Reading a mass spectrum: fragments and isotopes

A mass spectrum is a plot of ion abundance versus mass-to-charge ratio (m/z). For an organic molecule ionised in electron-impact (EI) mode, every peak is a positive ion the spectrometer detected — the intact molecular ion at highest mass, and a spray of fragments at lower masses. Three questions turn this bar chart into a structure: what is the molecular mass, which elements are present, and how did the molecule break apart? This guide answers all three, in order.

Prerequisites: molecular formulas and functional groups. If you can compute a molar mass from the molar-mass calculator, you can read a mass spectrum.

The three parts of a mass spectrum

Every EI mass spectrum has three tiers of information:

1. Molecular ion (M⁺). The peak at highest m/z that is not an isotope satellite. Its m/z equals the molecular mass (integer, using dominant isotopes: ¹²C, ¹H, ¹⁶O, etc.). Sometimes M⁺ is weak or absent — a common failure mode discussed below.

2. Fragment ions. Lower-mass peaks produced when the molecular cation loses neutral fragments. Each fragment ion carries information about a bond that broke.

3. Isotope satellites (M+1, M+2, sometimes M+4). Small peaks at m/z one, two, or four higher than M. Their intensities relative to M encode the elemental composition.

The base peak is the tallest peak (defined as 100% intensity); every other peak is reported as a percentage of the base. The base peak is often not M⁺ — it is usually the most stable fragment cation.

Molecular formula from M+1 and M+2

The isotope satellites are where you get the elemental composition, because different elements have different characteristic isotope distributions:

Carbon. ¹³C is 1.08% of natural carbon (IUPAC 2021). Every carbon in a molecule contributes 1.08% to M+1 relative to M. If M+1 is 6.5% of M, the molecule has about 6 carbons.

Hydrogen. Deuterium is 0.015% — negligible for most spectra unless the sample is labelled.

Nitrogen has one dominant isotope (¹⁴N, 99.6%), so its M+1 contribution is tiny. But nitrogen’s nominal mass is odd (14), which matters:

Nitrogen rule. A molecule with an odd number of N atoms has an odd nominal molecular mass; a molecule with zero or an even number of N atoms has an even nominal molecular mass. If M⁺ is odd, count on an odd number of nitrogens.

Chlorine. ³⁵Cl : ³⁷Cl = 3.13 : 1 (75.8% : 24.2%). One Cl gives M+2 at 32% of M. Two Cls give M and M+2 in about 3:2 with M+4 present. Three Cls give the famous 27:27:9:1 pattern.

Bromine. ⁷⁹Br : ⁸¹Br = 1.03 : 1. One Br gives M and M+2 at nearly equal intensity — the “1:1 doublet” fingerprint.

Sulfur. ³²S : ³³S : ³⁴S ≈ 95 : 0.75 : 4.2. One S adds ~4% to M+2.

If M+2/M is roughly 4%, suspect one sulfur. If it is ~32%, one chlorine. If it is ~100%, one bromine.

Worked example — identifying an unknown

The workflow is fragile in isolation. Combined with a fragmentation pattern, one can usually distinguish isomers (2-chloropropane vs 1-chloropropane) by which cleavages are preferred.

Fragmentation — how molecules break

When a molecule loses an electron in the EI source (70 eV, the standard energy), it becomes a radical cation with substantial excess energy. The excess energy is released by breaking a bond, and the bond most likely to break is the one that produces the most stable products. Three principles govern fragmentation:

1. Stevenson’s rule. In a homolytic cleavage that produces a cation and a radical, the charge tends to stay on the fragment with the lower ionisation energy — i.e., the fragment that is easier to ionise remains as the cation.

2. Stability of the cation. Tertiary carbocations beat secondary beat primary. Allyl and benzyl cations are especially stable. Acylium ions (R-C≡O⁺) are stable. So are oxocarbenium (R₂C=O-R’⁺) and iminium (R₂C=NR₂⁺) ions.

3. Rearrangements happen. The McLafferty rearrangement (γ-H transfer to a carbonyl via a 6-membered transition state, loss of a neutral alkene) is the most famous. It gives a diagnostic peak at m/z 58 for methyl ketones, 60 for carboxylic acids and esters.

Diagnostic fragment losses

Certain neutral losses are so common they are worth memorising:

M − 15 = loss of methyl. M − 18 = loss of water. Seeing both immediately suggests an alcohol. M − 43 with an intense peak at 43 itself suggests an acetyl fragment (CH₃CO⁺, m/z 43) — very common for methyl ketones.

Worked example — reading a ketone spectrum

The McLafferty rearrangement

Aldehydes, ketones, carboxylic acids, and esters with a γ-hydrogen undergo a specific 6-membered-transition-state rearrangement: the γ-H transfers to the carbonyl oxygen while a β-γ C-C bond breaks, ejecting a neutral alkene and leaving an enol-type radical cation.

For a methyl ketone RCH₂CH₂CH₂C(O)CH₃ with γ-H, the rearrangement gives a peak at m/z 58 — the enol tautomer of acetone (CH₃-C(OH)=CH₂⁺·). Any methyl ketone with a chain of ≥3 carbons attached shows this diagnostic 58 peak.

Carboxylic acids and methyl esters undergo the analogous rearrangement giving m/z 60 (acid) or m/z 74 (methyl ester). These “even-mass McLafferty peaks” are so reliable they are the second-fastest way to spot the functional-group class (after M − 17 / M − 45 for acids).

When the molecular ion is missing

Some molecules never show M⁺:

• Highly branched alkanes — the M⁺ fragments too readily.
• Alcohols — often M − H₂O (M − 18) is the highest visible peak, with true M⁺ under 1%.
• Amines — cleave α to nitrogen very efficiently.
• Halides — often the highest peak is M − X.

Chemical ionisation (CI), a softer ionisation method that produces [M+H]⁺ or [M+NH₄]⁺ adducts with far less excess energy, is the standard fix. Modern electrospray (ESI) and MALDI ionisation give [M+H]⁺, [M+Na]⁺, and other soft adducts that let you determine molecular mass even for macromolecules like proteins.

Common pitfalls

Confusing M+1 with the molecular ion. M+1 is the ¹³C-containing isotopologue of M, not a separate molecule. Do not treat it as a fragment.

Ignoring the nitrogen rule. An odd M⁺ mass with no nitrogens is impossible for a hydrocarbon. Recount your carbons and hydrogens; you have made an arithmetic error.

Missing the Br pattern. Two peaks nearly equal in intensity separated by 2 mass units is bromine. If you see a 1:1 doublet in the middle of a spectrum, one of the fragments contains Br.

Assigning a peak based on m/z alone without checking degrees of unsaturation. A peak at m/z 77 could be C₆H₅⁺ (phenyl, from a benzene derivative) or C₃H₅N₂⁺. Cross-check with the degrees of unsaturation calculator to see which makes chemical sense given the M⁺.

Reading high-resolution vs low-resolution as interchangeable. Nominal-mass MS gives integer m/z. High-resolution MS gives four-decimal m/z, enough to distinguish C₃H₇NO (m/z 73.0528) from C₃H₉N₂ (73.0766). Only HR-MS can determine a molecular formula unambiguously.

Practice

Where this leads

Mass spectrometry is the linchpin of modern analytical chemistry:

• Coupled techniques — GC-MS for volatile compounds, LC-MS for polar or thermally labile molecules, HPLC with MS as the detector.
• High-resolution MS — determines molecular formula from mass alone to 5 ppm accuracy.
• Tandem MS (MS/MS) — selecting one ion, fragmenting it further, mapping structure fragment by fragment. The workhorse of proteomics and metabolomics.
• Soft-ionisation methods — ESI, MALDI, and DART give [M+H]⁺ of intact proteins up to hundreds of kDa.

Every one of them starts by asking three questions: what is the mass, what elements are there, and how did it break.