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

La cromatografía en capa fina, o TLC, es una técnica utilizada para separar compuestos orgánicos en función de las diferencias en sus polaridades. Hay tres componentes de un sistema TLC: la fase estacionaria, el soluto y el solvente de desarrollo.
La fase estacionaria es la superficie en la que se separarán los compuestos, que suele estar compuesta por un gel de sílice extremadamente polar. Los compuestos o mezclas que se analizan son los solutos. Las soluciones diluidas de los solutos se ven en un borde de la placa utilizando capilares delgados.
A continuación, la parte inferior de la placa se sumerge en el disolvente de revelado, que suele ser un disolvente orgánico menos polar que la fase estacionaria. El disolvente se desplaza por la placa por capilaridad. A medida que el solvente pasa a través de las manchas de soluto, arrastra algo de cada soluto consigo. Esta mezcla soluto-disolvente es la fase móvil.
La parte superior de la fase móvil se denomina frente solvente. A medida que la fase móvil viaja por la placa, las moléculas de soluto y solvente se adhieren temporalmente a la fase estacionaria en función de su polaridad.
Los solutos polares más polares son atraídos fuertemente por la fase estacionaria, por lo que se adhieren con más frecuencia y permanecen atascados durante un tiempo. Los solutos menos polares se adhieren con menos frecuencia y no permanecen atascados por mucho tiempo. Esto significa que menos solutos polares pasan más tiempo en la fase móvil que más solutos polares. En otras palabras, los solutos menos polares viajan más rápido que los solutos más polares.
Para lograr una buena separación entre los compuestos, el solvente debe ser lo suficientemente polar como para mover el soluto más polar. Estos son algunos solventes comunes en orden creciente de polaridad. A veces, se combinan dos disolventes para lograr la polaridad deseada.
Debido a que la mayoría de los compuestos orgánicos son blancos o incoloros, las placas TLC generalmente contienen un compuesto fluorescente. Bajo la luz ultravioleta, los compuestos orgánicos aparecerán como manchas oscuras en la placa brillante.
Una vez que se visualizan los compuestos, se puede medir qué tan lejos viajaron y calcular el factor de retardo, o Rf, para cada uno. Este valor es la distancia recorrida por el soluto dividida por la distancia total recorrida por la fase móvil.
Si el solvente se elige correctamente, cada soluto tendrá una Rf claramente diferente. Esto nos permite identificar los compuestos componentes en las mezclas haciendo coincidir los valores de Rf con los compuestos conocidos.
En este laboratorio, usará TLC para determinar el Rf de varios compuestos conocidos y, a continuación, usará esos valores de Rf para determinar la identidad de un compuesto desconocido.

cromatografía en capa fina

cromatografía
La cromatografía es una técnica utilizada en química orgánica para separar compuestos en una mezcla en función de sus diferencias de solubilidad entre dos fases diferentes. El concepto es similar a la extracción líquido-líquido, excepto que en cromatografía, las dos fases consisten en la fase estacionaria y la fase móvil. La fase estacionaria es un sólido, generalmente una perla de hidrogel a microescala, mientras que la fase móvil es un solvente portador.
En la cromatografía tradicional, la fase estacionaria se empaqueta en una columna vertical y la mezcla de solución se introduce en la parte superior de la columna. A medida que la mezcla fluye a través de la fase estacionaria, los compuestos se dividen entre la fase estacionaria y la fase móvil en función de sus estructuras y polaridades, formando bandas discretas. Los solutos que interactúan débilmente con la fase estacionaria se mueven más rápidamente a través de la columna y salen, o eluyen, primero. Los solutos que interactúan más fuertemente con la fase estacionaria se mueven lentamente a través de la columna y se eluyen más tarde. Las bandas se pueden recolectar individualmente para aislar y purificar los compuestos en una mezcla.
Existen varios tipos de cromatografía, cada uno de los cuales aprovecha una propiedad química diferente para lograr la separación. Por ejemplo, en la cromatografía de intercambio iónico, las perlas de fase estacionaria pueden estar cargadas positiva o negativamente, atrayendo moléculas con carga opuesta solamente. En la cromatografía de exclusión por tamaño, la fase estacionaria porosa está compuesta por perlas como polímeros de agarosa o dextrano. Las moléculas más pequeñas pueden entrar en los poros más fácilmente, mientras que las moléculas más grandes fluyen a través de los poros y eluyen más rápido.
<h4>Cromatografía en capa fina</h4>
La cromatografía en capa fina (TLC) es un tipo de técnica de cromatografía que separa los compuestos en función de su polaridad. Al igual que la cromatografía tradicional, hay tres componentes de un sistema TLC: la fase estacionaria, la fase móvil y el soluto. Sin embargo, a diferencia de la cromatografía tradicional, la fase estacionaria está dispuesta en una capa delgada en una placa en lugar de empaquetarse en una columna. La TLC utiliza con mayor frecuencia gel de sílice polar, una forma de dióxido de silicio, como fase estacionaria. La fase estacionaria forma enlaces de hidrógeno debido a los grupos OH en su superficie.
Primero, se dibuja una línea de inicio en la parte inferior de la placa TLC con un lápiz. Los compuestos o mezclas que se están analizando se detectan en la línea de salida mediante un capilar fino. Luego, la parte inferior de la placa se sumerge en la fase móvil, que suele ser un solvente orgánico que es menos polar que la fase estacionaria. El disolvente viaja por la placa por capilaridad, pasando los puntos de soluto y arrastrando consigo parte de cada componente.
A medida que el solvente viaja por la placa, los componentes se dividen en la fase móvil o en la fase estacionaria. Si el componente es polar, interactúa más con la fase estacionaria polar. Viaja lentamente y solo se mueve una distancia corta en la placa TLC. Si el componente de la muestra es menos polar, y más soluble en la fase móvil que en la fase estacionaria, interactúa más con la fase móvil y viaja más lejos en la placa TLC. El alcance de la polaridad del componente y la fase móvil son esenciales para comprender y predecir la separación.
Las placas TLC suelen contener un tinte fluorescente reactivo a los rayos UV que brillará bajo una fuente UV de 254 nanómetros. Por lo tanto, las placas TLC se pueden analizar observándolas bajo luz ultravioleta. Los compuestos dentro de la placa TLC, como los solutos de interés, aparecerán como manchas oscuras en comparación con un fondo verde. Al rodear las manchas con un lápiz de grafito, se puede medir la distancia recorrida por los compuestos en relación con el frente del solvente. La mancha del compuesto orgánico, si no es fluorescente en sí misma, enmascara la fluorescencia de la placa y se muestra como una mancha oscura. Algunos compuestos orgánicos son activos a los rayos UV y emiten luz cuando se exponen a la luz ultravioleta. Por lo general, se trata de compuestos conjugados, es decir, con enlaces dobles y simples alternados, y se pueden identificar por la longitud de onda emitida.
<h4>Factor de retardo</h4>
Al analizar el factor de retardo (Rf) de un componente con un solvente específico, se puede determinar un soluto desconocido usando TLC. El factor de retardo es la relación entre la distancia recorrida por un componente y la distancia recorrida por la fase móvil.
<img src="/CDNSource/lm/labs/62/62_Concepts_1.jpg">
La distancia recorrida por el soluto se mide desde la línea de inicio hasta el punto central de la mancha, y la distancia recorrida por la fase móvil se mide desde la misma línea de inicio hasta el frente del solvente. El factor de retardo de un compuesto depende de la fase móvil utilizada. El factor de retardo es grande para los compuestos que son altamente apolares con una fase móvil no polar. Se observan valores bajos del factor de retardo para los componentes polares con una fase móvil no polar.
En el caso de una fase móvil altamente no polar, es posible que algunos componentes polares no se muevan en absoluto. Esto da como resultado un factor de retardo extremadamente bajo y una separación insuficiente. Una fase móvil altamente polar hace que el compuesto se mueva con el solvente y produce un factor de retardo extremadamente alto. Esto da como resultado muy poca separación entre los componentes.
Para que la separación sea efectiva, los factores de retardo de los componentes deben estar separados entre 0,3 y 0,7. Para encontrar la fase móvil eficiente, se emplea el ensayo y error. A menudo, una mezcla de dos disolventes resulta ser la más eficaz.
<h4>Referencias</h4>
<ol>
	<li style="font-size:17px">Harris, D.C. (2015). Análisis químico cuantitativo. Nueva York, NY: W.H. Freeman and Company.</li>
</ol>

Cromatografía en capa fina

Cromatografía
La cromatografía es una técnica utilizada en química orgánica para separar compuestos en una mezcla en función de sus diferencias de solubilidad ...

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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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TLC Separation by Polarity

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