1. EVALUATION SEMINAR
SUBJECT:
MODERN PHARMACEUTICAL ANALYSIS
TOPIC: ELECTROPHORESIS
Presented by
DEVI. S
I M. Pharm,
Department of Pharmaceutics,
M.S.R.C.P,
Bangalore-54.

2. CONTENTS
¤ Introduction
¤ Definition and Principle
¤ Theory
¤ Factors affecting the
migration of ions
¤ Types of electrophoresis
o Moving boundary electrophoresis
o Zone electrophoresis

3. INTRODUCTION
 Electrophoresis is a physical method of analysis
which involves separation of the compounds that
are capable of acquiring electric charge in
conducting electrodes.
ELECTROPHORESIS (Greek word)=BORNE BY ELECTRICITY
 It is a separation technique in which the components are
separated due to their varying behavior under the
influence of an applied electric field.
 The technique was pioneered in 1937 by the Swedish
chemist Arne Tiselius for the separation of proteins.

4. DEFINITION AND PRINCIPLE
Electrophoresis is defined as the “migration of
charged molecules under the influence of an
external electric field”
 In practical terms, a positive (anode) and negative (cathode)
electrode are placed in a solution containing ions.
 Then, when a voltage is applied across the electrodes, solute
ions of different charge, i.e., anions (negative) and cations
(positive), will move through the solution towards the
electrode of opposite charge.

5. THEORY
Ion migration velocity can be expressed as:
V=µeE
The applied field (Fef) (driving force) : Fef=qE
The friction (Ffr) drag is given by: Ffr=6ήπrv
At equilibrium: Fef=Ffr
Therefore: qE=6ήπrv

6. FACTORS AFFECTING THE MIGRATION OF
IONS
 Factors related to the sample
 Charge
 Size
 Shape
 Properties of electric field
 Potential difference
 Current
 Resistance
 Environmental characteristics
 pH
 Temperature
 Electrolyte concentration
 Composition and nature of supporting medium
 Buffer
 Supporting medium

7. TYPES OF
ELECTROPHORESIS
Free solution / frontal / moving boundary
electrophoresis: supporting medium is absent
Zone electrophoresis: supporting medium is present
1) Paper electrophoresis
2) Gel electrophoresis
3) Capillary electrophoresis
4) Continuous electrophoresis
5) Isotachophoresis
6) Iso electric focusing

8. MOVING BOUNDARY ELECTROPHORESIS

9. ZONE ELECTROPHORESIS
Paper electrophoresis
a) Open strip Paper Electrophoresis
Horizontal Vertical
b) Closed strip Paper Electrophoresis
Horizontal Vertical

10. c. Immersed strip Paper Electrophoresis
- +
Cooling
liquid
Filter paper
Buffer
d. Enclosed strip Paper Electrophoresis

11. GEL ELECTROPHORESIS

12. CAPILLLARY ELECTROPHORESIS

13. CONTINUOUS ELECTROPHORESIS

14. ISOTACHOPHORESIS
 Iso-equal , tachos- speed, phoresis-migration
 The technique of Isotachophoresis depends on the
development of potential gradient.
Principle:
 A leading electrolyte(chloride) with a higher mobility than the
analytes and a trailing electrolyte(glycinate) with a lower
mobility are used.
 After application of an electric potential a low electrical field
is created in the leading electrolyte and a high electrical field
in the terminating electrolyte.
 The pH at sample level is determined by the counter-ion of
the leading electrolyte that migrates in the opposite direction.

16. ISO ELECTRIC FOCUSINNG(IEF)
» Proteins carry both positive and negative charges, which is the PH when
molecule has no net charge.
» The pH which gives zero net charge is the isoelectric point or pH.
» Generally proteins readily crystallize at the isoelectric point.
» Most of the proteins have isoelectric point of 5-9.
» When electrophoresis is run in a solution buffered at constant PH
, proteins having a net charge will migrate towards the opposite
electrode so long as the current flows.
» The use of PH gradient across the supporting medium causes each
protein to migrate to an area of specific PH
» Proteins are found at the point of the gradient where they carry no net
charge.
» The PH of the protein equals the PH of the gradient, thus resulting in
sharp well defined protein bands.

17. REFERENCES
1. Instrumental methods of chemical analysis by B.K. sharma
pg.no C269-c281.
2. Electrophoresis by Melvin wiley publications.
3. http://en.wikipedia.org/wiki/Electrophoresis.
4. www.pharmainfo.net.
5. Theory of Electrophoresis-K.S. Pitre, Dr. Harisingh Gour
University, Sagar, India, Encyclopedia of Analytical
Science.
6. http://books.google.co.in
7. Fundamentals of Analytical chemistry-Skoog (pg.no-1003)

20. ‘winning doesn’t always mean
being first, means ur doing
better than you have done
before’
Bonnie
Blair