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

İnce tabaka kromatografisi veya TLC, organik bileşikleri polaritelerindeki farklılıklara göre ayırmak için kullanılan bir tekniktir. Bir TLC sisteminin üç bileşeni vardır: durağan faz, çözünen ve geliştirme çözücüsü.
Sabit faz, genellikle son derece polar silika jelden oluşan bileşiklerin ayrılacağı yüzeydir. Analiz edilen bileşikler veya karışımlar çözünen maddelerdir. Çözünen maddelerin seyreltik çözeltileri, ince kılcal damarlar kullanılarak plakanın bir kenarında lekelenir.
Plakanın alt kısmı daha sonra, genellikle sabit fazdan daha az polar olan organik bir çözücü olan geliştirme çözücüsüne daldırılır. Çözücü, kılcal hareket ile plakayı yukarı doğru hareket ettirir. Çözücü çözünen maddelerden geçerken, her bir çözünen maddenin bir kısmını beraberinde taşır. Bu çözünen-çözücü karışımı mobil fazdır.
Mobil fazın üst kısmına solvent cephesi denir. Mobil faz plakadan yukarı doğru hareket ederken, çözünen ve çözücü molekülleri, polaritelerine bağlı olarak geçici olarak sabit faza yapışır.
Daha fazla polar çözünen madde, durağan faza güçlü bir şekilde çekilir, bu nedenle daha sık yapışırlar ve bir süre takılı kalırlar. Daha az polar çözünen madde daha az yapışır ve uzun süre takılı kalmaz. Bu, daha az polar çözünen maddenin mobil fazda daha fazla polar çözünen maddeden daha fazla zaman harcadığı anlamına gelir. Başka bir deyişle, daha az polar çözünen, daha fazla polar çözünenden daha hızlı hareket eder.
Bileşikler arasında iyi bir ayrım elde etmek için, çözücü en polar çözünen maddeyi hareket ettirecek kadar polar olmalıdır. İşte artan polarite sırasına göre bazı yaygın çözücüler. Bazen, istenen polariteyi elde etmek için iki çözücü birleştirilir.
Organik bileşiklerin çoğu beyaz veya renksiz olduğundan, TLC plakalarının içinde genellikle bir floresan bileşiği bulunur. UV ışığı altında, organik bileşikler parlayan plaka üzerinde koyu lekeler olarak görünecektir.
Bileşikler görselleştirildikten sonra, ne kadar uzağa gittiklerini ölçebilir ve her biri için geciktirme faktörünü veya Rf'yi hesaplayabilirsiniz. Bu değer, çözünen maddenin kat ettiği mesafenin, mobil faz tarafından kat edilen toplam mesafeye bölünmesiyle elde edilir.
Çözücü uygun şekilde seçilirse, her çözünen madde belirgin şekilde farklı bir Rf'ye sahip olacaktır. Bu, Rf değerlerini bilinen bileşiklerle eşleştirerek karışımlardaki bileşen bileşiklerini tanımlamamızı sağlar.
Bu laboratuvarda, bilinen birkaç bileşiğin Rf'sini belirlemek için TLC'yi kullanacaksınız ve ardından bilinmeyen bir bileşiğin kimliğini belirlemek için bu Rf değerlerini kullanacaksınız.

İnce tabaka kromatografisi

Kromatografi
Kromatografi, organik kimyada, iki farklı faz arasındaki çözünürlük farklılıklarına dayalı olarak bir karışımdaki bileşikleri ayırmak için kullanılan bir tekniktir. Konsept, sıvı-sıvı ekstraksiyonuna benzer, kromatografi dışında, iki faz sabit faz ve mobil fazdan oluşur. Sabit faz bir katıdır - tipik olarak mikro ölçekli bir hidrojel boncuk - mobil faz ise bir taşıyıcı çözücüdür.
Geleneksel kromatografide, sabit faz dikey bir kolona paketlenir ve çözelti karışımı kolonun tepesine eklenir. Karışım durağan fazdan akarken, bileşikler yapılarına ve polaritelerine göre durağan faz ile mobil faz arasında bölünür ve ayrı bantlar oluşturur. Durağan faz ile zayıf bir şekilde etkileşime giren çözünen maddeler, kolon boyunca daha hızlı hareket eder ve önce çıkar - veya elute - . Durağan faz ile daha güçlü etkileşime giren çözünen maddeler, kolon boyunca yavaşça hareket eder ve daha sonra elüte olur. Bantlar, bir karışımdaki bileşikleri izole etmek ve saflaştırmak için ayrı ayrı toplanabilir.
Her biri ayırmayı sağlamak için farklı bir kimyasal özellikten yararlanan birkaç kromatografi türü vardır. Örneğin, iyon değişim kromatografisinde, sabit faz boncukları pozitif veya negatif yüklü olabilir ve yalnızca zıt yüklü molekülleri çekebilir. Boyut dışlama kromatografisinde, gözenekli sabit faz, agaroz veya dekstran polimerleri gibi boncuklardan oluşur. Daha küçük moleküller gözeneklere daha kolay girebilirken, daha büyük moleküller gözeneklerden geçer ve daha hızlı elüte olur.
<h4>İnce Tabaka Kromatografisi</h4>
İnce tabaka kromatografisi (TLC), bileşikleri polaritelerine göre ayıran bir tür kromatografi tekniğidir. Geleneksel kromatografi gibi, bir TLC sisteminin üç bileşeni vardır: sabit faz, mobil faz ve çözünen. Bununla birlikte, geleneksel kromatografiden farklı olarak, sabit faz, bir kolona paketlenmek yerine bir plaka üzerinde ince bir tabaka halinde düzenlenir. TLC çoğunlukla sabit faz olarak bir silikon dioksit formu olan polar silika jeli kullanır. Durağan faz, yüzeyindeki OH grupları nedeniyle hidrojen bağları oluşturur.
İlk olarak, bir kalem kullanılarak TLC plakasının altına bir başlangıç çizgisi çizilir. Analiz edilen bileşikler veya karışım, ince bir kılcal damar kullanılarak başlangıç çizgisinde tespit edilir. Daha sonra, plakanın alt kısmı, genellikle sabit fazdan daha az polar olan organik bir çözücü olan mobil faza daldırılır. Çözücü, kılcal hareketle plakayı yukarı doğru hareket ettirir, çözünen maddeleri geçer ve her bir bileşenin bir kısmını beraberinde taşır.
Çözücü plakadan yukarı doğru hareket ederken, bileşenler ya mobil faza ya da sabit faza ayrılır. Bileşen polar ise, polar sabit faz ile daha fazla etkileşime girer. Yavaş hareket eder ve TLC plakası üzerinde sadece kısa bir mesafe hareket eder. Numunenin bileşeni daha az polar ise - ve mobil fazda sabit fazda olduğundan daha fazla çözünürse - mobil faz ile daha fazla etkileşime girer ve TLC plakası üzerinde daha uzağa hareket eder. Bileşenin polaritesinin derecesi ve mobil faz, ayrılmayı anlamak ve tahmin etmek için çok önemlidir.
TLC plakaları tipik olarak 254 nanometrelik bir UV kaynağı altında parlayacak UV reaktif bir floresan boya içerir. Bu nedenle, TLC plakaları UV ışığı altında gözlemlenerek analiz edilebilir. İlgilenilen çözünen maddeler gibi TLC plakası içindeki bileşikler, yeşil bir arka plana kıyasla koyu lekeler olarak görünecektir. Noktaları bir grafit kalemle daire içine alarak, bileşiklerin çözücü cephesine göre kat ettiği mesafe ölçülebilir. Organik bileşiğin noktası, floresanın kendisi değilse, plakanın floresansını maskeler ve karanlık bir nokta olarak ortaya çıkar. Bazı organik bileşikler UV aktiftir ve UV ışığına maruz kaldıklarında ışık yayarlar. Bunlar tipik olarak konjuge bileşiklerdir, yani alternatif çift ve tek bağlara sahip olanlardır ve yayılan dalga boyu ile tanımlanabilirler.
<h4>Geciktirme Faktörü</h4>
Bir bileşenin geciktirme faktörünü (Rf) belirli bir çözücü ile analiz ederek, TLC kullanılarak bilinmeyen bir çözünen madde belirlenebilir. Geciktirme faktörü, bir bileşen tarafından kat edilen mesafenin, mobil faz tarafından kat edilen mesafeye oranıdır.
<img src="/CDNSource/lm/labs/62/62_Concepts_1.jpg">
Çözünen maddenin kat ettiği mesafe, başlangıç çizgisinden noktanın merkez noktasına kadar ölçülür ve mobil fazın kat ettiği mesafe, aynı başlangıç çizgisinden çözücü cephesine kadar ölçülür. Bir bileşiğin geciktirme faktörü, kullanılan mobil faza bağlıdır. Geciktirme faktörü, polar olmayan bir mobil faz ile yüksek oranda polar olmayan bileşikler için büyüktür. Polar olmayan mobil faza sahip polar bileşenler için düşük geciktirme faktörü değerleri görülür.
Yüksek derecede polar olmayan bir mobil faz durumunda, bazı polar bileşenler hiç hareket etmeyebilir. Bu, son derece düşük bir geciktirme faktörü ve yetersiz bir ayırma ile sonuçlanır. Oldukça polar bir mobil faz, bileşiğin çözücü ile hareket etmesine neden olur ve son derece yüksek bir geciktirme faktörü verir. Bu, bileşenler arasında çok az ayrım ile sonuçlanır.
Ayırmanın etkili olması için, bileşenlerin geciktirme faktörleri yaklaşık 0,3 - 0,7 aralıklı olmalıdır. Verimli mobil aşamayı bulmak için deneme yanılma yöntemi kullanılır. Genellikle, iki çözücünün karışımı en etkili olduğunu kanıtlar.
<h4>Başvuru</h4>
<ol>
	<li style="font-size:17px">Harris, D.C. (2015). Kantitatif Kimyasal Analiz. New York, NY: W.H. Freeman ve Şirketi.</li>
</ol>

İnce Tabaka Kromatografisi

Kromatografi
Kromatografi, organik kimyada, bir karışımdaki bileşikleri, aralarındaki çözünürlük farklılıklarına göre ayırmak için kullanılan bir tekniktir.

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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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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

İnce Tabaka Kromatografisi öğretimine bu 6 dakikalık eğitmen rehberiyle hazırlanın. JoVE Lab Manual üzerindeki Chemistry laboratuvarı için temel hazırlık adımlarını kapsar.

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15 dakikalık bu öğrenci protokol videosunda İnce Tabaka Kromatografisi prosedürlerini adım adım takip edin. JoVE Lab Manual üzerindeki Chemistry koleksiyonunun bir parçasıdır.

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TLC Separation by Polarity

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