Article pubs.acs.org/JPCA
Conformational Analysis of Quinine and Its Pseudo Enantiomer Quinidine: A Combined Jet-Cooled Spectroscopy and Vibrational Circular Dichroism Study Ananya Sen,† Aude Bouchet,† Valeria Lepère,† Katia Le Barbu-Debus,† D. Scuderi,‡ F. Piuzzi,§ and A. Zehnacker-Rentien*,† †
CNRS, Institut des Sciences Moléculaires d’Orsay (ISMO), UMR8214, Orsay F-91405, and Univ Paris-Sud, Orsay F-91405, France CNRS, Laboratoire de Chimie Physique (LCP), UMR8000, Orsay F-91405, and Univ Paris-Sud, Orsay F-91405, France § CEA, IRAMIS, Service des Photons, Atomes et Molécules (SPAM), and CNRS, Laboratoire Francis Perrin (LFP), URA 2453, 91191 Gif-sur-Yvette, France ‡
S Supporting Information *
ABSTRACT: Laser-desorbed quinine and quinidine have been studied in the gas phase by combining supersonic expansion with laser spectroscopy, namely, laser-induced fluorescence (LIF), resonance-enhanced multiphoton ionization (REMPI), and IR-UV double resonance experiments. Density funtional theory (DFT) calculations have been done in conjunction with the experimental work. The first electronic transition of quinine and quinidine is of π−π* nature, and the studied molecules weakly fluoresce in the gas phase, in contrast to what was observed in solution (Qin, W. W.; et al. J. Phys. Chem. C 2009, 113, 11790). The two pseudo enantiomers quinine and quinidine show limited differences in the gas phase; their main conformation is of open type as it is in solution. However, vibrational circular dichroism (VCD) experiments in solution show that additional conformers exist in condensed phase for quinidine, which are not observed for quinine. This difference in behavior between the two pseudo enantiomers is discussed.
1. INTRODUCTION Quinine and its pseudoenantiomer quinidine (see Scheme 1) are two of the Cinchona alkaloids found in the bark of Cinchona,
chemical activity. In this respect, the rotation around the C8C9 bond linking the quinuclidine and quinoline parts is of prime importance because it defines the geometry of the chiral pouch made from the two cyclic moieties. The molecules are very flexible along this rotation coordinate. Indeed, this angle has been shown to depend on the state charge of the Cinchona alkaloids, the presence of an interacting molecule, or that of a counterion.4,5 A large variety of conformations may exist along this coordinate, ranging from the open conformation in which the C4′C9C8N1 dihedral angle is close to 150° or 290° to the closed conformation in which C4′C9C8N1 is close to 50°. In neutral cinchonidine, for example, closed and open conformations are thought to coexist, depending on the polarity of the solvent. Polar solvents tend to stabilize the more polar closed form.6 The open form dominates in nonpolar solvents, and we therefore expect it in the gas phase as well.7 Moreover, the Cinchona alkaloids bear five chiral centers, namely, N1, C3, C4, C8, and C9. That of the C8 and C9 atoms plays an especially important role in their chemical and pharmaceutical activity, and their conformation leads to four different diastereoisomers. Among them, quinine and quinidine (see Scheme 1) are natural products and, despite being
Scheme 1. Molecules under Study and Numbering of the Atoms
which gained tremendous importance since its medical use against malaria, at the beginning of the 17th century.1 Besides their medical properties, Cinchona alkaloids play an important role in analytical chemistry as chiral stationary phases.2 They are also of wide use as enantioselective catalysts for the reduction of ketones.3 The conformation of Cinchona alkaloids is thought to play a decisive role in most of the aspects of their biological as well as © 2012 American Chemical Society
Received: May 17, 2012 Revised: July 26, 2012 Published: July 27, 2012 8334
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seeded supersonic beam was intersected by two laser beams superimposed in the ion-source region of a linear time-of-flight mass spectrometer (RM Jordan, Wiley−McLaren type). As the ionization potential is more than twice the energy of the S0−S1 energy gap, one-color ionization was not possible. The fourth harmonic of an Nd:Yag laser (BM Industrie) at 266 nm was therefore used for ionizing the system. Its fluence has been kept as low as possible to avoid ionization by two photons at 266 nm. Molecules were excited by a tunable frequency-doubled dye laser (Sirah equipped with DCM) pumped by a Nd:YAG laser (Quanta Ray, Spectra Physics) and ionized by the fourth harmonic of a Nd:YAG laser (BM Industrie). The dye laser output was frequency-doubled by a KDP crystal. Ions were detected by a microchannel plate detector (RM Jordan, 25 mm) mounted on top of a 1 m flight tube. The ion signal was averaged by an oscilloscope (Lecroy wavesurfer) and processed through a personal computer. Fluorescence excitation spectra were obtained by exciting the jet-cooled molecule by the same tunable dye laser. The fluorescence signal from the sample was collected perpendicularly to both the exciting light and the molecular beam by a two-lens collecting system and a 25 cm monochromator (Huet M25) used under broad band conditions and then detected by a Hamamatsu R2059 photomultiplier tube. The output electrical signal from the PMT was processed as described above. Vibrational spectra of jet-cooled species were obtained resorting to the IR-UV double resonance technique,15−17 using fluorescence detection. The experimental setup has been described in detail elsewhere18−22 and will be briefly described here. It rests on the use of two copropagating lasers focused by a 500 mm focal length lens on the cold region of the supersonic expansion. The UV laser wavelength is fixed on a transition due to a given conformer, while the IR laser is scanned in the region of the ν(OH) and ν(CH) stretch modes. When the IR laser is resonant with a vibrational transition arising from the same ground state as that being probed by the UV laser, decrease in the fluorescence signal is observed. A chopper has been introduced on the path of the IR so that its repetition rate (5 Hz) is half that of the UV, which allows monitoring the IR absorption by measuring the difference in fluorescence signal produced by successive UV laser pulses (one without and one with the IR laser present). The IR pulse reaches the supersonic beam 50 to 80 ns before the UV pulse. The 5 Hz IR laser can also be fixed on a vibrational transition observed in the IR-UV depletion, while the 10 Hz UV probe was tuned through the S1−S0 region of interest. Comparison between the fluorescence signal obtained with and without IR present allows one to determine whether the transitions observed in the S1−S0 spectrum arise from the same ground state as the transition origin. The tunable IR source is a tabletop IR Optical Parametric Oscillator/Amplifier (OPO/OPA) (LaserVision). The resolution of the UV is 0.02 cm−1, while that of the IR OPO is 3 cm−1. Synchronisation between the lasers is performed by a computer-controlled homemade gate generator. The solution-phase vibrational absorption spectra were measured using a FTIR spectrometer Vertex 70 (Bruker). The VCD spectra were measured with the same spectrometer equipped with a VCD module PMA 50 (Bruker). A spectral resolution of 4 cm−1 was used for both absorption and circular dichroism spectra. The IR radiation was polarized with a linear polarizer and modulated by a 50 kHz ZnSe photoelastic
diastereoisomers strictly speaking, are called pseudo enantiomers because only the configuration of the C8 and C9 atoms are opposite to each other. Despite being considered like true enantiomers for numerous applications,8 they show slightly different properties in their pharmaceutical use. For instance, quinidine binds slightly less than quinine to human serum albumin and is slightly more efficient than quinine. However, the behaviors of the two pseudo enantiomers are similar, while they are one to two orders of magnitude more active than the diastereoisomers having C8 and C9 of the same chirality, namely, 9-epiquinine and 9-epiquinidine.9 Crystallographic measurements show different packing properties between cinchonine and cinchonidine.10,11 Quinine and quinidine also show different solubility in usual solvents.12 These effects have been tentatively explained on the basis of ab initio calculations and 2D NOESY NMR spectra, in terms of a different stereo orientation of the peripheral ethylenic ligand in the two pseudo enantiomers. This results in subtle enthalpy and entropy changes between the two systems.12 In order to get better insight into the structural properties of quinine and its pseudo enantiomer quinidine, we have undertaken the first study reported so far of neutral quinine in the gas phase, thanks to a laser-ablation source, which allows desorbing the molecules intact. This study combines electronic and vibrational spectroscopy of the jet-cooled species in the gas phase with vibrational circular dichroism (VCD) in solution. We will first make use of the good resolution achieved by combining jet-cooled conditions with laser spectroscopy for getting information on the tiny differences between the two pseudo enantiomers. We will then check whether this difference also appears in solution, by using VCD, which is very sensitive to minor structural changes. Last, we will have an insight into the photophysics of the first excited state of quinine and quinidine. While protonated quinine has a high fluorescence quantum yield, which makes it a standard for fluorescence quantum yield measurement, neutral quinine does not fluoresce in nonpolar solvent due to a fast photoinduced charge transfer between the quinuclidine nitrogen to the aromatic ring.13 We will investigate in this work the fluorescence properties of quinine and quinidine isolated in the gas phase.
2. EXPERIMENTAL AND THEORETICAL METHODS Quinine and quinidine were purchased from Aldrich Chemicals and used without further purification. Quinine and quinidine were introduced in gas phase thanks to a homemade desorption source similar to that developed by Piuzzi et al.14 The source has been miniaturized to allow the excitation laser to be as close as a few mm from the nozzle, as required for laser-induced fluorescence experiments. A mixture of 75% analyte molecule and 25% graphite, used as a template, was prepared and pressed at 2.5* 108 Pa to make a pellet. A frequency-doubled Nd:YAG laser (532 nm) (Continuum Minilite) was used as the desorption source and propagated by an optical fiber to the surface of a half-pellet (diameter 13 mm) with 1.5 to 2 mJ energy per pulse and a repetition rate of 10 Hz. To increase its lifetime, the pellet is fixed on a linear translation system. This setup is attached to a pulsed valve (General Valve- Parker) with a nozzle diameter of 100 to 300 μm. The supersonic expansion is obtained by expanding argon or neon at 5 atm. The desorption plume crosses the cold jet at a distance of less than 1 mm from the nozzle. Mass-resolved S0−S1 spectra were obtained by two-color resonance-enhanced multiphoton ionization (2c-REMPI). The 8335
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Table 1. List of Gibbs Free Energies ΔG (kcal/mol), Boltzmann Populations (%), and Dihedral Angles (deg) of the Most Stable Conformers of Quinine and Quinidine Present at 300 K in Vacuum and in CHCl3 quinine in vacuum
ΔG
population
τ1 C3′C4′C9C8
τ2 C4′C9C8N
τ3 NC8C9O
T3 C9C8OH
cis-γ-open(3) cis-α-open(3) cis-γ-closed(1) trans-γ-open(3) quinine in CHCl3
0.00 1.11 1.24 1.28 ΔG
72 11 9 8 Population
99 104 −107 100 τ1 C3′C4′C9C8
152 151 58 154 τ2 C4′C9C8N
−83 −88 −178 −82 τ3 NC8C9O
174 80 −180 172 T3 C9C8OH
cis-γ-open(3) trans-γ-open(3) quinidine in vacuum
0.00 1.46 ΔG
92 8 population
100 100 τ1 C3′C4′C9C8
155 158 τ2 C4′C9C8N
−82 −78 τ3 NC8C9O
170 169 T3 C9C8OH
cis-γ-open(3) trans-γ-open(3) cis-γ-closed(1) quinidine in CHCl3
0.00 1.15 1.19 ΔG
78 11 11 population
−99 −100 107 τ1 C3′C4′C9C8
−153 −156 −59 τ2 C4′C9C8N
83 81 176 τ3 NC8C9O
−173 −171 178 T3 C9C8OH
cis-γ-open(3) cis-γ-closed(1) cis-α-closed(7) cis-γ-closed(2) trans-γ-closed(2)
0.00 0.91 0.96 1.05 1.36
59 13 12 10 6
−99 108 −17 −70 −69
−157 −57 −58 −59 −56
79 178 176 −179 −177
−170 −176 76 180 −178
Figure 1. (a) Most stable structures calculated for quinine at b3lyp/6-31G+(d,p) level of theory in CHCl3 solvent; see Table 1 for relative abundance. (b) Most stable structures calculated for quinidine at b3lyp/6-31G+(d,p) level of theory in CHCl3 solvent; see Table 1 for relative abundance.
included in the Maestro suite.23 The 20 most stable conformers have been optimized at the b3lyp/6-31+G(d,p) level of theory with Gaussian09.24 Dispersion-corrected density-functional theory (b3lyp-D/6-31+G(d,p) level) has been applied to the most stable conformers to check whether dispersion changes the relative stability of the conformers.25 As this is not the case, the results presented in what follows are those obtained at the b3lyp/6-31+G(d,p) level of theory. In what follows, the stability of the monomer is given in terms of free energy ΔG at 298.15 K (in kcal/mol) relative to the most stable conformation. Unless specified otherwise, we will limit the discussion to conformers with ΔG smaller than 1.5 kcal/mol. Indeed, those with ΔG above 1.5 kcal/mol have a population smaller than ∼5% of the main form. The vibrational spectrum has been simulated by calculating the harmonic frequencies at the same level of theory and convoluting the scaled frequencies by a Lorentzian line shape of
modulator (Hinds). The absorption spectra were detected using MCT IR detector with a BaF2 window, cooled with liquid nitrogen. A low-pass filter cutting at 2000 cm−1 was added before the linear polarizer to increase the dynamical response of the detector. The signal of the MCT detector was demodulated using a lock-in amplifier (Stanford Research Systems SR 830). The spectra were measured using 0.18 M solutions in CDCl3 in an adjustable BaF2 cell (Bruker) with a path length of 120 μm. The acquisition time was 3 h. The alignment of the spectrometer was carefully checked by verifying the mirrorimage relationship between the VCD spectra of the two enantiomers of camphor (0.3 M in CCl4) in the same cell. The spectra of quinine and quinidine were then corrected for baseline deviation by subtracting the solvent spectra recorded under identical conditions. A global exploration of the potential energy surface has been done for both molecules by means of the Macromodel program 8336
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3 cm−1 full width at half-maximum (fwhm). The calculated frequencies have been scaled by 0.960 in the ν(OH) and ν(CH) stretch region and 0.98 in the fingerprint region. The VCD spectra have been simulated including the solvent (CHCl3) at the same level of theory as the isolated species. The solvation model is the polarizable continuum model (PCM) using the integral equation formalism variant (IEFPCM) provided in the Gaussian09 package.
isoenergetic structure (cis-γ-closed(1)) is found, with the methoxy in the cis position; it corresponds to a closed geometry with the N atom of the quinuclidine ring standing above the quinoline ring. Results obtained when taking into account the CHCl3 solvent show that the most stable conformer obtained in vacuum is also the most stable in solution (see Figure 1b). However, some differences have to be noticed. While the most stable conformer has a population of 78% in vacuum, its population decreases to 59% in solution. Indeed, three other conformers (cis-γ-closed(1), cis-α-closed(7), and cis-γ-closed(2)) are found with population around 10% and one (trans-γclosed(2)) with a population of 6%. We can notice here that all those four conformers are closed forms, what was not observed for quinine. Indeed, quinine spectra in solution will mainly contain one conformer, while those of quinidine will contain contribution of five conformers. This contrasts with the gas phase conditions where the same cis-γ-open(3) structure is expected to be dominant for quinine and quinidine alike. 3.1.3. Vibrational Frequencies. Vibrational harmonic frequencies have been calculated for gas phase and solvated structures. The spectra calculated for the most stable conformations are presented in Figure S1, Supporting Information. All calculated spectra look very similar. For a given conformation, the calculated vibrational spectra of quinine and quinidine are identical. The observed differences between structures do not arise from the fact that these molecules are pseudo enantiomers but that they are characteristic of their conformations only. The ν(OH) stretch frequency has the same value (3822 cm−1) for all the γ conformers, whether they are cis or trans and open or closed. The α form displays a slightly higher ν(OH) stretch frequency (3863 cm−1). Indeed, there is a very weak interaction between the aromatic C3′H and the OH group of the linker, which is made possible in the α conformers and not in the γ, because the oxygen lone pair lies in the same plane for the former and not for the latter. This results in a small blue shift of the ν(OH) stretch in α relative to γ. This is reflected by a shorter distance (2.35 Å) in α relative to γ (2.47 Å). The ν(CH) stretch region is characteristic of the open or closed conformation. Indeed, a band appears at 3190 cm−1 in the spectrum of the closed form, which is absent in that of the others. It mainly corresponds to the C3′H stretching mode. Small differences between conformers also appear below 3100 cm−1, which are characteristic of the α or γ conformations, but are difficult to rationalize. 3.2. Gas Phase Electronic Spectroscopy. 3.2.1. REMPI Spectra. The time-of-flight spectra of jet-cooled quinine, with λexc= 318 nm and λion = 266 nm, is given in the Supporting Information (Figure S2). The main peak is assigned to the ion of the bare molecule (m/z 324). Besides, weaker peaks appear assigned to its dimer (m/z 648) and a photofragment (m/z 136). This fragment results from the loss of the quinuclidinyl radical ion, resulting from the homolytic rupture of the C8C9 bond in the α position of the quinuclidine nitrogen. This process is similar to that observed after photofragmentation of the protonated quinine dimer isolated in an ion trap.30,31 However, it is to be noted that the m/z 136 fragment bears the positive charge when it results from the dissociation of the quinine radical cation, while it is a neutral radical when it results from the dissociation of the protonated quinine dimer.
3. RESULTS 3.1. Calculated Structures. The most stable forms of quinine and quinidine have been calculated in the gas phase and in CHCl3. The calculated free energy differences at room temperature for the different conformers are given in Table 1. The most stable calculated forms in CHCl3 are shown in Figure 1. 3.1.1. Structure of Quinine. The results obtained for quinine resemble those obtained by Bürgi et al. for cinchonidine, and we will use the same nomenclature in what follows.6 The dihedral angles C3′C4′C9C8, C4′C9C8N1, and N1C8C9O are called τ1, τ2, and τ3, respectively. The values of τ1 and τ2 define the open or closed nature of the conformation, i.e., whether the quinuclidine ring is folded on the quinoline or not. We will add to this nomenclature the cis or trans position of the methoxy group relative to the C4′−C9 bond. We will also consider the C8−C9−O−H dihedral angle, T3, described by Caner et al.26 which will be named α, β, and γ for values of +60°, −60°, and 180°, respectively. The combined values of τ3 and T3 describe the relative orientation of the hydroxyl and tertiary amino groups, i.e., whether there is an internal hydrogen bond or not. In the gas phase, the most stable is the cis-γ-open(3) conformer. Its population amounts to 72%, and three other conformers with populations between 8 and 11% are found. Their geometries are cis-α-open(3) (11%), cis-γ-closed(1) (9%), and trans-γ-open(3) (8%). It is to be noted that no OH···N hydrogen bond takes place in these structures as the hydroxyl hydrogen atom is opposite the N atom, with the O and N atoms in an almost gauche position, (N1C8C9O dihedral of about −83° or −178°). It should be noted too that the methoxy substituent of the quinoline ring is for the most stable conformations in a cis conformation. Larger stability of cis relative to trans has been observed already in methoxynaphthalene and its derivatives.27−29 When introducing the CHCl3 solvent, only two conformers are obtained with a population above 5% (see Figure 1a). The cis-γ-open(3) conformer is still the most stable with a population of 92%, while the trans-γ-open(3) has a population of 8%. We can notice that no closed forms are obtained in that case. 3.1.2. Structure of Quinidine. The nomenclature for quinidine is the reverse of that of quinine as the chiral centers of interest, namely, C8 and C9, are of opposite configuration for quinine and quinidine. In other terms, τ1, τ2, and τ3 with opposite values for quinidine and quinine correspond to the same open(x) or closed(y) name. In the gas phase, the most stable form of quinidine corresponds to an open geometry (cis-γ-open(3)) with a C4′C9C8N1 dihedral angle of −153°. Following a Boltzmann distribution, this conformer should amount to 78% of the total population at room temperature. A second conformer (trans-γopen(3)) with the methoxy substituent of the quinoline ring in trans configuration, is calculated at 1.15 kcal/mol higher in energy and contributes to the population by 11%. An almost 8337
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lasers. At higher excitation energy, the signal-to-noise ratio of the REMPI spectra becomes poor because the lifetime of quinine becomes shorter and shorter. We have then resorted to laser-induced fluorescence to get better insight into the spectroscopy of quinine and quinidine. 3.2.2. Laser-Induced Fluorescence Excitation Spectra. The laser-induced fluorescence excitation spectrum of quinine and its pseudoenantiomer quinidine are shown in Figure 3. They
The mass-selected REMPI spectrum of quinine has been studied in the energy range from 30 300 to 31 630 cm−1 and is shown in Figure 2. An intense 0−0 transition appears at 30 621
Figure 3. Laser-induced fluorescence excitation spectrum of jet-cooled (a) quinine and (b) quinidine with Ne as a carrier gas. The zero of the scale is set at the transition origin located at 30 621 cm−1 for quinine and 30 641 cm−1 for quinidine, respectively.
Figure 2. REMPI S0−S1 spectrum of jet-cooled quinine with Ar as a carrier gas. Inset: variation in intensity of the region of the origin transition with change in carrier gas. The zero of the scale is set at the transition origin located at 30 621 cm−1.
cm−1; it has been assigned to the transition origin. Vibronic bands appear at higher energy at 288, 412, and 467 cm−1. The band 412 cm−1 is reminiscent of the band observed at 363 cm−1 in cis-2-methoxynaphthalene,27 which has been assigned to the lowest in-plane ring mode of naphthalene of b1u symmetry (8b1u). The band at 467 cm−1 corresponds to the 8b1g vibronic mode of naphthalene, which gains intensity via Herzberg− Teller coupling with the S2 state in naphthalene. It is usually active in the excitation spectrum of naphthalene derivatives even when substitution in the 2-position makes the transition Franck−Condon allowed.27,28,32−34 A band at 747 cm−1 has been assigned to the 8a1g mode of naphthalene. The observed frequencies as well as their assignment to calculated values are summarized in Table 2. The low-energy region displays a very rich pattern, which will be discussed later. Because of the very short lifetime of quinine (within the laser pulse), the intensity of the signal strongly depends on the temporal overlap between the excitation and the ionization
show strong resemblance to the REMPI spectrum. In what follows, we shall focus on the low-energy region of the LIF excitation spectrum and the low-frequency modes of quinine and assess the differences between quinine and quinidine. No attempt has been made at measuring the S1 lifetime as it is shorter than the laser pulse (
