John B. Westmore

Cleavage of interchain disulfide bonds following matrix-assisted laser desorption☆

Abstract The metastable decay of insulin produced by matrix-assisted laser desorption has been examined in a reflecting time-of-flight mass spectrometer. The [M-H]− molecular ion decays into negative A and B chain daughters with various numbers of sulfurs attached, corresponding to cleavage of the interchain disulfide bridges at different points. This type of decomposition can be identified by the characteristic 32 u spacing between the components.

Mass spectra of sterically crowded trialkylsilyl ether derivatives of steroids.

The electron-impact mass spectra of the title compounds have some important features which give these derivatives certain advantages over the widely studied trimethylsilyl analogues. There is significantly less extensive fragmentation, and abundant ions at (M - t.Bu)+ or (M - i.Pr)+ serve as indicators of molecular weight and should be useful for selected ion monitoring. From various precursors, the ease of elimination of HX2SiOH, via a proposed multi-centre transition state, appears to depend upon conformational and stereochemical factors, as well as the position of the parent silyloxy group, RX2SiO, on the steroid skeleton. This particular fragmentation appears to be a powerful diagnostic method for distinguishing between stereoisomers, being especially useful for differentiation between epimers. In addition, the presence of a 17-silyloxy function promotes a characteristic cleavage of ring B in a skeletally saturated steroid. Elimination of silanol, RX2SiOH, at various stages in the fragmentations of bis-silylated steroids is also an important process, but other familiar features of the spectra of steroid trimethylsilyl ethers, though usually present, are very much suppressed.

Secondary ion mass spectrometry of protected oligonucleotides

Abstract Positive and negative secondary ion mass spectra, produced by 8 or 28 keV Cs+ ions, have been measured for several protected oligoribonucleotides with masses up to 3452 u. Malecular weights can be deduced from the ions [M+H]+, [M+Na]+, [M+Cs]+ (for Cs+ -doped samples), [M-H]− and [M+Cl]−. In the positive ion spectra, fission of C5′-OP bonds yields carbocations while fission of C3′-OP bonds gives species with the positive charge on the phosphate-containing fragments, which may be protonated or cationized. The negative ion spectra reveal the presence of phosphate anions produced by loss of any of the three attached groups.

Injector port reactions in gas chromatography : Sources of error in the quantitative analysis of alkylsilyl derivatives of nucleosides

Abstract The direct monitoring, by gas chromatography, of rates of silylation of various substrates is complicated by accelerated reactions occurring in the hot injector port of the gas chromatograph, e.g. further silylation by excess reagent. Monitoring the hydrolysis of silyl derivatives is also prevented since desilylation by hydrolysis media occurs in the injector port. Techniques have been developed to avoid these injector port reactions and include: (a) deactivation of silylating reagent by reaction with excess methanol, (b) deactivation of substrate by further derivatization with an active reagent, and (c) removal of hydrolysis media in vacuo. The methods are illustrated by studies on the deoxynucleoside thymidine but should be capable of extension to a wide variety of suitable substrates.

Sterically crowded trialkylsilyl ether derivatives for the analysis of steroid metabolites.

The applications of sterically crowed trialkyksilyl ether derivatives to the analysis and characterization by thin-layer chromatography, gas chromatography and mass spectrometry, of metabolites of 2alpha, 3alpha-cyclopropano-5alpha-androstan-17beta-ol in the rabbit are described. These derivatives are complementary to the familiar trimethysilyl ether derivatives, but have greater hydrolytic stability (an advantage for TLC), generally give better GC separations, and have characteristic mass spectra. Isomer differentiation by GC and MS is also more readily achieved than via the TMSi ether derivatives. These properties should make SCTASi ethers useful derivatives for studies of steroid metabolism.

Metastable ion studies with a secondary ion time-of-flight mass spectrometer. Enhanced distinction between isomers of O-alkylated thymidines

Abstract First-order rate constants and half-lives have been measured for the decomposition of several metastable ions in the secondary ion time-of-flight mass spectra of 4- and 2- O -alkylthymidines. Some of the factors which influence the accuracy of these measurements have been analyzed. Relative values of the kinetic parameters appear to be a more reliable indicator of isomeric form than the “normal” mass spectra, which are sensitive to the method of sample preparation. In particular, the [M + Na] + ions of 4- O -alkylthymidines are significantly more stable than those of 2- O -alkylthymidines, an effect attributed to chelation of Na + by the 2, 4′, and 5′ oxygens of the former isomers. An interesting rearrangement involving loss of an alkene from the [B + 2H] + ion (B = pyrimidine base) has also been confirmed. The rate of this decomposion is much greater for O -isopropyl- than for O -ethylthymidines, an effect which is rationalized in terms of conformational preferences of the O -alkyl group.

Sequencing an «unknown» peptide by time-of-flight secondary ion mass spectrometry

Abstract The procedure in sequencing the peptide dynorphin 1–17 (mass 2146.20 u) with a reflecting time-of-flight mass spectrometer is reported in some detail. Samples (1.5 μg) of the peptide were distributed as unknowns to participating laboratories in preparation for a workshop at the 1990 meeting of the American Society for Mass Spectrometry. At the time of the workshop about half the sequence had been determined correctly from the secondary ion spectrum with the aid of enzymatic digestions carried out on the target. Subsequent measurements have defined the remainder, apart from a couple of uncertainties.

Secondary ion mass spectrometry of oligopeptides

Abstract Positive and negative secondary ion mass spectra produced by a beam of 8 to 28 keV Cs+ ions have been observed for underivatized oligopeptides with molecular weights up to 1347. Each compound gave prominent ions characteristic of the intact molecule, i.e. [M+H]+, [M+Na]+, [M-H]- and “sequence-characteristic” ions progressing from either end of the molecule. Many of the observed ions are metastable, so the relative intensities observed in the time-of-flight spectrometer used in these measurements may be different from those measured in sector-field or quadrupole instruments.

Ion-Neutral Correlations Following Metastable Decay

An important property of linear time-of-flight (TOF) mass spectrometers is their high efficiency for detecting the products of metastable decay [1,2]. Ions that decay in the field-free drift region contribute to the parent ion peak because the centre-of-mass velocity remains constant. However, the normal TOF spectrum provides no information on metastable decay paths or rate constants. Such data can contribute to structural elucidation and understanding of the desorption process.

Study of rearrangement reactions occurring during gas chromatography of tert.-butyldimethylsilyl ether derivatives of uridine

The gas chromatography of partial O-tert.-butyldimethylsilyl (TBDMS) derivatives of uridine is complicated by the occurrence of a 2′ ↔ 3′ rearrangement of the TBDMS group when uridine derivatives having a 2′- or 3′-O-TBDMS group and an underivatized OH group in the 3′- or 2′-position are injected into a gas chromatograph. When a 5′-O-TBDMS group is also present these rearrangements are accompanied by a thermal decomposition thought to involve elimination of methane. Further derivatization by acetylation or trimethylsilylation (but not trifluoroacetylation), both of which prevent the rearrangement and decomposition reactions, together with gas chromatography-mass spectrometry selected ion recording techniques, is suggested as an analytical procedure for the synthetically useful partial TBDMS derivatives of ribonucleosides.