2 PERKIN 498 J. Chem. Soc., Perkin Trans. 2, 2001, 498–506 DOI: 10.1039/b009019k This journal is © The Royal Society of Chemistry 2001 Collisionally-induced dissociation mass spectra of organic sulfate anions Athula B. Attygalle,* Silvina García-Rubio, Jennifer Ta and Jerrold Meinwald Baker Laboratory, Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853, USA Received (in Cambridge, UK) 8th November 2000, Accepted 18th January 2001 First published as an Advance Article on the web 1st March 2001 The collisionally-induced dissociation mass spectra of a variety of organic sulfate ester anions are described and mechanistically rationalized. A cyclic syn-elimination pathway, analogous to that of the Cope elimination, is postulated for the commonly observed formation of bisulfate anion (HSO 4  , m/z 97). Deuterium labeling experiments confirm that the proton transferred to oxygen during bisulfate elimination normally originates from the C-2 position, although examination of the spectra of polyfunctional steroids reveals that the proton abstracted may originate from more distant sites as well. Adamantyl, phenyl, and vinyl sulfate anions, which do not readily lend themselves to a cyclic syn-elimination, do not give rise to an m/z 97 ion. Instead, these sulfates undergo both heterolytic and homolytic S–O bond cleavages to yield an m/z M  80 anion, resulting from loss of neutral SO 3 , as well as an ion at m/z 80, corresponding to SO 3  , respectively. Sulfates that can give rise to a resonance stabilized radical by homolytic C–O bond fission, as exemplified by benzyl and linalyl sulfates, can be recognized by the formation of an m/z 96 (SO 4  ) ion. Introduction Sulfate esters constitute a widespread and highly diverse group of natural and non-natural compounds. The resonance- stabilized nature of sulfate mono-ester anions underlies their unique chemistry, providing an electrostatic component to specific interactions without giving rise to significantly basic or nucleophilic behavior. The industrial importance of anionic surfactants such as sodium dodecyl sulfate (SDS) has tended to overshadow the significance of the increasing recognized num- ber of biologically active natural products that contain a sulfate moiety. While once believed to occur exclusively as metabolites of marine organisms 1 (i.e. iejimalide D, 1, from the tunicate Eudistoma cf. rigida, 2 and squalamine, 2, from the dogfish shark 3 ), sulfate esters are now known from many other sources (e.g. uzarigenin 3-sulfate, 3, from the Ranunculaceae plant Adonis aleppica 4 ). 5 In addition, knowledge of the enzymes that catalyze sulfate ester synthesis has grown exponentially during the last decade. 6 The formation of sulfate esters 7 is an import- ant step in the elimination 8 or bioactivation 9 of xenobiotics and drugs, and in the biotransformations of many hormones and neurotransmitters. 6a,b,c,e Recent studies of sulfated carbo- hydrates have revealed that many of these compounds populate extracellular spaces, and mediate a diverse range of events that contribute to intercellular recognition in both normal and pathological processes. 10 Since encountering the first glycosylated nucleoside sulfate ester HF-7 (4) in the venom of a funnel-web spider, 11 we became interested in the possibility that similar anionic sulfate esters might have gone undetected in neurotoxic venoms. The ideal technique for searching for such compounds is clearly mass spectrometry. The facile ionization of sulfates obviously renders them amenable to negative-ion mass spectrometry. Par- ticularly in conjunction with electrospray ionization (ESI) and liquid chromatography (LC), mass spectrometry (MS) has already provided a powerful approach to the characterization of sulfated compounds in biological fluids. 12–16 Nevertheless, although several interesting features have been noted in the mass spectra of these sulfate esters, 12,14,15,17 little attention has been paid to the details of their fragmentation mechanisms. A prominent product ion peak clearly attributable to HSO 4  has been observed at m/z 97 in most spectra of organic sulfates. 17 The formation of this anion requires the transfer of a proton to oxygen, in addition to the cleavage of a C–O bond. The origin of this transferred proton, however, has not been established,