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Thin Layer Chromatography (TLC) is an analytical technique used for separating non-volatile compounds based on their movement between a stationary phase and a mobile phase. It has a rich history, evolving since its introduction in the early 20th century, and is characterized by its simplicity, speed, and effectiveness in various applications such as drug identification and purity testing. Despite its advantages, TLC has limitations including limited resolution and difficulties in quantification.

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6 views

Thin_Layer_Chromatography_PPT

Thin Layer Chromatography (TLC) is an analytical technique used for separating non-volatile compounds based on their movement between a stationary phase and a mobile phase. It has a rich history, evolving since its introduction in the early 20th century, and is characterized by its simplicity, speed, and effectiveness in various applications such as drug identification and purity testing. Despite its advantages, TLC has limitations including limited resolution and difficulties in quantification.

Uploaded by

hareshkrishna2k3
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© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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THIN LAYER CHROMATOGRAPHY

RITU PRAKASH
A4024007
THIN LAYER CHROMATOGRAPHY (TLC)
Thin Layer Chromatography (TLC) is a simple, fast,
and widely used analytical technique for separating
mixtures of non-volatile compounds. It is based on
the movement of substances between two phases: a
stationary phase, usually a thin layer of silica gel
coated on a glass or plastic plate, and a mobile
phase, typically a solvent or a mixture of solvents.
This method enables chemists to determine the
composition of a sample, assess purity, and monitor
the progress of chemical reactions.
History of TLC
TLC was first introduced in the early 20th century
and became widely used in the 1950s as an
alternative to paper chromatography. Its simplicity,
speed, and efficiency made it a popular choice for
qualitative analysis in both academic and industrial
laboratories. Over the decades, refinements in
plate material, visualization techniques, and
solvent systems have made TLC an indispensable
tool in modern analytical chemistry.
PRINCIPLE OF TLC
• Based on the differential adsorption of substances
between a stationary phase (usually silica gel) and a
mobile phase (solvent). When a mixture is spotted on a
TLC plate and placed in a solvent, different components
travel at varying speeds due to differences in their
polarity and affinity for the stationary phase. Less polar
compounds travel further up the plate, while more
polar compounds interact strongly with the stationary
phase and move slowly. This separation forms the basis
for identifying the different constituents of a mixture.
COMPONENTS OF TLC
• 1. Stationary phase (TLC plate)
• 2. Mobile phase (solvent)
• 3. Sample
• 4. Development chamber
TLC PLATE
• - Acts as a support for the stationary phase
• - Typically 5–20 cm in length
• - Made from glass, aluminum, or plastic
• - Should be clean and grease-free
• - Provides rigidity and uniform migration
TLC PLATE PREPARATION
ACTIVATION OF PLATE
MOBILE PHASE
• Solvent or solvent mixture used to carry the
sample components up the plate by capillary
action.
• Choice depends on polarity of substances
• Should not dissolve the stationary phase
• Non-polar (hexane) or polar (ethanol, methanol)
• Trial and error used to find ideal system
STAHL’S TRIANGLE FOR M.P
SAMPLE APPLICATION
DEVELOPMENT CHAMBER
• A sealed container where the TLC plate is
placed for development. Maintains solvent
vapor saturation.
DEVELOPMENT TECHNIQUES
•Ascending Development
•Descending Development
•Horizontal Development (Radial or Circular
TLC)
•Two-Dimensional Development
•Automated or Forced Development
TYPES OF DETECTION METHODS
1.NON-SPECIFIC DETECTION METHODS
• UV Light
• Iodine Vapor
2.SPECIFIC DETECTION METHODS
• Ninhydrin Spray – for amino acids and amines
• Dragendorff’s Reagent – for alkaloids
• Sulfuric Acid Spray – for general organic compounds
• Anisaldehyde Spray – for terpenes and steroids
• Vanillin Spray – for sugars and phenols
ANALYSIS
1. Direct Methods
• Visual Observation-Spot colour, intensity, and separation pattern.
• Rf Value Calculation- Rf = (Distance travelled by compound) / (Distance travelled by
solvent front)
• Comparison with Standards- Match unknown spots with known reference samples.
2. Indirect Methods
• Densitometry
• Quantitative analysis using a densitometer.
• Measures spot intensity and area.
• UV/Fluorescence Spectroscopy
• Detection based on UV absorbance or fluorescence.
• Chemical Derivatization
• Spots treated with reagents to enhance visibility or reactivity.
• Helps detect compounds not visible under normal conditions.
FACTORS AFFECTING TLC
1. Type of stationary phase
2. Solvent polarity
3. Sample concentration
4. Temperature and humidity
ADVANTAGES OF TLC
• Quick and simple
• Inexpensive
• Can analyze multiple samples simultaneously
• Minimal sample required
DISADVANTAGES OF TLC
• Limited resolution
• Difficult quantification
• Not suitable for volatile substances
APPLICATIONS OF TLC
• Drug identification
• Purity testing
• Food and cosmetic analysis
• Plant extract analysis
• Ink analysis
• Forgery
CONCLUSION
TLC is a versatile and essential technique in
chemical and pharmaceutical analysis, offering
simplicity and effectiveness.
REFERENCES

•Stahl, E. (1969). Thin-layer chromatography: A laboratory handbook.


Springer-Verlag.
•Touchstone, J. C. (1992). Practice of thin layer chromatography (3rd
ed.). Wiley-Interscience.
•Poole, C. F. (2012). The essence of chromatography. Elsevier.
•Srivastava, M. M. (2001). Chromatographic techniques. Alpha Science
International Ltd.
•Sherma, J., & Fried, B. (2003). Handbook of thin-layer chromatography
(3rd ed.). CRC Press.
•Skoog, D. A., Holler, F. J., & Crouch, S. R. (2013). Principles of
instrumental analysis (6th ed.). Cengage Learning.
THANK YOU

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