Theory — Mass Spectrometry
1. What does mass spec measure?
A mass spectrometer measures the mass-to-charge ratio (m/z) of charged species. For most organic chemistry, the charge is +1 (singly ionised cations), so m/z is essentially the mass of the ion in atomic mass units (Daltons, Da). The instrument has three parts: an ion source (where neutral molecules are converted to cations), a mass analyser (which separates ions by m/z), and a detector. The output is a spectrum: m/z on the x-axis, relative intensity (0-100%) on the y-axis. The most intense peak is called the base peak (set to 100% by convention).
2. Ionisation methods
The choice of ionisation method affects which ions you see:
| Method | Energy | What you get | Best for |
|---|---|---|---|
| EI (electron ionisation) | 70 eV (high) | Lots of fragments + sometimes a weak molecular ion | Small to medium organic molecules; the "classical" mass spec |
| CI (chemical ionisation) | ~10 eV (soft) | Mainly [M+H]⁺ or [M-H]⁻; little fragmentation | When EI loses the molecular ion; gives MW directly |
| ESI (electrospray) | Very soft | Ions from solution; multiply charged for large molecules | Polar molecules, salts, biomolecules, proteins |
| MALDI | Pulsed laser | Mostly [M+H]⁺ ions | Large molecules: proteins, polymers, lipids |
| APCI | Soft | Volatile molecules from LC eluent | LC-MS for small/medium polar molecules |
Most undergraduate organic chemistry uses EI mass spectra from GC-MS instruments. EI provides rich fragmentation patterns that reveal structure; the rest of this lab focuses on EI.
3. The molecular ion (M⁺)
In EI, the high-energy electron beam knocks one electron out of the molecule, producing the radical cation M⁺· (the molecular ion, with the same mass as the neutral molecule). This ion appears at the highest m/z value in the spectrum and tells you the molecular weight directly. Many EI molecular ions are unstable and fragment quickly; the M⁺ peak may be small or absent.
M⁺· (excited; if stable, observed; if not, fragments to give A⁺ + B·)
The M⁺ peak in the spectrum has m/z = molecular weight of the compound.
4. Isotope peaks
Most elements have several stable isotopes. The most abundant isotope gives the main peak; less abundant isotopes give peaks 1, 2, or more mass units higher (M+1, M+2, etc.).
| Element | Major isotope | Minor isotopes | Pattern |
|---|---|---|---|
| C | ¹²C (98.9%) | ¹³C (1.1%) | ~1.1% per C in M+1 peak |
| H | ¹H (99.99%) | ²H (deuterium, trace) | Negligible |
| N | ¹⁴N (99.6%) | ¹⁵N (0.4%) | Negligible |
| O | ¹⁶O (99.8%) | ¹⁷O (0.04%), ¹⁸O (0.2%) | Small M+2 contribution |
| Cl | ³⁵Cl (75.8%) | ³⁷Cl (24.2%) | 3:1 ratio (M:M+2) — very distinctive |
| Br | ⁷⁹Br (50.7%) | ⁸¹Br (49.3%) | 1:1 ratio (M:M+2) — very distinctive |
| I | ¹²⁷I (100%) | None significant | No isotope pattern |
| S | ³²S (95%) | ³⁴S (4.2%) | ~4% M+2 contribution |
The chlorine and bromine patterns are diagnostic. A 3:1 ratio of peaks separated by 2 mass units = chlorine. A 1:1 ratio of peaks separated by 2 mass units = bromine. These patterns let you identify halogen-containing compounds at a glance.
Two peaks 2 mass units apart, ratio 1:1 → one Br
Three peaks 2 mass units apart, ratio 9:6:1 → two Cl
Three peaks 2 mass units apart, ratio 1:2:1 → two Br
If you see "M and M+2 about equal" in the spectrum, the compound contains Br. If you see "M significantly larger than M+2 in 3:1 ratio", the compound contains Cl.
5. Fragmentation patterns — alpha cleavage
The most common fragmentation in EI is alpha cleavage: cleavage of a C-C bond NEXT to a heteroatom or a π bond. The result: the heteroatom keeps its lone pair on the cation side (so the cation is stabilised by resonance with the heteroatom), and a free radical is lost.
- Ketones and aldehydes: alpha cleavage gives an acylium cation R-C≡O⁺ (m/z 43 for acetyl, m/z 57 for propanoyl, etc.)
- Alcohols: alpha cleavage gives an oxocarbenium R-C(=OH⁺)- which rearranges to lose water (M-18 peak common)
- Amines: alpha cleavage gives an iminium cation R-C(=NHR\')-
- Ethers: similar to amines, gives an oxocarbenium
For acetone (CH₃COCH₃, MW 58), the EI spectrum shows: M⁺ at m/z 58 (small); alpha cleavage gives [CH₃CO]⁺ (acetyl cation, m/z 43, base peak); further loss of CO gives [CH₃]⁺ (m/z 15). The pattern 58 / 43 / 15 with 43 as base peak is classic ketone alpha cleavage.
6. McLafferty rearrangement
For carbonyl compounds with a γ-hydrogen (a hydrogen on the third carbon from the carbonyl), a six-membered cyclic transition state allows transfer of the γ-H to the carbonyl O, with simultaneous cleavage of the α-β bond. This gives an enol cation (preserves the carbonyl ion) and a neutral alkene.
The enol cation has m/z = [original M] - [neutral alkene mass]
Diagnostic for esters, ketones, aldehydes, carboxylic acids with γ-H. The McLafferty fragment is typically a strong peak.
7. Tropylium and other aromatic fragmentations
Aromatic compounds with benzylic CH₂ groups give a very stable cation: the tropylium cation (C⁷H⁷⁺, m/z 91), a 7-membered aromatic ring. Toluene\'s mass spectrum has m/z 91 as the base peak (!) because the initial benzyl cation ([PhCH₂]⁺, m/z 91) rearranges to the tropylium cation (also m/z 91) which is much more stable. Almost any benzyl-type cleavage product gives m/z 91 as a major peak. m/z 77 (phenyl cation, [C₆H₅]⁺) and m/z 65 (cyclopentadienyl cation from tropylium loss of acetylene) are also common.
8. Common fragment ion masses
| m/z | Fragment | Indicator of |
|---|---|---|
| 15 | [CH₃]⁺ | Methyl group present |
| 17 | [OH]⁺ | Hydroxyl loss (alcohols) |
| 18 | [H₂O]⁺ or M-18 | Water loss (alcohols, acids) |
| 28 | [CO]⁺, [N₂]⁺, or [C₂H₄]⁺ | Carbonyl, nitrogen, ethylene |
| 29 | [CHO]⁺ or [C₂H₅]⁺ | Aldehyde C-H of carbonyl, or ethyl |
| 43 | [CH₃CO]⁺ or [C₃H⁷]⁺ | Acetyl cation (ketones), propyl |
| 57 | [C₄H⁹]⁺ or [CH₃CH₂CO]⁺ | Butyl, propanoyl |
| 77 | [C₆H₅]⁺ | Phenyl cation |
| 91 | [C⁷H⁷]⁺ | Tropylium / benzyl — aromatic with -CH₂- |
| 105 | [C₆H₅CO]⁺ | Benzoyl cation (aromatic ketone) |
9. Real vs simulated spectra in this lab
The instrument bench in this lab uses a mix of real reference spectra (annotated as REAL) and simulated spectra (annotated as SIMULATED). Real spectra are computer renderings of published data from the NIST WebBook, SDBS, or commercial databases; simulated spectra are constructed from known fragmentation patterns and isotope ratios. Each spectrum in the lab is labeled at the top so you always know which you\'re looking at. Both modes preserve the diagnostic pattern (M⁺, base peak, isotope pattern) accurately.
10. The systematic MS analysis approach
- Find M⁺ (highest m/z peak). This gives the molecular weight. Check the M+1 and M+2 peaks for isotope clues (especially Cl/Br pattern).
- Identify the base peak (most intense peak). This is the most stable fragment; it tells you what kind of cleavage dominates.
- Calculate mass differences from M⁺. Common losses: 15 (CH₃), 17 (OH), 18 (H₂O), 28 (CO), 29 (CHO), 43 (CH₃CO).
- Check for diagnostic patterns: m/z 91 (tropylium = aromatic with CH₂); m/z 77 (phenyl); halogen 3:1 or 1:1 patterns; McLafferty fragments at predicted positions.
- Combine with IR + NMR + integration formula. MS gives mass; IR gives functional groups; NMR gives structure detail. The three together identify most unknowns.
Instructions
This lab\'s Simulation section has four parts. Complete them in order.
Prerequisite: Familiarity with basic organic functional groups; ideally completion of the Functional Group Tests, Aldehydes & Ketones, and IR Spectroscopy labs first. Mass spec is most useful when combined with IR and NMR for full structural identification.
Simulation
Four interactive parts. Use the ↺ Reset Simulation button at any time to clear all answers and start over.
Eight fragment-identification cases. For each: (a) m/z position type; (b) which fragment ion; (c) the compound class indicated.
Six unknown samples. Select a sample, click Inject & Ionize, then identify the compound class from the spectrum. Fragment labels appear after you answer.
Eight harder puzzles: McLafferty rearrangements, halogen isotope patterns, M⁺ identification, alpha cleavage prediction.
Round 1 — SDS interpretation
Four common MS materials and reagents. Each has 4 questions.
Round 2 — Microscale technique scenarios
Six scenarios. Identify the appropriate MS technique or sample prep.
Team Questions
Discuss with your team before answering.
Example Lab Notebook Entry
Use the format below as a template.
Mass Spectrometry — Lab Notebook Entry
Submitted by: [Student Name]
Course: Organic Chemistry I · Section: 201-A · Date: May 8, 2026
Objective
To learn how electron ionisation (EI) mass spectrometry produces molecular and fragment ions; to identify the molecular ion (M⁺) and base peak; to recognise diagnostic fragmentation patterns including alpha cleavage, McLafferty rearrangement, and tropylium formation; to recognise halogen isotope patterns (Cl 3:1, Br 1:1); to interpret mass spectra of unknown compounds using systematic analysis.
Instrument bench results (Section II)
| Sample | Compound | M⁺ | Base peak | Key diagnostic |
|---|---|---|---|---|
| 1 | Methanol | 32 | 31 | Loss of H from M⁺ (alcohol pattern) |
| 2 | Acetone | 58 | 43 (CH₃CO⁺) | Alpha cleavage of ketone |
| 3 | Hexane | 86 | 57 (C₄H⁹⁺) | Alkane series: 57, 43, 29 (loss of CH₂ units) |
| 4 | 2-Butanone | 72 | 43 (CH₃CO⁺) | Alpha cleavage + McLafferty (m/z 58) |
| 5 | Chlorobenzene | 112/114 (3:1) | 112 (M⁺) | Cl isotope pattern (3:1) |
| 6 | Toluene | 92 | 91 (C⁷H⁷⁺ tropylium) | Tropylium formation from benzyl cation |
Fragmentation summary (key m/z values memorised)
| m/z | Fragment | Indicator |
|---|---|---|
| 15 | [CH₃]⁺ | Methyl loss |
| 18 | M-18 (water) | Alcohol or carboxylic acid |
| 29 | [CHO]⁺ or [C₂H₅]⁺ | Aldehyde C-H or ethyl |
| 43 | [CH₃CO]⁺ | Methyl ketone (alpha cleavage) |
| 57 | [C₄H⁹]⁺ | Butyl, branched alkyl |
| 77 | [C₆H₅]⁺ | Phenyl cation |
| 91 | [C⁷H⁷]⁺ | Tropylium / aromatic with -CH₂- |
| 105 | [C₆H₅CO]⁺ | Aromatic ketone (PhCO⁺) |
Discussion
Mass spectrometry by electron ionisation gives two pieces of information from one spectrum: (1) molecular weight from M⁺, and (2) structural information from the fragmentation pattern. The molecular ion is the highest m/z peak (its abundance varies; some compounds have weak M⁺). Below it, fragment ions appear at lower m/z. The most intense peak is the base peak, normalised to 100%. Other peaks are reported as percentage of the base peak.
Alpha cleavage is the most common fragmentation: a C-C bond next to a heteroatom or pi system breaks, giving a stabilised cation on the heteroatom side. For ketones, this produces an acylium cation (R-C≡O⁺). Acetone (CH₃COCH₃) gives the acetyl cation (CH₃CO⁺) at m/z 43 as the base peak. Higher ketones give characteristic acylium fragments at predictable m/z values: 43 (acetyl), 57 (propanoyl), 71 (butanoyl), etc.
McLafferty rearrangement is a six-membered cyclic transition state that transfers a gamma-hydrogen to a carbonyl oxygen and cleaves the alpha-beta bond. The result is an enol cation (still containing the carbonyl) and a neutral alkene. McLafferty fragments are diagnostic of carbonyls with at least three carbons in the alpha chain. 2-butanone has a McLafferty fragment at m/z 58 (the enol of acetone, CH₂=C(OH)CH₃) plus the standard alpha cleavage at m/z 43.
Halogen isotope patterns are the most useful spectral feature for identifying Cl and Br. Chlorine\'s ³⁵Cl/³⁷Cl ratio is 3:1, so a chlorinated compound shows two peaks separated by 2 mass units in 3:1 ratio. Bromine\'s ⁷⁹Br/⁸¹Br ratio is roughly 1:1, so a brominated compound shows two peaks of nearly equal height separated by 2 mass units. Multiple halogens give characteristic patterns (e.g., two Cl gives three peaks 9:6:1).
Tropylium formation in aromatic compounds with benzylic CH₂ groups gives the diagnostic m/z 91 peak (often the base peak). The initial benzyl cation [PhCH₂]⁺ rearranges to the seven-membered tropylium cation, which is aromatic (6 pi e⁻) and exceptionally stable. Toluene, ethylbenzene, propylbenzene all show m/z 91 as the base peak. Other aromatic fragments include m/z 77 (phenyl cation) and m/z 65 (cyclopentadienyl from tropylium loss of acetylene).
The systematic approach to interpreting an MS: (1) identify M⁺ and check its isotope pattern for Cl/Br; (2) identify the base peak and what mechanism produces it; (3) calculate mass differences from M⁺ to identify common losses (15 = CH₃, 17 = OH, 18 = H₂O, 28 = CO, 43 = CH₃CO); (4) match the pattern to a compound class. Mass spec works best in combination with IR (functional groups) and NMR (connectivity); each gives different information, and the three together identify most undergraduate unknowns.
Conclusion
Mass spectrometry is the most direct way to determine molecular weight and a powerful tool for structural identification through fragmentation patterns. The key skills covered are: identifying M⁺, recognising halogen isotope patterns, predicting alpha cleavage and McLafferty fragments, and matching fragmentation patterns to compound classes. Combined with IR and NMR (which provide complementary functional group and connectivity information), mass spectrometry forms the structural identification toolkit used in research, pharmaceutical analysis, and quality control. The next labs in this Spectroscopy quartet (NMR and UV/Vis) build on the same systematic analysis logic.
References
1. Pavia, D. L.; Lampman, G. M.; Kriz, G. S.; Vyvyan, J. R. Introduction to Spectroscopy, 5th ed., Cengage, 2015, Ch 8.
2. Silverstein, R. M.; Webster, F. X.; Kiemle, D. J. Spectrometric Identification of Organic Compounds, 8th ed., Wiley, 2014, Ch 1.
3. NIST Mass Spectrometry Data Center, https://webbook.nist.gov/chemistry.
4. SDBS — Spectral Database for Organic Compounds, AIST Japan, https://sdbs.db.aist.go.jp.
5. McLafferty, F. W.; Turecek, F. Interpretation of Mass Spectra, 4th ed., University Science Books, 1993.
Practice Questions
Work through each before peeking at the hint.