Remember that the solute should dissolve only when the solvent is heated. Therefore, the solvent is heated to its boiling point remember to use boiling stones! If too much solvent is added, the solution will not be saturated upon cooling and no crystals will form. Dissolving the solute generally involves adding a small volume of hot solvent, swirling the flask or stirring the solution , and watching to see if the solute dissolves. Decolorize the solution. If the solute is supposed to be white in its pure solid state most organic solids are and the solution is colored after dissolving all the solute, it will be necessary to add decolorizing carbon to the solution.
This will cause the colored molecules to adsorb onto the surface of the decolorizing carbon, thereby ridding the solution of these impurities. Should these impurities remain in solution, they may become trapped in the developing crystal during cooling. Review the material about decolorizing carbon.
Filter any solids from the hot solution. If decolorizing carbon was used as in step 3 or undissolved impurities remain in the hot solution, it is necessary to gravity filter the solution while it is still hot.
Review the information about hot gravity filtration and decolorizing carbon. Under no circumstances should the hot solution be vacuum filtered with a Buchner funnel. This leads to premature crystal development as the solution passes through the vacuum filter.
The vacuum reduces the pressure, but also the temperature. Impurities will be trapped in the crystal lattice and steps 1 through 3 will need repeated! Crystallize the solute. This involves allowing the hot solution with the solute dissolved to return to room temperature slowly. The slower the cooling process, the less chance of trapping impurities in the developing crystal lattice.
For some compounds, you might need to wait until the solution boils before your compound completely dissolves. Q: After the solution has cooled down to room temperature, how long should I let it cool in the ice bath? You should let the solution cool in the ice bath for a minimum of 15 minutes to ensure that the bulk of the sample has recrystallized.
You need to cool the solution first to room temperature before placing it in the ice-water bath. Besides reducing the risk of breaking your flask and loosing your product in the ice-water, you will get better and purer crystals if you let the solution cool slowly. Q: When we are collecting our crystals using vacuum filtration, what solvent do we use to wash our crystals?
When vacuum filtering, wash your crystals with the solvent you used to recrystallize your compound. However, use ice-cold solvent to ensure that you do not dissolve any of your crystals. Q: I have a really lousy suction from that water aspirator. Make sure it is there and everything is sitting tight - The water trap is not closed to the atmosphere. Q: When using the two-solvent recrystallization method, why is it necessary to keep both solvents hot when adding?
For the single-solvent and the two-solvents recrystallization method it is essential that you prepare a hot, saturated solution. To do this, all solvents must be hot before you add them. Heating the solvents decreases the kinetic energy necessary to dissolve the compound. This also means that less solvent is needed to dissolve the compound, which makes the solution more saturated. The more saturated the solution, the easier the crystals will form once the solution cools down.
Q: Can we add the second solvent first? You must add a minimum amount of first hot solvent to dissolve your compound. Then you decrease the solubility of your solid by adding the second hot solvent to the first cloud. Reheat the solution to clear it again. This allows for a saturated solution and for crystallization to occur upon cooling. Q: If I can choose between the 1-solvent or 2-solvent method, which one should I choose?
The preferred method of recrystallization uses one solvent. When a suitable single solvent can not be found, the two-solvent method of recrystallization is used. Back to Techniques Page. FAQ: How many boiling stones should I use? FAQ: How long does it take for the crystals to grow?
FAQ: If we add too much solvent do we just boil it off? FAQ: Can I put my hot solution directly into the ice bath? FAQ: What should I put on the label when handing in my sample? Return to Top Recrystallization Theory Recrystallization is a purification technique. Insoluble impurities can be filtered by hot gravity filtration.
Return to Top Single Solvent Recrystallization Single solvent recrystallization is the most basic and commonly used recrystallization method. Process: 1. Use solubility tests to determine a suitable recrystallization solvent. Allow the crystals to dry. Return to Top Two Solvent Recrystallization Two solvent recrystallization is an alternative and very useful recrystallization method to single solvent recrystallization. Return to Top Q: How many boiling stones should I use?
Return to Top Q: When I tested the recrystallization solvent in a test tube it worked, but now my sample won't dissolve!
Return to Top Q: How long does it take for the crystals to grow? Return to Top Q: So - once the solvent has reached its boiling point and my crude solid is NOT dissolved, do I add more solvent or do I let it boil longer? Return to Top Q: If we add too much solvent, do we just boil it off? Return to Top Q: My sample has dissolved, but my solvent is just hot.
Return to Top Q: After the solution has cooled down to room temperature, how long should I let it cool in the ice bath? Return to Top Q: Can I put my hot solution directly into the ice bath? Return to Top Q: When we are collecting our crystals using vacuum filtration, what solvent do we use to wash our crystals?
Return to Top Q: I have a really lousy suction from that water aspirator. Some possible reasons for little suction are: - The black filter vac adapter between the filtering flask and the Buchner or Hirsch funnel is missing.
Return to Top Q: What should I put on the label when handing in my sample? On your sample label, you should write: - your name, - experiment number e. Return to Top Q: When using the two-solvent recrystallization method, why is it necessary to keep both solvents hot when adding? Return to Top Q: Can we add the second solvent first? However, if an impurity is less soluble, it will crystalize first, and can then be filtered out of the heated solution, prior to recrystallization of the solute.
If no single solvent has the necessary properties, a mixture of solvents can be used. For a solvent pair, the first solvent should readily dissolve the solid. The second solvent must have a lower solubility for the solute and be miscible with the first solvent. Common solvent pairs include ethyl acetate and hexane, toluene and hexane, methanol and dichloromethane, and water and ethanol. Now that you understand the principles of recrystallization, let's go through a procedure for purification of an organic compound by recrystallization.
Add 0. If the compound dissolves completely, the solubility in the cold solvent is too high to be used for recrystallization. Otherwise, heat the mixture in the test tube to boiling. If the compound does not dissolve completely in the boiling solvent, heat another portion of solvent to boiling. Add the boiling solvent dropwise to the test tube until the solid dissolves completely or until the test tube contains 3 mL of solvent.
If the solid still does not dissolve, then its solubility in this solvent is too low. Confirm that impurities are either insoluble in the hot solvent so they can be filtered out after dissolution or soluble in the cold solvent so they remain in solution after recrystallization is complete. If a solvent meets all criteria, it is suitable for recrystallization. To start recrystallization, heat the solvent to boiling on a hot plate in an Erlenmeyer flask with a stir bar. Place the compound to be recrystallized in another Erlenmeyer flask at room temperature.
Next, add a small portion of hot solvent to the compound. Swirl the mixture in the flask and then place it on the hot plate as well. Repeat this process until the sample has completely dissolved or until addition of solvent causes no further dissolution. Filter the solution to remove insoluble impurities. If crystals form during filtration, dissolve them with drops of hot solvent.
Cool the solution on the benchtop. Cover the flask to prevent solvent loss to evaporation and to keep particulates out of the solution. Leave the flask undisturbed until it has cooled to room temperature. Agitation during cooling may cause rapid crystallization, yielding less pure crystals. If no crystal formation is evident upon cooling, induce crystallization by gently scratching the inside walls of the flask with a glass rod or adding a small seed crystal of the compound being recrystallized.
If crystal formation cannot be induced, reheat the solution to boil off some of the solvent, and then cool the solvent to room temperature once more. Once crystals have formed, prepare an ice bath.
Keeping the solution covered, cool the solution in the ice bath until crystallization appears to be complete. Clamp a filtration flask to a ring stand and connect the flask to a vacuum line. Pour the mixture of solution and crystals into the funnel and begin vacuum filtration. Rinse any crystals remaining in the flask into the funnel with cold solvent. Wash the crystals on the funnel with cold solvent to remove soluble impurities.
Continue drawing air through the funnel to dry the crystals and then turn off the vacuum pump. If necessary, the crystals may be allowed to stand at room temperature to air dry or placed in a desiccator before storing the crystallized solid.
The yellow impurities present in the crude compound have been removed, yielding an off-white solid. Based on the identity of the compound and the impurities, the purity of the crystals can be verified by NMR spectroscopy, melting point measurements, or visual inspection. X-ray crystallography is a powerful characterization technique that identifies the three-dimensional atomic structure of a molecule.
This requires a pure single crystal, which is obtained by recrystallization. Some classes of molecules such as proteins are difficult to crystallize, but their structures are extremely important for understanding their chemical functions. With careful selection of recrystallization conditions, even these classes of molecules can be analyzed by X-ray crystallography. To learn more about this process, see this collection's video on growing crystals for crystallography.
Impure reactants can cause unwanted side reactions. Purifying reactants by recrystallization improves product purity and yield. Once a solid product has been isolated and washed, reaction yield can also be increased by removing volatiles from the filtrate and recrystallizing the product from the resulting solid. Antifreeze proteins, or AFPs, are expressed in many organisms that live in icy environments.
AFPs hinder internal ice growth by binding to ice planes, inhibiting recrystallization into larger ice crystals. Different AFPs bind to different types of ice crystal planes. Investigating AFP binding mechanisms involves adsorbing them onto single ice crystals. Proper growth of a single ice crystal is essential for clear and informative results. These proteins have applications from the engineering of cold-resistant crops to cryosurgery.
You've just watched JoVE's introduction to purifying compounds by recrystallization. You should now be familiar with the principles of the technique, a purification procedure, and some applications of recrystallization in chemistry.
An example of the results of recrystallization is shown in Figure 2. The yellow impurities present in the crude compound have been removed, and the pure product is left as an off-white solid. The purity of the recrystallized compound can now be verified by nuclear magnetic resonance NMR spectroscopy or, if it is a compound with a published melting point, by how similar its melting point is to the literature melting point.
If necessary, multiple recrystallizations can be performed until the purity is acceptably high. Figure 2. Recrystallization is a method of purifying a compound by removing any impurities that might be mixed with it. It works best when the compound is very soluble in a hot solvent, but very insoluble in the cold version of the same solvent. The compound must be a solid at room temperature. Recrystallization is often used as a final clean-up step, after other methods such as extraction or column chromatography that are effective at removing larger amounts of impurities, but that do not raise the purity of the final compound to a sufficiently high level.
Recrystallization is the only technique that can produce absolutely pure, perfect single crystals of a compound. These crystals can be used for X-ray analysis, which is the ultimate authority in determining the structure and three-dimensional shape of a molecule. In these cases, the recrystallization is allowed to proceed very slowly, over the course of weeks to months, to allow the crystal lattice to form without the inclusion of any impurities.
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