Since the development of mode-locked laser sources [1
], ultrafast optical spectroscopy has provided invaluable insight into the light-triggered dynamical processes in many different systems of interest for physics, materials science and (bio)chemistry [2
]. While several techniques with growing level of complexity and sophistication have been developed [4
], transient absorption (TA) still remains a very powerful and versatile approach, delivering a rich information content with a comparatively simple experimental setup. TA typically works in a stroboscopic fashion, using two synchronized pulses, the pump and the probe. The pump pulse is resonant with a transition of the system under study, triggering a photoinduced process whose time course is followed by measuring the absorption change of the time-delayed probe pulse, which should ideally be as broadband as possible in order to deliver the maximum amount of spectroscopic information on the system [6
]. The time resolution of TA spectroscopy is determined by the so-called instrumental response function (IRF). In TA spectrometers with relatively narrow-band (>100 fs duration) transform-limited (TL) pump and probe pulses, the IRF is the cross-correlation of the intensity profiles of pump and probe pulses [10
]. When dealing with broadband probe pulses, it can be shown that, independently from their chirp, the temporal resolution is determined by their TL duration, provided that the probe pulses are detected through a narrow-band filter [11
]. The ability to generate pump pulses with duration of just a few optical cycles [1
] and probe pulses with ultrabroad bandwidth has pushed the time resolution in the visible and near-infrared (NIR) ranges to the sub-10-fs domain, allowing the observation of primary light-induced processes in molecules and solids, which govern important biochemical processes such as photosynthesis [14
] and vision [15
The ultraviolet (UV) wavelength range is of great interest for ultrafast spectroscopic investigations due to several reasons. First of all, the UV range is resonant with electronic transitions of many small molecules with fairly simple excited state energy level structure, whose photoinduced dynamics can be accurately modeled using ab initio computational approaches; ultrafast TA spectroscopy in the UV can thus be used to benchmark the accuracy of such methods [16
]. In addition, the UV matches the absorption spectrum of many biomolecules, which present cyclic aromatic rings with strongly allowed π→π* transitions. Examples are the nucleobases (adenine, cytosine, guanine, thymine, and uracil) and their multimers up to the double-stranded DNA helix, as well as the aromatic amino acids (tryptophan, phenylalanine, and tyrosine), all of which have strong absorption bands in the 250–300 nm wavelength range (≈4–5 eV). While the main biological functions of these biomolecules are not directly related to their interaction with light, still the study of their ultrafast optical response is very important. In DNA the absorbed UV photon energy could initiate a variety of photoreactions, involving substantial rearrangements of the molecular structure, that corrupt the information encoded in the base sequence. Nevertheless, the quantum yield of such photoproducts is remarkably low, because DNA manages to dissipate the absorbed energy very efficiently, mainly through harmless ultrafast non-radiative decay processes [18
]. This self-protecting property of DNA is not just an interesting feature of its photophysics, but it is an essential requirement for the very existence and the replication of life, which relies on the stability of the hereditary genetic information. In proteins, aromatic amino acids have been proposed as built-in structural markers [22
] whose TA signal is able to deliver information on their secondary structure and on the misfolding processes that lead to aggregation, fibril formation and are the precursors of several neurodegenerative diseases.
Despite the strong scientific motivation and the great potential interest, pushing the time resolution of UV TA spectroscopy to the 10–20 fs domain, which is nowadays customary in the visible range, requires addressing several technological challenges:
the generation of few-optical-cycle pulses in the UV is difficult, due to the lack of suitable broadband gain media, either linear or nonlinear, in this frequency range;
standard methods for the temporal characterization of ultrashort pulses, based on nonlinear frequency up-conversion processes such as second harmonic generation (SHG), cannot be applied to the UV, and more complex techniques are required;
due to the proximity with the onset of electronic transitions, all transparent materials in the UV display a strong dispersion, causing a frequency chirp; this calls for a careful design of the spectroscopic system to minimize the number and the thickness of refractive optical elements and prevent the loss of temporal resolution.
This paper describes our recent efforts at developing an ultrafast TA spectroscopy system in the UV with high temporal resolution for the study of the ultrafast optical response of biomolecules. The system combines sub-20-fs pump pulses tunable in the 3.35–4.7 eV range (corresponding to 260–370 nm) with broadband probe pulses obtained by white-light-continuum (WLC) generation in different materials. The paper is organized as follows: Section 2
reviews the methods used for the generation and characterization of ultrashort UV pulses; Section 3
describes the experimental setup of the UV TA spectrometer; Section 4
characterizes the performance of the system and presents representative experimental results; finally, Section 5
draws conclusions and discusses prospects for applications and improvements of the system.
4. Experimental Results
a reports a sequence of UV pump pulses generated either by SHG or by SFG of the NOPA output, which cover a wavelength range from 260 to 370 nm (4.75 to 3.35 eV). The figure also plots the spectral phases of the pulses, retrieved by 2DSI, which are essentially flat: the corresponding temporal intensity profiles, shown as solid lines in Figure 4
b–d, reveal sub-20-fs pulses, with a FWHM duration very close to that of the corresponding TL pulses (with intensity profiles shown as dashed lines): 16.2 fs for the 270-nm pulse, 18.6 fs for the 290-nm pulse and 8.4 fs for the 340-nm pulse.
shows spectra of the WLC generated in the different materials pumping with the FW or with the SH, which -as previously discussed- cover different wavelength ranges. For each spectrum we calculated the wavelength dependent root-mean-square (rms) of fluctuations between consecutive pulses as:
) is the signal measured by the spectrometer at a given probe wavelength for the i
-th laser shot and
is the average probe signal.
The wavelength dependent rms fluctuations of the WLC measured for N = 300 consecutive pulse pairs are reported in Figure 5
as shaded areas. One can see that the WLC generated in sapphire has the best noise properties, with fluctuations below 0.2% over most of the detection range; the WLC generated in CaF2
driven by the FW has slightly higher fluctuations but adds a very significant spectral interval from 330 to 450 nm; finally, the WLC generated in CaF2
driven by the SH displays higher but still acceptable fluctuations which are lower than 0.4% over most of the detection range.
The overall stability of the WLC is quite good: in fact, a 0.2% rms fluctuation corresponds to a noise <10−4
for the ∆T/T signal when averaged over 500 consecutive pulse pairs, which correspond to 1 second measurement time. This high stability of the probe is crucial for TA measurements with UV pump pulses; in water solvent, in fact, two-photon absorption of the pump can produce hydrated electrons [80
], which display broad and long-lived absorption spectra in the visible [81
]. To avoid or minimize such spurious signals, the pump fluence should be kept as low as possible, thus calling for a high sensitivity of the TA setup.
In the following we present a few examples of ultrafast TA spectroscopy results using our high time resolution apparatus. As exemplary molecular system we use pyrene (C16
, chemical structure shown in Figure 6
), a polycyclic aromatic hydrocarbon consisting of four benzene rings. Pyrene purchased from Sigma-Aldrich was used as received, dissolved in methanol, and flown in a 200-μm-thick cuvette with 200-μm-thick entrance and exit windows. The absorption spectrum of pyrene, reported in Figure 6
, displays an intense band peaking at 3.7 eV (334 nm), with well-resolved vibronic replicas at 3.9 eV (319 nm) and 4.1 eV (306 nm), followed by a second band peaking at 4.6 eV (270 nm) with a well visible replica at 5 eV (250 nm). According to the well-known electronic structure of pyrene, the ground state has Ag
state), while the first excited state has B3u
state); however, the oscillator strength of the S0
transition is very weak so that it is essentially optically forbidden [82
]. The first “bright” excited singlet state of pyrene is of B2u
state) and the S0
transition peaks at 3.7 eV; a second bright excited state is the S3
state (or 21
state) with the S0
transition peaking at 4.55 eV (270 nm). Following excitation of the bright S2
states, rapid internal conversion (IC) to the dark S1
state occurs, mediated by conical intersections. While the S2
process has been studied in detail [83
], the S3
process is less explored. Here we exploit our tunable pump and broadband probe to follow the dynamics of both these processes.
shows TA maps obtained by pumping with SFG at 340 nm and probing in the UV and in the visible range respectively. We start by discussing the UV range (Figure 7
a). Around time zero we observe a short-lived signal that is assigned to the solvent response and is due to TPA of one pump and one probe photon. After this we note a strong positive peak at 334 nm, corresponding to photo-bleaching of the S0
transition, resonant with our pump, with the well resolved vibronic replicas at 320 and 306 nm. At longer wavelengths around 360–370 nm we observe a photoinduced absorption (PA) band, assigned to the S1
state, which is not formed instantaneously, but rather grows on the 200-fs timescale. In the visible range we observe a PA band, assigned to the S2
state, which peaks at 580 nm, forms instantaneously and decays on the 200 fs timescale, and two long-lived PA bands peaking at 515 and 470 nm, assigned to S1
. Taken together, these data confirm ultrafast IC from the photoexcited S2
to the S1
state; the S2
PA dynamics at 585 nm, displayed in Figure 8
b, shows a decay with ≈85-fs time constant, in very good agreement with previous results [83
a shows the 2D TA map, as a function of probe wavelength and delay, following excitation of the S3
state at 270 nm. At early times we observe a negative ∆T/T signal which peaks at 550 nm and is assigned to a PA from the photoexcited S3
state; this band rapidly shifts to the red at 580 nm (red spectrum in Figure 9
b), matching the PA band of S2
; at longer delays, the PA spectrum of S1
reappears, with characteristic peaks at 370, 470 and 515 nm. These data are consistent with a two-step S3
IC process; the TA dynamics, displayed in Figure 10
, show that this cascaded process occurs, as expected, on a longer 500 fs timescale.
Finally, Figure 11
a shows the ∆T/T dynamics at 385 nm, corresponding to the PA from the S1
state; the signal is clearly modulated by a long-lived oscillation, with 85-fs period (corresponding to a 392 cm-1
frequency, as evidenced in panel (b)), which is observed thanks to the high temporal resolution of our setup. A wavelength dependent Fourier transform (panel (c)) reveals a second mode at 410 cm-1
at wavelengths between 335 and 342 nm, which closely matches the Raman ground state mode of 408 cm−1
]. Since the 392 cm-1
mode shows a clear amplitude minimum at a wavelength of 370 nm, which corresponds to the peak of the S1
PA band, we assign the oscillation to a vibrational wavepacket on the S1
state, which survives the IC process and is indicative of a vibrationally coherent photophysics in pyrene [87
In this paper, we described a TA spectroscopy system in the UV spectral range, for the study of the ultrafast optical response of biomolecules. The setup, which is driven by a 100-fs regeneratively amplified Ti:sapphire laser, combines sub-20-fs UV pump pulses with broadband white-light-continuum probe pulses. The ultrashort UV pump pulses are generated by frequency up-conversion (SHG or SFG) of a visible ultrashort NOPA output, and are tunable in the 260–370 nm range. Direct frequency-doubling of the ultrashort visible light delivers broadband UV pulses with carrier wavelength from 260 to 290 nm. SFG between the NOPA pulses and a fraction of the laser fundamental wavelength produces pulses with bandwidth in the 320–370 nm range. After suitable compression stages, the pulse duration ranges from 8.4 fs (for the 340 nm pulse) to 18.6 fs (for the 290 nm pulse).
The broadband probe pulses are obtained by white-light continuum generation in sapphire or CaF2 plates, and cover the spectral ranges from 270 to 710 nm with energy fluctuations of 0.2–0.4% rms, corresponding to a noise <10−4 for the ∆T/T signal averaged over 1 s. The ultrashort duration of the pump pulses and the broad bandwidth of the probe pulses determine the extremely high temporal resolution of the pump-probe measurement.
The TA spectroscopy system was applied to study ultrafast processes in pyrene in methanol. After excitation of the bright optical transitions S0→ S2 (occurring at 3.7 eV) and S0→S3 (at 4.6 eV), the system undergoes ultrafast internal conversion from either S2 or S3 to the dark S1 state. Thanks to the ultrashort duration of the pump pulses and the ultrabroad bandwidth of the probe pulses, we could successfully track the internal conversion process, which occurs on hundreds of femtoseconds timescale. In addition, the temporal resolution allowed resolving an impulsively excited molecular vibration, with 85-fs period, which survives the internal conversion process, pointing to a vibrationally coherent photochemistry in pyrene.
The tunability of the pump pulses in the UV spectral range, the sub-20-fs temporal resolution and the broad spectral coverage of the probe contribute to make this TA system a unique, powerful, and versatile tool for the study of many biomolecules, which exhibit absorption in the 250–350 nm wavelength range. This system will allow tracking fundamental biochemical processes such as the photoprotection mechanisms in DNA and the secondary structure of proteins based on the response of aromatic amino acids.