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Organic Chemistry
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JoVE Science Education Organic Chemistry
Purifying Compounds by Recrystallization
  • 00:00Overview
  • 00:58Principles of Recrystallization
  • 03:41Selecting a Solvent
  • 04:50Recrystallization
  • 07:40Applications
  • 09:31Summary

再結晶により物質を浄化

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Overview

ソース: 博士ジミー フランコ – メリマック大学講座

再結晶は、固体の化合物を浄化するために使用される手法です。1固体はより冷たい液体でより熱い液体に溶解する傾向があります。再結晶化、ソリューションが飽和し、液体は冷却する許可まで、不純な固体化合物は熱い液体で溶解します。2化合物は、比較的純粋な結晶を形成する必要があります。理想的には、存在する不純物は、溶液中に残ってし、(図 1)、成長の結晶に組み込まれません。結晶は、濾過によりソリューションから除去することができます。回復可能なすべての化合物は-いくつかのソリューションに残ります、失われます。

再結晶は一般的に予想外の分離の手法としてむしろ、それは化合物から少量の不純物を除去する精製技術です。ただし、2 つの化合物の溶解度特性が十分に異なる場合、再結晶化は、ほぼ均等に存在している場合でも、それらを分離する使用できます。再結晶は、ほとんど不純物が抽出または列クロマトグラフィーなどの別の方法で既に削除された場合に最適です。

Figure 1
図 1再結晶の一般的なスキーム

Principles

成功した再結晶溶媒の適切な選択により異なります。化合物は、寒い時に熱い溶媒と同じ溶媒に不溶可溶必要があります。再結晶、目的間を考慮する 3 w/v の分割線水溶性と不溶性: 水溶性化合物の 3 g は、100 mL の溶媒に溶解する場合があります。再結晶溶媒、熱い溶解度と溶解性冷間違いが大きい、回復可能なより多くの製品を選択します。

冷却の率は、サイズと結晶の品質を決定します: 小さな結晶を支持する急速な冷却と大規模な一般的に純粋な結晶の成長を支持する遅い冷却します。再結晶化の率は通常、物質の融点以下約 50 ° C で最大最大結晶形成に融点以下約 100 ° C で発生します。

用語「結晶化」と「再結晶」が同じ意味で使用される時が、彼らは技術的に別のプロセスを参照してください。化学反応によって新たに不溶解性製品の形成は、結晶化この製品を多くのトラップ不純物を含む非晶質固体として反応液から沈殿させます。化学反応を伴わない再結晶粗生成物は単にソリューションに溶解し、再形成する結晶を許可する条件を変更し。再結晶より純粋な最終的な製品を生成します。このため、通常結晶化による強固な製品を作り出す実験プロシージャには純粋な化合物を与える最終的な再結晶のステップが含まれます。

Procedure

溶剤ガスへの暴露を防ぐためにヒューム フードのすべての手順を実行します。 1. 溶媒を選択します。 三角フラスコに 50 mg のサンプル (N ブロモスクシンイミド) を配置します。 0.5 mL の沸騰の溶媒 (水) を追加します。試料が完全に溶解する場合は、良い結晶溶媒に冷たい溶媒で溶解度が大きすぎます。 サンプルは冷たい溶媒中に溶解しない場合、は、溶媒は沸騰するまでテスト チューブを熱します。 サンプルが完全にこの時点で解散はない場合、は、すべての固体が溶解するまでにもっと沸点溶媒を drop-wise、追加しています。熱い溶媒中で試料を溶かす 3 mL 以上かかる場合、この溶媒の溶解性が良い再結晶溶媒にする低すぎるおそらく。 溶媒の最初の選択肢は良い再結晶溶媒で、他を試してください。動作単一溶剤が見つからない場合、2 つの溶媒系をしようと。 適切な単一溶剤システムを見つけることができない場合、溶媒ペアが必要かもしれません。溶媒のペアを識別するには、いくつかの重要な考慮事項 1 があります) 最初の溶媒は容易に固体を解散すべきです。2) 2 つ目の溶媒では、1st溶剤と混和性であるが、溶質のはるかに低い溶解度する必要があります。 一般的なルールとして「好きなものディゾルブ好き」極性化合物の極性溶媒と非極性化合物に溶解する傾向があることを意味は、しばしばより水溶性の非極性化合物。 一般的な溶剤ペア (表 1) 沸騰の溶媒および室内温度溶媒間の適度な温度差があるので、溶剤が、少なくとも 40 の ° C の沸点であることを確認します。 あることを確認、溶媒沸点以下約 120 ° C 結晶から溶媒の最後の痕跡を削除する方が簡単です。 また溶媒の沸点は化合物形成不水溶性廃油としてではなく、固体の結晶としてので、化合物の融点よりも低いであることを確認します。 不純物がどちらかであることを確認 (彼らは化合物を分解したら、ホット フィルターできます) ので熱い溶媒に不溶または (し、全体のプロセスの中に溶解したまま) 冷たい溶媒に不溶。 2. 熱い溶媒中でサンプルの溶解 三角フラスコで再結晶する化合物を配置します。これは、傾斜の側面トラップ溶剤蒸気を助けるおよび蒸発の速度が遅いので、ビーカーよりもより良い選択です。 別のエルレンマイヤー フラスコに溶媒 (水) を配置し、沸騰のチップやスムーズに沸騰を維持する攪拌棒を追加します。ホット プレートの上に沸騰にそれを熱する。 化合物が完全に溶けるまで、各付加の後で旋回、小さな部分に化合物を含有する室温でフラスコに熱い溶媒を追加します。 溶解処理中にソリューションを保持でホットすべての回でも、ホット プレートの上に休憩します。必要 – 以上より熱い溶媒を追加しない試料を溶かすだけで十分。 固体の部分が解消しない場合は熱い溶媒を追加すると後でさえもそうだ非常に不溶解性の不純物の存在のため。この場合、溶媒を追加するを停止し、続行する前に熱いろ過を行います。 熱いろ過を実行するためにフルーティングを施された円錐形フィルター紙を折るし、ガラスの茎のない漏斗に入れます。 手順で蒸発を許可する高温の溶液に熱い溶媒の 10-20% 過剰を追加します。 紙を溶液を注ぐ。結晶開始フォーム プロセス中にいつでも場合、は、それらを解消する温かみのある溶媒の小さな部分を追加します。 3. 冷却ソリューション フラスコは行われません熱を離れて早すぎる、または卓上セット ペーパー タオルなど表面に溶存化合物を含有を設定します。 軽く冷やす蒸発を防ぐために、ソリューションに陥るからほこりを防ぐためにフラスコをカバーします。 それは部屋の温度に冷却するまでフラスコを妨げられていない残します。 結晶を形成している、一度結晶の最大量が得られることを確保するための氷浴にソリューションを配置します。ソリューションは、30 分間氷浴で妨げられていない 1 h に委ねられるべきまたは化合物表示が完全に解決から結晶化するまで。 それが中を掻くことによって誘起すること結晶形成を明らかにしない場合ガラス棒または同じ化合物の小さな種結晶を追加することによってフラスコの壁。 これはまだ動作するように失敗した場合、あまりにも多くの溶媒は使用でしょう。ソリューションを再加熱、ボイルオフガス、溶媒のいくつかを許可し、それを冷却します。 4. 分離と水晶の乾燥 冷たいフラスコ、ベンチトップに新たに形成された結晶を含むを設定します。 軽く蒸発を防ぐために、ソリューションに陥るからほこりを防ぐためにフラスコをカバーします。 Büchner またはハーシュ漏斗 (クランプ リングにフラスコは最初スタンド) を使用して、真空ろ過で結晶を分離します。 結晶を Büchner 漏斗、冷たい新鮮な溶媒 (同じ溶媒再結晶に使用) の少量のリンスの結晶に固着する可能性のあるすべての不純物を除去します。 結晶を乾燥するには、漏斗フィルターでそれらを残すし、数分間それらを介して空気を描画します。結晶は、数時間または数日の帽子をかぶらないで済むも乾することができます。効率的な方法には、真空乾燥または乾燥器で配置することがあります。 極性溶媒 以下の極性溶媒 酢酸エチル ヘキサン メタノール 塩化メチレン 水 エタノール トルエン ヘキサン 表 1。一般的な溶剤のペア。

Results

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
Figure 2. 2a) A crude compound (left), 2b) recrystallized product before filtration (middle), and 2c) the same compound after recrystallization (right).

Applications and Summary

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. Special glassware is needed to allow the solvent to evaporate as slowly as possible during this time, or to allow the solvent to very slowly mix with another solvent in which the compound is insoluble (called antisolvent addition).

The pharmaceutical industry also makes heavy use of recrystallization, since it is a means of purification more easily scaled up than column chromatography.3 The importance of recrystallization in industrial applications has triggered educators to emphasize recrystallization in the laboratory curriculum.4 For example, the drug Stavudine, which is used to reduce the effects of HIV, is typically isolated by crystallization.5 Often, molecules have multiple different crystal structures available, so it is necessary for research to evaluate and understand which crystal form is isolated under what conditions, such as cooling rate, solvent composition, and so forth. These different crystal forms might have different biological properties or be absorbed into the body at different rates.

A more common use of recrystallization is in making rock candy. Rock candy is made by dissolving sugar in hot water to the point of saturation. Wooden sticks are placed into the solution and the solution is allowed to cool and evaporate slowly. After several days, large crystals of sugar have grown all over the wooden sticks.

Disclosures

  1. Mayo, D. W.; Pike, R. M.; Forbes, D. C., Microscale organic laboratory : with multistep and multiscale syntheses. 5th ed.; J. Wiley & Sons: Hoboken, NJ, p xxi, 681 p (2011).
  2. Armarego, W. L. F.; Chai, C. L. L., Purification of laboratory chemicals. 5th ed.; Butterworth-Heinemann: Amsterdam ; Boston; p xv, 609 p (2003).
  3. Ray, P. C.; Tummanapalli, J. M. C.; Gorantla, S. R., Process for the large scale production of Stavudine. Google Patents: (2011).
  4. Hightower, T. R.; Heeren, J. D., Using a Simulated Industrial Setting for the Development of an Improved Solvent System for the Recrystallization of Benzoic Acid: A Student-Centered Project. Journal of Chemical Education 83 (11), 1663 (2006).
  5. Rohani, S.; Horne, S.; Murthy, K., Control of Product Quality in Batch Crystallization of Pharmaceuticals and Fine Chemicals. Part 1: Design of the Crystallization Process and the Effect of Solvent. Organic Process Research & Development 9 (6), 858-872 (2005).

References

  1. Mayo, D. W.; Pike, R. M.; Forbes, D. C., Microscale organic laboratory : with multistep and multiscale syntheses. 5th ed.; J. Wiley & Sons: Hoboken, NJ, p xxi, 681 p (2011).
  2. Armarego, W. L. F.; Chai, C. L. L., Purification of laboratory chemicals. 5th ed.; Butterworth-Heinemann: Amsterdam ; Boston; p xv, 609 p (2003).
  3. Ray, P. C.; Tummanapalli, J. M. C.; Gorantla, S. R., Process for the large scale production of Stavudine. Google Patents: (2011).
  4. Hightower, T. R.; Heeren, J. D., Using a Simulated Industrial Setting for the Development of an Improved Solvent System for the Recrystallization of Benzoic Acid: A Student-Centered Project. Journal of Chemical Education 83 (11), 1663 (2006).
  5. Rohani, S.; Horne, S.; Murthy, K., Control of Product Quality in Batch Crystallization of Pharmaceuticals and Fine Chemicals. Part 1: Design of the Crystallization Process and the Effect of Solvent. Organic Process Research & Development 9 (6), 858-872 (2005).

Transcript

Recrystallization is a purification technique for solid compounds.

To perform recrystallization, an impure solid compound is mixed with hot solvent to form a saturated solution. As this solution cools, the solubility of the compound decreases, and pure crystals grow from solution.

Recrystallization is often used as a final step after other separation methods such as extraction, or column chromatography. Recrystallization may also be used to separate two compounds with very different solubility properties. This video will illustrate solvent selection for recrystallization, purification of an organic compound from solution, and will introduce a few applications in chemistry.

Crystallization begins with nucleation. Solute molecules come together to form a stable small crystal, which is followed by crystal growth. Nucleation occurs faster on nucleation sites such as seed crystals, scratches, or solid impurities than spontaneously in solution. Agitation may also encourage rapid nucleation. However, rapid growth can lead to incorporation of impurities if not grown in optimal conditions.

The solubility of a compound tends to increase with temperature, and is highly dependent on the choice of solvent. The greater the difference in solubility at high and low temperature, the more likely it is for the solute to come out of the solution as it cools, and form crystals.

The solvent chosen should have a boiling point of at least 40 °C so there is a significant temperature difference between boiling and room temperature. The solvent’s boiling point must also be below the melting point of the solute to enable crystallization. Rapid cooling of the solution induces the formation of many nucleation sites, thus favors the growth of many small crystals. However, slow cooling induces the formation of fewer nucleation sites, and favors larger and purer crystals. Thus, slow cooling is preferred.

Additionally, a solvent can be selected to minimize impurities. If a solution impurity is more soluble than the solute itself, it can be washed off of the fully formed crystals with cold solvent. 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.

To begin this procedure, place 50 mg of the sample in a glass test tube.

Add 0.5 mL of room temperature solvent. 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.

Add a 10% excess of hot solvent to the solution to account for evaporation. Place filter paper in a Büchner funnel setup. 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. Set a Büchner funnel and adapter in the mouth of the flask.

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.

Purification by recrystallization is an important tool for chemical synthesis and analysis.

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.

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JoVE Science Education Database. JoVE Science Education. Purifying Compounds by Recrystallization. JoVE, Cambridge, MA, (2023).