Partition Chromatography technique is defined as. the separation of components between two liquid phases viz original solvent and the film of solvent used in the column.
ION EXCHANGE CHROMATOGRAPHY
By M.Vharshini
B.Sc. Bio Medical Science
Sri Ramachandra University
ION EXCHANGE CHROMATOGRAPHY
Ion-exchange chromatography is a process that allows the separation of ions and polar molecules based on their affinity to the ion exchanger.
It can be used for almost any kind of charged molecule including large proteins, small nucleotides and amino acids.
Cations or Anions can be separated using this method.
PRINCIPLE
It is based on the reversible electrostatic interaction of ions with the separation matrix (i.e.)
The separation occurs by reversible exchange of ions between the ions present in the solution and those present in the ion exchange resin.
CLASSIFICATION OF RESINS
According to the chemical nature they classified as-
1. Strong cation exchange resin
2. Weak cation exchange resin
3. Strong anion exchange resin
4. Weak anion exchange resin
According to the Source they can -
Natural resins : Cation - Zeolytes, Clay
Anion - Dolomite
Synthetic resins: Inorganic & Organic resins
◘Organic resins are polymeric resin matrix.
The resin composed of –
Polystyrene (sites for exchangeable functional groups)
Divinyl benzene(Cross linking agent)-offers stability.
Ion exchange resin should have following requirements
»It must be chemically stable.
»It should be insoluble in common solvents.
» It should have a sufficient degree of cross linking.
»The swollen resin must be denser than water.
»It must contain sufficient no. of ion exchange groups.
Physical properties of ion exchange resins
Cross linking:
It affects swelling & strength & solubility
Swelling:
When resin swells, polymer chain spreads apart
Polar solvents → swelling
Non-polar solvents → contraction
Swelling also affected electrolyte concentration.
Particle size and porosity
Increase in surface area & decrease in particle size will increase the rate of ion exchange.
Regeneration
Cation exchange resin are regenerated by treatment with acid, then washing with water.
Anion exchange resin are regenerated by treatment with NaOH, then washing with water until neutral.
EXPERIMENTAL SETUP OF ION EXCHANGE CHROMATOGRAPHY
Metrohm 850 Ion chromatography system
Instrumentation of ion exchange chromatography
PRACTICAL REQUIREMENTS
1.Column
» glass, stainless steel or polymers
2.Packing the column
» Wet packing method:
A slurry is prepared of the eluent with the stationary phase powder and then carefully poured into the column. Care must be taken to avoid air bubbles.
3.Application of the sample
After packing, sample is added to the top of the stationary phase, use syringe or pipette.
This layer is usually topped with a small layer of sand or with cotton or glass wool to protect the shape of the organic layer from the velocity of newly added eluent.
4.Mobile phase
Acids, alkalis, buffers…
6.Stationary phase
The ionic
HPLC Principle,Instrumentation and ApplicationAlakesh Pradhan
HPLC Chromatography and its principle
Liquid chromatography
High Performance Liquid Chromatography ( HPLC )
The components of the high performance liquid chromatograph (HPLC).
The separation process.
The chromatogram
Column Chromatography: basics of chromatography and principle of chromatography, Classification, partition coefficient , chromatogram, retention time and volume, capacity and selectivity factors, plate theory, band broadening, rate theory, mass transfer, packed GLC column, open tubular column capillary columns, liquid chromatography column resolution.
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This presentation contains all the topics related to column chromatography. That includes introduction, principle,apparatus, experimental aspects of column chromatography, application of column chromatography, advantage and disadvantage of column chromatography with reference.
A presentation on column efficiency parameters in chromatography.. A part of gas chromatography in pharmacutical analysis..will be helpful for all mphrm students
ION EXCHANGE CHROMATOGRAPHY
By M.Vharshini
B.Sc. Bio Medical Science
Sri Ramachandra University
ION EXCHANGE CHROMATOGRAPHY
Ion-exchange chromatography is a process that allows the separation of ions and polar molecules based on their affinity to the ion exchanger.
It can be used for almost any kind of charged molecule including large proteins, small nucleotides and amino acids.
Cations or Anions can be separated using this method.
PRINCIPLE
It is based on the reversible electrostatic interaction of ions with the separation matrix (i.e.)
The separation occurs by reversible exchange of ions between the ions present in the solution and those present in the ion exchange resin.
CLASSIFICATION OF RESINS
According to the chemical nature they classified as-
1. Strong cation exchange resin
2. Weak cation exchange resin
3. Strong anion exchange resin
4. Weak anion exchange resin
According to the Source they can -
Natural resins : Cation - Zeolytes, Clay
Anion - Dolomite
Synthetic resins: Inorganic & Organic resins
◘Organic resins are polymeric resin matrix.
The resin composed of –
Polystyrene (sites for exchangeable functional groups)
Divinyl benzene(Cross linking agent)-offers stability.
Ion exchange resin should have following requirements
»It must be chemically stable.
»It should be insoluble in common solvents.
» It should have a sufficient degree of cross linking.
»The swollen resin must be denser than water.
»It must contain sufficient no. of ion exchange groups.
Physical properties of ion exchange resins
Cross linking:
It affects swelling & strength & solubility
Swelling:
When resin swells, polymer chain spreads apart
Polar solvents → swelling
Non-polar solvents → contraction
Swelling also affected electrolyte concentration.
Particle size and porosity
Increase in surface area & decrease in particle size will increase the rate of ion exchange.
Regeneration
Cation exchange resin are regenerated by treatment with acid, then washing with water.
Anion exchange resin are regenerated by treatment with NaOH, then washing with water until neutral.
EXPERIMENTAL SETUP OF ION EXCHANGE CHROMATOGRAPHY
Metrohm 850 Ion chromatography system
Instrumentation of ion exchange chromatography
PRACTICAL REQUIREMENTS
1.Column
» glass, stainless steel or polymers
2.Packing the column
» Wet packing method:
A slurry is prepared of the eluent with the stationary phase powder and then carefully poured into the column. Care must be taken to avoid air bubbles.
3.Application of the sample
After packing, sample is added to the top of the stationary phase, use syringe or pipette.
This layer is usually topped with a small layer of sand or with cotton or glass wool to protect the shape of the organic layer from the velocity of newly added eluent.
4.Mobile phase
Acids, alkalis, buffers…
6.Stationary phase
The ionic
HPLC Principle,Instrumentation and ApplicationAlakesh Pradhan
HPLC Chromatography and its principle
Liquid chromatography
High Performance Liquid Chromatography ( HPLC )
The components of the high performance liquid chromatograph (HPLC).
The separation process.
The chromatogram
Column Chromatography: basics of chromatography and principle of chromatography, Classification, partition coefficient , chromatogram, retention time and volume, capacity and selectivity factors, plate theory, band broadening, rate theory, mass transfer, packed GLC column, open tubular column capillary columns, liquid chromatography column resolution.
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
This presentation contains all the topics related to column chromatography. That includes introduction, principle,apparatus, experimental aspects of column chromatography, application of column chromatography, advantage and disadvantage of column chromatography with reference.
A presentation on column efficiency parameters in chromatography.. A part of gas chromatography in pharmacutical analysis..will be helpful for all mphrm students
• 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 : A seperation techniqueSHIVANEE VYAS
Chromatography is a method of seperating mixture of components into individual components through equlibrium distribution between two phases.
Each chromatographic method essentially consists of 2 phases a staionary phase and a mobile phase.
Stationary phase : solid or liquid
Mobile phase : liquid or gas
Paper chromatography is an analytical method used to separate colored chemicals or substances. It is primarily used as a teaching tool, having been replaced by other chromatography methods, such as thin-layer chromatography.
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.
Chromatography is a bioanalytical technique used for separation of analytes into pure components. Biomolecules such as amino acids, proteins and carbohydrates can be purified by different chromatographic methods.
Cancer is a group of diseases involving abnormal cell growth with the potential to invade or spread to other parts of the body. These contrast with benign tumors, which do not spread. Possible signs and symptoms include a lump, abnormal bleeding, prolonged cough, unexplained weight loss, and a change in bowel movements. While these symptoms may indicate cancer, they can also have other causes. Over 100 types of cancers affect humans.
AAS is an analytical technique used to determine how much of certain elements are in a sample. It uses the principle that atoms (and ions) can absorb light at a specific, unique wavelength. When this specific wavelength of light is provided, the energy (light) is absorbed by the atom.
Strain improvement is one element of fermentation process management. It is the process of increasing the productivity of a microorganism by improving or selecting for a more productive phenotype.
Eukaryotic transcription is carried out in the nucleus of the cell and proceeds in three sequential stages: initiation, elongation, and termination. Eukaryotes require transcription factors to first bind to the promoter region and then help recruit the appropriate polymerase.
A mutation is a change in the DNA sequence of an organism. Mutations can result from errors in DNA replication during cell division, exposure to mutagens or a viral infection.2
A DNA vaccine is a type of vaccine that transfects a specific antigen-coding DNA sequence into the cells of an organism as a mechanism to induce an immune response.
DNA vaccines work by injecting genetically engineered plasmid containing the DNA sequence encoding the antigen(s) against which an immune response is sought, so the cells directly produce the antigen, thus causing a protective immunological response.
Bioremediation is a branch of biotechnology that employs the use of living organisms, like microbes and bacteria, in the removal of contaminants, pollutants, and toxins from soil, water, and other environments.
radioactivity is the act of emitting radiation spontaneously. This is done by an atomic nucleus that, for some reason, is unstable; it "wants" to give up some energy in order to shift to a more stable configuration.
Hypersensitivity reactions are exaggerated or inappropriate immunologic responses occurring in response to an antigen or allergen. Type I, II and III hypersensitivity reactions are known as immediate hypersensitivity reactions because they occur within 24 hours of exposure to the antigen or allergen.
Diphtheria is a serious infection caused by strains of bacteria called Corynebacterium diphtheriae that make toxin. It can lead to difficulty breathing, heart rhythm problems, and even death. CDC recommends vaccines for infants, children, teens, and adults to prevent diphtheria. Causes and How It Spreads.
Clostridium is a genus of anaerobic, Gram-positive bacteria. Species of Clostridium inhabit soils and the intestinal tract of animals, including humans. This genus includes several significant human pathogens, including the causative agents of botulism and tetanus.
transposon, class of genetic elements that can “jump” to different locations within a genome. Although these elements are frequently called “jumping genes,” they are always maintained in an integrated site in the genome. In addition, most transposons eventually become inactive and no longer move.1
Gene regulation is the process used to control the timing, location and amount in which genes are expressed. The process can be complicated and is carried out by a variety of mechanisms, including through regulatory proteins and chemical modification of DNA.
Genetic recombination (genetic reshuffling) is the exchange of genetic material between different organisms which leads to production of offspring with combinations of traits that differ from those found in either parent. The process occurs naturally and can also be carried out in the lab.
Mycotoxins are naturally occurring toxins produced by certain moulds (fungi) and can be found in food.
The moulds grow on a variety of different crops and foodstuffs including cereals, nuts, spices, dried fruits, apples and coffee beans, often under warm and humid conditions.
Mycotoxins can cause a variety of adverse health effects and pose a serious health threat to both humans and livestock.
Microbiology essentially began with the development of the microscope. Although others may have seen microbes before him, it was Antonie van Leeuwenhoek, a Dutch draper whose hobby was lens grinding and making microscopes, who was the first to provide proper documentation of his observations.
We can make various products like soup powder, papad, nuggets, chips, preserve, candy etc. using different mushrooms. products like pasta, noodles etc. by supplementing with fresh or dried mushroom powder.
The microbes are highly useful for making vaccines and antibiotics for making medicines. It is a well-known fact that harmful pathogens that cause different diseases by infecting our body. The antibiotics and medicines would help us in fighting these diseases and infections.
A genetically modified organism (GMO) is any organism whose genetic material has been altered using genetic engineering techniques. The exact definition of a genetically modified organism and what constitutes genetic engineering varies, with the most common being an organism altered in a way that "does not occur naturally by mating and/or natural recombination". A wide variety of organisms have been genetically modified (GM), from animals to plants and microorganisms.
Biofertilizers are living microbes that enhance plant nutrition by either by mobilizing or increasing nutrient availability in soils. Various microbial taxa including beneficial bacteria and fungi are currently used as biofertilizers, as they successfully colonize the rhizosphere, rhizoplane or root interior.
Slide 1: Title Slide
Extrachromosomal Inheritance
Slide 2: Introduction to Extrachromosomal Inheritance
Definition: Extrachromosomal inheritance refers to the transmission of genetic material that is not found within the nucleus.
Key Components: Involves genes located in mitochondria, chloroplasts, and plasmids.
Slide 3: Mitochondrial Inheritance
Mitochondria: Organelles responsible for energy production.
Mitochondrial DNA (mtDNA): Circular DNA molecule found in mitochondria.
Inheritance Pattern: Maternally inherited, meaning it is passed from mothers to all their offspring.
Diseases: Examples include Leber’s hereditary optic neuropathy (LHON) and mitochondrial myopathy.
Slide 4: Chloroplast Inheritance
Chloroplasts: Organelles responsible for photosynthesis in plants.
Chloroplast DNA (cpDNA): Circular DNA molecule found in chloroplasts.
Inheritance Pattern: Often maternally inherited in most plants, but can vary in some species.
Examples: Variegation in plants, where leaf color patterns are determined by chloroplast DNA.
Slide 5: Plasmid Inheritance
Plasmids: Small, circular DNA molecules found in bacteria and some eukaryotes.
Features: Can carry antibiotic resistance genes and can be transferred between cells through processes like conjugation.
Significance: Important in biotechnology for gene cloning and genetic engineering.
Slide 6: Mechanisms of Extrachromosomal Inheritance
Non-Mendelian Patterns: Do not follow Mendel’s laws of inheritance.
Cytoplasmic Segregation: During cell division, organelles like mitochondria and chloroplasts are randomly distributed to daughter cells.
Heteroplasmy: Presence of more than one type of organellar genome within a cell, leading to variation in expression.
Slide 7: Examples of Extrachromosomal Inheritance
Four O’clock Plant (Mirabilis jalapa): Shows variegated leaves due to different cpDNA in leaf cells.
Petite Mutants in Yeast: Result from mutations in mitochondrial DNA affecting respiration.
Slide 8: Importance of Extrachromosomal Inheritance
Evolution: Provides insight into the evolution of eukaryotic cells.
Medicine: Understanding mitochondrial inheritance helps in diagnosing and treating mitochondrial diseases.
Agriculture: Chloroplast inheritance can be used in plant breeding and genetic modification.
Slide 9: Recent Research and Advances
Gene Editing: Techniques like CRISPR-Cas9 are being used to edit mitochondrial and chloroplast DNA.
Therapies: Development of mitochondrial replacement therapy (MRT) for preventing mitochondrial diseases.
Slide 10: Conclusion
Summary: Extrachromosomal inheritance involves the transmission of genetic material outside the nucleus and plays a crucial role in genetics, medicine, and biotechnology.
Future Directions: Continued research and technological advancements hold promise for new treatments and applications.
Slide 11: Questions and Discussion
Invite Audience: Open the floor for any questions or further discussion on the topic.
Richard's entangled aventures in wonderlandRichard Gill
Since the loophole-free Bell experiments of 2020 and the Nobel prizes in physics of 2022, critics of Bell's work have retreated to the fortress of super-determinism. Now, super-determinism is a derogatory word - it just means "determinism". Palmer, Hance and Hossenfelder argue that quantum mechanics and determinism are not incompatible, using a sophisticated mathematical construction based on a subtle thinning of allowed states and measurements in quantum mechanics, such that what is left appears to make Bell's argument fail, without altering the empirical predictions of quantum mechanics. I think however that it is a smoke screen, and the slogan "lost in math" comes to my mind. I will discuss some other recent disproofs of Bell's theorem using the language of causality based on causal graphs. Causal thinking is also central to law and justice. I will mention surprising connections to my work on serial killer nurse cases, in particular the Dutch case of Lucia de Berk and the current UK case of Lucy Letby.
THE IMPORTANCE OF MARTIAN ATMOSPHERE SAMPLE RETURN.Sérgio Sacani
The return of a sample of near-surface atmosphere from Mars would facilitate answers to several first-order science questions surrounding the formation and evolution of the planet. One of the important aspects of terrestrial planet formation in general is the role that primary atmospheres played in influencing the chemistry and structure of the planets and their antecedents. Studies of the martian atmosphere can be used to investigate the role of a primary atmosphere in its history. Atmosphere samples would also inform our understanding of the near-surface chemistry of the planet, and ultimately the prospects for life. High-precision isotopic analyses of constituent gases are needed to address these questions, requiring that the analyses are made on returned samples rather than in situ.
Comparing Evolved Extractive Text Summary Scores of Bidirectional Encoder Rep...University of Maribor
Slides from:
11th International Conference on Electrical, Electronics and Computer Engineering (IcETRAN), Niš, 3-6 June 2024
Track: Artificial Intelligence
https://www.etran.rs/2024/en/home-english/
Professional air quality monitoring systems provide immediate, on-site data for analysis, compliance, and decision-making.
Monitor common gases, weather parameters, particulates.
This presentation explores a brief idea about the structural and functional attributes of nucleotides, the structure and function of genetic materials along with the impact of UV rays and pH upon them.
A brief information about the SCOP protein database used in bioinformatics.
The Structural Classification of Proteins (SCOP) database is a comprehensive and authoritative resource for the structural and evolutionary relationships of proteins. It provides a detailed and curated classification of protein structures, grouping them into families, superfamilies, and folds based on their structural and sequence similarities.
2. SYNOPSIS
INTRODUCTION
TERMS RELATED TO CHROMATOGRAPHY
HISTORY OF CHROMATOGRAPHY
TYPES OF CHROMATOGRAPHY
PARTITION CHROMATOGRAPHY
TYPES
PRINCIPLE
PROCEDURE
APPLICATIONS
REFERENCES
3. Introduction
Chromatography is an analytical technique used for separation, identification, and analysis of
various components of a mixture. The sample components often vary in physical and/ or
chemical properties.
The compound that is separated during chromatography is called analyte.
Russian botanist - M. S. Tswett. He developed this useful technique in 1906 to separate plant
pigments under gravity using a calcium carbonate column. Tswett also coined the term
‘chromatography’, which comes from the words chroma (Greek) i.e. colour and graphein,
which means "to write". “Towrite with colors”
In fact, this technique can be used to distinguish between two
compounds that are quite similar in molecular mass or
charge; however, this requires an appropriate combination of
materials and operating conditions.
A wide range of soluble or volatile, organic or inorganic
compounds can be thus separated using chromatographic technique.
4. TERMS RELATED TO Chromatography
StationaryPhase - phasethat stays in placeinside the column
usually viscous liquid chemically bonded to the inside of acapillary
column
Mobilephase- solvent moving through the column and is either
liquid orgas.
Elution- processof passing liquid or gasthrough the column.
Eluent -fluid entering the column
Eluate-fluid leaving the column
5. History of Chromatography
• Chromatography was first employed in Russia by the Italian-born
scientist Mikhail Tswett in 1906.
• of Archer John Porter Martin and Richard Laurence Millington
Synge during the 1940s and 1950s, for which they won the 1952 Nobel
Prize in Chemistry.
• They established the principles and basic techniques of partition
chromatography, and their work encouraged the rapid development of
several chromatographic methods.
• Researchers found that the main principles of Tswett's chromatography
could be applied in many different ways.
7. PartitionChromatography
basedon athin film formed on the surfaceof a
solidsupport by aliquid stationary phase.
Solute equilibrates between the mobile phase&
the stationary liquid.
method of separation in which the components
present in the mixture get distributed more likely
into two liquid phasesbecauseof differences in partition coefficients during the
flow of mobile phasein the chromatographycolumn.
8. Partition Coefficient - the ratio of the concentrations of a solute in
two immiscible or slightly miscible liquids, or in two solids, when it is
in equilibrium acrossthe interface between them.
TYPESOFPARTITIONCHROMATOGRAPHY
Liquid –Liquid Chromatography
Gas- Liquid Chromatography
9. Liquid - LiquidChromatography
• Employs liquid mobile andstationary phases
• Usessmall particles with moleculesbonded to their surfaceto give a
thin filmthat hasliquid like properties
• PAPER CHROMATOGRAPHY
10. 1.PAPERADSORPTIONCHROMATOGRAPHY
Paperimpregnatedwith silicaor aluminaactsasadsorbent(stationary phase)and
solvent asmobilephase.
2. PAPER PARTITIONCHROMATOGRAPHY
Moisture / Water present in the pores of cellulose fibers present in filter paperacts
asstationary phase& another mobilephaseisused assolvent
Ingeneral,PaperChromatography=PaperPartitionChromatography
Typesof PaperChromatography
11. PaperPartitionChromatography
In standard method of analysis,wherein the paperisutilized asa support with
one solvent asmobile phaseand the other is the stationaryphase
Themigration of substancesisdueto the partition coefficients
separation of similar substances by repeated divisions between two
immiscible liquids, so that the substances, in effect, cross the partition
between the liquids in opposite directions; where one of the liquids is
boundasafilm onfilter paper.
• Ascending,
• Descending
• Circular
12.
13. Cellulose layers in filter paper contains moisture which acts as stationary
phase& organicsolvents/buffersareusedasmobile phase
STATIONARY PHASEAND PAPERS USED
Whatman filter papers of different grades like No.1, No.2,
No.3, No.4, No.20, No.40, No.42 etc are used.
In general this paper contains 98-99% of α-cellulose, 0.3–1%β -cellulose
Principle of PaperPartitionChromatography
14. Pure solutions can be applied direct on the paper but solids are always dissolved in small
quantity of asuitablesolvent.
Biological tissues are treated with suitable solvents and their extracts obtained.
Proteins can be precipitated with alcohol and salts can be removed by treatment with
ion exchangeresin.
APPLICATIONOF SAMPLE
• Thesampleto beapplied isdissolved in the mobile phaseandapplied asasmall spot
onthe origin line,usingcapillarytubeor micropipette.
• verylow concentrationisusedto avoidlargerzone.
• Thespotisdriedonthe filter paperandisplacedin developingchamber.
Preparation of thesolution
15.
16. • Glass tanks are preferred most.
They are available in various
dimensional size depending upon
paper length and development
type.
• The chamber atmosphere
should be saturated with
solventvapor.
Chromatographic Chamber
17. • Paper is flexible when compared to glass plate used in TLC, several types of
development are possible which increasesthe easeof operation.
• The paper is dipped in solvent in such a manner that the spots will not dip
completely into the solvent.
• The solvent will rise up and it is allowed to run 2/3rd of paper height for better
andefficient result.
Procedures
20. Temperature
Purity of the solventsused
Quality of the paper,adsorbents& impurities presentnthe
adsorbents
Chambersaturation techniques,method of drying & development
Distancetravelled bythe solute& solvent
Chemicalreaction between the substancesbeingpartitioned.
pH of thesolution
Factors affecting RfValue
21. Gas- LiquidChromatography
• mobilephaseisagasandthe stationary phaseisaliquid, usuallyon smallbeads
packedin along column
Pointstoremember:
Samplehasto beableto bevaporizedwithout
decomposition
Basedonboiling point/vapor pressure
• Mobilephase
o Inert carrier gaslike Helium or Nitrogen
• Stationaryphase
o Layer of liquid or polymer on inert solid support
o Insideaglassor metal tubing (COLUMN)
24. GasPressure Regulator
• Helium - It has an excellent thermal conductivity, low density,
inertness andit permits greater flow rates. It ishighly expensive
• Nitrogen - Itoffers reduced sensitivity and is inexpensive
• Hydrogen - It has a distinctly better thermal conductivity and lower
density. Demerits are its reactivity with unsaturated compounds and
hazardous explosivenature
• Air - It is employed only when the atmospheric O2 is beneficial to
the detectorseparation.
25. Applications
usedfor final purification natural extracts, synthetic mixtures and
biological matrices.
It is also usedfor fractionization of complex crude extracts eg.
Petroleum fractions
Determination ofwater quality
Separation of aroma moleculesof wine
Determination of pesticide residue
26. REFERENCES
• Biophysical Chemistry By Upadhyay & Upadhyay Nath
• Instrumental Methods Of Chemical Analysis By Dr. B.K.Sharma
• A Textbook Of Microbiology By R.C. Dubey & D.K. Maheshwari
• Google Search
• Slideshare
