Chromatography

Synthesising a compound is only half the job — you also need to separate it from the by-products and identify what you have made. Chromatography is the analytical technique that answers both questions. It is used everywhere: forensic drug identification, dope-testing at the Olympics, purifying biotech products, food safety, clinical diagnostics. At A-Level you meet three variants: thin-layer chromatography (TLC), gas chromatography (GC) and the combined GC-MS.

This lesson covers the OCR A-Level Chemistry A (H432) specification point 6.3.1: principles of chromatography, interpretation of TLC Rf values, GC retention times and use of GC-MS for identification.

1. The Big Idea — Partition Between Two Phases

All chromatographic techniques work on the same underlying principle:

Key Principle: A sample is split between a stationary phase (which stays put) and a mobile phase (which moves). Different components of the sample spend different amounts of time in each phase, so they travel at different speeds, and become separated along the direction of mobile-phase flow.

The strength of interaction between each component and each phase depends on the components' polarity, size and chemical affinities. So any two compounds with different structures will travel at different speeds, and can (in principle) be separated.

Technique	Stationary phase	Mobile phase
TLC	Silica or alumina on a plate	Liquid solvent (e.g. ethanol, cyclohexane)
GC	Viscous oil on an inert solid	Gas (N₂, He)
Column chromatography	Silica in a column	Liquid solvent

2. Thin-Layer Chromatography (TLC)

2.1 Setup

A small rectangular plastic or glass plate is coated with a thin layer of silica gel (SiO₂) or alumina (Al₂O₃) — this is the stationary phase.
A pencil line is drawn near the bottom (not ink — ink would dissolve and move with the solvent!).
A spot of the sample (dissolved in a volatile solvent) is applied to the line.
The plate is stood in a beaker containing a small volume of solvent (the mobile phase) — the solvent level must be below the pencil line or the sample washes away.
The beaker is covered with a lid or watch glass to prevent evaporation.

2.2 What happens

The solvent rises up the plate by capillary action, carrying the sample components with it. Each component partitions between:

Adsorption on the silica gel (stationary phase) — more polar compounds stick harder.
Dissolution in the solvent (mobile phase) — less polar compounds dissolve more.

Less polar components travel further up the plate because they spend more time in the moving solvent. More polar components travel less far because they are held more strongly on the silica.

When the solvent nears the top of the plate, the plate is removed and the solvent front is marked with pencil. The plate is then dried, and the spots are visualised (see Section 2.4).

graph LR
  A[Apply sample on pencil line] --> B[Stand in solvent, solvent level below line]
  B --> C[Solvent rises by capillary action]
  C --> D[Components separate by polarity]
  D --> E[Mark solvent front, dry, visualise]
  E --> F[Calculate Rf values]

2.3 Rf value

The retention factor (Rf) is the ratio of the distance travelled by the spot to the distance travelled by the solvent front:

$R_f = \frac{\text{distance moved by spot}}{\text{distance moved by solvent front}}$

Rf is always between 0 and 1.
Rf is characteristic of a compound under specified conditions (solvent, stationary phase, temperature). Two different solvents will give two different Rf values for the same compound.
Rf has no units — it is a ratio.

Example: A spot moves 3.6 cm while the solvent front moves 7.2 cm. The Rf is 3.6 / 7.2 = 0.50.

Exam Tip: Measure the distance to the centre of the spot, and always measure from the pencil line, not the bottom of the plate. OCR markschemes are strict on this.

2.4 Visualising colourless spots

Many organic compounds are colourless and invisible on a white silica plate. Common ways to visualise them:

UV lamp (254 nm or 365 nm): Most silica plates contain a fluorescent indicator; compounds that absorb UV show up as dark spots on a glowing background.
Iodine vapour: Many compounds adsorb iodine, giving brown spots.
Ninhydrin spray: Turns amino acids purple.
Chemical stains: e.g. KMnO₄ spray turns alkenes brown.

2.5 Uses of TLC

Reaction monitoring: Spot the starting material, the reaction mixture and the pure product side by side. Watch the starting material spot disappear as the reaction proceeds.
Identification: Spot an unknown and a suspected pure compound; if they give the same Rf in two different solvents, they are almost certainly the same compound.
Purity check: A pure compound should give a single spot.
Forensic and medical: Rapid screening of drugs, dyes, amino acids.

3. Gas Chromatography (GC)

3.1 Setup

Stationary phase: A long, thin, coiled capillary column (up to 100 m, wound into a coil) coated on the inside with a high-boiling-point oil (the stationary liquid). The column sits in a temperature-controlled oven.
Mobile phase: An inert carrier gas — usually nitrogen or helium — flowing through the column at a controlled rate.
A tiny volume of the liquid sample (microlitres) is injected into the hot injection port, where it vaporises.
The vapours are swept along the column by the carrier gas.

3.2 What happens

Each component of the sample partitions between:

Lesson 12: Chromatography