Rate theory is based on the random walk mechanism of molecules migrating through a chromatography column. It takes into account factors like band broadening, the effect of elution rate on band shape, and the different possible paths molecules can take through diffusion and availability. The symmetry factor As compares the widths of the first and second halves of a peak and should be close to 1 for a good separation. The height H of a packed bed should generally be 2-3 times the particle diameter for optimal performance.
This is a type of chromatography in which similar charged ions are separated by using ion exchange resin, that exchanges ions according to their relative affinities.
This is a type of chromatography in which similar charged ions are separated by using ion exchange resin, that exchanges ions according to their relative affinities.
ION EXCHANGE CHROMATOGRAPHY
ByM.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
Principles of Ion -exchange chromatography, High performance liquid chromatography (HPLC) , chromatography generally stands for a technique which separates mixtures based on different dynamic sharing of their components between two distinct physio-chemical environments called mobile and stationary phase by repeated absorption/desorption steps. Ion chromatography (IC) is a member of large family of liquid phase
chromatographic methods (that is a mobile phase is a liquid and a stationary phase is a
solid).
Ion exchange chromatography -SlideShareRIZWAN RIZWI
This ppt provide a good knowledge about ion exchange chromatography. I think this is very helpful for you .Here i have tried to explain a best way and simple method so guys you all enjoy this and gain your knowledge. And wish for me to provide more pptx for you all .at the end i want your experience give me suggestion if i made any mistake thank you .
Ion exchange chromatography may be defined as a reversible reaction in which free mobile ions of a solids called ion exchange are exchanged for different ions of similar charge present in solution.....................................................................
this slide contains all the basic about the topic ion exchange chromatography which contains all important information about topic in very easy language. it will be helpful for BSc, pharmacy and biomedical student.
ION EXCHANGE CHROMATOGRAPHY
ByM.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
Principles of Ion -exchange chromatography, High performance liquid chromatography (HPLC) , chromatography generally stands for a technique which separates mixtures based on different dynamic sharing of their components between two distinct physio-chemical environments called mobile and stationary phase by repeated absorption/desorption steps. Ion chromatography (IC) is a member of large family of liquid phase
chromatographic methods (that is a mobile phase is a liquid and a stationary phase is a
solid).
Ion exchange chromatography -SlideShareRIZWAN RIZWI
This ppt provide a good knowledge about ion exchange chromatography. I think this is very helpful for you .Here i have tried to explain a best way and simple method so guys you all enjoy this and gain your knowledge. And wish for me to provide more pptx for you all .at the end i want your experience give me suggestion if i made any mistake thank you .
Ion exchange chromatography may be defined as a reversible reaction in which free mobile ions of a solids called ion exchange are exchanged for different ions of similar charge present in solution.....................................................................
this slide contains all the basic about the topic ion exchange chromatography which contains all important information about topic in very easy language. it will be helpful for BSc, pharmacy and biomedical student.
INSTRUMENTAL METHODS OF ANALYSIS, B.PHARM 7TH SEM. AND FOR BSC,MSC CHEMISTRY. This is Geeta prasad kashyap (Asst. Professor), SVITS, Bilaspur (C.G) 495001
Dr.S.Karthikumar
Asst. Prof., Dept. of Biotechnology
Kamaraj College of Engineering and Technology
S.P.G.C.Nagar, Virudhunagar, Tamilnadu, India
skarthikumar@gmail.com
How to Split Bills in the Odoo 17 POS ModuleCeline George
Bills have a main role in point of sale procedure. It will help to track sales, handling payments and giving receipts to customers. Bill splitting also has an important role in POS. For example, If some friends come together for dinner and if they want to divide the bill then it is possible by POS bill splitting. This slide will show how to split bills in odoo 17 POS.
Operation “Blue Star” is the only event in the history of Independent India where the state went into war with its own people. Even after about 40 years it is not clear if it was culmination of states anger over people of the region, a political game of power or start of dictatorial chapter in the democratic setup.
The people of Punjab felt alienated from main stream due to denial of their just demands during a long democratic struggle since independence. As it happen all over the word, it led to militant struggle with great loss of lives of military, police and civilian personnel. Killing of Indira Gandhi and massacre of innocent Sikhs in Delhi and other India cities was also associated with this movement.
We all have good and bad thoughts from time to time and situation to situation. We are bombarded daily with spiraling thoughts(both negative and positive) creating all-consuming feel , making us difficult to manage with associated suffering. Good thoughts are like our Mob Signal (Positive thought) amidst noise(negative thought) in the atmosphere. Negative thoughts like noise outweigh positive thoughts. These thoughts often create unwanted confusion, trouble, stress and frustration in our mind as well as chaos in our physical world. Negative thoughts are also known as “distorted thinking”.
This is a presentation by Dada Robert in a Your Skill Boost masterclass organised by the Excellence Foundation for South Sudan (EFSS) on Saturday, the 25th and Sunday, the 26th of May 2024.
He discussed the concept of quality improvement, emphasizing its applicability to various aspects of life, including personal, project, and program improvements. He defined quality as doing the right thing at the right time in the right way to achieve the best possible results and discussed the concept of the "gap" between what we know and what we do, and how this gap represents the areas we need to improve. He explained the scientific approach to quality improvement, which involves systematic performance analysis, testing and learning, and implementing change ideas. He also highlighted the importance of client focus and a team approach to quality improvement.
The Indian economy is classified into different sectors to simplify the analysis and understanding of economic activities. For Class 10, it's essential to grasp the sectors of the Indian economy, understand their characteristics, and recognize their importance. This guide will provide detailed notes on the Sectors of the Indian Economy Class 10, using specific long-tail keywords to enhance comprehension.
For more information, visit-www.vavaclasses.com
Students, digital devices and success - Andreas Schleicher - 27 May 2024..pptxEduSkills OECD
Andreas Schleicher presents at the OECD webinar ‘Digital devices in schools: detrimental distraction or secret to success?’ on 27 May 2024. The presentation was based on findings from PISA 2022 results and the webinar helped launch the PISA in Focus ‘Managing screen time: How to protect and equip students against distraction’ https://www.oecd-ilibrary.org/education/managing-screen-time_7c225af4-en and the OECD Education Policy Perspective ‘Students, digital devices and success’ can be found here - https://oe.cd/il/5yV
How to Create Map Views in the Odoo 17 ERPCeline George
The map views are useful for providing a geographical representation of data. They allow users to visualize and analyze the data in a more intuitive manner.
Model Attribute Check Company Auto PropertyCeline George
In Odoo, the multi-company feature allows you to manage multiple companies within a single Odoo database instance. Each company can have its own configurations while still sharing common resources such as products, customers, and suppliers.
Unit 8 - Information and Communication Technology (Paper I).pdfThiyagu K
This slides describes the basic concepts of ICT, basics of Email, Emerging Technology and Digital Initiatives in Education. This presentations aligns with the UGC Paper I syllabus.
1. Rate Theory
• Based on a random walk mechanism
for the migration of molecules through a
column
• takes into account:
– band broadening
– effect of rate of elution on band shape
– availability of different paths for different
solute molecules to follow
– diffusion of solute along length
2.
3.
4. The symmetry factor (As) is expressed as:
As = b/a
where
a = 1st half peak width at 10% of peak height
b = 2nd half peak width at 10% of peak height
As should be as close as possible to 1. A
reasonable As value for a short column as
used in IEX is 0.80–1.80.
As a general rule, a good H value is about two to three times the
average particle diameter of the medium being packed. For a 90 µm particle,
this means an H value of 0.018–0.027 cm.
5. MATRIX
•Rigid Solids: Based on silica matrix, with stand high pressures
(4000-6000 psi)
•Hard gels: based on highly porous/non porous particles of
polystyrene cross-linked with divinyl benzene
• Soft gels: Such as cellulose/agarose, dextran, polyamide and
other hydrophilic polymers. Separation of proteins etc.
Typical Particle size used in Chromatography
Analytical Applications 3-10 µm
Preparative Separations 10-40 µm
Low-pressure/large-scale applications 40-150 µm
Very large scale operations ~300 µm
7. •Biomolecules are purified using chromatography techniques that
separate them according to differences in their specific properties.
•Ion exchange chromatography (IEX) separates biomolecules according
to differences in their net surface charge.
Gel filtration Hydrophobic Ion Exchange Affinity Chromatography
9. Ion-Exchange Chromatography
•IEX for the separation of biomolecules was introduced in the
1960s and continues to play a major role in the separation and
purification of biomolecules.
•Today, IEX is one of the most frequently used techniques for
purification of proteins, peptides, nucleic acids and other charged
biomolecules, offering high resolution and group separations with
high loading capacity.
•The technique is capable of separating molecular species that have
only minor differences in their charge properties, for example two proteins
differing by one charged amino acid.
•These features make IEX well suited for capture, intermediate purification or
polishing steps in a purification protocol and the technique is used from
microscale purification and analysis through to purification of kilograms
of product.
10. Ion Exchange Chromatography (IEX)
• IEX separates molecules on the basis of differences in their net surface
charge.
• Molecules vary considerably in their charge properties and will exhibit
different degrees of interaction with charged chromatography media
according to differences in their overall charge, charge density and
surface charge distribution.
• A protein that has no net charge at a pH equivalent to its isoelectric
point (pI) will not interact with the charged medium.
• At a pH above its isolectric point, a protein will have net negative
surface charge and will bind to positively charged medium or
anion exchanger
• At a pH below its isolectric point, a protein will have net positive
surface charge and will bind to negatively charged medium or cation
exchanger
11. IEX Chromatography is the main means of protein purification
both at laboratory and industrial scale
• IEX matrices are relatively cheap
• IEX matrices have very high capacity (up to 100 mg protein per
ml gel)
• IEX-chromatography has high resolution
• Versatile, the same column can be used for purification of
different proteins
• Optimization and scale up is rather straightforward
12. Isoeletric point (pI) and Ion Exchangers
• At a pH above its isoelectric point, a protein will bind to a positively
charged medium or anion exchanger.
• At a pH below its pI, a protein will bind to a negatively charged
medium or cation exchanger.
13. Ion Exchange Matrices- Functional groups
Anion Exchangers Functional group
----------------------------------------------------------------------------------------------------
Quaternary ammonium (Q) strong -O-CH2N+(CH3)3
Diethylaminoethyl (DEAE) weak -O-CH2CH2N+H(CH2CH3)2
Diethylaminopropyl (ANX) weak -O-CH2CHOHCH2N+H(CH2CH3)2
Cation Exchangers Functional group
----------------------------------------------------------------------------------------------------------------
Sulfopropyl (SP) strong -O-CH2CHOHCH2OCH2CH2CH2SO3-
Methyl sulfonate (S) strong -O-CH2CHOHCH2OCH2CHOHCH2SO3-
Carboxymethyl (CM) weak -O – CH2COO-
___________________________________________________________________________
14. STRONG/WEAK – What does it mean?
• strong and weak do not refer to the strength with which
the functional groups bind to proteins.
• Strong ion exchangers show no variation in ion exchange
capacity with change in pH.
• These exchangers do not take up or lose protons with changing pH
and so have no buffering capacity, remaining fully charged over
a broad pH range.
• Strong ion exchangers include Q (anionic), S and
SP (cationic) (pH 2 - 12).
• Weak ion Exchangers: DEAE (anion exchange) and CM (cation exchange)
are fully charged over a narrower pH range (pH 2 - 9 and pH 6 - 10,
respectively).
15. Strong Ion-Exchangers
Titration curves show the ion exchange capacity of strong ion exchangers
Q and S.
Approximately 5 ml of Q or S Sepharose Fast Flow are equilibrated in
1 M KCl and titrated with 0. 1 M NaOH.
20. Elution profile of three different proteins at various pH
Protein A
Protein B
Protein C
The pH vs. net surface charge curves for three different proteins are shown. Schematic chromatograms
for a CM and a DEAE ion exchanger are shown at the top and bottom respectively. At the most acidic pH
value, all three proteins are positively charged and adsorb only to the CM ion exchanger. They are then
eluted in the order of net charge. At the next pH value chosen, the protein has passed its isoelectric point
and is now negatively charged, while the other two still retain positive charges. The blue protein will
consequently adsorb to a DEAE ion exchanger, but not to a CM ion exchanger the other two do. At the
next highest pH value the only one positively-charged protein still adsorbs to the CM ion exchanger.
Because of their negative net charges, the two other proteins adsorb to the DEAE ion exchanger. At
the most alkaline pH, all three proteins are adsorbed to the DEAE ion exchanger only. Thus, by varying
the pH of the mobile phase, one can greatly influence the selectivity in ion exchange chromatography.
21. Ion Exchange Matrices
Trade name Material Mean particle size
___________________________________________________________
MiniBeads Polystyrene/divinyl benzene 3 µm
MonoBeads ----do--- 10 µm
SOURCE 30 ---do--- 30 µm
Sepharose High Performance Agarose 6% 34 µm
Sepharose Fast Flow Agarose 6% 90 µm
Sepharose 4 Fast Flow Agarose 4% 90 µm
Sepharose XL Agarose 6%, dextran chains 90 µm
coupled to agarose
Sepharose Big Beads Agarose 6% 200 µm
___________________________________________________________________
22. MonoBeads showing spherical, Uniform size distribution of SOURCE
monodispersed particles. monodispersed particles.
Structure of cross-linked agarose
media (Sepharose).
Packed columns and resins
24. Practical considerations for IEX separation
1. Equilibrate column with 5–10 column volumes of start buffer
or until the baseline, eluent pH and conductivity are stable.
2. Adjust the sample to the chosen starting pH and ionic strength and
apply to the column.
3. Wash with 5–10 column volumes of start buffer or until the baseline,
eluent pH and conductivity are stable i.e. when all unbound material has
washed through the column.
4. Begin elution using a gradient volume of 10–20 column volumes with an
increasing ionic strength up to 0.5 M NaCl (50%B).
5. Wash with 5 column volumes of 1 M NaCl (100%B) to elute any remaining
ionically bound material.
6. Re-equilibrate with 5–10 column volumes of start buffer or until eluent
pH and conductivity reach the required values.