Table of Contents

Cover

Title Page

Preface

References

Abbreviations

Chapter 1: Introduction To LC–MS Technology

1.1 Introduction
1.2 Analyte Ionization: Ion Sources
1.3 Mass Spectrometer Building Blocks
1.4 Tandem Mass Spectrometry
1.5 Data Acquisition
1.6 Selected Literature on Mass Spectrometry
References

Chapter 2: Interpretation of Mass Spectra

2.1 Mass Spectrometry: A Nuclear Affair
2.2 Isomers, Isotones, Isobars, Isotopes
2.3 Masses in MS
2.4 Isotopes and Structure Elucidation
2.5 Nitrogen Rule, Ring Double-Bond Equivalent, and Hydrogen Rule
2.6 Resolving Power, Resolution, Accuracy
2.7 Calculating Elemental Composition from Accurate
2.8 Protonated and Deprotonated Molecules and Adduct Ions
References

Chapter 3: Fragmentation of Even-Electron Ions

3.1 Introduction
3.2 Analyte Ionization Revisited
3.3 Fragmentation of Odd-electron Ions
3.4 High-energy Collisions of Protonated Molecules
3.5 Fragmentation of Protonated Molecules
3.6 Characteristic Positive-ion Fragmentation of Functional Groups
3.7 Fragmentation of Deprotonated Molecules
3.8 Fragmentation of Metal-ion Cationized Molecules
3.9 Generation of Odd-electron Ions in ESI-MS, APCI-MS, and APPI-MS
3.10 Useful Tables
References

Chapter 4: Fragmentation of Drugs and Pesticides

4.1 Fragmentation of Drugs for Cardiovascular Diseases and Hypertension
References
4.2 Fragmentation of Psychotropic or Psychoactive Drugs
References
4.3 Fragmentation of Analgesic, Antipyretic, and Anti-Inflammatory Drugs
References
4.4 Fragmentation of Drugs Related to Digestion and the Gastrointestinal Tract
References
4.5 Fragmentation of Other Classes of Drugs
References
4.6 Fragmentation of Steroids
References
4.7 Fragmentation of Drugs of Abuse
References
4.8 Fragmentation of Antimicrobial Compounds
References
4.9 Fragmentation of Antimycotic and Antifungal Compounds
References
4.10 Fragmentation of Other Antibiotic Compounds
References
4.11 Pesticides
References

Chapter 5: Identification Strategies

5.1 Introduction
5.2 Confirmation of Identity in Following Organic Synthesis
5.3 Confirmation of Identity in Targeted Screening by SRM-based Strategies
5.4 Confirmation of Identity by High-resolution Accurate-mass MS Strategies
5.5 Library Searching Strategies in Systematic Toxicological Analysis
5.6 Dereplication and Identification of Natural Products and Endogenous Compounds
5.7 Identification of Structure-related Substances
5.8 Identification of Known Unknowns and Real Unknowns
References

Compound Index

Subject Index

End User License Agreement

List of Illustrations

Chapter 1: Introduction To LC–MS Technology

Figure 1.1 Schematic diagram of an electron ionization (EI) source.
Figure 1.2 Scheme for the generation of ionizing electrons in an EI source.
Figure 1.3 Relationship between ion current and electron energy.
Figure 1.4 Ground electronic state of a neutral homodiatomic molecule (a). Vertical transitions depicting the ionization process in an EI source (b).
Figure 1.5 Devices for ion transmission from the EI ion source to the mass analyzer.
Figure 1.6 Ongoing processes inside a chemical ionization (CI) source during reagent gas ionization (methane) in a CI experiment (a). Main chemical reactions involved in the ionization of methane reagent gas in an EI source during a CI experiment (b).
Figure 1.7 Approximate range of molecular mass and polarity for the most common ionization sources in MS.
Figure 1.8 Schematic representation of an atmospheric-pressure ionization (API) source with an electrospray ionization (ESI) inlet.
Figure 1.9 (a) Electrospray setup in a positive-ion ESI-MS experiment. (b) Positively charged meniscus formation at the electrospray emitter tip.
Figure 1.10 Taylor cone formation and charged aerosol generation in an ESI experiment.
Figure 1.11 Analyte ion formation illustrating evaporation-coulombic fission events in positive-ion ESI-MS.
Figure 1.12 Schematic representation of an atmospheric-pressure ionization (API) source with an atmospheric-pressure chemical ionization (APCI) inlet.
Figure 1.13 Schematic representation of the solute ionization process in positive-ion APCI-MS.
Figure 1.14 Schematic representation of analyte evaporation and ionization in a MALDI-MS experiment.
Figure 1.15 Flow chart of a typical MS experimental setup.
Figure 1.16 GC–MS analysis of a mixture of organochlorine insecticides. (a) Total-ion chromatogram (TIC), (b) profile or continuum mass spectrum for isodrin, and (c) centroid mass spectrum for isodrin.
Figure 1.17 Different graphical representations of the chromatography–MS data. (a) Conventional 2D TIC, (b) zoom-in showing individual spectra as a function of time, (c) 3D TIC, and (d) contour plot of the 3D TIC shown in (c).
Figure 1.18 Schematic diagram of an electron multiplier for ion detection.
Figure 1.19 Schematic diagram of a quadrupole mass analyzer.
Figure 1.20 Stability diagram for a quadrupole mass filter.
Figure 1.21 Schematic diagram of an ion-trap mass analyzer.
Figure 1.22 Principle of time-of-flight mass spectrometry.
Figure 1.23 Operation principle of kinetic-energy focusing in a reflectron time-of-flight mass spectrometer.
Figure 1.24 Schematic diagram on a linear-ion-trap–orbitrap hybrid instrument.
Figure 1.25 Schematic representation of selected-reaction monitoring (SRM) in a tandem-quadrupole instrument.

Chapter 2: Interpretation of Mass Spectra

Figure 2.1 Formation of ¹⁴C and its incorporation into the carbon cycle.
Figure 2.2 The electron ionization (EI) mass spectrum of phenyl benzoate.
Figure 2.3 The EI mass spectrum of CH₃Br showing isotopic peaks due to ¹²CH₃⁺ (m/z 15):¹³CH₃⁺ (m/z 16) and CH₃⁷⁹Br^+• (m/z 94):CH₃⁸¹Br^+• (m/z 96) with a relative intensity of 1:0.011 and 1:0.97, respectively.
Figure 2.4 Combined effect of isotopes on the relative intensity of isotope containing ions.
Figure 2.5 Expected isotopic patterns for ions containing Cl + Br atoms.
Figure 2.6 Parameters for the characterization of resolving power and resolution.
Figure 2.7 Effect of resolution on the identification of 28 Da isobars.
Figure 2.8 Relationship between peak width and resolution. The isotope patterns are simulated for the [M+3H]³⁺ of the 1–24 fragment of follicle-stimulating hormone (MDYYRKYAAI FLVTLSVFLH VLHS) with m/z 962.9.
Figure 2.9 Variation of the number of possible elemental formulae as the mass accuracy changes.
Figure 2.10 H⁺/Na⁺ exchange by the acid functional groups of adenosine triphosphate in positive-ion and negative-ion ESI mass spectra.

Chapter 3: Fragmentation of Even-Electron Ions

Figure 3.1 Mechanism of the α-cleavage in OE^+•, illustrated for amphetamine (M^+• with m/z 135) and its trifluoroacetyl derivative (M^+• with m/z 231).
Figure 3.2 Mechanism of the McLafferty rearrangement. For the trifluoroacetyl derivative of amphetamine, the McLafferty rearrangement results in an ion with m/z 118 (Figure 3.1).
Figure 3.3 Mechanism of the retro-Diels–Alder fragmentation for OE^+•, illustrated for 4-methyl-cyclohexene (M^+• with m/z 96) and 4-phenyl-cyclohexene (M^+• with m/z 158).
Figure 3.4 Proposed mechanisms for EE⁺ fragmentation of a tertiary amine, with (a) the inductive cleavage, (b) the concerted four-center fragmentation mechanism for the acyclic EE⁺ cleavage involving βH-rearrangement, and (c) the fragmentation mechanism of the acyclic EE⁺ cleavage via an ion–molecule complex.
Figure 3.5 Product-ion mass spectra for eight protonated C₅-dialkyl- and trialkylamines.
Figure 3.6 Two fragmentation mechanisms for an alkene loss from alkyl iminium ions via a McLafferty-type γH-rearrangement. As the top mechanism involving homolytic cleavages (Djerassi & Fenselau, 1965) is a violation of the even-electron rule, an alternative mechanism involving heterolytic cleavages is shown at the bottom.
Figure 3.7 Fragmentation of [M+H]⁺ of a peptide, with (a) cleavage of the peptide bond to generate either an acylium ion or a (shorter) protonated peptide, and (b) simplified mechanism, assuming an inductive cleavage to generate an acylium ion and an acyclic EE⁺ cleavage (H-rearrangement) to generate the (shorter) protonated peptide.
Figure 3.8 Nomenclature of peptide sequence ions. The N-terminal sequence ions a, b, and c and the C-terminal ions x, y, and z are due to backbone cleavages. Substituent-chain cleavage to ions d, v, and w only occurs in high-energy collisions and with specific amino acids, which helps to determine the identity of the peptide.
Figure 3.9 Characteristic EE⁺ fragmentation of esters, secondary amines, ethers, and amides. For ethers and secondary amines, only the results from the cleavages of the CO and CN bonds, respectively, of the heteroatomR²-moiety are shown; the results from the cleavage in the heteroatomR¹-moiety are not shown. For each compound class, the top reaction involves a four-center H-rearrangement (acyclic EE⁺ cleavage) and the bottom reaction involves an inductive cleavage.
Figure 3.10 Product-ion mass spectrum of bromhexine ([M+H]⁺ with m/z 375; the ion of the isotopologue with ⁷⁹Br,⁸¹Br with m/z 377 was selected as precursor ion.
Figure 3.11 Product-ion mass spectra of (a) n-butyl 4-hydroxybenzoate, (b) meperidine, and (c) anisodamine.
Figure 3.12 Mass spectra of the triazine herbicide propazine: (a) EI-MS spectrum, and (b) ESI-MS–MS spectrum.
Figure 3.13 Fragmentation mechanisms for p-hydroxycinnamoyl putrescine (N-(4-aminobutyl)-3-(4-hydroxyphenyl)prop-2-enamide), with (a) general mechanism according to the four-center cleavage with H-rearrangement and (b) proposed mechanism involving neighboring-group participation.
Figure 3.14 Fragmentation of [M+H]⁺ of peptides via an oxazolone intermediate, generating a protonated oxazolone and a (shorter) protonated peptide. Illustration of mobile-proton hypothesis.
Figure 3.15 The retro-Diels–Alder (RDA) fragmentation illustrated for flavonoids, with (a) the nomenclature system for flavonoid fragment ions, (b) RDA fragmentation in flavonoids, resulting in two complementary fragment ions, ^1,3A and ^1,3B, and (c) RDA fragmentation illustrated for two isomeric dimethoxy-flavan-3-ols, that is, 5,7-dimethoxy-catechin and 3′,4′-dimethoxy-catechin ([M+H]⁺ with m/z 319).
Figure 3.16 Fragmentation of 3-nitro-tyrosine.
Figure 3.17 ortho-Effect in the fragmentation of (a) protonated 2-acetamido-5-aminobenzenesulfonic acid and (b) protonated 5-acetamido-2-aminobenzenesulfonic acid.
Figure 3.18 Fragmentation of deprotonated dimethyl succinate, resulting in two pairs of CO ions from direct cleavages.
Figure 3.19 Two fragmentation pathways in high-energy CID of ethane-1,2-diyl diacetate, with (a) ion formation of a direct-cleavage pair of ions via an ion–molecule complex and (b) additional fragment ions after a Claisen-type rearrangement.
Figure 3.20 Fragmentation pathways for deprotonated amides, illustrated for xipamide ([M−H]⁻ with m/z 353) and lornoxicam ([M−H]⁻ with m/z 370).
Figure 3.21 Fragmentation scheme of [M−H]⁻ of a peptide.
Figure 3.22 Comparison of the fragmentation of the iridoid glycoside globularin using (a) the ESI-generated [M+H]⁺ as precursor ion and (b) the ESI-generated [M+Na]⁺ as precursor ion.
Figure 3.23 Proposed mechanism for the loss of the C-terminal amino-acid residue from [M+Na]⁺ of a peptide.
Figure 3.24 Fragmentation of carotenes: (a) retro-Diels–Alder fragmentation of the ionone ring of the ESI-generated M^+• of α-carotene, showing the loss of C₄H₈, (b) Edmunds–Johnstone fragmentation mechanism for M^+•, leading to the loss of C₆H₅CH₃, and (c) various fragment ions observed from [M+H]⁺ of β-carotene (* indicates positions of [¹³C]-labels in [¹³C₆]-β-carotene).

Chapter 4: Fragmentation of Drugs and Pesticides

Figure 4.1.1 Typical structures of β-blockers.
Figure 4.1.2 Fragmentation schemes for protonated propranolol and timolol.
Figure 4.1.3 Fragmentation schemes for the deprotonated β-blockers acebutolol, carvedilol, and celiprolol.
Figure 4.1.4 Proposed structures for the fragment ions of protonated nimodipine.
Figure 4.1.5 Proposed structures and fragmentation mechanisms for the formation of the ions with m/z 294 and 238 of protonated amlodipine.
Figure 4.1.6 Proposed structures for the fragment ions of deprotonated nitrendipine.
Figure 4.1.7 Fragmentation schemes for protonated and deprotonated captopril.
Figure 4.1.8 Fragmentation scheme for protonated fosinopril.
Figure 4.1.9 General fragmentation pathways for deprotonated thiazide diuretic drugs.
Figure 4.1.10 Fragmentation scheme for protonated buthiazide.
Figure 4.1.12 Fragmentation schemes for deprotonated and protonated xipamide.
Figure 4.1.13 Fragmentation scheme for protonated torasemide.
Figure 4.1.14 Fragmentation scheme for protonated amiloride.
Figure 4.1.15 Structures and characteristic class-specific fragment ions for protonated angiotensin II receptor antagonists (sartans).
Figure 4.1.16 Proposed structures for the fragment ions of the deprotonated angiotensin II receptor antagonist losartan.
Figure 4.1.17 Fragmentation schemes for protonated terazosin and tamsulosin.
Figure 4.1.18 Fragmentation scheme for protonated urapidil.
Figure 4.1.19 Fragmentation schemes for protonated butalamine and ajmalicine.
Figure 4.1.20 Fragmentation pathways for protonated bosentan.
Figure 4.1.21 Fragmentation schemes for protonated and deprotonated sildenafil.
Figure 4.1.22 Fragmentation scheme for protonated flecainide.
Figure 4.1.23 Fragmentation schemes for protonated amiodarone and benziodarone.
Figure 4.1.24 Fragmentation schemes for protonated verapamil and diltiazim.
Figure 4.2.1 Fragmentation schemes for protonated perazine and promazine.
Figure 4.2.2 Fragmentation schemes for protonated risperiodone, pyritinol, flupentixol, sulpiride, and tiapride.
Figure 4.2.3 Fragmentation schemes and proposed structures for some of the fragment ions of protonated haloperidol and pipamperone.
Figure 4.2.4 Fragmentation scheme for protonated clozapine.
Figure 4.2.5 Fragmentation scheme and proposed structures for some fragment ions of protonated amitriptyline and protriptyline.
Figure 4.2.6 Proposed structures for some of the fragment ions of protonated mirtazepine.
Figure 4.2.7 Fragmentation schemes and proposed structures for some of the fragment ions of protonated citalopram and paroxetine.
Figure 4.2.8 Fragmentation schemes and proposed structures for some of the fragment ions of protonated moclobemide and venlafaxine. For venlafaxine, both ion-trap MSⁿ fragment ions and TQ MS–MS fragment ions are shown.
Figure 4.2.9 Fragmentation pathways for protonated diazepam.
Figure 4.2.10 Fragmentation pathways for protonated nordiazepam.
Figure 4.2.11 Fragmentation pathways for protonated oxazepam. The ion with m/z 194 is not observed for oxazepam, but it is observed for its analogue compounds (Section 4.2.2).
Figure 4.2.12 Proposed structures for the fragment ions of protonated alprazolam.
Figure 4.2.13 Fragmentation schemes for protonated lidocaine and procaine.
Figure 4.2.14 Proposed structures for the fragment ions of deprotonated amobarbital.
Figure 4.2.15 Proposed structures for the fragment ions of protonated phenytoin and sultiame.
Figure 4.2.16 Proposed structures for the fragment ions of protonated primidone and mesuximide.
Figure 4.2.17 Fragmentation scheme for protonated zopiclone.
Figure 4.3.1 Fragmentation scheme for protonated acetaminophen (paracetamol).
Figure 4.3.2 Fragmentation schemes for protonated and deprotonated acetylsalicylic acid (aspirin).
Figure 4.3.3 Proposed structures for some of the fragment ions of deprotonated olsalazine.
Figure 4.3.4 Fragmentation schemes for deprotonated mefenamic acid and for protonated and deprotonated glafenine.
Figure 4.3.5 Proposed structures for the fragment ions of protonated and deprotonated phenylbutazone.
Figure 4.3.6 Fragmentation schemes for protonated piroxicam and deprotonated lornoxicam.
Figure 4.3.7 Fragmentation schemes and proposed structures for some of the fragment ions of protonated and deprotonated valdecoxib. The framed structure shows the interpretation of the negative-ion product-ion mass spectrum proposed elsewhere (Zhang et al., 2003).
Figure 4.4.1 Fragmentation schemes for protonated and deprotonated carbutamide.
Figure 4.4.2 Fragmentation schemes for protonated glyburide and deprotonated glimepiride.
Figure 4.4.3 Fragmentation schemes for protonated metformin, nateglinide, and rosiglitazone.
Figure 4.4.4 Fragmentation schemes for protonated omeprazole, cimetidine, ranitidine, and famotidine.
Figure 4.4.5 Fragmentation schemes for protonated and deprotonated bezafibrate and for protonated fenofibrate.
Figure 4.4.6 Fragmentation scheme for deprotonated etiroxate.
Figure 4.4.7 Proposed structures for the fragment ions of protonated simvastatin and deprotonated simvastatin dihydroxy acid.
Figure 4.4.8 Fragmentation schemes for protonated atorvastatin and deprotonated fluvastatin.
Figure 4.4.9 Fragmentation schemes for protonated and deprotonated dolasetron.
Figure 4.4.10 Fragmentation scheme for deprotonated domperidone.
Figure 4.4.11 Fragmentation scheme for deprotonated metoclapramide.
Figure 4.5.1 Predicted fragmentation scheme for protonated fenoterol. The fragment ions related to the indicated cleavage, that is, α-CN bond relative to the hydroxy-substituted C-atom, are not observed.
Figure 4.5.2 Fragmentation schemes for protonated pheniramine, clemastine, and cinnarizine.
Figure 4.5.3 Fragmentation schemes for protonated tripelenamine and methaphenilene.
Figure 4.5.4 Fragmentation scheme for protonated desloratadine.
Figure 4.5.5 Fragmentation schemes for protonated drofenine, oxybutynin, and propiverine and for the propantheline cation.
Figure 4.5.6 Proposed structures for the fragment ions of protonated tacrine.
Figure 4.5.7 Proposed structures for the fragment ions of protonated physostigmine.
Figure 4.5.8 Proposed structures for the fragment ions of protonated rivastigmine.
Figure 4.5.9 Proposed structures for the fragment ions of protonated donepezil.
Figure 4.5.10 Fragmentation pathways for protonated galantamine as proposed by Jegorov et al.
Figure 4.5.11 Alternative structure proposals for some of the fragments observed for protonated galantamine.
Figure 4.5.12 Proposed structures for some of the fragment ions of protonated levodopa (l-DOPA).
Figure 4.5.13 Fragmentation schemes for protonated selegiline, ropinirole, cabergoline, and metixene.
Figure 4.5.14 Proposed structures for the fragment ions of protonated anthracycline daunorubicin and fragmentation scheme for mitoxanthrone.
Figure 4.5.15 Proposed structures for the fragment ions of protonated ifosfamide, cyclophosphamide, and trofosfamide.
Figure 4.5.16 Proposed structures for the fragment ions of protonated nimustine.
Figure 4.5.17 Fragmentation schemes for protonated chlorambucil and melphalan.
Figure 4.5.18 Proposed structures for the fragment ions of protonated demecolcin.
Figure 4.5.19 Fragmentation scheme for protonated paclitaxel.
Figure 4.5.20 Fragmentation scheme for protonated teniposide.
Figure 4.5.21 Fragmentation schemes for protonated mycophenolic acid and deprotonated mycophenolic acid, leflunomide, and teriflunomide.
Figure 4.5.22 Fragmentation schemes for the protonated X-ray contrast agents iopromide and ioglicic acid.
Figure 4.5.23 Fragmentation schemes for protonated coumafuryl and deprotonated warfarin.
Figure 4.6.1 General structure and carbon numbering in steroids with (a) cholesterol and (b) testosterone.
Figure 4.6.2 Proposed nomenclature for steroid fragment ions.
Figure 4.6.3 Proposed structures for the fragment ions of protonated testosterone.
Figure 4.6.4 Proposed structures for the fragment ions of protonated stanozolol.
Figure 4.6.5 Proposed structures for the fragment ions of norethisterone in positive-ion MSⁿ in an ion-trap–time-of-flight hybrid instrument.
Figure 4.6.6 Proposed structures for the fragment ions of protonated betamethasone, showing characteristic fragment ions of 3-keto-Δ^1,4-corticosteroids.
Figure 4.6.7 Fragmentation pathways for protonated beclomethasone.
Figure 4.6.8 Proposed structures for the fragment ions of deprotonated estrogens, illustrated for estradiol.
Figure 4.6.9 Proposed structures for the fragment ions of protonated estrone.
Figure 4.7.1 Proposed structures for the fragment ions of protonated tetrahydrocannabinol.
Figure 4.7.2 Proposed structures for the fragment ions of deprotonated tetrahydrocannabinol.
Figure 4.7.3 Proposed structures for the fragment ions of protonated cocaine.
Figure 4.7.4 Fragmentation scheme for protonated dimethocaine.
Figure 4.7.5 Proposed structures for the fragment ions of protonated lysergic acid diethylamide.
Figure 4.7.6 Fragmentation schemes for two protonated synthetic cannabinoids, AB-FUCINACA and JWH-018.
Figure 4.7.7 Proposed structures for the fragment ions of protonated 25I-NBOMe, that is, 2-(4-iodo-2,5-dimethoxyphenyl)-N-(2-methoxybenzyl)ethanamine.
Figure 4.8.1 Fragmentation scheme and proposed structures for the fragment ions of protonated sulfadimidine (sulfamethazine).
Figure 4.8.2 Proposed structures for the fragment ions of deprotonated sulfamethoxazole.
Figure 4.8.3 Proposed structures for the fragment ions of deprotonated chloramphenicol.
Figure 4.8.4 Fragmentation scheme and proposed structures for the fragment ions of protonated penicillin G.
Figure 4.8.6 Fragmentation scheme and proposed structures for the fragment ions of protonated cefalexin.
Figure 4.8.5 Proposed structures for the fragment ions of deprotonated dicloxacillin.
Figure 4.8.7 Fragmentation scheme and proposed structures for the fragment ions of protonated cefotaxime.
Figure 4.8.8 Proposed structures for the fragment ions of deprotonated cefalexin.
Figure 4.8.9 Proposed structures for the fragment ions of deprotonated cefoxitin.
Figure 4.8.10 General structure of (fluoro)quinolone antibiotics.
Figure 4.8.11 Proposed structures for the fragment ions of the protonated quinolone antibiotics nalidixic acid, oxolinic acid, piromedic acid, and pipemidic acid.
Figure 4.8.12 Proposed structures for the fragment ions of the protonated fluoroquinolone antibiotic ciprofloxacin.
Figure 4.8.13 Proposed structures for the fragment ions of protonated gentamicin C1.
Figure 4.8.14 Fragmentation scheme and proposed structures for some fragment ions of protonated tetracycline.
Figure 4.8.15 Analysis of furazolidone antibiotics: release of the 3-amino-1,3-oxazolidin-2-one unit (AOZ) from protein adducts and subsequent derivatization using nitrobenzaldehyde.
Figure 4.8.16 Proposed structures for the fragment ions of protonated o-nitrobenzene derivatives of protein-released firazolidone, furaltadone, nitrofurantoin, and nitrofurazone.
Figure 4.8.17 Fragmentation scheme and proposed structures for some fragment ions of protonated erythromycin A.
Figure 4.8.18 Proposed structures for some fragment ions of protonated trimethoprim.
Figure 4.8.19 Proposed structures for the fragment ions of protonated spectinomycin.
Figure 4.9.1 Proposed structures for the fragment ions of some protonated imidazole fungicides.
Figure 4.9.2 Proposed structures for the fragment ions of protonated enilconazole. The fragment ions observed in ion-trap MSⁿ are indicated. Other ions are observed under collision-cell CID conditions.
Figure 4.9.3 Proposed structures for the fragment ions of some protonated triazole fungicides.
Figure 4.9.4 Proposed structures for the fragment ions of protonated triadimefon.
Figure 4.9.5 Proposed structures for the fragment ions of the protonated benzamidazole fungicides carbendazim and rabendazole. In the frame, structures proposed elsewhere are given for the fragment ions of carbendazim.
Figure 4.9.6 Fragmentation schemes for the protonated fungicides naftifine, terbinafine, and buprimate.
Figure 4.9.7 Proposed structures for the fragment ions of the protonated fungicide quinomethionate.
Figure 4.9.8 Fragmentation schemes and proposed structures for some of the fragment ions of protonated tolnaftate and tolciclate.
Figure 4.9.9 Fragmentation schemes for some deprotonated fungicides.
Figure 4.10.1 Fragmentation scheme and proposed structures for the fragment ions of protonated anthelmintic drug doramectin.
Figure 4.10.2 Proposed structures for the fragment ions of the protonated anthelmintics pyrantel and morantel (m/z of its fragment ions in parenthesis).
Figure 4.10.3 Proposed structures for the fragment ions of protonated dimetridazole.
Figure 4.10.4 Fragmentation schemes for the protonated chemical coccidiostats amprolium, robenidine, and halofuginone and for deprotonated halofuginone.
Figure 4.10.5 Fragmentation scheme for sodiated maduramicin, [M+Na]⁺.
Figure 4.10.6 Fragmentation scheme and fragment ions for sodiated salinomycin, [M+Na]⁺.
Figure 4.10.7 Structures for the fragment ion with m/z 479 from monensin A. The structure with m/z 479.262 was proposed as a fragment ion from [M+Na]⁺ (Volmer & Lock, 1998). Accurate-mass data from FT-ICR-MS indicates an alternative fragment ion with m/z 479.298 (Lopes et al., 2002b). The fragment ion with m/z 479.337 was observed from [M+H]⁺ (Lopes et al., 2002c).
Figure 4.10.8 Fragmentation scheme for deprotonated lasalocid A.
Figure 4.10.9 Fragmentation schemes for protonated chloroquine, piperaquine, and proguanil and for deprotonated proguanil.
Figure 4.10.10 Fragmentation schemes for protonated indinavir and nelvinavir.
Figure 4.10.11 Fragmentation schemes for deprotonated indinavir and saquinavir.
Figure 4.10.12 Fragmentation schemes for protonated moroxydine and oseltamivir.
Figure 4.10.13 Proposed structures for the fragment ions of protonated benzylcinnamate.
Figure 4.10.14 Fragmentation scheme for protonated chlorhexidine.
Figure 4.11.1 Proposed structures for the fragment ions of the protonated chlorotriazine herbicide atrazine.
Figure 4.11.2 Proposed structures for the fragment ions of the protonated methylthiotriazine herbicide ametryn.
Figure 4.11.3 Proposed structures for the fragment ions of the protonated methoxytriazine herbicide atraton.
Figure 4.11.4 General structures for the carbamate pesticide classes.
Figure 4.11.5 Proposed structures for the fragment ions of the protonated N-methyl carbamate pesticide carbofuran.
Figure 4.11.6 Proposed structures for the fragment ions of the protonated N-oxime carbamate pesticide aldicarb.
Figure 4.11.7 Proposed structures for the fragment ions of the protonated N,N-dimethyl carbamate insecticide pirimicarb.
Figure 4.11.8 Proposed structures for the fragment ions of the quaternary ammonium herbicide chlormequat.
Figure 4.11.9 Ions observed for the quaternary ammonium herbicides paraquat and diquat, and proposed structures for the fragment ions based on [M−H]⁺.
Figure 4.11.10 Examples of organophosphorus pesticides: bromfenvinphos-methyl is a dimethyl phosphate, fenthion a dimethyl phosphorothioate, azinphos-methyl a dimethyl phosphorodithioate, omethoate a dimethyl phosphorothioate, bromfenvinphos a diethyl phosphate, diazinon a diethyl phosphorothioate, and azinphos-ethyl a diethyl phosphorodithioate.
Figure 4.11.11 Proposed structures for the fragment ions of (a) dimethyl phosphates, (b) diethyl phosphates, (c) dimethyl phosphorothioates, (d) diethyl phosphorothioates, (e) dimethyl phosphorodithioates, and (f) diethyl phosphorodithioates, described in Table 4.11.2.
Figure 4.11.12 Fragmentation scheme and proposed structures for the fragment ions of the protonated phosphorothioate pesticide diazinon.
Figure 4.11.13 Fragmentation schemes for deprotonated organophosphorus pesticides quinalphos and bromophos methyl.
Figure 4.11.14 Proposed structures for the fragment ions of the protonated and deprotonated organophosphorus pesticide dimethoate. The framed structure shows the interpretation of the negative-ion product-ion mass spectrum proposed elsewhere (Fernández et al., 2001a).
Figure 4.11.15 Fragmentation scheme and proposed structures for fragment ions of the protonated phenylurea herbicide diuron.
Figure 4.11.16 Fragmentation scheme for the protonated benzoylphenylurea herbicides illustrated for flufenoxuron. The F₄ fragment ion is only observed for fluazuron and chlorflurazone.
Figure 4.11.17 Fragmentation schemes for the protonated sulfonylurea herbicides foramsulfuron and chlorsufuron.
Figure 4.11.18 Fragmentation schemes for the deprotonated sulfonylurea herbicides nicosulfuron and chlorsufuron.
Figure 4.11.19 Fragmentation scheme for the protonated halogenated aryloxyphenoxypropionic acid ester herbicide fluazifop-butyl.
Figure 4.11.20 Proposed structures for the fragment ions of the protonated anilide herbicides chloranocryl, flufenacet, and mefenacet.
Figure 4.11.21 Proposed structures for the fragment ions of the protonated chloracetanilide herbicide alachlor.
Figure 4.11.22 Proposed structures for the fragment ions of the protonated cyclohexanedione oxime herbicide cycloxydim.
Figure 4.11.23 Proposed structures for the fragment ions of the protonated cyclohexanedione oxime alloxydim.
Figure 4.11.24 Proposed structures for the fragment ions of the protonated dinitroanaline herbicide benfluralin.
Figure 4.11.25 Proposed structures for the fragment ions of the protonated imidazolinone herbicide imazapyr.
Figure 4.11.26 Fragmentation scheme for the deprotonated nitrophenyl ether herbicide acifluorfen.

Chapter 5: Identification Strategies

Figure 5.1 Structure and major fragments of nefazodone. In the top structure, both profile groups and hydrolysis positions are indicated.
Figure 5.2 MS, MS², and MS³ mass spectra of the “known unknown” discussed, which is identified as 1,3-diphenylguanidine.
Figure 5.3 Hits for the search of C₁₃H₁₃N₃ in the Sigma-Aldrich database/catalog. Our prediction on the ability to generate the fragment ions with m/z 195 (−NH₃), 119, and 94 is indicated.

List of Tables

Chapter 1: Introduction To LC–MS Technology

Table 1.1 Proton affinities of compounds commonly used in GC–MS and LC–MS
Table 1.2 Common reagent gases used in positive-ion CI and adducts formed thereof
Table 1.3 Anions used for neutral analyte negative ionization in GC–MS and LC–MS

Chapter 2: Interpretation of Mass Spectra

Table 2.1 Standard atomic weights and isotopic abundances for atoms commonly present in bioorganic MS (NIST^*)
Table 2.2 Elements commonly encountered in bioorganic MS and tabulated by type and percentage (%) relative abundance
Table 2.3 Expected signal isotopic ratios for ions containing Cl + Br atoms as shown in Figure 2.5
Table 2.4 Summary of the nitrogen rule for OE^+•/OE^−• and EE⁺/EE⁻ ions or molecules of a given nominal mass

Chapter 3: Fragmentation of Even-Electron Ions

Table 3.1 Fragmentation (expressed as percentage relative abundance, %RA) of protonated C₅-dialkyl- and trialkylamines ([C₅H₁₃N+H]⁺; [M+H]⁺ with m/z 88)
Table 3.2 Proton affinities (PA in kJ mol⁻¹) for a number of compounds
Table 3.3 Small neutral losses in positive-ion mode
Table 3.5 Small neutral losses in negative-ion mode
Table 3.4 Characteristic fragment ions in positive-ion mode
Table 3.6 Characteristic product ions in negative-ion mode

Chapter 4: Fragmentation of Drugs and Pesticides

Table 4.1.1 Fragmentation of β-blockers with a 1-(iso-propylamino)-3-phenoxypropan-2-ol skeleton
Table 4.1.2 Fragmentation of β-blockers with a 1-(tert-butylamino)-3-phenoxypropan-2-ol skeleton
Table 4.1.3 Dihydropyridine calcium channel blockers
Table 4.1.4 Fragmentation of angiotensin II converting enzyme (ACE) inhibitors
Table 4.1.5 Structures and negative-ion fragmentation of the benzothiadiazine diuretics studied
Table 4.2.1 Characteristic fragments of various classes of phenothiazines
Table 4.2.2 Fragmentation of benzodiazepines containing the N¹-desmethyl acetamide skeleton in the diazepine ring: m/z values of the ions observed
Table 4.2.3 Fragmentation of benzodiazepines containing the N¹-desmethyl-α-hydroxy acetamide skeleton in the diazepine ring: m/z values of the ions observed
Table 4.4.1 Class-specific fragmentation for second-generation sulfonylurea antidiabetics
Table 4.5.1 Fragmentation of β-adrenergic receptor agonists
Table 4.6.1 Structures of glucocorticosteroids
Table 4.6.2 Fragmentation of 17 representative corticosteroids in negative-ion mode (structures in Table 4.6.1)
Table 4.6.3 Fragmentation of estrogen in negative-ion MS–MS
Table 4.7.1 Class-specific and major compound-specific fragment ions of amphetamine and related compounds (designer drugs or NPSs)
Table 4.7.2 Class-specific and major compound-specific fragment ions of 3,4-methylenedioxyamphetamine (MDA) and related compounds (designer drugs or NPSs)
Table 4.7.3 Class-specific and major compound-specific fragment ions of x-(2-aminopropyl)benzofurans and related compounds (designer drugs or NPSs), with x = 4, 5, 6, or 7
Table 4.7.4 Class-specific and major compound-specific fragment ions of cathinones (designer drugs or NPSs), with (a) cathinones based on the 2-amino-1-phenylpropan-1-one skeleton, and (b) cathinones based on other 2-amino-1-phenylalkan-1-one skeletons
Table 4.7.5 Class-specific and major compound-specific fragment ions of pyrrolidinophenones (designer drugs or NPSs)
Table 4.7.6 Fragmentation of cocaine and its metabolites (Figure 4.7.3)
Table 4.7.7 Structure and major fragmentation of synthetic cannabinoids of the naphthoylindole class
Table 4.7.8 m/z values for characteristic fragments of 25X-NBOMe compounds, with 2′-methoxy unless otherwise indicated
Table 4.8.1 Characteristic fragment ions for protonated sulfonamide antibiotics
Table 4.8.2 Class-specific fragment ions of protonated penicillin β-lactam antibiotics
Table 4.8.3 Structures and fragmentation of deprotonated penicillin β-lactam antibiotics
Table 4.8.4 Structures and class-specific fragment ions of cephalosporin β-lactam antibiotics with a 2-amino-2-phenylacetyl substituent chain at the amide C⁷ of the β-lactam ring
Table 4.8.5 Structures and class-specific positive-ion fragment ions of cephalosporin β-lactam antibiotics with a 2-(2-amino-1,3-thiazol-4-yl)-2-(methoxyimino)acetyl substituent chain at the C⁷amide of the β-lactam ring and a substituent (ester, (thio)ether, or quaternary ammonium (4^o N)) on the methylene (CH₂) group at C³ of the β-lactam ring
Table 4.8.6 Fragmentation of aminoglycosides in MS–MS (for annotation, Figure 4.8.13)
Table 4.10.1 Molecular formulae and characteristic fragment ions frequently used in SRM transitions for avermectin anthelmintics
Table 4.10.2 Fragmentation of chlorinated and brominated phenol derivatives used as antiseptics and disinfectants
Table 4.11.1 Fragmentation of (a) chlorotriazine herbicides (Figure 4.11.1), (b) methylthiotriazine herbicides (Figure 4.11.2), and (c) methoxytriazine herbicides (Figure 4.11.3)
Table 4.11.2 Fragmentation of protonated N-methyl carbamates
Table 4.11.3 Class-specific fragments in positive-ion product-ion mass spectra of OPP
Table 4.11.4 Class-specific fragment ions and neutral losses in negative-ion MS–MS spectra of phosphorothioate and phosphorodithioate OPPs
Table 4.11.5 Structures and major fragment ions for selected phenylurea herbicides
Table 4.11.6 Structures and major fragment ions for selected urea herbicides
Table 4.11.7 Fragmentation of benzoylphenylurea herbicides. Cleavage sites are illustrated for hexaflumuron
Table 4.11.8 Fragmentation of sulfonylurea herbicides, illustrated for the pyrimidinyl sulfonylurea foramsulfuron and the triazinyl sulfonaylurea chlorsulfuron (Figure 4.11.16)
Table 4.11.9 Structures and fragmentation of chlorinated phenoxy acid herbicides (CPAs)
Table 4.11.10 Structure and major fragment ions of chloracetanilide herbicides

Chapter 5: Identification Strategies

Table 5.1 Mass shifts and mass-defect shifts due to Phase I biotransformation reactions
Table 5.2 Mass shifts and mass-defect shifts due to Phase II biotransformation
Table 5.3 Selected compound databases either commercially available or publicly available on the internet
Table 5.4 Selected software tools for in silico prediction of fragmentation of small molecules in MS–MS or MSⁿ
Table 5.5 Mass Frontier prediction of the five characteristic fragment ions of the unknown with an [M+H]⁺ with m/z 212 ([C₁₃H₁₃N₃+H]⁺) for the five structure proposals obtained from searching the Sigma-Aldrich catalog (Figure 5.3). Mass Frontier (version 5) was used

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Names: Niessen, W. M. A. (Wilfried M. A.), 1956- author. | Correa C., Ricardo A., 1961- author.

Title: Interpretation of MS-MS mass spectra of drugs and pesticides / Wilfried M.A. Niessen, Ricardo A. Correa C.

Description: Hoboken, New Jersey : John Wiley & Sons, 2016. | Series: Wiley series on mass spectrometry | Includes bibliographical references and index.

Identifiers: LCCN 2016031593 (print) | LCCN 2016046526 (ebook) | ISBN 9781118500187 (cloth) | ISBN 9781119294245 (pdf) | ISBN 9781119294252 (epub)

Subjects: LCSH: Tandem mass spectrometry. | Liquid chromatography. | Drugs--Analysis. | Pesticides–Analysis.

Classification: LCC QD96.M3 N525 2016 (print) | LCC QD96.M3 (ebook) | DDC 543/.65--dc23

Preface

In the 1980s, tandem mass spectrometry was introduced for the structural elucidation of even-electron ions (protonated or deprotonated molecules) generated by soft ionization techniques such as fast-atom bombardment, thermospray, and electrospray. When compared to the fragmentation of odd-electron ions generated by electron ionization, scientists were well aware of the fact that different rules apply to the fragmentation of even-electron ions. Surprisingly, no major fundamental research was carried out on trying to understand and describe these differences. More effort was placed on the development of improved instrumentation and advanced applications for the emerging technologies. This particular effort paid off, as exemplified by tandem mass spectrometry which, often in combination with gas or liquid chromatography, has been a major contributor to the progress of many scientific disciplines, for example, pharmaceutical, biochemical, and environmental sciences; food safety; sports doping analysis; clinical diagnostics; forensics; and toxicology.

This work is an attempt to add to the understanding of the fragmentation of even-electron ions. This has been done by studying the fragmentation of a wide variety of compounds, with a special focus on chemical structure similarities, that is, from the same class. The basic data set used comprises a number of mass spectral libraries developed for general unknown screening in toxicology. In this respect, we need to thank Dr Wolfgang Weinmann (originally at the Institute of Legal Medicine, University of Freiburg, Germany, and currently at the Institute of Forensic Medicine, University of Bern, Switzerland) for providing public access to his toxicology library and the library of designer drugs via the Internet (http://www.chemicalsoft.de/index.html); Dr Pierre Marquet (of the Faculty of Medicine, Department of Pharmacology, Toxicology, and Pharmacovigilance at the University Hospital of Limoges, France) for providing his mass spectral library of negative-ion mass spectra; and Dr Bernhard Wüst of Agilent Technologies for his help with using the Agilent Broecker, Herre & Pragst PCDL for forensic toxicology. The information from these libraries and other data sets is complemented by data from the scientific literature.

The origins of this book can be found in two publications describing the fragmentation of toxicologically relevant drugs in both positive-ion tandem mass spectrometry (Niessen, 2004) and negative-ion tandem mass spectrometry (Niessen, 2005). Soon after, the authors decided to develop the project further by extending the number of compounds covered and the detail of the information provided. The fragmentation of some 1300 compounds and the product-ion mass spectra of even more are studied and interpreted in this book.

This volume consists of five chapters. Chapters 3 and 4 are the main chapters, where proposed fragmentation rules for the “Fragmentation of Even-Electron Ions” (Chapter 3) are derived from the behavior of the “Fragmentation of Drugs and Pesticides” (Chapter 4) pertaining to many different classes of compounds. Chapter 1, “Introduction to LC–MS–MS Technology”, provides a concise introduction to mass spectrometry technology. Chapter 2, “Interpretation of Mass Spectra” gives the basic concepts and definitions related to the information that can be extracted from mass spectra. Finally, Chapter 5, “Identification Strategies” gives an overview of the different classes of unknowns and identification strategies that exist as well as how they relate to multiple areas of application.

Last but not least, special thanks go to our families, and the many people who have inspired us to continue working on this project. We hope that you, as our reader, find this material useful and inspirational to further extend our understanding of the fragmentation of even-electron ions in tandem mass spectrometry.

Wilfried M. A. Niessen
hyphen MassSpec
Herenweg 95, 2361 EK Warmond, The Netherlands
mail@hyphenms.nl; www.hyphenms.nl

Ricardo A. Correa C.
Trans-Laboratory
Rue François Stroobant 41, 1050 Brussels, Belgium
ricardo.correa@translaboratory.com; www.translaboratory.com

References

Niessen WMA. 2011. Fragmentation of toxicologically relevant drugs in positive-ion liquid chromatography–tandem mass spectrometry. Mass Spectrom Rev, 30: 626–663.
Niessen WMA. 2012. Fragmentation of toxicologically relevant drugs in negative-ion liquid chromatography–tandem mass spectrometry. Mass Spectrom Rev, 31: 626–665.