Think of this whole chapter as a detective toolkit. You're given an unknown organic compound and you need to identify it. Each technique gives you a different clue:

• High Resolution Mass Spectrometry — tells you the exact molecular formula (to 4 decimal places of mass), letting you rule out "look-alike" formulas that have almost the same mass.
• ¹³C NMR — counts how many different environments exist, and the chemical shift tells you what's attached to each carbon (C=O, C-O, C-Cl, etc).
• Low resolution ¹H NMR — counts how many different environments exist, and the peak area ratio tells you how many H atoms are in each environment.
• High resolution ¹H NMR — zooms in further: the splitting pattern of each peak (singlet, doublet, triplet, quartet…) tells you how many H atoms are on the carbon.
• TLC (Thin-Layer Chromatography) — separates a mixture into spots and uses the R_f value to identify each component by comparing distance travelled.
• HPLC & GC — more powerful, automated versions of chromatography that separate mixtures and use to identify each component, often used in forensics and drug testing.

By the end, you should be able to look at a set of spectra (MS + NMR) and the full structure of an unknown molecule piece by piece — that's the classic exam question in this topic.

At AS level, mass spectrometry gave you the molecular ion peak (M⁺) rounded to the nearest whole number, which tells you the approximate M_r. The problem is that lots of different molecular formulas can add up to almost the same whole-number mass. For example, both C₄H₁₀ and C₃H₆O round to an M_r of "58" if you're only working to whole numbers.

A high resolution mass spectrometer measures mass accurate to 4 decimal places. Because every element's isotope has a very slightly different exact mass (not a nice round number), formulas that look identical at low resolution become distinguishable at high resolution.

Notice carbon is exactly 12.0000 — that's because the entire relative atomic mass scale is against carbon-12. Every other element's mass is ever-so-slightly off a whole number, and that's exactly the "wobble" high resolution MS is designed to detect.

A compound has an accurate M_r of 58.0417. Is it C₄H₁₀, C₂H₆N₂, or C₃H₆O?

Step 1 — Calculate each candidate:

• C₄H₁₀ = (4 × 12.0000) + (10 × 1.0078) = 58.0780
• C₂H₆N₂ = (2 × 12.0000) + (6 × 1.0078) + (2 × 14.0031) = 58.0530
• C₃H₆O = (3 × 12.0000) + (6 × 1.0078) + 15.9949 = 58.0417 ✓

Step 2 — Match: C₃H₆O matches exactly, so that's the molecular formula. Note: this only tells you the formula — it can't tell you whether it's or . You'd need fragmentation data or NMR for that.

Most carbon atoms in nature are carbon-12, which has an even mass number and does show up on NMR. But a small natural fraction of carbon atoms are carbon-13 — an isotope with an odd mass number, which produce a signal.

In ¹³C NMR, we measure the magnetic environment of every carbon-13 nucleus in the molecule, plot it on a spectrum, and compare it all against a standard reference compound: Tetramethylsilane (TMS), which is set at 0 ppm.

Notice the pattern: the more electronegative / double-bonded the neighbours, the further right (higher ppm) the signal appears. Carbonyls (C=O) sit way out at 190–220 ppm because oxygen pulls electron density away hard.

• Displays sharp single signals — no splitting into sub-peaks like ¹H NMR shows.
• The height of each peak is NOT proportional to the number of carbons in that environment — unlike ¹H NMR, you cannot use peak size to count atoms.
• Different chemical environments give different chemical shifts.
• TMS is still the 0 ppm reference.

Propanone is CH₃–CO–CH₃. Both CH₃ groups are chemically identical (the molecule is symmetrical), so there are only 2 molecular environments: the two equivalent CH₃ carbons, and the single C=O carbon.

Result: the ¹³C NMR spectrum shows 2 peaks — one around 30 ppm (CH₃, alkyl) and one around 205 ppm (C=O, ketone).

¹H NMR works on exactly the same principle as ¹³C NMR, but now we're detecting hydrogen nuclei (protons) instead of carbon-13. Again, TMS is the reference at 0 ppm — chosen because it's non-toxic, doesn't react with the sample, has a low boiling point (easy to remove afterwards), and gives one single sharp peak.

The big difference from ¹³C NMR: in ¹H NMR, the area under each peak IS proportional to the number of protons in that environment. This is a huge advantage — it means peak area (measured by an "integration trace") tells you the ratio of H atoms directly.

You'll be given a data sheet with these values in the actual exam — you don't need to memorise exact numbers, but you should recognise the rough order: alkyl CH is furthest left (low ppm), carboxylic acid OH is furthest right (high ppm, most deshielded).

Ethanol, CH₃CH₂OH, has 3 chemical environments: −CH₃, −CH₂, and −OH.

So the low resolution ¹H NMR spectrum shows 3 peaks, at roughly:

• 1.2 ppm → −CH₃ (3 protons)
• 3.7 ppm → −CH₂ (2 protons)
• 5.4 ppm → −OH (1 proton)

The areas of these three peaks are in the ratio 3 : 2 : 1, matching the number of H atoms in each environment. This ratio is exactly what a real spectrum's integration trace would show you.

A low resolution spectrum only tells you exist and are in each. A high resolution spectrum reveals something extra: each peak is actually made of tiny sub-peaks clustered together, called a splitting pattern.

This splitting is caused by a proton's nuclear spin "feeling" the spins of protons on the adjacent (neighbouring) carbon atom. It's like each nearby proton nudges the signal slightly, and the number of nudges tells you exactly how many protons are next door.

Notice the intensity ratios follow Pascal's triangle — a nice pattern-spotting shortcut if you already know it from maths!

Take the same three environments from before and now look at splitting:

• −OH peak: appears as a singlet. This is a special exception — because the OH proton rapidly exchanges with trace water in the sample, it doesn't experience consistent coupling with its neighbours, so it never splits (you just need to know this as a fact).
• −CH₂− peak: the adjacent carbon (CH₃) has 3 protons, so n=3, giving 3+1 = 4 peaks (quartet).
• −CH₃ peak: the adjacent carbon (CH₂) has 2 protons, so n=2, giving 2+1 = 3 peaks (triplet).

This quartet + triplet combination is extremely common — it's the signature of an ethyl group, CH₃CH₂−, and you should learn to spot it instantly.

A complete high resolution spectrum combines three pieces of information for each signal:

1. Chemical shift (δ) — tells you the type of environment (from the data table)
2. Integration / relative peak area — tells you how many H atoms are in that environment
3. Splitting pattern — tells you how many H atoms are on the adjacent carbon

For methyl chloroethanoate, ClCH₂COOCH₃, the integration ratio is 2:3 (2H in CH₂, 3H in CH₃) — but note these two groups aren't adjacent to each other (there's a C=O and O in between), so neither peak splits the other — both would appear as singlets!

Every chromatography technique (TLC, column, HPLC, GC) works on the same idea: a mixture is dissolved in a mobile phase (a fluid that moves), which flows through or over a stationary phase (a fixed material). Different components of the mixture interact with the stationary phase to different extents — some cling on tightly (high retention), some barely interact at all and race ahead with the mobile phase.

1. Prepare a beaker with a small quantity of solvent.
2. On the TLC plate, draw a horizontal baseline near the bottom edge — in pencil, never pen (pen ink would dissolve and interfere with the results).
3. Spot pure reference compounds and the unknown sample mixture onto the baseline, and let them air dry.
4. Stand the plate in the beaker of solvent, making sure the solvent level stays below the spots. Cover with a lid so the solvent atmosphere stays saturated.
5. As the solvent rises up the plate by capillary action, it carries the dissolved components with it at different rates. When the solvent nears the top, remove the plate and immediately mark the solvent front in pencil.
6. If the spots are colourless, visualise them using UV light, iodine vapour, or ninhydrin (a carcinogen, used carefully).

In column chromatography, a vertical tube (like a burette) is packed with an inert solid stationary phase (silica gel or alumina), saturated with solvent. The sample is carefully added to the top without disturbing the packed surface, then more solvent (eluent) is continually added on top to keep pushing everything through.

Gravity pulls everything down through the column — components with the strongest affinity for the stationary phase move the slowest and come out last; components with the weakest affinity flow through fastest and come out first.

HPLC is essentially column chromatography, souped up:

• The sample is pumped through under pressure rather than relying on gravity — much faster.
• The stationary phase particles are much smaller, giving a bigger surface area and therefore sharper, better separation.
• A detector sits at the end of the column and measures retention time — the time from injection to detection.
• The whole process is automated and linked to a computer that can instantly compare results against a database of known compounds.

GLC is used for analysing gases, volatile liquids, and solids that can be vaporised. Key differences from liquid-based methods:

• Stationary phase: a thin film of non-volatile liquid coated inside a long coiled column.
• Mobile phase: an inert carrier gas (e.g. helium or nitrogen) — inert so it doesn't react with the sample.
• The sample is injected through a self-sealing disc and vaporises, then travels through the column carried by the gas.

Just like HPLC, each component reaches the detector at its own characteristic retention time, and the resulting chromatogram shows a series of peaks — one per compound present.

A GLC trace of a compound shows four peaks: A, B, C, D — with D being the tallest and appearing at the longest retention time, and B and C being equal-sized smaller peaks.

• Which compound is present in the greatest quantity? → D (largest peak area = greatest quantity)
• Which compounds are present in equal amounts? → B and C (equal peak sizes)
• Which compound interacts most strongly with the stationary phase? → D (longest retention time = spent the most time "stuck" to the stationary phase)

Common uses: forensic evidence, drug testing in sport, environmental pollution monitoring, detecting explosives in baggage.

The big limitation: HPLC and GC alone cannot give in a legal sense, because:

• Different compounds can coincidentally have the same retention time.
• Conditions like temperature and pressure are hard to keep perfectly controlled, which shifts retention times.
• Some unknown compounds simply have no reference data in the database to compare against.

That's why HPLC and GC are almost always coupled with mass spectrometry — giving HPLC-MS and GC-MS. This lets you separate the mixture AND get a definitive mass-spectrum fingerprint of each component, all in one machine run.