Ultrafast Circular Dichroism Study of the Ring Opening of 7

Jan 4, 2012 - A great deal of effort has been expended to investigate mechanisms for improving product yield,(2-4) but understanding the dynamics of t...
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Ultrafast Circular Dichroism Study of the Ring Opening of 7-Dehydrocholesterol Julia Meyer-Ilse,†,‡ Denis Akimov,‡ and Benjamin Dietzek*,†,‡ †

Institute of Physical Chemistry and Abbe Center of Photonics, Friedrich-Schiller-University, Jena, Helmholtzweg 4, D-07743 Jena, Germany ‡ Institute of Photonic Technology Jena (IPHT) e.V., Albert-Einstein-Straße 9, D-07745 Jena, Germany S Supporting Information *

ABSTRACT: UV femtosecond time-resolved circular dichroism (TRCD) spectroscopy has been used to study the ultrafast changes of chirality in a small molecular biological paradigm sample, 7-dehydrocholesterol (7-DHC). Upon UV-photoexcitation, 7-DHC undergoes a ring opening to produce previtamin D3, and two of the chiral centers of 7-DHC are removed, which impacts the overall chirality of the molecule. Here, measurements of this chirality change connected to the ring opening of 7-DHC with a time resolution of 280 fs in the UV are reported. With this method, a previously described discrepancy concerning the photophysics of 7-DHC was clarified. With our setup, the relaxation time of the chirality change was measured to be 1−2 ps, which corresponds to the shortest time constant in the transient absorption (TA) measurements, allowing us to assign that time constant to the ring opening. SECTION: Kinetics, Spectroscopy

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circular dichroism (CD), the difference in absorption between left- and right-circular light (quantified by the difference in extinction coefficients Δε = εL − εR7). However, the intrinsically small CD signal (Δε/ε < 10−3) makes timeresolved circular dichroism (TRCD) measurements, that is, tracking the changes in chirality, difficult, especially with ultrafast time resolution. In this Letter, we present a study of the ring opening of 7-DHC using the only recently developed technique of ultrafast TRCD. Subpicosecond time resolution in the UV using TRCD has only been achieved so far, utilizing the pump−probe approach by Hache and colleagues with 800 fs.8,9 In the visible spectral range, experiments with 65010,11 and 400 fs12,13 have been reported by these authors. Trifonov et al.,14 on the other hand, used supramolecular samples with giant CD signals on the order of 400 mdeg to report transient chirality changes in the visible with 250 fs time resolution. With the setup presented, an unprecedented time resolution of 280 fs in the UV for CD measurements was achieved, opening up a previously unexplored and experimentally very challenging parameter space. This time resolution was accomplished by inserting a grating compressor to compensate for the dispersion of the UV pulses in optical media. The UV sub-280-fs transient CD spectrometer was applied to study the ring opening of 7DHC, as laid out above (Scheme 1).

he reaction sequence to synthesize vitamin D3 involves a UV-irradiation-induced photochemical ring opening of 7-dehydrocholesterol (7-DHC) to previtamin D3 and subsequent thermal rearrangement.1 The photochemical ring opening occurs with a quantum yield of 0.26;1,2 however, the absorption band of 7-DHC lies in the same spectral region as the absorption bands of its products, causing formation of both reversible and irreversible undesirable products. A great deal of effort has been expended to investigate mechanisms for improving product yield,2−4 but understanding the dynamics of the photochemical ring opening might ultimately lead to information to control the reaction to produce only the desired products.5 The ultrafast absorption dynamics of 7-DHC have been studied by femtosecond transient absorption (TA) spectroscopy to gain insight into the synthesis of previtamin D3.5,6 In the literature available, time constants around 5 and 120 ps were detected, and it was agreed upon that the 120 ps time constant is connected with the cis−trans isomerization of previtamin D3. The time constant of 5.2 ps was assigned by some authors to the ring opening,6 whereas others assigned the 5 ps time constant to vibrational cooling and the ring opening to a subpicosecond time constant,5 which, due to the solvent spike, could not be further specified in their UV TA measurements. During the ring opening, two chirality centers from the 7-DHC are removed, whereas during vibrational cooling, no massive structural rearrangements such as isomerization or ring opening occur. Therefore, tracking the photoinduced change in chirality could provide an exclusive measure for investigating the ring opening. The chirality of a molecule is often detected via © 2012 American Chemical Society

Received: November 5, 2011 Accepted: December 27, 2011 Published: January 4, 2012 182

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Scheme 1. Schematic Presentation of the Ring-Opening Reaction of 7-DHC to Form Previtamin D3a

a

Figure 2. TA measurements of the photoexcitation of 7-DHC with a pump wavelength of 275 nm and probed at two different wavelengths, 300 (green) and 280 nm (blue). Symbols and solid lines represent experimental data and multi-exponential fits, respectively.

The reaction is initiated by irradiation with UV light.

For all of our time-resolved measurements, a 280 fs, 275 nm pump pulse was used to induce the ring opening in 7-DHC. Here, the pump wavelength coincides with the maximum of the steady-state absorption spectrum of 7-DHC (Figure 1A), which

Table 1. Summary of the Time Constants of the TA Measured in the Literature in Comparison with Our Measurements (in bold) probe wavelength τ1 [ps] τ2 [ps] τ3 [ps]

262 nm6

261−285 nm5

300 nm

280 nm

5.2 ± 0.5 125 ± 20

∼0.4−1.0 5.0 ± 0.7 100 ± 20

1.0 ± 0.1 6.2 ± 0.1 192 ± 102

1.5 ± 0.1 6.3 ± 1.4 105 ± 41

centered at 300 nm (285 nm), whose polarization was continuously switched from right- to left-circular for consecutive pulses via a Pockels cell (see the Experimental Section for details). For different delay times between the pump and probe pulses, the circular dichroism, that is, the difference in absorption for two consecutive pulses, was recorded. A delay time interval between −3 and 18 ps was scanned to cover the range of both possible time constants for the ring opening, as discussed in literature, that is, either subpicosecond5 or 5 ps.6 At each time point, an average of 100 pulse pairs was recorded, and these measurements were averaged over about 20 repetitions to increase the signal-to-noise ratio. The integral measurement time was chosen so that no accumulation of undesired photoproducts was observed. Also, to make sure that the detected signal was not due to a Pockels cell drift over the long averaging time, the same measurement was performed on pure methanol, and there no drift was detected. From the measured CD data, a steady-state offset was subtracted before analyzing the data further, as described in the literature.14 The steady-state offset, that is, the CD value before time zero, occurs because 7-DHC is chiral and therefore already produces a CD signal before photoexcitation. After time zero, that is, upon photoexcitation, a change in the CD signal is detected, and the resultant data can be fit with a single exponential with time constants of 1.1 ± 0.2 and 1.9 ± 0.3 ps at probe wavelengths of 300 and 285 nm, respectively (Figure 3). The CD does not decay back to zero in the time window measured and appears as an offset in our fit. This apparent offset can be associated with the photoproduct (previtamin D3) that is created by the ring opening because the excited state, which might have a different CD signal than the ground state, has decayed back to the ground state within this time frame.21 Therefore, the ring opening has to occur before, and the time constants measured represent the induced ring opening in 7-DHC upon photoexcitation. To make sure that the produced photoproduct does not cause a depletion of the 7-DHC sample and therefore introduce a change in the CD signal, steady-state

Figure 1. Steady-state absorption (A) and CD (B) measurements of the 7-DCH in the near-UV (200−350 nm).

was recorded in methanol, the solvent also used in the transient CD measurements. The spectrum agrees with the spectra previously reported,2,5,6,15 indicating the purity of the sample used. This spectrum differs from the reported spectra of previtamin D3,5,6,15 which is less structured than 7-DHC and has its absorption maximum at 260 nm. The steady-state CD spectrum of 7-DHC shows a structured negative band in the wavelength region of the π−π* transition (Figure 1B) and differs from the steady-state CD spectra of the previtamin D3 reported in the literature, which is a smooth curve having a negative maximum at around 260 nm.16 The differences in the steady-state spectra of 7-DHC and previtamin D3 will manifest as a dynamics change of the spectroscopic properties after the photoexcitation of 7-DHC that can be followed by timeresolved spectroscopy. In particular, ultrafast transient CD measurements (see below) will prove valuable to resolve the persistent discussion on the nature of the 5 ps time constant observed in the photophysics of 7-DCH.5,6 TA measurements for two different probe wavelengths, 280 and 300 nm, are shown in Figure 2. After the coherent artifact at time zero, a triexponential kinetic is observed for both probe wavelengths. The coherent artifact stems from the interaction of the weak probe pulse in the presence of a strong pump pulse in the solvent17−20 and was therefore excluded from the exponential fit. The time constants of our triexponential fits are summarized in Table 1 along with those found in the literature. The time constants reported here coincide with those reported in the literature, which provides an effective cross-validation of our time-resolved spectrometer. For the TRCD measurements, the configurational state of the 7-DHC was probed with a 170 fs (222 fs) probe pulse 183

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tests the change in the difference in absorption between left- and right-circular polarization. This is accomplished by rotating the polarization of every other pulse with the help of a Pockels cell and calculating the absorption difference between two consecutive pulses. The setup is based on a commercial femtosecond 1-kHz laser system (LIBRA Coherent) operating at 800 nm that drives two independently tunable noncollinear optical parametric amplifiers (TOPAS-white, Light Conversion Ltd.) to produce the pump and probe pulses. The TOPAS-white produces UV pulses with 170−220 fs and 1−2 mW of average power, depending on the wavelength. The UV pulse lengths at the sample position were measured using difference frequency generation (DFG) with the fundamental 800 nm LIBRA in a BBO crystal. The difference frequency was then detected either on a photodiode to measure the pulse length or spectrally dispersed on a spectrometer to analyze the chirp. The detected DFG signals for the pump and the two probe wavelength can be viewed in the Supporting Information. The 275 nm wavelength was chosen for the pump because it lies at the absorption maximum of 7-DHC. The pump was aligned over a motor-driven delay stage so that the timing between the pulses at the sample could be regulated. To avoid polarization absorption effects due to the pump polarization, a depolarizer was used, which broadened the pulse to 280 fs (Figure SI1, Supporting Information); however, experimentally, the exclusion of possible signal contributions due to linear dichroism has not yet been investigated. For the probe, two different wavelengths were chosen, 285 and 300 nm. These where prechirped via a grating compressor. A Berek compensator (NewFocus 5540) was used as a quarter waveplate to create the circular polarization. The polarization phase was retarded by 90° for every other pulse via a Pockels cell (Series 1147-RTP, Döhrer Elektrooptik), that is, the polarization was switched from right- to left-circular for every other pulse. A sketch of the experimental setup and timing scheme of the applied voltage for the Pockels cell can be seen in Figure 4. The Pockels cell was aligned as described by

Figure 3. Measured transient CD kinetics of 7-DHC at probe wavelengths of 300 (green) and 285 nm (blue) after subtracting the steady-state CD value. The single exponential fit of the data is represented by the red curves and has time constants of 1.1 ± 0.2 (300 nm) and 1.9 ± 0.3 ps (285 nm).

measurements were performed before and after irradiation to ensure the integrity of the sample. The rapidness of the decay of the photoinduced CD signal change measured provides evidence that the model presented by Anderson et al. describes the ring opening of 7-DHC. The slight discrepancy in our time constants for the two probe wavelengths is probably due to the nonoptimal signal-to-noise ratio in our measurements as the change in CD is such a weak event. Also, a precise determination of the subpicosecond time constant in the TA measurements in ref 5 was hindered by the strong solvent spike that overlaps with the TA signal in that time region. Hence, associating the single-exponential time constant found in our TRCD data with the shortest one found by Anderson et al. and that in our own TA measurements is reasonable. With the novel femtosecond TRCD setup, which allows us to record transient chirality changes of small molecules in the UV range with sub-280 fs temporal resolution, the ultrafast process of the ring opening of 7-DHC was detected. This provided further evidence that the subpicosecond time constant observed in TA measurement is associated with the ring opening. The emerging tool, TRCD, is ideally suited to complement standard time-resolved studies by supplying information on global configurational changes in molecules. The choice of pumping and probing in the near-UV is also well-suited because the absorption of many biological samples lies in this wavelength range. Therefore, the work presented was not only able to clarify the specific photophysical question of the ring opening of 7-DHC, but with, it we were able to further establish a tool for investigating ultrafast processes in the UV that influence the chirality.

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EXPERIMENTAL SECTION The 7-dehydrocholesterol (7-DHC) was purchased (SigmaAldrich) and used without further purification. The 7-DHC was dissolved in methanol of spectroscopic grade at an optical density between 0.35 and 0.7 OD at 283 nm and filled into a standard 1 mm quartz cuvette. Steady-state absorption and CD measurements prior to and after each time-resolved experiment ensured sample integrity. The steady-state absorption (steadystate circular dichroism) measurements were performed on a JASCO-530 (JASCO-715) spectrometer. Time-resolved circular dichroism (TRCD) measurements are based on a pump−probe setup, where the depolarized pump induces the chiral change in the sample. Subsequently, the probe

Figure 4. A schematic representation of the experimental setup.

Dartigalongue and Hache,22 and the grating compressor was used to compensate for the length of the Pockels cell so that the pulses were optimally compressed at the sample position. The pump and the probe beams were focused into the sample by means of 75 cm focal length UV fused silica plano-convex 184

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(14) Trifonov, A.; Buchvarov, I.; Lohr, A.; Würthner, F.; Fiebig, T. Broadband femtosecond Circular Dichroism Spectrometer with White-Light Polarization Control. Rev. Sci. Instrum. 2010, 81, 043104/1−043104/6. (15) Gliesing, S.; Reichenbächer, M.; Ilge, H.-D.; Fassler, D. Bestimmung der Extinktionskoeffizienten der Photoisomeren von 3β-Hydroxy-cholest-5,7-dien (Provitamin D3). J. Prakt. Chem. 1987, 329, 311−320. (16) Maessen, P. A.; Jacobs, H. J. C.; Cornelisse, J.; Havinga, E. Photochemistry of Previtamin D3 at 92 K: Formation of an Unstable Tachysterol3 Rotamer. Angew. Chem., Int. Ed. Engl. 1983, 22, 718−719. (17) Rasmusson, M.; Tarnovsky, A. N.; Åkesson, E.; Sundström, V. On the Use of Two-Photon Absorption for Determination of Femtosecond Pump−Probe Cross-Correlation Functions. Chem. Phys. Lett. 2001, 335, 201−208. (18) Kovalenko, S. A.; Ernsting, N. P.; Ruthmann, J. Femtosecond Hole-Burning Spectroscopy of the Dye DCM in Solution: The Transition from the Locally Excited to a Charge-Transfer State. Chem. Phys. Lett. 1996, 258, 445−454. (19) Kovalenko, S. A.; Dobryakov, A. L.; Ruthmann, J.; Ernsting, N. P. Femtosecond Spectroscopy of Condensed Phases with Chirped Supercontinuum Probing. Phys. Rev. A: At., Mol., Opt. Phys. 1999, 59, 2369−2384. (20) Dietzek, B.; Pascher, T.; Sundström, V.; Yartsev, A. Appearance of Coherent Artifact Signals in Femtosecond Transient Absorption Spectroscopy in Dependence on Detector Design. Laser Phys. Lett. 2007, 4, 38−43. (21) Tang, K.-C.; Rury, A.; Orozco, M. B.; Egendorf, J.; Spears, K. G.; Sension, R. J. Ultrafast Electrocyclic Ring Opening of 7-Dehydrcholesterol in Solution: The Influence of Solvent on Excited State Dynamics. J. Chem. Phys. 2011, 134, 104503/1−104503/13. (22) Dartigalongue, T.; Hache, F. Precise Alignment of a Longitudinal Pockels Cell for Time-Resolved Circular Dichroism Experiments. J. Opt. Soc. Am. B 2003, 20, 1780−1787.

lenses (Thorlabs LA4716). After the sample, the pump beam was blocked, and the probe beam was detected by a photodiode and analyzed with a computer (system commercially available from Pascher Instruments). The TA measurements were done with the same setup except that the Pockels cell was turned off and the Berek in the probe was rotated to produce linear polarization.

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ASSOCIATED CONTENT

S Supporting Information *

Difference frequency cross-correlation curves for the determination of the UV pulse lengths. This material is available free of charge via the Internet at http://pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Phone: +49 (0)3641 206332. Fax: +49 (0)3641 206399.

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ACKNOWLEDGMENTS This research is financially supported by a grant from the German Science Foundation (Di 1517/2-1). Furthermore, we thank Prof. Dr. Jürgen Popp, Prof. Dr. Michael Schmitt, and Ronald Siebert for helpful discussions.

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REFERENCES

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