Chrominfo

Saturday, April 4, 2020

What is the difference between planar and column chromatography?

Column chromatography and planar chromatography are types of chromatography both are generally used methods for separating the molecules from complex mixtures.

The key difference between planar and column chromatography is that in planar chromatography the stationary phase is on a flat plate it is either paper/ plate of an alumna or silica, and column chromatography is packed with silica or suitable material which used as the stationary phase.

Thin-layer chromatography and paper chromatography are examples of planar chromatography, which both follow very similar principles and methods. Gas chromatography, high-performance liquid chromatography example of column chromatography, they work on parallel principle.

Here are some points of difference between planar and column chromatography.

The mobile phase of planar chromatography is run against gravity.
The mobile phase of column chromatography is run under gravity.
Planar chromatography requires a more polar mobile phase compared to the column chromatography.
Column chromatography requires a less polar mobile phase compared to the planar chromatography.
Planar chromatography rapidly separates the analytes.
Column chromatography takes time to separate the analytes.
In planar chromatography the stationary phase is short.
In column chromatography the stationary phase is long.
Planar chromatography requires a low amount of solvents and samples.
Column chromatography requires a high amount of solvents and samples.
For analytical purposes, the planar chromatography is used.
For preparatory purposes, the column chromatography is used.

Advantages and disadvantages of planar chromatography

Planar chromatography is a type of chromatography method used to separate a mixture of sample components, in which the stationary phase is on a flat plate and the mobile phase travel through capillary movement by stationary phase. There are two types of planar chromatography which are paper chromatography and thin-layer chromatography (TLC). Both techniques work on the same principles and methods, they employ different planes for the stationary phase.
The advantages of planar chromatography are as follows.

The analytes are separated by this method within a short period.

A complex sample mixture can be easily separated as it is sensitive.

Planar chromatography requires fewer sample volumes for analysis.

This can be automated in the form of high-performance thin-layer chromatography HPTLC).

Compared with other separation techniques it is an economical method.

The use and setup of the planer chromatography are simple.

This technique requires very few tools to carry out the process.

In this method, organic as well as inorganic molecules can be possible to identify

The disadvantages of planar chromatography are as follows.

The use of the planar chromatography method cannot separate volatile components.

Since it is an open system, humidity and temperature can affect the results.

Another disadvantage of planar chromatography is that the molecules must be soluble.

Planar chromatography isolates only small sample volumes. This has less accuracy compared with HPTLC and HPLC.

Advantages and disadvantages of flash chromatography

Flash chromatography is a method used to separate sample mixtures into their components. Column chromatography is classified into two categories, based on how the liquid flows down the column. When the mobile phase is allowed to flow down the column by gravity, it is gravity column chromatography. When the mobile phase is allowed to flow down by the column by the pressure it is called flash chromatography. In flash chromatography a controlled air pressures combination and a shorter column chromatography which has been designed for particularly quick separation.
The advantages of flash chromatography are as follows.

Large amounts of samples can be separated using flash chromatography.

It is a rapid and economical method of separation.

We can use thin-layer chromatography (TLC) to find out and implement a suitable mobile phase.

There is no expensive tool to perform the process.

If high resolution is needed, it is performed before high-performance liquid chromatography (HPLC) to prevent any contamination of the HPLC column.

A wide range of mobile phases can be used in flash chromatography.

In preparatory style chromatography, the analytes can be isolated and reused.

The disadvantages of flash chromatography are as follows.

Due to the open system, it can be affected by many factors like temperature, humidity, etc.

Another disadvantage of flash chromatography is the irreversible adsorption of solutes.

The drawbacks of long stationary phase in the column are time-consuming.

More amount of solvents is required to separate, which is more costly.

Manual loading of silica can cause errors in the process.

Advantages and disadvantages of gas-liquid chromatography

The gas-liquid chromatography is a type of chromatography used for separation and identification of volatile compounds. The stationary phase consists of small volumes of non-volatile liquid, placed on a finely divided solid inert support, and a mobile phase (carrier gas) is an inert gas such as nitrogen or helium used in gas-liquid chromatography. Gas-liquid chromatography works on the principle of partition. In GC the vaporized molecules of sample mixture fractionated caused by the partition among a liquid stationary phase and a gaseous mobile phase held in the column.
The advantages of gas-liquid chromatography are as follows.

Qualitative and quantitative analysis is possible with high sensitivity in gas-liquid chromatography.

GC column length is extra as compared with high-performance liquid chromatography and thus the more complex sample mixture can be separate with high resolution.

GLC provides high sensitivity, precision and resolution of power compared to other chromatographic methods.

We can use it with a thermal detector or with a mass detector.

Sampling analysis is very fast so it saves time.

Very small quantities of samples are required for analysis.

The disadvantages of gas-liquid chromatography are as follows.

The detectors used in GC are destructive.

The disadvantage of the FID is that we cannot recover the sample emanating from the detector.

The major disadvantage of gas-liquid chromatography is that only volatile samples can be analyzed.

In HPLC or TLC we can change the composition of the mobile phase, but in GC, we cannot change the mobile phase since it has a constant flow of carrier gas.

Advantages and disadvantages of partition chromatography

Partition chromatography is a kind of chromatography, in that sample components are distributed in two liquids due to variation in the partition coefficient. In this technique, both stationary and mobile phases are in the same phase and in which the molecules are separated. Thus the analytes are distributed at any phage. "Partition" is basic principle that is used in several chromatographic methods such as high-performance liquid chromatography (HPLC), thin-layer chromatography (TLC), gas chromatography (GC), and paper chromatography. Gas-liquid chromatography and liquid-liquid chromatography are the types of partition chromatography
The advantages of partition chromatography are as follows.

The advantages of partition chromatography consist of an easy-to-operate and economical method for separation.

The partition chromatography technique can isolate both organic and inorganic compounds.

It saves time because in a short duration it separates compounds.

It provides high efficiency.

This technique provides accurate results.

Selectivity is better in partition chromatography because a mobile phase can be simply changed.

The disadvantages of partition chromatography are as follows.

The high volume of the mobile phase is sometimes required for separation

Data cannot be stored long in certain types of partition chromatography.

Automation has made it more difficult and costly.

In gas-liquid chromatography, only volatile compounds can be separated.

It is possible to separate only soluble analytes of sample mixtures.

Advantages and disadvantages of chiral chromatography

Chiral chromatography is a type of chromatographic technique used to separate the enantiomers of a chiral compound. It can use to determine the ratio of the enantiomers found in a sample or to separate different enantiomers. It consists of a mobile phase, either liquid or gas passing over a chiral stationary phase in a cylindrical column. A sample component injected into the mobile phase and as per their different affinities for the chiral stationary phase, they separated. The chiral chromatography is used when some analytes have a chiral center and they not isolate by the reversed-phase chromatography and normal phase chromatography.
The advantages of chiral chromatography are as follows.

The majorities of inorganic molecules are chiral and are analyzed by this method.

Board ranges of the chiral column are available.

This chromatographic method provides fast and accurate results.

You can get different selectivity’s from the chiral phases.

The disadvantages of chiral chromatography are as follows.

It is used only to separate chiral compounds. This is the major disadvantage of chiral chromatography.

we need to be an expert to handle the system.

The major disadvantage of chiral chromatography is the need for pure chiral substances and their relative slowness.

Several chiral additives are expensive, and must often be synthesized.

Advantages and disadvantages of adsorption chromatography

The adsorption chromatography is a form of chromatography and it is the most widely used method in the form of column chromatography, gas-solid chromatography, and thin-layer chromatography. Adsorption chromatography consists of a stationary phase (fine solid partitioned) and a mobile phase (solvent or a mixture of solvents). The analyte differently interacts with the stationary phase because every analyte has a different affinity towards the adsorbent. Therefore, the separation of molecules occurs at different retention times.
The advantages of adsorption chromatography are as follows.

It has a wide range of mobile phases for the separation of compounds.

Adsorption chromatography is an important method to separate many components that are not separated by other techniques.

The complex sample mixtures can be easily separated by this method.

Very few types of apparatus or types of equipment are required for isolation.

The disadvantages of adsorption chromatography are as follows.

Obtained results from some methods of adsorption chromatography are complex to reproduce.

This can cause catalytic variations.

Some solutes take retention time to separate this is a major disadvantage of adsorption chromatography.

It is more complicated and costly when make automation.

Advantages and disadvantages of ion-exchange chromatography

Ion-exchange chromatography is a type of HPLC chromatography and is used for purification of proteins and other charged molecules. The molecules are separated in this method, depending on their charge. In this type of chromatography, the negatively charged analytes are attracted with a positive charge to solid supports. Conversely, in cation-exchange chromatography positive charged analytes are pulled into a negatively charged solid support. Cation exchange chromatography and anion exchange chromatography are two types of ion-exchange chromatography
The advantages of ion-exchange chromatography are as follows.

Ion-exchange chromatography is one of the most powerful methods of separating charged particles.

Using this method the inorganic ions can also be separated.

It can use more commonly for both analytical and preparative purposes.

Almost all charged molecules such as small amino acids, nucleotides and large proteins can separate using this method.

This is very effective and powerful water softening method.

Resins have a long life.

It has cheap maintenance.

The disadvantages of ion-exchange chromatography are as follows.

The buffer requirement is the major disadvantage of ion-exchange chromatography.

It has a high working cost since the buffer is used for the separation of components.

This method can only be used to isolate charged molecules.

Sodium ions entering the soft water will increase the acidity level in the water.

Advantages and disadvantages of size exclusion chromatography

The size exclusion chromatography or gel filtration chromatography is one of the high-performance liquid chromatography (HPLC) separation types. This technique is used to separate particles based on size, larger molecules travel faster throughout the column as compared to smaller molecules, as they penetrate deep into the holes. The gel permeation chromatography (GPC) and gel filtration chromatography (GFC) are two basic types of size exclusion chromatography (SEC). GPC uses a hydrophobic stationary phase and a non-aqueous mobile phase and GFC uses a hydrophilic stationary phase and an aqueous mobile phase for the separation of components.
The advantages of size exclusion chromatography are as follows.

Good separation of large components from the small sample components with a less amount of solvent is the major advantage of size exclusion chromatography.

The separation in gel filtration chromatography doesn't rely on the buffer composition, pH, temperature and ionic strength.

Size exclusion chromatography provides a well-defined separation of analytes.

This technique requires a short analysis time.

There is no sample loss in this technique.

The flow rate of the system can be set as per requirement.

It gives a narrow band and good sensitivity.

The lower amount of mobile phase required for isolation compared to other techniques.

The disadvantages of size exclusion chromatography are as follows.

Mobile phase filtering is necessary to prevent the columns and interfere with detectors.

The major disadvantage of size exclusion chromatography is that the limited number of peaks can be resolved since the run time is short.

It has poor selectivity compared to other chromatographic techniques.

The load capacity of the sample is another major disadvantage of gel filtration chromatography.

Tuesday, March 31, 2020

Advantages and disadvantages of reversed-phase chromatography

The reversed-phase chromatography is a type of HPLC chromatography; it is working on the principle of hydrophobic interactions and is one of the most popular techniques, as it applies to a wide variety of analytes. The RP- chromatography is a commonly used separation technique, in which the molecules are separated based on polarity. The polar mobile phase and a non-polar stationary phase are used in RP-chromatography. The more hydrophobic molecules are the more strongly attaches to the column and the more volume of solvent required to elute the molecule. Hence, the retention times are longer for non-polar components, while polar compounds are more readily elute from the stationary phase.
The advantages of reversed-phase chromatography are as follows.

It is an economical method compared to other chromatographic techniques.

RP-HPLC allows water to be used in the composition of the mobile phase with other solvents.

Another advantage of using reversed-phase chromatography is that it provides accurate results with small amounts of sample.

RP-chromatography also has the advantage of being able to use pH selectivity to improve the separation.

The hydrophobic stationary phase in reverse-phase columns works well for the retention of most organic molecules.

In RP-chromatography we can use pH selectivity to get better separations.

About 75 percent of all HPLC methods use reversed-phase chromatography.

The disadvantages of reversed-phase chromatography are as follows.

Water-insoluble compounds and amines can be more difficult to analyze.

In RP-HPLC need to create pressure.

It requires technical capability and skill to handle the system.

The silica of the reversed-phase column can be dissolution at pH > ~7.5.

The eluted sample from the column cannot be recovered.

Additional techniques are needed to confirm the identity of the analytes.

Thursday, March 19, 2020

Principle and Application of Polarimetry

What is a polarimeter?

Polarimetry can be defined as the analysis of the rotation of polarized light by transparent components. Polarimetry is a responsive, non-destructive technique to measure the optical activity of organic and inorganic compounds. If linearly polarized light rotates when passing, it is assumed that a compound is optically active. The concentration of the chiral molecules and the molecular composition of the material determine the amount of optical rotation. The direction and extent of rotation are useful for qualitative and quantitative analysis and are also useful for the elucidation of chemical composition. Optical behavior is the ability to polarize light rotation in the determination of certain compounds.

Table of Contents

What is a polarimeter?
Principle of polarimeter
Working principle of polarimeter
Types of polarimeter
Procedure of polarimeter
Applications of polarimeter
Advantages of polarimeter
Disadvantages of polarimeter

Polarimeter principle:

The basic operation principle of a polarimeter includes a source that produces light with a specifically prepared linear polarization state, typically by going through a polarizer. The light is transmitted by an optically active sample that often rotates the polarization direction. After going through the sample, another measure of the changed angle of the polarization by transferring light through analyze, that can be rotated around the beam axis. The easiest technique is to move the analyzer to the place for which the transmission of optical power is at its lowest.

Working principle of polarimeter:

Polarimetry determines the rotation of polarized light as it travels through an optically active sample solution. A polarimeter includes a source of polarized light, filter, sample cell, analyzer, and a detector to measure the rotation angle. The determined rotation can be used to measure the concentrations of the sample particularly sugar, vitamins, peptides, and volatile oils.

Types of polarimeter:

Different types of polarimeters are mentioned below.

Manual polarimeter, fully-automatic, semi-automatic polarimeter, quartz-wedge polarimeter, biquartz polarimeter, lippich polarimeter, and Laurent’s half shade polarimeter, etc.

Polarimeter procedure:

Before beginning a polarimeter experiment, we must recognize the various components necessary to execute the process. The optical activity is calculated by the Polarimetry that consists of the source, filter, sample cell, analyzer, and detector.

Source: Sodium vapor lamps are usually used to produce wavelengths greater than 450 nm.

Filter: The filter is used to absorbing the undesired radiation of polychromatic light into monochromatic light.

Sample cell: Sample cells are used to place the sample in the sample compartment, there is made up of glass and has long tube-shaped.

Analyzer: It is mostly employed to analyze the samples when they rotate to the right or left side of the plane of polarized light.

Detector: The photomultiplier tube is broadly used for wavelength detection. The optically active component present in the solution rotates the plane-polarized light either in the clockwise direction or anti-clockwise direction. After that analyze calculates the angle of rotation and the detector detects it.

Experimental procedure of polarimeter:

Prepare the sample solutions and standard solutions as you required.
Switch the instrument to turn on and warm up for around 5 minutes.
Rotate the Polaroid wheel so that display reads “ZERO”.
Put the Polarimeter tube inside the instrument and close the cover.
Rotate the wheel back and forth to display minimal intensity. Note down the readings of degrees (X1) and don’t disturb the system.
Then take out the tube and refill it with the test solution, and put it back in its place. The intensity of the sample will rise due to the rotation of the plane of polarization.
Rotate the wheel in the direction of decreasing intensity until the intensity reaches the minima and note down the readings of degrees (X2).

Polarimeter applications:

Polarimetry is used to determine the specific rotation and optical rotation of products such as amino acids, cocaine, antibiotics, dextrose, carbohydrate, analgesics, vitamins, steroids, sugars, serums, diuretics, and codeine, etc.
For structural determination, it can be used. In this application, changes in optical rotation about the substance chemical change are estimated.
It is used to ensure product quality by determining the purity and concentration of compounds in sugar-based food, syrups, and cereals
The applications of polarimeter consist, quantitative and qualitative analysis of optically active components.
Polarimetry is used in the analysis of fragrance, flavor, and essential oil manufacturer
Chiral compounds can also be determined by polarimetry.

Advantages of polarimeter:

Polarimetry is an easy technique to operate and needs no experts to handle.
It is not affected by changes in laser intensity.
The temperature and pH within the eye remain stable.
The advanced version of the polarimeter has a broad wavelength emission range over conventional light sources.
The analysis is very easy and rapid, which means this a cost-effective technique.

Disadvantages of polarimeter:

To determine the specific and optical rotation of a substance, it required large sample volumes with high concentrations.
The major disadvantage of polarimetry is that the only optically active components can be analyzed by polarimetry.
It has low sensitivity compared to other techniques.
It is very sensitive to motion and scattering.

Commonly asked questions on polarimeter are as follows.

What is Polarimeter?

A polarimeter is an analytical tool that is used to determine the angle of rotation due to the passing of polarized light through the optically active molecule.

How does a polarimeter measure optical rotation?

A polarimeter is a tool that determines the angle of rotation bypassing the source of polarized light through the optically active material.

Which light source is used in the polarimeter?

Sodium (Na) vapor lamp is used in a polarimeter as a light source, since it produces monochromatic light and with high-energy output

What is the major advantage of a polarimeter?

The main advantage of a polarimeter is its analysis of a wide range of compounds.

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