Chromatography is a laboratory technique used to separate components of a mixture through differential partitioning between a stationary and mobile phase. The key aspects are:
- Mixtures are separated based on how components partition between a mobile liquid or gas phase and a stationary solid or liquid phase.
- There are various types including adsorption chromatography which uses interactions between components and a solid stationary phase, partition chromatography which relies on differing solubilities in mobile and stationary liquid phases, and ion-exchange chromatography which separates based on charge.
- Factors like pH, salt concentration, temperature and column properties influence the separation in chromatography. It has many applications in analyzing compounds like drugs, proteins, sugars and more.
Content include basic introduction to chromatography. Brief view of Liquid Chromatography. HPLC introduction, other names, types of HPLC, detailed instrumentation with image of each part, and applications. Sources of content described in 'References' entitled slide. This presentation was prepared for the partial fulfillment of Master of Pharmacy.
Presenting a presentation on the topic of Column chromatography with including basics of chromatography, principles, equations, graphs and data related to it.
Topics which covered in this ppt is
Principle of chromatography
classification of chromatography
partition coefficient
chromatogram
Resolution
plate theory
determination of N
band zone broadening
rate theory
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Content include basic introduction to chromatography. Brief view of Liquid Chromatography. HPLC introduction, other names, types of HPLC, detailed instrumentation with image of each part, and applications. Sources of content described in 'References' entitled slide. This presentation was prepared for the partial fulfillment of Master of Pharmacy.
Presenting a presentation on the topic of Column chromatography with including basics of chromatography, principles, equations, graphs and data related to it.
Topics which covered in this ppt is
Principle of chromatography
classification of chromatography
partition coefficient
chromatogram
Resolution
plate theory
determination of N
band zone broadening
rate theory
https://www.linkedin.com/in/preeti-choudhary-266414182/
https://www.instagram.com/chaudharypreeti1997/
https://www.facebook.com/profile.php?id=100013419194533
https://twitter.com/preetic27018281
Please like, share, comment and follow.
stay connected
If any query then contact:
chaudharypreeti1997@gmail.com
Thanking-You
Preeti Choudhary
What is Chromatography?
Applications of Chromatography
Types of Chromatography
1- Column Chromatography
2- Planar chromatography
Paper Chromatography
Gas Chromatography
Detectors
• Chromatography is a method of separation in which the components to be separated are distributed between two phases, one of these is called a stationary phase and the other is a mobile phase which moves on stationary phase in a definite direction
Chromatography is an important biophysical technique that enables the separation, identification, and purification of the components of a mixture for qualitative and quantitative analysis.
The Russian botanist Mikhail Tswett coined the term chromatography in 1906.
The first analytical use of chromatography was described by James and Martin in 1952, for the use of gas chromatography for the analysis of fatty acid mixtures.
A wide range of chromatographic procedures makes use of differences in size, binding affinities, charge, and other properties to separate materials.
It is a powerful separation tool that is used in all branches of science and is often the only means of separating components from complex mixtures.
hromatography is based on the principle where molecules in mixture applied onto the surface or into the solid, and fluid stationary phase (stable phase) is separating from each other while moving with the aid of a mobile phase.
The factors effective on this separation process include molecular characteristics related to adsorption (liquid-solid), partition (liquid-solid), and affinity or differences among their molecular weights.
Because of these differences, some components of the mixture stay longer in the stationary phase, and they move slowly in the chromatography system, while others pass rapidly into the mobile phase, and leave the system faster.
Three components thus form the basis of the chromatography technique.
1. Stationary phase: This phase is always composed of a “solid” phase or “a layer of a liquid adsorbed on the surface solid support”.
2. Mobile phase: This phase is always composed of “liquid” or a “gaseous component.”
3. Separated molecules
Types of Chromatography
Substances can be separated on the basis of a variety of methods and the presence of characteristics such as size and shape, total charge, hydrophobic groups present on the surface, and binding capacity with the stationary phase.
This leads to different types of chromatography techniques, each with their own instrumentation and working principle.
For instance, four separation techniques based on molecular characteristics and interaction type use mechanisms of ion exchange, surface adsorption, partition, and size exclusion.
Other chromatography techniques are based on the stationary bed, including column, thin layer, and paper chromatography.
Applications of Chromatography
Pharmaceutical sector
To identify and analyze samples for the presence of trace elements or chemicals.
Separation of compounds based on their molecular weight and element composition.
Detects the unknown compounds and purity of mixture.
In drug development.
Chemical industry
In testing water samples and also checks air quality.
HPLC and GC are very much used for detecting various contaminants such as polychlorinated biphenyl (PCBs) in pesticides and oils.
In various life sciences applications.
In forensic pathology and crime scene testing like analyzing blood and hair samples.
powerpoint presentation on high performance liquid chromatography which include its definition, classification, principles of seperation, instrumentation and application.
2024.06.01 Introducing a competency framework for languag learning materials ...Sandy Millin
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Published classroom materials form the basis of syllabuses, drive teacher professional development, and have a potentially huge influence on learners, teachers and education systems. All teachers also create their own materials, whether a few sentences on a blackboard, a highly-structured fully-realised online course, or anything in between. Despite this, the knowledge and skills needed to create effective language learning materials are rarely part of teacher training, and are mostly learnt by trial and error.
Knowledge and skills frameworks, generally called competency frameworks, for ELT teachers, trainers and managers have existed for a few years now. However, until I created one for my MA dissertation, there wasn’t one drawing together what we need to know and do to be able to effectively produce language learning materials.
This webinar will introduce you to my framework, highlighting the key competencies I identified from my research. It will also show how anybody involved in language teaching (any language, not just English!), teacher training, managing schools or developing language learning materials can benefit from using the framework.
This presentation includes basic of PCOS their pathology and treatment and also Ayurveda correlation of PCOS and Ayurvedic line of treatment mentioned in classics.
How to Build a Module in Odoo 17 Using the Scaffold MethodCeline George
Odoo provides an option for creating a module by using a single line command. By using this command the user can make a whole structure of a module. It is very easy for a beginner to make a module. There is no need to make each file manually. This slide will show how to create a module using the scaffold method.
Safalta Digital marketing institute in Noida, provide complete applications that encompass a huge range of virtual advertising and marketing additives, which includes search engine optimization, virtual communication advertising, pay-per-click on marketing, content material advertising, internet analytics, and greater. These university courses are designed for students who possess a comprehensive understanding of virtual marketing strategies and attributes.Safalta Digital Marketing Institute in Noida is a first choice for young individuals or students who are looking to start their careers in the field of digital advertising. The institute gives specialized courses designed and certification.
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This slide is special for master students (MIBS & MIFB) in UUM. Also useful for readers who are interested in the topic of contemporary Islamic banking.
it describes the bony anatomy including the femoral head , acetabulum, labrum . also discusses the capsule , ligaments . muscle that act on the hip joint and the range of motion are outlined. factors affecting hip joint stability and weight transmission through the joint are summarized.
A workshop hosted by the South African Journal of Science aimed at postgraduate students and early career researchers with little or no experience in writing and publishing journal articles.
Acetabularia Information For Class 9 .docxvaibhavrinwa19
Acetabularia acetabulum is a single-celled green alga that in its vegetative state is morphologically differentiated into a basal rhizoid and an axially elongated stalk, which bears whorls of branching hairs. The single diploid nucleus resides in the rhizoid.
MATATAG CURRICULUM: ASSESSING THE READINESS OF ELEM. PUBLIC SCHOOL TEACHERS I...NelTorrente
In this research, it concludes that while the readiness of teachers in Caloocan City to implement the MATATAG Curriculum is generally positive, targeted efforts in professional development, resource distribution, support networks, and comprehensive preparation can address the existing gaps and ensure successful curriculum implementation.
June 3, 2024 Anti-Semitism Letter Sent to MIT President Kornbluth and MIT Cor...Levi Shapiro
Letter from the Congress of the United States regarding Anti-Semitism sent June 3rd to MIT President Sally Kornbluth, MIT Corp Chair, Mark Gorenberg
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The US House of Representatives is deeply concerned by ongoing and pervasive acts of antisemitic
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2. CHROMATOGRAPHY
A laboratory technique that separates
components within a mixture by using the
differential affinities of the components
for a mobile medium and for a stationary
adsorbing medium through which they
pass.
3. CHROMATOGRAPHY
Introduced first by the Russian botanist
Mikhail Semenovich Tswett.
Mixtures of solutes dissolved in a common
solvent are separated from one another by
a differential distribution of the solutes
between two phases.
4. CHROMATOGRAPHY
Two phases in chromatography are:
mobile phase is part of the chromatographic
system which carries the solutes through the
stationary phase.
stationary phase is the part which the mobile
phase flows where the distribution of the
solutes between the phases occurs.
5. PRINCIPLE
Fractionalism of mixtures of
substances
In the operation of the
chromatogram, a mobile gaseous or
liquid phase is use to wash the
substances to be separated through a
column of a porous material.
6. PRINCIPLE
Capillary Action – the movement of
liquid within the spaces of a material
due to the forces of adhesion,
cohesion, and surface tension.
7. PRINCIPLE
The rate of migration of the solute
depends upon the rate of interaction
of the solute with the two phases, one
being the mobile phases and the
other stationary phase as the
compounds travel through the
supporting medium.
8. PRINCIPLE
The rate of migration of the solute
depends upon the rate of interaction
of the solute with the two phases, one
being the mobile phases and the
other stationary phase as the
compounds travel through the
supporting medium.
9. MECHANISMS OF SEPARATION
IN CHROMATOGRAPHY
• Based on the interactions of solutes with
mobile and stationary phases.
10. ADSORPTION (LIQUID-SOLID)
CHROMATOGRAPHY
Based on the competition
between the sample and the
mobile phase for binding
sites of the solid (stationary)
phase. Molecules that are
soluble in the mobile phase
move fastest.
11. ADSORPTION (LIQUID-SOLID)
CHROMATOGRAPHY
Advantages
An extensive separation
literature is available on thin
layer chromatography
methods that are readily
transferable to adsorption
The flexibility, speed, and low
cost of TLC allow its use in
experimental development.
12. ADSORPTION (LIQUID-SOLID)
CHROMATOGRAPHY
Advantages
TLC has great value for use in
the preliminary investigation
of samples of unknown
constituents
Adsorption chromatography,
particularly with silica gel, has
been widely used for the
separation of drugs in both the
HPLC and TLC modes.
13. PARTITION (LIQUID-LIQUID)
CHROMATOGRAPHY
separates molecules on
the basis of sample
volatility.
Depends on the solubility
of the solute in nonpolar
(organic) or polar
(aqueous) solvents
14. PARTITION (LIQUID-LIQUID)
CHROMATOGRAPHY
Advantage: the
stationary phase does not
leave the solid support
and bleed into the
detector, and a uniform
monomolecular layer of
the stationary phase is
obtained through the
bonding procedure.
15. PARTITION (LIQUID-LIQUID)
CHROMATOGRAPHY
Since the chemical
influence of the solid
support may be largely
ignored, the adhering
film behaves essentially
like a liquid stationary
phase.
16. ION-EXCHANGE CHROMATOGRAPHY
∞ Based on the net charge
of molecules
∞ It is one of the common
types of separation
mechanism which
depends on the nature of
the stationary phase.
∞ It has 2 prinicipal types
of ion- exchanger is
cationic and anionic.
17. ION-EXCHANGE CHROMATOGRAPHY
∞ Ion exchange matrices
can be further
categorized as either
strong or weak.
∞ It separates amino acid
by electric charges based
on their respective
changes
18. ION-EXCHANGE CHROMATOGRAPHY
∞ Usually performed in
columns
∞ Uses a charged
stationary phase to
separate charged
compounds including
anions, cations, amino
acids, peptides and
proteins.
20. ION-EXCHANGE CHROMATOGRAPHY
∞ PRINCIPLE:
∞ Once the solutes are bound, the
column is washed to equilibrate it in
the starting buffer,which should be of
low ionic strength
∞ Then the bound molecules are eluted
off using a gradient be of low ionic
strength
22. ION-EXCHANGE CHROMATOGRAPHY
Factors to be considered in Ion Exchange
Chromatography:
⓭ Buffers – use anionic buffers for cation exchange
and cationic buffers for anion exchange to avoid
difficulty.
⓭ pH- influence the charge on the macromoloecules
in solution.
23. ION-EXCHANGE CHROMATOGRAPHY
Factors to be considered in Ion Exchange
Chromatography:
⓭ Salts to use for elution
Ions of the eluting salt must displace other
molecules from the charged groups on the stationary
phase with either a gradient or step in the 0 to 1.0
range.
25. ION-EXCHANGE CHROMATOGRAPHY
Most molecules have a net
charge within a pH range of 2 to
10. When the pH is altered, the
net charge on molecules can
change drastically.
In this experiment, a mixture of two
chemicals is absorbed onto a solid
support ion-exchange column and
separated during elution under
conditions that influence their net
charge.
28. ION-EXCHANGE CHROMATOGRAPHY
ADVANTAGES: DISADVANTAGES:
Long Life of Resins Nature and properties of
Cheap maintenance ion exchange resins
Environmental friendly Nature of exchanging ions
because it deals only with There are substances (such
substances occurring in as organic matter or Fe3+
water. occurring in some water
which can foul the resin.
30. Applications:
Serves as liquid chromatography detectors
and as quality control monitors in drug
manufactures
Also occurs in air and water quality, medical
and clinical laboratories and industrial
laboratories
32. By chromatographic bed shape
A. Column Chromatography
• A separation technique in which the
stationary bed is within a tube
• It works on a much larger scale by
packing the same materials into a
vertical glass column.
34. By chromatographic bed shape
B. Plane Chromatography
• A separation that takes place on a flat surface or
a plane
Example:
• Paper Chromatography
• Thin Layer Chromatography
35. Paper chromatography
Based on nature of
solvent, solubility of
solute and rate of
diffusion.
Uses paper as the
stationary phase and a
solvent as the mobile
phase.
36. Paper chromatography
Solvent moves through
the paper by a capillary
action
Separation depends on
the solubility of solute
and solvents, the polarity
of solvent, and polarity of
solutes in the sample.
38. Paper chromatography
Considered to be the simplest
and the most widely used of
the chromatographic
techniques because its
APPLICABILITY TO THE
FOLLOWING:
ISOLATION
IDENTIFICATION
AND QUANTITATIVE
DETERMINATION OF
ORGANIC AND INORGANIC
COMPOUNDS
39. Instrumentation
of Paper chromatography
1) Lid
2) Paper
3) Solvent
Front
4) Solvent
40. THIN-LAYER CHROMATOGRAPHY
Used as a semi-quantitative screening test
screening test
Uses as thin layer of silica gel, alumina gel,
polyacrylamide gel, or starch gel attached
to glass plate as stationary phase and the
mobile phase is liquid solvent.
41. THIN-LAYER CHROMATOGRAPHY
Fractions in the sample are generally quite
soluble in the solvent and move with it up the
stationary phase by capillary action.
Separated fractions are also developed in TLC by
applying a chemical reaction with the separated
fractions to produce color changes
43. THIN-LAYER CHROMATOGRAPHY
ADVANTAGE: DISADVANTAGE:
Simple and economical Spots are often faint
Easy to perform since it TLC is difficult to
only involves spotting
reproduce
the stationary phase
with the sample & Not typically
placing one edge of the automated
stationary phase plate in
the mobile phase
reservoir.
45. By Physical State of Mobile Phase
Gas Chromatography
• It can separate nanograms or pictograms of
volatile substances.
• It is principally a method for the separation
and quantitative determination of gases and
volatile liquids and substances.
46. Gas Chromatography
Volatile compounds can be separated in a
gas chromatograph, in which the mobile
phase is usually a relatively unreactive
carrier gas such as helium, nitrogen or
hydrogen.
Separations can be carried out in the vapor
phase, most parts of a gas chromatograph
are temperature controlled; selection of
temperature is based on the composition of
the sample.
47. Gas Chromatography
It uses a special detector according to
the different kinds of compound and
the most widely used are:
A. Mass spectrophotometer
B. Thermal Conductivity
C. Flame Ionization Detector
D. Electron Capture Detector
48. GAS CHROMATOGRAPHY
Applications:
Most effectively used for analyses of organic
compounds, space related, complex mixtures
of volatile substances at column temperature
of less than -40 °C to greater than 550° C.
Geochemical research projects such as
determination of various environmental
pollutants at extremely low concentrations.
49. GAS CHROMATOGRAPHY
ADVANTAGES: DISADVANTAGES:
Ability to provide LIMITED to volatile
qualitative information and samples
quantitative information Not suitable for
thermally labile samples
FAST ANALYSIS
Fairly difficult for large
Efficient, providing high preparative samples
resolution Requires spectroscopy
Sensitive usually mass
Nondestructive spectroscopy for
confirmation of peak
Requires small samples identity
Inexpensive
50.
51. COMPONENTS
Autosampler- provides the means to introduce a
sample automatically into the inlets. Automatic
insertion provides better reproducibility and time-
optimization.
Column inlet (or injector)- provides the means to
introduce a sample into a continuous flow of
carrier gas. The inlet is a piece of hardware
attached to the column head.
52. COMPONENTS
Carrier Gas (mobile phase) - must be
chemically inert, include helium, hydrogen
and nitrogen. It should be of high purity, and
the flow must be tightly controlled to ensure
optimum column efficiency and
reproducibility of test results.
Detector
53. GAS CHROMATOGRAPHY
GAS-LIQUID GAS-SOLID
CHROMATOGRAPHY CHROMATOGRAPHY
Separates molecules on Uses a solid material as an
the basis of sample absorbent
volatility Based upon a solid
Mobile phase is a gas such
stationary phase on which
retention of analytes is the
as helium and the consequence of physical
stationary phase is a high adsorption
boiling point liquid
Relies upon a large
absorbed onto a solid. granular surface to aid in
the separation of the
substances.
55. Application
Use in biomedical research, routine clinical
determination and drug researching
programs
56. LIQUID CHROMATOGRAPHY
The mobile phase is percolated through the
column by means of either gravity , under
pressure generated by a suitable pump or
centrifugal force.
57. SIZE EXCLUSION CHROMATOGRAPHY
Particles of different size
will elute (filter) through
a stationary phase at
different rates. This
results in the separation
of a solution of particles
based on size. Provided,
that all the particles are
loaded simultaneously or
near – simultaneously of
the same size should
elute together.
58. SIZE EXCLUSION CHROMATOGRAPHY
The support material has certain range of
pore sizes. As solutes travel through, the
small molecules can enter the pores, whereas
the larger ones cannot and will elute first
from column.
The determination of molecular weight, e.g. ,
of enzymes, and estimation of equilibrium
constants can be achieved with relative ease
59. SIZE EXCLUSION CHROMATOGRAPHY
ADVANTAGES DISADVANTAGES
RAPID ROUTINE ANALYSIS FILTRATIONS MUST BE
PERFORMED BEFORE USING
IDENTIFYING HIGH MASS THE INSTRUMENT BAD
COMPONENTS EVEN IN RESPONSE FOR VERY
LOW CONCENTRATION SMALL MOLECULAR
WEIGHTS
CAN ANALYZE STANDARDS ARE NEEDED
POLYDISPERSED
SENSITIVE FOR FLOW RATE
SAMPLES,BRANCHING VARIATION. INTERNAL
STUDIES CAN BE DONE, STANDARD SHOULD BE
ABSOLUTE MOLECULAR USED WHENEVER POSSIBLE
WEIGHTS CAN BE HIGH INVESTMENT COST
OBTAINED.
62. LIQUID CHROMATOGRAPHY
High-performance liquid
chromatography (HPLC)
Uses a pressure for the pumping of aqueous
or organic solution through a column.
The mobile phase is forced under pressure
through a long, narrow column, yielding an
excellent separation in a relatively short time.
Highly sensitive and specific.
64. LIQUID CHROMATOGRAPHY
High-Performance Liquid Chromatography
ADVANTAGES: DISADVANTAGES:
An automated process that Difficult to detect coelution,
takes only a few minutes to which may lead to inacurrate
produce results. compound categorization.
Uses gravity instead of high High cost for equipment
speed pump to force
compounds through the needed to conduct HPLC.
densely packed tubing. Operation is complex, requiring
Results are of high resolution a trained technician to operate.
and are easy to read. Equipment has low sensitivity
Can be reproduce easily via to some compounds because
automated process. of the speed of the process.
65. Application
Use in biomedical research, routine clinical
determination and drug researching
programs
66. HIGH PERFORMANCE LIQUID
CHROMATOGRAPHY
Instruments
Fraction Collector
Auto Sampler
Pumping systems
Columns & Packing
Detectors
Control Data & Processing
68. Application
Use in monitoring the use of therapeutic
drugs and detecting drug abuse.
also use to separate compounds contributing
to the fragrance of flowers