Application of Capillary

Electrophoresis
By- Group B
Abhay Verma
Aniket Roshan
Anshuman Patankar
Kriti Agrawal
Electrophoresis
Electrophoresis is the movement of particles
relative to a fluid under the influence of
spatially uniform electric field.

Anions move towards anode and vice versa.

Electrophoresis
Separation of components in a
mixture under the influence of
electric field depends on velocity of
the particles
● v=Eq/F
○ v= Velocity of molecule
○ E= Electric Field
○ q= Charge of the particle
○ F= Friction coefficient
Types of Electrophoresis:
● Gel Electrophoresis
● High resolution
Electrophoresis
● Capillary Electrophoresis
● Isoelectric Focussing
● Immunochemical
● Pulse Field
● 2D Electrophoresis
Capillary Electrophoresis
Various Separation Modes of CE

Capillary Zone Capillary Gel Electrophoresis

Electrophoresis (CZE)
● Simplest form of CE
● Separation based on charge and
ionic radius of analytes
● Fundamental to CZE are
homogeneity of buffer solution and
constant field strength along the
capillary
● Application: Separation of
hemoglobin from hemolysate
● Used for size and shape separation
● Separation based on solute sizes
● Detection by UV absorbance of
chromophore
● Application: DNA Sequencing and
Protein Analysis
Capillary Isoelectric Immunoaffinity Capillary
Focussing Electrophoresis

● Depends on PH buffer gradient ● Combine immunoassay and CE

● Capillary is coated inside with an ● Application:
ampholyte, when the field is applied ○ DNA Analysis
create a PH gradient ○ Pharmacological
● Application: ○ Forensics
○ Separation of proteins
○ Separation of peptides
○ Separation of amino Acid
○ Separation of drugs
Application in Cosmetics
❖ The demand of cosmetic products due to skin care, has increased remarkably.
❖ One major use of capillary electrophoresis in cosmetics is to determine the metal
impurity in it.
❖ According to FDA(Food and Drug Administration) the cosmetics should not
contain more than 10 ppm of Lead(Pb), 1 ppm of Mercury(Hg) and 20 ppm for
Cadmium(Cd)
Determining metal impurity

❖ One can determine most of metal using suitable

ligand as chelating agent
❖ Mercury(Hg), Cadmium(Cd), and Lead(Pb) can be
determined by EDTA.Because these elements shows
high affinity toward it
❖ The technique used to determine the concentration of
metal using capillary electrophoresis is dynamic
chelation mechanism
❖ Firstly, the capillary was rinsed by a 0.6 mM TTAB solution to reverse the EOF
❖ The sample solution, which was diluted using 25 mM ammonium acetate (pH
6.0), was injected into the capillary
❖ Then, EDTA buffer was injected at −25 kV for 1 min
❖ Finally, the separation was performed at −20 kV using a separation buffer
❖ A small quantity of chelating agents introduced into the capillary could yield
33, 50 and 100-fold detection improvements for Pb, Cd and Hg, respectively
Application in Pharmaceutics
❖ Capillary electrophoresis (CE) offers fast and highly efficient
separation of molecules with a broad range of properties making it
well suited for the analyses of biological samples
❖ Drug making process involve detection and extraction of essential
proteins like insulin
❖ Insulin is essential for glucose digestion and exists as a Zn2+
containing hexamer stored in the pancreas
Environmental Applications:

❖ An important precursor of photochemical smog

is aldehyde.
❖ Aldehydes of low molecular mass are the most
abundant organic gas in atmosphere.
❖ Aldehydes has been found responsible for
formation of harmful free radicals, ozone, and
peroxyacetyl nitrate.
❖ As aldehydes are very harmful, capillary
electrophoresis can be used for its
environmental analysis.
Applications in Environment & Nutritional field
● Continuous monitoring of pesticide, with
unquestionable impact on crop production
and the residues in environmental samples
is important. Capillary electrophoresis has
proven be a resourceful alternative for the
analysis of pesticides.
● Capillary electrophoresis can make an
important contribution in wine production.
It is used to find the contents of mono- , di-
and tri-carboxylic acids in wines, beers , etc.
● CE is also used in food analysis of various
carbohydrate and fatty acids. Figure
presents the CE separation of carbohydrates
present in yogurt.
Application in clinical analysis
Hemoglobins in hemolysate

● Used as determination of several

metabolites related to Diabetes Mellitus
● Detects high levels of glucose are found
in blood, which promote the glycosylation
of hemoglobin giving glycated
hemoglobin (HbA1c), a strong indicative
of the glucose concentration
● Figure shows separation of HbA1c among
other hemoglobins and variants using
capillary isoelectric focusing (cIEF) with
chemical mobilization
Application in clinical analysis
ketoacid mtetabolites in serum

● Use metabolic disorder associated with Diabetes

Mellitus is the excessive production of ketonic
bodies (pyruvic, acetoacetic acids, acetone and
the reduced forms).
● Detects and analyses imbalance in anion gap
caused by ketonic bodies, [Na+] + [K+] – [Cl-] –
[HCO3-] and ketonic bodies
● Fig illustrates the separation of pyruvate, lactate,
acetoacetate and b-hydroxybutyrate in human
blood serum by CE as a single analytical
technique for the evaluation of the ketoacidosis
condition in diabetic patients serum.
Application in clinical/therapeutic analysis
opiates in hair
● The opiates like morphine, codeine, heroin and
fentanyl can be detected using CE.
● Hair is a popular specimen for drug analysis:
non-invasive to collect, provides a historical
record of exposure
● Drug analysis by Figure shows analysis of
morphine in hair samples of patients suffering
from medullar lesion, receiving morphine.
● A three-step methodology was developed on hair
analysis of such a patient: hydrolysis,
ion-exchanger extraction followed by CE analysis
in a low pH phosphate buffer. Peak identity was
confirmed by spiking techniques.
Conclusion

● At the present time, capillary electrophoresis (CE) encompasses a family of

electrodriven techniques with distinct separation mechanisms and
selectivities.
● Relevant aspects of the technique such as high efficiency, high resolving
power, high speed, full automation and a variety of injection based
pre-concentration schemes and detection modes have all been extensively
investigated.
● CE continues to prove to be an important and efficient tool for drug analysis,
clinical/forensic , cosmetological, environmental, nutritional and
pharmaceutical applications. Further rountine applications are extensively
researched in present times.