RICERCA ITALIANA     

Research program

Vibrational dynamics and relaxation in densified glasses and confined disordered systems

University Co-ordinator

Università degli Studi di PERUGIA - FISICA - ()

Research Unit Leader

Francesco Sacchetti

Description

The Research Unit (RU) of Perugia is internationally known for the competence gained in the use of neutron scattering. It is well known that the RU of Perugia substantially contributed to both the design and the construction and commissioning of the spectrometer BRISP (BRIllouin SPectrometer), installed at the high flux reactor of the Institut Laue-Langevin (ILL, Grenoble), which is the most powerful steady flux neutron source in the world. This spectrometer is a unique tool since it allows for the measurement of the dynamic structure factor in the momentum transfer range between 1 nm^-1 and 20 nm^-1 with an energy transfer of tens of meV. This dynamic region is of fundamental importance for the study of collective modes in condensed matter, that is density fluctuations at characteristic time of ps and relatively short wavelength between 0.5 nm and 5 nm. The present authors contributed to this field by studying the collective dynamics in liquid metals [27] and water [24]. In addition, the RU of Perugia has also a considerable experience in the study of single particle dynamics using elastic, quasi-elastic and inelastic neutron incoherent scattering. The instruments devoted to these last studies are mainly time of flight spectrometers and back-scattering spectrometers which cover a frequency domain from GHz up to THz with momentum transfer between 1 nm^-1 and 20 nm^-1. Some of the most relevant results obtained at the RU are related to the dynamics and relaxation of biomolecules in a glassy matrix and the protein hydration water in the time range from ps to ns [20,28]. In general, the scientists of the RU of Perugia performed some 150 experiments of neutron scattering in last 10 years at the European large scale facilities and at ILL (Grenoble) in particular. This performance shows the capability of performing neutron scattering experiments in a constant way at the most advanced neutron sources.
From this starting point we propose a detailed neutron scattering study of the dynamics and relaxation, with frequency between GHz and THz, of the confined water by focusing on two rather different specific systems which have great interest for technological and bio-technological applications: hydration water of nafion® and hydration water of proteins.

I) The first activity field of the present project is devoted to the collective and single particle dynamics of the water absorbed by nafion. In this system the water molecules mainly interact with the “sulfonic” groups which carry about 6 water molecules. At room temperature at the maximum hydration (50% by weight, that is about 30 molecules per sulfonic group), the collective dynamics of water in nafion showed the existence of the typical fast sound which is present in bulk water [26]. This is an effect which gives rise to a collective mode velocity in the THz range much higher than that one observes at low frequency, for instance, by means of the light Brillouin scattering. Light scattering experiments, previously performed in collaboration with the RU of L’Aquila and with the LENS in Florence, showed that the collective excitations with frequency in the GHz range propagate with a velocity of 1250 m/s against 3040 m/s observed in the THz range with neutron scattering [24]. It should be also observed that the velocity of 1250 m/s we determined at low frequency is smaller than both the velocity in dry nafion (about 1700 m/s) and in bulk water (about 1500 m/s). Starting from these observations the collective dynamics will be investigated as a function of both the hydration level and the temperature, to understand the relationship between confinement/interaction with the matrix and the undercooling and the connected glass formation.
In this sort of experiments we will employ heavy water D2O to emphasize the coherent (collective) signal.
Since the hydrogen atoms show a very large incoherent neutron cross section (about 80 barn) as compared to other atoms, the single particle dynamics, which will be also studied as a function of hydration level and temperature, can be easily enhanced in the samples hydrated by means of H2O.
The bulk of the information on the collective and single particle dynamics should allow to get some experimental evidence useful to understand the glass formation and the possibly present liquid-liquid transition mechanisms [23].


II) The second activity field is devoted to the study of the collective and single particle dynamics of the Maltose Binding Protein, which is a model protein important for the maltodestrine transport in the Escherichia coli through the external cell membrane. More important, when this protein bounds to the maltose, it changes its conformation with the closure of its domains giving rise to an open-close transition. This special effect has a potential application in the field of bio-sensors for instance[29]. The presence of hydration water is of fundamental importance for the conformation change to happen, therefore it is very important to study its fast and slow dynamics. The Maltose Binding Protein is not commercially available, particularly if one considers the rather large amount one needs in neutron scattering experiments (mass of 1 g and above). In order to solve this problem a collaboration with the Deuteration Laboratory (D-Lab), a facility established as a collaboration between the Institut Laue-Langevin and the European Molecular Biology Laboratory of Grenoble to produce deuterated samples of biological interest to perform neutron scattering experiments, has been established. Within this collaboration, D-Lab will produce for the present project a sample of Maltose Binding Protein in the hydrogenated and deuterated forms. It should be observed that the production of the deuterated samples implies very long production time and the cost of the precess is fairly high and implies also the set up of very complex procedures of molecular biology.
The single particle dynamics of the biomolecule can be easily determined by experiments performed on the hydrogenated sample by hydration with heavy water to make negligible the water signal, while the single particle dynamics of the water can be observed by using a deuterated sample hydrated with H2O.
Recently, the collective dynamics of the hydration water of the protein Ribonuclease [25] at room temperature and 70% hydration level (by weight of water with respect to the dry protein) has been determined using the BRISP spectrometer. Similarly to the case of the water confined in nafion, we observed the typical fast sound originally observed in bulk water [26]. It is important to extend the study at low hydration degree, as it can help in identifying the role of the direct interaction between protein and water. Indeed at low hydration level this interaction must be more important. In the recent experiment [25] it has been observed that at the 70% hydration level the collective mode velocity is higher than that in bulk water since we found a velocity of 3500 m/s against 3040 m/s observed in water [26]. The experiment at low hydration level is quite difficult because of the reduced signal due to the interfacial (confined) water. Therefore to this purpose it is quite important to perform the experiment using a deuterated protein which will help in determining the contribution of water against that of the protein.
As we said in the section “State of the art”, the first hydration shell shows dynamic properties similar to those characteristic of a glassy system. We will perform different experiments as a function of temperature to investigate the dynamic mechanism which governs the glass transition and the fragile-strong transition in these systems.
We recall also that a collaboration with Dr. M. Tarek (CNRS, Nancy) and Dr. D.J. Tobias (University of California) has been already established to perform molecular dynamics simulations on the Maltose Binding Protein as a function of hydration level and temperature. Tarek and Tobias are among the major experts in the simulation studies of the protein dynamics and the contribution from this numerical technique is invaluable to the purpose of interpreting neutron scattering results at the microscopic level.

The experiments devoted to the study of the collective dynamics of the interfacial/confined water in nafion and proteins will be performed on the BRISP spectrometer, and possibly on the three axis spectrometers available at the ILL. The experiments performed on BRISP allow for an extended and fast survey of the sample dynamics, while the experiments performed on a three axis like IN1 and IN8 allow for some “surgical cut” of the frequency and momentum space. The experiments to study the single particle dynamics will be performed mainly on the time of flight spectrometers (IN5, IN6) and backscattering spectrometers (IN10, IN13, IN16), again installed at the Institut Laue-Langevin. The use of different instruments allows to access a wide frequency and momentum region.
Since the study of hydration water is rather complex, particularly at low hydration level, it is important to get a good signal to noise ratio. Therefore both the single particle experiments and those devoted to the collective mode study must be designed in order to maximize the statistics of the data. As a consequence the construction of the cell, the sample preparation and the accuracy of the experimental set up on the instrument one is using (sample environment, measurement conditions) are essential steps. The long experience in the use of neutron scattering at the RU of Perugia gives the necessary added value for the success of these experiments.
The long experience gained at the RU in neutron scattering is also specially useful in the data treatment in particular at low momentum and maximum possible energy transfer. In energy-momentum region, where the collective modes develop, it is important to calculate in the best way the multiple scattering contribution, which can alter in a sizable way the collective mode signal, even if it has a rather broad energy band. To this purpose a properly adapted Monte Carlo simulation software has been developed at the RU of Perugia. This software can evaluate the multiple scattering contribution in complex systems, taking into account also the sample container. In addition, a dedicated software written for MATHLAB has been developed to analyse the data produced by the BRISP spectrometer, which is a rather complex instrument with a multidetector having about 40 Mega-pixel in space and time. This software will be further developed to fully exploit the instrument characteristics.
Finally the results obtained in all the systems will be compared with the results obtained in the model glassy systems, even densified, which will be also studied by means of neutron scattering, in collaboration with the other RU of the present project. Also in this case the long experience in performing accurate neutron scattering measurements of inelastic and quasi-elastic scattering will be important in order to design the experiments and to analyse the data.
The comparison between the rather exotic systems previously considered and the model systems should be useful in providing precious information on the relationship between relaxation (and damping) and velocity of the collective modes in connection also with the confinement of the glassy systems.

To the purpose of completing the described research program it is necessary to obtain the following results:

a) Construction of adequate neutron scattering cells to optimise the signal to noise ratio and minimize the multiple scattering. This technical part is rather complex, indeed it is necessary to design cells having a good stability under vacuum, using a not very strong material like aluminium, employ properly designed cadmium shielding to reduce the multiple scattering. In addition the cells should be designed in such a way that the filling is simple and fast, vacuum tight in order to avoid hydration change during the experiment.
b) One should be able to prepare the appropriate samples (both in the case of nafion membranes and in the case of hydrated proteins) in controlled conditions in order to determine the hydration with an accuracy of the order of 1 %. The hydration degree will be controlled using thermo-gravimetric measurements. In the case of nafion an appropriate protocol will be developed in order to clean the membrane if necessary, before the hydration. In the case of biological samples their native state will be controlled during the preparation phase using measurements of circular dichroism and absorption at the RU of Perugia.
c) Nafion. It will be necessary to perform characterization tests using X ray on the dry membrane and on the hydrated membrane with D2O and H2O as necessary. The characterization tests will be performed as a function of the hydration level and as a function of the temperature between 100 K and 300 K. These samples will be employed for the neutron scattering experiments. The Brillouin light scattering experiments in the visible and UV range will be performed in collaboration with the other RU’s to determine the collective dynamics at low frequency and calorimetry experiments will be also performed to complement the neutron inelastic scattering data.
d) MBP. The deuterated protein samples must be characterized using similar procedures as the nafion samples employing X ray and spectroscopic techniques other then light scattering and calorimetry, to be used for neutron scattering experiments.