leu, Val are very hydrophobic
o Arg, Asp, Lys, Glu, Asn, Gln, and His are quite hydrophilic
o The rest are in between -- neither very polar nor very hydrophobic

Reversible oxidation of 2 cysteine side chain thiols to form cystine, or rereduction to 2 thiols
o
o
o
disulfide bonds between 2 Cys residues in a (usually extracellular)
protein
often a critical structural feature in extracellular proteins (stabilize
folded structures, in interior of protein structure)
When found in intracellular proteins, usually have a functional role.
Ionization Properties of Amino Acid Functional Groups (in PEPTIDES AND
PROTEINS)
 weak conjugate acid/base groups in peptides and proteins crucial to functions
o only one -amino and one -carboxyl group on a peptide or proteins
(at the termini of the chain) because the rest of the -amino and carboxyl groups are tied up in amide bonds holding monomers together in polymer (more later)
o side chain ionizable groups (only 7 of the 20 amino acids)
 PDF of the acid dissociation reactions for functional groups of amino acid
residues in peptides and proteins
 ionization states of side chain weak acid groups control charges on protein
 Note: local environment in peptide or protein determines actual pKa of that
specific group, so the ranges shown below (and the rather arbitrary "generic"
values, rounded off for simplicity) are only the usual expected ranges for
pKa values for the functional groups in peptides and proteins; the pKa of a
specific group in a specific protein can lie significantly outside the expected
range if the local environment is unusual.
 links in table below are to titration curves for that amino acid or functional
group
Group
usual pKa range, in peptides & proteins (approx."generic"pKa )
a-Carboxyl (terminal group
of peptide or protein)
~3.0 - 4.0 (generic 3.0)
Asp, Glu (side chain carboxyl)
~4.0 - 4.5 (generic 4.0)
His (imidazole)
~6.0 - 7.4 (generic 6.5)
Cys (thiol, SH)
~8.5 - 9.0 (generic 8.5)
Tyr (phenolic OH)
a-Amino (terminal group
of peptide or protein)
Lys
-amino)
Arg (guanidino)
~9.5 - 10.5 (generic 10.0)
~8.0 - 9.0 (generic 8.0)
~9.8 - 10.4 (generic 10.0)
~12.0 - 12.5 (generic 12.0)
Isoelectric point (pI)
 pI = "isoelectric pH" = "isoelectric point" = pH at which the NET charge on
a molecule is ZERO.
o If pH < pI, net charge is positive (more + than - charges)
o If pH > pI, net charge is negative (more - than + charges)
 pI = the pH exactly halfway between the two pKa values surrounding the zero net charge equivalence point on the titration curve (examples to be analyzed in class: Gly and His)
 Fig. 5-10. Titration curve of glycine (Nelson & Cox: Lehninger Principles of
Biochemistry, 3rd ed.)






Molecular separations based on charge properties (paper electrophoresis of
amino acids as an example)
paper strip soaked in buffer, in contact with 2 reservoirs with electrodes
connected to a power supply
Buffer reserBuffer reservoir #1
voir #2
_
+
O
(anode; ani(cathode; catons move toions move
ward it)
toward it)
^
Ultraviolet absorbance of amino acid side chains
Aromatic amino acids (Trp, Tyr, Phe) absorb light in the near ultraviolet region of the spectrum (250-300 nm).
Trp has highest molar absorptivity, followed by Tyr, with Phe making only a
small contribution.
Disulfide bonds (between Cys residues in proteins) also absorb in the uv
range, but much less than the aromatics.
Fig. 5-6 (Nelson & Cox, Lehninger Principles of Biochemistry, 3rd ed.):
Absorbance of ultraviolet light by aromatic amino acids
Posttranslational modifications of amino acid side chains
 chemical modifications AFTER biosynthesis of proteins
 occur for a few amino acid residues in some proteins
 Some examples (see also Fig. 5-8, Nelson & Cox: Lehninger Principles of
Biochemistry, 3rd ed.)):
O-Phosphoserine 4-Hydroxyproline 5-Hydroxylysine
-carboxyglutamate
 reversible phosphorylation and dephosphorylation of Ser, Thr, and Tyr residues very important in covalent regulation of activity of some enzymes and
many biosignalling proteins, including some hormone receptors and transcription factors
 4-hydroxyproline & 5-hydroxylysine important in structure of collagen (fibrous protein in connective tissue)

-carboxyglutamate important in a number of proteins whose function involves Ca2+ binding, including several proteins involved in blood clotting
Chemical Reactions of Amino Acids
 All amino acids have at least two reactive groups: the
amino and carboxyl groups and these groups can react with a variety of reagents. Here
are two examples:


A particularly interesting example is the green fluorescent protein (GFP)
from the Pacific Northwest jellyfish Aequorea victoria, which has generated
intense interest as a marker for gene expression and localization of gene
products. The chromophore, which results from the spontaneous cyclization
and oxidation of the sequence -Ser65-Tyr66-Gly67- , is unusual because it
does not involve a non-protein chromophore, as is usually the case for colored proteins. The chromophore is buried in the interior of GFP.

The Peptide Bond
 Peptides and proteins:polymers of amino acids joined bypeptide bonds
 amide linkages from condensation of -carboxyl group of one amino acid
with -amino group of another amino acid

process repeated many times --> linear chain of amino acids, a polypeptide
chain

convention: sequence written from left to right starting with residue with
free -amino group (the N-terminal or amino terminal amino acid residue)
and ending with the residue containing the free -carboxyl group (the Cterminal or carboxyl terminal residue),
e.g., NH2-Glu-Gly-Ala-Lys-COOH = EGAK
 average residue mass ~110 (average Mr of the 20 amino acids minus Mr of
H2O)
 a polypeptide chain with 100 amino acid residues would have a
Mr of about 11,000)
 small peptides (a "few" amino acid residues) = oligopeptides
Peptide bond formation endergonic ( Go' ~21 kJ/mol)
 (How would a cell make the reaction go in the direction of condensation in
an aqueous environment? no details needed here for biochemical mechanism
-- that's covered in BIOC 411)
 peptide bonds metastable in aqueous environment -- equilibrium lies far in
direction of hydrolysis, but RATE of hydrolysis very slow in absence of catalyst
 Enzymes that catalyze peptide bond hydrolysis
= peptidases or proteases, e.g., (specific examples of proteases) your digestive proteases like trypsin and pepsin
Ionization properties of peptides
 analyzed the same way as for free amino acids
 one -amino group (pKa approx. 8) and one -carboxyl group
(pKa approx. 3), plus any ionizable side chains on residues in the peptide
 To figure out approximate net charge of a peptide at a given pH:
o make yourself notes on the sequence to keep track of what you're doing
o add up charges on all the ionizable groups
Example: Fig. 5-14 (Nelson & Cox: Lehninger Principles of Biochemistry, 3rd
ed.): pentapeptide SGYAL = Ser-Gly-Tyr-Ala-Leu
= Serylglycyltyrosylalanylleucine
Amino Acid Analysis
 Sequence of amino acids in a protein is dictated by the sequence of nucleotides in the gene encoding that protein:
(from Berg, Tymoczko & Stryer, Biochemistry, 5th ed., p. 28)



Each protein (unique sequence) has unique amino acid composition.
Can chemically hydrolyze (hot 6N HCl) a pure protein to generate the free
amino acids and determine its amino acid composition chromatographically
Because side chains of the amino acids have different properties, can separate and quantitate all 20 amino acids using a variety of chromatographic
techniques, as illustrated below.
Peptide bond has resonance structures --> partial double bond character
 Due to the partial double bond character of the peptide bond, the O, C, N
and H atoms are nearly planar and there is no rotation about the peptide
bond (peptide). As we shall see later, the planarity of the these elements has
important consequences for the three dimensional structure of proteins.

Generally, the two C groups are in a trans configuration, which minimizes
steric interaction (cis/trans).
Modul 3. Proteins.
Primary structure of proteins. Secondary, tertiary and quaternary structures. Chemical properties and methods of definition of primary structure of
proteins. Classification of proteins. The role of proteins in a food.
Peptides and Protein Primary Structure
 Peptide bond formation: Note that a peptide bond is simply an amide bond
between the alpha carboxyl and amino groups of amino acids. If we write
the reacting groups in their unionized (acid and amine) forms, then we can
see the reaction takes place with the loss of the elements of water, via an attack of the lone-pair electrons of the amine on the carbonyl carbon of the
carboxyl group:
Now that we have looked at peptide bond formation, we next want to look at the
structure of this bond and the sequence of amino acid residues (primary structures)
of proteins. (Note that "residue" refers to the remainder of a molecule after it is incorporated into a polymer.)
 The peptide bond is formed with the elimination of water, giving a planar
bond between the carboxyl carbon and the amino nitrogen. [overhead 5.8
MvH] This is due to the partial double bond character on the amide/peptide
bond as seen in the shorter bond length (0.133 nm vs. 0.146 nm). [overhead
7-2, V&V] This bond is nearly always trans in proteins due to steric interactions of the amide hydrogen and oxygen, except for proline.
 Linear peptides will have free amino- and carboxy- terminal groups. Thus
they will exhibit titration curves similar to a free amino acid, but with the
pKa values shifted closer to simple acid and amine values (there will be no
charge stabilization).
 By convention the amino terminal residue is written on the left progressing
to the carboxyl terminal residue on the right: +H3N-aa-aa-aa-aa-CO2-.
 Can determine the composition of a peptide by acid hydrolysis and amino
acid analysis.
 Can sequence proteins by specific enzyme and chemical hydrolysis to give
peptides which can then be run through sequenators (up to about 100 aa's).
 Amino acid sequences have been used to help determine relatedness of organisms.
3-D Structure of Proteins
Overview: Proteins are commonly large (MW > 6,000), globular molecules serving many functions.
Proteins are complex systems - difficult to understand at a fundamental structural
level. Thus we search for patterns using normal perceptual tools: regularity, clustering, cleavage/separation/emptiness.
We are then able to discern alpha helices, beta sheets, beta turns, and "random" regions. 310 helical regions show up with computer searches. None of these is necessarily more or less random than others, they are simply easier or more difficult for
us to perceive as ordered. They exist through our rationalization. Often structural
elements also appear to serve a functional role, thou this is through our dissection
of the molecular machine.
Look at theoretical possibilities resulting from the available bond angles around the
peptide bond system
 Most peptide bonds are trans because of reduced steric hindrance. Most exceptions are with proline which has nearly equal hindrance in both cis and
trans [overhead 5.8 P]
 Any rotation in the peptide chain will therefore take place around the two
bonds of the alpha carbon, referred to as the phi (f) and psi (y) bonds. There
are a restricted number of angles which these bonds can achieve (Figure 4.8)
[overhead 5.9 P, V&V 7.6]. Of course the range of angles will be further reduced due to side chains.
 If we assume hard spherical atoms with van der Waals radii, we can determine the accessible phi (f) and psi (y) angles. This procedure was followed
by Ramachandran to produce the Ramachandran plot, an example is seen
in Figure 4.9 of your text [overhead 6.2, MvH; 7.7 V&V].
o There are only a few regions of possible angles available to the alpha
carbon bonds as shown on this plot.
o Note that the common secondary structures, the alpha helix, the beta
strand, and the collagen triple helix all occur in these regions.
o Of course real atoms are somewhat compressible and real bonds can
bend a little, so we might wonder how this plot stacks up to reality. A
study of the distribution of conformation angles of a thousand amino
acid residues in eight proteins as determined by x-ray diffraction
showed that most of the values do indeed fall in the predicted regions.
Most of the residues outside of these regions are glycines, with the
least restriction.
Let's go back and look at overall shape and interpret it. Look for substructures that
recur in various molecules. Perhaps we see a globule is made of subglobules. Look
closer and we see alpha helices and beta structures. Finally we can discern aa residues.
In order to understand and categorize their organization, protein structure has been
divided into four hierarchical levels and a couple of sublevels:
 Primary structure (1°) : the linear order or sequence of peptide bonded
amino acid residues, beginning at the N-terminus. (Characteristic bond type:
covalent.)
 Secondary structure (2°): the steric relations of residues nearby in the primary structure which give rise to local regularities of conformation. These
structures are maintained by hydrogen bonds between peptide bond carbonyl
oxygens and amide hydrogens. The major secondary structural elements are
the alpha helix and the beta strand. (Characteristic bond type: hydrogen.)
 Tertiary structure (3°): the steric relations of residues distant in the primary sequence; the overall folding pattern of a single covalently linked molecule. (Characteristic bond type: hydrophobic; others: hydrogen, ion-pair, van
der Waals, disulfide.)
Super secondary structure (motifs): defined associations of secondary structural elements. (Characteristic bond type: hydrogen &
hydrophobic.)
o Domains: independent folding regions within a protein. The
group/pattern of secondary structures forming a Domain's tertiary
structure is called a Fold. (Characteristic bond type: hydrophobic;
others: hydrogen, ion-pair, van der Waals.)
 Quarternary structure (4°): the association of two or more independent
proteins via non-covalent forces to give a multimeric protein. The individual
peptide units of this protein are referred to as subunits, and they may be
identical or different from one another. (Characteristic bond type: hydrophobic; others: hydrogen, ion-pair, van der Waals.)
3-D Structure of Proteins 2
Secondary Structure
Tertiary structure (3°): the steric relations of residues distant in the primary sequence; the overall folding pattern of a single covalently linked molecule. (Characteristic bond type: hydrophobic; others: hydrogen, ion-pair, van der Waals, disulfide.)
 Super secondary structure (motifs): defined associations of secondary
structural elements. (Characteristic bond type: hydrogen & hydrophobic.)
 Domains: independent folding regions within a protein. The group/pattern
of secondary structures forming a Domain's tertiary structure is called
a Fold. (Characteristic bond type: hydrophobic; others: hydrogen, ion-pair,
van der Waals.)
o
Last time looked at what is possible given the bond angles etc. between amino acid
residues. Now can look at specific structures.
Alpha helix: (Figure 4.10, pg 90 of your text) [overhead 2.31 S, 5.15 P] The most
frequent secondary structure is the right-handed a-helix.
 In this cylinder-like structure the amino acid residues curl around in a
spring/rod-like structure.
 There is a rise/residue (movement along the axis) of 0.15 nm and a pitch
(rise/turn) of 0.54 nm.
 There are 3.6 residues per turn and 13 atoms/H-bonded "ring" - this makes it
a 3.613 helix.
 Very importantly, the H-bonds are nearly linear and therefore of near maximum strength. The side chains of the helix stick out from the sides.
o The stability of the helix is determined in part by the side chains. Thus
glycine allows too much rotational freedom to favor this structure,
while very large or like charged side chains can also destabilize it.
 As you might expect a proline residue stops a helix abruptly since proline' s
angles are not accommodated in the helix.
Beta Strand: (Figure 4.15, pg 93 of your text) [overhead 5.19 P] The next secondary structural element is the beta-strand, which is seen in the supersecondary structures called parallel and anti-parallel beta sheets [overheads 7.16 & 17 V&V].

The beta strand is in a sense an abstract structure, since, unlike the a-helix, a
beta-strand does not exist alone, there is always another strand to make a
sheet.
 In the older literature beta-sheets are considered secondary structures, but
they are more consistently considered super secondary with the current nomenclature.
 Beta strands are nearly fully extended, thus they have very little extensibility
(stretch).
 Beta strands are stabilized by hydrogen bonding to adjacent beta-strands.
Thus they are stabilized by inter-strand H-bonds whereas a-helices are stabilized by intra-strand H-bonds.
Aside: Fibrous proteins: alpha-keratin (hair etc., alpha-helix based) [overhead 711 V&V, 7-25 & 26]; stretched alpha-keratin (parallel b-pleated sheet) [overhead,
Figure 7-26].
3-D Structure of Proteins 3
Secondary Structure, cont.
Collagen strand: This is a specialized structure occurring in only a particular family of fibrous proteins. It does not occur in globular proteins that I am aware of.
 Collagen triple helix. Note repeating sequence of -(gly-x-y)- where x is usually proline and y is usually hydroxyproline. (Fig 4.36) [overheads: 118&10, S; 4-10 to 12]
Non-repetitive secondary elements: Proteins can also have non-repetitive secondary structures which consist of a few residues in a turn or loop. Among these
are:
 beta-turns:
o Type I turns: Fig. 4.18, left [overhead 7.22, V&V] four amino acid
residues in a 180° turn, usually H-bonded between the carbonyl O of
the first residue and the amide N of the fourth. Proline is often the
second residue. [overhead, 7-22 V&V]
o Type II turns: Fig. 4.18 [overhead 7.22, V&V] four amino acid residues in a 180° turn, usually H-bonded between the carbonyl O of the
first residue and the amide N of the fourth. Glycine is most frequently
the third residue and proline is often the second residue. [overhead, 722 V&V]
 A partial turn of a 310 helix. Short sections of this helix often occur at the
ends of alpha-helixes as transitional elements.
Tertiary Structures
The Tertiary structure describes the overall folding of a single covalent structure.
 Lysozyme model [overhead, model]
As the number of known protein structures increased additional patterns became
obvious within the tertiary level of structure: Motifs & Domains.
Super Secondary structures (Motifs)
Recall the two classical structures based on the beta-strand:

Anti-parallel b-pleated sheet: strong, linear H-bonds spaced adjacent, then
R grp, then single, then R grp, then adjacent etc. (Fig 4.15b) [overhead 7-17
V&V, 5.19 P]
 Parallel b-sheet: evenly spaced, but slanted H-bonds (less stable), (Fig
4.15a) [overhead 5.19 P]
Let's next look at some of the other more common motifs found in globular proteins (Fig 4.19 of your text):
 Hairpin - b-strand-short loop-b-strand
 b-meander - an anti-parallel beta sheet with short connecting loops
 aa motif - two successive alpha-helixes with slightly inclined axis to give
better contact between side chains
 bab unit: alternate pattern of beta-strands and alpha-helixes
 Greek Key
 b-sandwich
Domains
Large proteins (>200 aa's) usually fold up in smaller pieces of 100-200 aa's called
domains. Recall that we define a Domain as an independent folding region in a
protein. Often defined by clefts in 3D structure giving globular elements connected
by "hinges" (single strand segments connecting the domains). Domains have the
advantages of speeding up the folding process (fold domains independently, then
assemble resultant folded domains - effectively processing folding of domains in
parallel). Another advantage of domain structure is that nature can take bits of
DNA specifying particular domains with particular functions and assemble them in
new combinations to get new activities (e.g. combine an ATP binding site and a
sugar binding site to give a sugar phosphorylating protein).
Example: IgG , domains, exons and evolution. [overheads: IgG/proteins; 7.23
MvH]
 IgG made up of four independently synthesized proteins, 2 heavy chains
with 4 domains each, and 2 light chains with 2 domains each.
o Domain types: b-meander [anti-parallel b-sheet], b-barrel. (Note that
Motifs and Domains often use the same nomenclature, and indeed often overlap. Can in fact have Motif = Domain = Tertiary structure!)
 Domains correspond to exons of DNA (frequently, but not always the case)
o The domains are all apparently related through gene duplication in the
remote past.
 The active site of IgG (2/IgG) is made up between two domains, one from a
heavy chain and one from a light chain.
 When immune system is developing individual cells express single IgG
molecules made from randomly expressed heavy and light chains.
In a similar manner we see that many enzymes have active sites created between
two domains, often one domain binds one substrate while the second binds a second substrate.
Its as if these proteins were designed by taking "off-the-shelf" components, assembling them, and then over time (and generations) tuning the combination up.
3-D Structure of Proteins 4
Domains, cont.
Note that domains will have their own tertiary structures, made up of secondary
and frequently supersecondary elements. Domains can be categorized into four
main groups:
1. All alpha
2. All beta
3. alpha/beta (have alternating alpha and beta structures, such as in the betaalpha-beta motif)
4. alpha + beta (local clusters of alpha and beta in same chain with each cluster
consisting of contiguous primary structure).
Groups of motifs forming the core of the tertiary structures of domains are referred
to as Folds. (p 99) Over 600 folds have been discovered, with an expectation that
about 1,000 exist. (a bunch, but well below the infinite number possible!) Common
examples include (Fig 4.24) [overhead]:
 Parallel twisted sheet.
 Beta barrel.
 Alpha/Beta barrel.
 Parallel twisted sheet .
Folds/Motifs are often more highly conserved than sequences, and so are used
along with sequences to trace relatedness among molecules and thus organisms.
An example of conservation for a domain is seen in Cytochrome c as shown in
your text in Figure 4.21.
Quaternary Protein Structure
ternary (4°) structures (Fig. 4.25; overheads: MvH 6.26, Fig 25): Geometrically
specific associations of protein subunits; the spatial arrangement of protein subunits.
Folding Hierarchy Overview
Rationale for quaternary: There are a variety of advantages to large structures:
 Increasing the size of a protein allows better "fits" for catalysis and binding many weak bonds are needed to maintain specific structures.
 Can bring sequential active sites of metabolic pathways into close proximity.
 However, large peptides have some problems:
The process of folding slows tremendously with increasing size, thus
folding individual subunits, and assembling these subunits can greatly
enhance folding efficiency.
3
o Get about 1 error / 10 aa residues due to the precision of the translation of messenger RNA to protein. Thus need to keep residue number
down.
 Interacting subunits provide mechanisms for regulation.
Quaternary structures allows the assembly of large to extremely large structures.
o
Protein Folding
Primary structure specifies tertiary (& therefore quaternary) structure. This is
known from in vitro denaturation/renaturation studies of small proteins.
 Denaturation means to unfold to non-functional state, often achieve a "random coil" in solution,
 Renaturation means to return to the properly folded, natural, and functional
state.)
The classic study involved Ribonuclease: Reduce (break) -S-S- bonds, denature
with urea to random coil. Now can renature by gently removing denaturant (urea)
and oxidize -S-S- bonds. [overhead 5.41, P] Enzyme activity fully recovered. Xray diffraction image same! Note - no gremlins, no magic, done in "test tube."
Other small proteins, such as Myoglobin and proinsulin, fold up spontaneously in
the same manner as Ribonuclease. However, insulin fails to fold correctly, since a
peptide essential to folding has been cleaved off.
Accesory Folding Proteins. The ribonuclease renaturation-type experiment has
not been repeated with large proteins, which seem to require the participation of
"folding catalysts," the chaperones, to aid their folding.
Modul 4. Enzymes.
The nomenclature and classification of ferments. Frame and catalytic properties of ferments. Temperature effect, рН, concentration of ferment and substrate for speed of enzymatic reactions. Regulation of activity of ferments
Enzymes are found all around us, they are found in every plant and animal.
Any living organism needs enzymes for its functioning. All living being are controlled by chemical reactions. Chemical reactions that are involved in growth,
blood coagulation, healing, combating disease, breathing, digestion, reproduction,
and everything else are catalyzed by enzymes. Our body contains about 3,000 enzymes that are constantly regenerating, repairing and protecting us.
Enzymes are powerhouses that are able to perform variety of functions in the
human body. Enzymes are wondrous chemicals of nature. Enzymes are used in
supplement form in medical arena. Although our bodies can make most of the enzymes, our body can wreak havoc the body's enzyme system and cause enzyme
depletion due to poor diet, illness, injury and genetics.
Enzymes Definition
Enzymes are large biomolecules that are responsible for many chemical reactions that are necessary to sustain life. Enzyme is a protein molecule and are biological catalysts. Enzymes increase the rate of the reaction. Enzymes are specific,
they function with only one reactant to produce specific products. Enzymes have a
three-dimensional structure and they utilize organic molecules like biotin and inorganic molecules like metal ions (magnesium ions) for assistance in catalysis.
Substrate is the reactant in an enzyme catalyzed reaction. The portion of the
molecule that is responsible for catalytic action of enzyme is the active site.
Characteristics of Enzymes
Characteristics of enzymes are as follows:
 Enzymes possess great catalytic power.
 Enzymes are highy specific.
 Enzymes show varying degree of specificities.
 Absolute specificity where the enzymes react specifically with only one substrate.
 Stereo specificity is where the enzymes can detect the different optical isomers and react to only one type of isomer.
 Reaction specific enzymes, these enzymes as the name suggests reacts to
specific reactions only.
 Group specific enzymes are those that catalyze a group of substances that
contain specific substances.
 The enzyme activity can be controlled but the activity of the catalysts can
not be controlled.
 All enzymes are proteins.
 Like the proteins, enzymes can be coagulated by alcohol, heat, concentrated
acids and alkaline reagents.
 At higher temperatures the rate of the reaction is faster.
 The rate of the reaction invovlving an enzyme is high at the optimum temperature.
 Enzymes have an optimum pH range within which the enzymes function is
at its peak.
 If the substrate shows deviations larger than the optimum temperature or pH,
required by the enzyme to work, the enzymes do not function such conditions.
 Increase in the concentration of the reactants, and substrate the rate of the
reaction increase until the enzyme will become saturated with the substrate;
increase in the amount of enzyme, increases the rate of the reaction.
 Inorganic substances known as activators increase the activity of the enzyme.
 Inhibitors are substances that decrease the activity of the enzyme or inactivate it.



Competitive inhibitors are substances that reversibly bind to the active site
of the enzyme, hence blocking the substrate from binding to the enzyme.
Incompetitive inhibitors are substances that bind to any site of the enzyme
other than the active site, making the enzyme less active or inactive.
Irreversible inhibitors are substances that from bonds with enzymes making
them inactive.
Enzyme Classification
The current system of nomenclature of enzymes uses the name of the substrate or the type of the reaction involved, and ends with "-ase". Example:'Maltase'substrate is maltose. 'Hydrolases'- reaction type is hydrolysis reaction.
Classification of enzymes
Enzymes are classified based on the reactions they catalyze into 6
groups: Oxidoreductases, transferases, hydrolases, lyases, isomearses, ligases.
Oxidoreductases - Oxidoreductase are the enzymes that catalyze oxidationreduction reactions. These emzymes are important as these reactions are responsible for the production of heat and energy.
Transferases - Transferases are the enzymes that catalyze reactions where
transfer of functional group between two substrates takes place.
Hydrolases - Hydrolases are also known as hydrolytic enzymes, they catalyze the hydrolysis reactions of carbohydrates, proteins and esters.
Lyases - Lyases are enzymes that catlayze the reaction invvolving the removal of groups from substrates by processes other than hydrolysis by the formation of double bonds.
Isomerases - Isomerases are enzymes that catalyze the reactions where interconversion of cis-trans isomers is involved.
Ligases - Ligases are also known as synthases, these are the enzymes that
catalyze the reactions where coupling of two compounds is involved with the
breaking of pyrophosphate bonds.
Structure of Enzymes
Enzymes are proteins, like the proteins the enzymes contain chains of amino
acids linked together. The characteristic of an enzyme is determined by the sequence of amino acid arrangement. When the bonds between the amino acid are
weak, they may be broken by conditions of high temperatures or high levels of acids. When these bonds are broken, the enzymes become nonfunctional. The enzymes that take part in the chemical reaction do not undergo permanent changes
and hence they remain unchanged to the end of the reaction.
Enzymes are highly selective, they catalyze specific reactions only. Enzymes have a part of a molecule where it just has the shape where only certain kind of substrate can bind to it, this site of activity is known as the 'active
site'. The molecules that react and bind to the enzyme is known as the 'substrate'.
Most of the enzymes consists of the protein and the non protein part called
the 'cofactor'. The proteins in the enzymes are usually globular proteins. The pro-
tein part of the enzymes are known 'apoenzyme', while the non-protein part is
known as the cofactor. Together the apoenzyme and cofactors are known as the
'holoenzyme'.
Cofactors may be of three types: prosthetic groups, activators and coenzymes.
Prosthetic groups are organic groups that are permanently bound to the enzyme. Example: Heme groups of cytochromes and bitotin group of acetyl-CoA
carboxylase.
Activators are cations- they are positively charged metal ions. Example: Fe cytochrome oxidase, CU - catalase, Zn - alcohol dehydrogenase, Mg - glucose - 6 phosphate, etc.
Coenzymes are organic molecules, usually vitamins or made from vitamins.
they are not bound permanently to the enzyme, but they combine with the enzymesubstrate complex temporarily. Example: FAD - Flavin Adenine Dinucleotide,
FMN - Flavin Mono Nucleotide, NAD - Nicotinamide Adenine Dinucleotide,
NADP - Nicotinamide Adenine Dinucleotide.
Function of Enzymes
Biological Functions of Enzymes:
 Enzymes perform a wide variety of functions in living organisms.
 They are major components in signal transduction and cell regulation, kinases and phosphatases help in this function.
 They take part in movement with the help of the protein myosin which aids
in muscle contraction.
 Also other ATPases in the cell membrane acts as ion pumps in active
transport mechanism.
 Enzymes present in the viruses are for infecting cell.
 Enzymes play a important role in the digestive activity of the enzymes.
 Amylases and proteases are enzyme sthat breakdown large molecules into
absorbable molecules.
 Variuos enzymes owrk together in a order forming metabolic pathways.
Example: Glycolysis.
Industrial Application of Enzymes:
 Food Processing - Amylases enzymes from fungi and plants are used in production of sugars from starch in making corn-syrup.
 Catalyze enzyme is used in breakdown of starch into sugar, and in baking
fermentation process of yeast raises the dough.
 Proteases enzyme help in manufacture of biscuits in lowering the protein
level.
 Baby foods - Trypsin enzyme is used in pre-digestion of baby foods.
 Brewing industry - Enzymes from barley are widely used in brewing industries.
 Amylases, glucanases, proteases, betaglucanases, arabinoxylases, amyloglucosidase, acetolactatedecarboxylases are used in prodcution of beer industries.










Fruit juices - Enzymes like cellulases,pectinases help are used in clarifying
fruit juices.
Dairy Industry - Renin is used inmanufacture of cheese. Lipases are used in
ripening blue-mold cheese. Lactases breaks down lactose to glucose and
galactose.
Meat Tenderizes - Papain is used to soften meat.
Starch Industry - Amylases, amyloglucosidases and glycoamylases converts
starch into glucose and syrups.
Glucose isomerases - production enhanced sweetening properties and lowering calorific values.
Paper industry - Enzymes like amylases, xylanases, cellulases and liginases
lower the viscosity, and removes lignin to soften paper.
Biofuel Industry - Enzymes like cellulases are used in breakdown of cellulose into sugars which can be fermented.
Biological detergent - proteases, amylases, lipases, cellulases, asist in removal of protein stains, oily stains and acts as fabric conditioners.
Rubber Industry - Catalase enzyme converts latex into foam rubber.
Molecular Biology - Restriction enzymes, DNA ligase and polymerases are
used in genetic engineering, pharmacology, agriculture, medicine, PCR
techniques, and are also important in forensic science.
Examples of Enzymes
A few well known examples of enzymes are as follows: Lipases, Amylases,
Maltases, Pepsin, Protease, Catalases, Maltase, Sucrase, Pepsin, Renin, Catalases,
A few examples of foods that are rich in enzymes:
Enzymes are available in the food we eat. Foods that are canned, or processed food like irradiation,drying, and freezing make the foods enzyme dead. Refined foods are void of any sort of nutrition. Food that is whole, uncooked and unpasteurized milk will provide enough enzymes. There are two basic ways to increase enzyme intake. First is to eat more fresh foods, cooking tends to kill enzymes. Raw fruits and vegetables are a good source of enzymes. Fermented food
like yoghurt, intake improves body's enzyme status. The other way to increase enzyme status of the body is by intake of enzyme supplements.
Here is a list of foods rich in enzymes - Apples, apricots, asparagus, avocado, banana, beans, beets, broccoli, cabbage, carrots, celery, cherries, cucumber,
figs, garlic, ginger, grapes, green barley grass,kiwi fruit, etc.
Modul 5. Vitamins
The definition, constitution and classification of vitamins and their role
in enzymatic reactions and in exchange processes.
Vitamins are natural substances found in plants and animals and known as
Essential nutrients for human beings. The name vitamin is obtained from "vital
amines" as it was originally thought that these substances were all amines. Human
body uses these substances to stay healthy and support its many functions. There
are
two
types
of
vitamins:
water-soluble
and
fat-soluble.
The body needs vitamins to stay healthy and a varied diet usually gives you all the
vitamins you need. Vitamins do not provide energy (calories) directly, but they do
help regulate energy-producing processes. With the exception of vitamin D and K,
vitamins cannot be synthesized by the human body and must be obtained from the
diet. Vitamins have to come from food because they are not manufactured or
formed by the body.
There are several roles for vitamins and trace minerals in diseases:
o
Deficiencies of vitamins and minerals may be caused by disease
states such as mal absorption;
o
Deficiency and excess of vitamins and minerals can cause disease in and of themselves (e.g., vitamin A intoxication and liver disease);
o
Vitamins and minerals in high doses may be used as drugs (e.g.,
niacin for hypercholesterolemia).
Vitamins are essential for the normal growth and development of a multicellular organism. The developing fetus requires certain vitamins and minerals to
be present at certain times. If there is serious deficiency in one or more of these nutrients, a child may develop a deficiency disease. Deficiencies of vitamins are classified as either primary or secondary.
o
Primary Deficiency: A primary deficiency occurs when you do
not get enough of the vitamin in the food you eat.
o
Secondary Deficiency: A secondary deficiency may be due to
an underlying disorder that prevents or limits the absorption or use of the
vitamin.
Types of Vitamins
Vitamins, one of the most essential nutrients required by the body and can
be broadly classified into two main categories i.e., water-soluble vitamins and fatsoluble vitamins.
Water-Soluble Vitamins
B-complex Vitamins
Eight of the water-soluble vitamins are known as the vitamin B-complex group:
thiamin (vitamin B1), riboflavin (vitamin B2), niacin (vitamin B3), vitamin B6
(pyridoxine), folate (folic acid), vitamin B12, biotin and pantothenic acid. The B
vitamins are widely distributed in foods,and their influence is felt in many parts of
the body. They function as coenzymes that help the body obtain energy from food.
The B vitamins are also important for normal appetite, good vision, and healthy
skin, nervous system, and red blood cell formation.
Thiamin: Vitamin B1
What is Thiamin. Thiamin, or vitamin B1, helps to release energy from foods,
promotes normal appetite, and is important in maintaining proper nervous system
function.
Food Sources for Thiamin. Sources include peas, pork, liver, and legumes. Most
commonly, thiamin is found in whole grains and fortified grain products such as
cereal, and enriched products like bread, pasta, rice, and tortillas. The process of
enrichment adds back nutrients that are lost when grains are processed. Among the
nutrients added during the enrichment process are thiamin (B1), niacin (B3), riboflavin (B2), folate and iron.
Thiamin Deficiency. Under-consumption of thiamin is rare in the United States due
to wide availability of enriched grain products. However, low calorie diets as well
as diets high in refined and processed carbohydrates may place one at risk for thiamin deficiency. Alcoholics are especially prone to thiamin deficiency because excess alcohol consumption often replaces food or meals. Symptoms of thiamin deficiency include: mental confusion, muscle weakness, wasting, water retention
(edema), impaired growth, and the disease known as beriberi. Thiamin deficiency
is currently not a problem in the United States.
Too much Thiamin. No problems with overconsumption are known for thiamin.
Riboflavin: Vitamin B2
What is Riboflavin. Riboflavin, or vitamin B2, helps to release energy from foods,
promotes good vision, and healthy skin. It also helps to convert the amino acid
tryptophan (which makes up protein) into niacin.
Food Sources for Riboflavin. Sources include liver, eggs, dark green vegetables,
legumes, whole and enriched grain products, and milk. Ultraviolet light is known
to destroy riboflavin, which is why most milk is packaged in opaque containers instead of clear.
Riboflavin Deficiency. Under consumption of riboflavin is rare in the United
States. However, it has been known to occur with alcoholism, malignancy, hyperthyroidism, and in the elderly. Symptoms of deficiency include cracks at the corners of the mouth, dermatitis on nose and lips, light sensitivity, cataracts, and a
sore, red tongue.
Too much Riboflavin. No problems with overconsumption are known for riboflavin.
Niacin: Vitamin B3, Nicotinamide, Nicotinic Acid.
What is Niacin. Niacin, or vitamin B3, is involved in energy production, normal
enzyme function, digestion, promoting normal appetite, healthy skin, and nerves.
Food Sources for Niacin. Sources include liver, fish, poultry, meat, peanuts, whole
and enriched grain products.
Niacin Deficiency. Niacin deficiency is not a problem in the United States. However, it is known to occur with alcoholism, protein malnourishment, low calorie
diets, and diets high in refined carbohydrates. Pellagra is the disease state that occurs as a result of severe niacin deficiency. Symptoms include cramps, nausea,
mental confusion, and skin problems.
Too much Niacin. Consuming large doses of niacin supplements may cause flushed
skin, rashes, or liver damage. Over consumption of niacin is not a problem if it is
obtained through food.
Vitamin B6: Pyridoxine, Pyridoxal, Pyridoxamine
What is Vitamin B6. Vitamin B6, otherwise known as pyridoxine, pyridoxal or pyridoxamine, aids in protein metabolism and red blood cell formation. It is also involved in the body’s production of chemicals such as insulin and hemoglobin.
Food Sources for Vitamin B6. Sources include pork, meats, whole grains and cereals, legumes, and green, leafy vegetables.
Vitamin B6 Deficiency. Deficiency symptoms include skin disorders, dermatitis,
cracks at corners of mouth, anemia, kidney stones, and nausea. A vitamin B6 deficiency in infants can cause mental confusion.
Too much Vitamin B6. Over consumption is rare, but excess doses of vitamin B6
over time have been known to result in nerve damage.
Folate: Folic Acid, Folacin
What is Folate. Folate, also known as folic acid or folacin, aids in protein metabolism, promoting red blood cell formation, and lowering the risk for neural tube
birth defects. Folate may also play a role in controlling homocysteine levels, thus
reducing the risk for coronary heart disease.
Food Sources for Folate. Sources of folate include liver, kidney, dark green leafy
vegetables, meats, fish, whole grains, fortified grains and cereals, legumes, and citrus fruits. Not all whole grain products are fortified with folate. Check the nutrition
label to see if folic acid has been added.
Folate Deficiency. Folate deficiency affects cell growth and protein production,
which can lead to overall impaired growth. Deficiency symptoms also include
anemia and diarrhea. A folate deficiency in women who are pregnant or of child
bearing age may result in the delivery of a baby with neural tube defects such as
spina bifida.
Too much Folate. Over consumption of folate offers no known benefits, and may
mask B12 deficiency as well as interfere with some medications.
Vitamin B12: Cobalamin
What is B12. Vitamin B12, also known as cobalamin, aids in the building of genetic material, production of normal red blood cells, and maintenance of the nervous
system.
Food Sources for Vitamin B12. Vitamin B12 can only be found only in foods of
animal origin such as meats, liver, kidney, fish, eggs, milk and milk products, oysters, shellfish. Some fortified foods may contain vitamin B12.
Vitamin B12 Deficiency. Vitamin B12 deficiency most commonly affects strict
vegetarians (those who eat no animal products), infants of vegan mothers, and the
elderly. Symptoms of deficiency include anemia, fatigue, neurological disorders,
and degeneration of nerves resulting in numbness and tingling. In order to prevent
vitamin B12 deficiency, a dietary supplement should be taken. Some people develop a B12 deficiency because they cannot absorb the vitamin through their stomach
lining. This can be treated through vitamin B12 injections.
Too much Vitamin B12. No problems with overconsumption of vitamin B12 are
known.
Biotin
What is Biotin. Biotin helps release energy from carbohydrates and aids in the metabolism of fats, proteins and carbohydrates from food.
Food Sources for Biotin. Sources of Biotin include liver, kidney, egg yolk, milk,
most fresh vegetables, yeast breads and cereals. Biotin is also made by intestinal
bacteria.
Biotin Deficiency. Biotin deficiency is uncommon under normal circumstances, but
symptoms include fatigue, loss of appetite, nausea, vomiting, depression, muscle
pains, heart abnormalities and anemia.
Too much Biotin. No problems with overconsumption are known for Biotin.
Pantothenic Acid
What is Pantothenic Acid. Pantothenic Acid is involved in energy production, and
aids in the formation of hormones and the metabolism of fats, proteins, and carbohydrates from food.
Food Sources for Pantothenic Acid. Sources include liver, kidney, meats, egg yolk,
whole grains, and legumes. Pantothenic Acid is also made by intestinal bacteria.
Pantothenic Acid Deficiency. Pantothenic Acid deficiency is uncommon due to its
wide availability in most foods.
Too much Pantothenic Acid. No problems with overconsumption are known for
Pantothenic Acid. Rarely, diarrhea and water retention will occur with excessive
amounts.
Vitamin C: Ascorbic Acid, Ascorbate
What is Vitamin C
The body needs vitamin C, also known as ascorbic acid or ascorbate, to remain in
proper working condition. Vitamin C benefits the body by holding cells together
through collagen synthesis; collagen is a connective tissue that holds muscles,
bones, and other tissues together. Vitamin C also aids in wound healing, bone and
tooth formation, strengthening blood vessel walls, improving immune system function, increasing absorption and utilization of iron, and acting as an antioxidant.
Since our bodies cannot produce or store vitamin C, an adequate daily intake of
this nutrient is essential for optimum health. Vitamin C works with vitamin E as an
antioxidant, and plays a crucial role in neutralizing free radicals throughout the
body. An antioxidant can be a vitamin, mineral, or a carotenoid, present in foods,
that slows the oxidation process and acts to repair damage to cells of the body.
Studies suggest that vitamin C may reduce the risk of certain cancers, heart disease, and cataracts. Research continues to document the degree of these effects.
Food Sources for Vitamin C. Consuming vitamin C-rich foods is the best method
to ensure an adequate intake of this vitamin. While many common plant foods contain vitamin C, the best sources are citrus fruits. For example, one orange, a kiwi
fruit, 6 oz. of grapefruit juice or 1/3 cup of chopped sweet red pepper each supply
enough vitamin C for one day.
Vitamin C Deficiency. Although rare in the United States, severe vitamin C deficiency may result in the disease known as scurvy, causing a loss of collagen
strength throughout the body. Loss of collagen results in loose teeth, bleeding and
swollen gums, and improper wound healing. More commonly, vitamin C deficiency presents as a secondary deficiency in alcoholics, the elderly, and in smokers.
The following conditions have been shown to increase vitamin C requirements
(Table 1):
 Environmental stress, such as air and noise pollution
 Use of certain drugs, such as oral contraceptives
 Tissue healing of wounds

Growth (children from 0- 12 months, and pregnant women)
 Fever and infection
 Smoking.
Too Much Vitamin C. Despite being a water-soluble vitamin that the body excretes
when in excess, vitamin C overdoses have been shown to cause kidney stones,
gout, diarrhea, and rebound scurvy.
Fat-Soluble Vitamins
The fat-soluble vitamins, A, D, E, and K, are stored in the body for long periods of
time and generally pose a greater risk for toxicity when consumed in excess than
water-soluble vitamins. Eating a normal, well-balanced diet will not lead to toxicity in otherwise healthy individuals. However, taking vitamin supplements that contain megadoses of vitamins A, D, E and K may lead to toxicity. The body only
needs small amounts of any vitamin.
While diseases caused by a lack of fat-soluble vitamins are rare in the United
States, symptoms of mild deficiency can develop without adequate amounts of vitamins in the diet. Additionally, some health problems may decrease the absorption
of fat, and in turn, decrease the absorption of vitamins A, D, E and K. Consult a
medical professional about any potential health problems that may interfere with
vitamin absorption.
Vitamin A: Retinol
What is Vitamin A
Vitamin A, also called retinol, has many functions in the body. In addition to helping the eyes adjust to light changes, vitamin A plays an important role in bone
growth, tooth development, reproduction, cell division, gene expression, and regulation of the immune system. The skin, eyes, and mucous membranes of the mouth,
nose, throat and lungs depend on vitamin A to remain moist. Vitamin A is also an
important antioxidant that may play a role in the prevention of certain cancers.
Food Sources for Vitamin A
Eating a wide variety of foods is the best way to ensure that the body gets enough
vitamin A. The retinol, retinal, and retinoic acid forms of vitamin A are supplied
primarily by foods of animal origin such as dairy products, fish and liver. Some
foods of plant origin contain the antioxidant, betacarotene, which the body converts to vitamin A. Beta-carotene, comes from fruits and vegetables, especially
those that are orange or dark green in color. Vitamin A sources also include carrots, pumpkin, winter squash, dark green leafy vegetables and apricots, all of
which are rich in beta-carotene.
Compared to vitamin A, it takes twice the amount of carotene rich foods to meet
the body’s vitamin A requirements, so one may need to increase consumption of
carotene containing plant foods.
Recent studies indicate that vitamin A requirements may be increased due to hyperthyroidism, fever, infection, cold, and exposure to excessive amounts of sunlight. Those that consume excess alcohol or have renal disease should also increase
intake of vitamin A.
Vitamin A Deficiency
Vitamin A deficiency in the United States is rare, but the disease that results is
known as xerophthalmia. It most commonly occurs in developing nations usually
due to malnutrition. Since vitamin A is stored in the liver, it may take up to 2 years
for signs of deficiency to appear. Night blindness and very dry, rough skin may indicate a lack of vitamin A. Other signs of possible vitamin A deficiency include
decreased resistance to infections, faulty tooth development, and slower bone
growth.
Too much Vitamin A
In the United States, toxic or excess levels of vitamin A are more of a concern than
deficiencies. The Tolerable Upper Intake Level (UL) for adults is 3,000 mcg RAE
(Table 2). It would be difficult to reach this level consuming food alone, but some
multivitamin supplements contain high doses of vitamin A. If you take a multivitamin, check the label to be sure the majority of vitamin A provided is in the
form of beta-carotene, which appears to be safe. Symptoms of vitamin A toxicity
include dry, itchy skin, headache, nausea, and loss of appetite. Signs of severe
overuse over a short period of time include dizziness, blurred vision and slowed
growth. Vitamin A toxicity also can cause severe birth defects and may increase
the risk for hip fractures.
Vitamin D
What is Vitamin D
Vitamin D plays a critical role in the body’s use of calcium and phosphorous. It
works by increasing the amount of calcium absorbed from the small intestine, helping to form and maintain bones. Vitamin D benefits the body by playing a role in
immunity and controlling cell growth. Children especially need adequate amounts
of vitamin D to develop strong bones and healthy teeth.
Food Sources for Vitamin D
The primary food sources of vitamin D are milk and other dairy products fortified
with vitamin D. Vitamin D is also found in oily fish (e.g., herring, salmon and sardines) as well as in cod liver oil. In addition to the vitamin D provided by food, we
obtain vitamin D through our skin which produces vitamin D in response to sunlight.
Vitamin D Deficiency
Symptoms of vitamin D deficiency in growing children include rickets (long, soft
bowed legs) and flattening of the back of the skull. Vitamin D deficiency in adults
may result in osteomalacia (muscle and bone weakness), and osteoporosis (loss of
bone mass).
Recently published data introduces a concern that some adults and children may be
more prone to developing vitamin D deficiency due to an increase in sunscreen
use. In addition, those that live in inner cities, wear clothing that covers most of the
skin, or live in northern climates where little sun is seen in the winter are also
prone to vitamin D deficiency. Since most foods have very low vitamin D levels
(unless they are enriched) a deficiency may be more likely to develop without adequate exposure to sunlight. Adding fortified foods to the diet such as milk, and for
adults including a supplement, are effective at ensuring adequate vitamin D intake
and preventing low vitamin D levels.
Vitamin D deficiency has been associated with increased risk of common cancers,
autoimmune diseases, hypertension, and infectious disease. In the absence of adequate sun exposure, at least 800 to 1,000 IU of vitamin D3 may be needed to reach
the circulating level required to maximize vitamin D’s benefits.
Who is at Risk — These populations may require extra vitamin D in the form of
supplements or fortified foods:
 Exclusively breast-fed infants: Human milk only provides 25 IU of vitamin D
per liter. All breast-fed and partially breast-fed infants should be given a vitamin D supplement of 400 IU/day
 Dark Skin: Those with dark pigmented skin synthesize less vitamin D upon
exposure to sunlight compared to those with light pigmented skin.
 Elderly: This population has a reduced ability to synthesize vitamin D upon
exposure to sunlight, and is also more likely to stay indoors and wear sunscreen which blocks vitamin D synthesis.
 Covered and protected skin: Those that cover all of their skin with clothing
while outside, and those that wear sunscreen with an SPF factor of 8, block
most of the synthesis of vitamin D from sunlight.
 Disease: Fat malabsorption syndromes, inflammatory bowel disease (IBD),
and obesity are all known to result in a decreased ability to absorb and/or use
vitamin D in fat stores.
Vitamin E: Tocopherol
Vitamin E benefits the body by acting as an antioxidant, and protecting vitamins A
and C, red blood cells, and essential fatty acids from destruction. Research from
decades ago suggested that taking antioxidant supplements, vitamin E in particular,
might help prevent heart disease and cancer. However, newer findings indicate that
people who take antioxidant and vitamin E supplements are not better protected
against heart disease and cancer than non-supplement users. Many studies show a
link between regularly eating an antioxidant rich diet full of fruits and vegetables,
and a lower risk for heart disease, cancer, and several other diseases. Essentially,
recent research indicates that to receive the full benefits of antioxidants and phytonutrients in the diet, one should consume these compounds in the form of fruits and
vegetables, not as supplements.
Food Sources for Vitamin E
About 60 percent of vitamin E in the diet comes from vegetable oil (soybean, corn,
cottonseed, and safflower). This also includes products made with vegetable oil
(margarine and salad dressing). Vitamin E sources also include fruits and vegetables, grains, nuts (almonds and hazelnuts), seeds (sunflower) and fortified cereals.
Vitamin E Deficiency
Vitamin E deficiency is rare. Cases of vitamin E deficiency usually only occur in
premature infants and in those unable to absorb fats. Since vegetable oils are good
sources of vitamin E, people who excessively reduce their total dietary fat may not
get enough vitamin E.
Too much Vitamin E
The Tolerable Upper Intake Level (UL) for vitamin E is shown in Table 2. Vitamin
E obtained from food usually does not pose a risk for toxicity. Supplemental vita-
min E is not recommended due to lack of evidence supporting any added health
benefits. Megadoses of supplemental vitamin E may pose a hazard to people taking
blood-thinning medications such as Coumadin (also known as warfarin) and those
on statin drugs.
Vitamin K
What is Vitamin K
Vitamin K is naturally produced by the bacteria in the intestines, and plays an essential role in normal blood clotting, promoting bone health, and helping to produce proteins for blood, bones, and kidneys.
Food Sources for Vitamin K
Good food sources of vitamin K are green, leafy-vegetables such as turnip greens,
spinach, cauliflower, cabbage and broccoli, and certain vegetables oils including
soybean oil, cottonseed oil, canola oil and olive oil. Animal foods, in general, contain limited amounts of vitamin K.
Vitamin K Deficiency
Without sufficient amounts of vitamin K, hemorrhaging can occur. Vitamin K deficiency may appear in infants or in people who take anticoagulants, such as
Coumadin (warfarin), or antibiotic drugs. Newborn babies lack the intestinal bacteria to produce vitamin K and need a supplement for the first week. Those on anticoagulant drugs (blood thinners) may become vitamin K deficient, but should not
change their vitamin K intake without consulting a physician. People taking antibiotics may lack vitamin K temporarily because intestinal bacteria are sometimes
killed as a result of long-term use of antibiotics. Also, people with chronic diarrhea
may have problems absorbing sufficient amounts of vitamin K through the intestine and should consult their physician to determine if supplementation is necessary.
Too much Vitamin K
Although no Tolerable Upper Intake Level (UL) has been established for vitamin
K, excessive amounts can cause the breakdown of red blood cells and liver damage. People taking blood-thinning drugs or anticoagulants should moderate their
intake of foods with vitamin K, because excess vitamin K can alter blood clotting
times. Large doses of vitamin K are not advised.
Modul 6.Carbohydrates
Classification of carbohydrates and their most important reactions. Disaccharides and polysaccharides: lactose, maltose, sucrose, starch, glycogen, cellulose, quinine. The role of carbohydrates in a food.
CARBOHYDRATES
The carbohydrates, or sugars, are our third group of biomolecules. They are characterized by having a carbonyl carbon (aldehyde or ketone) and multiple hydroxyl
groups. The smallest sugars are thus the three carbon trioses, glyceraldehyde (aldotriose) and dihydroxyacetone (ketotriose).
Note that sugars occur in both D and L forms. As we shall see the natural sugars
are generally D.
CARBOHYDRATES, cont.
Note that sugars occur in both D and L forms. As we shall see the natural sugars
are generally D. Let's look at the two families, aldoses and ketoses. The important
aldoses (Figure 8.3, p 234) [overhead 9.4 P] include the five carbon aldopentose,
ribose:
which commonly occurs in the cyclic furanose form.The six carbon aldohexoses,
glucose, mannose, and galactose.
which commonly occur in the cyclic pyranose form (as shown for glucose) [glucose model], and the six carbon ketohexose, fructose.
which commonly occurs in a cyclic furanose form. The important ketoses include
dihydroxyacetone, D-Xylulose, D-Ribulose, and D-Fructose [overhead 9.7 P]Note
the relationship between the Fischer projections and the cyclic Haworth projections, using the example of glucose.
The ring is then sealed via a hemiacetal bond. [overhead 9.10 P] This would normally be quite unstable, however the closeness of the two reacting centers in the
same chain makes them poor leaving groups, thus the hemiacetal is in fact the stable form of the six carbon aldoses. Thus the expected aldehyde chemistry for glucose is not seen (glucose is stable to oxygen etc.).Note that if drawn in the proper
conformations (Figure 8.11, p 239), or if constructed as models it will be seen that
the chair conformation should be more stable. In addition, the beta configuration of
the hemiacetal -OH will be equatorial and should thus be preferred steriochemically as is in fact the case. Interestingly organisms can generally only use the alpha
form, so isomerases are provide to interchange the two.
An important reaction is the Lobry-de-Bruyn-van Ekenstein Transformation. This
base catalyzed reaction sequence interconverts three of the major hexoses, and will
be used later in understanding some isomerase enzyme mechanisms. The mechanism is symmetrical. You should finish the second half on your own.
DISACCHARIDES
Can link sugars via acetal bonds, known as glycosidic bonds.
There are four common disaccharides (Fig 8.20, p 244) [overhead 9.24, P]:
 maltose [ -D-Glucopyranosyl-(1,4)- -D-glucopyranose]
 cellobiose [ -D-Glucopyranosyl-(1,4)- -D-glucopyranose]
 lactose [beta-D-Galactopyranosyl-(1,4)-beta-D-glucopyranose], and
 sucrose [alpha-D-Glucopyranosyl-(1,2)-beta-D-fructofuranoside]
The first three are reducing sugars, that is they have "free" aldehyde groups,
whereas sucrose has both carbonyl groups tied up in the relatively stable glycosidic
bond. Maltose and fructose are joined in alpha-glycosidic bonds. In general the al-
pha-glycosidic bond is easily cleaved (it is less stable chemically and organisms
have enzymes to cleave it) whereas the beta-glycosidic bond is very difficult to
break down.
Thus for cellobiose, and more importantly, cellulose which is also linked by betabonds, essentially only bacteria can digest this bond.
So animals can't digest cellulose! You may ask, What about Cows and things?
Well they use bacteria. Cows for instance are basically walking fermentation tanks.
Cool biological examples of cellulose use by animals: Desert Iguana consume feces to maintain culture; Rabbits eat and reprocess first pass feces (soft) to take advantage of fermentation; Multiple stomachs in Ruminants; Ultimate symbiosis in
some termites: protozoans in gut have bacteria in gut, and use spirochetes as "cilia"
(rowers).
An exception for mammals is the ability of nursing animals to digest lactose.
Note that this ability is generaly lost at the age of weaning, at which time the
animal becomes lactose intolerant. POLYSACCHARIDES
Can have both homo- and heteropolysaccharides. We will focus on homopolysaccharides as most central, but will mention some heteropolysaccharides to illustrate
their functions. Homopolysaccharides have a single type of residue. Most common
polysaccharides contain glucose. Used for energy (food) storage (starches and glycogen) and structure (cellulose).
Starch (energy storage in plants). Two kinds
 Amylose: linear, alpha-1,4 glycosidic links. MW: 4,000-150,000. (Figure
8.22) [overhead 8.21 MvH] (Gives char. deep blue color with iodine due to
coiled complex enclosing iodine-color is lost with heating, returns as
cooled).
 Amylopectin: branched every 30 or so units - linear alpha-1,4 chain of 30
glu residues then alpha-1,6 branch point. of course get branches on branches
as well. MW: ->500,000. (Figure 8.23, 8.24) [overhead 10-18, V&V] Broken down by alpha-amylase (pancreas and salivary glands; random cleavage
of alpha-1,4 links) to give glucose and maltose; or beta-amylase (plants; hydrolyses from reducing ends to give maltose). When either of these enzymes
attack amylopectin they are blocked when they reach or are near a branch,
thus end up with a "limit dextrin."
Glycogen: animal starch. Just like amylopectin, but more highly branched (every
8-12 residues). This allows more free ends for more rapid breakdown-important in
animals.
STRUCTURAL POLYSACCHARIDES
Cellulose: beta-1,4 linkages, thus resistant to breakdown (including acid hydrolysis) as want for structure (don't want to digest self). Multiple, extended strands
come together as fibrils held together with H-bonds (Figure 8.25, 8.26) [overhead
10.15, V&V], laid down in cell wall in criss-cross pattern, glued together with polyalcohols (lignin).
Chitin: Serves similar role to cellulose, but in animals (crustaceans and insects),
fungi, and some algae. Homopolymer of N-acetyl-D-glucosamine. Like cellulose ,
it has beta-1,4 linkages, and is thus resistant to breakdown. (Figure 8.27)
Among the heteropolysaccharides are the glycans such as Hyaluronic acid, an alternating polysaccharide of D-glucuronic acid and N-acetyl-D-glucosamine; MW
to 5,000,000 (Figure 8.28) which serves as a lubricant in joints and is a component
of the vitreous humor. Again we see beta-1,4 linkages.
Also very important are the glycans conjugated to proteins and peptides to give
proteoglycans (Figure 8.29).
Modul 7.Lipids
Biomedical value of lipids. Frame and classificationof lipids. The constitution
and transport properties of cellular membranes.
LIPIDS
Recall the lipid definition: The portion of an organism which will partition into a
non-polar solvent.
 Types of Lipids: (FiFatty acids: long chain carboxylic acids. (Figure 9.2)
[overhead 11.2, P; 12-3, S].Three of the most common are:
o Palmitic acid: (You can view an on-line model by clicking the Palmitic acid button at this site.)



o
Stearic acid:
o
Oleic acid: C9-C10 (Delta9double bond; You can view an on-line
model by clicking the Oleic acid button at this site.)
Triacylglycerols: Three fatty acids esterified to glycerol. (Figure 9.5)
[overhead 11.6]
Glycerophospholipids: Two fatty acids and a phosphate esterified to glycerol. (Table 9.2; Figure 9.7) [overhead 11.8; Table 11.2, P; 11-4 V&V]:
Sphingolipids: (Figure 9.10) [overhead 11.16 P; 11-6, V&V]
 Cholesterol: (Figure 9.15) [overhead 11-9, V&V] (You can view an on-line
model by clicking the Cholesterol button at this site.)
Lipid Properties: An important consideration for lipids of all sorts is their fluidity. Thus membranes must be fluid enough to allow the diffusion of proteins,
transport processes etc. but not so fluid as to weaken the membranes structure. For
storage want fat to be fluid enough to flow to fill out body shape at normal operat-
ing temperatures. A number of strategies are used by organisms to adjust lipid fluidity:
 Fatty acid chain length: longer chains have higher melting points (less fluid
at a given temperature).
 Unsaturation: double bonds introduce a "kink" in the chain, harder to stack,
so less van der Waals contact and thus lower melting points (more fluid).
 Branched chains (bacterial only): Again, less van der Waals contact and thus
lower melting points (more fluid).
 Cholesterol: Its planar shape enables it to stiffen bilayers
Lipid Bilayers
Detergents & Micelles: Polar heads of detergents and soaps (such as long chain
fatty acids) tend to associate with polar solvents such as water, while non-polar
"tails" are excluded by water and are forced to associate with themselves making
globules known as micelles.
Lipid Bilayer: Figures 9.20 [overhead 11-12, V&V; 12-11]:
The lipid bilayer forms the core for the lipid bilayer membrane as seen in the Fluid
Mosaic Model of biological membranes.
gure 9.1) [overhead 11.1, P]
Modul 8. Hormones
All multicellular organisms produce hormones. Plant hormones are known
as phytohormones. Blood is the medium of transport of animal hormones. The
term hormones is derived from Greek word homao which means to excite. It was
first used by William M. Bayliss and Ernest H. Starling, both of the London University College, in the year 1904. They showed a chemical substance - secretin, secreted in the intestine can stimulate the action of a pancreatic secretion. These substances were known as 'chemical messengers'.
The tissues that produce the hormones are known as the effectors while
those tissues are that are influenced by the effectors are called as targets. Hormones are of low molecular weight and they diffuse readily. The effects brought
about by the hormones are not permanent as the get readily oxidized.
Hormones Definition
Hormones definition - Hormones are organic chemical substances released
by a cell, or organ or a gland or any body part of plants and animals that functions
in the regulation of physiological activities and to maintain homeostasis. The
chemical discharged from one part affects the cells in other part of the organism.
Hormones are released in very minute quantities. Minute quantity of hormones carry out functions evoking responses from the target organs or tissues. The target organs or tissues are adapted to the minute quantities of the hormones. Hormones
acts as chemical messengers that transport signal from one cell to the other.
Hormones are transmitted to their target organs in the blood stream after
they are discharged from the glands secreting them. Cells express a specific receptor molecule to the hormone molecules to which they respond. Endocrine secretions is the mode of discharge directly into the bloodstream.
Characteristics of Hormones
General characteristics of hormones are as follows:
 Hormones are secreted by endocrine cells.
 Hormones are chemical messengers.
 The are chemical signals that circulate in the body fluids.
 The hormones regulate the behavior of the target cells.
 Hormones, unlike enzymes do not catalyze any reaction.
 They are secreted only when needed, they are not stored prior to requirement.
 Hormones may be proteinaceous or non-proteinaceous in nature (aminoacids or steroids).
 The secretion of hormones is regulated by the nervous system through the
feed back effect.
 Hormones usually cause long term effects like change in behavior, growth,
etc.
 The hormones function is to stimulate or inhibit the target organs.
Classification of Hormones
Traditional Classification
Hormones are classified traditionally into three types:
Classical hormones - These hormones are secreted from the endocrine cells
into the interstitial fluid. These hormones diffuse into the bloodstream and are distributed to all body parts by the circulatory system.
Neurohormones - These hormones are synthesized by the neuroendocrine
cells and are secreted at the nerve terminals. They are transported around the body
through the blood vessels, into which they were diffused.
Local hormones - These hormones are secreted into the interstitial fluid and
they act locally in two ways. Some hormones act on the neighboring cells and are
known as paracrine hormones and some hormones act on the cells from which they
were secreted, they are autocrine hormones.
Structural Classification
Hormones are structurally classified into four groups steroids, peptides, amino acids and fatty acids.
Steroid hormones are derived from cholesterol and are soluble in lipids.
The steroid hormones include the sex hormones and the hormones produced by the
adrenal gland. The sex hormone include androgens, estrogens and progesterone.
The adrenal hormones are mineralcorticosteroids and glucocorticosteroids.Steroids
hormones are important as they take part in important functions including water
balance, sexual development and stress response.
Amino acid derivative hormones - These hormones are derived from amino acids like tyrosine and tyroptophan. Two types of tyrosine derived hormones,
they are thyroid hormone and catecholamines.
Thyroid hormone is the most important as it regulates the develpment of organs and metabolism.
Catecholamines - Norepoinephrine and epinephrine are catecholamines.
They are stress hormones and ar neurotransmiiters.,
Tryptophan amino acid is the precursor of hormones like serotonin and melatonin. Serotonin regulates the movement of the intestines and is also associated
with mood and low levels of this hormones often result in depression.
Peptide hormones - These hormones are derived from peptides. Prohormones are the precursors of for peptide hormones. The prohormones are synthesized by the endoplasmic reticulum. Proper structural configuration is necessary
for their functioning. The peptide hormones are stored in the cell vesicle until there
is stimuli signals for their release into the blood stream. Examples of peptide hormones are TSH (thyroid stimulating hormone),insulin, prolactin, vassopressin.
Fatty acids derived hormones - Hormones derived from the fatty acids are
called eicosanoids, they are derived from arachidonic acid. These hormones are
produced by every cell in the body. They have important roles in the body including inflammation, blood pressure and blood clotting.
On the Basis of Mode of Action
Based on the mode of action hormones are classified into quick acting hormones and short acting hormones.
Quick acting hormones - These hormones initiate immediate response from
their target cells. These hormones have outer plasma membrane receptors on the
target cell, they are large sized. Example: Protein and amine hormones.
Short acting hormones - These hormones initiate a delayed response. These
hormones are small in size and they bind to the protein receptors present in the cytosol. Example: steroid hormones of reproductive organs and sdrenal cortex.
Functions of Hormones
Effects of hormones in mammals:
 They stimulate or inhibit growth.
 Hormones control the wake-up cycle and the circadian rhythms.
 Are responsible for mood swings.
 Induces or suppresses apoptosis.
 Activates or inhibits the immune system.
 Regulates metabolism.
 Prepares body for mating, fleeing, flighting and other activity.



Also prepares body for new mode of life like puberty, parenting and menopause.
It controls the activity of the reproductive cycle.
Controls hunger.
Functions of steroid hormones
The male sex hormone - testosterone is produced by the testis and is responsible for male characteristics like deep voice, facial hair during puberty.
Estradiol is the female sex hormone and is responsible for the development
of the secondary sexual characteristics in females. It is also participates in control
of menstrual cycle.
Progesterone is responsible for preparing the uterus for implantation of the
fertilized egg. It also plays a important role as birth control agents.
Hormones of Adrenal Gland
Mineral corticoids are made from different cells of the adrenal cortex. This
hormone is concerned with the water-salt balance in the body. It also regulates the
NaCl content of blood and causes excretion of potassium in urine.
Glucocorticoids are made by the adrenal cortex, these hormones modify certain metabolic reactions and has and anti-inflammatory effect.
Function of Peptide hormones
Peptide hormones like insulin take part in carbohydrate metabolism. It increases penetrations of cell membranes to facilitate entry of glucose. Hence decreasing the concentration of glucose int eh blood. Insulin is often referred as hypoglycemic factor.
Deficiency of insulin causes diabetes mellitus.
Functions of Amino Acid hormones
Thyroid hormones thyroxin and tri-iodothyroxine, they affect the general
metabolism of the body. The thyroid hormones are known as the pace setter of the
endocrine system.
List of Hormones
List of vertebrate hormones and the secretory organs is as follows:
S. No. Secretory Organ
Hormone
Estrogens
ß-estradiol
Ovary
Estriol
Estrone
1.
Androgens
Testosterone
Testes
Androsterone
DHEA
2.
Adrenal
Cortex Corticosteroids
Aldosterone
Cortisone
Cortisol
Corticosterone
Medulla
3.
Epinephrine
Norepinephrine
Progesterone
Corpus Leuteum
Relaxin
Islets of Langerhans
4.
ß-cells
Insulin
α- cells
Glucagon
Pars
5.
6.
7.
8.
distalis Thyrotropin
Corticotropin
Gonadotropins
FSH
LH
LTH
Somatotropin
Pars
inermedia Intermedins
Pars nervosa
Oxytocin
Vasopressin
Secretin
Gastrointestinal
Pancreozymin
Tract
Gastrin
Parathyroid
Parathormone
T3,
Thyroid
Thyroxine,
T4
Hormone Regulation
Controlling how much hormones are secreted and released from the cells is
hormones regulation. This regulatory activity is done in two mechanisms - the negative feedback mechanism and positive feedback mechanism and also by counter
regulatory hormones.
Negative Feedback Mechanism - Mostly hormones regulation is done by
negative feedback mechanism. In this mechanism hormone causes an effect, the
cells that make hormones detect this effect and the production of hormones is
ceased.
Example of negative feedback mechanism is with the hormone insulin. Insulin hormone is produced by ß-cells of pancreas. The release of insulin by the pancreas is the response to the consumption of glucose. Rise in the glucose levels in
blood, is detected by the pancreas and secretes insulin into the blood. Insulin increase the uptake of glucose in the target cells. Some of the glucose is used by the
cells and the other is converted and stored in the form of glycogen. The uptake of
glucose by the cells decreases glucose levels, this decrease is detected by the pancreas and as a response to the decrease in glucose levels,it stops secreting insulin
into the bloodstream. Decrease in insulin levels in the blood decreases glucose uptake by the cells. This negative feedback mechanism helps to maintain normal
blood glucose levels and prevents extremities.
Positive Feedback Mechanism - A few hormones are regulated through positive feedback mechanism. In the positive feedback mechanism the effect of hormones, make the gland secrete more hormones. This is the opposite of negative
feedback mechanism.
Example of positive feedback mechanism is the hormone that causes childbirth. The hormones is oxytocin which is made by the pituitary gland. The onset of
labor stretches the muscles in the cervix, the nerves here sends signals to the pituitary. This signal makes the pituitary release more oxytocin. The oxytocin hormone
causes the muscles of the uterus to contract which causes more stretching in the
cervix. This stretching causes even more secretion of oxytocin. The levels of oxytocin keeps rising until the contractions leads to childbirth.
Counter regulatory hormones - Activities in the body is sometimes controlled by two or more hormones.
Example of counter regulatory hormones - Glucose levels in blood is very
important to an organism. This is just not controlled by one hormone, other hormones also make glucose levels to increase or decrease. If the levels of glucose is
too low, the body release hormones that function opposite to the activity of insulin.
These hormones do not let the cells uptake of glucose from the blood. They make
the cells put back glucose into the blood. These hormones that work opposite to
other hormones are called counter regulatory hormones. Counter regulatory hormones for insulin are glucagon and epinephrine.
Modul 9. Nucleicacids
Biological value of nucleic acids. Deoxyribonucleicand ribonucleic acids.
Nucleotides. A constitution and functions in alive organisms. Transfer of ancestral features. Biosynthesis of protein. Processes of replication, a transcriptional and translation. A mutagenesis and hereditary diseases. Biotechnology
and gene engineering.
Modul 10. Metabolism
Concept about a metabolism and metabolic paths: the katabolism and
anabolism.
A metabolism and reception of biochemical energy.
Role of АТР in an energy exchange.
Proteins metabolism.
Carbohydrates metabolism.
Metabolism: chemical reactions in cells
Countless chemical reactions take place in cells and are responsible for all
the actions of organisms. Together, these reactions make up an organism's metabolism. The chemicals taking part in these reactions are
called metabolites.
In all reactions:
 chemical bonds in the reacting molecules are broken; this takes in energy
 new chemical bonds form to make the products; this gives out energy
When a chemical reaction takes place energy is either taken in or released.
This depends on the relative strengths of bonds being broken and bonds being
formed.
In an exergonic reaction, energy is released to the surroundings. The bonds
being formed are stronger than the bonds being broken.
In an endergonic reaction, energy is absorbed from the surroundings. The
bonds being formed are weaker than the bonds being broken.
You may also come across the terms exothermic and endothermic reactions.
These describe exergonic and endergonic reactions when the energy released or
absorbed is heat energy. In an exothermic reaction the temperature of the surroundings increases. In an endothermic reaction the temperature of the surroundings decreases.
Anabolism and catabolism
Two types of metabolic reactions take place in the cell: 'building up' (anabolism) and 'breaking down' (catabolism).
Anabolic reactions use up energy. They are endergonic. In an anabolic reaction small molecules join to make larger ones. For example, the following condensation reactions that occur in cells are anabolic:

amino
acids
join
together
to
make
dipeptides:
e.g. NH2CHRCOOH + NH2CHRCOOH
NH2CHRCONHCHRCOOH + H2O
and the process continues as large protein molecules are built up

small sugar molecules join together to make dissacharides:
e.g.
C6H12O6 +
C6H12O6
C12H22O11 +
and the process continues as large polysaccharide molecules are built up

glycerol
reacts
with
fatty
acids
to
make
e.g. CH2OHCH(OH)CH2OH + C17H35COOH
H2O
lipids:
CH2OHCH(OH)CH2OOCC17H35
and the process continues as the trigyleride is produced via similar reactions with
the other two hydroxyl groups of the glycerol molecule

during photosynthesis carbon dioxide and water are used to
produce
glucose
and
oxygen:
e.g. 6CO2 + 6H2O
C6H12O6 + 6O2
Catabolic reactions give out energy. They are exergonic. In a catabolic reaction large molecules are broken down into smaller ones. For example, the reverse
of the condensation reactions described above, i.e. hydrolysis reactions, are catabolic.

A simple example of a catabolic reaction that occurs in cells is
the decomposition of hydrogen peroxide into water and oxygen:
2H2O2
2H2O + O2

The conversion of glucose during respiration to produce carbon
dioxide
and
water
is
another
common
example:
C6H12O6 + 6O2
6CO2 + 6H2O
How chemical reactions occur
Chemical reactions that occur during metabolism are affected by temperature. Many animals maintain a constant temperature which results in relatively stable rates of metabolic reactions. Cold-blooded animals are particularly influenced
by the temperature of their environment - they are livelier when warm. In the cold
their metabolism slows dramatically, and this is why some cold-blooded animals
hibernate. Surgery is sometimes carried at low temperatures to slow the patient's
metabolic rate, for example, during operations on the heart or brain.
Molecules move and collide
Molecules are constantly moving. Their bonds vibrate and rotate. In gases,
liquids and solutions molecules move around, bumping into one another. When
molecules collide there is the possibility of a reaction taking place, but only if the
colliding molecules:

Have enough energy

Are aligned correctly
The more collisions there are between molecules with sufficient energy and
correct alignment, the faster the reaction takes place. This is called collision
theory.

The more molecules present, the faster the reaction. Therefore
reactions take place faster in concentrated solutions than in solutions that are
more dilute.

At high temperatures molecules have more energy than at lower
temperatures. Therefore collisions are more frequent and the likelihood of
the molecules having enough energy is greater. Consequently the rate of
chemical reactions increases with increasing temperature.
Activated complexes and activation energy
Some reactions take place in a single step. We can represent this using
an energy profile. An activated complex (or transition state) forms between reactant and product. This is not a 'real' substance in the sense that it can be isolated
and put in a test tube. But based on various pieces of experimental evidence it is
the chemist's model of how the reaction occurs. The energy 'hump' shows how
much energy reacting molecules must have for a 'successful' collision, i.e. one that
leads to reaction. The formation of an activated complex requires energy to bring
molecules together in the correct orientation. Therefore, it is always an endergonic
reaction. The energy required is called the activation energy (Ea).
It is more common for reactions between molecules to take place in a series
of consecutive steps. After each step a reaction intermediate forms. Unlike an activated complex this has a real existence. For each step an activated complex is
formed and there is an associated activation energy. The step with the highest activation energy is the rate-determining step in the reaction and controls how fast
the overall reaction is.
3. Practical work
Practical work № 1
Topic: Biochemistry subject
Qualitative analysis of fat-soluble vitamins
The purpose of the work: Consider the qualitative reactions tofat-soluble vitamins.
Qualitative reactions to vitamin A
Reaction with sulfuric acid
Principle of the method. Vitamin A gives color reaction with sulphuric acid. The
received bond is painted in blue-violet color. Chemism of reaction it is final on examinations.
Devices: Rack with test tubes(test tubes must be dry).
Reactants: Vitamin A solution in oil (industrial medicine). Сoncentrated sulfuric
acid.
Work description. In a test tube pour 3 drops of Solutio oleosa of vitamin A and
add 1 drop of sulphuric acid. In a test tube the blue-violet staining appears.
Qualitative reactions to vitamin D
Reaction with aniline
Principle of the method. Vitamin D at interaction with an aniline reactant when
warming is painted in red color.
Devices: Rack with test tubes.
Reactants: Aniline reactant (mix 15 parts of aniline and 1 part of the concentrated
hydrochloric acid).
Work description. In a dry test tube pour 1 ml of working fat, 1 ml of an admixture
of aniline with the hydrochloric acid, mix, carefully heat at constant stirring of a
dokipeniya and boil half-minute. In the presence of vitamin D turning yellow
emulsions gains green, and then red color in the beginning.
Qualitative reactions to vitamin Е
Reaction with nitric acid.
Work description: In 2 test tubes pour 2-3 drops of Solutio oleosa of vitamin E. To
the first test tube add 1-2 ml of water, to the second – the same amount of the concentrated nitric acid. Both test tubes heat on the boiling water bath of 10 min. In a
test tube with nitric acid the oil layer of vitamin is painted in orange-red color.
To make out results of works in table 1.
№
The name
of vitamin
А
D
Е
1.
2.
3.
1.
2.
3.
4.
5.
6.
7.
Reactants
The observed
coloration
Note
Table 1
Conclusions
Questions for self-preparation of students:
Subject of biological chemistry.
Use of achievements of biochemistry in livestock production and other fields
of agriculture.
General characteristic of vitamins. The mechanism of their action on a metabolism.
Classification of vitamins, their nomenclature.
Chemical nature of vitamins.
A concept about hypo, hyper, poly-avitaminoses. Reasons of their
emergence.
Characteristic of vitamins of the A,D,E,K group.
Practical work № 2
Topic: Amino acids: classification, structure, stereochemistry, physical and chemical properties and classification amino acids forming proteins.
Qualitative analysis of water-soluble vitamins
The purpose of the work: Consider the qualitative reactions to water-soluble vitamins.
Qualitative reaction to vitaminВ1
Principle of the method.In an alkaline environment thiazole cycle ofthiamine becomes unstable and easily cleaved to form reactive compounds - Thiamine-thiol.
Thiamine - thiol is reacted with p-diazobenzenesulfonic acid (diazoreagent) to
form a compound which colored to yellow- pink color.
Devices: Rack with test tubes.
Reactants: Fresh whole milk. Vitamin В1, 0,001%-aqueous solution.Sodium hydroxide, 5% solution.
Solution A (0.9 g a phenyl-sulfo acid dissolved in 9 ml of conc. HCl acid in a volumetric flask of 100 ml, and after some time is adjusted to the mark with water. In
a dark bottle solution is stored for a long time).
Solution B (sodium nitrite), 5% solution, freshly prepared).A freshly prepared diazo reagent (in a volumetric flask of 50 ml which immersed in a container with ice,
metered 1,5 ml of solution A and added in small portions to 7.5 mL of solution B.
Volume of the solution adjusted to the mark with cold water; after 15 minutes the
reagent is ready for use).
Work description. In test tube mixed 2 ml of sodium hydroxide solution and 3 ml
of a diazo reagent. Half of the resulting mixture is cast into another tube. Added to
the tube 1-2ml milk. In the tube develops yellow-pink color.
Qualitative reaction to vitamin В2
Principle of the method.Vitamin B2reduced by hydrogen, which is released by reacting metallic zinc with hydrochloric acid.
Devices: Rack with test tubes. Pipettes. Rubber stoppers for tubes.
Reactants: Vitamin B2, 0,005%-aqueous solution; hydrochloric acid,
diluted (1:1), metallic zinc (granular).
Work description. Pour intothe test tube 2 ml solution of vitamin B2, 1 ml of hydrochloric acid and stirred, and cast into an equal volume of another tube. In one of
the tubes cast piece of zinc metal and both tubes are stoppered. In the tube with
zinc discoloration of solution occurs within 5-10 min.
Qualitative reaction to vitamin В6(РР)
Principle of the method. Under the action of sodium hydrosulfite is restored nicotinamide (nicotinic acid) to give 1,4 - dihydro pyridine derivative, a yellow colored.
Devices: Rack with test tubes.
Reactants: Nicotinic acid or nicotinic acid amide (powder). Sodium bicarbonate,
10% solution.Sodium hydrosulfite (Na2S2O4 * 2H2O), a 5% solution, freshly prepared.
Work description. Inthe test tube is placed a little vitamin B5 powder (nicotinic acid or nicotinamide), add 1.2 ml of sodium carbonate solution and after stirring, 2.1
ml of a solution of sodium hydrosulfite. The liquid in the tube becomes yellow.
Qualitative reaction to vitamin С.
Principle of the method. Ascorbic acid reduces iron in the complex ion - ferrocyanide, turning it into a hexacyanoferrate, painted in blue color (Prussian Blue).
Devices: Rack with test tubes.
Reactants: Potassium hexacyanoferrate (6К3[Fe(CN)6]), 1% solution; ferric chloride, 1% solution; Vitamin C, 0.5% solution.
Work description. Into the test tube pour 2.3 ml of water, poured into another test
tube 3.2 ml of a solution of vitamin C. In both tubes add a few drops of a potassium hexacyanoferrate solution and ferric chloride. In the tube with Vitamin C solution appears blue color.
The results write to the table 2.
№
1.
2.
3.
4.
The name
of vitamin
В1
В2
В5
С
Reactants
Theobservedcoloration
Note
Table 2
Conclusions
Questions for self-preparation of students:
1. Characteristicsof vitamins B:
-B2/ riboflavin/
- В12/ cobalamin/
-В3/ pantothenic acid/
- Вс/ folic acid/
-В5/ РР, nicotinamide/
- В15/ pangamic acid/
2. Characteristicsof vitamins С:
- С / ascorbic acid/
- Р/ flavone/
Practical work № 3
Topic: Primary structure of proteins. Secondary, tertiary and quaternary structures.
The general properties of enzymes – thermolability, specificity and influence рН
on activity of enzymes.
The purpose of the work: To study properties of enzymes.
А. Thermolability of enzymes.
Principle of the method. Influence of change of temperature of the external environment on activity of enzyme of amylase of saliva is investigated.
Devices: Support with test tubes; glass (50 ml); spirit-lamp; thermostat (37 ͦC); a
glass with ice.
Reactants: The diluted saliva (rinse a mouth with the distilled water, and then,
having gained 20-25 ml of water in a mouth, bring together her in a glass); sodium
chloride; 0,3% solution of chloride of sodium; the Lugol reactant
(приготовление:в 100 ml of water dissolve 20 g of iodide of potassium and 101 g
of iodine. For reaction with starch the received solution is dissolved with the distilled water 1:5).
Work description. In three test tubes pour 2-3 ml of the diluted saliva (amylase).
Saliva in a test tube 1 is boiled within 1-2 min. Then add 4-5 ml of starch to all test
tubes. Test tubes 1 and 3 put in the thermostat / 37 ͦ For 10 min. The test tube 3 is
immersed for 10 min. in ice. After the specified time add to all test tubes on 1 drop
of a reactant of Lugol. Results of experience enter 3 thermolabilities of enzymes in
the table and draw conclusions.
Thermolability of enzymes
№
Enzyme
1.
Amylase
2.
3.
Amylase
Amylase
Experimental
conditions
The
denatured
enzyme
Enzymatic
Enzymatic
Substrate
Incubation
Starch
10 min
Starch
Starch
10 min 37ͦ с
10 min 0ͦ с
Table 3
Coloring
with iodine
B. Influence of рН on activity of enzymes.
Principle of a method. Activity of enzyme of amylase of saliva at various values
of the environment is investigated.
Devices: A support with test tubes. Pipettes. Thermostat /37ͦ С/.
Reactants: Disubstituted format of sodium, 0,2 M solution/B/; buffer solutions (рН
5,0: To mix 515 ml of solution A from 485 ml of solution B; рН 6,8: To mix 772,5
ml of solution A from 27,5 ml of solution B). The diluted saliva (Starch, 1% solution. Lugol reactant).
Work description. In three test tubes flow 2-3 ml of buffer solutions with various
rn/5,0; 6,8; 8,0/. Add to all test tubes on 2-3 ml of the diluted saliva / amylase(s) on
4-5 ml of solution of starch, 10 min. in the thermostat / 37 ͦ With / mix and incubate. Then add to each test tube on 1 drop of a reactant of Lugol. Results of observation enter in table 4 showing influence рН to activity of amylase of saliva.
№ of test
tubes
1.
Enzyme
рН
Substrate
Incubation
Amylase
5,0
Starch
2.
Amylase
6,8
Starch
3.
Amylase
8,0
Starch
10 min. 37ͦ
С
10 min. 37ͦ
С
10 min. 37ͦ
С
Table 4
Coloring
with iodine
C. Specificity of enzymes.
Principle of a method. It is investigated impacts of enzymes of amylase and sucrose on various substrata - starch and sucrose.
Devices: A support with test tubes. Pipettes. Thermostat (37 ͦ C). Spirit-lamp.
Reactants: Starch, 1% solution. Sucrose, 2% solution, the diluted saliva/. Sakharaza, solution / 10 g of yeast homogenize in 100 ml vody./. Lugol reactant/. Reactant of Felinga/preparation:
Solution 1. 31,65 g of not removed crystals of a copper vitriol (CuSO4·5 H2O)
dissolve in a flask on 500 ml at first in a small amount of the distilled water, and
after crystals bring to a tag.
Solution 2. 173 g of segnetovy salt (potassium – sodium – tartrate) Dissolve in a
flask on 500 ml in a small amount of the distilled water, add 62,5 g of the sodium
hydroxide dissolved in 100 ml of the distilled water well mix and bring to a tag.
Before the use solutions 1 and 2 mix in different volumes.
Work description. In test tubes 1 and 2 pour 4-5 ml of solution of starch, in test
tubes 3 and 4 – on 4-5 ml of solution of sucrose. To test tubes 1 and 3 add 2-3 ml
of the diluted saliva (amylase), to test tubes 2 and 4 on 2-3 ml of solution of sucrose. Contents of test tubes 10 min. in the thermostat mix and incubate (37· C).
Then 1 and 2 add to test tubes on 1 drop of a reactant of Lugol, in test tubes 3 and
4 – on 1-2 ml of a reactant of Felinga and heat. Observation is written down in the
table of the 5th specificity of enzymes of amylase and sucrose.
Specificity of enzymes.
№
Substrate
Enzyme
Incubation
1.
Starch
Amylase
10 min. 37ͦ
С
Coloring
with iodine
Table 5.
Reaction of
Feling
2.
Starch
Sakharaza
3.
Starch
Amylase
4.
Starch
Sakharaza
10 min. 37ͦ
С
10 min. 37ͦ
С
10 min. 37ͦ
С
Questions for self-preparation of students:
1. The general concept about the fermetakh.
2. Short story of development of the doctrine about enzymes.
3. Nomenclature and classification.
4. The most important methods of definition, receiving and cleaning.
5. Main properties of enzymes:
− Thermolability,
− Specificity,
− Influence рН Wednesdays,
− Colloidal properties.
Practical work № 4
Topic: Classification of proteins. The role of proteins in a food.
Mechanism of effect of enzymes. Influence of activators and inhibitors.
Purpose: To study the mechanism of effect of enzymes, their activators and inhibitors.
А. Influence of activators and inhibitors.
Principle of a method. In 2 test tubes pour 4-5 ml of solution of starch, 1 add 1-2
ml of solution of sulfate of copper to a test tube. In both test tubes flow 1-2 ml of
the diluted saliva, mix contents of test tubes and test tubes incubate 10 min. in the
thermostat /37ͦ С /. Then add to both test tubes on 1 drop of a reactant of Lugol.
Observations enter in table 6 showing influence of chloride of sodium and sulfate
of copper on activity of amylase.
Influence of activators and inhibitors.
№
Enzyme
Effector
Substrate
Incubation
1.
Amylase
NaСl
Starch
2.
Amylase
CuSO4
Starch
10 min. 37ͦ
С
10 min. 37ͦ
С
Table 6.
Coloring
with iodine
B. Definition of activity of a catalase of blood.
Principle of a method. An indicator of activity of enzyme is releases of the molecular oxygen which is formed at hydrogen peroxide splitting by a blood catalase.
Devices: A support with test tubes. Pipettes. Spirit-lamps.
Reactants: Blood integral, tsitratny. Hydrogen peroxide, 3% solution.
Work description. In two test tubes pour 2-3 ml of the distilled water, add 1-2
drops of blood and 1/kontrol/heat contents of a test tube to boiling for enzyme destruction. In a test tube 2/experience / enzyme are active. Add 1 ml of solution of
peroxide of hydrogen to both test tubes. In a test tube 2 observe rough release of
oxygen. Results of experience enter in table 8.
Definition of activity of a catalase of blood.
№
Reactants:
Observed coloring
Table 8.
Conclusions
1
2
Questions for self-preparation of students:
1. Chemical nature and structure of enzymes:
− Coenzymes,
− Active centers,
− Substratny center,
− Allosterichesky center
2. Functions of enzymes and their localization in a cage (kompartmentalization).
3. Multifermental systems. Isoenzymes.
4. The factors influencing the speed of enzymatic reactions:
− Concentration of a substratum and enzyme,
− Activators and inhibitors,
− Konkuretny and noncompetitive inhibitors,
− nature of packaging of polypeptide chains.
5. Mechanism of effects of enzymes:
− Theory of the intermediate relations,
− the Adsorptive theory.
Practical work № 5
Topic: The nomenclature and classification of ferments. Frame and catalytic properties of ferments.
Qualitative reactions to insulin and adrenaline.
Purpose: To study qualitative reactions of hormones.
А. Qualitative reactions to insulin
Biuret reaction.
Principle of a method. In alkaline solution at copper sulfate addition such substances as a biuret, an oxalamide, polypeptides and proteins, form the complex
salts painted in violet color.
Devices: A support with test tubes. Pipettes. Spirit-lamps.
Reactants: Insulin, solution / patent drug of insulin in ampoules with concentration
of 40 PIECES/ml dilute with a double amount of water. Sodium hydroxide, 10%
solution. Copper sulfate, 1% solution.
Work description. Add the peer volume of solution of sodium hydroxide and 1-2
drops of copper sulfate solution to 1-2 ml of solution of insulin. In a test tube the
violet staining appears.
Reactions with the concentrated nitric acid.
Devices: A support with test tubes. Pipettes. Spirit-lamps.
Reactants: Insulin, solution / patent drug of insulin in ampoules with concentration
of 40 PIECES/ml dilute with double amount of water/. The concentrated nitric acid.
Work description. In a test tube pour 1 ml of the concentrated nitric acid and strictly on a wall flow 1 ml of solution of insulin on drops. On the section of two liquids
accurately expressed ring is formed. To write down the observed phenomena in
table 9.
Table 9.
№
Reactants
Observed coloring Principle of the
method
B. Reaction of adrenaline with iodine
Principle of the method. In case of adrenaline solution heating with iodine the
adrenaline oxidation products painted in red color are formed.
Devices: A support with test tubes. Pipettes. Spirit-lamps.
Reactants: Adrenaline, solution / patent medicine of adrenaline in ampoules with
concentration 1:1000 dilute with double amount of water/. Iodine, 0,1 N spirtovy
solution.
Work description. In one test tube pour 1-3 ml of water, and in another - 1-2 ml of
solution of adrenaline. Add 2 drops of solution of iodine to both test tubes and
slightly warm up. In a test tube with adrenaline red coloring appears.
To enter results in table 10.
Reaction of adrenaline with iodine.
Table 10.
№
Reactants
Observed coloring Principle of the
method
C. Reaction of adrenaline with chloric iron.
Principle of a method. At addition to solution of adrenaline of solution of chloric
iron green coloring which emergence is caused by existence of the rest of pyrocatechin in an adrenaline molecule develops.
Devices: Devices: A support with test tubes. Pipettes. Spirit-lamps.
Reactants: Solution adrenaline. Chloric iron, 1% solution.
Work description. In one test tube pour 1-2 ml of water, in another-1-2 ml of solution of adrenaline. Add 2-3 drops of solution of chloric iron to both test tubes. In a
test tube with adrenaline green coloring appears. To issue observation in table 11.
Reaction of adrenaline with iron.
Table 11.
№
Reactants
Observed coloring Principle of the
method
Practical work № 6
Topic: Temperature effect, рН, concentration of ferment and substrate for speed of
enzymatic reactions. Regulation of activity of ferments.
Metabolism and energy. Biological oxidation.
Purpose: To get acquainted with method of studying of a metabolism and energy.
To reveal understanding students of the main questions of biological oxidation.
Questions for self-preparation of students:
1. A concept about a metabolism and energy.
2. A metabolism and energies - the single interconnected process of an organism.
3. Main stages of a metabolism.
4. Methods of studying of a metabolism on stages.
5. Biological oxidation:
− Bach's Theory,
− Palladin-Wiland's Theory,
− Modern representation about biological an okislenii.dykhatelny chain
− Oxidizing phosphorylation and free oxidation. Coefficient Р/О
− Overall effectiveness of accumulation of energy in case of an okislitelnomfosforilrovaniye.
Practical work № 7
Topic: The definition, constitution and classification of vitamins and their role in
enzymatic reactions and in exchange processes.
Metabolism of carbohydrates (anaerobic breakdown).
Purpose: To study the mechanism of intermediate exchange of carbohydrates. Refraktometric determination of dairy sugar in milk.
Principle of method. The method is based on dependence of index of refraction of
a beam on content in solution of dairy sugar. For this purpose from milk previously
emit serum in which determine index of refraction. Determination is made at a
fixed temperature.
Reactants and equipment:
1. Refractometer
2. A dropper from 4% solution of chloride calcium
3. The boiling bath
4. Piece test tubes-2
5. Pipettes on 5 ml
6. Funnel
7. Filters
Work description.
1. To measure 5 ml of the researched milk in a test tube, to add 5-6 drops of 4% of
solution of chloride calcium.
2. A test tube with mix to mix and deliver in the boiling bath for 10 min.
3. To take out a test tube from a bath and to cool it to room temperature.
4. To filter content of a test tube via the dry filter.
5. To apply a drop of the received filtrate on the lower prism of the refractometer.
6. To lower the upper prism of the refractometer and to make counting on a scale
of index of refraction.
7. Using the size of index of refraction, to determine percentage of dairy sugar by
the table.
Refractivity
1,3400
1,3401
Dairy sugar in %
3,52
3,57
Refractivity
1,3420
1,3421
Table 18
Dairy sugar in %
4,49
4,54
1,3402
1,3403
1,3404
1,3405
1,3406
1,3407
1,3408
1,3409
1,3410
1,3411
1,3412
1,3413
1,3414
1,3415
1,3416
1,3417
1,3418
1,3419
3,61
3,65
3,69
3,73
3,77
3,82
3,87
3,93
3,98
4,03
4,08
4,13
4,18
4,23
4,28
4,33
4,36
4,44
1,3422
1,3423
1,3424
1,3425
1,3426
1,3427
1,3428
1,3429
1,3430
1,3431
1,3432
1,3433
1,3434
1,3435
1,3436
1,3437
1,3438
1,3439
4,59
4,64
4,69
4,74
4,79
4,84
4,89
4,95
5,00
5,05
5,10
5,15
5,20
5,25
5,30
5,35
5,40
5,45
Questions for self-preparation of students:
1. Value of carbohydrates in a metabolism and energy of an animal organism.
2. Mono - di - polysaccharides, their structure, distribution and a role.
3. Digestion and absorption of carbohydrates in digestive tract of animals. Cellulose splitting.
4. Education and disintegration of a glycogen in muscles and a liver. Blood sugar.
5. Features of transformation of carbohydrates in a hem of ruminant.
6. Intermediate exchange of carbohydrates. Glycolysis and гликогенолиз. Similarity and distinction of these processes.
7. Anaerobic splitting of carbohydrates.
8. Similarity and distinction between glycolysis and spirit fermentation.
9. Balance of energy.
Practical work № 8
Topic: Classification of carbohydrates and their most important reactions. Disaccharides and polysaccharides: lactose, maltose, sucrose, starch, glycogen, cellulose, quinine. The role of carbohydrates in a food.
Metabolism of carbohydrates (aerobic breakdown).
Purpose: Study the mechanism of intermediate exchange of carbohydrates. Definition of glucose by Bertrán's method.
Principle of a method. The method is based on ability of free carbonyl group of
glucose to restore in alkaline solution oxide medi-CuO (II) in copper protoxide –
Cu2O (I).
Reactants and equipment:
1. Liquid of Felinga
2. Sulfate iron (II) solution
3. 0,1H KMnO4 solution
4. Hot water
5. A glass on 150 ml
6. Bunsen's flask
7. Shot's filter
8. Pipettes, measured cylinders
9. Water-jet pump or Kamovsky's pump
10. Rangette
Work description. In a flask with a capacity at 150 ml pour in 20 ml of the studied solution, then add 40 ml of liquid of Felinga. After that contents of a flask are
heated to boiling, and boiled exactly 3 minutes. To the received red deposit of protoxide of copper (I) filter a decantation via the glass filter in Bunsen's flask for suction. After that whenever possible, without losing a deposit of protoxide of copper
(I), wash it from alkali two-three times the hot distilled water. The washed-out deposit of protoxide of copper (I) is dissolved in small (5 ml) by quantities of
Fe2(SO4)3 and merge on the glass filter. The filter 2-3 times small wash out quantities (2-3 ml) of Fe2(SO4)3 and H2SO4, so that on the filter there is no copper protoxide left. Then the filter and a flask are washed out hot water. The received astvor of
protoxide of copper is titrut by 0,1 N KMnO4 solution before emergence of pink
coloring. Reaction proceeds on the equation:
Cu2O+ Fe2(SO4)3 + H2SO4 = 2 CuSO4 +2FeSO4 +H2O+ 10FeSO4 +2KMnO4+
8H2SO4 = 5Fe2(SO4)2+K2SO4+2MnSO4+8H2O
The number of ml 0,1H of KMnO4 solution, left by titration is multiplied on 6,36
and according to the table find the content of glucose in the studied solution.
Table 19
Definition of glucose by Bertrán's method
Glucose, mg
Copper, mg
Glucose, mg
Copper, mg
10
20,4
43
82,9
11
22,4
44
84,7
12
24,3
45
86,4
13
26,3
46
88,2
14
28,3
47
90,0
15
30,2
48
91,8
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
32,2
34,2
36,2
38,1
40,1
42,0
43,9
45,8
47,7
49,6
51,5
53,4
55,3
57,2
59,1
60,9
62,8
64,6
66,5
68,3
70,1
72,0
73,8
75,7
77,5
79,3
81,1
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
93,6
95,4
97,1
98,9
100,6
102,3
104,1
105,8
107,6
109,3
111,1
112,8
114,5
116,2
117,9
119,6
121,3
123,0
124,7
126,4
128,1
129,8
131,4
133,1
134,7
136,3
138,9
Questions for self-preparation of students:
1. Aerobic metabolism of lactic acid (a lactate to a piruvat and CoA acetyl.
2. Cycle trikarbonovykh of acids or Krebs's cycle, its biological value.
3. Balance of energy.
4. Regulation and pathology of carbohydrate exchange.
Practical work № 9
Topic: Biomedical value of lipids. Frame and classificationof lipids. The constitution and transport properties of cellular membranes.
Carbohydrates
A. The proof of the restoring ability at glucose and absence her at fructose.
a) Trommer's reaction
Place 0.5 ml of 0.5% solution of glucose and 6-8 drops in a big test tube 2n.naoh.
Then on drops add 0.2 N of CuSO4 solution, his dissolution won't stop yet. Carefully heat a test tube on a spirit-lamp. The blue not soluble deposit of hydrate of an
oxide of copper in water gradually passes in yellow, and then into a red deposit of
protoxide of copper. Do similar experience with 0.5% solution of fructose.
B. Proof of existence of a cyclic form of glucose in solution
Bring 6 drops of solution of glucose and 3 drops of solution of fuksinosernisty
acid in a test tube. To stir up contents.
The pink coloring characteristic of aldehydes doesn't arise. Explain the received
result.
C. Discovery of fructose (river of Selivanov).
In the first test tube bring 10 drops of 0.5% solution of fructose, in the second
test tube-10 of drops of 0.5% solution of glucose. Add to both test tubes in equal
volumes a freshly cooked reactant of Selivanov (0.5% solution of resorcin in 20%
hydrochloric acid). Carefully heat on a spirit-lamp. In a test tube with fructose
gradually there is a red coloring.
D. The proof of lack of the restoring ability of sucrose.
Bring 0.5 ml of 0.5% solution of sucrose and 6-8 drops in a test tube 2n.rastvora
a caustic natr. Then on drops add 0.2 N of solution of sulfate of copper (II), his dissolution won't stop yet. Will carefully heat a test tube on a spirit-lamp. The blue
deposit doesn't pass in yellow, and then – in red that proves lack of the restoring
ability at sucrose.
E. The proof of the restoring ability of lactose.
Bring 0.5 ml of 1% solution of lactose and 6-8 drops of 2 N of solution of a caustic natr in a test tube. Then on drops add 0.2n. solution of sulfate of copper (II) before full dissolution. Carefully heat on a spirit-lamp: the blue deposit of hydrate of
an oxide of copper passes into yellow-orange.
Practical work № 10
Topic: Classification of hormones
Metabolism of lipids
Purpose: To study transformation of lipids in an organism of animals. Definition
of iodic number of fat across Margoshes
Principle of the method. The Iodny number of fat is amount of iodine (in) which
can join 100 g of fat, it shows degree of nonsaturation of fat. The method is based
on reaction of accession of halogens to the remains of the nonlimiting fatty acids
which are a part of fat.
Reactants and equipment:
1. Fat
2. Ethanol
3. 5% spirit solution of iodine
4. 1% solution of starch
5. 0,1 N solution of hyposulphite of sodium (Na2S2O3)
6. A flask with a cover on 300 ml-2 of piece
7. The burette for titration-1 of piece.
8. The cylinder on 200 ml-1 of piece
9. A pipette on 10 ml-2 of piece
10. Water bath
11. Scales
Work description.
1. A fat hinge plate (0,1-,12 g) place a flask and dissolve in 10 ml of ethyl alcohol.
The flask is placed in a water bath (50-60 °C) for fat dissolution (to heat to an ischeznovaniye of drops of fat). At the same time blind experience becomes for what
in other flask pour only 10 ml of ethyl alcohol. Further, with this flask perform the
same operations that from skilled.
2. In both flasks add 5% of spirit solution of iodine on 5 ml, well stir up and flow
200 ml of warm water (20-30 °C). Flasks close a cover, stir up and leave to stand
for 5 min.
3. Excess of iodine in both flasks is ottitrovyvat by 0,1 N hyposulphite solution. As
the indicator add 10-12 drops of 1% of solution of starch. Titration will see to total
disappearance of blue coloring.
4. The iodic number of the studied fat is determined by a formula:
X = (and - in) * 0,0127*100/C, where
H-iodine number of fat,
and - the number of ml of hyposulphite which has gone for titration of blind experience
in - the number of ml of hyposulphate which has gone for titration of a flask with
fat
Fat S-hinge plate in, 0,0127 amount of iodine, corresponding 1 ml 0,1 N of solution of hyposulphite.
Questions for self-preparation of students:
1. Lipids. Neutral fats. Chemical structure, properties and role.
2. Saturated and nonsaturated fatty acids.
3. Zhiropodobny substances. Letsitina, kefalina, sterida, their value and formulas.
4. Value of fats in a metabolism and energy in an animal organism.
5. Digestion and absorption of fats in digestive tract.
6. A role of gastric acids in these processes.
7. Synthesis and adjournment of fats in fabrics of an animal organism.
8. Oxidizing disintegration of fats in fabrics:
− glycerin Oxidation,
− Oxidation of fatty acids.
9. Formation of ketone bodies.
10. Power balance.
11. Transformation of fosfatid, sterid and sterols.
Practical work № 11
Topic: Nucleicacids
Proteins. Physico-chemical properties.
Color reactions to proteins.
Principle of the method. Color reactions are reactions to structural elements of protein - peptide communication, on various amino acids in a protein molecule.
Devices: Test tubes. Water bath or spirit-lamp.
Reactants: Protein solution. NaOH, 20% rastvor.naoh, 30% solution. Sulfate copper, 1% solution. The concentrated nitric acid. Acetic lead 5% solution. Solution of
an ingidrin, 0,5%.
1. Biuretny reaction. This reaction is the general reaction to proteins. She points
to existence in a molecule of protein of peptide communications - CO-NH-. To 1
ml of solution of protein add the equal volume of 20% of NaOH and 3-4 drops of
1% of solution of sulfate CuSO4 copper, and mix. Liquid is painted in violet color.
Coloring of liquid depends on the number of peptide communications. All proteins
give this reaction.
2. Xanthoproteic reaction. By means of this reaction open at presence to a molecule of protein of cyclic amino acids - Thyrosinum, a tryptophan and phenylalanine.
To 1 ml of solution of protein add 0,5 ml конц. nitric acid, heat to boiling within
1-2 minutes. At the same time the formed deposit of the overthrown protein turns
yellow, / there is a reaction of a nitridation of a benzolnogokolets of aromatic amino acids to formation of nitrobonds of yellow color that indicates presence of cyclic amino acids/.
3. Reaction Fouling. By means of this reaction open presence at a protein molecule sulfur of the containing amino acids - Cysteinum, cystine and methionine.
Take 1-2 ml of acetic lead in a test tube and on drops add solution of diabrotic
sodium before dissolution of the formed deposit of hydrate of an oxide of lead.
Flow several (4-5) drops of protein and gradually heat an admixture. Observe
gradual darkening of solution.
4. Ninhydrin reaction. In a test tube would flow 5-6 drops of solution of solution
of an ingidrin. Bring to boiling. There is a violet coloring.
This reaction is characteristic of α-amino acids.
To issue conclusions on works in table 14.
Qualitative reactions to proteins.
Table 14.
№
Reactions
Observed coloring What groups open
in protein
1.
Biuretny
2.
Xanthoproteic
3.
Fouling
4.
Ninhydrin
Questions for self-preparation of students:
1. General characteristic of proteins.
2. Main functions of proteins.
3. Physical and chemical properties of proteins.
4. Methods of allocation and purification of proteins.
Practical work № 12
Topic: Concept about a metabolism and metabolic paths
Metabolism of proteins
Purpose: Determination of ammonia in blood with Nessler's reagent.
Principle of the method. The method is based on compound of ammonia with sulfuric acid from education sulfate ammonit which with Nessler's reagent gives yellow coloration. Intensity of a painting of solution depends on concentration of
ammonia in blood and is defined spectrophotometric.
Reagents and equipment
1. Nessler's reagent: 34,9 g of KI and 45,5 g of HgI2 dissolve in a small amount of
bidistillirovanny water, add 112 g of KOH and bring volume to 1 l water.
2. Carbonate and bicarbonate buffer рН =10,2:
And. 6M to add K2CO3-414g water to 500 ml
Б.3М To add KHCO3-150g water to 500 ml.
For preparation of the buffer it is necessary to mix solutions and/and/.
3. Penicillinic bottles.
4. Corks with polished sticks.
5. Rotor
6. Spectrophotometer
7. 1H sulfuric acid (27,2 ml konts.h2so4 to 1 l)
Operation course. To enter 0,5 ml of the buffer to penicillinic bottles рН =10,2, to
surge 1 ml of blood and to close immediately cork with a tugootshlifovanny stick
which end shall be moistened 1H H2SO4 (after the ground stick was dipped in a
glass with 1H sulfuric acid, it is necessary to remove excesses of acid percussion
about hour glass.
To insert bottles into holes of a rotor and to rotate 60 min. Then carefully to open
cork, to wash away the end of a stick of 5 ml of a reagent of Nesseler diluted (1 ml
of solution of Nesseler of +5 ml of a redistillator). To fill a basin and to fotometrirovat on the spectrophotometer in case of wavelength 420 nanometers, against
monitoring in which instead of 1 ml of blood water, 1 ml is poured. On a gage
curve find the content of ammonia in blood in % mg.
Questions for self-preparation of students:
1. Value of proteins in feeding of animals. Nitrogenous balance.
2. Biologicheskayapolnetsennost of proteins.
3. Znamenimy and irreplaceable amino acids.
4. Digestion of proteins in digestive tract.
5. Features of digestion of proteins at the poligastrichnykh of animals.
6. Products of rotting of proteins in intestines and a way of their neutralization.
7. Dezaminiraniye of amino acids, a vosstanovitelnoyeaminirovaniye of ketoacids
in tissues of animals.
8. Reactions reamination and decarboxylations of amino acids
9. Ammonia binding reaction.
10. Urea synthesis - M. V. Nentsky's theory.
11. Modern idea of urea biosynthesis (Krebs's theory)
Practical work № 13
Topic: Preparation of stand, posters about healthy lifestyles.
The exchange of water and minerals
Purpose: To study exchange of water and mineral substances in an organism of animals. To master a technique of definition of calcium in blood serum.
Principle of a method. The calcium which is contained in blood serum in the form
of soluble salts is besieged shchavelevokisly ammonium.
CaCl2+(NH4)2C2O4=CaC2O4+2NH4Cl
The formed deposit of shchavelevokisly calcium is washed ammonia solution from
excess of shchavelevokisly ammonium and dissolved in sulfuric acid.
CaC2O4+H2SO4= H2C2O4+CaSO4
The released oxalic acid is ottitrovyvat 0,01H KMnO4 solution
5H2C2O4+2KMnO4+3H2SO4= K2SO4+2MnO4+8H2O+10CO2
The quantity of KMnO4 which has gone for titration of oxalic acid is equivalent to
amount of calcium in serum.
Reactants and equipment:
1. (NH4)2C2O4 (saturated solution).
2. Blood serum.
3. 2% water solution of ammonia.
4. 0,01H KMnO4 solution.
5. The centrifuge on 1500 rpm.
6. Water bath.
7. Pipettes (2 ml, 1 ml, 5 ml).
8. Microburette (2 ml).
9. Glass stick.
Work course. The centrifugal test tube is poured by 2 ml of the distilled water, 2
ml of serum of blood, 1 ml of shchavelevokisly ammonium, stir up and leave in a
support for 15 min. In parallel put check experiment where instead of serum take 2
ml of the distilled water.
Further experience with serum of blood and check experiment are carried out
equally.
Centrifuge test tubes of 5 min. At 1500 rpm. At the bottom of a test tube with serum the dense deposit is besieged. Liquid of test tubes is poured out. In test tubes
pour 2% of NH4OH solution on 4 ml, stir up and again centrifuge 5 minutes.
Washing by ammonia is repeated by 3 times, merging liquid after each centrifugation. After draining of the last portion of ammonia, liquid from walls is deleted
with a strip of filter paper.
In both test tubes pour 2 ml 1H of H2SO4 solution, mix a deposit a thin glass stick
and immerse in a glass with the boiling water (carefully) for 2 min.
More hotly solution is titrut in a test tube 0,01H KMnO4 solution before not disappearing pink coloring.
Content of calcium is calculated on a formula:
X=(а-в)*0,2*100/2 мг% Са, where:
a - the number of ml of KMnO4 which has gone for titration of a test tube with serum.
b- the number of ml of KMnO4 which has gone for titration of a control test tube.
Questions for self-preparation of students:
1. Content of water in fabrics, her physiological role and exchange.
2. Pathology and regulation of water exchange.
3. Macro - and minerals, their biological role, exchange and its pathology, regulation.
4. Interrelation of mineral elements in a metabolism.
5. Biogeochemistry and problem of mineral food of animals.
LIST OF THEMES FOR STUDENT SELF-INDEPENDENT WORK
1. Biotechnology and gene engineering.
2. General characteristics of the vitamins.
3. Cell is a basic structural element of a live matter.
4. History of biochemistry.
РЕКОМЕНДАЦИИ ПО ВЫПОЛНЕНИЮ САМОСТОЯТЕЛЬНОЙ
РАБОТЫ И СРОКИ СДАЧИ ЗАДАНИЯ
Самостоятельные работы сдаются в соответствии с тематическим
планом и сроками, указанными в электронном журнале.
ТРЕБОВАНИЯ
К
ОФОРМЛЕНИЮ
РЕЗУЛЬТАТОВ
САМОСТОЯТЕЛЬНОЙ РАБОТЫ
ТЕЗИСНЫЙ КОНСПЕКТ
Описываются основные понятия, касающиеся темы, или делается план,
опираясь на который, студент формирует структуру устного ответа.
КРОССВОРД
На первом (титульном) листе пишутся тема (по которой пишется
кроссворд), название факультета студента, его группа, Ф.И.О. На второй
странице пишутся вопросы к кроссворду, отдельно по вертикали и по
горизонтали. Затем изображается непосредственно сам кроссворд. Последняя
страница должна содержать ответы (ключ) к вопросам: По вертикали: 1) ..., 2)
... и т.д.; По горизонтали: 1) .... 2) ... . Кроссворд должен включать не менее
30 слов.
ГЛОССАРИЙ
Титульный лист (см. оформление кроссворда). Затем в алфавитном
порядке пишутся основные понятия, относящиеся к данной теме, и даются
им четкие определения. Понятия должны быть выделены другим шрифтом
или подчеркнуты.
БУКЛЕТ
Титульный лист (см. оформление кроссворда). Например, по теме
«ООПТ Казахстана», буклет должен включать основные категории особоохраняемых природных территорий (заповедники, заказники, природные
парки, ботанические сады, памятники природы и др.), перечень всех
заповедников Республики Казахстан с указанием их места расположения,
года и цели создания. После чего подробно описывается один из
заповедников республики: место расположения заповедника, цель его
создания, климат, рельеф, виды животных и растений, занесенных в Красную
книгу. Буклет необходимо оформить соответствующим иллюстративным
материалом и обозначить список использованной литературы.
ТЕСТЫ
Титульный лист (см. оформление кроссворда). Тесты должны
содержать не менее 25-30 вопросов с предлагаемыми 5 вариантами ответов,
один из которых верный. Последняя страница должна содержать ответы
(ключ) к вопросам.
БЛОК-СХЕМА
Титульный лист (см. оформление кроссворда). Блок-схема
представляет собой схематично представленный материал, содержащий
краткие, емкие определения основных понятий с указанием взаимосвязей
между ними.
ПЛАН-КОНСПЕКТ ПРАКТИЧЕСКОГО ЗАНЯТИЯ
Титульный лист (см. оформление кроссворда). План-конспект должен
содержать не менее 5 заданий по данной теме, которые представляют собой
различные виды работ. Формулируется цель практической работы, план,
задания. Также предоставляется выполненный материал по данной
практической работе.
ЭССЕ
Изложение своих размышлений, соображений на актуальные
социально-экономические проблемы, в различных жанрах – критики,
публицистики и др.
КОЛЛОКВИУМ
Форма контроля самостоятельной работы обучающегося, проводимая в
виде собеседования по изученным разделам конкретной дисциплины с
целью определения качества освоения учебного материала.
КОНСПЕКТ
Последовательная фиксация информации, отобранной и обдуманной в
процессе чтения. Конспект бывают четырех типов: плановые (каждому
вопросу плана соответствует определенная часть конспекта); текстуальные
(состоящие из цитат); свободные (сочетающие выписки, цитаты, тезисы);
тематические (содержащие ответ на поставленный вопрос по нескольким
источникам).
КОНСПЕКТ-СХЕМА
Схематическая
запись
прочитанного
материала.
Наиболее
распространенными являются схемы «генеалогическое древо» и «паучок».
В схеме «генеалогическое древо» выделяются основные составляющие
наиболее сложного понятия, ключевые слова и т.п. и располагаются в
последовательности «сверху вниз» — от общего понятия к его частным
составляющим.
В схеме «паучок» название темы или вопроса записывается и
заключается в овал, который составляет «тело паучка». Затем
продумывается, какие понятия являются основными, их записывают на схеме
так, что они образуют «ножки паучка». Для того чтобы усилить устойчивость
«ножки», к ним присоединяют ключевые слова или фразы, которые служат
опорой для памяти. Составление конспектов-схем способствует не только
запоминанию материала. Такая работа развивает способность выделять самое
главное, существенное в учебном материале, классифицировать
информацию.
ОБЗОР ПО ТЕМЕ
Подготовка краткого письменного литературного обзора на 1-2
страницы по рекомендуемой теме с привлечением дополнительного
материала из печати, информационных ресурсов.
ПЛАН
«Скелет» текста, он компактно отражает последовательность
изложения материала. План как форма записи обычно значительно более
подробно передает содержание частей текста, чем оглавление книги или
подзаголовки статей. Форма записи в виде плана чрезвычайно важна для
восстановления в памяти содержания прочитанного, для развития навыка
четкого формулирования мыслей, умения вести другие виды записей. Удачно
составленный план говорит об умении анализировать текст, о степени
усвоения его содержания.
ПРЕЗЕНТАЦИЯ
Устное выступление по заданной проблеме с представлением основных
положений выступления в виде слайдов, видеороликов и другое.
РАСЧЕТНЫЕ ЗАДАЧИ
Решение расчетных задач с составлением алгоритма решения.
РЕЦЕНЗИЯ
Критический отзыв о конкретном произведении (статье), где автор
высказывается о качестве изложения материала, дает развернутую научно
обоснованную оценку ведущих идей рецензируемого источника.
Последовательно излагая свою позицию по ключевым вопросам исследуемой
проблемы, автор рецензии высказывает свое отношение, свои взгляды на
статью в целом и на главные ее положения.
РЕЗЮМЕ
Краткий вывод из сказанного, написанного.
СИТУАТИВНЫЕ ЗАДАНИЯ
Описание различных ситуаций и перечень заданий к ним.
СХЕМА АНАЛИТИЧЕСКОГО АНАЛИЗА
Составление плана проведения контрольного лабораторного занятия,
последовательно излагая ступени тех или иных действий с анализом
выводов.
ВЫПИСКА
В толковом словаре говорится: «Выписать - значит списать какоенибудь нужное, важное место из книги, журнала, сделать выборки» (от слова
«выбрать»). Вся сложность выписывания заключается как раз в умении найти
и выбрать нужное из одного или нескольких текстов. Выписки особенно
удобны, когда требуется собрать материал из разных источников.
ГРУППОВОЙ ПРОЕКТ
Задание по разработке проекта группой в количестве
3-5
обучающихся.
ДЕЛОВЫЕ ИГРЫ
Имитация любого процесса, направленная на выработку у
обучающихся навыков, необходимых для будущей профессиональной
деятельности, и требующая предварительной подготовки и самостоятельного
завершения.
ДЕРЕВО ЦЕЛЕЙ
Задание, предполагающее выбор проблемы, обозначение целей по
уровням; определение задания на всех уровнях и алгоритма их выполнения;
определение способов достижения целей; выбор измерителей качества
работы и формы контроля заданий.
ДОКЛАД
Развернутое устное сообщение на какую-либо тему, сделанное
публично, т. е. в присутствии слушателей, зрителей. Обычно в качестве тем
для докладов преподавателем предлагается тот материал учебного курса,
который не освещается в лекциях, а выносится на самостоятельное изучение
студентами. Поэтому доклады, сделанные студентами на семинарских
занятиях, с одной стороны, позволяют дополнить лекционный материал, а с
другой — дают преподавателю возможность оценить умения студентов
самостоятельно работать с учебным и научным материалом.
ИНДИВИДУАЛЬНЫЙ ПРОЕКТ
Задание по разработке исследовательского проекта по актуальной теме,
рассчитанное на наиболее подготовленных обучающихся.
6. List of recommended literature
6.1. Main Reading
6.1.1 D. Hames, N. Hooper Biochemistry - Taylor & Francis e-Library, 2006
6.1.2. Graff,S. Essays in Biochemistry/ S. Graff.- [репринтноеизд-е].- New York:
Book Renaissance, 2014 [1956].- 345 p.
6.1.3 Сеитов З.С. Биохимия. –Алматы: Агроуниверситет, 2000.-897 с.
6.1.4. Биологическая химия/под ред. Н.И. Ковалевской.-М.: Академия, 2009.255 с.
6.1.5 Berg,J.M. Biochemistry /Jeremy M. Berg, John L. Tymoczko, LubertStryer;
with Gregory J. Gatto, Jr.-International seventh edition/ J.M. Berg.- New York:
W.H. Freeman and Company, 2012.- 1098 с.
6.1.6 Voet,D. Biochemistry /Donald Voet, Judith G. Voet.-4-th edition/ D. Voet.UnitedStatesofAmerica: JohnWiley&Sons, Inc, 2011.- 1428 с.
6.1.7 Кайрханов К.К. Методические указания к лабораторным занятиям по
биохимии /издание третье, переработанное и дополненное/ Семипалатинск,
ЦНТИ, 2003.
6.2 Additional literature
6.2.1 Березов Т.Т., Коровкин Б.Ф., Биохимия. -Москва: 2000.-520 с.
6.2.2 Строев Е.А. Биологическая химия. -М.: 1986.-480 с.
6.2.3 Николаев А.Я. Биохимия. -Москва: 1989.-495 с.
6.2.4 Строев Е. А., Макарова А. А., Практикум по биологической химии. -М.:
«Высшая школа», 1986. -231с.
6.3 Internet-resources
6.3.1 Подред. Е.С. Северина. Биохимия: Учеб. для вузов, 2003. 779 с.
http://biochemistry.ru/biohimija_severina/B5873Content.html
6.3.2 Ridgway N., McLeod R., Vance J.E., Vance D. Biochemistry of Lipids, Lipoproteins and Membranes, 624 pages. Elsevier, 2012.
http://www.elsevier.com/books/biochemistry-of-lipids-lipoproteins-andmembranes/ridgway/978-0-444-53219-0
6.3.3 John W. Pelley, Edward F. Goljan Rapid Review Biochemistry//Chapter 3 –
Membrane biochemistry and Signal Transduction, 208 pages. Elsevier, 2011.
http://www.us.elsevierhealth.com/biochemistry/rapid-review-biochemistrypaperback/9780323068871/