LPL - Laboratoire de Physique des Lasers

Biomolecules and spectroscopy (BMS)

Toward a characterization of the molecular origin of the indirect effects in the radiation damage

C. Desfrançois, F. Lecomte, N. Nieuwjaer and B. Manil

 

The last experimental achievement of the BMS team has been the development of an alternative technique to get biomolecules and their complexes directly under vacuum in conditions as close as the ones encountered in solution. This molecular source is new, does not exist in France and is based on ultra-soft laser desorption from liquid microdroplets under vacuum. The desorbed complexes, initially present in the liquid, can thus be identified and characterized in the gas phase under carefully chosen conditions in a controlled environment, close to their native forms in solution. This source is based on a concept originally developed by B. Brutschy (University of Frankfurt): large biomolecules are put in the gas phase by non-resonant laser desorption on microdroplets. We have initiated the development of such a source and we have obtained some first promising results. In our device, liquid microdroplets (50 micrometers diameter) are generated on demand by a commercial droplet generator (Microdrop) and injected from 300 mbar into a high vacuum chamber (10-6 mbar) through differential pumping stages. Upon irradiation by mid-IR broad band laser pulses (centered on an absorption band of the solvent), analyte ions (and neutrals) are ejected directly under vacuum. The desorption occurs at the entrance of a Paul Trap into which ions are guided by a weak electrostatic field, in order to avoid any fragmentation or activation during collisions with the buffer gas in the trap. Ions are then ejected from the trap, accelerated, focused and detected by microchannel plates in a second vacuum chamber (10-7 mbar). The preliminary results show mass-selected non-covalent complexes, which proves that biomolecular desorption is soft enough to preserve weak molecular interactions. This source then represents an important breakthrough in the field of gas-phase analysis techniques applied to relevant biological systems and will give us the unique opportunity to get insights in the damage processes at the molecular level by the use of gas phase analysis techniques.

 

By coupling of this experimental set-up with an irradiation platform (see figure), we will irradiate the microdroplets (solvent + biomolecules) by simply charged ions with a kinetic energy of some tenths of keV. This irradiation will create a secondary particles radiolysis cascade (solvated electrons and radicals), only into a superficial layer (first hundreds of nanometers) of the droplet. This corresponds to the physico-chemical stage of the irradiation, which is well-known through the study of water radiolysis. During the next (chemical) stage, the free radicals are going to react with biomolecules in the non-irradiated area. This second stage is far less well characterized for water radiolysis, and almost completely not understood in the case of biomolecules. By mass-analyzing the biomolecular products after irradiation and radiolysis inside the microdroplets, our experimental method will give us the possibility to study, for the first time, these radical chemistry processes linked to indirect effects, being free from any other radiation contribution.

 

Schematic layout of the experimental apparatus

Schematic layout of the experimental apparatus

 

Contact
Bruno Manil

 

Reference
R. Lozada Garcia, S.D. Leite, F. Lecomte, N. Nieuwjaer, C. Desfrançois and B. Manil (in preparation)

 

Main financial support: BIORAD (2015-2018, INCa-INSERM)

 

Structural characterization of biomolecules

C. Desfrançois, F. Lecomte, N. Nieuwjaer and B. Manil

 

In the last years, the BMS team has been involved in fruitful collaborations with other experimental groups, in order to extend our research activity to the study of model biomolecules through a combined laser-spectroscopy and mass-spectrometry approach. To that end, we perform IR spectroscopy of neutral isomer-selected or ionic mass-selected systems (nucleic acid bases, amino acids, peptides, drugs) and their non-covalent complexes in the gas-phase for which structural assignment is done through comparison with high-level quantum chemistry calculations. Recently, we performed an experiment with the group of G. Grégoire (ISMO-Orsay) to study the vibronic spectra of protonated hydroxypyridines (HPH), whose derivatives are endogenous or synthetic photosensitizers which could contribute to solar radiation damages. The study of their excited states could lead to a better understanding of their action mechanisms. We obtained the ultraviolet (UV) spectra of the protonated 2-, 3- and 4-hydroxypyridine. They display well-resolved vibrational structures, with a clear influence of the position of the OH group. These results are interpreted with excited states calculations at the coupled cluster CC2 level.

 

We aims also to study liganded nano-clusters (see figure), after mass-selection in the gas phase, using IRMPD laser spectroscopy at CLIO facility (IR Free Elecron Laser in Orsay). The advantage of the mass-selection is to avoid all contribution from the solvent or from multiple cluster sizes and to simplify the comparison with quantum chemical calculations. The IR spectra (performed in a wide range of IR wavelengths: 3 to 100 µm) will hopefully give an unambiguous signature for the structural assignments. The goal is to get a better understanding of the nature of the metal-ligand interactions connected with some physical and chemical processes, which are involved in various biological applications (grafting, functionalization, phototherapy, radiosensitizers …) of nanoparticles.

 

Calculated structure of a liganded gold nanocluster (Au11(((PC6H4-CONH-CH3)3)7Cl3)

Calculated structure of a liganded gold nanocluster (Au11(((PC6H4-CONH-CH3)3)7Cl3)

 

Contact
Charles Desfrançois

 

References

Lozada Garcia R.R., Nieuwjaer N., Desfrançois C., Lecomte F., Leite S.D., Manil B., Broquier M., Grégoire G.,
Vibronic spectra of protonated hydroxypyridines: contributions of prefulvenic and planar structures,
Physical Chemistry Chemical Physics, 19, 12, 8258-8268, (2017)
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T.N. Le, J-C. Poully, F. Lecomte, N. Nieuwjaer, B. Manil, C. Desfrançois, F. Chirot, J. Lemoine, P. Dugourd, G. van der Rest, G. Grégoire,
Journal of the American Society for Mass Spectrometry, 24, 1937, (2013).

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