Column chromotography | Principle, Technique, procedure, applications | Bio science

Bio science
21 Nov 201914:02
EducationalLearning
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TLDRThis educational video script delves into the principles and procedures of column chromatography, a technique used for separating molecules in a mixture. It explains the roles of the stationary phase (often silica or alumina) and the mobile phase (a solvent), and describes the apparatus setup, including a glass tube, cotton wool, and a plug or tap. The script outlines the steps for packing the column using either the dry or wet method, adding the sample, and eluting the components. It also touches on the mechanism of separation based on polarity and adsorption capacity, and concludes with applications such as separating geometrical isomers, enzymes, and tautomeric mixtures, encouraging viewers to subscribe for more updates.

Takeaways
  • 🔍 Column chromatography is a technique used for the separation of molecules in a chemical compound mixture.
  • 🧪 The stationary phase in column chromatography is typically a solid, such as silica or alumina, while the mobile phase is a liquid, such as a solvent.
  • 📐 The apparatus for column chromatography includes a glass tube with a large inlet and a small outlet, a plug or tap to prevent leakage, and cotton wool to prevent the mixture from flowing out.
  • 📦 The packing of the column involves adding the stationary phase (adsorbent) and the mobile phase (solvent) together, which can be done using either the dry or wet method.
  • 💧 In the dry method, dry silica powder is added to the glass tube followed by the addition of a suitable solvent, causing the stationary phase to dissolve into the mobile phase.
  • 🌊 The wet method involves creating a slurry of silica powder and solvent before adding it to the column, using a funnel or injection.
  • 📈 The procedure of column chromatography involves adding the sample mixture to the stationary phase, adding solvent to initiate separation, and then adding more solvent to complete the separation process.
  • 🔬 The separation mechanism is based on the polarity and adsorption capacity of the molecules, with low polarity molecules eluting first, followed by medium and high polarity molecules.
  • 🌈 The script provides an example of how different colored components represent molecules with varying polarities and how they are separated during the process.
  • 📝 Applications of column chromatography include the separation of geometrical isomers, enantiomers, and tautomeric mixtures, as well as the use of polymers, starch, and powdered sugar as stationary phases for separating different types of compounds.
  • 🔔 The video encourages viewers to subscribe for updates and to comment with any doubts for further clarification.
Q & A
  • What is the main aim of chromatography?

    -The main aim of chromatography is the separation of molecules from a given mixture or sample using a specific technique.

  • What distinguishes column chromatography from other chromatographic techniques?

    -Column chromatography is distinguished by its use of a solid stationary phase, such as adsorbent, and a liquid mobile phase, such as a solvent.

  • What are the two phases used in column chromatography?

    -The two phases used in column chromatography are the stationary phase, which is solid and often an adsorbent like silica or alumina, and the mobile phase, which is a liquid such as a solvent.

  • What is the role of the glass tube in column chromatography?

    -The glass tube serves as the container for the column chromatography setup, with a large inlet for adding the mixture and a smaller outlet for the elution of the separated components.

  • Why is cotton wool or glass wool used in the glass tube during column chromatography?

    -Cotton wool or glass wool is used to prevent the flow of mixture components into the outer environment and to ensure that the stationary phase is held in place within the glass tube.

  • What are the two methods for packing the column with the stationary phase in column chromatography?

    -The two methods for packing the column with the stationary phase are the dry method, where dry silica powder is added followed by a suitable solvent, and the wet method, where a slurry of silica powder and solvent is prepared and then added to the column.

  • What is the purpose of adding the sample mixture to the stationary phase in column chromatography?

    -The purpose of adding the sample mixture to the stationary phase is to initiate the separation process, where the components of the mixture will interact with the stationary phase and begin to separate based on their affinity to it.

  • How does the addition of solvent affect the separation process in column chromatography?

    -The addition of solvent increases the pressure within the column, which helps to elute the separated components out of the column. The solvent also interacts with the stationary phase, aiding in the separation of the components based on their polarity or adsorption capacity.

  • What is the significance of the order in which components are eluted in column chromatography?

    -The order of elution is significant as it indicates the relative polarity or adsorption capacity of the components. Typically, components with lower polarity or adsorption capacity are eluted first, followed by those with higher polarity or adsorption capacity.

  • What are some applications of column chromatography mentioned in the script?

    -Some applications of column chromatography include the separation of geometrical isomers, separation of enantiomers, separation of tautomeric mixtures, and the use of polymers like nylon for separating sugars and fatty acids, starch for separating nucleotides and the power of sugar for separating pigments.

Outlines
00:00
🧪 Introduction to Column Chromatography

This paragraph introduces the concept of column chromatography, a technique used for separating molecules within a mixture. It explains the principle behind chromatography, emphasizing the use of a stationary phase in solid form, such as silica or alumina, and a liquid mobile phase, typically a solvent. The video will cover the principle, experimentation, packing of the column, the procedure, mechanism, and applications of column chromatography. The stationary phase is packed into a glass tube, which is prepared with specific features like a large inlet for adding mixtures and a small outlet for eluting the separated components. Cotton wool is used to prevent leakage and ensure the mixture remains within the tube.

05:01
🔬 Experimentation and Packing of the Column

The second paragraph delves into the experimental setup for column chromatography, detailing the preparation of the glass tube and the process of packing the column with the stationary phase. Two methods for packing the column are discussed: the dry method, where dry silica powder is added directly to the tube followed by a suitable solvent, and the wet method, which involves creating a slurry of silica powder and solvent before adding it to the column. The paragraph also explains the steps involved in the procedure of column chromatography, including adding the sample mixture to the stationary phase and using solvent to facilitate the separation of components, which are initially in a crowded form but eventually separate clearly as more solvent is added.

10:01
🌟 Mechanism and Applications of Column Chromatography

The final paragraph explains the mechanism behind the separation process in column chromatography. It describes how particles with different polarities are affected by the addition of solvent, leading to the elution of low polarity molecules first, followed by medium and then high polarity molecules. The paragraph also provides examples of applications for column chromatography, such as the separation of sugars and fatty acids using polymers like nylon, the separation of resonances using starch, and the separation of pigments using powdered sugar. Additionally, it mentions the technique's use in separating geometrical isomers, enantiomers, and tautomeric mixtures. The video concludes by encouraging viewers to subscribe for more updates and to leave comments with any questions.

Mindmap
Keywords
💡Column Chromatography
Column chromatography is a technique used for separating the components of a mixture based on their affinity to the stationary phase versus the mobile phase. In the video, it is the main topic discussed, including its principle, procedure, and applications. The script explains that the stationary phase is typically an adsorbent like silica or alumina, while the mobile phase is a liquid solvent.
💡Chromatographic Techniques
Chromatographic techniques are a set of methods used to separate mixtures into their individual components. The video script introduces column chromatography as the second technique in a series, emphasizing its importance in the study of chemical compounds and mixtures.
💡Stationary Phase
The stationary phase in chromatography is the medium that remains stationary throughout the process, providing a surface for the separation of compounds. In the script, it is mentioned that the stationary phase in column chromatography is in solid form, such as silica or alumina, which acts as an adsorbent.
💡Mobile Phase
The mobile phase is the component of the chromatographic system that moves through the stationary phase, carrying the sample mixture with it. The script describes the mobile phase as a liquid solvent that interacts with the stationary phase to facilitate the separation of molecules.
💡Adsorbent
An adsorbent is a material that has the ability to adsorb molecules onto its surface, which is crucial for the separation process in column chromatography. The script specifies silica or alumina as examples of adsorbents used in the stationary phase.
💡Glass Tube
The glass tube is the physical container used in column chromatography to hold the stationary phase and allow the mobile phase to pass through. The script describes its dimensions and features, such as a large inlet for adding the mixture and a small outlet for the elution of separated components.
💡Cotton Wool
Cotton wool is used in the glass tube to prevent the leakage of the stationary phase and to ensure that the mixture does not escape the system. The script mentions that it is placed at the base of the glass tube before the addition of the stationary phase.
💡Dry Method
The dry method is one of the two methods described in the script for packing the column with the stationary phase. It involves adding dry silica powder directly to the glass tube, followed by the addition of a suitable solvent to form a slurry within the tube.
💡Wet Method
The wet method is the alternative to the dry method for packing the column in column chromatography. As described in the script, it involves creating a slurry of silica powder and solvent before adding it to the column, which helps to evenly distribute the stationary phase.
💡Elution
Elution is the process by which the separated components are carried out of the column by the mobile phase. The script explains that after the components have been separated within the column, they are eluted out through the outlet, which is a key step in obtaining the purified substances.
💡Polarity
Polarity refers to the property of molecules that have an uneven distribution of electron charge, leading to the formation of partial positive and negative charges. In the context of the script, polarity affects how molecules interact with the stationary phase and thus influences their elution order in column chromatography, with less polar molecules eluting before more polar ones.
Highlights

Introduction to column chromatography as a technique for separating molecules within a mixture.

Explanation of the principle behind chromatography, focusing on the separation of molecules.

Distinction of column chromatography from other chromatographic techniques by its use of a solid stationary phase and a liquid mobile phase.

Description of the apparatus required for column chromatography, including a glass tube with specific dimensions and features.

Importance of the plug or tap in preventing leakage of solvents and mixtures during the chromatography process.

Role of cotton wool or glass wool in the column to prevent the flow of mixture components into the environment.

Process of packing the column with stationary phase adsorbents like silica or alumina and the addition of the mobile phase.

Two methods of column packing: the dry method and the wet method, each with its own technique for adding silica and solvent.

Detailed procedure of column chromatography involving four steps, starting with the addition of the sample mixture to the stationary phase.

Mechanism of separation in column chromatography, where components separate based on their polarity and adsorption capacity.

Explanation of how low polarity molecules are eluted first, followed by medium and high polarity molecules.

Practical applications of column chromatography in the separation of various compounds, such as sugars, fatty acids, pigments, and more.

Use of different adsorbents like polymers, starch, and powdered sugar for specific separation tasks.

Discussion on the separation of geometrical isomers, enantiomers, and tautomeric mixtures as applications of column chromatography.

Invitation to subscribe to the channel and press the bell icon for updates on the latest videos.

Encouragement for viewers to comment with any doubts regarding the video content for further clarification.

Transcripts
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