classification and analysis

1. CARBOHYDRATES
DEVIPRIYA P V
M PHARM
PHARMACEUTICAL ANALYSIS

2. DEFINITION
Carbohydrates may be defined as
polyhydroxy aldehydes or ketones or
compounds which produce them on
hydrolysis.

3. CLASSIFICATION
 Based on number of sugar units:
1. Monosaccharides
: aldoses
: ketoses
2. Oligosaccharides
: disaccharides
: trisaccharides
3. Polysaccharides
: homopolysaccharies
: heteropolysaacharides

4. CARBOHYDRATES
MONOSACCHARIDES
(1 SUGAR MOLECLE)
GLUCOSE
FRUCTOSE
GALACTOSE
DISACCHARIDES
(2 SUGAR
MOLECULE)
SUCROSE
LACTOSE
MALTOSE
OLIGOSACCHARIDES
(2-10 SUGAR
MOLECULES)
RAFFINOSE
STACHYOSE
POLYSACCHARIDES
(10 OR MORE)
STARCH
GLYCOGEN
CELLULOSE

5. Monosaccharide:
 Simple sugars.
 Further classification based on functional
group :
1. Aldoses:
 Functional group is aldehyde.
Eg: glyceraldehyde, glucose
2. Ketoses:
 Functional group is ketose.
Eg: dihydroxyacetone , fructose

6.  Classification based on no: of c-atoms:
:Trioses(3 C)
:Tetroses (4C)
:Pentoses(5C) etc.
Oligosaccharides:
 Liberate 2-10 monosaccharide units on
hydrolysis.
 Division based on no: of monosaccharide
units
Disaccharide :
 2 monosaccharide units(similar or
dissimilar)
:reducing & non reducing

7.  Reducing: free aldehyde / keto group.
-Eg: maltose, lactose
 Non reducing : no aldehyde /keto group.
-Eg: sucrose, trehalose.
Polysaccharides
 10 or more monosaccharide units.
Homopolysaccharies:
 Hydrolysis yield only single type of
monosaccharide
Heteropolysaacharides:
 Hydrolysis yield a mixture of a few
monosaccharide or their derivatives.

8. Analysis of
carbohydrates1. Sample preparation
:Extraction
:Removal of interfering compounds
:Clarification
2. Calculation by difference
3. Chemical methods:
:Lane-Eynon method
:Munson walker method
:Nelson Somogyi method
:Alkaline ferricyanide method
:Phenol sulfuric acid method.
:Anthrone method.

9. 4.Enzymatic methods:
:D-glucose/D-fructose/D-sorbitol
method.
:Lactose/D-galactose method
:Maltose/sucrose/D-glucose method.
:Raffinose method
:Oxidase method.
5. Physical method:
:polarimetry
:Refractive index measurement.
:Specific gravity measurement.
6.Modern analytical methods:
:HPLC
:GC
:NMR

10. Extraction of mono and
oligosaccharides:
Finely ground material + redistilled alcohol +
pptd CaCO3.
↓
Heated on steam bath for 30 min
↓
Filter
↓
Re -extract using soxhlet apparatus
↓
Combine the filtrate & made up to 80%
alcohol

11. Removal of interfering compounds:
 Treatment with
: lead acetate
: Ion exchange resins
Calculation by difference:
Total carbohydrate in food
=100% -(% moisture + %
protein +% fat+%ash )

12. Lane- Eynon method
 Determine concentration of reducing sugars
Sample solution
↓
Added to reduce Cu in soxhlet apparatus
↓
Mixture boiled for 2 min
↓
Add methylene blue.
↓
Titrate until decolouration (within 3 min)

13. Munson walker method
 Oxidation of carbohydrate in the presence
of heat and excess of cupric sulphate and
alkaline tartrate under controlled conditions.
 Cupric oxide precipitate as carbohydrates
are oxidized.
 Determined : gravimetrically
: electrolytic deposition
: titrimetrically

14. Alkaline ferricyanide method
 ferricyanide carbohydrates ferrocyanide.
 Basic pH.
 Ferrocyanide + ferric ions gives prussian blue
colour , measured spectrophotometrically at
700 nm.
Phenol sulfuric acid method:
 Carbohydrate strong acid furfural+hydroxy
methyl
heat(20min) furfural.
 Condense with phenol & form yellow orange
color(490 nm).

15. Anthrone method:
Carbohydrate + Anthrone Blue green
color
heat in water
bath 15 min
cool in
dark (30 min)
absorbance at 620 nm stable color.
Somogyi method:
 Based on the reduction of cu (2)ion to cu (1)
ions by reducing sugars.
 Reduction of complex produces an intense
stable blue color which is determined
spectrophotometrically.

16. Enzymatic methods
1. Total change method.
2. Rate assay method.
D-glucose method:
D- glucose + ATP Hexokinase G-6-P + ADP
2 G-6-P +NAD G-6-P dehydrogenase Gluconate
6 phosphate +NADPH
 NADPH is measured
Spectrophotometrically at 340nm.

17. Lactose / D- galactose method:
 Lactose + H2O β galactosidase
D-glucose +D-galactose.
 D-galactose+NAD+ β galactosidase dehydrogenase
D-galactonic acid +NADH +H+
 The amount of NADH is stoichiometric with the
amount of lactose.
Maltose / sucrose method:
Maltose + H2O α-glucosidase 2-D glucose.
Sucrose +H2O α glucosidase D-glucose+D-fructose

18. Oxidase method:
 Combination of enzymatic and
instrumental method.
 Oxidase is used.
 H2O2 produced by enzymatic oxidation
is subjected to electrochemical
oxidation and then measured.

19. Physical methods
1.Polarimetry :
 Carbohydrates rotate the plane polarised light
through an angle .
 Polarimeter is used.
[α]D
20
=100α lc
2.Refractive index measurement:
RI= sine of angle of incidence/ sine of angle of
refraction
3. Specific gravity method:
S=wt of x ml of substance/wt of x ml of water.

20. QUALITATIVE TEST FOR
ANALYSIS
 Monosaccharides:
1. Molisch’s test
2.Reduction test
:Fehling’s test
:Benedict’s test
:Barford's test
:Picric acid test
3.Selivanoff’s test
4.Elson Morgan test
5.Tauber test
6.Glucose Oxidase test

23. Oligosaccharides:
 Disaccharides
Maltose= glucose + glucose(reducing)
Lactose= glucose+ galactose(reducing)
Sucrose=glucose +fructose(non reducing)
 inversion test for sucrose.
Polysaccharides:
:Iodine test
:Osazone test

27. THANK YOU