COLUMN CHROMATOGRAPHY

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hai dear bloggers,

i m here with my blog titled as 'COLUMN CHROMATOGRAPHY from team "PHARMA WARRIORS" .

COLUMN CHROMATOGRAPHY "

column chromatography is a technique in which a column of
stationary phase is used.based upon the stationary phase nature,this
column chromatography is further divided into 2 types.they are if a
column of solid stationary phase is used it is called as column
adsorption chromatography.if a column of liquid stationary phase is used
then it is called as column partition chromatography.column adsorption
chromatography is widely used.(1)

PRINCIPLE:
adsorption is the principle of separation and here a solid
stationary phase and a liquid mobile phase is used.when a mixture of
different components which are dissolved in the mobile phase is
introduced in the column,the different components moves with different
rates depending upon their different relative affinities.the lesser
affinity compound which is towards the stationary phase moves faster and
it is eluted first.the greater affinity compound which is towards the
stationary phase is eluted later.thus in this way depending upon their
relative affinities the separation is done.(2)
the rate of movement of the compound(R)is given by the following formula:
R=rateof movement of a component/rate of movement of mobile phase (or)
R=distance travelled by the solute/distance travelled by the solvent(2)
if the mobile phase is liquid then
R=Am/Am+P.As
P=partition coefficient
Am=average cross section of mobile phase
As=average cross section of stationary phase(2)
INSTRUMENTATION:(3)
  • stationary phase
  • mobile phase
  • column characteristic
  • preparation of column
  • ntroduction of the sample
  • development technique
  • detection of components
  • recovery of components
STATIONARY PHASE:(ADSORBENT)(4)
The adsorbent used in this must have the following characteristics:
  • particle size and geometry:60-200microns
  • should have high mechanicsandhyasravyaal stability
  • should be inert
  • insoluble in the solvents.
  • it should be colorless
  • it should allow free flow of mobile phase
  • inexpensive
  • freely available
  • It should separate wide variety of components
TYPES OF ADSORBENTS:(4)
  • weak adsorbents:sucrose,starch,inulin,talc,sodium carbonate
  • medium adsorbentscaco3,mgco3,calcium hydroxide
  • strong adsorbents:activated alumina,act.charcoal,act magnesia
selection of stationary phase:
the selection depends of the following:
  • removal of impurities
  • no. of components to be separated
  • affinity differenced between components
  • length of the column used
  • quantity of the adsorbent used
MOBILE PHASE:(4)
the functions of the mobile phase are
  • to introduce the mixture into the column as a solvent
  • to develop the zones of separation as developing agent
  • to remove pure component out of the column as an eluent
different mobile phases used are
petroleum ether,cyclohexane,carbondisulphide,toluene,chloroform,benzene,acetone,alcohols etc.....
COLUMN CHARACTERISTICS:(5)
the column must be neutral such that it shoul not be affected by
acids ,alkalies or solvents.the length :diameter must range from
10:1-30:1.It must be 100:1 for more efficiency.
the length of the column depends upon
  • affinity of compounds towards the adsorbent used.
  • number of compounds to be separated
  • type of adsorbent used
  • quantity of the sample
PREPARATION OF THE COLUMN:(6)
the preparation of the column is done by 2 methods
  • dry packing tecnique
  • wet packing technique
in dry packing technique.the required quantity of adsorbent is
packed in the column in dry form and the solvent is allowed to flow
through the column.the demerits are formation of airbubbles and cracking
due to the complete dry packing of the coulmn.
in wet packing technique,the adsorbent is mixed with the mobile
phase solvent in a beaker and it is poured into the column.this is an
ideal process.
INTRODUCTION OF THE SAMPLE:(6)
The sample which consists of mixture of components is dissolved in
minimum quantity of mobile phase used for separation of the column.from
here samples can be separated individually by elution process.
DEVELOPMENT TECHNIQUE:(ELUTION)(7)
the elution techniques are
1.isocratic elution technique:in this technique,the same solvent composition is used throughout the separation process.
2.gradient elution techique:solvents of gradually increasing
polarity or increasing elution strength are used during separation
process.initially low polar solvent is used,gradually higher polar
solvent is used.
DETECTION OF COMPONENTS:(7)
Different properties that can be used are:
  • absorption of light using UV/VISIBLE detector
  • flourescence detector
  • flame ionisation detector
  • refractive index detector
  • evaporation of the solvent
RECOVERY OF THE COMPONENTS:(7)
earlier the recovery can be done by cutting the column into
distinct zones by means of a plunger.the best technique for recovery is
elution.the components are called as eluate and the solvent used is
called as eluent and the process is called a s elution.
FACTORS AFFECTING COLUMN EFFICIENCY:(5)
  • dimensions of the column
  • particle size of the adsorbent
  • nature of the solvent
  • temperature of the column
  • pressure
APPLICATIONS OF COLUMN CHROMATOGRAPHY:(8)
  • separation of different mixture of compounds
  • removal of impurities
  • isolation of active constituents
  • isolation of metabolite from biological fluids
  • estimation of drugs in formulations
ADVANTAGES OF COLUMN CHROMATOGRAPHY:(9)
  • Any type of mixture can be separated
  • any quantity of the mixture can be separated
  • wider choice of mobile phase
  • automation is possible
DISADVANTAGES OF COLUMN CHROMATOGRAPHY:(9)
  • time consuming method
  • more amount of solvents are required which are expensive
  • automation technique makes complication
"THIS BLOG DOES NOT CONTAIN ANY PLAGIARISED MATERIAL"
references:
3.a text book of pharmaceutical analysis by ravi shankar(accessed on 16th march,8:45p.m)
5,a text book of pharmaceutical analysis by ravi shankar(accessed on 16th march,8:45p.m)
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Comments

Ankur sood's picture

NICE BLOG AND VERY GOOD PRSENTATION . 1.CAN YOU TELL ME THE BEST MOBILE PHASE FOR COLUMN CHROMATOGRAPHY, GIVE ITS COMPOSITION WITH RATIO? WITH REGARDS ANKUR SOOD
Sandhya Sravya malla's picture

usually the mobile phase used serves as an eluent.the selection of mobile phase depends upon the separation technique employed and It is chosen in such a way that the retention factor value of the compound of interest is around the value 0.2 - 0.3 in order to reduce the time and the amount of eluent to run the chromatographic process. The eluent is also chosen so that the different compounds can be separated in an efficient manner. m.sandhyasravya@gmail.com PHARMA WARRIORS
Venkata Ram Sudhir Kakarlapudi's picture

specific application of column chromatography in pharmacy. how column chromatography useful in separation and identification of natural products.can u explain with examples regards kvr sudhir

KVRSUDHIR

Sirisha Pingali's picture

Hello sandhya I appreciate your effort in giving a good glance of the topic. Its very difficult to cover the entire topic in a single blog but you managed well. In packing of column, is dry packing preferred over wet packing? If yes, where it is used? Can you cite an example.

Sirisha Pingali

http://www.pharmainfo.net/sirisha

Viswanadha Institute of Pharmaceutical Sciences.

www.vnips.edu.in

Sandhya Sravya malla's picture

thank you very much for your comment and when coming to your question i mentioned that dry packing is not an ideal process when compared to wet packing,because when compared to wet packing dry packing has many demerits.hence dry packing is less preferred over wet packing.wet packing is an ideal process hope you got my point. m.sandhyasravya@gmail.com PHARMA WARRIORS

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