Identification of the related substances of tilmicosin by liquid chromatography/ion trap mass spectrometry

Structures of seven impurities of the veterinary drug tilmicosin have been elucidated by multiple fragmentation with ion trap tandem mass spectrometry. All related compounds possess the main lactone ring of tilmicosin. The differences in their structures are due to the hydroxyl, mycaminose, 3,5-dimethylpiperidine and mycinose groups connected to C(3), C(5), C(6), C(14) of the lactone ring, respectively. The following compounds of the impurity profile of tilmicosin were identified: B - tilmicosin with a hydroxyl group at C(3); C - tilmicosin without a methyl group at the N-atom connected to C(3) of the mycaminose ring; D - tilmicosin with a hydroxyl group at C(6) of the mycaminose ring; E - tilmicosin with a methoxy group at C(3), F - desmicosin; G - 20-dihydrodesmicosin; and H - tilmicosin without a mycaminose ring. Isomers of the compounds B, C, D, E and H were identified by their mass chromatograms and retention times. The concentrations of the impurities varied in the range of 0.1% to 2.9%.     

Mass spectra of some naturally occurring stictane triterpenoids and their trimethylsilyl derivatives

The mass spectra of ten di- and tri-oxygenated stictane triterpenoids and their trimethylsilyl derivatives have been studied in detail. Gas chromatography mass spectrometry of the TMS derivatives on OV-17 and OV-101 columns provided useful separations and identifications of mixtures of stictanes triterpenoids in lichen extracts. The major fragmentations involve ring C, but specific cleavages in rings A, B and E are also observed which allow the substitution pattern of the skeleton to be readily determined. The formation of an intense [M – C₅H₁₁?]⁺ ion for 22α-OTMS stictanes appears specific to their ring E configuration.     

Selective reagents in chemical ionization mass spectrometry: Tetramethylsilane

Mixtures of 50% tetramethylsilane (TMS) and methane have been found to give [M+73]⁺ adduct ions and structurally useful fragment ions for many oxygen- and nitrogen-containing organic compounds. All of the reagent ions in TMS react with polar compounds. The high-pressure TMS chemical ionization spectra of many simple oxygenated compounds are in agreement with predictions from ion chemistry of (CH₃)₃Si⁺ obtained by ion cyclotron resonance experiments at very low pressures, but differences are noted. Sensitivities for oxygen-, nitrogen-, and sulfur-containing compounds with TMS as the reagent gas appear to be approximately the same.     

Influence of substituent groups at the 3‐position on the mass spectral fragmentation pathways of cephalosporins

The structural fragment ions of nine cephalosporins were studied by electrospray ionization quadrapole trap mass spectrometry (Q‐Trap MSⁿ) in positive mode. The influence of substituent groups in the 3‐position on fragmentation pathway B, an α‐cleavage between the C7? C8 single bond, coupled with a [2,4]‐trans‐Diels‐Alder cleavage simultaneously within the six‐membered heterocyclic ring, was also investigated. It was found that when the substituent groups were methyl, chloride, vinyl, or propenyl, fragmentations belonging to pathway B were detected; however, when the substituents were heteroatoms such as O, N, or S, pathway B fragmentation was not detected. This suggested that the [M–R₃]⁺ ion, which was produced by the bond cleavage within the substituent group at the 3‐position, had a key influence on fragmentation pathway B. This could be attributed to the strong electronegativity of the heteroatoms (O, N, S) that favors the production of the [M–R₃]⁺ ion. Moreover, having the positive charge of the [M–R₃]⁺ ion localized on the nitrogen atom in the 1‐position changed the electron density distribution of the heterocyclic structure, which prohibits a [2,4]‐reverse‐Diels‐Alder fragmentation and as a result fragmentation pathway B could not occur. The influence of the substituent group in the 3‐position was determined by the intensity ratio (e/d) of ions produced by fragmentation pathway A, a [2,2]‐trans‐Diels‐Alder cleavage within the quaternary lactam ring, including the breaking of the amide bond and the C6? C7 single bond (ion d), and fragmentation pathway B (ion e). The results indicate that the electronegativity of the substituent group was a key influencing factor of pathway B fragmentation intensity, because the intensity ratio (e/d) is higher for a chlorine atom, a vinyl, or a propenyl group than that of a methyl group. This study provided some theoretical basis for the identification of cephalosporin antibiotics and structural analysis of related substances in drugs. Copyright © 2010 John Wiley & Sons, Ltd.     

On the stereospecificity of the structurally diagnostic m/z 84 ion in the mass spectra of gibberellins and kaurenes with a 15-hydroxy substituent

The presence of a 15-hydroxy substituent in naturally-occurring kaurenes and gibberellins gives rise to a structurally diagnostic ion at m/z84 (C₅H₈O) in their electron-impact mass spectra, which shifts to m/z156 for the TMS derivative, and is unaffected by differences in the A and B rings. Studies on a model tricyclic system have shown that this ion is more intense when the stereochemistry of the B/C ring junction is cis than when it is trans. In accordance with a previously proposed pathway for the formation of this ion, deuterium labelling confirms that there is a site-specific transfer of the β-hydrogen from the cis ring junction to the C₅ fragment ion, although a minor modification of the earlier mechanism is proposed.     

Fragmentation of 2‐aroylbenzofuran derivatives by electrospray ionization tandem mass spectrometry

We investigated the gas‐phase fragmentation reactions of a series of 2‐aroylbenzofuran derivatives by electrospray ionization tandem mass spectrometry (ESI‐MS/MS). The most intense fragment ions were the acylium ions m/z 105 and [M+H–C₆H₆]⁺, which originated directly from the precursor ion as a result of 2 competitive hydrogen rearrangements. Eliminations of CO and CO₂ from [M+H–C₆H₆]⁺ were also common fragmentation processes to all the analyzed compounds. In addition, eliminations of the radicals •Br and •Cl were diagnostic for halogen atoms at aromatic ring A, whereas eliminations of •CH₃ and CH₂O were useful to identify the methoxyl group attached to this same ring. We used thermochemical data, obtained at the B3LYP/6‐31+G(d) level of theory, to rationalize the fragmentation pathways and to elucidate the formation of E , which involved simultaneous elimination of 2 CO molecules from B .     

Cyclic ions in the mass spectra of trimethylsilyl derivatives of substituted o-dihydroxybenzenes

The mass spectra of trimethylsilyl (TMS) ethers/methyl esters of phenolic acids containing o-dihydroxybenzene groups have base peaks at [M?119]⁺ instead of the usual [M?15]⁺ and [M?31]⁺ that are characteristic of TMS/methyl esters of monohydroxyphenolic acids. These ions, formed by the loss of 31+88 u from the parent ion, possess a cyclic moiety as proven by substitution of deuterium atoms for hydrogen atoms in the TMS groups of the methyl esters of 3,4,5-trihydroxybenzoic (gallic), 3,4-dihydroxybenzoic (protocatechuic) and β-(3,4-dihydroxyphenyl)propenoic (caffeic) acids. Although these cyclic ions are the base peaks in TMS-derivatized o-dihydroxyphenolic acid esters, similar ions represent intense peaks but not necessarily the base peak in other derivatized compounds such as 1,2-dihydroxybenzene, 1,2-dihydroxy-3-methyl- and 1,2-dihydroxy-4-methyl-benzenes and flavan-3-ols that possess o-dihydroxybenzene groups. Compounds possession m- or p-dihydroxybenzene groups do not form these cyclic ions; therefore, this procedure for derivatization and interpretation of mass spectra is valuable for the identification of compounds containing o-dihydroxybenzene groups in complex mixtures of isomeric compounds.     

Mass spectrometry of isoxazole derivatives—III: Skeletal rearrangement ions in the mass spectra of isoxazoles having a heteroatom at C5

Two skeletal rearrangements of the type[ABC]^+.→[AC]^+. + B previously found in the mass spectra of 5-alkoxyisoxazoles are also seen for 5-amino- and 5-alkylmercapto-isoxazoles. They are discussed in terms of formation of an azirine ion and its ring expansion to an odd electron ion having a five-membered ring by means of bond formation involving nitrogen or sulphur. Fragmentation via an oxazole radical ion, which has a photochemical analogy, is also discussed.     

Gas chromatography electrospray ionization mass spectrometry analysis of trimethylsilyl derivatives

A method based on the analysis of trimethylsilyl (TMS) derivatives by capillary gas chromatography electrospray ionization mass spectrometry (GC–ESI/MS) was proposed. To improve separation, analytes were derivatized to their TMS derivative. During ESI analysis, TMS derivatives may hydrolyze back to their polar native form and are thus suitable for ESI analysis. Several types of analytes were studied to investigate the potential of the approach. Not all TMS derivatives hydrolyzed back to their native form as anticipated. Incomplete hydrolysis was observed for TMS‐organic acids and TMS‐nonchlorinated phenols. For TMS‐chlorophenols, the observation of only the [M ? H]^? ion suggested that these phenols were hydrolyzed back to their native form. For TMS‐beta agonists, the hydrolysis rate was low; therefore, the hydrolysis product was not detected. Both TMS‐chlorophenols and TMS‐beta agonists provide a sensitivity in the range of low parts per billion (0.25–5 ng/ml and 0.5–10 ng/ml respectively). Copyright © 2016 John Wiley & Sons, Ltd.     

Mass spectrometry in structural and stereochemical problems—CXCIX Contrasting fragmentations of singly- and doubly-charged o-, m- and p-hydroxyalkylphenones and their trimethylsilyl ethers

High resolution mass spectrometry, metastable defocusing and deuterium labeling of trimethylsilyl (TMS) ethers have been used to study the electron-impact induced fragmentations of o-, m- and p-hydroxyalkylphenones and their TMS ether derivatives. These derivatives have proven useful in contrasting the fragmentation patterns of singly- and doubly-charged ions because of the competing fragmentations: α-cleavage and a McLafferty rearrangement from the ketone moiety and methyl cleavage from the TMS group. A proximity effect was responsible for a markedly increased methyl radical loss from the o-TMS ether. This fragmentation was minor with the m- and p-isomers. Significantly intense doubly-charged ions were formed from ketonic cleavage and by the loss of a TMS methyl radical. The sequence of fragmentation depended on the size of the alkyl group attached to the ketone carbonyl. There was no evidence found for a McLafferty rearrangement occurring from the doubly-charged molecular ion of the TMS ethers of the hydroxyalkylphenones but the rearrangement occurred from the doubly-charge molecular ion of bis-3-(1-oxopentyl)-4-hydroxy-phenyl-methane and, of course, from the singly charged [M]^+. The bis-p-hydroxyphenylmethane derivatives were studied in an effort to increase the intensity of the doubly-charged ions as it was expected that the charges would be separated by a longer distance.     

Tetramethylsilane chemical ionization mass spectrometry of some isomeric unsaturated dicarboxylic acids and esterst

The tetramethylsilane (TMS) chemical ionization (CI) mass spectra of some geometrical isomers of unsaturated dicarboxylic acids, esters and isomeric phthalic acids reveal explicit differences. The (E)-acids show an abundant [M + 73 ? CH₄]⁺Ion whereas the (Z)-acids exhibit a strong [M + 73 ? H₂O]⁺ ion in their TMS CI spectra. The loss of CH₄ from the adduct of fumaric acid has been confirmed by the study of fumaric acid-d₂ and B/E linked scan studies. In the case of esters, the TMS CI spectra of (E)-isomers contain abundant [M + 73]⁺ adduct ions, whereas these are weakly abundant in the TMS CI of the (Z)-isomers.     

Generation of CsI cluster ions for mass calibration in matrix-assisted laser desorption/ionization mass spectrometry

A simple method was developed for the generation of cesium iodide (CsI) cluster ions up to m/z over 20,000 in matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). Calibration ions in both positive and negative ion modes can readily be generated from a single MALDI spot of CsI₃ with 2-[(2E)-3-(4-tert-butylphenyl)-2-methylprop-2-enylidene] malononitrile (DCTB) matrix. The major cluster ion series observed in the positive ion mode is [(CsI)_nCs]⁺, and in the negative ion mode is [(CsI)_nI]⁻. In both cluster series, ions spread evenly every 259.81 units. The easy method described here for the production of CsI cluster ions should be useful for MALDI MS calibrations.     

Investigation of 4-(nitrophenylamino)pent-3-en-2-ones and 4-(nitrobenzylamino)pent-3-en-2-ones by electron ionization mass spectrometry. Observation of characteristic ortho effects

The compounds 4-(phenylamino)pent-3-en-2-ones (1-3) and 4-(benzylamino)pent-3-en-2-ones (4-6) substituted with a nitro group on the aromatic ring were studied by electron ionisation mass spectrometry (EIMS). It was deduced that the compounds 1-3 are converted into the tautomeric 4-(arylimino)pentan-2-one during the EI process. Mass spectrometric decompositions of ortho-substituted derivatives (1 and 4) were found to be different from those observed for meta- and para-isomers. The fragmentation pathways are discussed on the basis of data from linked B/E and B(2)/E scans, mass-analysed ion kinetic energy (MIKE) spectra, accurate mass measurements and isotope labelling.     

The electron impact mass spectra of some cevine orthoacetates

The most important fragmentation in the electron impact mass spectra of dihydrocevine orthoacetate and some analogues involves loss of an acetoxyl radical from the molecular ion, and is associated with strain in the orthoacetate group bridging a six-membered ring. The process is much less important in the less-strained dihydrocevine isoorthoacetate, which gives the ion m/z 112, characteristic of cevine alkaloids, as the most abundant. A further important difference concerns an ion involving the loss of C(20) and its substituents together with a hydrogen atom from M⁺˙, also relatively unimportant in the isoorthoacetate.     

The mass spectrum of 1-(2-thienyl) hexane-1-13C

The mass spectrum of 1-(2-thienyl) hexane-1-¹³C is reported. The principal fragmentation routes ofthe parent ion are delineated. Beta cleavage of the alkyl chain predominates, and the label retention indicates that the alpha carbon atom remains with the charged ring moiety. A substantial part of the m/e 97 ion undergoes a ring expansion to a six membered ring. A small amount of alpha cleavage of the alkyl chain occurs with the expected loss of the label. Further fragmentation of the ions from initial alpha or beta cleavage produces similar fragment ions. It is noted that many of the neutral particles lost in the formation of the fragment ions are typical of those encountered in the alkylbenzenes or other aromatic ion systems.     

Strained heterocyclic systems. VII. The mass spectral fragmentations of dihydrocycloalka[b]quinolines and quinoline N-Oxides

High resolution mass spectra of compounds 1-6 were investigated. The quinolines ( 1-3 ) all exhibit a prominent (M-1)⁺ peak and subsequent loss of HCN. These processes are consistent with azatropylium ion intermediates. The N-oxides ( 4-6 ) all exhibit major peaks at (M-16)⁺; the abundance of the latter is related to the geometry of the molecule. The four-membered ring compounds (3 and 6) give more complex spectra, which reflect the influence of the fused strained ring.     

Structure and Rearrangements of 7-(Heptaphenylcycloheptatrienyl)isochalcogencyanates

According to DFT B3LYP/6-31G(d,p) calculations, the intramolecular migration of NCX groups (X = O, S, Se, Te) in 7-(heptaphenylcycloheptatrienyl)isochalcogencyanates in the gas phase proceeds via the dissociation–recombination mechanism. Structures were found of transition states preceding the formation of tight ion pairs. High activation barriers to the migration of isochalcogencyanate groups (\(\Delta E{^\neq_{\rm{ZPE}}}\) 26.7, 21.3, 20.5, 19.3 kcal mol^–1) originate from the existence of stable conformation of molecules with NCX groups located in the equatorial position and are in agreement with the data of dynamic NMR. For the transition of the molecule in the form favorable for the NCX group migration an inversion of the seven-membered ring and the rotation of the phenyl group are necessary.     

Structural analysis of rapamycin and related compounds using [M + Li]+ ions generated by liquid secondary ion mass spectrometry

Cationization of the macrocyclic immunosuppressant rapamycin with lithium ion upon liquid secondary ion mass spectrometric ionization yields a number of fragment ions, which are observable in the full-scan spectrum. These are clearly assigned using B/E linked scanning (fragment ion scanning), B²/E linked scanning (precursor ion scanning) and peak matching for accurate mass measurement. Many of the fragments are produced by processes that open the macrocyclic ring, and it is possible to observe several different pieces of the molecule as fragment ions. The diversity of fragments produced facilitates the elucidation of new rapamycin-like structures through mass spectrometry. Three structurally modified rapamycin analogues have been examined by this technique, and the modifications to the molecule may be located based on the nominal masses of their fragments.     

Electron ionization mass spectrometric study of substituted alloxazine‐5‐oxides and iso‐alloxazine‐5‐oxide

The fragmentation pathways in electron ionization (EI) mass spectra of a series of new N(5)‐oxides of alloxazines and iso‐alloxazine are presented, and compared with those of substituted alloxazines and iso‐alloxazine. The EI mass spectra of these compounds showed characteristic fragmentation pathways A, B and C, started by the ejection of atomic oxygen, a HNCO molecule and an OH ^. radical, respectively. On the basis of B/E and B²/E spectra, the mechanism of elimination of the OH ^. radical is discussed. The influence of the methyl substituent in the benzene ring of alloxazine on the mass fragmentation pathways is described. Copyright © 2009 John Wiley & Sons, Ltd.