What is Chromatography: Definition and 49 Discussions

Chromatography is a laboratory technique for the separation of a mixture.
The mixture is dissolved in a fluid (gas, solvent, water, ...) called the mobile phase, which carries it through a system (a column, a capillary tube, a plate, or a sheet) on which is fixed a material called the stationary phase. The different constituents of the mixture have different affinities for the stationary phase. The different molecules stay longer or shorter on the stationary phase, depending on their interactions with its surface sites. So, they travel at different apparent velocities in the mobile fluid, causing them to separate. The separation is based on the differential partitioning between the mobile and the stationary phases. Subtle differences in a compound's partition coefficient result in differential retention on the stationary phase and thus affect the separation.Chromatography may be preparative or analytical. The purpose of preparative chromatography is to separate the components of a mixture for later use, and is thus a form of purification. Analytical chromatography is done normally with smaller amounts of material and is for establishing the presence or measuring the relative proportions of analytes in a mixture. The two are not mutually exclusive.

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  1. I

    Chiral Column Chromatography

    "Resolution of enantiomers can be accomplished using column chromatography. When enantiomers are passed through a traditional column, they travel at the same rate because their properties are identical. However, if a chiral adsorbent is used, the enantiomers interact with the adsorbent...
  2. S

    Question about antibody affinity chromatography

    I am learning about protein purification in my Biochemistry class, and I have a question about protein elution in antibody affinity chromatography. My understanding of the mechanism for the technique is that proteins that do not bind to the antibody will be separated first as it runs down the...
  3. B

    How does one prepare standards for headspace gas chromatography?

    I need to measure trichloroethylene (TCE) in water samples using headspace gas chromatography. Obviously, I will need a set of standard solutions to make a standard curve for concentration vs area. How exactly do you prepare the standard solutions? What I don't understand is that, say I prepare...
  4. M

    MATLAB Solving Chromatography PDE with MOL and ode15s

    Hello all I am using the method of lines to solve the following PDE: ## \frac {\partial C} {\partial t} + F\frac {\partial q} {\partial t} + u \frac {dC} {dz} = D_{ax} \frac{\partial^2 C} {\partial z^2} ## ## \frac {\partial q} {\partial t} = k (q^{*}-q) ## With these initial conditions: ##...
  5. T

    Which chromatography test fits best?

    Homework Statement A purified protein fraction containing buffer with salt can be desalted by one of the following techniques: (1) Gel filtration chromatography (3) Paper chromatography (2) Affinity chromatography (4) Thin layer chromatography2. The attempt at a solution Well, the analyte...
  6. T

    Which chromatography technique is most suitable?

    Homework Statement [/B] Two proteins of similar molecular weight (<900 Da), same net charge in the solution but with different amino acid composition is to be separated. Which one should be used? a. Cation exchange b. Anion exchange c. gel filtration d. Reverse phase 2. My attempt: Since the...
  7. R

    Temperature(Pre) -- Gas Chromatography Question

    Does anyone here have any experience with GC? I am developing a method for analyses and I understand all the settings but there is one related to the TCD called Temperature(pre). Does anyone have any ideas? Thanks for any suggestions!
  8. T

    Chromatography retention time question

    Homework Statement For this chromatography graph there are 2 peaks: One of them is propanoic acid and another one is pentanoic acid. The more polar compound will have a longer retention time. Which peak belongs to which compound. Homework EquationsThe Attempt at a Solution The answers says...
  9. E

    Latent Heat and Bond Energy

    The kinetic theory says that temperature is the measure of the average kinetic energy. That would mean then : The temperature rises with the rise in kinetic energy. We know that when matter changes state 'latent heat' helps in 'breaking the bonds' at the transition phase and that this 'extra...
  10. E

    Chromatography: The difference in kinetic energy

    In chromatography, the upper portion of the moving water solvent has more kinetic energy, than the lower portion of the moving water solvent because the lower portion of water molecules have to face more obstacles in the way because of the absorbed water molecules. Is it true?
  11. MarcozXD

    Should carbon boiling stones be used in O-chem reactions?

    I performed column cromatography on an oil, and used carbon boiling chips but got some weird IR. I used hexanes and acetone as solvents, I am trying to figure out some of the spikes, soo I am thinking that maybe the boiling stone being carbon could acted as activated carbon and absorbed some...
  12. C

    Gas chromatography question -- instrument not turning back on....

    My gas chromatography instrument ran out of air while cooling down and now it won't turn on, nor the computer it is hooked up to? Can this be fixed?
  13. D

    Separation Through chromatography

    Dear all, We all know that chromatography is a separation technique. In paper or TLC,the mixture is differentiated according to their relative affinities with the solvent on the paper or glass plate. How do we really separate the mixture. Kindly, help me how to extract the components...
  14. H

    Gel filtration chromatography

    Hi, I am studying for my biochemistry lab exam. While going over one of the questions... I came across one that I can't answer...help will be appreciated. Question under analysis is related to a previous question. (Previous question) A gel filtration (size exlcusion) is set up and the...
  15. D

    Protocol and solvent help needed for gel-permeation chromatography

    So I have been asked to separate a polymer (polyurethane) into high molecular weight and low molecular weight components. I have access to Sephadex LH-20, a simple glass column, acetone (a List II chemical), ethyl acetate, and hexane. I may be able to get access to other solvents, as long as it...
  16. P

    Sodium borohydride reductions and chromatography

    Homework Statement Hi, I did a lab where benzophenone was dissolved in ethanol and reacted with sodium borohydride (NaBH4) and chromatography was done on the reaction at different points in time. There was no HCl work up of this though. When I developed the TLC (silica) plate, it looked like...
  17. C

    Chromatography - What is a scouting gradient?

    I'm reading about gas chromatography at the moment and the notes I'm reading mentioned a "generic scouting gradient" but didn't explain what it is. I've been googling it and found a few HPLC tutorials (in GC its temperature gradient whereas in the HPLC tutorials they're talking about mobile...
  18. J

    Why during chromatography must the paper NOT touch the sides

    Homework Statement why during chromatography must the paper NOT touch the sides of the test tubes in which the experiment is being carried out? Homework Equations The Attempt at a Solution I am not sure but could our results get smudged or distroted? if so I still can't work out how? could it...
  19. E

    Column Chromatography ketone/silica

    I recently ran a baylis-hillman reaction of 2nitrobenzaldehyde and acrylonitrile (DABCO catalyst) and after my reaction had gone to completion (determined by tlc) i extracted with chloroform/washed with brine, dried, filtered, rotovaped etc etc I ran a crude NMR and determined I did have my...
  20. C

    Chromatography - How does flow rate affect peak width?

    Today in a lecture on chromatography the lecturer gave an explanation which seems pure ******** to me. The example she used was a handful of solutes, each taking a different path through the stationary phase, obviously some paths will be longer than others and thus each solute will have a...
  21. Saladsamurai

    Gas Chromatography: Stray Peak and Beginner's Questions

    Hey guys! It seems that there are a few users here, so I will post the same question I posted over at chromforum.org: -------------------------------------------------- Hello All! :smile: I am quite new here and to GC in general, so I hope you can bear with me. I will try to keep this...
  22. Saladsamurai

    Gas Chromatography: Varian CP 3800 GC

    Hello people! I am wondering if anyone uses this machine? I am a mechanical engineering student and I have one in my laboratory. I have quite a bit of documentation on the machine, however, a lot of it is like a foreign language to me. Before I start asking any questions, I wanted to...
  23. H

    Ion exchange chromatography experiment involving transitional metals

    I did an experiment in which I used an anion exchange resin to separate nickel from cobalt but I don't understand the theory behind it. What I know is that the ion exchange resin uses quaternary ammonium groups as its cations but that's about it. First we packed a column with the resin then...
  24. V

    Cleaning chromatography columns and hydrodynamics

    If you had to prove that your method for cleaning chromatography columns works and you had different sizes and shapes of columns, which one would you pick as a worst case to focus on? You have tall, skinny columns and short, fat columns, as well as different sizes. These are cylindrical glass...
  25. U

    How would you use chromatography to determine whether an unknown pure liquid....

    Homework Statement How would you use chromatography to determine whether an unknown pure liquid is 1-pentene, cis-2-pentene, or trans-2-pentene? Homework Equations The Attempt at a Solution I looked up the boiling points of the three and see that 1-pentene is lower, but the cis...
  26. S

    Chromatography Scale-up

    Homework Statement The problem requires the scale up of a linear gradient ion exchange chromatography column by a factor of 150 (throughput). The lab scale column is 1 cm (ID) x 20 cm (bed height) with 20 micron particle size and 30 cm/hr superficial velocity. Resolution must be kept constant...
  27. S

    Gas Chromatography - Organic Chemistry

    Why do ten molecules of toluene that enter a GC column at exactly the same time not reach the detector at the same time? Why are London Dispersion forces of the sample molecules strongly influencing GC measurements? And why are they negligible in TLC experiments on silica gel? I have no...
  28. P

    Chromatography as a separation techinique

    Homework Statement In detail, explain the theory behind the use of chromatography as a a.)separation technique. b.) identification technique. Homework Equations The Attempt at a Solution I know that chromatography can be used as a separation technique and also indetification...
  29. L

    Why are Pigment Spots Different Sizes?

    1. Why are some pigment spots larger (area and volume) than others? I understand that pigments move up the porous paper by capillary action, from research. But why are some pigment colours (areas) larger? THanks
  30. O

    Thin Layer Chromatography

    Homework Statement How does heating affect the spots on the TLC? Homework Equations None The Attempt at a Solution I have no clue, unless heating evaporates the solution and therefore the spots disappear? or Perhaps the elutent evaporated due to heat rather then the solution?
  31. L

    Building Gas Chromatograph with Capillary Column: Tips & Guidelines

    I have a project that I want to do; really it is an experiment that would prove useful to another project of mine. I want to build a gas chromatograph. The specs. Capillary column Helium carrier Thermal Conductivity Detector The analyte is a mixture of CH4,N2 (for now) I already...
  32. H

    Chromatography: Please help

    Chromatography: Please help! What is the visualizing reagent used for Cu(II), Cd (II), Ni(II) and Co(II) ions when performing paper chromatography? Are there any special visualizing reagents for transition metals?
  33. F

    Chromatography - Net Charge

    Homework Statement What procedures would you use to obtain gram quantities of each of the following compounds, free of each of the other compounds? Glycine - monoamino, monocarboxylic amino acid Glucose - non-ionic monosaccharide Starch - non-ionic large polysaccharide Glucose -...
  34. P

    Ion Chromatography Analysis: Determining Anion Identity

    In our analytical lab we were given an unknown solution containing up to 4 anions - fluoride, chloride, nitrate, sulphate and phosphate. It was diluted, run through the chromatograph and a chromatogram was printed for 3 trials. Then we printed chromatograms for two different standard solutions...
  35. P

    Optimized Developing Mn+2 - Best Reagent Explained

    [SOLVED] paper chromatography Homework Statement Which reagent is the best to develop Mn+2? Briefly explain. Homework Equations Mn^+2 + 2OH- -> Mn(OH)_2 (s)(pale pink) Mn^+2 + C_2O_4^-2 -> MnC2O4 (s)(pale pink) 2Mn^+2 + [Fe(CN)_6]^-4 -> Mn_2Fe(CN)_6 (s pale green) 2Mn^+2 +...
  36. G

    Thin-layer chromatography

    Is it possible to separate cis/trans isomers of diazo compound using Thin-layer chromatography?
  37. chem_tr

    Thin Lead Chromatography help

    Hello everyone, I was poor on TLC before, and am poor now :smile: I want to ask a question to you guys. Please look at the attached zip file with a tif image. My question is very simple: Is it possible to fully isolate the impurities (yellow spot and light olive-mustard-colored...
  38. M

    Separtion by paper chromatography

    Hi does anyone know why it is imporatnt not to permit the solvent front to reach the top of the chromatogram?
  39. K

    A Question about Chromatography

    I calculated the total exchange capacity of a column per gram of dry weight of resin to be 2.74 meq/g. This value comes from experimental values. However, the manufacturer's specifications for the number of exchange sites is 3.4 meq/g. 1. What's meq? :blushing: I know this means...
  40. S

    Good links about Gas Chromatography

    i hope for good links about GC. thanks!
  41. S

    Gas Chromatography: Polar Compounds & Retention Time

    it says when i choose the liquid stationary phase, then i have to use the rule; "like dissolves like". i wonder if the compound i am interested in is polar, this means that the stationary phase i choose must also be polar, right? how would you predict the retention time of my compound? would it...
  42. M

    Gas Chromatography: Example of Molar Proportion of A & B

    what does it mean when it says; "the areas under the two peaks are directly proportional to the molar amounts of A and B in the mixture? directly proportional? can anyone give an example of it? many thanks!
  43. M

    Rf values and Chromatography

    Hi for biology we had to do a lab which we separated the plant pigments by chromatography of spinach. We used a solvent of 90% petroleum ether:10% acetone. After we did the experiment we ran out of time in class so the teacher went and recorded everyones results on there paper. The problem with...
  44. I

    Chromatography using acetone

    chromatography...using acetone! the protocol: Vg = volume not accessable to solvent (volume of the gel matrix) Vg cannot be measured directly. However, it should apparent that: Vg = Vt - volume accessable to solvent We can measure the volume accessable to the solvent, using a small...
  45. M

    Chromatography Problem: Setting a "Clen After Gradient

    hi guys! as you know i am doing the chromatography stuff and am now having the problem with understanding the protocol. it says; "set a clen after gradient value to 100% buffer B over 2 CV”? :bugeye: does any of you know what it tries to tell me? thanks!
  46. S

    How Do I Calculate Column Cross-Sectional Area for Chromatography Scaling Up?

    hi guys! i am doing the chromatography. in scaling up i have to increase my volumetric flow rate but maintain my linear flow rate. my problem is i don't know how to calculate the column cross sectional area (cm2). i hope you guys who work with this area can tell me how i can calculate it...
  47. M

    About ionic chromatography

    is bed length the heigh of the column or is that the heigh of the matrix/gel? i don't know where to put this question, but since it is about proteins then i guess here is the place :approve: thanks!
  48. D

    Understanding the Roles of Carotene and Xanthophyll in Chromatography

    I did an experiment in class on Chromatography and extracting pigment material from a leaf and such. We took a piece of some special paper put a base line in pencil at the bottom and covered it with leaf extract. Then putting a pin through the top set it in a beaker of acetone and the...
  49. S

    High pressure chromatography?

    What are the applications and advantages/disdvantages of using gas and high pressure chromatography? Why does the pharmaceutial industry use chromatography? Any info would be much appreciated Thanx