Chromatography and Rf values

Two black felt-tip lines look identical, so how could a strip of paper and a little water tell the two inks apart? Put a spot of each ink on paper, dip the bottom edge in water, and one ink falls apart into blue, pink and yellow. The other climbs the paper as a single black spot. It's tempting to say the second ink must be one substance, but one test in one solvent can't prove that. Testing it fairly takes a second solvent, and a number with no unit.

Paper chromatography separates the substances dissolved in a mixture by letting a solvent carry them up a sheet of paper. It has two phases. The mobile phase is the solvent, which moves up through the paper. The stationary phase is the paper itself, which stays where it is. Mobile means able to move, and stationary means standing still, so the names say which is which. Now a check. Water soaks up a stick of chalk, carrying spots of ink with it. Which is the stationary phase? The chalk. It stays put, so it's the stationary phase, and the water moving through it is the mobile phase. One examiner's report on a Foundation paper says most students did not choose the terms mobile phase and stationary phase, and those who did often labelled them the wrong way round. The habit is to label both phases on the apparatus before explaining anything about it. Each substance in the mixture is pulled two ways. It dissolves in the solvent, which carries it up, and it's attracted to the paper, which holds it back. A substance that is more soluble in the solvent travels further, and one that is more attracted to the paper travels less. Now picture a dye that barely dissolves in the solvent. Does it finish near the bottom or the top? Near the bottom. The solvent can hardly carry it, so it barely leaves the starting line.

Setting up a chromatogram has four rules, and each one has a reason. First, draw the starting line, called the baseline, in pencil. Pencil doesn't dissolve in the solvent, while an ink line would run up the paper with the samples. Second, put the spots on the baseline and keep the solvent level below it, so the spots travel up the paper instead of dissolving into the solvent in the container. Third, put a lid on the container, so the solvent doesn't evaporate from the paper as it climbs. Now a reason to find. Why is the solvent front marked the moment the paper comes out? The solvent front is the furthest point the solvent reached. As the paper dries, that wet edge disappears, so it's marked straight away. Not every course includes this next part, and the screen shows which. Thin layer chromatography swaps the paper for a plate coated in a thin layer of solid, and that layer is the stationary phase. Its solvent can be water, called aqueous, or another liquid such as ethanol, called non-aqueous. A colourless substance is shown up with a locating agent, a chemical that makes invisible spots visible.

An Rf value turns a chromatogram into a number. The Rf value is the distance moved by the substance, divided by the distance moved by the solvent. Both distances are measured from the baseline. The top of the fraction goes to the centre of the spot, and the bottom goes to the solvent front. Now think about the two distances. Can a spot ever travel further than the solvent front? No. The solvent carries it, so the spot always stays behind the front, and an Rf value is always less than one. And because both distances are in centimetres, the units cancel, so an Rf value has no unit. No course gives you this formula in the exam, so it has to be learned by heart. A Foundation report from another year says the Rf value was not well known, and often only the distance the spot moved was given. The fix is to write the whole fraction down first, with both distances in their places, before any number goes in. Here's a chromatogram. A green spot has travelled three point six centimetres from the baseline, and the solvent front is eight point nought centimetres from it. Calculate the Rf value of the green spot to two decimal places, writing the fraction before you divide. Pause it there and work it out for yourself. Check first: three point six is less than eight point nought, so the answer has to be less than one. The fraction is three point six centimetres over eight point nought centimetres, and the centimetres cancel. Three point six divided by eight point nought is nought point four five. So the Rf value of the green spot is nought point four five, with no unit. An examiner's report on a different Foundation paper says an Rf question was generally well answered, with about two thirds of students gaining two or more marks. The same report notes a large variation in the solvent distances given, which suggests students did not know what the solvent front was. So measure the bottom distance to the marked solvent front, never to the top of the paper or the water level.

The formula can also be turned round, to find a distance from an Rf value. Now, with an Rf value and a solvent distance, how do you get the distance the spot moved? Multiply. The Rf value is the fraction of the solvent's distance that the spot travelled, so the distance moved by the substance is the Rf value times the distance moved by the solvent. Try it on a new chromatogram with a known substance on it. With an Rf value of nought point four five and the front at twelve point nought centimetres, how far has it moved? Nought point four five times twelve point nought centimetres is five point four centimetres. And five point four is less than twelve point nought, so it passes the check. Now a student's answer comes out much larger than that. Why can't twenty-six point seven centimetres be right, without any arithmetic? It's further than the solvent front, at twelve point nought centimetres, and a spot can't outrun the solvent that carries it. One examiner's report on a Higher paper says a common incorrect answer came from dividing the distance moved by the solvent front by the Rf value. That's exactly where twenty-six point seven comes from: twelve point nought divided by nought point four five. The fix is to multiply, then check the answer is shorter than the solvent distance before writing it down.

A chromatogram also identifies what's in a mixture, by running it beside known substances on the same paper. A pure substance gives one spot in every solvent. A mixture usually gives more than one, and spots that overlap in one solvent can be pulled apart by a different solvent. A four-lane chromatogram shows a blue, a yellow and a red food colouring, then the coating from a green sweet. The blue gives one spot, the yellow one spot, and the red two spots, while the coating gives two spots, one level with the blue and one level with the yellow. What's in the coating, which lanes could be pure, and how would you check with another solvent? The coating contains the blue and the yellow colourings, because its two spots sit level with theirs. The blue and the yellow each give one spot, so they could be pure. The red gives two spots, so it's a mixture. To check, run it all again in a different solvent. If the blue and yellow still give one spot each, and the coating's spots still match them, the answer stands. Two different questions get mixed up here. A spot below the solvent level spoils the run, while changing the solvent or the paper changes the Rf values. Now a different one. Does the amount of solvent in the container change an Rf value? No. The Rf value depends on which solvent and which stationary phase you use, not on how much solvent is in the container. One examiner's report on a Higher paper names a common misconception that the water level was too low and would therefore affect the Rf value calculated. So keep the two questions apart: what spoils the run, and what changes the Rf value.

Now a last round of four. A spot moves two point four centimetres and the solvent front six point nought centimetres. What is the Rf value?

Two point four over six point nought is nought point four, or nought point four nought to two decimal places, with no unit.

Now turn it round. The Rf value is nought point six and the front is at ten point nought centimetres. How far did the spot move? Nought point six times ten point nought is six point nought centimetres, which is less than ten, so it passes the check.

Now a single spot. A sample gives one spot in water. Is it definitely pure? Not yet. A pure substance gives one spot in every solvent, so run it in another one.

Now the formula itself: how does the Rf formula go, in words? The Rf value is the distance moved by the substance, divided by the distance moved by the solvent.

Back to the two felt tips. The ink that split is a mixture of at least three dyes. The one that climbed as one spot might be a single substance, but only one solvent has tested it so far. Run it in a second solvent and measure its Rf value each time. If it still gives one spot, the case that it's one substance gets much stronger. And if its Rf values match a known dye's in both solvents, you can say which dye it is.

If you can get an Rf value under one with no unit and say why, you own this one, so give it a thumbs-up. If not, give it a day and look again.

That's the end of this chapter on states of matter and separating mixtures. The next chapter is Bonding: ionic, covalent and metallic. Hopefully that's next in your feed, but if not, search for ScholaFly C H oh four, oh oh.

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