DEPARTMENT OF MEDICAL BIOCHEMISTRY

FACULTY OF BASIC MEDICAL SCIENCES

EBONYI STATE UNIVERSITY

MBC 201: INTRODUCTORY TO MEDICAL BIOCHEMISTRY

(ANAT, PHYSIO, MLS, N.Sc)

DR. ORINYA O. F.

TOPIC: PRIMARY, SECONDARY AND COMPLEX CARBOHYDRATES, THEIR STRUCTURES AND

CLINICAL CORRELATION. BASIC PRINCIPLES OF TEST FOR CARBOHYDRATES

OBJECTIVES: At the end of this lecture you should be able to

1. Define carbohydrates
2. State different classifications of carbohydrates with examples
3. Describe simple and complex carbohydrates
4. Discuss the structures of complex carbohydrates
5. Explain the clinical correlations of complex carbohydrates
6. Explain the basic principles of common tests for carbohydrates

DEFINITION

Carbohydrates are biomolecules containing carbon, hydrogen and oxygen in the same ratio as
water (2:1) and are used as structural materials and for energy storage in living tissues. They
have the empirical formula Cn(H2O)n. However, not all carbohydrates conform to this
stoichiometric definition definition (uronic acids, deoxy-sugars such as fucose) and not all
compounds that conformed to the definition are classified as carbohydrates (eg. Formaldehyde,
acetic acid).

Therefore, carbohydrates are broadly defined as polyhydroxy aldehydes or ketones and their
derivatives or any substance that yield one of the compounds on hydrolysis.

FUNCTIONS OF CARBOHYDRATES

1. Carbohydrates are the main sources of energy in the body. Brain cells and RBCs are almost
wholly dependent on carbohydrates as the energy source. Energy production from
carbohydrates will be 4 k calories/g (16 k Joules/g).

2. Storage form of energy (starch and glycogen).

3. Excess carbohydrate is converted to fat.

4. Glycoproteins and glycolipids are components of cell membranes and receptors.

5. Structural basis of many organisms: Cellulose of plants; exoskeleton of insects, cell wall of
microorganisms, mucopolysaccharides as ground substance in higher organisms.

Carbohydrates are broadly classified into:

a. Monosaccharides: These are simple sugars that cannot be further hydrolysed into
smaller units. Eg. glucose, fructose, galactose, ribose
b. Disaccharides: These are formed when two monosaccharides are combined together by
a glycosidic linkage. The important disaccharides are Sucrose, Maltose, and Lactose.

c. Oligosaccharides: These are carbohydrate chains containing 3–10 sugar units. However,
some authors also include carbohydrates with up to 20 residues. Oligosaccharides can
be made of any sugar monomers, but most research has been carried out on
fructooligosaccharides (e.g., oligofructose) and galactooligosaccharides (e.g., raffinose,
human milk oligosaccharides). Few oligosaccharides are hydrolyzed and absorbed in the
small intestine (e.g., maltotriose), but nearly all enter the colon intact (nondigestible
oligosaccharides).
d. Polysaccharides: These are complex carbohydrates of high molecular weight formed by
polymerization of multiple monossacharide units, and/or their derivatives. They could
be homo- or heteroglycans.

Carbohydrate can also be generally grouped into simple carbohydrates and complex
carbohydrates.

SIMPLE CARBOHYDRATES

Simple carbohydrates are made of one or two sugar molecules eg glucose, fructose, maltose,
sucrose etc. Simple carbohydrates are digested quickly and spike blood sugar faster and
higher. They are common in Soft drinks, candy, cookies and other sweet snacks. These foods
are often made with white sugar, a form of processed sugar. Simple carbohydrates also are
found in natural sugars. Fruit, milk and vegetables contain natural sugars.

COMPLEX CARBOHYDRATES

Complex carbohydrates are made of multiple sugar molecules, called polysaccharides, and
include glycogen, starch and fiber etc. Complex carbohydrates represent an important energy
source for the body. They are digested more slowly and release glucose into the blood stream
more gradually. Polysaccharides contain hundreds and thousands of monosaccharides. Since
complex carbs are higher in fiber and digest more slowly. This also makes them more filling,
which means they’re a good option for weight control.They’re ideal for people with type 2
diabetes because they help manage blood sugar spikes after meals.

POLYSACCHARIDES:

POLYSACCHARIDES:

They are polymerized products of high molecular weight in which many monosaccharide units
or their derivatives are linked by glycosidic bond eg starch, glycogen, cellulose, chitin etc.

They may be homogenous or heterogenous in nature. The ealier is composed of a single type
of monosaccharide and is known as homopolysaccahrides or homoglycans e.g. starch, glycogen
and cellulose, while the later is made up of two or more different monosaccharides and is called
heteropolysaccharide or heteroglycans e.g. hyaluronic acid, chondroitin sulphate.

CHARACTERISTICS OF POYSACCHARIDES

1. Do not have sweet taste

2. Many are insoluble in water
3. Do not form crystals on desiccation
4. Can be extracted to form white powder
5. Inside the cell they are compact and osmotically inactive
6. Mostly contain C, H, O, where H and O are in the ratio (2:1) as seen in water

FUNCTIONS OF POYSACCHARIDES

1. Storage: they store energyin the form of sugars in living organisms eg glycogen, starch,
dextran
2. Structural function: they are the structural components of the cell walls of plants eg
cellulose, and exoskeleton in insects and fungi eg chitin
3. Serve as lubricants to connective tissues and joints eg chondroitin sulphate
Q. State other functions of polysaccharides

HOMOPOLYSACCHARIDES

1. STARCH: It is the reserve carbohydrate of plant kingdom and composed of amylose and
amylopectin. When starch is treated with boiling water, 10-20% is solubilized; this part is called
amylase(mw=400, 000 D). Amylose is made up of unbranched chain of glucose units with alpha-
1,4 glycosidic linkages. The insoluble part absorbs water and forms paste like gel; this is called
amylopectin. It is made up of glucose units, but highly branched with molecular weight more
than 1 million. The branching points are made by alpha-1,6 linkages.
HYDROYSIS OF STARCH

Hydrolysis by amylase: Salivary amylase and pancreatic amylase are alpha-amylases, which act
at random on alpha-1,4- glycosidic bonds to split starch into smaller units (dextrins), and finally
to alpha-maltose. Beta-amylases are of plant origin and converts starch to beta-maltose. They
act on amylose to split maltose units consecutively. Thus the enzyme starts its action from one
end. When beta-amylase acts on amylopectin, maltose units are liberated from the ends of the
branches of amylopectin, until the action of enzyme is blocked at the 1,6-glycosidic linkage. The
action of beta-amylase stops at branching points, leaving a large molecule, called limit dextrin
or residual dextrin. A debranching enzyme, α-1,6-glycosidase hydrolyzes the α-1,6- linkages at
the branches and thereby completes the hydrolysis.

When starch is hydrolysed by mild acid, smaller and smaller fragments are produced. Thus
hydrolysis for a short time produces amylodextrin which gives violet color with iodine and is
nonreducing. Further hydrolysis produces erythrodextrin which gives red color with iodine and
mild reduction of Benedict's solution. Later achrodextrins (no color with iodine, but reducing)
and on continued hydrolysis, maltose (no color with iodine, but reducing).

NB: Starch will form a blue colored complex with iodine; this color disappears on heating and
reappears when cooled. This is a sensitive test for starch. Starch is non reducing because it
has negligible number of free sugar groups.
2. GLYCOGEN: This is the glucose reserve in liver and muscle. Excess carbohydrates are stored
as glycogen. Glycogen is composed of glucose units joined by alpha-1,4 links in the straight
chains. It also has alpha-1,6 glycosidic linkages at the branching points. Branching points of
glycogen occurs at about every 8 to 12 glucose residues. The molecular weight is about 5
million. Innermost core of glycogen contains a primer protein, Glycogenin. Glycogen is more
branched and more compact than amylopectin. It gives red violet color with iodine.

3. CELLULOSE: This is the supporting tissues of plants. It is made up of glucose units joined with
beta-1,4 linkages. It has a straight line structure, with no branching points. Molecular weight is
about 2 to 5 million. Beta-1,4- bridges are hydrolysed by the enzyme cellulase. But this enzyme
is absent in human digestive system, hence cellulose cannot be digested by man. Herbivorous
animals have large caecum, which harbor bacteria. These bacteria can hydrolyse cellulose.
White ants (termites) also digest cellulose with the help of intestinal bacteria. Commercial
applications of cellulose includes, use as starting material to produce fibres, celluloids,
nitrocellulose and plastics.

4. INULIN: A long chain homoglycan composed of D-fructose units with repeating beta-1,2
linkages. It is the reserve carbohydrate present in various bulbs and tubers such as chicory,
dahlia, onion, garlic etc. It is clinically used to find renal clearance value and glomerular
filtration rate.

5. DEXTRANS: These are highly branched homopolymers of glucose units produced by

microorganisms. (yeast, bacteria). It has α- 1-4 linked glucose units and vary in their branch
points which may be -1,2-, -1,3-, -1,6- linkages in different species . They have molecular weight
1 million to 4 millions.

They do not easily go out of vascular compartment, so can be used for intravenous infusion as
plasma volume expander for treatment of hypovolemic shock.

6. CHITIN: It contains units of N-acetylglucosamine with beta-1,4 glycosidic linkages. It is a

structural polysaccharide present in exoskeletons of crustacea and insects.

HETEROPOLYSACCHARIDES

These are polysaccharides containing more than one type of sugar residues.

1. AGAR. It contains galactose, glucose and other sugars. It is dissolved in water at 100°C,
and upon cooling sets into a gel. Agar cannot be digested by bacteria and hence used
widely as a supporting agent to culture bacterial colonies. Agar is used as a supporting
medium for immuno-diffusion and immuno-electrophoresis. Agarose is made up of
 galactose combined with 3,6-anhydrogalactose units; it is used as matrix for
electrophoresis.

2. MUCOPOLYSACCHARIDES: Mucopolysaccharides or glycosaminoglycans (GAG) are polymers

of sugar derivatives carrying charges. They are heteropolysaccharides containing uronic acid
and amino sugars. Acetylated amino groups, sulfate and carboxyl groups are also present.
Because of the presence of these charged groups, they attract water molecules and so they
produce viscous solutions. They combine with proteins to form proteoglycans (mucoproteins).
They include:

A. Hyaluronic Acid: It is the most abundant mucopolysaccharides. They are found in the
ground substance of connective tissues, tendons, synovial fluid of the joint and vitreous humor
of the eye. It serves as a lubricant in joint cavities. It is composed of repeating units of N-Acetyl-
glucosamine → beta-1, 4-Glucuronic acid → beta-1-3-N-Acetyl glucosamine.

B. Heparin. Made up of repeating units of sulphated glucosamine, alpha-1, 4-L-iduronic acid.

Idose is the 5th epimer of glucose. Iduronic acid is the oxidized form of idose. Heparin is an
anticoagulant widely used when taking blood in vitro for clinical studies. It is also used in vivo in
suspected thrombo-embolic conditions to prevent intravascular coagulation. It activates
antithrombin III, which in turn inactivates thrombin, factor X and factor IX. It is found in liver,
lungs, spleen and monocytes. Commercial preparation of heparin is from animal lung tissues.
Heparan sulfate is also present in tissues.

C. Chondroitin Sulphate. It is one of the sulphated GAGs with repeating disaccharide units of β-
1,4-linked d-glucuronic acid (GlcA) and β-1,3-linked N-acetyl galactosamine (GalNAc)
(glucuronic acid → beta-1,3-N-acetyl galactosamine sulphate → β-1,4-). It is normally found in
the ground substance of connective tissues widely distributed in cartilage, bone, tendons,
cornea and skin. Chondroitin Sulphate has been found to possess antioxidant,
antiartherosclerosic, anticoagulant, antithrombosis, and negative immunogenic properties.
They have been one of the most widely used materials in the treatment of osteoarthritis.
Moreover, they play a predominant role in neural signal transduction and improve the activity
of central nervous system. Furthermore, tight packing of chondroitin sulfate with highly
charged sulfated groups could restrict the compression of cartilage and aid in cartilage
regeneration. Moreover, the ability of chondroitin sulfate to target the CD44 receptors on the
cancer cells made them to be explored in active targeting for cancer therapy. Etc.

Chondroitin-4-sulfate: R1 = H; R2 = SO3H; R3 = H. Chondroitin-6-sulfate: R1 = SO3H; R2, R3 = H

Based on the position of sulfur groups substituted on the chondroitin sulphate backbone,
different chondroitin sulphate has been designated (chondroitin sulphate a, b, c), chondroitin
sulphate b contains aldonic acid instead of glucuronic acid.

D. Keratan Sulphate: It is the only GAG which does not contain any uronic acid. The repeating
units are galactose and N-acetyl glucosamine in beta linkage. It is found in cornea and tendons.

E. Dermatan Sulphate: It contains L-iduronic acid and N-acetyl galactosamine in beta -1, 3
linkages. It is found in skin, blood vessels and heart valves.

NB: Lysosomal storage disorders due to defective degradation of GAG are called
Mucopolysaccharidoses.

AMINO SUGARS

When the hydroxyl groups of sugars are substituted with amino groups amino sugars are
formed. Generally, the amino group is added to the second carbon atom of hexoses. Amino
sugars have no reducing properties and do not produce osazones. Glucosamines are present in
hyaluronic acid, heparin and blood group substances. Galactosamines are found chondroitin of
cartilage, bone and tendons. Mannosamine is a constituent of glycoproteins.
 The amino group in the sugar may be further acetylated to produce N-acetylated sugars such
as N-acetyl-glucosamine (GluNac), N-acetyl-galactosamine (GalNac), etc. which are important
constituents of glycoproteins, mucopoly-saccharides and cell membrane antigens. The amino
sugars, N-acetyl glucosamine, N-acetyl galacto-samine and N-acetyl neuraminic acid are
synthesized from fructose-6-phosphate. The amino group is derived from amide group of
glutamine. The reaction is catalyzed by an amido transferase. This is irreversible; it is the rate
limiting step in amino sugar synthesis. The glucosamine-6-phosphate is then acetylated by
acetyl CoA to form N-acetyl glucosamine-6-phosphate. The active form of amino sugar, namely,
UDP-N-acetyl glucosamine is formed from N-acetyl glucosamine-1-phosphate, with the help of
a mutase and a pyrophosphorylase. It is then epimerized to galactose derivative.

Both these are used for synthesis of glycosaminoglycans and gangliosides. Mannose (6C) and
pyruvate (3C) combines to form 9C compound, N-acetyl neuraminic acid (NANA).

Importance of amino sugar • Amino sugars are components of glycolipid (ganglioside),

glycoprotein and proteoglycans (glycosaminoglycans). • Several antibiotics, e.g. erythromycin,
carbomycin contain amino sugar

GLYCOPROTEINS AND MUCOPROTEINS

When the carbohydrate chains are attached to a polypeptide chain it is called a proteoglycan. If
the carbohydrate content is less than 10%, it is generally named as a glycoprotein. If the
carbohydrate content is more than 10% it is a mucoprotein. They are seen in almost all tissues
and cell membranes. About 5% of the weight of the cell membrane is carbohydrates. The
carbohydrate containing uppermost layer of plasma membrane is known as glycocalyx.
Glycoproteins play a role as enzymes, hormones, transport proteins, structural proteins and
receptors. As the carbohydrate content is increased, viscosity is increased and solubility is
decreased. The carbohydrate content of mucins is generally more than 50%. Mucus consists of
5-10% of mucins. Mucins will form a protective barrier on epithelial surfaces. They are also
found in secretions of the gastrointestinal, respiratory and urogenital tract.

Glycophorin is the major membrane glycoprotein of erythrocytes. It is a transmembrane

protein. Carbohydrate chains are attached to the amino terminal portion, outside the cell
surface.

Glycoproteins have a protein backbone to which oligosaccharide groups are attached. The
oligosaccharide chains of glycoproteins are composed of varying numbers of the following
carbohydrate residues: Glucose (Glu); mannose (Man); galactose (Gal); Nacetyl glucosamine
(GluNAc); N-acetyl galactosamine (GalNAc); arabinose (Ara); Xylose (Xyl); L-fucose (Fuc) and N-
acetylneuraminic acid (NANA), also called sialic acid..
Depending on the sialic acid content, the electric charges are also varied. Glycoproteins are
widely distributed in tissues. All plasma proteins are glycoproteins (Q. list examples of
glycoproteins).

In glycoproteins, the carbohydrate groups are attached to the polypeptide chain by the
following types of linkages:

a. Through the amide group of asparagine to N-acetyl glucosamine (N-glycosidic linkages) (The
biosynthesis of N-linked glycoproteins involves Dolichol phosphate. Dolichol is a polyisoprenol,
containing about 20 repeating isoprenoid units). Dolichol is activated to Dol-P-P-GluNAc; this is
the starting point, over which other carbohydrate units are added. Tunicamycin specifically
inhibits Nglycosylation by inhibiting the transferase which adds GluNAc to dolichol-phosphate.

b. Through hydroxyl group of serine, threonine, hydroxylysine and hydroxyproline to N-acetyl

glucosamine or galactose or xylose (beta-O-glycosidic linkages). Glycosylation occurs in golgi
bodies and only proteins that are glycosylated and properly folded are exported.

c. Linked to carboxyl terminal amino acid of the protein via phosphoryl-ethanolamine, which is
linked to a phosphatidyl inositol, which in turn is linked to a glucosamine in the glycan group.
This is called glycosyl phosphatidyl inositol (GPI) linkage. Some proteins are anchored to the
plasma membrane by glycosyl phosphatidyl inositol (GPI) linked proteins.

Enzymatic addition of specific carbohydrate unit on the protein is called glycosylation, while
nonenzymatic spontaneous addition is called glycation. Proteins with identical amino acid
sequences, but having different oligosaccharide sequences are called glycoforms.

NB: Major constituents of prokaryotic (bacterial) cells are heteropolysaccharides, consisting of

repeating units of N-acetyl muramic acid (NAM) and N-acetyl glucosamine (NAG). This
polysaccharide provides mechanical strength. Synthesis of this complex polysaccharide is
blocked by penicillin. This inhibition is responsible for the bactericidal action of penicillin

Aquasomes: They are one of the most recently developed delivery systems that are making a
niche as the peptide/protein carriers. These are nanoparticulate carrier systems. They comprise
the central solid nanocrystalline core coated with polyhydroxy oligomers onto which
biochemically active molecules are adsorbed. The solid core provides the structural stability.
The carbohydrate coating acts as dehydroprotectant and stabilizes the biochemically active
molecules. Aquasomes are being proposed as a carrier system for delivery of peptide based
pharmaceuticals. The delivery system has been successfully utilized for the delivery of insulin,
hemoglobin and various antigens. Oral delivery of enzymes like serratiopeptidase has also been
achieved.
BLOOD GROUP SUBSTANCES (ANTIGENS)

The RBC membrane contains several antigenic substances, based on which persons are
classified into different blood groups. More than 160 different antigens are known. Of these
ABO system and Lewis system are known to involve glycoproteins. ABO system is associated
with 3 blood group substances on RBCs designated A, B and H antigens. The H antigen is the
basic structure. It has the following structure: Fucose—Gal—GalNAc—Protein. The RBCs
carrying such H antigen are denoted as blood group O. A and B antigens differ from this in
having additional sugar residues.

A antigen: – Fucose—Gal—GalNAc—Protein

GalNAc

B antigen: – Fucose—Gal—GalNAc—Protein

Gal

The H locus codes for fucosyl transferase. In a person belonging to blood group A, N-acetyl-
galactosaminyl transferase is present. In group B person, galactosyl transferase is seen. Lack of
both leads to blood group O, while AB persons have both enzymes.

NB: Blood group A contains A-antigen with anti-B antibodies in the plasma. Blood group B
contains B-antigen with anti-A antibodies in the plasma. Blood group AB contains both
antigens (i.e AB-antigen) with neither A nor B antibodies in the plasma. It is called the universal
acceptor/recipient in transfusion because it lacks the antibodies and does not form
agglutination with other blood groups. Blood group O has no antigen but anti-A and antibodies.
It is termed the universal donor.

BASIC PRINCIPLES OF TESTS FOR CARBOHYDRATES

1. Glucose Oxidase Test

Principle: Glucose is oxidized by oxygen in the presence of glucose oxidase to hydrogen
peroxide and gluconic acid. A second enzyme, peroxidase converts H2O2 to H2O and
O2. The molecular oxygen oxidizes the chromogen to a coloured complex whose
intensity is directly proportional to the amount of glucose present in the test solution. A
chromogen is a compound that becomes coloured on acceptance of oxygen eg O-
toulidine, benzidine, 4-amino atipyrine.

2. Benedict's Test
Principle: The Principle is that when a solution of reducing sugar is heated with Benedict's
Reagent, the alkaline sodium carbonate converts the sugar into enediol* and this enediol
further reduces the cupric ions of the Reagent into cuprous ions. The resulting precipitate of
cuprous oxide is brick-red in a colour that confirms the presence of reducing sugar. Lactose,
maltose, and glucose give a positive reaction to this Test. Benedict reagents:-Anhydrous
Sodium Carbonate, Sodium Citrate (forms a complex with cupric ions so that they are not
reduced to cuprous ions during storage), Copper (II) Sulphate Pentahydrate (gives the cupric
ions) and Distilled water (solvent).

Reaction

3. Molisch's Test
Principle: It is a general Test for Carbohydrates. In Molisch's Test experiment, Carbohydrates
are reacted with Molisch's Reagent and concentrated sulphuric acid; carbohydrate dehydrates
to form furfural and its derivatives. The products further react with sulfonated alpha-naphthol
to give a purple coloured complex. All Carbohydrates, that is, monosaccharides, disaccharides,
and polysaccharides give a positive result for Molisch's Test. Molisch's Reagent consists of α-
naphthol dissolved in ethanol.

4. Iodine Test

Principle: The Test gives positive results only with polysaccharide (starch and Glycans. The
Principle is that on reacting with starch, Iodine gets trapped in the helical coils of the
polysaccharide chain via a coordinate complex**. Due to the complex formation, blue/black
colour is observed, which confirms the presence of starch. The blue/black colour disappears on
the addition of an alkali or heating; this is due to the uncoiling of the polysaccharide network
and release of the Iodine molecules.

5. Seliwanoff’s Test
Principle: This test is used to distinguish aldoses from ketoses. Ketoses undergo
dehydration to give furfural derivatives, which then condense with resorcinol to form a
red complex. Prolonged heating will hydrolyze disaccharides and other monosaccharides
will also eventually give color.

Reaction

6. Bial’s test
Principle: This test is useful in the determination of pentose sugars.
Reaction is due to formation of furfural in the acid medium which
condenses with orcinol in presence of ferric ions to give a blue-green
colored complex which is soluble in butyl alcohol.

Others include: Anthrone test for general Carbohydrates, Barfoed’s test

(distinguish between mono- and disacch) it depends on the time of
reaction, mucic test for galactose, Fehling’s test for reducing sugars,
picric acid test for reducing sugars etc.
Ref.

1. Vasudevan, D.M., Sreekumari, S., Vaidyanathan, K. (1994). Textbook of Biochemistry for

Medical Students. 6th edition. Jaypee Brothers Publishers (P) Limited.
2. Robert K. Murray, Kathleen M. Botham, Peter J. Kennelly, Victor W. Rodwell, P. Anthony
Weil (2012). Harper’s illustrated biochemistry 29th Edition. Mc Rraw Hill Medical
3. Nelson, D. L., and Cox, M. M. (2000). Lehninger Principles of biochemistry. 5th edition.
PAL GRAVE
4. https://www.sciencedirect.com/topics/biochemistry-genetics-and-molecular-biology/
amino-sugar Retrieved 2025
5. Xiong, S., Li, A., and Hou, D(2012). Progress in structural analysis of glycosaminoglycans
and their applications in biomaterials. African Journal of Biotechnology Vol. 11(19), pp.
4311-4316, Available online at http://www.academicjournals.org/AJB
6. Qaulitative test for Carbohydrates.
http://14.139.61.83/BioChemicalEstimations/qualitativetests_carbohydrates.htm
retrieved Feb 2025
7. Practical Manual in Medical Biochemistry by Diribe C.O., Elom S.O. and Edeogu, C.O.
Department of Medical Biochemistry, Ebonyi state University.