Paper & TLC Chromatography

Dr Deep Sir

1. ‹#› Paper Chromatography + Thin Layer Chromatography Dr. Deepkumar Joshi Assistant Professor Chemistry Department Sheth M. N. Science College, Patan

2. Classification of Chromatography Chromatography is generally classified on different basis, but the most popular is on the shape of the stationary phase. 1. Planar Chromatography. 2. Column Chromatography. Planar Chromatography is further divided into Paper Chromatography and Thin layer Chromatography. ‹#›

3. Planar Chromatography Planar Chromatography is a technique in which stationary phase is present on a plane; which can be paper or a glass of variant size. If the plane used is a paper i.e. Cellulose then it is called Paper Chromatography, where as if glass is used then it is called Thin Layer Chromatography, popularly TLC. In TLC the stationary phase is generally Silica gel G. G in silica gel G stands for Gypsum. ‹#›

4. This Chromatography involves placing a small drop of sample solution on a paper, the paper serves as a stationary phase . The paper is placed in a jar containing small layer of mobile phase at the bottom. The mobile phase slowly travels along the paper and carries the sample in the direction and thus separates components from the mixture. Further this chromatography technique is subdivided into 3 types. Paper Chromatography ‹#›

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6. This is a sub type of paper chromatography, where a paper in a square shape is kept on a paltry dish filled with mobile phase which travels through the paper from the centre to the periphery of the circular paper. A wick made from the same paper is used for the displacement of mobile phase from reservoir to paper. Wick Circular Chromatography ‹#›

7. In ascending paper chromatography a paper is kept hanging from a support into a jar filled with mobile phase such that the end of paper just touches the mobile phase. The mobile phase runs upward carrying with it the sample up till a particular height. All the components in a mixture gets separated depending upon their polarity. Analytes are Separated According To polarity Mobile phase Ascending Chromatography ‹#›

8. A process with same application as ascending chromatography is demonstrated here only with a change in flow direction of mobile Phase. Here the reservoir is placed at the top of the jar and the mobile phase flows upside down. The sample here gets separate at the bottom of the paper. In all types of paper chromatography the sample that has more attraction towards Cellulose takes more time to separate . Descending Chromatography ‹#›

9. R f values : Retardation Factor Calculating the R f value ‹#›

10. Calculate R f Value ‹#›

11. The R f value needs to be between 0.0 and 1.0 If the value is over 1.0 or less than 0.0, the calculation is wrong If the R f value is greater than 0.8 or lower than 0.2 the values are hard to interpret, thus creating a larger error The best R f values are 0.3 to 0.6 R f Value ‹#›

12. R f Value What affects the R f value? Temperature Solvent Thickness of spot Concentration of the sample impurities in sample Type of Paper (material, porosity, etc.) ‹#›

13. Compare to paper chromatography, Thin Layer Chromatography is more beneficial. In TLC the mobile phase runs faster than Paper. Here the choice of using variety of mobile phase is easy as silica gel is more inert than cellulose. Separation is more better than Paper chromatography. Analytes are Separated According To polarity Mobile phase Comparing TLC with Paper Chromatography ‹#›

14. Components Affinity to Stationary Phase Affinity to Mobile Phase Blue ---------------- Insoluble in Mobile Phase Black ✓ ✓ ✓ ✓ ✓ ✓ ✓ ✓ Red ✓ ✓ ✓ ✓ ✓ ✓ ✓ Yellow ✓ ✓✓✓ ✓ ✓ ✓ ✓ ✓ ✓ ✓ ✓ Mixture Components Separation Stationary Phase Mobile Phase Similar concept for TLC ‹#›

15. Preparing A TLC??? Mix Water or suitable solvent to the absorbent. Spread a thin layer of absorbent on an unreactive hard surface (Glass, plastic, thick aluminium) Heat in oven at 110°C for 30 mins to activate and dry the plate Place a small amount of solvent in a beaker Place a drop of sample solution on the line ‹#›

16. Solvents Choose a solvent depending on the polarity of the compound Least Polar More polar Petroleum ether Cyclohexane Toluene Chloroform Acetone Ethanol Methanol ‹#›

17. Visualization Destructive visualization Spray plate with H 2 SO 4 , and then bake in the oven at 110ºC for 15-20 minutes. Compound is destroyed but all spots will be visible Non-destructive visualization – because of the use of a UV light the sample will not be destroyed. Although, not all of the spots on the plate will be visible. Long wave UV Short wave UV Semi-destructive visualization ‹#›

18. Chromatography is an infinite techno………………………. Column Planar Chromatography HPLC UPLC Paper TLC GC ‹#›

19. Books for Chromatography Introduction to modern liquid chromatography by Llyod R Synder Principles and Practice of Chromatography by Raymond P. W. Scott, Useful e-links biopharminternational.findpharma.com/... Basics ... Chromatography /.. www.youtube.com/watch?v=g3m2OgXmnYM ‹#›

20. Experiment to Separate Silver , Mercury and mix ture!!! Take the solution containing the mixture of these two metals in form of their corresponding compounds. Drop a spot on the paper and keep into the mobile phase and run the water for the given time. Dry it and then spry with dichromate solution to make the spots appear clearly. The three spots will be observed corresponding to the metals…. Calculate the retardation factor value for each…….. ‹#›