Name: tlc separation by polarity
Uploaded: 2020-03-26T07:22:28.000Z
Duration: 3 min 7 s
Channel: JoVE

薄層クロマトグラフィー(TLC)は、極性の違いに基づいて有機化合物を分離するために使用される技術です。TLCシステムには、固定相、溶質、現像溶媒の3つのコンポーネントがあります。
固定相は、化合物が分離される表面であり、通常は極性の高いシリカゲルで構成されています。分析される化合物または混合物は溶質です。溶質の希薄溶液は、細いキャピラリーを使用してプレートの一方の端にスポットされます。
次に、プレートの底部を現像溶媒(通常は固定相よりも極性の低い有機溶媒)に浸します。溶媒は毛細管現象によってプレートを上に移動します。溶媒が溶質スポットを通過すると、各溶質の一部が一緒に運ばれます。この溶質-溶媒混合物が移動相です。
移動相の上部はソルベントフロントと呼ばれます。移動相がプレートを上に移動すると、溶質分子と溶媒分子は、その極性に基づいて一時的に固定相に付着します。
より多くの極性溶質は固定相に強く引き付けられるため、より頻繁に付着し、しばらくの間留まります。極性の低い溶質は、くっつく頻度が少なくなり、長く固まることはありません。これは、極性の低い溶質は、極性が高い溶質よりも移動相で過ごす時間が長いことを意味します。言い換えれば、極性の低い溶質は、極性が高い溶質よりも速く移動します。
化合物間の良好な分離を達成するためには、溶媒は最も極性の高い溶質を移動させるのに十分な極性である必要があります。ここでは、一般的な溶媒を極性が増す順に示します。場合によっては、2つの溶媒を組み合わせて目的の極性を達成することもあります。
ほとんどの有機化合物は白色または無色であるため、TLCプレートには通常、蛍光化合物が含まれています。紫外線の下では、有機化合物は光るプレート上に暗いスポットとして現れます。
化合物が視覚化されたら、化合物がどれだけ移動したかを測定し、各化合物の遅延係数(Rf)を計算できます。この値は、溶質が移動した距離を移動相が移動した合計距離で割った値です。
溶媒が適切に選択されている場合、各溶質は明確に異なるRfを持ちます。これにより、Rf値を既知の化合物と一致させることにより、混合物中の成分化合物を同定することができます。
このラボでは、TLC を使用していくつかの既知の化合物の Rf を決定し、次にそれらの Rf 値を使用して未知の化合物の同一性を判別します。

薄層クロマトグラフィー

クロマトグラフィー
クロマトグラフィーは、有機化学で使用される手法で、2つの異なる相間の溶解度の違いに基づいて混合物中の化合物を分離します。この概念は液液抽出と似ていますが、クロマトグラフィーでは、2つの相が固定相と移動相で構成される点が異なります。固定相は固体(通常はマイクロスケールのハイドロゲルビーズ)であり、移動相はキャリア溶媒です。
従来のクロマトグラフィーでは、固定相を垂直カラムに充塡し、溶液混合物をカラムの上部に導入します。混合物が固定相を流れると、化合物はその構造と極性に基づいて固定相と移動相の間で分配され、離散的なバンドを形成します。固定相と弱く相互作用する溶質は、カラム内をより速く移動し、最初に出る(溶出する)ことができます。固定相とより強く相互作用する溶質は、カラム内をゆっくりと移動し、後で溶出します。バンドは、混合物中の化合物を分離および精製するために、個別に収集できます。
クロマトグラフィーにはいくつかの種類があり、それぞれが異なる化学的特性を利用して分離を実現します。例えば、イオン交換クロマトグラフィーでは、固定相ビーズを正または負に帯電させ、反対の電荷を持つ分子のみを引き付けることができます。サイズ排除クロマトグラフィーでは、多孔質固定相はアガロースポリマーやデキストランポリマーなどのビーズで構成されています。小さな分子は細孔に入りやすく、大きな分子は細孔を通過してより速く溶出します。
<h4>薄層クロマトグラフィー</h4>
薄層クロマトグラフィー(TLC)は、化合物の極性に基づいて化合物を分離するクロマトグラフィー技術の一種です。従来のクロマトグラフィーと同様に、TLCシステムには、固定相、移動相、溶質の3つのコンポーネントがあります。ただし、従来のクロマトグラフィーとは異なり、固定相はカラムに充填されるのではなく、プレート上の薄い層に配置されます。TLCは、ほとんどの場合、二酸化ケイ素の一種である極性シリカゲルを固定相として使用します。固定相は、その表面のOH基により水素結合を形成します。
まず、TLCプレートの底面に鉛筆でスタートラインを描きます。分析対象の化合物または混合物は、細いキャピラリーを使用してスタートラインで検出されます。次に、プレートの底部を移動相(通常は固定相よりも極性の低い有機溶媒)に浸漬します。溶媒は毛細管現象によってプレートを上に移動し、溶質スポットを通過して各成分の一部を運びます。
溶媒がプレートを上に移動すると、成分は移動相または固定相のいずれかに分配されます。成分が極性の場合、極性固定相とより相互作用します。ゆっくりと移動し、TLCプレート上を短い距離しか移動しません。サンプルの成分が極性が低く、固定相よりも移動相に溶けやすい場合、サンプルは移動相とより相互作用し、TLCプレート上をより遠くに移動します。成分と移動相の極性の範囲は、分離を理解し予測するために不可欠です。
TLCプレートには通常、254ナノメートルのUV光源の下で光るUV反応性蛍光色素が含まれています。そのため、TLCプレートは紫外線下で観察することで分析することができます。目的の溶質など、TLCプレート内の化合物は、緑色の背景と比較して暗いスポットとして表示されます。グラファイトペンシルでスポットを一周することにより、化合物が溶媒前面に対して移動した距離を測定できます。有機化合物のスポットは、それ自体が蛍光でない場合、プレートの蛍光を覆い隠し、ダークスポットとして現れます。有機化合物の中には、紫外線に強いものがあり、紫外線を浴びると発光します。これらは通常、共役化合物、つまり二重結合と単結合が交互に発生する化合物であり、放出される波長によって識別できます。
<h4>遅延係数</h4>
特定の溶媒で成分の遅延係数(Rf)を分析することにより、TLCを使用して未知の溶質を決定できます。遅延係数は、移動相が移動した距離に対するコンポーネントが移動した距離の比率です。
<img src="/CDNSource/lm/labs/62/62_Concepts_1.jpg">
溶質の移動距離は開始線からスポットの中心点まで測定され、移動相の移動距離は同じ開始線から溶媒フロントまで測定されます。化合物の遅延係数は、使用する移動相によって異なります。遅延係数は、非極性移動相を持つ非極性が高い化合物で大きくなります。リタデーション係数の値は、非極性移動相を持つ極性コンポーネントで見られます。
非常に非極性な移動相の場合、一部の極性成分がまったく移動しないことがあります。これにより、遅延係数が非常に低く、分離が不十分になります。高極性の移動相により、化合物は溶媒とともに移動し、非常に高い遅延係数が得られます。これにより、コンポーネント間の分離がほとんどなくなります。
分離を効果的に行うには、コンポーネントの遅延係数を約0.3〜0.7離す必要があります。効率的な移動相を見つけるために、試行錯誤を重ねます。多くの場合、2つの溶剤を混合すると最も効果的であることが証明されています。
<h4>参照</h4>
<ol>
	<li style="font-size:17px">ハリス、ワシントンD.C.(2015年)。定量的化学分析。ニューヨーク州ニューヨーク:WHフリーマンアンドカンパニー。</li>
</ol>

クロマトグラフィー
クロマトグラフィーは、有機化学で使用される手法で、混合物中の化合物を、...

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JoVE Lab Manual

Chemistry

化学

Thin-Layer Chromatography

tlc separation by polarity

Thin-layer chromatography (TLC) separates compounds in a mixture by polarity. It uses the same basic parts as other chromatography methods: a stationary phase, a mobile phase, and the solute. In TLC, the stationary phase is a thin coating on a plate instead of material packed into a column.

The stationary phase in most TLC systems is polar silica gel, a form of silicon dioxide. Its surface has OH groups that can form hydrogen bonds. The sample is first spotted on a pencil start line with a thin capillary. Then the bottom of the plate is placed in the mobile phase, which is usually an organic solvent that is less polar than the stationary phase.

As the solvent moves up the plate by capillary action, it carries the sample components with it. Each compound then partitions between the mobile phase and the stationary phase. A polar component interacts more with the polar stationary phase, so it moves more slowly and stays near the start. A less polar component is more soluble in the mobile phase, so it travels farther up the plate.

TLC plates often include a UV-reactive fluorescent dye that glows under 254 nm UV light. The compounds appear as dark spots against the green background, and the spots can be circled with a graphite pencil so their movement can be measured. Some organic compounds are UV-active and emit light when exposed to UV light. These are often conjugated compounds, which have alternating double and single bonds, and they can be identified by the wavelength they emit.

The retardation factor, or Rf, helps identify an unknown solute from TLC results. It is the ratio of the distance traveled by the compound to the distance traveled by the solvent front. The compound distance is measured from the start line to the center of the spot, and the solvent distance is measured from the same start line to the solvent front. Rf depends on the mobile phase used.

Rf values are high for highly non-polar compounds with a non-polar mobile phase, and low for polar compounds with a non-polar mobile phase. If the mobile phase is too non-polar, some polar compounds may not move at all. If it is too polar, the compound may move with the solvent and the separation may be poor. For an effective separation, the Rf values of the components should be about 0.3 to 0.7 apart. Choosing the best mobile phase usually takes trial and error, and a mixture of two solvents often works well.

TLC Separation by Polarity

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chemistry

入門化学ラボコースの指導者と学生のための、包括的でカリキュラムに焦点を当てたビデオ。

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Learn how thin-layer chromatography separates compounds by polarity, how spots are seen under UV light, and how to use Rf values to compare samples.

TLC Separation by Polarity | JoVE K12

薄層クロマトグラフィー を教える準備を、この 6 分間の教員向けガイドで行いましょう。JoVE Lab Manual の Chemistry ラボの主要な準備手順を解説します。

Chemistry の 薄層クロマトグラフィー 教員準備 - 8.8K+ 回の視聴 | JoVE Lab Manual

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TLC Separation by Polarity

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