Techniques and tools
Synthesis and chromatographies
Synthesis
In our laboratories, in addition to classical organic chemistry apparatus, we are equipped with three leading-edge instruments which allow:
- Automated synthesis of peptides under microwave activation thanks to Liberty 1® instrument from CEM. This apparatus is based on solid-phase peptide synthesis and leads to higher yields and purity compared to traditional peptide synthesis. Moreover, the microwave activation enables a much faster access to peptides.
- Microwave synthesis using Discover® SP system from CEM. During a synthesis, the thermal effect associated to microwave irradiation induces a constant heat on each molecule of the reaction mixture. This mechanism leads to much faster kinetics of reaction, and typically induces higher yields and even potentially different selectivity.
- Continuous-flow hydrogenation thanks to H-CUBE® from ThalèsNano. This instrument is able to produce hydrogen in situ thanks to water electrolysis, which limits hydrogen storage and associated risks. The hydrogenation reaction is then performed in a continuous flow manner using supported catalysts (CatCart®).
Chromatographies
In order to analyze and purify synthetic products, our lab is equipped with two high performance liquid phase chromatography instruments:
- An analytical ACQUITY® UPLC® from Waters equipped with a photodiode array (PDA) detector and a simple quadrupole mass spectrometer (SQD). This instrument is mainly used to monitor reactions or to characterize combinatorial mixtures. In particular, the coupling to mass spectrometry allows verifying masses of the different products formed during a reaction. Compared to a classical HPLC, the UPLC® system can be used at higher flow rate, leading to shorter time of analyses (~5 minutes) without altering chromatographic performances.
- A dual analytical/preparative Autopurification-HPLC instrument from Waters equipped with a UV-Vis detector (single or double wavelengths) and a simple quadrupole mass spectrometer (3100). This machine presents mainly two interesting features: reaction mixtures are first analyzed using analytical chromatography in order to determine optimal conditions for their purification on preparative-scale. These purifications can then be performed on several hundreds of milligrams for a single injection. This instrument is particularly suited to purify polar compounds with solubility in cater or organic solvents such as acetonitrile or methanol.
Self-assembly characterization by scattering techniques
Scattering techniques are used in collaboration with physicists Prof. E. Buhler (Paris Diderot University) et Dr. M. Rawiso (ICS) in order to determine sizes and shapes of our supramolecular self-assemblies.
- Light scattering experiments (both static and dynamic) are performed on a 3D LS spectrometer in the laboratory of Prof. Buhler. Static light scattering (SLS) is used to determine, for instance, the molecular weight Mw and the gyration radius Rg of self-assembled structures and supramolecular polymers. Dynamic light scattering (DLS) is used at a fixed angle to measure the mean particle size of our assemblies (hydrodynamic radius RH). This last experiment can also be performed at the ICS on a Malvern Zetasizer Nano ZS apparatus.
- Small angle neutron scattering (SANS) experiments are typically performed on large instruments such as laboratory Léon Brillouin (LLB) located at CEA Saclay (nearby Paris) or Institute Laue-Langevin (ILL) in Grenoble. These experiments indeed require the use of a nuclear reactor in order to generate neutrons which will interact with the atom nuclei of the studied structure and detectors are typically more than 5 meter long. With such experiments, internal structures but also size and mass of our self-assemblies can be determined. They can be complemented by static light scattering experiments.
- Small and wide angle X-ray scattering (SAXS and WAXS, respectively) experiments are usually performed at ICS in the group of Dr. Rawiso, who is equipped with two diffractometers : Nanostar from Bruker (small and wide angles) and MicroMax with a rotating anode generator from Elexience. Unlike neutrons, X-rays interact with electrons which surround the atomic nucleus. Therefore, although SAXS and WAXS also provide information on the size, the mass and the internal structure of the supramolecular objects, these experiments are often complementary to neutron scattering ones. X-ray scattering experiments can also be performed on large Synchrotron instruments such as Soleil in Gif sur Yvette (nearby Paris) or ESRF in Grenoble.
Microscopic characterization
Scanning probe microscopy
Our Lab has a variety of scanning probe microscopy techniques:
- Atomic Force Microscopy (AFM) in force spectroscopy mode. It’s house-made AFM combined with fluorescence optical microscope. This technique is used to manipulate a single molecules and to measure the interaction forces at single molecule level.
- Multimode AFM Nanoscope 8 imagery, in tapping mode and Peak-Force mode.
We have both expertise and a wealth of experience in the development of innovative solutions to a broad range of problems in the domain of self-assembly systems, biopolymers, gel,… and the imagery at nanometer and micrometer scales in liquids (buffer solutions) and controlled environment.
Transmission Electron Microscopy
Transmission Electron Microscopy (TEM) is frequently used in our team in order to probe the morphology of the self assembled objects synthesized. As it requires a small amount of material and can be performed very fast, we often use it as a first screening analytical method in order to select the samples of interest that can then be analyzed with other more time consuming techniques. Combined with scattering techniques such as DLS, SANS or SAXS as well as imaging techniques such as AFM, it allows to obtain structural insight on the self-assembled systems from the team. When the sample to study displays a gel behaviour, Freeze Fracture Electron Microscopy is used in order to image the gel in its native state. The samples are analyzed on a Phillips CM12 electron microscope operating at 120kV.
Spectroscopic characterization
In our institute, several spectroscopic apparatus, managed by the polymer characterization service, are shared between the different research groups. Among them, some are used to determine the purity of our synthetic organic products (NMR 400 MHz Spectrometer – Bruker and MALDI-TOF Autoflex – Bruker). Some others are used to study the optical properties of our molecules and self-assemblies. In particular, our institute is equipped with the following instruments:
- FT-IR spectrometer Vertex 70 from Bruker. This apparatus can be used either in the ATR mode or for the study of self-assemblies in solution. Therefore, we possess several IR cells that can be used either with organic solvents or aqueous ones.
- UV-Vis-NIR Cary 5000 spectrometer from Agilent. This instrument can be used to carry out variable-temperature experiments, to measure optical properties of thin films, or even to perform reflectance measurements on our samples.
- FluoroMax-4 spectrofluorometer from Horiba, which is used to determine the steady-state fluorescence properties of our molecules and self-assemblies in solution.
