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Friday, January 31, 2020

Principle and Procedure of Column Chromatography

Learn the principle of column chromatography which is works on four different mechanisms. Also, learn its types, applications, advantages, and how to do column chromatography.

What is column chromatography?

Column chromatography is one of the most effective methods for separating and purifying solid and liquid samples, in which the stationary phase is a solid and a liquid is a mobile phase. In column chromatography, the column is filled with solid support, (silica gel) and a mobile phase, the sample component is placed on top of the column, and then it moves from the solid support under the force of gravity. 

The different sample components to be separated migrate at different rates through the column. The components are separated based on their varying affinity for the stationary phase, hence they are separated into fractions.
Column chromatography

Column chromatography principle:

The principle involved in column chromatography is that the adsorption of the solutes of a mobile phase through a stationary phase and separates the combination into individual analytes. It is based on the affinity towards the stationary phase and mobile phase. The molecules that have a lower affinity for the stationary phase that they elute rapidly, and molecules that have more affinity for the stationary phase elute slowly.


Before starting the column chromatography experiment, we must understand the various steps required to perform the process.
Adsorbent or Stationary Phase:
The alumina and silica gel is two commonly used adsorbents for column chromatography. Both are the solid material that has a good adsorption property, are chemically inert, have high mechanical stability and are suitable for separating different molecules.

Mobile Phase or solvent mixture:
The mobile phase is a solvent or solvent mixture, it acts as an eluting agent and it helps separate the sample molecules.


Types of column packing in column chromatography:

Typically, two methods are used to prepare columns.

Dry method: In this process, the column is filled with dry silica powder. Subsequently, an appropriate solvent or mobile phase is flushed through it until all the silica is wet and settled. The column still needs to keep wet with solvent from this point until the end.

Wet method: In the wet packing, silica and solvent slurry is prepared and then transfer into the column.

Experimental procedure of column chromatography:

  • Put the column in a vertical position with the help of an appropriate stand.
  • Place a glass wool plug at the bottom of the column.
  • There are different ways to prepare columns but most analysts preferred slurry methods. Prepare the silica gel slurry with an appropriate solvent, and then gently pour it into the column.
  • Open the tap and drain out the certain solvent. The mobile phase layer should always cover the adsorbent; if not, cracks in the column will form.
  • Add the sample mixture to the column in such a way that it does not disturb its upper layer.
  • Cautiously open the stop-cock and maintain the solvent level. The solvent needs to be repeatedly added as needed during the entire process.
  • The components of the mixture run down the column and separate according to their polarity (forming separate bands).
  • Collect the constituents individually when it arrives at the end of the column.

Types of column chromatography:

There are four types of column chromatography available based on the stationary phase, and they operate on various mechanisms.
1. Adsorption chromatography: This is a separation process, in which the mixture components are adsorbed on the adsorbent surface.

2. Ion-exchange chromatography: In which ion exchange resin is the stationary phase.

3. Partition chromatography: For partition chromatography both the stationary phase and the mobile phase is liquid.

4. Gel chromatography: In which the separation is carried out in a column filled with gel.

Column chromatography applications:

  • Separating compound mixtures make column chromatography very effective.
  • The column chromatography can be used to separate drugs and components of several classes such as aggregates, plant extracts, amino acids, glycosides, and alkaloids.
  • Impurities can be isolated in a formula, using the correct mobile phase and stationary phase.
  • The pharmaceutical industries usually employ column chromatography to purify compounds.
  • It is used for estimating drugs from drug formulations.
  • It is possible to separate essential constituents from the crude extract active constituents, plant extracts, and formulations.
  • Column chromatography is a flexible method of purification used to separate compounds within a solution.

Advantages of column chromatography:

  • Column chromatography can be used to separate all kinds of complex mixtures.
  • It is an inexpensive, robust, and versatile method of separation.
  • Column chromatography can isolate both small and large quantities of mixtures.
  • It has a wide range of mobile phages.
  • In preparative type chromatography, the molecules can be separated and reused.
  • Automation can be implemented.

Disadvantages of column chromatography:

  • It takes longer to separate the compounds by this method.
  • You need the expertise to handle the equipment and process.
  • It is less accurate and precise than other chromatographic techniques.
  • Automation makes things more complex and expensive.
  • This requires more solvent to separate the components.

The questions commonly asked about chromatography are as follows.

What is column chromatography?
Column chromatography is a commonly used method of separating the compounds from complex mixtures. Chromatography, when performed in a column, is called column chromatography.

What is the basic principle of Column Chromatography?
The basic principle involved in column chromatography is to adsorb solutes of the solution using a stationary phase and separate the sample mixture further into individual components.

What is the principle of adsorption chromatography?
Adsorption chromatography is based on the principle that some adsorbents (solid material) will hold the molecule on their surface.

What is the major advantage of Column Chromatography?
The main advantage of column chromatography is that any type and amount of mixture can be separated by a wider choice of mobile Phase.

What is the  difference between column chromatography and paper chromatography?
The major difference between column chromatography and paper chromatography is that paper chromatography works under gravity and column chromatography works against gravity. another major difference between column and paper chromatography is that the stationary phase used in the separation.

Why TLC used before column chromatography?
Column chromatography works like TLC and uses both similar stationary and mobile phases. Therefore the thin layer chromatography is used to obtain a suitable solvent system and Rf value for column chromatography.


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Wednesday, January 29, 2020

What are the good things about junk food?

Junk food is a kind of food that is high in calories from fat or sugar. It also contains some vitamins, minerals, dietary fiber, protein and other important forms of nutritional value.
Junk food and fast food are generally known for their unhealthy effects, although it also has several good benefits. They do not only give nutrients to the body but also providing economic benefits to the people. They appear to be the simplest and quickest way to get food. Junk food tends to spoil good eating habits by most adolescents, especially in colleges. A healthy body requires well-balanced diets that increase the proper building of the body's muscles.
  • There is no bad or unhealthy food for us, it is bad for our health if we have habitual to it and people forget to eat healthily and work out.
  • Many fast-food chains are changing their menus since consumers are more health-conscious.
  • Although most foods are good to eat occasionally, several fasts or junk foods menu options are healthier selections than others.
  • The availability of nutritional information is one of the major advantages of the food chain as compared to other formal eateries.
  • Knowing the nutrition of the foods we take, you can choose a healthy option to stay fit.
  • Cooking in fast-food chains is cooked using healthy methods. This is another positive change in the fast-food world.
  • In addition to foods prepared with fat reduction methods, healthy items are often available in fast food menus.
  • Apart from food prepared with fat reduction methods, healthy items are often available in fast food menus.
  • Junk food is often available anywhere and anytime; it can eradicate our hunger and give instant energy.
  • Homemade healthy food takes longer to cook than junk food, so people like it more in the workplace.
  • Junk food is infamous because it causes obesity; it is good if we take it occasionally.
  • Many alcoholic beverages are rich in antioxidants. This helps to keep the body healthy if consumed in the right amount.
some examples of healthy junk food popcorn, dark chocolate brownies, processed cheese, chocolates, ketchup, red wine, butter, cheese, salad, peanut butter, juice (fruits), milk or dairy products, low carb, whole wheat, and vegetable oil, etc.


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What is fast food?
What are the types of fast food?
Fast food and obesity
Effects of fast food on health
Harmful effects of fast food on health
What are the advantages of fast food?
Advantages and disadvantages of fast food
Advantages and Disadvantages of Junk Food
What are the Advantages of Junk Food
What are the Disadvantages of Junk Food?
Why Junk Food is so Popular?
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Why Junk Food is so Popular?

Taste, cost, availability, and convenience are the most prominent reasons that make junk food more popular among people. There is no bad food for you, just you should not try to eat regularly and make it a habit. Since junk and fast food are high in cholesterol, saturated fat, Trans fat, and sodium you should not eat often. In the early years, junk food and fast food are becoming very popular, although many tests confirm that it can harm our health. Junk food is tasty, filling, and economical but it is associated with different problems such as obesity, high blood pressure, heart problems, and diabetes.

Why junk food popular


Here are some points mentioned as to why junk food is so popular.
  • It is easy to make, and many of them taste good, consuming them becomes a habit.
  • Junk food is more popular than healthy food because it is cheaper.
  • Since fast foods are easily available nearby or around the market, they are more preferred by people.
  • Homemade food takes longer to cook than junk food.
  • Junk food is also popularized by its visual appeal and the good service provided by the restaurant.
  • Junk food attracts consumers, especially children, with its visual appearance.
  • Many people do not have time to prepare food by themselves; some are away from their family for work, in such situations they like fast food.
  • Nowadays fast-food chains have many nutritious food options.
  • They are convenient to take you anywhere and eat.
  • Food is promoted in various ways such as advertisements with animations and cartoons that attract people.


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What is fast food?
What are the types of fast food?
Fast food and obesity
Effects of fast food on health
Harmful effects of fast food on health
What are the advantages of fast food?
Advantages and disadvantages of fast food
Advantages and Disadvantages of Junk Food
What are the Advantages of Junk Food
What are the Disadvantages of Junk Food?

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What are the Disadvantages of Junk Food?

The Disadvantages of Junk Food are as Follows.
  • The more we consume junk food, the fewer intakes of nutrients that a healthy body needs.
  • The fast-food can harm our health; obesity is one of the most common side effects of intake such type of food.
  • Junk food contains calories and fat which contributes to weight gain. Obesity can cause many health problems such as heart diseases, joint pain, and diabetes.
  • Excess intake of junk food causes the body to lose important nutrients and amino acids.
  • Low-quality junk foods are inadequate of vitamins and nutrients necessary for proper growth and development of the body.
  • It is a processed food that is low in nutritional value. Excessive consumption can lead to high consumption of calories, sodium, and saturated fat.
  • These types of foods often have an unbalanced ratio of fat, refined carbohydrates, and sugar.
  • Eating in excess can increase the "bad" cholesterol in your bloodstream which endangers your heart health.
  • Junk food can cause people to suffer from obesity because it has an unbalanced ratio of sugars, fats, and carbohydrates.
  • Fast food can be rich in sugar it may lead to developing cavities (tooth decay) in children as well as elders.
  • Eating too much fast food can cause digestion problems.
  • Refined carbohydrates present in junk food that alter blood sugar levels can also cause fatigue, anxiety, confusion, and blood pressure.
  • Nowadays a major problem with junk food is that people can like it a lot more than healthy food and that tend to replace healthy foods.
Some examples of junk food are Chips, Burgers, Snacks, Cold drinks, Chocolates, Fried, Stuff, Ice Cream, Potato Chips, Pizza, Cakes, Donuts, Candies, Noodles, Cheese Stuff, and Soft Drinks, etc.


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What is fast food?
What are the types of fast food?
Fast food and obesity
Effects of fast food on health
Harmful effects of fast food on health
What are the advantages of fast food?
Advantages and disadvantages of fast food
Advantages and Disadvantages of Junk Food
What are the Advantages of Junk Food
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What are the Advantages of Junk Food

Junk food is a kind of food high in calories (fat, sugar). It has some amount of minerals, vitamins, protein, dietary fiber, and significant nutritional value. Junk foods are processed foods consisting of high calories, though not all fast foods are junk foods. Junk foods are processed foods consisting of high calories it may cause obesity, but if you take them carefully you can maintain your body weight.
The Advantages of Junk Food are as Follows.
  • Fast food is very good in taste and they are also available in different types that are why it is addictive for us.
  • Junk food is cheaper than other foods. This is the biggest benefit of junk food.
  • As compared to cooking lunch or dinner at home, selecting junk food saves 50% of the time.
  • The fast-food industry provides significant employment and economic boost.
  • It is a time-saving food that helps you manage work schedules more efficiently.
  • When you go to eat at fast-food restaurants, they are served more beautiful.
  • Junk food items are cheaper foods than other foods that give you more calories.
  • Nowadays fast food restaurants provide a healthier option than ever before.
  • Fast food packing looks good and they provide you with all the nutritional information for each item.
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What are the causes of broad peaks in HPLC?

The presence of unusually broad peaks is sometimes an occurring problem in liquid chromatography separation. This problem is regularly seen in isocratic separation; however, it can also occur with gradient elution. The peak broadening in HPLC is influenced by the flow rate of the mobile phase and particle size of the columns packing material. The smaller the particle with the latter, the better is the peak resolution. It is recommended that the chromatographers can try and optimize the composition and flow rate of the mobile phase and check the type of column used.
The peak broadening in high-performance liquid chromatography is due to several factors mentioned below.
  • In a very short time, several steps of gradient elution are changing from polar to non-polar.
  • Some precipitate in the sample solution.
  • Overloading of column (too much sample load)
  • Too long Retention time (RT)
  • Extra-column volume too large
  • Detector time too slow
  • Poor solubility of the sample
  • Incompletely filled sample loops
  • Saturation of the Detector
  • Contaminated column
  • Incorrect pH of the buffer or mobile phase 
 
 
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Monday, January 27, 2020

Principle and Procedure of Thin Layer Chromatography

Learn the principle of TLC, types of TLC, applications of TLC and how its works.

What is Thin Layer Chromatography?

Thin-layer chromatography is a technique in which the analytes of a mixture are separated by differential migration through the stationary phase. TLC is performed on glass, aluminum or plastic sheets. It is coated with a thin layer of adsorbent material, typically made using silica gel, aluminum oxide, or cellulose and is known as a stationary phase. After applying the sample to the TLC plate, a mobile phase (solvent mixture) is drawn up to the plate by capillary action. Since each analyte has a different affinity to the TLC plate; hence separation is achieved at different rates.

Thin Layer Chromatography Principle:

TLC is a separation technique where the molecules of mixtures separate using differential migration through a stationary phase, the solvent mixture flowing through the virtue of capillary forces. After chromatography is complete, solutes are detected on the surface of a TLC plate by visualizing the reagents and using a UV cabinet. TLC is a type of planar chromatography, such as paper chromatography, but the stationary phase in TLC is a thin-split Sorbent, which spreads as a thin layer of aluminum, glass, or plastic supporting flat plate.

Thin Layer Chromatography Diagram:


Principle and Procedure of Thin Layer Chromatography (TLC)

Thin Layer Chromatography Procedure:

Before beginning a TLC experiment, we must recognize the various components essential to perform the process.
Thin Layer Chromatography Chamber: To develop the plates, the TLC chamber is used. This maintains a stable environment and prevents solvent evaporation, which helps spots develop.
Thin Layer Chromatography Plates: The separation occurs on the TLC plate, in which the stationary phase is applied as a thin layer on its surface. Nowadays, ready plates are available in the market. They are chemically inert, stable and of high quality.
Thin Layer Chromatography Mobile phase: A solvent or mixture of solvents with suitable polarity is used as the mobile phase to achieve the required separation.

Experimental Procedure of TLC:

  1. The adequate mobile phase is poured into a TLC chamber to give liquid about 0.5 cm deep. Subsequently, cover the chamber for a few minutes to allow the atmosphere to become saturated with solvent vapor.
  2. The small amount of liquid sample is “spotted” on the TLC plate with close to an edge and parallel with an edge. The plate is then placed in contact with the mobile phase in the chamber and is covered.
  3. The mobile phase is drawn from the edge of the TLC plate through the Sorbent bed, mainly by capillary action, Allow the development process until the mobile phase is approximately half a centimeter below the top of the TLC plate. The mobile phase and sample are displaced in parallel passages in the direction of flow, and the components separate according to their affinity.
  4. Remove the plate from the chamber and mark it immediately in front of the solvent.
  5. Allow the plate to dry with warm air or room temperature.
  6. Circle the visual spot lightly with a pencil. Most analytes are not colored they are to be visualized by spraying chromogenic reagent or in UV cabinet. Mark the sample spot and calculate the Rf value. The retention factor (Rf value) will always be in the range 0 to 1.

Thin Layer Chromatography Applications:

  • It is used to separate and identify the number of components present in a sample mixture.
  • TLC can be used to select the appropriate mobile phase system in column chromatography.
  • It is used for the separation and identification of sweetening agents, preservatives, dyes, and different cosmetic products.
  • It is used for the separation and identification of food products, pharmaceutical formulations, and natural products.
  • Thin-layer chromatography can be used to monitor the progress and the rate of reactions at particular intervals.

Advantages of Thin Layer Chromatography:

  • TLC is a very easy and sensitive mode of separating analytes.
  • The molecules are separated in a short period, as they are quickly eluted.
  • Very small amounts of the sample need to be analyzed.
  • A complex mixture of sample solution can be easily separated and recovered.
  • This can be automated using high-performance thin-layer chromatography (HPTLC).
  • This requires very few types of apparatus.

Disadvantages of Thin Layer Chromatography:

  • Only non-volatile compounds are separated by this method.
  • It is difficult to reproduce to obtain from TLC.
  • The separation length in TCL is insufficient, as it does not have a long stationary phase.
  • Humidity and Temperature can affect TLC results.
  • Another disadvantage of TLC is that it only analyzes soluble analytes.
Commonly asked questions on chromatography are as follows.

What is the principle of TLC?
TLC comprises a stationary and mobile phase, in which a thin layer of alumina or silica is coated on glass, metal, or plastic that is a stationary phase and a solvent or mixture of solvents is a mobile phase. The sample mixture travels through the stationary phase with the mobile phase and separates according to the degree of their adhesion.

What type of chromatography is TLC?
Thin-layer chromatography is a type of planar chromatography.

What is the TLC plate made of?
Generally, TLC plates are made using silica gel, cellulose, aluminum oxide or similar material. A thin layer of special granular ground matrix is coated as on a glass plate, aluminum or a plastic film.

What does TLC Rf value mean?
It is the proportion of separation traveled by an analyte to distance traveled by a mobile phase front. It is used to identify analytes due to their specificity for each compound.

What is the major advantage of thin-layer chromatography?
The major advantage of TLC is that many samples can be analyzed simultaneously over rapid time.

What is the major difference between TLC and HPTLC?
The major difference between TLC and HPTLC is that the particle size of the absorbing material. An HPTLC plate contains very small particles of adsorbent material while the TLC contains larger particles.


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