Reading a Chromatogram

Introduction

Once the paper has dried, a chromatogram looks like a simple pattern of coloured dots. Reading it well is a skill. How many substances are in each sample? Do two samples share an ingredient? Did anything fail to move at all? Every spot and its position carries information, and learning to read that information is what makes chromatography useful in laboratories, factories and courtrooms.

Core explanation

Layout. A chromatogram has a baseline at the bottom where the samples were placed, a solvent front near the top, and above each starting point a vertical column of spots. Each column belongs to one sample.

Counting spots. Each separate spot above a sample represents a different substance . So:

- a sample giving three spots contains at least three components; - a sample giving one spot may be a single pure substance.

The phrase "at least" matters. Two different substances might happen to travel exactly the same distance in the chosen solvent and appear as one spot. Colourless substances may also be invisible unless the paper is treated. So the number of visible spots is a minimum, not a guaranteed total.

Comparing positions. Samples run on the same paper with the same solvent can be compared directly. If a spot in one sample is at exactly the same height as a spot in another and has the same colour, the two spots are probably the same substance. If the heights differ, they are different substances.

Spots on the baseline. A spot that has not moved from the baseline is a substance that is insoluble in the solvent used. It is still a component of the mixture. Trying a different solvent may make it move.

Quality of spots. Tidy, round spots are easy to compare. Long streaks usually mean the original spot was too large or too concentrated. Faint spots mean there is only a little of that substance. The size and intensity of a spot give a rough idea of how much of a substance is present, but the height tells you what it is.

Putting it together. Suppose sample X gives a red spot high up and a blue spot low down, and reference dye R gives a single red spot at the same height as X's red spot. Then X contains dye R plus at least one other substance.

Step-by-step reasoning

To read a chromatogram:

1. Find the baseline and the solvent front. 2. For each sample, count the spots, including any on the baseline. 3. Note the colour and height of each spot. 4. Match spots at the same height and colour across samples. 5. State conclusions using "at least" where appropriate.

Visual explanation

Picture five columns of spots. The first, a known dye, has one yellow spot halfway up. The fourth, an unknown sweet colouring, has three spots: one yellow at exactly the same height, one pink near the top and one purple just above the baseline. The matching yellow spot stands out immediately.

Real-world analogy

Reading a chromatogram is like comparing bar codes at a supermarket. Each product has lines at particular positions. If two labels have lines in exactly the same places, they belong to the same product; a line in a different place means something is different.

Real-world example

Quality-control scientists at an ink factory run each new batch beside a sample of the approved ink. If the new batch shows the same spots at the same heights, it has the correct blend. An extra spot or a missing one signals a mistake in the recipe.

Why?

Why can samples only be compared directly if they are on the same paper? The distance travelled depends on the solvent, the paper, the temperature and how far the solvent ran. Running samples side by side makes all these conditions identical, so any difference in height is due to the substances themselves.

Common misconception

"A large spot means that substance travelled further or is a different substance." Spot size reflects how much of the substance was present or how big the starting drop was. Identity is judged by position (and colour), not by size.

Worked example

Question: On one chromatogram, food colouring F gives spots at heights of 2.0 cm, 4.5 cm and 6.0 cm. Reference dyes P, Q and S give single spots at 4.5 cm, 3.0 cm and 6.0 cm. Which reference dyes are in F?

Reasoning: Compare heights. F's spots at 4.5 cm and 6.0 cm match P and S. No spot in F is at 3.0 cm, so Q is absent. F's spot at 2.0 cm matches none of the references.

Answer: F contains P and S, does not contain Q, and contains at least one other unknown substance.

Quick check

1. A sample gives two spots. What is the smallest number of substances it contains? Answer: Two.

Exam focus

Questions usually show a chromatogram and ask how many substances are in a sample or which known substances a mixture contains. Count every spot, including one left on the baseline, and match by height across the same paper. Use "at least" when stating the number of components.

Advanced insight

When two substances co-run in one solvent, scientists can use two-way chromatography: they run the paper in one solvent, turn it through 90° and run it again in a different solvent. Substances that overlapped in the first direction usually separate in the second, spreading the spots across the paper like a map.

Summary

Each spot above a sample is a different substance, so the number of spots gives the minimum number of components. A spot left on the baseline is a substance insoluble in that solvent. Spots at the same height and colour on the same chromatogram are probably the same substance. Spot size indicates amount, while position indicates identity.

Practice questions

1. What does a spot remaining on the baseline tell you? Answer: That component is insoluble in the solvent used, so it did not move. 2. Why do we say a sample with one spot "may" be pure rather than "is" pure? Answer: Two substances could travel the same distance and overlap, or a colourless substance may be invisible, so one spot is not absolute proof. 3. Sample A has spots at 3 cm and 5 cm; sample B has one spot at 5 cm on the same paper. What can you conclude? Answer: A probably contains the substance in B, plus at least one other substance. 4. Why should samples to be compared be run on the same chromatogram? Answer: So the solvent, paper, temperature and solvent distance are identical, making the heights directly comparable.