4. BONUS: Lab Techniques and Procedures
Chromatography uses differences in molecular attraction to separate components within a mixture.
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So when we say chromatography, we're going to say that this is a technique that involves the separation of components, mainly solids and liquids within a mixture because of a different and molecular attractions, we're going to say in the procedure, a mixture is spotted on a silica plate and the progress of the components on that plate is based on the affinity to the solvent or the plate itself. Now with this process we have two phases. one represents a silica plate, which doesn't move and it holds the mixture because it doesn't move. We call it the stationary phase. The other phase represents the solvent, the portion that's moving the liquid portion. So we call it the mobile phase so it moves up the silica plate and it moves by capillary action. So here I have a TLC plate, I draw a line, I have a mixture and I take a sample of this mixture and I spot the TLC plate, I place it within my mobile phase, which is my solvent. And here let's say that the solvent is 50% ethanol which is CH three ch 20 H, ethanol is slightly polar and then we have 50% hexagons which is C six H 14, it's a hydrocarbon, so it's non polar. Now the dots can start to separate and move up. So here as the solvent phase is moving up the TLC plate, we can see that this dot which is made up of a mixture of two different compounds. Art starts to split, it splits into red dots and green dots to show the separation of the mixture that I originally took from my sample. Now, if the dots themselves, if the the attraction to the plate is greater than the attraction to the solvent, then the dots are not going to move very far up the plate. So there's gonna be low movement. Now let's say that the dots have a greater affinity for the solvent than the plate. So since they have a greater affinity or attraction to the solvent, the mobile phase, they're gonna move with it. So there's gonna be high movement or higher movement. Looking at this, we can see that the at the end the solvent is reached up this high. In terms of the plate, I take it up, we can see that the green dots have moved up higher then the red dot. So the green dots had a higher affinity for the solvent. They moved up with it. The red dots had less of an affinity for the solvent, so they don't move up as high here, I mark 123 and 41 being the original position. The starting starting line two was the position where the Red Dots stopped. Three is the position where the green dot stopped and four is my mobile front. That's where my uh solvent stopped where I took it took the paper out from these numbers. I can determine my R. F. Value. So the distance traveled by the components is a method we use to we use we can use to find the R. F. Value. This helps us in the identification of the compound. You're, our F value equals distance traveled by compound divided by distance traveled by solvent. So how would I do this? Well, if we're looking at the red dots here, we'd say the red dots traveled to and the solvent traveled for. So the R. F. Value of red is 0.50 or green. They traveled three, Solvent traveled four, so that's .75. So these are the R. F values of the red dot and the green dot. Usually you'd have a manual and associated with these numbers, you'd have uh names of actual compounds with our F values and you choose which compound from that list matches closely with 0.15 point 75. And in that way you'd identify what is the red dot represents, what compound and what does the green dot represent. So that's how we can use basically TLC plates in spotting mobile phases and stationary phases to determine the identity of these two different spots. Okay. And we incorporate the R. F value to do so.
The mixture that is spotted on the silica gel plate has its movement based on its affinity to either the solvent or to the plate itself.