Biochemical OF CHROMATOGRAPHY Paper chromatography It is a beneficial

Biochemical techniques refer to a set of  assays, procedure and approaches that enable
scientists to analyze the materials found inside organisms and the chemical
reactions underlying life techniques.

Biochemistry related to applied field :

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Biochemistry is mixed with both
clinical and applied discipline that involves in  is both analytical & quantitative methods
in order  to explain the impact of
relationship in molecular level.

In biochemical research, experimental models are first
subjected to qualitative evaluation i.e., the complex device is damaged down
& components are separated & identified.

Application of biochemical techniques

It gives an opportunity for researchers and scientists to
discover the modern research techniques within the discipline of biochemistry .

It could be used each for separation & identity of
macromolecules & micro molecules.  In
biochemistry, the term macromolecule is applied to the four biopolymers (
nucleic acids, proteins , carbohydrates & lipids ) in addition to non-polymeric
molecules with high weight.

Biochemical techniques

1.       DIALYSIS

2.       CHROMATOGRAPHY

3.       ELECTROPHORESIS

4.       SPECTROPHOTOMETER

5.     
ULTRA-VIOLET
VISIBLE SPECTROSCOPY:

6.       CENTRIFUGATION

DIALYSIS:

This method involves small solutes diffuse from a low
concentration solution to high conc. solution across a semi permeable membrane
when equilibrium is reached.

principle:

The porous membrane selectively  pass small solute  with  maintaining of large solute in solution . it
is separation process  of molecules primarily
based on size .

Chromatography:

Chromatography is an analytical approach dealing with the
separation of closely related compound from a aggregate mixture . These include
protein, peptides, amino acid, lipids, carbohydrates, vitamins .

PRINCIPLE:

Chromatography 
generally consists of a cellular phase and a stationary phase. The
mobilized phase (cellular phase) refers to the combination of substances( to be
separate), dissolved in a liquid or a gas. The 
immobilized phase is a porous strong matrix via which the sample  pattern contained in the cell segment
percolates. The interaction between the mobile and stationary stages results
within the separation of the compounds from the mixture.

 TYPES OF  CHROMATOGRAPHY

Paper chromatography

It is  a beneficial
method in oreder to seprate the combination of amino acid, sugar, chemical
substances, lipid, urea in solution.  The
paper are used to aid a immobilized phase section at the same time as mobile
phase  moves down the suspended paper
strip in a beaker. Separation is primarily based on a liquid-liquid partition
of the compounds.

High performance Liquid Chromatography (HPLC)

It is used to separate 
elements from complex solution  by
the use of chemical interactions between the materials being analyzed & the
chromatography column.

Electrophoresis:

The motion of charged ions under the effect of electrical
field  that results in migration towards
the oppositely charged electrode is called electrophoresis

TYPES OF ELECTROPHORESIS

1.            Paper
electrophoresis,

2.            Gel
electrophoresis,

3.            isoelectric
focussing.

PAPER ELECTROPHORESIS

 Separation of charged
particles is determined by using differences in their migration size which
varies with electric chage , length of particle & form of particles.

GEL ELECTROPHORESIS,

It is used for the separation of proteins & nucleic
acid.  This technique helps for  separation of molecules based on their size
and charge.

ISOELECTRIC FOCUSSING.

Basically used for purification of proteins.  This technique is primarily based on the
immobilization of the molecules at isoelectric pH at some point of
electrophoresis.

SPECTROPHOTOMETER

It is a method to measure how much a chemical absorbs light,
by measuring the intensity of light as a beam of light passes through sample
solution.

 Also it gives the
quantitative dimension of the mirrored image molecules.

Ultra -violet visiblespectroscopy:

It refers to absorption spectroscopy or reflectance
spectroscopy in the ultraviolet spectra.

Principle :

It makes use of the visible spectrum .The absorption or
reflectance within the visible range witho impacts the perceived coloration of
the chemicals . on this region of electromagnetic spectrum , molecules go
through transition state. The absorption measures energy  transition from the low energy state to the high
energy state.

Centrifugation

Is a device for separating   particles from a solution according to there
length, shape, viscosity, density of the medium.

 It is used
to separate subcellular components and macromolecules by means of sedimenting
them below greater and varying gravitational pressure. The approach is important
for dedication of molecular weight of macromolecules as the sedimentation  size depends upon

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