This document provides an introduction to gas chromatography including its components, advantages, and applications. It discusses the basic process of separating components using an inert gaseous mobile phase and immobilized liquid or solid stationary phase. Key components are described including the carrier gas, sample injection port, columns, and common detectors like FID and TCD. Applications include qualitative and quantitative analysis of compounds like oils, fatty acids, foods, drugs, and pollutants.
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
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
Gas chromatography and its instrumentationArgha Sen
Gas chromatography is an unique technology which helps us in separating volatile analytes. Its is an easy and reproduciple method for detecting residual solvents found in APIs.
In this slide contains principle, instrumentation, methodology, and application of gel chromatography.
Presented by: SATHEES CHANDRA (Department of pharmaceutical analysis).
RIPER, anantapur
HPLC is a High Performance liquid Chromatography.
High Pressure Liquid Chromatography.
High Priced Liquid Chromatography.
It is column chromatography.
It is Liquid Chromatography.
It is modified from of gas chromatography, it is applicable for both Volatile as well as Non volatile compound.
It can mainly divided by two types 1. Normal phase HPLC 2. Reversed Phase HPLC.
It is having a high resolution and separation capacity.
This presentation gives you thorough knowledge about the IR Spectroscopy. This include basic principle, type of vibrations, factors influencing vibrational frequency, instrumentation and applications of IR Spectroscopy. This is the most widely used technique for identifying unknown functional group depending on the vibrational frequency.
Detectors are the brain of any chromatograhic system. It help us to record the chromatogram based on certain characteristics of the analyte and help us in identifying that compound both qualitatively and quantitatively.
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
A separation technique in which the mobile phase is a gas. Gas chromatography is always carried out in a column.
Separating mixtures of gases or volatile materials based primarily on their physical properties.
Gas chromatography and its instrumentationArgha Sen
Gas chromatography is an unique technology which helps us in separating volatile analytes. Its is an easy and reproduciple method for detecting residual solvents found in APIs.
In this slide contains principle, instrumentation, methodology, and application of gel chromatography.
Presented by: SATHEES CHANDRA (Department of pharmaceutical analysis).
RIPER, anantapur
HPLC is a High Performance liquid Chromatography.
High Pressure Liquid Chromatography.
High Priced Liquid Chromatography.
It is column chromatography.
It is Liquid Chromatography.
It is modified from of gas chromatography, it is applicable for both Volatile as well as Non volatile compound.
It can mainly divided by two types 1. Normal phase HPLC 2. Reversed Phase HPLC.
It is having a high resolution and separation capacity.
This presentation gives you thorough knowledge about the IR Spectroscopy. This include basic principle, type of vibrations, factors influencing vibrational frequency, instrumentation and applications of IR Spectroscopy. This is the most widely used technique for identifying unknown functional group depending on the vibrational frequency.
Detectors are the brain of any chromatograhic system. It help us to record the chromatogram based on certain characteristics of the analyte and help us in identifying that compound both qualitatively and quantitatively.
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
A separation technique in which the mobile phase is a gas. Gas chromatography is always carried out in a column.
Separating mixtures of gases or volatile materials based primarily on their physical properties.
La cromatografía de gases es una técnica cromatográfica en la que la muestra se volatiliza y se inyecta en la cabeza de una columna cromatográfica. La elución se produce por el flujo de una fase móvil de gas inerte. A diferencia de los otros tipos de cromatografía, la fase móvil no interacciona con las moléculas del analito; su única función es la de transportar el analito a través de la columna.
The investigation of thermodynamic properties and reactivity yields interesting insights into the chemistry of newly synthesized substances. With thermal analysis extensive information can be gained from small samples (often only a few milligrams). In addition, the data obtained by thermal analysis can be used to plan and optimize a synthesis. Among the most important applications are identification and purity analysis, and the determination of characteristic temperatures and enthalpies of phase transitions (melting, vaporization), phase transformations, and reactions. Investigations into the kinetics of consecutive reactions and decomposition reactions are also possible. With the instruments available today such analyses can usually be performed quickly and easily. In this review the fundamentals of thermoanalytical methods are described and illustrated with selected examples of applications to low and high molecular weight compounds.
In DSC the heat flow is measured and plotted against temperature of furnace or time to get a thermo gram. This is the basis of Differential Scanning Calorimetry (DSC).
The deviation observed above the base (zero) line is called exothermic transition and below is called endothermic transition.
Low amount of sample
Complex mixture.
Gas chromatography is a process of separating component(s) from the given crude drug or mixture by using stationary phase (solid or liquid) and gaseous mobile phase. It involves a sample being vaporized and injected onto the head of the chromatographic
column. The sample is transported through the column by the flow of inert, gaseous
mobile phase. The column itself contains a solid or liquid stationary phase which is adsorbed onto the
surface of an inert solid.
Gas Chromatography in Analytical Analysis.pptxRAHUL PAL
Gas chromatography is a common type of chromatography used in analytical chemistry for separating and analyzing compounds that can be vaporized without decomposition. Typical uses of GC include testing the purity of a particular substance, or separating the different components of a mixture.
Introduction to gas Chromatography
,Principle of gas chromatography
Instrumentation of gas Chromatography
Type of detectors of gas chromatography
Advantages of gas chromatography
Disadvantages of gas chromatography
Applications of gas chromatography
Gas Chromatography is an analytical techniques, used for the separation of volatile substances on the basis of their partition coefficient . In this slide you will find out different instrumentation of gas chromatography, its advantages, disadvantages and moreover its applications.
Gas chromatography- “It is a process of separating component(s) from the given crude drug by using a gaseous mobile phase.”
Principle- The principle of separation in GC is “partition.”
The mixture of components to be separated is converted to vapor and mixed with the gaseous mobile phase.
The component which is more soluble in the stationary phase travels slower and eluted later.
The component which is less soluble in the stationary phase travels faster and eluted out first.
No two components have the same partition coefficient conditions.
So the components are separated according to their partition coefficient.
The partition coefficient is “the ratio of solubility of a substance distributed between two immiscible liquids at a constant temperature.’
It involves a sample being vaporized and injected onto the head of the chromatographic column.
The sample is transported through the column by the flow of inert, gaseous mobile phase.
The column itself contains a liquid stationary phase which is adsorbed onto the surface of an inert solid.
Two major types:
1. gas-solid chromatography: Here, the mobile phase is a gas while the stationary phase is a solid.
Used for separation of low molecular gases,
e.g., air components, H2S, CS2, CO2, rare gases, CO, and oxides of nitrogen.
2.Gas-liquid chromatography: The mobile phase is a gas while the stationary phase is a liquid retained on the surface as an inert solid by adsorption or chemical bonding.
Advantages-
Both qualitative and quantitative analyses are possible.
The instrument is simple, time of analysis is short.
High sensitivity.
The method is applicable to about 60% of organic compounds.
Very small sample sizes can be used.
Analysis can be highly accurate and precise.
Applications-
Quality control and analysis of drug products like antibiotics (penicillin), antivirals (amantadine), general anesthetics (chloroform, ether), sedatives/hypnotics (barbiturates), etc.
Assay of drugs – purity of a compound can be determined for drugs like :
Atropine sulfate
Clove oil
Stearic acid
In determining the levels of metabolites in body fluids like plasma, serum, urine, etc
Estimation of spoilage components, such as histamine and carbonyls, that cause rancidity.
Theme of Pharmacist Day 2021 - Trust building process in patients at hospitals and community pharmacies. It briefly explain the education and skills of pharmacists in healthcare. It may useful for practicing pharmacists with D Pharm , B Pharm and Pharm D .
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Just an attempt to promote pharmacy practice in India. It helps for the students and professionals of pharmacy in healthcare, especially Doctor of Pharmacy and Pharmacy Practice students
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https://alandix.com/academic/papers/synergy2024-epistemic/
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Length: 30 minutes
Session Overview
-------------------------------------------
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- What out-of-the-box solutions are available for real-time monitoring JMeter tests?
- What are the benefits of integrating InfluxDB and Grafana into the load testing stack?
- Which features are provided by Grafana?
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To view the webinar recording, go to:
https://www.rttsweb.com/jmeter-integration-webinar
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Kubernetes & AI - Beauty and the Beast !?! @KCD Istanbul 2024
Gas chromatography
1. K V GOPINATH M Pharm PhD,CPhT
Tirumala Tirupati Devasthanams
TIRUPATI
e-mail:gopinath.karnam@gmail.com
GAS CHROMATOGRAPHY
2. Introduction
Gas chromatography – It is a process of separating component(s)
from the given crude drug by using a gaseous mobile phase.
It involves a sample being vaporized and injected onto the head of
the chromatographic column. The sample is transported through the
column by the flow of inert, gaseous mobile phase. The column itself
contains a liquid stationary phase which is adsorbed onto the surface
of an inert solid.
Two major types
• Gas-solid chromatography
(stationary phase: solid)
• Gas-liquid chromatography
(stationary phase: immobilized liquid)
3. Advantages of Gas Chromatography
The technique has strong separation power and even complex
mixture can be resolved into constituents
The sensitivity of the method is quite high
It gives good precision and accuracy
The analysis is completed in a short time
The cost of instrument is relatively low and its life is generally long
The technique is relatively suitable for routine analysis
4. Components of Gas chromatography
Carrier gas
- He (common), N2, H2, Argon
Sample injection port
- micro syringe
Columns
2-50 m coiled stainless steel/glass/Teflon
Detectors
-Flame ionization (FID)
-Thermal conductivity (TCD)
-Electron capture (ECD)
-Nitrogen-phosphorus
-Flame photometric (FPD)
-Photo-ionization (PID)
6. Carrier gas
The carrier gas must be chemically inert.
Commonly used gases include nitrogen, helium, argon, and carbon
dioxide.
The choice of carrier gas is often dependant upon the type of
detector which is used.
The carrier gas system also contains a molecular sieve to remove
water and other impurities.
- P inlet 10-50 psig
-F=25-150 mL/min packed column
-F=1-25 mL/min open tubular column
7. Sample injection- Direct Injection
1)Direct injection :into
heated port (>T oven)
using micro syringe
- (i) 1-20 uL
packed column
-(ii) 10-3
uL
capillary column
8. Sample injection- rotary sample valve with
sample loop
Split injection: routine method
- 0.1-1 % sample to column
- remainder to waste
Split less injection: all sample to column
- best for quantitative analysis
- only for trace analysis, low [sample]
On-column injection:
-for samples that decompose above boiling
Point ( no heated injection port)
-column at low temperature to condense
sample in narrow band
-heating of column starts chromatography
9. Gas Chromatography - Columns
There are two general types of column, packed and capillary (also known
as open tubular).
Packed columns contain a finely divided, inert, solid support material
( diatomaceous earth) coated with liquid stationary phase. Most packed
columns are 1.5 - 10m in length and have an internal diameter of 2 - 4mm.
Capillary columns have an internal diameter of a few tenths of a
millimeter. They can be one of two types; wall-coated open
tubular (WCOT) or support-coated open tubular (SCOT).
- Wall-coated columns consist of a capillary tube whose walls are
coated with liquid stationary phase. In support-coated columns, the inner
wall of the capillary is lined with a thin layer of support material such as
diatomaceous earth, onto which the stationary phase has been adsorbed.
- SCOT columns are generally less efficient than WCOT columns.
Both types of capillary column are more efficient than packed columns.
11. G C - DETECTORS
There are many detectors which can be used in gas chromatography.
Different detectors will give different types of selectivity.
Detectors can be grouped into concentration dependant
detectors and mass flow dependant detectors.
The signal from a concentration dependant detector is related to the
concentration of solute in the detector, and does not usually destroy
the sample Dilution of with make-up gas will lower the detectors
response.
Mass flow dependant detectors usually destroy the sample, and the
signal is related to the rate at which solute molecules enter the
detector. The response of a mass flow dependant detector is
unaffected by make-up gas
12. G C – IDEAL DETECTORS
Sensitive (10-8
-10-15
g solute/s)
Operate at high T (0-400 °C)
Stable and reproducible
Linear response
Wide dynamic range
Fast response
Simple (reliable)
Nondestructive
Uniform response to all analytes
13. Flame Ionization Detector (FID)
It operates by the principle that by
change in conductivity of the flame as
the compound is burnt. The change in
conductivity of the flame does not arise
by simple ionization of the compound ,
it is partial or complete stripping of the
compound to give charged hydrogen-
deficient polymers or aggregates of
carbon of low ionization potential.
Rugged; Sensitive (10-13
g/s) ; Wide dynamic range
(107
) ;Signal depends on # C atoms in organic
analyte - mass sensitive; not concentration sensitive;
Weakly sensitive to carbonyl, amine, alcohol,
amine groups; Not sensitive to non-combustibles -
H2O, CO2, SO2, Nox; Destructive
14. Thermal Conductivity Detector (TCD)
It is based upon the alteration of the
thermal conductivity of the carrier gas
in the presence of an organic
compound. The platinum wires are
heated electrically and assume
equilibrium conditions of temperature
and resistance when carrier gas alone
passes over them. They are mounted in
a whetstone bridge arrangement and
when a compound emerges, the
thermal conductivity of the gas
surrounding wire alters, and hence the
temperature and resistance of the wire
change with a concomitant out of
balance signal which is amplified and
recorded.
Rugged ;Wide dynamic range
(105
);Nondestructive; Insensitive
(10-8
g/s) - non-uniform
15. Electron Capture Detector (ECD)
The ECD ionizes the carrier
gas by means of a radioactive
source. The potential across
two electrodes is adjusted to
collect all the ions and a steady
saturation current, is therefore,
recorded.
Electrons from b-source ionize carrier
molecules capture electrons and
decrease current ; Simple and reliable ;
Sensitive (10-15
g/s) to electronegative
groups (halogens, peroxides) ;Largely
non-destructive ; Insensitive to amines,
alcohols and hydrocarbons ; Limited
dynamic range (102
)
16. Summary of common GC detectors
Detector Type Support gases Selectivity Detectability Dynamic range
Flame ionization (FID) Mass flow Hydrogen and air Most organic cpds. 100 pg 107
Thermal conductivity
(TCD)
Concentration Reference Universal 1 ng 107
Electron capture (ECD) Concentration Make-up
Halides, nitrates,
nitriles, peroxides,
anhydrides,
organometallics
50 fg 105
Nitrogen-phosphorus Mass flow Hydrogen and air Nitrogen, phosphorus 10 pg 106
Flame photometric
(FPD)
Mass flow
Hydrogen and air
possibly oxygen
Sulphur, phosphorus,
tin, boron, arsenic,
germanium, selenium,
chromium
100 pg 103
Photo-ionization (PID) Concentration Make-up
Aliphatics, aromatics,
ketones, esters,
aldehydes, amines,
heterocyclics,
organosulphurs, some
organometallics
2 pg 107
17. Summary of common GC detectors
The effluent from the column is mixed with hydrogen and air, and
ignited.
Organic compounds burning in the flame produce ions and electrons
which can conduct electricity through the flame.
A large electrical potential is applied at the burner tip, and a
collector electrode is located above the flame. The current resulting
from the pyrolysis of any organic compounds is measured.
FIDs are mass sensitive rather than concentration sensitive; this gives
the advantage that changes in mobile phase flow rate do not affect
the detector's response.
The FID is a useful general detector for the analysis of organic
compounds; it has high sensitivity, a large linear response range, and
low noise. It is also robust and easy to use, but unfortunately, it
destroys the sample
18. Temperature Programming
As column temperature raised, vapor pressure analyte increases,
eluted faster
Raise column temperature during separation – temperature
programming - separates species with wide range of polarities or
vapor pressures
19. Evaluation
HETP- It is the distance on the column in which equilibrium is
attained between the solute in the gas phase and the solute in liquid
phase. Larger the number of theretical plates/ smaller the HETP, the
more efficient the column is for separation.
HETP = Length of column/n ; Where n = number of theretical plates= 16 * x2/y2
Retention Time: Time in minute from the point of injection to the
peak maximum.
Retention Volume: (1) VR = tR ×F (retained) (2) VM = tM ×F (non-
retained)
average volumetric flow rate (mL/min) F can be estimated by
measuring flow rate exiting the column using soap bubble meter
(some gases dissolving in soap solution)
But measured VR and VM depend on
- pressure inside column
20. Applications of Gas Chromatography
Qualitative Analysis – by comparing the retention time or volume of
the sample to the standard / by collecting the individual components
as they emerge from the chromatograph and subsequently identifying
these compounds by other method
Quantitative Analysis- area under a single component elution peak is
proportional to the quantity of the detected component/response
factor of the detectors.
Volatile Oils, official monograph gives chromatography profile for
some drugs. E.g. to aid distinction between anise oil from star anise
and that from Pimpinelle anisum
Separation of fatty acids derived from fixed oils
21. Applications of Gas Chromatography
Miscellaneous-analysis of foods like carbohydrates, proteins, lipids,
vitamins, steroids, drug and pesticides residues, trace elements
Pollutants like formaldehyde, carbon monoxide, benzen, DDT etc
Dairy product analysis- rancidity
Separation and identification of volatile materials, plastics, natural
and synthetic polymers, paints, and microbiological samples
Inorganic compound analysis