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FOOD ANALYSIS (MPA 104T)

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

Carbohydrates: classification and properties of food carbohydrates, General methods of analysis of food carbohydrates, Changes
in food carbohydrates during processing, Digestion, absorption and metabolism of carbohydrates, Dietary fibre, Crude fibre and
application of food carbohydrates.

Introduction
- Carbohydrates are good source of energy. Carbohydrates (polysaccharides) are long chains of sugars.
Monosaccharides are simple sugars that are composed of 3-7 carbon atoms.
- They have a free aldehyde or ketone group, which acts as reducing agents and are known as reducing sugars.
Disaccharides are made of two monosaccharides. The bonds shared between two monosaccharides are the
glycosidic bonds.
- Monosaccharides and disaccharides are sweet, crystalline and water-soluble substances. Polysaccharides are
polymers of monosaccharides. They are un-sweet and complex carbohydrates. They are insoluble in water and
are not in crystalline form.
- Example: glucose, fructose, sucrose, maltose, starch, cellulose etc.
- Carbohydrate is an organic compound, it comprises of only oxygen, carbon and hydrogen.
- The oxygen: hydrogen ratio is usually 2:1.
- The empirical formula being Cn(H2O)n .
- Carbohydrates are hydrates of carbon; technically they are polyhydroxy aldehydes and ketones.
- Carbohydrates are also known as saccharides; the word saccharide comes from Greek word sakkron which
means sugar.
Classification and nomenclature of carbohydrates
- The carbohydrates are divided into three major classes depending upon whether or not they undergo hydrolysis
and if they do, on the number of products formed.
1. Monosaccharides: The monosaccharides are polyhydroxy aldehydes or polyhydroxy ketones which cannot be
decomposed by hydrolysis to give simpler carbohydrates. e.g. Glucose, fructose, Galactose etc.
2. Oligosaccharides: The oligosaccharides (Oligo: few) are carbohydrates which yield a definite number (2-9) of
monosaccharide molecules on hydrolysis.
a) Disaccharides - Which yield two monosaccharides’ molecules on hydrolysis. Which have molecular formula is
C12H22O11.e.g. Sucrose, maltose etc

b) Trisaccharide’s - Which yield three monosaccharides’ molecules on hydrolysis and have molecular formula is
C18H32O16.

c) Tetrasaccharides - Which yield four monosaccharides’ molecules on hydrolysis and have molecular formula is
C22H42O21. eg: Stachyose [gal(α1→6)gal(α1→6)glu(α1↔2β)fru]
3. Polysaccharides: The carbohydrates which have higher molecular weight, which yield many monosaccharide
molecules on hydrolysis. E.g. Starch, glycogen, Dextrin, Cellulose etc.

- In general monosaccharides and oligosaccharides are crystalline solids, soluble in water and sweet to taste, they
are collectively known as sugars, the polysaccharides on the other hand are amorphous, insoluble in water and
tasteless, they are called non-sugars.

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

Different between monosaccharaides, oligosaccharides and Polysaccharides

Character Monosaccharaides Oligosaccharides Polysaccharides

No. of sugar molecules 1 2-9 More than 9

Glycoside bond Absent Present Present

Molecular Weight Low Moderate High

Taste Sweet Minimally sweet taste No taste

Solubility Soluble Soluble Insoluble

Nature Always reducing sugar May or may not be Always non reducing sugar

Glucose, fructose, Starch, Glycogen, Dextrin,

Example Sucrose, Maltose
Galactose Cellulose

Test for carbohydrate:

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Different between reducing and non-reducing sugar

Sl. No. Reducing sugar Non-reducing sugar
1 Such sugar bear a free aldehyde (-CHO) or These sugars do not have such groups
ketonic [RC(=O)R'] group
2 Reducing sugars have the capacity to reduce Non reducing sugars fail to reduce cupric ions of
cupric ions of benedicts or Fehling solution to Benedict’s or Fehling solution to cuprous ion
cuprous ion
3 Example: all monosaccharides and Example: Sucrose, Trehalose
disaccharides (like lactose, Melibiose,
Cellobiose, Gentiobiose)

Maltose and Cellobiose

Both maltose and cellobiose are composed of two glucose molecules. But in maltose the glucose molecules are in
the α configuration and joined with an α(1→4) link while in cellobiose they are in the β configuration and joined
with a β(1→4).

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Gentiobiose
It is a disaccharide composed of two units of D-
glucose joined with a β(1->6) linkage. Trehalose
It is also known as mycose or tremalose, is a natural alpha-
linked disaccharide formed by an α,α-1,1-glucoside bond
between two α-glucose units. In 1832, H.A.L. Wiggers
discovered trehalose in an ergot of rye

❖ Pentose Sugar:
• A pentose is a monosaccharide with five carbon atoms.
• Pentoses are organized into two groups.
i. Aldopentoses have an aldehyde functional group at position 1.
ii. Ketopentoses have a ketone functional group in position 2 or 3.
a) Aldopentoses: The aldopentoses have three chiral centers and therefore eight different stereoisomers are
possible.

b) Ketopentoses: The 2-ketopentoses have two chiral centers, and therefore four different stereoisomers are
possible. The 3ketopentoses are rare.

• The carbohydrates may also be classified as either reducing or non-reducing sugars. All those carbohydrates which
have ability to reduce Fehiling’s solution and Tollen’s reagents are referred to as reducing sugars, while others are
non-reducing sugars. All monosaccharides and disaccharides other than sucrose are reducing sugars.
• Carbohydrates are also said to be saccharides (Latin: Saccharon - Sugar).
• Note By: Synthetic sweetening agents

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

Polysaccharides
• Monosaccharides can undergo a series of condensation reactions, adding one unit after another to the chain until
very large molecules (polysaccharides) are formed. This is called condensation polymerisation, and the building
blocks are called monomers.
• The majority of carbohydrate material in nature occurs in the form of polysaccharides (also known as glycans).
Other molecules that contain polysaccharide structures are also referred to as polysaccharides.
- Homopolysaccharide. A polysaccharide whose monosaccharide units are identical.
- Heteropolysaccharide. A polysaccharide made up of different monosaccharides
• The most common constituent of polysaccharides is D-Glucose
• Homopolysaccharides composed of glucose are called glucans; Those composed of mannose are called mannans;
etc.
• Glycosidic linkages involve only one anomeric carbon (keto or aldehyde) per monosaccharide. However, there
are several potential hydroxyl acceptor groups on a given monosaccharide. Thus, polysaccharides can be
branched as well as linear.
- The characterization of polysaccharides includes their length, the type of monosaccharide units, the linkages
and branched nature
- Starch is a glucan that has two polysaccharide forms: one is a linear molecule with -1-4 linkages called
amylose. The other is a branched form of amylose, where the branches have -1-6 linkages, called
amylopectin.
• The properties of a polysaccharide molecule depend on:
i. Its length (though they are usually very long)
ii. The extent of any branching (addition of units to the side of the chain rather than one of its ends)
iii. Any folding which results in a more compact molecule
iv. Whether the chain is 'straight' or 'coiled'
• Starch, Glycogen and Cellulose are all polymers of glucose. They differ in the type of glucose present and the
bonds which link thru glucose monomers together, whereas Insulin is the polymer of glucose and fructose
• Starch and Glycogen are made from alpha-glucose. This is an isomer of glucose in which the hydroxyl (-OH)
group attached to carbon number 1 is below the plane of the ring. Whereas Cellulose are β-glucose monomer.
• The major form of storage polysaccharide in animals is glycogen (found in the liver and skeletal muscle).
• The most common storage polysaccharide in plants is starch.
• Yeast and bacteria make dextran as a storage polysaccharide. It is composed of -1-6 linkages of Dglucose, with
branched chains (1-2, 1-3 or 1-4 linkages between glucose units). Important component of dental plaque (from
bacteria).
Properties of Carbohydrates
- General properties of carbohydrates
Carbohydrates act as energy reserves, also stores fuels, and metabolic intermediates.
✓ Ribose and deoxyribose sugars forms the structural frame of the genetic material, RNA and DNA.
✓ Polysaccharides like cellulose are the structural elements in the cell walls of bacteria and plants.
✓ Carbohydrates are linked to proteins and lipids that play important roles in cell interactions.
✓ Carbohydrates are organic compounds; they are aldehydes or ketones with many hydroxyl groups.
- Physical Properties of Carbohydrates
✓ Stereoisomerism: Compound shaving same structural formula, but they differ in spatial
configuration. Example: Glucose has two isomers with respect to penultimate carbon atom. They are D-
glucose and L-glucose.
✓ Optical Activity: It is the rotation of plane polarized light forming (+) glucose and (-) glucose.
✓ Diastereoisomers: It the configurational changes with regard to C2, C3, or C4 in glucose. Example:
Mannose, galactose.
✓ Annomerism: It is the spatial configuration with respect to the first carbon atom in aldoses and second
carbon atom in ketoses.

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

- Chemical Properties of Carbohydrates

✓ Osazone formation with phenylhydrazine.
✓ Benedict’s test.
✓ Oxidation
✓ Reduction to alcohols
• Structure of Carbohydrates
- There are three types of structural representations of carbohydrates:
(i) Open chain structure.
(ii) Hemi-acetal structure.
(iii) Haworth structure.

Difference between Starch, Glycogen & Cellulose

Characters Starch Glycogen Cellulose
α-glucose
Amylose: (60 to 300 α-glucose
α-glucose β-glucose
Monomer Unit)
(6000 to 30,000 α-glucose Unit) (300 to 2500 β-glucose Unit)
Amylopectine: (300 to 6000 α-
glucose Unit)
Type of
α(1→4) glycosidic bond
bond α(1→4) and α(1→6) glycosidic
(Amylose) + α(1→4) and α(1→6) β(1→4) glycosidic bond
between bonds
glycosidic bond (Amylopectin)
monomers
Amylose is coiled unbranched Straight, long unbranched
Nature of Short many branched chains,
Amylopectin is long branched chains form H-bonds, with
chain some coiling
chains, some coiling adjacent chains
Occurrence In plants In animals and fungi In plants
Function Carbohydrate energy store Carbohydrate energy store Structural
General
Grains Small granules Fibres
form
Chemical
Test or Deep Blue-Violet color Reddish brown color Negative test
Iodine test

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Functions of Carbohydrates
✓ Carbohydrates are chief energy source, in many animals; they are instant source of energy. Glucose is broken
down by glycolysis/ kreb's cycle to yield ATP.
✓ Glucose is the source of storage of energy. It is stored as glycogen in animals and starch in plants.
✓ Stored carbohydrates act as energy source instead of proteins.
✓ Carbohydrates are intermediates in biosynthesis of fats and proteins.
✓ Carbohydrates aid in regulation of nerve tissue and are the energy source for brain.
✓ Carbohydrates get associated with lipids and proteins to form surface antigens, receptor molecules, vitamins
and antibiotics.
✓ They form structural and protective components, like in cell wall of plants and microorganisms.
✓ In animals they are important constituent of connective tissues.
✓ They participate in biological transport, cell-cell communication, and activation of growth factors.
✓ Carbohydrates those are rich in fiber content help to prevent constipation.
✓ Also, they help in modulation of immune system.
• Example of Carbohydrates
- Monosaccharides: Glucose, galactose, glycerose, erythrose, ribose, ribulose, fructose.
- Oligosaccharides: Maltose, lactose, sucrose, raffinose, stachyose.
- Polysaccharides: Starch, glycogen, cellulose, pectin, inulin, hyaluonic acid.
✓ Food’s rich in carbohydrates are referred to as strachy foods. They are found in legumes, starchy vegetables,
whole-grain breads and cereals. They also occur naturally with vitamins and minerals in foods like milk, fruits,
and milk products. They are also found in refined and processed products like candy, carbonated beverages,
and table sugar.
• Examples of Polysaccharides
Name of the
Composition Occurrence Functions
Polysaccharide
Polymer of glucose containing a straight
In several plant species as
chain of glucose molecules (amylose) and a storage of reserve
Starch main storage
branched chain of glucose molecules food
carbohydrate
(amylopectin)
Animals (equivalent of Storage of reserve
Glycogen Polymer of glucose
starch) food
In roots and tubers (like Storage of reserve
Inulin Polymer of fructose
Dahlia) food
Cellulose Polymer of glucose Plant cell wall Cell wall matrix
Pectin Polymer of galactose and its derivatives Plant cell wall Cell wall matrix
Hemi cellulose Polymer of pentoses and sugar acids Plant cell wall Cell wall matrix
Plant cell wall (dead cells
Lignin Polymer of glucose Cell wall matrix
like sclerenchyma)
Exoskeleton
Bodywall of arthropods.
Chitin Polymer of glucose Impermeable to
In some fungi also
water
Cell wall of prokaryotic
Murein Polysaccharide cross linked with amino acids Structural protection
cells
Connective tissue matrix,
Ground substance,
Hyaluronic acid Polymer of sugar acids Outer coat of mammalian
protection
eggs
Heparin Closely related to chrondroitin Connective tissue cells Anticoagulant
Gums and Gums - bark or trees. Retain water in dry
Polymers of sugars and sugar acids
mucilage’s Mucilage’s - flower seasons

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 FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

Properties of food carbohydrates

- Monosaccharide exists in both as straight chain structure and cyclic structure. Sugars with five membered rings
and with six membered rings are most stable. Cyclic structures are the result of hemiacetal formation by
intermolecular reaction between carbonyl group and a hydroxyl group.

Structure of α-D-glucose: A. Straight chain B. Cyclic form

(i) Chiral centre
All monosaccharide except dihydroxy acetone contain one or more asymmetric (chiral) carbon atoms thus are
optically active isomers (enantiomers). A molecule with n chiral centres can have 2n sterioisomers. Glyceraldehyde
with one chiral centre has 21=2 and glucose with four chiral centres, have 24=16 stereoisomers.

(ii) D and L isomerism

• The designation of a sugar isomer as the D form or of its mirror image as the L form.
• The L and D forms of the sugar depends on the orientation of the -H and -OH groups around the carbon atom
adjacent to the terminal primary alcohol carbon (carbon 5 in glucose) determines whether the sugar belongs to
the D or L series.
• When the -OH group on this carbon is on the right, then sugar is the D-isomer; when it is on the left, then it is
the L-isomer.

• Most of the monosaccharide occurring in mammals is D sugars, and the enzymes responsible for their
metabolism are specific for this configuration. In solution, glucose is dextrorotatory—hence the alternative
name dextrose.
• The presence of asymmetric carbon atoms also confers optical activity on the compound. When a beam of
plane-polarized light is passed through a solution of an optical isomer, it will be rotated either to the right,
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 FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates
dextrorotatory (+); or to the left, levorotatory (-). The direction of rotation is independent of the
stereochemistry of the sugar, so it may be designated D (-), D (+), L (-), or L (+). For example, the naturally
occurring formof fructose is the D (-) isomer.

D-Tagatose is an epimer of D-fructose inverted at C-4

(iii) Alpha and beta anomers
• The anomeric centre of a sugar is a stereocentre created from the intramolecular formation of an acetal (or ketal)
of a sugar hydroxyl group and an aldehyde (or ketone) group. The two stereoisomers formed from the two
possible stereochemistry at the anomeric centre are called anomers. They are diastereoisomers of one another.
• Cyclic Structures and Anomeric Forms
- Alcohols react readily with aldehydes to form hemiacetals:
✓ This reaction is promoted in the presence of either acid or base
✓ One of the subtleties of this reaction involves the stereochemistry of the aldehyde and keto carbon it
goes from being achiral to chiral.

- Similarly alcohols react with ketones to produce hemiketals:

- Since aldoses and ketoses contain alcohol groups, in addition to their aldehyde or ketone groups,
they have the potential to react to form cyclic forms.

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

• Cyclization creates an anomeric carbon (the former carbonyl carbon), generating the α and β configurations of
the sugar, for example, α-D-glucopyranose and β-D-glucopyranose.
• These two sugars are both glucose but are anomers of each other.
• In the α configuration, the OH on the anomeric C projects to the same side as the ring in a modified Fischer
projection formula. In a Haworth projection formula of the α configuration, this OH is trans to the CH 2OH
group.
• Since the α and β forms are not mirror images, they are referred to as diastereomers.
• Enzymes are able to distinguish between these two structures and use one or the other preferentially. For
example, glycogen is synthesized from α-D-glucopyranose, whereas cellulose is synthesized from β-D-
glucopyranose.
• The cyclic α and β anomers of a sugar in solution are in equilibrium with each other and can be spontaneously
interconverted which is known as mutarotation.

Note By:
Structures of important disaccharides: The α and β refer to the configuration at the anomeric carbon atom (asterisk).
When the anomeric carbon of the second residue takes part in the formation of the glycosidic bond, as in sucrose, the
residue becomes a glycoside known as a furanoside or pyranoside. As the disaccharide no longer has an anomeric carbon
with a free potential aldehyde or ketone group, it no longer exhibits reducing properties, where as the anomeric end of the
glucose residue is not involved in the glycosidic linkage it and, therefore, lactose remains a reducing sugar.

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 FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates
• Re ducing suga rs: If the oxygen on the anomeric carbon of a sugar is not attached to any other structure, that
sugar can act as a reducing agent and is termed a reducing sugar. Such sugars can react with chromogenic
agents (for example, Benedict's reagent or Fehling's solution) causing the reagent to be reduced and colored,
with the anomeric carbon of the sugar becoming oxidized. [Note: Only the state of the oxygen on the anomeric
carbon determines if the sugar is reducing or nonreducing—the other hydroxyl groups on the molecule are not
involved.

(iv) Pyranose and furanose ring structures

• The stable ring structures of monosaccharides are similar to the ring structures of either pyran (a six-
membered ring) or furan (a five-membered ring). For glucose in solution, more than 99% is in the pyranose
form.

• "Pyranose" is used to refer to the pyran ring structure (6membered ring with 5 carbons and 1 oxygen)
• For five membered rings (four carbons and 1 oxygen) the structure is a furanose ring.
• Cyclization is reversible.
• The resulting chirality of the aldehyde carbon (or keto carbon in ketoses) in the cyclic structure can be either
the - or -form. This carbon is termed the anomeric carbon, and the - and -forms are anomers.
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• The ring structure representations are termed "Haworth Projections".
• Cyclic structures can also be represented using Fischer diagrams

• Alternative ring structures

- Glucose has a variety of alcohol functional groups, and is able to form alternative hemiacetal ring structures.
- The 5C-OH can react with the aldehyde forming the above pyranose ring
- The 4C-OH can react with the aldehyde forming a five membered furanose ring
• Ring geometry
- The carbon atoms in the ring are sp3 hybrid orbitals, and tetrahedral geometry.
- The oxygen in the ring is also sp3, and tetrahedral.
- Thus, the ring structures cannot be planar (that would require trigonal planar geometry, or sp2 hybrid
orbitals, for the atoms in the ring)
- The rings actually look like

• There is a "boat" and a "chair" structure. There are two different orientations of the functional groups bonded
to the ring carbons:
- If they point in the general plane of the ring, they are said to be in the "equatorial" position
- If they point in an orientation perpendicular to the general plane of the ring, they are side to be in the
"axial" position
- The chair can have two different conformations. The principle effect of the alternative conformations is to
change the axial versus equatorial orientations for the carbon functional groups.
• Functional groups in the axial position are somewhat crowded together, whereas, the same functional
groups in the equatorial position are separated from each other.
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Fischer Projection Haworth Projection Configuration Chair Conformation

Right Down α Axial
Left Up β Equatorial

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(v) Isomers and epimers
• Compounds that have the same chemical formula but have different structures are called isomers. For example,
fructose, glucose, mannose, and galactose are all isomers of each other, having the same chemical formula,
C6H12O6.
• Carbohydrate isomers that differ in configuration around only one specific carbon atom are defined as epimers
of each other. For example, glucose and galactose are C-4 epimers—their structures differ only in the position of
the -OH group at carbon 4. [Note: The carbons in sugars are numbered beginning at the end that contains the
carbonyl carbon—that is, the aldehyde or keto group]
• Glucose and mannose are C-2 epimers. However, galactose and mannose are not epimers—they differ in the
position of -OH groups at two carbons (2 and 4) and are, therefore, defined only as isomers.

Isomers and Epimer of Fructose

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(vi) Enantiomers
• A special type of isomerism is found in the pairs of structures that are mirror images of each other. These mirror
images are called enantiomers and the two members of the pair are designated as a
D- and an L-sugar.
• The vast majority of the sugars in humans are D-sugars.
• In the D isomeric form, the OH group on the asymmetric carbon (a carbon linked to four different atoms or
groups) farthest from the carbonyl carbon is on the right, while in the L-isomer it is on the left.
• Enzymes known as r acem ases are able to interconvert D- and L-isomers.

(vii) Glycosidic bonds

• Monosaccharides can be joined to form disaccharides, oligosaccharides and polysaccharides.
• Important disaccharides include lactose (galactose + glucose), sucrose (glucose + fructose) and maltose (glucose
+ glucose).
• The bonds that link sugars are called glycosidic bonds. These are formed by enzymes known as
g ly cosy ltransferases that use nucleotide sugars such as UDP-glucose as substrates.
• Glycosidic bonds between sugars are named according to the numbers of the connected carbons and also with
regard to the position of the anomeric hydroxyl group of the sugar involved in the bond.
• If this anomeric hydroxyl is in the α configuration, the linkage is an α-bond. If it is in the
β configuration, the linkage is a β-bond.
• Lactose, for example, forming a glycosidic bond between carbon 1 of β-galactose and carbon 4 of glucose. The
linkage is, therefore, a β (1→4) glycosidic bond.

(viii) Mutarotation In Sugars

- Mutarotation is the change in the optical rotation because of the change in the equilibrium between two
anomers, when the corresponding stereocenters interconvert. Cyclic sugars show mutarotation as α and β
anomeric forms interconvert.
- The optical rotation of the solution depends on the optical rotation of each anomer and their ratio in the
solution.
- Mutarotation was discovered by French chemist Sir Dubrunfaut in 1846, when he noticed that the specific
rotation of aqueous sugar solution changes with time. [The organic fructose molecule was subsequently
discovered by Dubrunfaut in 1847].

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates
- Sugars in the ring form can exist in two states, one where the C-1 hydroxy group is above the plane of the ring
(β) and one where it is below (α).
- In aqueous solution there is a constant interchange between the various conformations via the breaking open
of the hemi acetal structure and its subsequent reforming.

- Glucose is a mixture of α- and β-anomers, primarily the α-anomer. The optical rotation of the α-anomer is
+112.2° (c = 10% in water, 20 °C) and the β-anomer is +18.7° (c = 10% in water, 20 °C). When D-glucose is
dissolved in water, the optical rotation gradually changes (mutarotates) with time and approaches a final
equilibrium value of +52.7° (c = 10%, 20 °C) due to the formation of an equilibrium mixture consisting of
approximately one-third α- and two-thirds β-D-glucose.

General methods of analysis of food carbohydrates

- Carbohydrates are a significant component of many meals and raw materials. Food carbohydrate analytical
techniques are important for food quality assurance and product uniformity.
- The classical methods are mainly divided into three categories:
(i) Total sugar analysis (phenol-sulfuric acid assay, anthrone-sulfuric acid methods, and methodology for
uronic acid and reducing sugar analysis)
(ii) Monosaccharide’s analysis (enzymatic methods, anion-exchange chromatography, and gas-liquid
chromatography)
(iii) Structure characterization (partial acid hydrolysis, smith degradation, and methylation analysis).

Total carbohydrate:
By difference: 100 - (weight in grams [protein + fat + water + ash + alcohol] in 100 g of food)
By direct analysis: weight in grams (monosaccharides + disaccharides + oligosaccharides + polysaccharides,
including fibre)
Available carbohydrate:
By difference: 100 - (weight in grams [protein + fat + water + ash + alcohol + fibre] in 100 g of food)
By direct analysis: weight in grams (monosaccharides + disaccharides + oligosaccharides + polysaccharides,
excluding fibre)*
*May be expressed as weight (anhydrous form) or as the monosaccharide equivalents (hydrous form including
water).

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

❖ Carbohydrate Assay
- The enzymatic assay of sucrose, fructose and starch are based on the enzymatic determination of glucose. The
first step is the conversion or degradation of the carbohydrate to glucose or glucose derivate. The glucose
concentration is then determined according to the two different detection systems: glucose oxidase (GO) or
hexokinase (HK) in combination with the glucose-6-phosphate dehydrogenase reaction.

Detection of glucose via glucose oxidase (GO) and peroxidase, monitored at 540 nm.

Detection of glucose via hexokinase (HK) and glucose-6-phosphate dehydrogenase by formation of NADH,
monitored at 340 nm.

Sucrose assay: Sucrose is hydrolysed to glucose and fructose by invertase. Glucose and fructose are phosphorylated
with ATP in the hexokinase reaction. Glucose-6- phosphate and NAD are then converted to 6-phosphogluconate and
NADH via glucose-6-phosphate dehydrogenase, monitored at 340 nm.

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

Fructose assay: Fructose is phosphorylated by ATP in a reaction catalysed by hexokinase. The resulting fructose-6-
phosphate is then converted to glucose-6- phosphate by phosphoglucose isomerase. Glucose-6-phosphate and NAD is
converted to 6-phosphogluconate and NADH, monitored at 340 nm.

- General methods of analysis of food carbohydrates

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

• Method for measuring the degree of starch gelatinization

• Estimation of reducing sugar by Nelson-Somogyi method

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates
• Total dietary fibre assay
- The total dietary fibre content is determined by a combination of enzymatic and gravimetric methods.
- Samples of dried, fat-free foods are gelatinised with heat-stable amylase and then enzymatically digested with
protease and amyloglucosidase to remove protein and starch in the sample.
- Ethanol is added to precipitate the soluble dietary fibre.
- The residue is then filtered and washed with ethanol and acetone.
- After drying, the residue is weighed.
- Half of the sample is analysed for protein and the other half is ashed.
- Total dietary fibre equals weight of residue minus weight of protein and ash.

• Method for the determination of resistant starch

100 mg sample
Add 10 ml enzyme solution in buffer (pH 6.9); pancreatic a -amylase (500U)
0.1 M tris-maleate buffer solution (calcium chloride 4mM)

Mix 16 hours at 37°C; then add 40 ml ethanol

Leave 1 hour; then centrifuge

Wash residue twice with 80% EtOH, dry at 60°C
Add 1.56 ml H2O, then add 1.5 ml 4M KOH

Mix 0.5 hours at room temperature
Add 12 ml H2O

To 1.5 ml dispersion, add approximately 0.65 ml 2M acetic acid (to obtain pH 4.5)
and 0.1 ml amyloglucosidase (20/0.1 ml 0.1 M Na acetate buffer pH 4.5)
Shake 90 min. at 65°C

Determine glucose using glucose oxidase assay
The result is resistant starch

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

Dietary fibre, Crude fibre and application of food carbohydrates

Dietary fiber
- Dietary fiber (British spelling fibre) or roughage is the portion of plant-derived food that cannot be completely
broken down by human digestive enzymes.
- Dietary fibers are diverse in chemical composition, and can be grouped generally by their solubility, viscosity,
and fermentability, which affect how fibers are processed in the body.
- Dietary fiber has two main components: Soluble fiber and insoluble fiber, which are components of plant foods,
such as legumes, whole grains and cereals, vegetables, fruits, and nuts or seeds.
- All types of soluble fibers slow digestion, so it takes longer for our body to absorb sugar (glucose) from the foods
you eat. This helps prevent quick spikes in our blood sugar levels -- an important part of managing diabetes.
Soluble fibers also bind with fatty acids, flushing them out of the body and helping to lower LDL (bad)
cholesterol.
- Insoluble fibers help hydrate and move waste through your intestines. That's one thing it does that helps prevent
constipation and keeps regular.
- Benefits of a Dietary Fiber: Normalize the bowel movement, Promote bowel health, Lower the cholesterol level [reduce
Low-Density Lipoprotein (LDL) in total cholesterol], Control blood sugar level [Soluble fiber reduces the absorption
of sugar and keeps the blood sugar level normal. Insoluble fiber also helps to reduce typeII diabetes], Support to
maintain healthy weight [Dietary fiber provides less energy dense by facilitating few calories from the same volume
of foods]
Crude Fiber
- Crude fiber is a part of insoluble fiber found in the edible portion of the plant cell wall. It is basically cellulose
material obtained as a residue of the chemical analysis of vegetable substances.
- The main health benefit of crude fiber is that it facilitates regular bowel movement. Leafy greens, whole grains,
and beans (black beans) are some common examples of crude fiber.

The difference between Dietary Fiber and Crude Fiber

Dietary Fiber Crude Fibers
Dietary fiber is the sum of both soluble and Crude fiber is a part of insoluble fiber found in
Origin
non-soluble fiber groups. the edible portion of the plant cell wall.
Dietary fiber is either soluble or non-soluble Crude fiber is not soluble in water.
Solubility
in water
Some dietary fibers are subjected to Crude fibers are not fermented inside the
Fermentation
fermentation inside the digestive system. digestive tract.
Nature Inside Dietary fiber might be relatively intact when Crude fiber is relative intact through a period of
The Digestive it passes through the digestive tract, but this passage.
Tract is based on the type of dietary fiber.
Dietary fiber contains pectins, gums, and Crude fibers do not contain pectins, gums, and
Composition
mucilages. mucilages.

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

Soluble or
Types of Fiber Sources Health Benefits
Insoluble

Naturally found in nuts, whole "Nature's laxative": Reduces constipation,

Cellulose, some
Insoluble wheat, whole grains, bran, seeds, lowers risk of diverticulitis, can help with
hemicellulose
edible brown rice, skins of produce. weight loss.

Extracted from onions and

Inulin byproducts of sugar production from May increase "good" bacteria in the gut and
Soluble
oligofructose beets or chicory root. Added to enhance immune function.
processed foods to boost fiber.

Good for heart health and possibly

Found naturally in flax, rye, some
Lignin Insoluble immune function. Use caution if you
vegetables.
have celiac disease or are gluten intolerant.

Naturally found in oats, oat bran, Helps lower bad LDL cholesterol, reduces
Mucilage, beta- beans, peas, barley, flaxseed, berries, risk of coronary heart disease and Type II
Soluble
glucans soybeans, bananas, oranges, apples, diabetes. Use caution if you have celiac
carrots. disease or are gluten intolerant.

Naturally found in fruits, berries, and

Soluble (some Slows the passage of food through the
seeds. Also extracted from citrus peel
Pectin and gums pectins can be intestinal GI tract, helps lower blood
and other plants boost fiber in
insoluble) cholesterol.
processed foods.

Added to processed foods as a

Polydextrose bulking agent and sugar substitute. Adds bulk to stools, helps prevent
Soluble
polyols Made from dextrose, sorbitol, and constipation. May cause bloating or gas.
citric acid.

Extracted from rushed seeds or

husks of Plantago ovata plant. Used in Helps lower cholesterol and prevent
Psyllium Soluble
supplements, fiber drinks, and added constipation.
to foods.

Starch in plant cell walls naturally

May help manage weight by increasing
found in unripened bananas,
fullness; helps control blood sugars. It
Resistant starch Soluble oatmeal, and legumes. Also extracted
increases insulin sensitivity and may
and added to processed foods to
reduce the risk of diabetes.
boost fiber.

Helps lower cholesterol (LDL and total

Extracted from wheat starch, and cholesterol) and may lower blood sugar
Wheat dextrin Soluble widely used to add fiber in processed and reduce risk for heart disease; more
foods. research is needed. Avoid if you have celiac
disease or are gluten intolerant.

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

Changes in food carbohydrates during processing

- Carbohydrates can be said to be relatively stable to processing and storage compared to sensitive nutrients such
as vitamins.
- There is no significant loss in the nutritional value of carbohydrates in frozen, canned or dried foods.
- Infact, some carbohydrates are made more digestible through processing and thus nutritionally available and
complex carbohydrates are converted to simple sugars.
- Adverse brown pigments occur in frozen and dried foods when carbohydrates react with proteins.
- During wet heat treatment as in blanching and boiling; there is considerable loss of low molecular weight
carbohydrates as well as micro nutrients into the processing water. The loss of glucose and fructose at boiling is
higher than that of sucrose. The loss of low molecular weight carbohydrates also may vary between species and
cultivars.
- Maillard reactions: Non enzymatic browning reactions occur between reducing sugars and amino groups in
foods during processing and storage. These reactions are temperature dependent and most extensive at
intermediate water activities. They are important nutritionally as they may diminish the bio availability of amino
acids especially lysine and thus diminishing the protein nutritional value. The carbohydrate content and
availability are influenced only marginally.

Maillard reaction (MR), also known as glycation

- Starch - heat-induced effects: When starch is heated in the presence of water, the starch granules ruptures and
form gels. The gelatinization increases the availability of starch for digestion by amylolytic enzymes. Gelatinized
starch is not in thermodynamic equilibrium, therefore a progressive re-association of the starch molecules upon
ageing. This recrystallization is referred to as retrogradation and may reduce the digestibility of the starch.
- Par-boiling: During par-boiling of rice, the kernels are subjected to a pre-treatment involving heating and drying.
This process reduces the stickiness of the rice, possibly by allowing leached amylose to retrograde and/or form
inclusion complexes with polar lipids on the kernel surface. Parboiling also affects the final cooking properties of
the rice.
- Starch–texturization: Chemicals are used in foods to carry out specialized functions. As foods are processed for
alteration in selected products, additives are usually used. The demand for starch-based texturizing agents is
increasing day by day. Among all the additives, starch-based texturizing agents are used in a large quantity.
Starch-based texturizing agents are meant to add or modify the whole texture of the food products. The primary
starch-based texturizing agents are emulsifiers, stabilizers, phosphates, dough conditioners, etc., each one has its
own unique functions. Apart from improving the texture of the foods, some of the other applications of starch-
based texturizing agents are dispersion of oil and flavours of the foods, prevent evaporation of volatile flavour
oil, and enhance the water-holding capacity of the food components.
- Milling and peeling: During milling of cereal grains to refined flours the outer fibre-rich layers are removed,
resulting in a lower content of total dietary fibre.
- Heat-treatment: Processes involving heat-treatment may affect the dietary fibre in different ways. An increased
temperature leads to a breakage of weak bonds between polysaccharide chains. Also, glycosidic linkages in the
dietary fibre polysaccharides may be broken. These changes are important from analytical, functional, and
nutritional points of view.

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

Digestion, absorption and metabolism of carbohydrates

❖ Digestion of Carbohydrates
- The primary goal of carbohydrate digestion is to break down polysaccharides and disaccharides into
monosaccharides that can then be converted to glucose.
- The major source of carbohydrates is found in plants.
- Glucose is the universal fuel for human cells. The glucose concentrations in the body are maintained within limits
by various metabolic processes.
- The principal sites of carbohydrate digestion are the mouth and small intestine. The dietary carbohydrate consists
of:
i. Polysaccharides: Starch, glycogen and cellulose.
ii. Disaccharides: Sucrose, maltose and lactose.
iii. Monosaccharides: Mainly glucose and fructose.
- Monosaccharides need no digestion prior to absorption, whereas disaccharides and polysaccharides must be
hydrolysed to simple sugars before their absorption.

• Digestion in Mouth
- Digestion of carbohydrates begins in the mouth. Salivary glands secrete α-amylase (ptyalin), which initiates the
hydrolysis of a starch.
- During mastication, salivary α-amylase (glucose-polymer cleavage enzyme) acts briefly on dietary starch in
random manner breaking some α-(1 → 4) bonds, α-amylase hydrolysis starch into dextrin’s.
• Digestion in Stomach
- Carbohydrate digestion halts temporarily in the stomach because the high acidity inactivates the salivary
α-amylase.
• Digestion in Intestine
- Further digestion of carbohydrates occurs in the small intestine by pancreatic enzymes.
- There are two phases of intestinal digestion.
i. Digestion due to pancreatic α-amylase
ii. Digestion due to intestinal enzymes: sucrase, maltase, lactase, isomaltase.

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates
• Digestion due to pancreatic α-amylase
- The function of pancreatic α-amylase is to degrade dextrin’s further into a mixture of maltose, isomaltose and
α-limit dextrin.
- The α-limit dextrin’s are smaller oligosaccharides containing 3 to 5 glucose units.
• Digestion due to intestinal enzymes
- Enzymes responsible for the final phase of carbohydrate digestion are located in the brush-border membrane.
- The end products of carbohydrate digestion are glucose, fructose and galactose which are readily absorbed
through the intestinal mucosal cells into the bloodstream.

❖ Absorption of Carbohydrates
- Carbohydrates are absorbed as monosaccharides from the intestinal lumen.
- Two mechanisms are responsible for the absorption of monosaccharides:
i. Active transport against a concentration gradient, i.e., from a low glucose concentration to a higher
concentration.
ii. Facilitate transport, with concentration gradient, i.e., from a higher concentration to a lower one.

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

✓ Active Transport
- A sodium dependent glucose transporter
(SGLT-1) binds both glucose and Na+ at
separate sites and transports them both
through the plasma membrane of the
intestinal cell.
- The Na+ is transported down its concentration
gradient (higher concentration to lower
concentration) and at the same time glucose is
transported against its concentration gradient.
- The free energy required for this active
transport is obtained from the hydrolysis of
ATP linked to a sodium pump that expels Na+
from the cell in exchange of K+.

✓ Facilitate Transport
- Fructose and mannose are transported across the brush border by a Na+ independent facilitative diffusion
process, requiring specific glucose transporter, GLUT-5.
- The same transport can also be used by glucose and galactose if the concentration gradient is favourable.
- The sodium independent transporter, GLUT-2 that facilitates transport of sugars out of the mucosal cells,
thereby entering the portal circulation and being transported to the liver.

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FOOD ANALYSIS (MPA 104T) Unit I: Carbohydrates

❖ Metabolism of Carbohydrates
- Metabolism of carbohydrates includes the process where in complex carbohydrates with 3 or more sugar units in
a chain are broken down into simple mono- and disaccharides like glucose and sucrose which are important
components of several metabolic pathways.
- Glycolysis is an important pathway which occurs in almost every living cell, both in prokaryotes and eukaryotes.
In this pathway glucose is broken down into 2-3 carbon unit molecules like pyruvate and the energy generated in
this pathway is stored in the form of ATP and NADH.
- Depending on whether the subsequent steps are aerobic or anaerobic several end products like acetyl CoA and
lactic acid, ethanol, acetic acid may be generated.

Overall Carbohydrate Metabolism

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