Your SlideShare is downloading. ×
  • Like
IB Chemistry on High Performance Liquid Chromatography and Gas Liquid Chromatography
Upcoming SlideShare
Loading in...5
×

Thanks for flagging this SlideShare!

Oops! An error has occurred.

×

Now you can save presentations on your phone or tablet

Available for both IPhone and Android

Text the download link to your phone

Standard text messaging rates apply

IB Chemistry on High Performance Liquid Chromatography and Gas Liquid Chromatography

  • 1,465 views
Published

IB Chemistry on High Performance Liquid Chromatography, Gas Liquid Chromatography, Thin Layer and Column Chromatography.

IB Chemistry on High Performance Liquid Chromatography, Gas Liquid Chromatography, Thin Layer and Column Chromatography.

Published in Education
  • Full Name Full Name Comment goes here.
    Are you sure you want to
    Your message goes here
    Be the first to comment
    Be the first to like this
No Downloads

Views

Total Views
1,465
On SlideShare
0
From Embeds
0
Number of Embeds
26

Actions

Shares
Downloads
88
Comments
0
Likes
0

Embeds 0

No embeds

Report content

Flagged as inappropriate Flag as inappropriate
Flag as inappropriate

Select your reason for flagging this presentation as inappropriate.

Cancel
    No notes for slide

Transcript

  • 1. Chromatography Techniques Partition Chromatography Adsorption Chromatography Partition Chromatography Adsorption Chromatography • Components distribute bet TWO immisible liquid phase • Component adsorb on solid stationary phase • Depends on relative solubility bet TWO phase • Depends on polarity of stationary phase, mobile phase and solutes • Solutes bonds to stationary phase or mobile phase • Stationary phase is polar – polar solutes adsorb strongly • Stationary phase is non polar – non polar solutes adsorb strongly • Mobile phase is polar – polar solutes stay in mobile phase • Mobile phase is non polar – non polar solutes stay in mobile phasePaper Chromatography GLC HPLC Thin Layer Chromatography Column Chromatography Y adsorb strongly X adsorb strongly Mobile phase – liquid Mobile phase – gas Mobile phase – liquid Mobile phase – liquid Mobile phase – liquid Stationary phase - liquid Stationary phase - liquid Stationary phase - liquid Stationary phase - solid Stationary phase - solid
  • 2. Paper Chromatography,Thin Layer Chromatography Colum Chromatography
  • 3. Chromatography Techniques Chromatography Techniques • Separation technique of mixture into their pure components • Identify if sample is mixture or pure both quantitatively and qualitatively • Interaction of substances between 2 phases - Stationary phase and Mobile phase • Separation is based on Partition or Adsorption Separation analysis Chromatography Partition Chromatography Adsorption Chromatography Partition Chromatography Adsorption Chromatography• Components distribute bet TWO immisible liquid phase • Component adsorb on solid stationary phase• Depends on relative solubility bet TWO phase • Depends on polarity of stationary phase, mobile phase and solutes• Solutes bonds to stationary phase or mobile phase • Stationary phase is polar – polar solutes adsorb strongly • Stationary phase is non polar – non polar solutes adsorb strongly • Mobile phase is polar – polar solutes stay in mobile phase • Mobile phase is non polar – non polar solutes stay in mobile phase Paper Chromatography Thin Layer Chromatography Column Chromatography Y adsorb strongly X adsorb strongly Application Application Application • Detection of amino acids in a mixture • Detection of amino acids in a mixture • Preparative/collection of sample • Different dyes in food colouring • Different dyes in food colouring of pigments • Separation of plant pigments • Separation of plant pigments
  • 4. Chromatography Techniques ChromatographyPartition Chromatography Adsorption Chromatography• components distribute between 2 immisible liquid phase • components adsorp on solid stationary phase• relative solubility in 2 phase• bonds strongly to mobile phase – move faster Stationary Liquid phase Mobile Y Mobile Y Y Stationary phase liquid phase X O- Stationary phase liquid phase X has a layer of liquid Y • solid containing containing Y O- • AI2O3 X and Y X X and Y X Y O- • SiO2 Y X X O- O- Mobile Mobile Y Y O- Y adsorb strongly XX Y Stationary phase Iiquid phase XX Y has a layer of liquid liquid phase Y O- containing X Y containing X XX Y X X Y O- X X XX X X Separation of X and Y X Y YYY Y X X Y YPartition –distribution of solute X and Y between 2 liquid phase Adsorption– solute X and Y adsorb temporary on the solid• X more soluble in mobile phase (move with mobile liquid phase) • Y adsorb strongly on solid phase, eluted slower• Y less soluble in mobile phase (stay on stationary liquid phase) • X in liquid mobile phase, eluted faster
  • 5. Chromatography Techniques Paper Chromatography Thin Layer Chromatography Partition chromatography Adsorption chromatography • Distribution solute bet both liquid phase • Solute adsorb on stationary solid phase • Depends on relative solubility O- Stationary phase solid Stationary phase - Cellulose paper X O- • AI2O3 or SiO2 • absorb water on its surface Y Y Y O- X Y O- Y Y X O- X Mobile Liquid phase with solute X and YMobile Liquid phase with solute X and Y Aqueous liquid phase on surface of stationary phase (paper) Stationary phase - solid silica SiO2 or alumina AI2O3 Mobile phase – liquid solvent move by capillary action Mobile liquid phase - solvent Separation depends on how strongly solute adsorb on solid Solvent move by capillary action Adsorption of solute on stationary solid phase
  • 6. Chromatography Techniques Column Chromatography Thin Layer Chromatography Adsorption chromatography Adsorption chromatography• Solute adsorb on stationary solid phase • Solute adsorb on stationary solid phase Mobile Liquid phase (solvent) with solute X and Y X Stationary phase solid O- Stationary phase solid X X AI2O3 or SiO2 Y Y O- • AI2O3 or SiO2 X Y Y X O- X O- X O- Y Y Y Y Y Mobile Liquid phase with solute X and Y Stationary phase - solid silica SiO2 or alumina AI2O3 Y Y Y Mobile phase – liquid solvent move by capillary action Mobile phase – liquid solvent added from top (Eluant) Adsorption of solute on stationary solid phase Solute in mobile phase adsorb on solid X - Adsorb strongly – travel slowly X - Strongly adsorb – travel slowly, eluted last Y – Adsorb weakly – travel fast with solvent Y - Weakly adsorb – travel fast with solvent, eluted first
  • 7. Chromatography Techniques Components separated by Paper/TLC can be identified using Rf value • Rf = Retention factor for a given eluent. • Measured the distance from original spot to the centre of a particular component to the solvent front Rf value for green spot Rf value for blue spot = 3cm/12cm = 6cm/12cm = 0.25 = 0.5 Uses of Paper/TLC/Column chromatography •Separate dyes in food colourings/plant pigments • Separation/Preparative/Collection of pigments • Detection of amino acids in mixture For colour formation of amino acids • Spray with ninhydrin for colour formation • Shine UV light for fluorescent compounds to formhttp://home.earthlink.net/~dayvdanls/photolab/photolab7.htm
  • 8. High Performance Liquid ChromatographyGas Liquid Chromatography
  • 9. Chromatography Techniques High Performance Liquid Chromatography Partition Chromatography • Components distribute between 2 immisible liquid phase • Mobile phase (liquid) with stationary phase (liquid) • Uses high pressure liquid to force through a column Normal HPLC Reverse phase HPLCMobile phase O- Stationary phase Mobile phase O- Stationary phaseNon-polar liquid Y •solid. AI2O3 and SiO2 Polar liquid Ywith solute X and Y O- •layer of polar liquid with solute X and Y X X O- •solid. AI2O3 and SiO2 Y •layer of non polar liquid Y X O- hydrocarbon Y Y O- X Y O- X O- X O- Y X O- O- O- Non-polar solute X in Y O- Polar Y solute distribute X X O- non polar stationary phase YY in polar stationary phase Y O- (elute later) XX O- (elute last) XNon-polar solute X XX O- Polar Y solute in polar Y O-in non polar mobile X phase (elute first) X O- mobile phase (elute first) YY Y O- Y XXX YYY YYY XXX Normal HPLC Reverse phase HPLC • non polar solvent (mobile phase) and polar stationary phase • polar solvent (mobile phase) and non polar stationary phase • polar solute – stay in polar stationary phase- elute later • non polar solute – stay in non polar stationary phase- elute later • non polar solute – stay in non polar mobile phase – elute first • polar solute – stay in polar mobile phase – elute first
  • 10. Chromatography Techniques Gas Liquid Chromatography Partition Chromatography• Components distribute between TWO immisible phase• Mobile phase (gas) with stationary phase (liquid)• Mobile phase – inert gas, helium, nitrogen• Incresing Temp – causes elution of solutes from stationary phase GLC Mobile gas phase O- Stationary phase with solute X and Y Y •solid. AI2O3 and SiO2 O- •layer of non volatile liquid Y Y X O- hydrocarbon Y X O- X O- O- Y O- Less volatile solute Y distribute YY in liquid stationary phase Y O- (elute later/longer retention time) Solute X (volatile) XX X O- in mobile gas phase (elute first) X O- XXX YYY GLC• Gas (mobile phase) and non volatile liquid on stationary phase• Less volatile solute Y – stay in stationary phase - elute later• Volatile solute X – stay in mobile phase – elute first
  • 11. Chromatography Techniques GLC vs HPLC Partition Chromatography • Components distribute between TWO immisible phase • Mobile phase (gas) with stationary phase (liquid) • Mobile phase – inert gas, helium, nitrogen • Incresing Temp – causes elution of solutes from stationary phase GLC HPLC Stationary phaseMobile gas phase O- Stationary phase Mobile phase O- •solid. AI2O3 and SiO2with solute X and Y Y •solid. AI2O3 and SiO2 Non-polar liquid Y O- •layer of non volatile liquid with solute X and Y X X O- •layer of polar liquid Y hydrocarbon Y X O- hydrocarbon Y Y O- X Y O- X O- X O- Y X O- O- O- Y O- Less volatile solute Y distribute Y Y O- Polar Y solute distribute YY in liquid stationary phase in polar stationary phase Y O- (elute later/longer retention time) YY O- (elute later)Solute X (volatile) XX X O- Non-polar solute X X XX Y O- in mobile gas phase in mobile phase(elute first) X O- (elute first) XX O- XXX YYY XXX YYY GLC Normal HPLC • Gas (mobile phase) and non volatile liquid on stationary phase • non polar solvent (mobile phase) and polar stationary phase • Less volatile solute Y – stay in stationary phase- elute later • polar solute – stay in polar stationary phase- elute later • Volatile solute X – stay in mobile phase – elute first • non polar solute – stay in non polar mobile phase – elute first
  • 12. Chromatography Techniques How GLC works How HPLC works vs Polar column Injection and vaporization of volatile sample in Column chamber Injection of non volatile sample Sample move with mobile gas phase Sample move with mobile liquid phase Elution of sample by temp change Elution of sample by using diff solvent polarity Detection by flame ionization detector FID (more sensitive) Detection by UV (less sensitive) Chromatogram - retention time for diff samples Chromatogram - retention time for diff samples More volatile – shorter retention time Non polar solute – Polar solute – short retention time greater retention time Less volatile – greater retention time Both can be use for identification, separation and quantification Quantification - Area under curve – represent amount solutehttp://lab-training.com/landing/free-hplc-training-programme-6/
  • 13. Chromatography Techniques Partition Chromatography Adsorption Chromatography Partition Chromatography Adsorption Chromatography • Components distribute bet TWO immisible liquid phase • Component adsorb on solid stationary phase • Depends on relative solubility bet TWO phase • Depends on polarity of stationary phase, mobile phase and solutes • Solutes bonds to stationary phase or mobile phase • Stationary phase is polar – polar solutes adsorb strongly • Stationary phase is non polar – non polar solutes adsorb strongly • Mobile phase is polar – polar solutes stay in mobile phase • Mobile phase is non polar – non polar solutes stay in mobile phasePaper Chromatography GLC HPLC Thin Layer Chromatography Column Chromatography Y adsorb strongly X adsorb strongly Mobile phase – liquid Mobile phase – gas Mobile phase – liquid Mobile phase – liquid Mobile phase – liquid Stationary phase - liquid Stationary phase - liquid Stationary phase - liquid Stationary phase - solid Stationary phase - solid Uses of GLC Uses of HPLC • Analysis of steroids, vegetable oils, alcohol in blood • Analysis of sugar, additives, pesticides, herbicides • Volatile/thermally stable compounds • Non volatile/thermally unstable/sensitive to heat • Small amt volatile liquid, not sensitive to heat vs • Compounds decomposes upon heating ( Vit C, E , antioxidants) • More sensitive – Flame Ionization Detector • Less sensitive – UV detector • Use for identification/separation/quantification • Use for identification/separation/quantification
  • 14. Chromatography Techniques Technique Stationary Phase Mobile Phase Application Paper Liquid on paper Liquid/Solvent - Detection of amino acids in a mixture Chromatography - Different dyes in food colouring - Separation of plant pigments Thin Layer Solid oxide coat Liquid/Solvent - Detection of amino acids in a mixture Chromatography - Different dyes in food colouring - Separation of plant pigments Column Solid oxide coat Liquid/Solvent - Preparative/collection of pigments/dyes. Chromatography Gas Liquid Liquid on solid coat Gas - Analysis of steroids, vegetable oils, alcohol in blood Chromatography - Volatile/thermally stable compounds - Small amt volatile liquid, not sensitive to heat - More sensitive – Flame Ionization Detector - Use for identification/separation/quantification High Performance Liquid on solid oxide Liquid/Solvent - Analysis of sugar, additives, pesticides, herbicidesLiquid Chromatography coat - Non volatile/thermally unstable/sensitive to heat - Compounds decomposes upon heating ( Vit C, antioxidants, vitamin E ) - Less sensitive – UV detector - Use for identification/separation/quantification
  • 15. Click here to view TLC and Column ChromatographySeparation using TLC TLC techniques step by stepColumn Chromatography Column chromatography separation
  • 16. Click here to view HPLC and Gas Liquid Chromatography HPLC works Gas Liquid Chromatography works Click here to view Click here to viewClick here on column chromatography Click here to view amino acid separation