Common Adducts, Neutral Losses, and PTM Mass Shifts

This appendix collects the mass reference tables used repeatedly in metabolite annotation (Chapters 10–11) and peptide/PTM identification (Chapters 8–9, 14). All values are monoisotopic and given in daltons (Da). Adduct m/z is computed from the neutral monoisotopic mass M of the molecule; the electron mass (0.000549 Da) is accounted for in the ion masses below.

Note

In R, compute these from formulas rather than transcribing constants: MetaboCoreUtils::mass2mz() and adductNames() for small molecules, and PSMatch/MsCoreUtils plus Unimod for peptide modifications. The tables here are for quick lookup and sanity-checking.

Reference constants (monoisotopic, Da)

Quantity	Value
Proton (H⁺)	1.0072765
Electron (e⁻)	0.0005486
Hydrogen atom (¹H)	1.0078250
¹³C − ¹²C spacing	1.0033548

Positive-mode adducts

For a neutral mass M, the ion m/z is given below. Multiply-charged and cluster ions divide/multiply M as shown.

Adduct	Charge	m/z formula	Mass shift (Da)
[M+H]⁺	1+	M + 1.007276	+1.007276
[M+NH₄]⁺	1+	M + 18.033823	+18.033823
[M+Na]⁺	1+	M + 22.989218	+22.989218
[M+K]⁺	1+	M + 38.963158	+38.963158
[M+CH₃OH+H]⁺	1+	M + 33.033489	+33.033489
[M+ACN+H]⁺	1+	M + 42.033823	+42.033823
[M+H−H₂O]⁺	1+	M − 17.003289	−17.003289
[M+H−2H₂O]⁺	1+	M − 35.013854	−35.013854
[M+2H]²⁺	2+	(M + 2.014552)/2	—
[M+3H]³⁺	3+	(M + 3.021828)/3	—
[M+H+Na]²⁺	2+	(M + 23.996494)/2	—
[2M+H]⁺	1+	2M + 1.007276	—
[2M+Na]⁺	1+	2M + 22.989218	—

Negative-mode adducts

Adduct	Charge	m/z formula	Mass shift (Da)
[M−H]⁻	1−	M − 1.007276	−1.007276
[M+Cl]⁻	1−	M + 34.969402	+34.969402
[M+HCOO]⁻ (formate)	1−	M + 44.998201	+44.998201
[M+CH₃COO]⁻ (acetate)	1−	M + 59.013851	+59.013851
[M−H−H₂O]⁻	1−	M − 19.017841	−19.017841
[M+Na−2H]⁻	1−	M + 20.974666	+20.974666
[M+K−2H]⁻	1−	M + 36.948606	+36.948606
[M−2H]²⁻	2−	(M − 2.014552)/2	—
[2M−H]⁻	1−	2M − 1.007276	—

Common neutral losses

Observed as the mass difference between a precursor and a fragment (or between related adducts). Useful for de-adducting and structural interpretation.

Neutral loss	Formula	Mass (Da)	Typical origin
Water	H₂O	18.010565	Alcohols, sugars, general
Ammonia	NH₃	17.026549	Amines, amino acids
Carbon monoxide	CO	27.994915	Carbonyls
Formaldehyde	CH₂O	30.010565	Methylenedioxy, methoxy
Methanol	CH₃OH	32.026215	Methyl esters
Carbon dioxide	CO₂	43.989830	Carboxylic acids
Formic acid	CH₂O₂	46.005479	Carboxylic acids
Acetic acid / ketene	C₂H₄O₂	60.021129	Acetates
Metaphosphate	HPO₃	79.966331	Phosphate esters
Sulfur trioxide	SO₃	79.956815	Sulfates
Phosphoric acid	H₃PO₄	97.976896	Phosphorylation
Pentose (−H₂O)	C₅H₈O₄	132.042259	Glycosides
Hexose (−H₂O)	C₆H₁₀O₅	162.052824	Glycosides
Glucuronic acid (−H₂O)	C₆H₈O₆	176.032088	Phase-II conjugates

Isotope patterns

The natural isotopic envelope helps confirm identity and read charge state.

Isotope pair	Δmass (Da)	Notes
¹³C / ¹²C	1.003355	Spacing of the M+1 peak; intensity ≈ 1.1 % × (number of C)
³⁷Cl / ³⁵Cl	1.997050	M+2 ≈ 32 % of M per Cl
⁸¹Br / ⁷⁹Br	1.997954	M+2 ≈ 97 % of M per Br (near 1:1)
³⁴S / ³²S	1.995796	M+2 ≈ 4.4 % of M per S
¹⁵N / ¹⁴N	0.997035	Relevant for ¹⁵N metabolic labelling

The M+2 abundance from Cl and Br is diagnostic: a strong, roughly 1:1 M/M+2 doublet indicates one bromine; a ~3:1 doublet indicates one chlorine.

Charge-state determination

For a multiply charged ion, adjacent isotope peaks are separated by 1.003355 / z Da in m/z. Therefore:

z = \frac{1.003355}{\Delta(m/z)_{\text{isotopes}}}

Observed isotope spacing (Δ m/z)	Charge state z
1.003	1+
0.502	2+
0.334	3+
0.251	4+
0.201	5+

Given a measured m/z and inferred z, the neutral monoisotopic mass is M = z \times (m/z) - z \times 1.007276 (positive mode).

Peptide post-translational modification (PTM) mass shifts

Monoisotopic mass added to the residue (Unimod values). Chapters 8–9 and 14 use these when auditing PSMs and interpreting modified peptides.

Modification	Composition	Δmass (Da)	Target residue(s)
Carbamidomethyl (fixed, IAA)	C₂H₃NO	+57.021464	C
Oxidation	O	+15.994915	M (also W, P)
Phosphorylation	HPO₃	+79.966331	S, T, Y
Acetylation	C₂H₂O	+42.010565	K, protein N-term
Methylation	CH₂	+14.015650	K, R
Dimethylation	C₂H₄	+28.031300	K, R
Trimethylation	C₃H₆	+42.046950	K
Deamidation	— (N→D, Q→E)	+0.984016	N, Q
GG remnant (ubiquitin/di-glycine)	C₄H₆N₂O₂	+114.042927	K
Pyro-glutamate (from Q)	−NH₃	−17.026549	N-term Q
Pyro-glutamate (from E)	−H₂O	−18.010565	N-term E
Carbamylation	CHNO	+43.005814	K, N-term

Isobaric labelling reporter ions and tag masses

Reporter-ion m/z values (Chapter 14). Each channel’s reporter appears in the low-m/z region of the MS/MS spectrum; the intact tag adds a fixed mass to labelled peptides.

Intact tag masses (added to peptide)

Reagent	Δmass per label (Da)
TMT / TMT-6/10/11-plex	+229.162932
TMTpro (16/18-plex)	+304.207146
iTRAQ 4-plex	+144.102063
iTRAQ 8-plex	+304.205360

TMT reporter-ion m/z (10-/11-plex)

Channel	Reporter m/z	Channel	Reporter m/z
126	126.127726	129N	129.131471
127N	127.124761	129C	129.137790
127C	127.131081	130N	130.134825
128N	128.128116	130C	130.141145
128C	128.134436	131N	131.138180
—	—	131C	131.144500

The 6-Da-apart “N/C” pairs (e.g. 127N vs. 127C) differ by ¹³C vs. ¹⁵N substitution (Δ ≈ 0.00632 Da) and require a high-resolution analyzer (Orbitrap/TOF) to resolve.

iTRAQ reporter-ion m/z

4-plex	8-plex
114, 115, 116, 117	113, 114, 115, 116, 117, 118, 119, 121

Reference

Unimod protein-modification database: https://www.unimod.org/.
MetaboCoreUtils::adducts() and mass2mz() for the authoritative, version-matched adduct definitions used in the book.

# Common Adducts, Neutral Losses, and PTM Mass Shifts {#sec-appendix-adducts .unnumbered} This appendix collects the mass reference tables used repeatedly in metabolite annotation (Chapters 10–11) and peptide/PTM identification (Chapters 8–9, 14). All values are **monoisotopic** and given in daltons (Da). Adduct *m/z* is computed from the neutral monoisotopic mass $M$ of the molecule; the electron mass (0.000549 Da) is accounted for in the ion masses below. ::: {.callout-note} In R, compute these from formulas rather than transcribing constants: `MetaboCoreUtils::mass2mz()` and `adductNames()` for small molecules, and `PSMatch`/`MsCoreUtils` plus Unimod for peptide modifications. The tables here are for quick lookup and sanity-checking. ::: **Reference constants (monoisotopic, Da)** | Quantity | Value | |---|---| | Proton (H⁺) | 1.0072765 | | Electron (e⁻) | 0.0005486 | | Hydrogen atom (¹H) | 1.0078250 | | ¹³C − ¹²C spacing | 1.0033548 | ## Positive-mode adducts For a neutral mass $M$, the ion $m/z$ is given below. Multiply-charged and cluster ions divide/multiply $M$ as shown. | Adduct | Charge | $m/z$ formula | Mass shift (Da) | |---|---|---|---| | [M+H]⁺ | 1+ | $M + 1.007276$ | +1.007276 | | [M+NH₄]⁺ | 1+ | $M + 18.033823$ | +18.033823 | | [M+Na]⁺ | 1+ | $M + 22.989218$ | +22.989218 | | [M+K]⁺ | 1+ | $M + 38.963158$ | +38.963158 | | [M+CH₃OH+H]⁺ | 1+ | $M + 33.033489$ | +33.033489 | | [M+ACN+H]⁺ | 1+ | $M + 42.033823$ | +42.033823 | | [M+H−H₂O]⁺ | 1+ | $M − 17.003289$ | −17.003289 | | [M+H−2H₂O]⁺ | 1+ | $M − 35.013854$ | −35.013854 | | [M+2H]²⁺ | 2+ | $(M + 2.014552)/2$ | — | | [M+3H]³⁺ | 3+ | $(M + 3.021828)/3$ | — | | [M+H+Na]²⁺ | 2+ | $(M + 23.996494)/2$ | — | | [2M+H]⁺ | 1+ | $2M + 1.007276$ | — | | [2M+Na]⁺ | 1+ | $2M + 22.989218$ | — | ## Negative-mode adducts | Adduct | Charge | $m/z$ formula | Mass shift (Da) | |---|---|---|---| | [M−H]⁻ | 1− | $M − 1.007276$ | −1.007276 | | [M+Cl]⁻ | 1− | $M + 34.969402$ | +34.969402 | | [M+HCOO]⁻ (formate) | 1− | $M + 44.998201$ | +44.998201 | | [M+CH₃COO]⁻ (acetate) | 1− | $M + 59.013851$ | +59.013851 | | [M−H−H₂O]⁻ | 1− | $M − 19.017841$ | −19.017841 | | [M+Na−2H]⁻ | 1− | $M + 20.974666$ | +20.974666 | | [M+K−2H]⁻ | 1− | $M + 36.948606$ | +36.948606 | | [M−2H]²⁻ | 2− | $(M − 2.014552)/2$ | — | | [2M−H]⁻ | 1− | $2M − 1.007276$ | — | ## Common neutral losses Observed as the mass difference between a precursor and a fragment (or between related adducts). Useful for de-adducting and structural interpretation. | Neutral loss | Formula | Mass (Da) | Typical origin | |---|---|---|---| | Water | H₂O | 18.010565 | Alcohols, sugars, general | | Ammonia | NH₃ | 17.026549 | Amines, amino acids | | Carbon monoxide | CO | 27.994915 | Carbonyls | | Formaldehyde | CH₂O | 30.010565 | Methylenedioxy, methoxy | | Methanol | CH₃OH | 32.026215 | Methyl esters | | Carbon dioxide | CO₂ | 43.989830 | Carboxylic acids | | Formic acid | CH₂O₂ | 46.005479 | Carboxylic acids | | Acetic acid / ketene | C₂H₄O₂ | 60.021129 | Acetates | | Metaphosphate | HPO₃ | 79.966331 | Phosphate esters | | Sulfur trioxide | SO₃ | 79.956815 | Sulfates | | Phosphoric acid | H₃PO₄ | 97.976896 | Phosphorylation | | Pentose (−H₂O) | C₅H₈O₄ | 132.042259 | Glycosides | | Hexose (−H₂O) | C₆H₁₀O₅ | 162.052824 | Glycosides | | Glucuronic acid (−H₂O) | C₆H₈O₆ | 176.032088 | Phase-II conjugates | ## Isotope patterns The natural isotopic envelope helps confirm identity and read charge state. | Isotope pair | Δmass (Da) | Notes | |---|---|---| | ¹³C / ¹²C | 1.003355 | Spacing of the M+1 peak; intensity ≈ 1.1 % × (number of C) | | ³⁷Cl / ³⁵Cl | 1.997050 | M+2 ≈ 32 % of M per Cl | | ⁸¹Br / ⁷⁹Br | 1.997954 | M+2 ≈ 97 % of M per Br (near 1:1) | | ³⁴S / ³²S | 1.995796 | M+2 ≈ 4.4 % of M per S | | ¹⁵N / ¹⁴N | 0.997035 | Relevant for ¹⁵N metabolic labelling | The M+2 abundance from Cl and Br is diagnostic: a strong, roughly 1:1 M/M+2 doublet indicates one bromine; a ~3:1 doublet indicates one chlorine. ## Charge-state determination For a multiply charged ion, adjacent isotope peaks are separated by $1.003355 / z$ Da in $m/z$. Therefore: $$ z = \frac{1.003355}{\Delta(m/z)_{\text{isotopes}}} $$ | Observed isotope spacing (Δ m/z) | Charge state z | |---|---| | 1.003 | 1+ | | 0.502 | 2+ | | 0.334 | 3+ | | 0.251 | 4+ | | 0.201 | 5+ | Given a measured $m/z$ and inferred $z$, the neutral monoisotopic mass is $M = z \times (m/z) - z \times 1.007276$ (positive mode). ## Peptide post-translational modification (PTM) mass shifts Monoisotopic mass added to the residue (Unimod values). Chapters 8–9 and 14 use these when auditing PSMs and interpreting modified peptides. | Modification | Composition | Δmass (Da) | Target residue(s) | |---|---|---|---| | **Carbamidomethyl** (fixed, IAA) | C₂H₃NO | +57.021464 | C | | **Oxidation** | O | +15.994915 | M (also W, P) | | **Phosphorylation** | HPO₃ | +79.966331 | S, T, Y | | **Acetylation** | C₂H₂O | +42.010565 | K, protein N-term | | **Methylation** | CH₂ | +14.015650 | K, R | | **Dimethylation** | C₂H₄ | +28.031300 | K, R | | **Trimethylation** | C₃H₆ | +42.046950 | K | | **Deamidation** | — (N→D, Q→E) | +0.984016 | N, Q | | **GG remnant** (ubiquitin/di-glycine) | C₄H₆N₂O₂ | +114.042927 | K | | **Pyro-glutamate (from Q)** | −NH₃ | −17.026549 | N-term Q | | **Pyro-glutamate (from E)** | −H₂O | −18.010565 | N-term E | | **Carbamylation** | CHNO | +43.005814 | K, N-term | ## Isobaric labelling reporter ions and tag masses Reporter-ion *m/z* values (Chapter 14). Each channel's reporter appears in the low-*m/z* region of the MS/MS spectrum; the intact tag adds a fixed mass to labelled peptides. **Intact tag masses (added to peptide)** | Reagent | Δmass per label (Da) | |---|---| | TMT / TMT-6/10/11-plex | +229.162932 | | TMTpro (16/18-plex) | +304.207146 | | iTRAQ 4-plex | +144.102063 | | iTRAQ 8-plex | +304.205360 | **TMT reporter-ion m/z (10-/11-plex)** | Channel | Reporter m/z | Channel | Reporter m/z | |---|---|---|---| | 126 | 126.127726 | 129N | 129.131471 | | 127N | 127.124761 | 129C | 129.137790 | | 127C | 127.131081 | 130N | 130.134825 | | 128N | 128.128116 | 130C | 130.141145 | | 128C | 128.134436 | 131N | 131.138180 | | — | — | 131C | 131.144500 | The 6-Da-apart "N/C" pairs (e.g. 127N vs. 127C) differ by ¹³C vs. ¹⁵N substitution (Δ ≈ 0.00632 Da) and require a high-resolution analyzer (Orbitrap/TOF) to resolve. **iTRAQ reporter-ion m/z** | 4-plex | 8-plex | |---|---| | 114, 115, 116, 117 | 113, 114, 115, 116, 117, 118, 119, 121 | ## Reference - Unimod protein-modification database: <https://www.unimod.org/>. - `MetaboCoreUtils::adducts()` and `mass2mz()` for the authoritative, version-matched adduct definitions used in the book.