Please cite this article in press as: A. Filippi, et al., Int. J. Mass Spectrom. (2013), http://dx.doi.org/10.1016/j.ijms.2013.05.016 ARTICLE IN PRESS G Model MASPEC-14939; No. of Pages 8 International Journal of Mass Spectrometry xxx (2013) xxx–xxx Contents lists available at SciVerse ScienceDirect International Journal of Mass Spectrometry j ourna l ho me page: www.elsevier.com/locate/ijms Protonated pyrimidine nucleosides probed by IRMPD spectroscopy Antonello Filippi a , Caterina Fraschetti a , Flaminia Rondino a , Susanna Piccirillo b , Vincent Steinmetz c , Leonardo Guidoni d , Maurizio Speranza a,∗ a Dipartimento di Chimica e Tecnologie del Farmaco, Sapienza-Università di Roma, P.le A. Moro 5-00185 Roma, Italy b Dipartimento di Scienze e Tecnologie Chimiche, Università di Roma “Tor Vergata”, Rome, Italy c Laboratoire Chimie Physique, UMR8000 CNRS, Université Paris Sud 11, Orsay, France d Dipartimento di Scienze Fisiche e Chimiche, Università degli Studi dell’Aquila, L’Aquila, Italy a r t i c l e i n f o Article history: Received 1 March 2013 Received in revised form 14 May 2013 Accepted 15 May 2013 Available online xxx In the memory of Detlef Schroeder. Keywords: RNA/DNA pyrimidine nucleosides Protomers IRMPD spectroscopy Conformational preference Ab initio MD simulations a b s t r a c t The ESI-formed protonated 2  -deoxycytidine, cytidine, cytarabine, and gemcitabine have been probed using infrared multiphoton dissociation (IRMPD) spectroscopy performed in the 900–2000 cm −1 region at CLIO, the Orsay Free Electron Laser facility, and in the 2800–3800 cm −1 region using a YAG-laser coupled to a table-top optical parametric oscillator/amplifier (OPO/OPA). The IRMPD spectra are compared of the protonated nucleosides with the IR spectra of their B3LYP/6-311++G(d,p)-optimized isomeric forms. The stability at room temperature of some conformers has been investigated by means of ab initio molecular dynamics simulations. The IRMPD spectra are consistent with the formation in the ESI source of both the N3- and the O2-protonated nucleosides. The most favoured members of both families are characterized by the pyrimidine base oriented anti to the furanose moiety. Concerning the O2-protonated nucleosides, IRMPD spectra and thermochemical considerations support the predominant formation of the structures with the proton oriented up relative to the furanose moiety. © 2013 Elsevier B.V. All rights reserved. 1. Introduction Nucleosides are fundamental DNA/RNA components [1] which exhibit a variety of specialized functions including long-range electron transport over the RNA and DNA molecules and repair mechanism after their radiation damage [2,3]. Their structure con- sists in a free nucleobase linked to a furanose-type ring (sugar) by the N-glycosidic bond. Besides the RNA nucleosides adenosine, guanosine, cytidine, thymidine, and uridine, numerous naturally occurring and chemically synthesized or modified nucleosides (nucleoside analogues) do exist which are used in medicinal and pharmaceutical sciences as prodrugs [4–6]. Compounds with a wide variety of modifications of the furanose ring have been synthesized and tested for activities. Nucleoside analogues are cytotoxic and have found expanding therapeutic use as antiviral and antitumor agents and most antimetabolites of nucleoside ana- logues possess the skeletal chemical structure of cytidine, such as cytarabine and gemcitabine (Fig. 1) [1,7]. Assessment of the dynamical structure and conformation of nucleosides and of their physicochemical properties is a funda- mental requirement for unravelling their intimate mechanism of functioning in living matter. Cleavage of the N-glycosidic bond is ∗ Corresponding author. Tel.: +39 06 4991 3497; fax: +39 06 4991 3602. E-mail address: maurizio.speranza@uniroma1.it (M. Speranza). a repair mechanism for damaged RNA and DNA molecules [8–13] which has been found to be sensitive to the conformation of the furanose ring [14] as well as to environmental and chemical agents, such as pH, metal cations, and alkylating compounds [15–19]. The nucleobase is available at prototropic equilibria which may favour on point mutations during the replication of the nucleic acid [20,21]. Protonated nucleobases are known to be involved in RNA catalysis [22]. The location of the protonation site of the nucle- obases plays a role in the stabilization of triplex structures as well [23]. In this frame, gas-phase studies on protonated nucleosides can appear of some interest, since they may provide precious information on the intrinsic properties of the selected species by eliminating the influence from solvent and counter-ions and any conformational ensemble averaging effects. Besides, gas-phase studies allow for a simpler direct comparison between experimen- tal results and quantum mechanical calculations. Mass spectrometry (MS), coupled to high-level theoretical calculations, is the method of choice to characterize charged species in the dilute gas state and to investigate their behaviour towards specific reactants. The gas-phase thermochemistry of fun- damental DNA/RNA components has been the matter of intense investigation by MS. Despite several MS approaches have been employed to measure the gas-phase basicity and proton affinity of nucleobases [24–28] and nucleosides [28–34], positive infor- mation on the actual structure of their protonated forms remains 1387-3806/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.ijms.2013.05.016