Steven Daly

Alexander Kulesza

Geoffrey Knight

Luke Macaleese

Rodolphe Antoine

Philippe Dugourd

The Gas-Phase Photophysics of Eosin Y and its Maleimide Conjugate

ABSRACT:

The use of the xanthene family of dyes as fluorescent probes in a wide range of applications has provided impetus for the studying of their photophysical properties. In particular, recent advances in gas-phase techniques such as FRET that utilize such chromophores have placed a greater importance on the characterization of these properties in the gas-phase. Additionally, the use of synthetic linker chains to graft the chromophores in a site-specific manner to their target system is ubiquitous. There is, however, often limited information on how the addition of such a linker chain may affect the photophysical properties of the chromophores, which is of fundamental importance for interpretation of experimental data reliant on grafted chromophores. Here, we present data on the optical spectroscopy of different protonation states of Eosin Y -a fluorescein derivative. We compare the photophysics of Eosin Y to its maleimide conjugate, and to the thioether product of the reaction of this conjugate with cysteamine. Comparison of the mass spectra following laser irradiation shows that very different relaxation takes place upon addition of the maleimide moiety, but that the photophysics of the bare chromophore are restored upon addition of cysteamine. This radical change in the photophysics is interpreted in terms of charge-transfer states, whose energy relative to the S 1  S 0 transition of the chromophore is dependent on the conjugation of the maleimide moiety. We also show that the shape of the absorption band is unchanged in the gas-phase as compared to the solution-phase, showing a maximum with a shoulder towards the blue, and examination of isotope distributions of the isolated ions show that this shoulder cannot be due to the presence of dimers. Consideration of the fluorescence emission spectrum allows a tentative assignment of the shoulder to be due to a vibrational progression with a high Frank-Condon factor.

Introduction

The xanthene-based dye family -which includes the rhodamines and fluorones -are almost ubiquitous as fluorescent probes with a high quantum yield in a vast range of subject areas, and as such there has been a wealth of studies probing their solution phase properties. [START_REF] Arbeloa | Photophysics of Rhodamines -Molecular-Structure and Solvent Effects[END_REF][START_REF] Arbeloa | Luminescence Properties of Rhodamines in Water-Ethanol Mixtures[END_REF][START_REF] Arden | Fluorescence and Lasing Properties of Rhodamine Dyes[END_REF][START_REF] Magde | Solvent Dependence of the Fluorescence Lifetimes of Xanthene Dyes[END_REF][START_REF] Karstens | Rhodamine-B and Rhodamine-101 as Reference Substances for Fluorescence Quantum Yield Measurements[END_REF] Eosin Y (EY), one such xanthene derivative, has been studied particularly intensively since it has found a use as a probe for trace amounts of peptide, as a source of singlet molecular oxygen, and as an absorber in dye-sensitized photo-cells. [START_REF] Waheed | Estimation of Submicrogram Quantities of Protein Using the Dye Eosin Y[END_REF][START_REF] Waheed | Mechanism of Dye Binding in the Protein Assay Using Eosin Dyes[END_REF][START_REF] Amatguerri | Singlet Oxygen Photogeneration by Ionized and Un-Ionized Derivatives of Rose-Bengal and Eosin Y in Diluted Solutions[END_REF][START_REF] Tai | Sensitized Nanostructured SnO2/TiO2 Solar Cells[END_REF][START_REF] Kim | Improved Performance of a Dye-Sensitized Solar Cell Using a TiO2/ZnO/Eosin Y Electrode[END_REF][START_REF] Jin | 5.1% Apparent Quantum Efficiency for Stable Hydrogen Generation over Eosin-Sensitized CuO/TiO2 Photocatalyst under Visible Light Irradiation[END_REF] The solution phase absorption spectroscopy of Eosin Y has been studied in some detail as a function of the solvent or pH. 12- 15 Eosin Y can exist in a number of different protonation states in solutions, ranging from singly protonated [EY+H] + to double deprotonated [EY-2H] 2-due to the presence of both phenolic and carboxylic acid moieties, Figure 1.

Batistela et al. found second and third pK a values of 2.02 and 3.80 to yield [EY-H] -and [EY-2H] 2-respectively. [START_REF] Batistela | pKa Determinations of Xanthene Derivates in Aqueous Solutions by Multivariate Analysis Applied to UV-Vis Spectrophotometric Data[END_REF] By studying the absorption spectroscopy in H 2 O as a function of the pH, they were able to extract values of the absorption maximum of 470 nm, 519 nm and 515 nm for [EY], [EY-H] -and [EY-2H] 2-respectively. The profile of the absorption spectrum for the negatively charged species shows a main peak with a shoulder at higher excitation energies. The origin of this shoulder is most commonly posited to be due to the presence of dimers. [START_REF] Chakraborty | Spectral Behaviour of Eosin Y in Different Solvents and Aqueous Surfactant Media[END_REF][START_REF] Chambers | Effect of Dimer Formation on Electronic Absorption and Emission-Spectra of Ionic Dyes -Rhodamines And Other Common Dyes[END_REF][START_REF] De | Environmental Effects on the Aggregation of Some Xanthene Dyes Used in Lasers[END_REF][START_REF] Suradkar | CMC Determination of an Odd Carbon Chain Surfactant (C13E20) Mixed with Other Surfactants Using a Spectrophotometric Technique[END_REF] They also found a pronounced change in the absorption band as the pH is increased from 0.31 to 2.95, which they attributed to the coexistence of multiple charge states in the solution. This coexistence of charge states makes the interpretation of solution phase spectroscopy difficult; particularly since the absorption maxima are close -making their deconvolution difficult.

Alternative to studying pH-dependent absorption in solution, the use of mass spectrometry allows the isolation and subsequent measurement of absorption properties by actionspectroscopy of different charge states independently. The gas-phase spectroscopy of xanthene derivatives has become a common technique in the past decade, and in particular the absorption and fluorescence properties as related to gas-phase Förster Resonance Energy Transfer (FRET). [START_REF] Daly | Visible and Ultraviolet Spectroscopy of Gas Phase Rhodamine 575 Cations[END_REF][START_REF] Sagoo | The Fluorescence Properties of Cationic Rhodamine B in the Gas Phase[END_REF][START_REF] Bian | Gas-Phase Fluorescence Excitation and Emission Spectroscopy of Mass-Selected Trapped Molecular Ions[END_REF][START_REF] Yao | Fluorescence and Electronic Action Spectroscopy of Mass-Selected Gas-Phase Fluorescein, 2′,7′-Dichlorofluorescein, and 2′,7′-Difluorofluorescein Ions[END_REF][START_REF] Mcqueen | On the Intrinsic Photophysics of Fluorescein[END_REF] Moreover, Eosin Y is potentially attractive as an acceptor chromophore for studies of anion action-FRET since electron detachment has been shown to be an important mechanism in related dyes. [START_REF] Yao | Fluorescence and Electronic Action Spectroscopy of Mass-Selected Gas-Phase Fluorescein, 2′,7′-Dichlorofluorescein, and 2′,7′-Difluorofluorescein Ions[END_REF][START_REF] Mcqueen | On the Intrinsic Photophysics of Fluorescein[END_REF] Additionally, the characterization of any changes in photophysical properties of chromophores following conjugation to a synthetic linker chainwhich is used for grafting onto specific sites in proteins or other systems -is vital for interpretation of FRET data. The maleimide moiety is one such linker. It reacts specifically with thiols to form a thioether product, and has been previously shown to influence the photophysics of chromophores, acting as a fluorescence quencher. [START_REF] Guy | Convergent Preparation and Photophysical Characterization of Dimaleimide Dansyl Fluorogens: Elucidation of the Maleimide Fluorescence Quenching Mechanism[END_REF] In this paper, we aim to unravel the mechanisms leading to the different charge-state dependent photo-processes in conjugated chromophore-maleimide molecules prior to and following the grafting reaction. We address this question by solution-phase absorption measurements of Eosin Y, gas-phase action-spectroscopy measurements on the mono-and dianions of Eosin Y (EY), its maleimide conjugate 5-maleimido-Eosin Y (EYM), and the product of the reaction of EYM with cysteamine (EYM-CA) and excited state computations. The experimental setup used to perform the experiments presented in this paper has been described in detail previously [START_REF] Daly | Visible and Ultraviolet Spectroscopy of Gas Phase Rhodamine 575 Cations[END_REF] . Briefly, a linear quadrupole ion trap mass spectrometer (LTQ Velos, Thermo Fisher Scientific, San Jose, CA) was used to generate, mass select, and trap ions in a first, high pressure trap for a controlled period during which time they may be activated by laser irradiation (Laser Induced Dissociation, LID). Fragment ions are subsequently transmitted to a second, lower pressure ion trap, and are mass analyzed. A fused silica window (3 mm thick, 1 in. diameter) is positioned at the back end of the instrument, and 1-2 mm diameter circular openings in the trapping electrodes allow coupling of light source and trapped ions. In order to optimize laser transmission, the central hole of the electrode closest to the fused silica window was enlarged to 5 mm in diameter.

Method

The light source used was a Horizon OPO pumped by the third harmonic of a Surelite II Nd:YAG laser (Continuum, Santa Clara, CA). A repetition rate of 10 Hz and a pulse width of 5 ns were used. The beam was focused into the first ion trap using a 1000 mm focal length convergent lens. A mechanical shutter, synchronized with the mass spectrometer, was used to stop the beam at all times except the "ion activation window", that is, the time after ion accumulation and before the mass analysis. A single laser pulse was used for the irradiation of the trapped ions, and when irradiating ions the normalized collision energy is kept at zero. To record action spectra, the laser wavelength was scanned between 590 and 420nm, in 0.1 nm steps and with a 1 s dwell time at each wavelength. During the scan, each data point is the average of 9 mass spectra, which corresponded to on average one data point per second. Laser power was recorded by performing a second scan under identical conditions immediately following acquisition of the action spectrum. To analyze the data, each mass spectrum was assigned a wavelength value given by 𝜆 𝑠𝑡𝑎𝑟𝑡 -((𝑖 -1) * ( Δ𝜆 𝑛 )) where 𝜆 𝑠𝑡𝑎𝑟𝑡 = 590 nm is the starting wavelength, 𝑖 the mass spectrum number, Δ𝜆 = 170 nm is the total wavelength change, and 𝑛 the total number of mass spectra acquired during the acquisition. The same procedure was used to provide a wavelength range for the laser power (P), and a linear interpolation performed to give values of the laser power at the wavelengths assigned to each mass spectrum. The action spectrum is defined as -log(𝐼 𝑝𝑎𝑟𝑒𝑛𝑡 𝐼 𝑡𝑜𝑡𝑎𝑙 ⁄ ) (𝜆 * 𝑃) ⁄ where 𝐼 𝑝𝑎𝑟𝑒𝑛𝑡 and 𝐼 𝑡𝑜𝑡𝑎𝑙 are the parent and total ion signal respectively at each point in the acquisition.

Solution phase spectroscopy was performed on Eosin Y diluted to 5 µM in H 2 O, and the pH was controlled by addition of either ammonium hydroxide or acetic acid. Absorption spectra were performed using an Avantes AvaLight DHS light source and an AvaSpec 2048 spectrometer. The fluorescence excitation spectra were measured on a Fluoromax-4 spectrophotometer (Horiba) by monitoring the fluorescence at 555 nm. The fluorescence emission spectra were measured on the same instrument following excitation at 520 nm.

Theoretical

Gas-phase optimizations and excited state calculations were performed using density functional theory (DFT) and time-dependent DFT (TDDFT) as implemented in Gaussian09

Rev D.01. 25 The hybrid CAM-B3LYP [START_REF] Becke | Density Functional Thermochemistry III The Role of Exact Exchange[END_REF][START_REF] Perdew | Generalized Gradient Approximation Made Simple[END_REF][START_REF] Yanai | A New Hybrid Exchange-Correlation Functional Using the Coulomb-Attenuating Method (CAM-B3LYP)[END_REF] exchange-correlation functional that provides long-range corrections with the Coulomb attenuation method was used. For all atoms, Ahlrichs triple-ζ plus polarization atomic basis sets were used (def2-TZVP). Optimizations included Grimme's empirical dispersion correction including Becke-Johnson damping D3(BJ) [START_REF] Grimme | Effect of the Damping Function in Dispersion Corrected Density Functional Theory[END_REF][START_REF] Grimme | A Consistent and Accurate Ab Initio Parametrization of Density Functional Dispersion Correction (DFT-D) for the 94 Elements H-Pu[END_REF].

Results

Eosin Y (EY) and 5-maleimido-Eosin Y (EYM) -Figure 1 -both contain two possible deprotonation sites; the phenolic acid and carboxylic acid moieties. Previous measurement of EY in solution phase has shown the former as having the lower pK a value (2.02) compared to the later (3.80). [START_REF] Batistela | pKa Determinations of Xanthene Derivates in Aqueous Solutions by Multivariate Analysis Applied to UV-Vis Spectrophotometric Data[END_REF] As noted above, given that EY likely exists in multiple charge states in solution, it is difficult to measure the absorption spectrum of individual charge state.

However, it is possible in solution to produce predominantly monoanions or dianions of EY by controlling the pH of the solution. [START_REF] Batistela | pKa Determinations of Xanthene Derivates in Aqueous Solutions by Multivariate Analysis Applied to UV-Vis Spectrophotometric Data[END_REF] As such, the solid blue curves in Figure 2 shows the solution phase spectra of EY in H 2 O at pH 2.53 (a) and 11.87 (b), which have been shown to produce predominantly [EY-H] -and [EY-2H] 2-respectively. The absorption band shows a similar profile under both conditions, with an absorption maximum at 520 nm for the pH 2.53 solution and 517 nm for the pH 11.87 solution, and a shoulder is observed for both solutions at 492 nm, Table 1. The conventional explanation for the presence of this shoulder in xanthene derivatives is the presence of dimers. [START_REF] Chakraborty | Spectral Behaviour of Eosin Y in Different Solvents and Aqueous Surfactant Media[END_REF][START_REF] Chambers | Effect of Dimer Formation on Electronic Absorption and Emission-Spectra of Ionic Dyes -Rhodamines And Other Common Dyes[END_REF][START_REF] De | Environmental Effects on the Aggregation of Some Xanthene Dyes Used in Lasers[END_REF][START_REF] Suradkar | CMC Determination of an Odd Carbon Chain Surfactant (C13E20) Mixed with Other Surfactants Using a Spectrophotometric Technique[END_REF] These values are in good agreement with the values of the absorption maximum determined by Batistela et al. of 519 and 515 nm respectively for [EY-H] -and [EY-2H] 2-respectively. [START_REF] Batistela | pKa Determinations of Xanthene Derivates in Aqueous Solutions by Multivariate Analysis Applied to UV-Vis Spectrophotometric Data[END_REF] Upon electrospray ionization of EY and EYM in H 2 O, the resulting mass spectrum shows both mono-and dianionic species. 531 nm, whilst [EYM-H] -shows a single peak at 532 nm with a shoulder at 507 mn (Table 1).

Similar absorption profiles to that of [EY-H] -have been observed in previous action-spectroscopy studies of fluorescent xanthene dyes, and can be interpreted in terms fluorescence rather than fragmentation being the dominant relaxation mechanism following photoexcitation. [START_REF] Daly | Visible and Ultraviolet Spectroscopy of Gas Phase Rhodamine 575 Cations[END_REF][START_REF] Daly | Action-FRET: Probing the Molecular Conformation of Mass-Selected Gas-Phase Peptides with Förster Resonance Energy Transfer Detected by Acceptor-Specific Fragmentation[END_REF] The fluorescence quantum yield may depend strongly on the excitation energy; low excitation energies leading to ions with minimal vibrational excitation in the excited state is where the fluorescence quantum yield tends to be highest. As the energy is increased, the increased vibrational energy of the excited state can lead to an increased rate of non-fluorescent relaxation pathways -such as direct photofragmentation or an increased rate of internal conversion or intersystem crossing.

The fluorescence quantum yield of EY has been determined to be between 0.15 and 0.22 in water, 0.44 and 0.63 in methanol and 0.4 and 0.72 in ethanol. [START_REF] Penzkofer | Intersystem-Crossing and Excited-State Absorption in Eosin Y Solutions Determined by Picosecond Double Pulse Transient Absorption Measurements[END_REF] Additionally, measurements of the gas-phase fluorescence emission spectrum of various halogenated fluorescein derivatives have shown that the monoanions possess a weak but measureable fluorescence quantum yield, and indicated that increased halogenation of fluorescein leads to an increase in the gas-phase fluorescence. [START_REF] Yao | Fluorescence and Electronic Action Spectroscopy of Mass-Selected Gas-Phase Fluorescein, 2′,7′-Dichlorofluorescein, and 2′,7′-Difluorofluorescein Ions[END_REF] We can thus assign the band at 531 nm in the action spectrum of [EY-H] -to be the true gas-phase absorption maximum, where the fluorescence quantum yield is high and hence other relaxation channels which may lead to fragmentation are suppressed. Similarly, we assign the band at 498 nm to be the shoulder observed in the solution-phase absorption spectrum. This assignment is supported by the [EYM-H] -action spectrum, where the addition of the maleimide moiety is known to cause a fluorescence quenching effect. As such, the shape of the absorption band closely resembles that seen in solution, and the absorption maximum of 532 nm is very close in energy to the first band in [EY-H] -.

This difference in photophysical behavior with addition of the maleimide moiety is even starker when the dianions are considered. It has been previously observed that dianionic fluorescein and its derivatives are non-fluorescent in the gas phase, and instead relax via photodetachment. [START_REF] Yao | Fluorescence and Electronic Action Spectroscopy of Mass-Selected Gas-Phase Fluorescein, 2′,7′-Dichlorofluorescein, and 2′,7′-Difluorofluorescein Ions[END_REF][START_REF] Mcqueen | On the Intrinsic Photophysics of Fluorescein[END_REF] occurring in the two systems. It is known that the maleimide moiety is able to act as a fluorescence quencher of chromophores, and that upon reaction with a sulfhydryl group to form a thioalkyl succinimide this quenching effect is lost. [START_REF] Guy | Convergent Preparation and Photophysical Characterization of Dimaleimide Dansyl Fluorogens: Elucidation of the Maleimide Fluorescence Quenching Mechanism[END_REF] Guy et al. posited that the quenching mechanism is a photo-induced electron transfer from the chromophore to the maleimide, which is consistent with the relaxation mechanism observed here being in direct competition with photo-detachment, a process that occurs on the femtosecond to picosecond timescale. [START_REF] Guy | Convergent Preparation and Photophysical Characterization of Dimaleimide Dansyl Fluorogens: Elucidation of the Maleimide Fluorescence Quenching Mechanism[END_REF] In order to determine that the observed quenching was quantitatively similar to that The results of this analysis are given in Figure 4 (see also Table S1 in the SI). CT states can be identified by inspection of the excitations between occupied Kohn-Sham orbitals φ i and unoccupied ones φ a . Contrary to the local π-π* excitation (e.g. S 1 for [EY-2H] 2-in Figure 4), CT excitations involve orbitals localized at different moieties. This locality can be quantified using the spatial overlap O ia defined as the inner product of the moduli of the two orbitals φ i and φ a (See SI, Table S1, used also in Tozer's diagnostic test for charge transfer states). It is thus possible to posit an explanation for differences in photophysical behavior of EY attached to either maleimide or succinimide moieties. In [EYM+CA-2H] 2-the excitation is localized on the xanthene moiety and photo-detachment is likely to occur from here, producing a single radical center. In [EYM-2H] 2-, the fast internal conversion to the CT state followed by photo-detachment will lead to the radical center being in a different location, which may give a more reactive monoanion radical species and hence lead to enhanced unimolecular fragmentation of the resulting system.

Finally, it is interesting to consider the origin of the shoulder which is observed in both solution-and gas-phase spectra. This shoulder is commonly attributed to the presence of dimers in solution-phase absorption spectroscopy, but in the gas-phase it is possible to unambiguously check for the presence of dimers. As the mass-to-charge ratio of, for example,

[EYM-nH] n-and [EYM-nH] 2 2n-will be the same, in order to discount the possibility of there being any dimers present during our isolation step it is important to examine the isotopic distribution we obtain when isolating both [EYM-H] -and [EYM-2H] 2-. Figure 5 There is no evidence for the presence of dimers during the isolation of either species, and thus the origin of the shoulder in the gas-phase action spectra cannot be attributed to the presence of dimers. This is consistent with previous gas phase measurements of rhodamine derivatives, where similar profiles to those observed in solution are also found, and indeed with other poly-aromatic dyes, indicating this may be a more general phenomenon. [START_REF] Sagoo | The Fluorescence Properties of Cationic Rhodamine B in the Gas Phase[END_REF][START_REF] Yao | Fluorescence and Electronic Action Spectroscopy of Mass-Selected Gas-Phase Fluorescein, 2′,7′-Dichlorofluorescein, and 2′,7′-Difluorofluorescein Ions[END_REF][START_REF] Mcqueen | On the Intrinsic Photophysics of Fluorescein[END_REF][START_REF] Loison | Microsolvation Effects on the Optical Properties of Crystal Violet[END_REF][START_REF] Drink | Optical Properties of Prodigiosin and Obatoclax: Action Spectroscopy and Theoretical Calculations[END_REF] Additionally, gas-phase action spectroscopy performed upon pseudo-dimers of rhodamine 575, where the dimerization is created by enforced proximity of the dyes following grafting onto a tri-peptide, shows there is only a small shift in the position of the absorption maximum relative to the monomeric species. [START_REF] Daly | Action-FRET: Probing the Molecular Conformation of Mass-Selected Gas-Phase Peptides with Förster Resonance Energy Transfer Detected by Acceptor-Specific Fragmentation[END_REF] Moreover, recent solution phase measurements of pyronin B dimers in solution found their absorption and fluorescence properties to differ only very slightly from the monomer. [START_REF] Zhang | The Fluorescence Properties of Three Rhodamine Dye Analogues: Acridine Red, Pyronin Y and Pyronin B[END_REF] Where then does the origin of this shoulder then lie? The possibility of their being a second singlet excitation close to the S 1 S 0 transition, or that there be a second tautomer (for example the lactone tautomer which is found in solution for xanthene dyes with underivatized carboxylic acid moieties) can be discounted by examination of the fluorescence spectrum.

Following excitation at the absorption maximum of EY at pH 2.43 and 11.87, the fluorescence emission spectrum possesses a red shoulder in both cases (Figures S1 and S2

respectively). Similar behavior is found in a range of different xanthene-based chromophores, in both solution-and gas-phases, strongly indicating that the shoulder is an intrinsic property of the S 1  S 0 transition. [START_REF] Zhang | Fluorescence Properties of Twenty Fluorescein Derivatives: Lifetime, Quantum Yield, Absorption and Emission Spectra[END_REF][START_REF] Sagoo | The Fluorescence Properties of Cationic Rhodamine B in the Gas Phase[END_REF][START_REF] Forbes | Gas-Phase Fluorescence Excitation and Emission Spectroscopy of Three Xanthene Dyes (Rhodamine 575, Rhodamine 590 and Rhodamine 6G) in a Quadrupole Ion Trap Mass Spectrometer[END_REF][START_REF] Klonis | Spectral Properties of the Prototropic Forms of Fluorescein in Aqueous Solution[END_REF][START_REF] Sjoback | Absorption and Fluorescence Properties of Fluorescein[END_REF] The fact that the mirror-image rule is obeyed by xanthene derivatives and that the Stoke's shift is small (21 nm for both mono-and di-anions of Eosin Y measured here) indicates that the ground and excited states geometries are similar, and that the shoulder may be ascribed to vibronic transitions in either absorption or emission spectra with strong Franck-Condon factors, see Figure S3.

There are previous solution-and gas-phase measurements of the ground state vibrations of fluorescein and some of its halogenated derivatives such as EY in solution by FTIR and Raman spectroscopy, and in the gas-phase by IRMPD. [START_REF] Narayanan | Vibrational Sprectral Analysis of Eosin Y and Erytheosin B -Intensity Studies for Quantiative Detection by Dyes[END_REF][START_REF] Wang | Raman and FTIR Spectroscopies of Fluorescein in Solution[END_REF][START_REF] Yao | Infrared Multiple Photon Dissociation Action-Spectroscopy and Computational Studies of Mass-Selected Gas-Phase Fluorescein and 2',7'-Dichlorofluorescein Ions[END_REF] Narayaman et al. found that the FTIR of EY in solution contains many intense absorption features between 1000 and 2000 cm -1 . Yao et al. measured gas-phase fluorescein and 2',7'-dichlorofluorescein by IRMPD action spectroscopy and found a similar series of intense vibrational transitions in the same energy range. They also performed vibrational calculations to assign the observed bands to normal modes, and found that the most intense bands can be assigned to xanthene-based vibrational modes. [START_REF] Yao | Infrared Multiple Photon Dissociation Action-Spectroscopy and Computational Studies of Mass-Selected Gas-Phase Fluorescein and 2',7'-Dichlorofluorescein Ions[END_REF] That the shoulder may be due to a vibrational progression based on the xanthene ring system is supported by the fact that the S 1 absorption profile of xanthene derivatives generally exhibit similar profiles, which would indicate that the origin of the shoulder lies in a common element of this dye family. Particularly, the presence of a shoulder in the solution phase absorption spectrum of acridine red -which contains only the xanthene moiety with no carboxyphenyl moiety -provides support for this assignment. [START_REF] Zhang | The Fluorescence Properties of Three Rhodamine Dye Analogues: Acridine Red, Pyronin Y and Pyronin B[END_REF] Hence, it seems that the shape of the S 1 absorption band of xanthene derivatives can be tentatively assigned as being due to the vibrational profile of the S 1 ← S 0 transition.

Conclusions

The from the EY to the maleimide π systems and enables alternative deactivation channels to be accessed. This situation is only present with the maleimide providing an exo-energetic electron attachment energy, while reaction with cysteamine no longer allows this "pull"-effect to take place, and thus quenches this electron-transfer. This correlates with the observation that upon grafting of cysteamine to EYM, the photophysical properties of the resultant species are very similar to those of the bare chromophore. These observations are of high importance in all situations where such linker chains are used for specific grafting of chromophores, where the photophysics of the chromophore is of paramount importance for interpretation of data.

The solution-phase absorption spectrum and gas-phase action spectrum both show an absorption maximum with a shoulder to higher energy. The isotope distribution for massselected anion and dianion EY derivatives in the gas-phase showed no evidence of dimers, to which this shoulder is commonly attributed, which in turn indicates that the presence of dimers is equally unlikely to be the origin of the shoulder in the solution-phase. Analysis of the fluorescence emission spectrum following excitation at the absorption maximum in solution indicated that the shoulder is an intrinsic property of the S1  S0 transition, and thus could be tentatively assigned to be due to a vibrational progression with a high Franck-Condon factor, with xanthene based vibrational normal modes potentially implicated. This enhanced understanding may be of importance in solution phase where xanthene derivatives are ubiquitous in their use as fluorescent reporters.

ASSOCIATED CONTENT

Details of the TDDFT calculations, and solution phase fluorescence excitation and emission spectra for Eosin Y at pH 2.53 and pH 11.87 may be found in the supplementary information.

This material is available free of charge via the Internet at http://pubs.acs.org.

Experimental 2 mg

 2 of Eosin Y (EY) (Sigma Aldrich) and 5-maleimido-Eosin Y (EYM) (Sigma Aldrich) were independently dissolved in 200 µL DMSO before subsequent dilution in 5 ml H 2 O to a final concentration of 30 µM for use in the electrospray source. 8.2 mg of Cysteamine hydrochloride was dissolved in 1 ml H 2 O at a concentration of 7.2 mM. For the reaction of EYM with cysteamine, an equimolar solution of the two reactants was prepared in H 2 O at a concentration of 30 µM and left to react at room temperature for 1 hour, after which it was directly used in the electrospray source.

Figure 2

 2 shows the gas phase action spectra of mass selected (a) [EY-H] -and [EYM-H] -and (b) [EY-2H] 2-and [EYM-2H] 2-as black squares and red circles respectively. There is a clear difference in the action spectrum measured for the two species, with [EY-H] -showing a broad absorption band with two features at 498 nm and

  observed by Guy et al., EYM was reacted with cysteamine to give the thioether EYMcysteamine product [EYM+CA], Figure 1. The action spectra for the mono-and dianion is shown in Figure 2, and the LID mass spectrum of the dianion at the absorption maximum are shown in panel (c) of Figure 3. The LID mass spectrum shows that photo-detachment is again the dominant relaxation channel of [EYM+CA-2H] 2-, closely resembling the case of [EY-2H] 2-, rather than [EYM-2H] 2-. This indicates that the quenching mechanism of the maleimide group is significantly reduced upon reaction of the maleimide with a thiol -as observed by Guy et al. -although some fragmentation pathways are still observed. Furthermore the absorption maximum of [EYM+CA-2H] 2-is found at 513 nm, which is intermediate to the values for [EY-2H] 2-and [EYM-2H] 2-and hence shows the thioalkyl succinimide perturbs the S 0  S 1 transition less than the unreacted maleimide. Equally, the shape of the absorption band of [EYM+CA-H] -more closely resembles [EY-H] -, with a similar suppression of fragmentation yield at low excitation energies being indicative of fluorescence. What is the origin of the quenching mechanism of the maleimide moiety? As noted above, the large reduction of the branching ratio of photo-detachment may indicative of an electron transfer process in [EYM-2H] 2-, which is switched off by the reduction of maleimide to succinimide by the addition of cysteamine to the maleimide. To better understand how this chemical transformation quenches the electron transfer process, calculations on the electron transfer characteristics were performed. The singlet excited states of [EYM-2H] 2-and [EYM+CA-2H] 2-were calculated and the transitions with dominant charge transfer character (CT) identified (long-range corrected hybrid functionals such as CAM-B3LYP have been designed for accurately describing CT types of transitions).

  [START_REF] Peach | Excitation Energies in Density Functional Theory: An Evaluation and a Diagnostic Test[END_REF] A nonlocal CT excitation usually has little spatial overlap (O ia < 0.1) as identified also in the leading excitations in the CT states in [EYM-2H] 2-(S 1 ) and [EYM+CA-2H] 2-(S 4 ). Both CT transitions involve excitation from the EY π-system to a π* orbital centered on the maleimide moiety. In the case of EYM this orbital is of pure π* character, whilst for EYM+CA a partial σ* character induced by the reduction of the maleimide to succinimide due to the reaction with cysteamine shifts the entire excited state higher in energy. Thus the CT state is found at 2.3 eV in [EYM-2H] 2-and at about 4.1 eV for the [EYM+CA-2H] 2-. The difference in the energy of the CT states is driven by changes in the electron affinity of the maleimide (-0.725 eV) and succinimide (+1.000 eV) moieties. The position of the CT state in relation to the initially excited EY π-π* governs the final deactivation pathway (the system relaxes from the lowest excited state to the ground state in analogy to Kasha's rule). TDDFT calculations fail to accurately reproduce the transition energy of the first singlet excitations of xanthene derivatives -although it is more successful in the UV. 19,34 The experimentally determined EY π-π* transition energy of 2.44 eV is therefore taken as a reference value for the energetic position of [EYM-2H] 2-and [EYM+CA-2H] 2-. Despite of the apparent systematic shift, the trends of TDDFT are still valuable for a comparison across the compounds and included into Figure 4. This comparison reveals that the π-π* transition energy is unaffected by the functionalization while the energy of the CT state significantly depend on the substitution at the maleimide (see above). In particular, the CT state in [EYM-2H] 2-is found below the π-π* transition by ~ 0.1 eV. This renders the conversion to the CT state possible for [EYM-2H] 2- but not for [EYM+CA-2H] 2-suggesting an electron-transfer enabled deactivation pathway for [EYM-2H] 2-.

  shows the experimental and simulated isotopic distributions for [EYM-H] -(a and b) and [EYM-2H] 2-(c and d), where the broadening of the distribution is due to the isotopes of bromine.

  photo-fragmentation and photo-detachment in the dianion of EYM supports the idea of photo-

Figures and Tables.

  Figures and Tables.

Figure 1 .

 1 Figure 1. The structure of Eosin Y (EY), 5-maleimido-Eosin Y (EYM), and the product of

Figure 2 .Figure 3 .

 23 Figure 2. Gas phase action spectra of monoanions (a) and dianions (b) of EY (black squares),

Figure 4 .

 4 Figure 4. Analysis of the theoretically obtained excited charge-transfer (CT) states of [EYM-2H] 2-(green) and [EYM+CA-2H] 2-(purple). CT states were identified by a

Figure 5 .

 5 Figure 5. Experimental and theoretical isotopic distribution for mass-selected [EYM-H] -(a) and (b) respectively and [EYM-2H] 2-(c) and (d) respectively

  This is exactly what is observed in [EY-2H] 2--Figure 3a -and the action spectrum now resembles very closely the solution-phase spectrum for EY (solid blue curve), with an absorption maximum of 509 nm and a shoulder at 482 nm observed. The absorption band of [EYM-2H] 2-is very similar in appearance to [EY-2H] 2-, but with slightly redshifted values of 515 nm and 494 nm for the absorption maximum and shoulder respectively.This shift already hints that the presence of the maleimide moiety is influencing the first π  π* transition of the Eosin Y chromophore. Examination of the LID mass spectrum at the absorption maximum shows that relaxation no longer takes place only via photo-detachment but instead by a combination of photo-detachment and photofragmentation (Figure3b) with fragments at m/z 691, 614 (losses of 45 and 123 Da and corresponding to loss of a charge plus CO 2 H and CO 2 Br respectively) and 79 (corresponding to Br -) being the dominant species observed. This difference in behavior indicates that very different excited state dynamics are

AUTHOR INFORMATION

Corresponding Author

* To whom correspondence should be addressed. Email: philippe.dugourd@univ-lyon1.fr. Telephone: +33 4 72 44 83 21

Author Contributions

All authors have given approval to the final version of the manuscript.

ACKNOWLEDGMENT

The research leading to these results has received funding from the


Valerio Basile
email: v.basile@rug.nl

Johan Bos
email: johan.bos@rug.nl

Aligning Formal Meaning Representations with Surface Strings for Wide-coverage Text Generation

Statistical natural language generation from abstract meaning representations presupposes large corpora consisting of text-meaning pairs. Even though such corpora exist nowadays, or could be constructed using robust semantic parsing, the simple alignment between text and meaning representation is too coarse for developing robust (statistical) NLG systems. By reformatting semantic representations as graphs, fine-grained alignment can be obtained. Given a precise alignment at the word level, the complete surface form of a meaning representations can be deduced using a simple declarative rule.

Introduction

Surface Realization is the task of producing fluent text from some kind of formal, abstract representation of meaning [START_REF] Reiter | Building Natural Language Generation Systems[END_REF]. However, while it is obvious what the output of a natural language generation component should be, namely text, there is little to no agreement on what its input formalism should be [START_REF] Evans | What is NLG?[END_REF]. Since open-domain semantic parsers are able to produce formal semantic representations nowadays [START_REF] Bos | Wide-Coverage Semantic Analysis with Boxer[END_REF][START_REF] Butler | Banking meaning representations from treebanks[END_REF], it would be natural to see generation as a reversed process, and consider such semantic representations as input of a surface realization component.

The idea of using large text corpora annotated with formal semantic representations for robust generation has been presented recently [START_REF] Basile | Towards generating text from discourse representation structures[END_REF][START_REF] Wanner | Towards a surface realization-oriented corpus annotation[END_REF]. The need for formal semantic representations as a basis for NLG was expressed already much earlier by [START_REF] Power | Controlling logical scope in text generation[END_REF], who derives semantic networks enriched with scope information from knowledge representations for content planning. In this paper we take a further step towards the goal of generating text from deep semantic representations, and consider the issue of aligning the representations with surface strings that capture their meaning.

First we describe the basic idea of aligning semantic representations (logical forms) with surface strings in a formalism-independent way (Section 2). Then we apply our method to a well-known and widely-used semantic formalism, namely Discourse Representation Theory (DRT), first demonstrating how to represent Discourse Representation Structures (DRSs) as graphs (Section 3) and showing that the resulting Discourse Representation Graphs (DRGs) are equivalent to DRSs but are more convenient to fulfill wordlevel alignment (Section 4). Finally, in Section 5 we present a method that generates partial surface strings for each discourse referent occurring in the semantic representation of a text, and composes them into a complete surface form. All in all, we think this would be a first and important step in surface realization from formal semantic representations.

Aligning Logic with Text

Several different formal semantic representations have been proposed in the literature, and although they might differ in various aspects, they also have a lot in common. Many semantic representations (or logical forms as they are sometimes referred to) are variants of first-order logic and share basic building blocks such as entities, properties, and relations, complemented with quantifiers, negation and further scope operators.

A simple snapshot of a formal meaning representation is the following (with symbols composed out of WordNet (Fellbaum, 1998) synset identifiers to abstract away from natural language):

blue#a#1(x) ∧ cup#n#1(x)
How could this logical form be expressed in natural language? Or put differently, how could we realize the variable x in text? As simple as it is, x describes "a blue cup", or if your target language is Italian, "una tazza blu", or variants hereof, e.g. "every blue cup" (if x happens to be bound by universally quantified) or perhaps as "una tazza azzurra", using a different adjective to express blueness. This works for simple examples, but how does it scale up to larger and more complex semantic representations?

In a way, NLG can be viewed as a machine translation (MT) task, but unlike translating from one natural language into another, the task is here to translate a formal (unambiguous) language into a natural language like English or Italian. Current statistical MT techniques are based on large parallel corpora of aligned source and target text. In this paper we introduce a method for precise alignment of formal semantic representations and text, with the purpose of creating a large corpus that could be used in NLG research, and one that opens the way for statistical approaches, perhaps similar to those used in MT.

Broadly speaking, alignments between semantic representations and surface strings can be made in three different ways. The simplest strategy, but also the least informative, is to align a semantic representation with a sentence or complete text without further information on which part of the representation produces what part of the surface form. This might be enough to develop statistical NLG systems for small sentences, but probably does not scale up to handle larger texts. Alternatively, one could devise more complex schemes that allow for a more fine-grained alignment between parts of the semantic representation and surface strings (words and phrases). Here there are two routes to follow, which we call the minimal and maximal alignment.

In maximal alignment, each single piece of the semantic representation corresponds to the words that express that part of the meaning. Possible problems with this approach are that perhaps not every bit of the semantic representation corresponds to a surface form, and a single word could also correspond to various pieces in the semantic representation. This is an interesting option to explore, but in this paper we present the alternative approach, minimal alignment, which is a method where every word in the surface string points to exactly one part of the semantic representation. We think this alignment method forms a better starting point for the development of a statistical NLG component. With sufficient data in the form of aligned texts with semantic representations, these alignments can be automatically learned, thus creating a model to generate surface forms from abstract, logical representations.

However, aligning semantic representations with words is a difficult enterprise, primarily because formal semantic representations are not flat like a string of words and often form complex structures. To overcome this issue we represent formal semantic representations as a set of tuples. For instance, returning to our earlier example representation for "blue cup", we could represent part of it by the tuples blue#a#1,arg,x and cup#n#1,arg,x . For convenience we can display this as a graph (Figure 1). Note that in this example several tuples are not shown for clarity (such as conjunction and the quantifier). We show below that we can indeed represent every bit of semantic information in this format without sacrificing the capability of alignment with the text. The important thing now is to show how alignments between tuples and words can be realized, which is done by adding an element to each tuple denoting the surface string, for instance cup#n#1,arg,x,"tazza" , as in Figure 2. We can further refine the alignment by saying something about the local order of surface expressions. Again, this is done by adding an element to the tuple, in this case one that denotes the local order of a logical term. We will make this clear by continuing with our example, where we add word order encoded as numerical indices in the tuple, e.g. cup#n#1,arg,x,"tazza",2 , as Figure 3 shows.

x blue#a#1 "blue" "a" cup#n#1 "cup" x blue#a#1 "blu" "una" cup#n#1 "tazza"
From these graphs we can associate the term x with the surface strings "a blue cup" and "una tazza blu". But the way we express local order is not limited to words and can be employed for partial phrases as well, if one adopts a neo-Davidsonian event semantics with explicit thematic roles. This can be achieved by using the same kind of numerical indices already used for the alignment of words. The example in Figure 4 shows how to represent an event "hit" with its thematic roles, preserving their relative order. We call surface forms "partial" or "incomplete" when they contain variables, and "complete" when they only contain tokens. The corresponding partial surface form is then "y hit z", where y and z are placeholders for surface strings. This is the basic idea of aligning surface strings with parts of a deep semantic representation. Note that precise alignment is only possible for words with a lexical semantics that include first-order variables. For words that introduce scope operators (negation particles, coordinating conjuncts) we can't have the cake and eat it: specifying the local order with respect to an entity or event variable directly and at the same time associating it with an operator isn't always possible. To solve this we introduce surface tuples that complement a semantic representation to facilitate perfect alignment. We will explain this in more detail in the following sections.

x blue#a#1 "blue" 2 "a" 1 cup#n#1 "cup" 3 x blue#a#1 "blu" 3 "una" 1 cup#n#1 "tazza" 2

Discourse Representation Graphs

The choice of semantic formalism should ideally be independent from the application of natural language generation itself, to avoid bias and spe-cific tailoring the semantic representation to one's (technical) needs. Further, the formalism should have a model-theoretic backbone, to ensure that the semantic representations one works with actually have an interpretation, and can consequently be used in inference tasks using, for instance, automated deduction for first-order logic. Given these criteria, a good candidate is Discourse Representation Theory, DRT [START_REF] Kamp | From Discourse to Logic; An Introduction to Modeltheoretic Semantics of Natural Language, Formal Logic and DRT[END_REF], that captures the meaning of texts in the form of Discourse Representation Structures (DRSs).

DRSs are capable of effectively representing the meaning of natural language, covering many linguistic phenomena including pronouns, quantifier scope, negation, modals, and presuppositions. DRSs are recursive structures put together by logical and non-logical symbols, as in predicate logic, and in fact can be translated into first-order logic formulas [START_REF] Muskens | Combining Montague Semantics and Discourse Representation[END_REF]. The way DRSs are nested inside each other give DRT the ability to explain the behaviour of pronouns and presuppositions ( [START_REF] Van Der Sandt | Presupposition Projection as Anaphora Resolution[END_REF].

Aligning DRSs with texts with fine granularity is hard because words and phrases introduce different kinds of semantic objects in a DRS: discourse referents, predicates, relations, but also logical operators such as negation, disjunction and implication that introduce embedded DRSs. A precise alignment of a DRS with its text on the level of words is therefore a non-trivial task.

To overcome this issue, we apply the idea presented in the previous section to DRSs, making all recursion implicit by representing them as directed graphs. We call a graph representing a DRS a Discourse Representation Graph (DRG, in short). DRGs encode the same information as DRSs, but are expressed as a set of tuples. Essentially, this is done by reification over DRSs -every DRSs gets a unique label, and the arity of DRS conditions increases by one for accommodating a DRS label. This allows us to reformulate a DRS as a set of tuples.

A DRS is an ordered pair of discourse referents (variables over entities) and DRS-conditions. DRS-conditions are basic (representing properties or relations) or complex (to handle negation and disjunction). To reflect these different constructs, we distinguish three types of tuples in DRGs:

• K,referent,X means that X is a discourse referent in K (referent tuples); With the help of a concrete example, it is easy to see that DRGs have the same expressive power as DRSs. Consider for instance a DRS with negation, before and after labelling it (Figure 6): Now, from the labelled DRS we can derive the following three referent tuples: K 1 ,referent,x , K 1 ,referent,y , and K 2 ,referent,z ; the following four condition tuples: K 1 ,relation,c 1 :r , K 1 ,unary,c 2 :¬ , K 2 ,concept,c 3 :p , and K 2 ,relation,c 4 :s ; and the following argument tuples: c 1 :r,internal,x , c 1 :r,external,y , c 2 :¬,scope,K 2 , c 3 :p,instance,x , c 4 :s,internal,z , and c 4 :s,external,y .

From these tuples, it is straightforward to recreate a labelled DRS, and by dropping the labels subsequently, the original DRS resurfaces again.

For the sake of readability we sometimes leave out labels in examples throughout this paper. In addition, we also show DRGs in graph-like pictures, where the tuples that form a DRG are the edges, and word-alignment information attached at the tuple level is shown as labels on the graph edges, as in Figure 9. In such graphs, nodes representing discourse referents are square shaped, and nodes representing conditions are oval shaped.

Note that labelling conditions is crucial to distinguish between syntactically equivalent conditions occurring in different (embedded) DRSs. Unlike Power's scoped semantic network for DRSs, we don't make the assumption that conditions appear in the DRS in which their discourse referents are introduced [START_REF] Power | Controlling logical scope in text generation[END_REF]. The example in Figure 6 illustrates that this assumption is not sound: the condition p(x) is in a different DRS than where its discourse referent x is introduced. Further note that our reification procedure yields "flatter" representations than similar formalisms [START_REF] Copestake | Translation using Minimal Recursion Semantics[END_REF][START_REF] Reyle | Dealing with Ambiguities by Underspecification: Construction, Representation and Deduction[END_REF], and this makes it more convenient to align surface strings with DRSs with a high granularity, as we will show below.

Word-Aligned DRGs

In this section we show how the alignment between surface text and its logical representation is realized by adding information of the tuples that make up a DRG. This sounds more straightforward than it is. For some word classes this is indeed easy to do. For others we need additional machinery in the formalism. Let's start with the straightforward cases. Determiners are usually associated with referent tuples. Content words, such as nouns, verbs, adverbs and adjectives, are typically directly associated with one-place relation symbols, and can be naturally aligned with argument tuples. Verbs are assigned to instance tuples linking its event condition; likewise, nouns are typically aligned to instance tuples which link discourse referents to the concepts they express; adjectives are aligned to tuples of attribute conditions. Finally, words expressing relations (such as prepositions), are attached to the external argument tuple linking the relation to the discourse referent playing the role of external argument.

Although the strategy presented for DRG-text alignment is intuitive and straightforward to implement, there are surface strings that don't correspond to something explicit in the DRS. To this class belong punctuation symbols, and semantically empty words such as (in English) the infinitival particle, pleonastic pronouns, auxiliaries, there insertion, and so on. Furthermore, function words such as "not", "if", and "or", introduce semantic material, but for the sake of surface string generation could be better aligned with the event that they take the scope of. To deal with all these cases, we extend DRGs with surface tuples of the form K,surface,X , whose edges are decorated with the required surface strings. Figure 7 shows an example of a DRG extended with such surface tuples.

k 1 unary ¬ ¬ scope k 2 k 2 referent e 1 k 2 referent x 1 1 A k 2 event pay k 2 concept customer k 2 role agent customer instance x 1 2 customer pay instance e 1 4 pay agent internal e 1 1 agent external x 1 k 2 surface e 1 2 did k 2 surface e 1 3 not k 2
surface e 1 5 .

Figure 7: Word-aligned DRG for "A customer did not pay." All alignment information (including surface tuples) is highlighted.

Note that surface tuples don't have any influence on the meaning of the original DRS -they just serve for the purpose of alignment of the required surface strings. Also note in Figure 7 the indices that were added to some tuples. They serve to express the local order of surface information.

Following the idea sketched in Section 2, the total order of words is transformed into a local ranking of edges relative to discourse referents. This is possible because the tuples that have word tokens aligned to them always have a discourse referent as third element (the head of the directed edge, in terms of graphs). We group tuples that share the same discourse referent and then assign indices reflecting the relative order of how these tuples are realized in the original text. Illustrating this with our example in Figure 7, we got two discourse referents: x 1 and e 1 . The discourse referent x 1 is associated with three tuples, of which two are indexed (with indices 1 and 2). Generating the surface string for x 1 succeeds by traversing the edges in the order specified, resulting in [A,customer] for x 1 . The referent e 1 associates with six tuples, of which four are indexed (with indices 1-4). The order of these tuples would yield the partial surface string [x 1 ,did,not,pay,.] for e 1 . Note that the manner in which DRSs are constructed during analysis ensures that all discourse referents are linked to each other by taking the transitive closure of all binary relations appearing in a DRS, and therefore we can reconstruct the total order from composing the local orders. In the next section we explain how this is done.

Surface Composition

In this section we show in detail how surface strings can be generated from word-aligned DRGs. It consists of two subsequent steps. First, a surface form is associated with each discourse referent. Secondly, surface forms are put together in a bottom-up fashion, to generate the complete output. During the composition, all of the discourse referents are associated with their own surface representation. The surface form associated with the discourse unit that contains all other discourse units is then the text aligned with the original DRG.

Surface forms of discourse referents are lists of tokens and other discourse referents. Recall that the order of the elements of a discourse referent's surface form is reflected by the local ordering of tuples, as explained in the previous section, and tuples with no index are simply ignored when reconstructing surface strings.

The surface form is composed by taking each tuple belonging to a specific discourse referent, in the correct order, and adding the tokens aligned with the tuple to a list representing the surface string for that discourse referent. An important part of this process is that binary DRS relations, represented in the DRG by a pair of internal and external argument tuple, are followed unidirectionally: if the tuple is of the internal type, then the discourse referent on the other end of the relation (i.e. following its external tuple edge) is added to the list. Surface forms for embedded DRSs include the discourse referents of the events Typically, discourse units contain exactly one event (the main event of the clause). Phenomena such as gerunds (e.g. "the laughing girl") and relative clauses (e.g. "the man who smokes") may introduce more than one event in a discourse unit. To ensure correct order and grouping, we borrow a technique from description logic [START_REF] Horrocks | A description logic with transitive and inverse roles and role hierarchies[END_REF] and invert roles in DRGs. Rather than representing "the laughing girl" as [girl(x) ∧ agent(e,x) ∧ laugh(e)], we represent it as [girl(x) ∧ agent -1 (x,e) ∧ laugh(e)], making use of R(x,y) ≡ R -1 (y,x) to preserve meaning. This "trick" ensures that we can describe the local order of noun phrases with relative clauses and alike.

To wrap things up, the composition operation is used to derive complete surface forms for DRGs. Composition puts together two surface forms, where one of them is complete, and one of them is incomplete. It is formally defined as follows:

ρ 1 : τ ρ 2 : Λ 1 ρ 1 Λ 2 ρ 2 : Λ 1 τ Λ 2 (1)
where ρ 1 and ρ 2 are discourse referents, τ is a list of tokens, and Λ 1 and Λ 2 are lists of word tokens and discourse referents. In the example from Figure 7, the complete surface form for the discourse unit k 1 is derived by means of composition as formulated in (1) as follows:

k2 : e1 x1 : A customer e1 : x1 did not pay e1 : A customer did not pay . k2 : A customer did not pay .

The procedure for generation described here is reminiscent of the work of [START_REF] Stuart | A uniform architecture for parsing and generation[END_REF] who also employs a deductive approach. In particular our composition operation can be seen as a simplified completion.

Going back to the example in Section 4, substituting the value of x 1 in the incomplete surface form of e 1 produces the surface string [A,customer,did,not,pay,.] for e 1 .

Selected Phenomena

We implemented a first prototype using our alignment and realization method and tested it on examples taken from the Groningen Meaning Bank, a large annotated corpus of texts paired with DRSs [START_REF] Valerio Basile | Developing a large semantically annotated corpus[END_REF]. Naturally, we came across phenomena that are notoriously hard to analyze. Most of these we can handle adequately, but some we can't currently account for and require further work.

Embedded Clauses

In the variant of DRT that we are using, propositional arguments of verbs introduce embedded DRSs associated with a discourse referent. This is a good test for our surface realization formalism, because it would show that it is capable of recursively generating embedded clauses. Figure 9 shows the DRG for the sentence "Michelle thinks that Obama smokes." Here the surface forms of two discourse units (main and embedded) are generated. In order to generate the complete surface form, first the embedded clause is generated, and then composed with the incomplete surface form of the main clause. As noted earlier, during the composition process, the complete surface form for each discourse referent is generated (Figure 8), showing a clear alignment between the entities of the semantic representation and the surface forms they represent.

Coordination

Coordination is another good test case for a linguistic formalism. Consider for instance "Subsistence fishing and commercial trawling occur within refuge waters", where two noun phrases are coordinated, giving rise to either a distributive (introducing two events in the DRS) or a collective interpretation (introducing a set formation of discourse referents in the DRS). We can account for both interpretations (Figure 10).

Note that, interestingly, using the distributive interpretation DRG as input to the surface realization component could result, depending on how words are aligned, in a surface form "fishing occurs and trawling occurs", or as "fishing and trawling occur".

Long-Distance Dependencies

Cases of extraction, for instance with WHmovement, could be problematic to capture with our formalism. This is in particular an issue when extraction crosses more than one clause boundary, as in "Which car does Bill believe John bought". Even though these cases are rare in the real world, a complete formalism for surface realization must be able to deal with such cases. The question is whether this is a separate generation task in the domain of syntax [START_REF] White | Towards broad coverage surface realization with CCG[END_REF], or whether the current formalism needs to be adapted to cover such long-distance dependencies. Another range of complications are caused by discontinuous constituents, as in the Dutch sentence "Ik heb kaartjes gekocht voor Berlijn" (literally: "I have tickets bought for Berlin"), where the prepositional phrase "voor Berlijn" is an argument of the noun phrase "kaartjes". In our formalism the only alignment possible would result in the sentence "Ik heb kaartjes voor Berlijn gekocht", which is arguably a more fluent realization of the sentence, but doesn't correspond to the original text. If one uses the original text as gold standard, this could cause problems in evaluation. (One could also benefit from this deficiency, and use it to generate more than one gold standard surface string. This is something to explore in future work.)

Control Verbs

In constructions like "John wants to swim", the control verb "wants" associates its own subject with the subject of the infinitival clause that it has as argument. Semantically, this is realized by variable binding. Generating an appropriate surface form for semantic representation with controlled variables is a challenge: a naive approach would generate "John wants John to swim". One possible solution is to add another derivation rule for surface composition dedicated to deal with cases where a placeholder variable occurs in more than one partial surface form, substituting a null string for a variable following some heuristic rules. A second, perhaps more elegant solution is to integrate a language model into the surface composition process.

Related work

Over the years, several systems have emerged that aim at generate surface forms from different kind of abstract input representations. An overview of the state-of-the-art is showcased by the submissions to the Surface Realization Shared Task [START_REF] Belz | The surface realisation task: Recent developments and future plans[END_REF]. [START_REF] Bohnet | Broad coverage multilingual deep sentence generation with a stochastic multilevel realizer[END_REF], for instance, employ deep structures derived from the CoNLL 2009 shared task, essentially sentences annotated with shallow semantics, lemmata and dependency trees; as the authors state, these annotations are not made with generation in mind, and they necessitate complex preprocessing steps in order to derive syntactic trees, and ultimately surface forms. The format presented in this work has been especially developed with statistical approaches in mind.

Nonetheless, there is very little work on robust, wide-scale generation from DRSs, surprisingly perhaps given the large body of theoretical research carried out in the framework of Discourse Representation Theory, and practical implementations and annotated corpora of DRSs that are nowadays available [START_REF] Valerio Basile | Developing a large semantically annotated corpus[END_REF]. This is in contrast to the NLG work in the framework of Lexical Functional Grammar [START_REF] Guo | Dependency-based n-gram models for general purpose sentence realisation[END_REF].

Flat representation of semantic representations, like the DRGs that we present, have also been put forward to facilitate machine translation [START_REF] Schiehlen | Verbmobil interface terms (vits)[END_REF] and for evaluation purposes [START_REF] Allen | Deep Semantic Analysis of Text[END_REF], and semantic parsing ( [START_REF] Le | Learning compositional semantics for open domain semantic parsing[END_REF] just because they're easier and more efficient to process. Packed semantic representations (leaving scope underspecified) also resemble flat representations [START_REF] Copestake | Translation using Minimal Recursion Semantics[END_REF][START_REF] Reyle | Dealing with Ambiguities by Underspecification: Construction, Representation and Deduction[END_REF] and can be viewed as graphs, however they show less elaborated reification than the DRGs presented in this paper, and are therefore less suitable for precise alignment with surface strings.

Conclusion

We presented a formalism to align logical forms, in particular Discourse Representation Structures, with surface text strings. The resulting graph representations (DRGs), make recursion implicit by reification over nested DRSs. Because of their "flat" structure, DRGs can be precisely aligned with the text they represent at the word level. This is key to open-domain statistical Surface Realization, where words are learned from abstract, syntactic or semantic, representations, but also useful for other applications such as learning semantic representations directly from text [START_REF] Le | Learning compositional semantics for open domain semantic parsing[END_REF]. The actual alignment between the tuples that form a DRG and the surface forms they represent is not trivial, and requires to make several choices. Given the alignment with text, we show that it is possible to directly generate surface forms from automatically generated word-aligned DRGs. To do so, a declarative procedure is presented, that generates complete surface forms from aligned DRGs in a compositional fashion. The method works in a bottom-up way, using discourse referents as starting points, then generating a surface form for each of them, and finally composing all of the surface form together into a complete text. We are currently building a large corpus of word-aligned DRSs, and are investigating machine learning methods that could automatically learn the alignments.

Surprisingly, given that DRT is one of the best studied formalisms in formal semantics, there isn't much work on generation from DRSs so far. The contribution of this paper presents a method to align DRSs with surface strings, that paves the way for robust, statistical methods for surface generation from deep semantic representations.

Figure 1 :

 1 Figure 1: Logical form graph.

Figure 2 :

 2 Figure 2: Logical form graphs aligned with surface forms in two languages.

Figure 3 :

 3 Figure 3: Encoding local word order.

Figure 4 :

 4 Figure 4: Graph for a neo-Davidsonian structure.

Figure 6 :

 6 Figure 6: From DRS to DRG: labelling.

  Michelle e1 : x1 thinks p1 e1 : Michelle thinks p1 p1 : that e2 x2 : Obama e2 : x2 smokes . e2 : Obama smokes . p1 : that Obama smokes . e1 : Michelle thinks that Obama smokes . k1 : Michelle thinks that Obama smokes .

Figure 8 :

 8 Figure 8: Surface composition of embedded structures.

Figure 9 :

 9 Figure 9: Word-aligned DRG for the sentence "Michelle thinks that Obama smokes."

Figure 10 :

 10 Figure 10: Analysis of NP coordination, in a distributive (left) and a collective interpretation (right).