Final Report Summary - CRYSTILS (Crystallization in Ionic Liquid Solutions)
The aim of the project was to establish fundamental relationships between composition (thus structure and properties) of the ILs and the crystallization mechanism in aqueous solution. The particular approach of the project consisted in looking at ILs from the perspective of their effect on water structure dynamics and respective consequences of aqueous solvent modification for crystal growth. The methodological approach intended to take advantage from the possibility to fine-tune hydration characteristics of ILs in order to perform a systematic study on crystal behaviour dependence on solvent structure and propose new and/or advance existing crystallization strategies for inorganic and protein crystals. The applications could be diverse, from the engineering of materials or nanotechnology devices to manufacture of large-tonnage commodity products or high-value specialty chemicals such as pharmaceuticals. Advanced methods for crystallization of proteins in particular, are of special importance for the scientific community because obtaining a good quality crystal of a specific protein often represents a bottleneck in structural genomics.
The specific goals to be achieved were:
- derive some fundamental relationships between composition of ILs and precipitation of inorganic crystals from the perspective of the new model relating action of crystallization additives with their effect on solvent structure dynamics,
- apply principles developed for inorganic systems to control crystallization of model proteins in ILs solutions and verify/determine relationship between composition of IL and precipitation of organic crystals.
The purpose to exploit principles determined for inorganic solids to organic systems was based on the findings that crystallization of inorganic minerals and biomacromolecules from solution can be described by the same fundamental laws13. Furthermore, recent studies imply that kinetics of incorporation of growth units into a crystal surface is diffusion limited for the solution-solid phase transition of common small-molecule ionic substances and for organic macromolecules14. Diffusion in aqueous solution is determined by the water structure and dynamics in solvation shells15. This supported the possibility to apply the approach that considers impact of additives on crystal behavior in terms of the structural properties of the aqueous solvent to understand and control the effect of ILs on crystallization of inorganic solids and as well as protein crystals. Implementation of ILs in industrial crystallization processes could have potential beneficial environmental impact if these liquid salts were used as alternative to commonly employed volatile organic solvents. It is worth noting that this project intended to use ILs in order to manipulate water structure and water is the greenest and the most universal solvent on the Earth.
According to the objectives of the project, novel mechanisms underlying the response of inorganic minerals and protein crystals to the presence of ionic liquid (IL) additives have been established. The revealed apparent similarities between the effect of ILs on the crystallization process of small molecule ionic substances and of biomacromolecules have been explained by considering the general feedback mechanism between water structure dynamics (modified by ILs) and crystal growth. The response of the crystallizing systems to the presence of ILs enabled also to extract some information about particular properties of ILs. Therefore, the model systems studied gave more insight on the action of ILs on the liquid-solid phase transition processes and on the intrinsic features of ILs in aqueous solutions. The unraveled relationships can help in designing new IL-based processes and crystallization strategies.
As a first step, a systematic dependence between the size of the precipitating crystalline particles and the conductivity of ionic liquids (ILs) has been found. The unraveled correlation has been interpreted in the light of the effect of ILs on water structure dynamics and respective consequences for crystal nucleation. The response of the crystal formation process to the presence of specific ILs has been used to extract information about some particular properties of ILs (ion association and hydration) that modify the crystal hydration environment.
Our model system; precipitation of barium sulfate in IL containing solutions; provided further insights into the fundamentals of the nucleation phenomena in aqueous ionic solutions on one hand, and served as a relatively simple probe of hydration and association characteristics of ionic liquids on the other. To the best of our knowledge, this is the first study showing a systematic dependence of a nucleation rate on a given measurable property (conductivity) of IL solutions and providing a theoretical explanation for the observed correlation. The revealed relationship defines the effect of specific properties of ILs (hydration and association) on the hydration characteristics of other ionic species present in solution. The established correlation should be treated as a model that bears qualitative information and it is important to realize that the response of the system to changes in hydration induced by ILs depends on the properties of the particular system.
The principles developed for inorganic system have been applied to control crystallization of model proteins in ILs solutions and verify/determine relationship between composition of IL and precipitation of organic crystals. The influence of complex ionic liquid ions on protein crystallization has been systematized and rationalized by considering the hydration-dependent affinity of the IL ions to the protein surface and the effect of ILs on water structure dynamics in macromolecular solvation shell and in a solution bulk. The recognized distinct effects of IL-surface and IL-bulk water interactions (e.g. initial protein solubilization due to binding and its subsequent salting-out by increasing water surface tension) could be beneficial for example in those systems where crystallization of biomacromolecules is restricted due to their limited solubility. To the best of authors knowledge, this work (published as invited contribution in a special issue of CrystEngComm and selected as Highlight from RSC themed issues on ionic liquids) reports also for the first time hydration modification (caused by induction, via the presence of an additive) of protein charged compartments, with this mechanism being partially responsible for the success of crystallization, in the same way as previously unraveled for inorganic ionic crystals. The general feedback mechanism between ion hydration-water structure dynamics-and crystallization (self assembly) proved to be valid from inorganic to biomacromolecular systems and opens new horizons to control order formation in complex systems.
Using the potential of ionic liquids to shape water in crystallization processes, inorganic crystals with potential functionality have been synthesized. These refer to hollow core calcite rhombohedra with composite (eventually multishell) walls [9], fluorescent vaterite microcapsules with permeable walls and composite calcium carbonate crystals with fluorescent vaterite cores encapsulated inside calcite shells (unique chemistry, mixed structure) [10]. Rhombohedral crystals with internal cavities capable of occupying more that 70 % of crystal volume have been grown encompassing ionic-liquid stabilized gas bubbles in solution. Stabilization of bubbles and bubble aggregates is determined by the effect of ions on water structure dynamics. Composite walls of those crystals could grow in gel-like environment (environment with restricted water mobility or in other words structured water) generated by ionic liquids. Loadable vaterite capsules and mixed-structure composite crystals have been grown in a system where inorganic ions (crystal building units) induce spontaneous emulsification of the water-immiscible ionic liquid and water and build a porous crystal around an emulsion droplet. Unstable (emulsifying) ionic liquid-water interfaces were used to template formation of porous films (COIL). The designed crystalline structures can find practical applications as delivery carriers for the controlled release of compounds, photonic crystals, lightweight fillers, or chemical storage devices for active species. Cavities enable loading and potential use in smart delivery systems. Compositional zoning can be especially advantageous for the affinity-based loading with different target materials and for solubility-controlled release. Fluorescence can be potentially employed for tracking the load or for imaging purposes.
It is important to note, that in all crystallization experiments the concentration of ILs in aqueous solution was kept relatively low, so that water could be considered the main solvent (according to the objectives of the project) and its local structure modification by IL additives was affecting process of crystal nucleation and grow. Although the importance of the water mediated interactions in crystalline order formation has been appreciated (though mainly with respect to protein crystals), there has been no comprehensive theory explaining the effect of other solutes on hydration of crystal building units and respective consequences for crystal nucleation and growth. In CRYSTILS project water has been always considered as a crucial player. And it was shown that character of interaction of ILs with water molecules, that defines partitioning of IL ions to the interfaces (e.g. air/water) and IL ions´ mutual affinity in aqueous solution, determines also effect of those liquid salts on hydration of other solutes (here crystal building units) and therefore ability of the latter to form crystalline order in solution. Therefore outcome of this project creates some common framework connecting chemistry and molecular structure of multicomponent aqueous solution and crystallization processes. Recognition of the relationship between IL-water interactions and crystal formation opens possibility to employ those designer chemicals in crystallization strategies for potential scientific and engineering applications.
As a result of this project knowledge from the field of ILs is transferred to the field of biomacromolecular crystallization in the form of a joint project between Laboratory of Molecular Thermodynamics at ITQB (the Host laboratory of the ERG grant beneficiary) and Laboratory of X-ray Crystallography of Proteins at Faculdade de Ciencias e Tecnologia. The Marie Curie ERG beneficiary is a Principal Investigator in this project.
The specific goals to be achieved were:
- derive some fundamental relationships between composition of ILs and precipitation of inorganic crystals from the perspective of the new model relating action of crystallization additives with their effect on solvent structure dynamics,
- apply principles developed for inorganic systems to control crystallization of model proteins in ILs solutions and verify/determine relationship between composition of IL and precipitation of organic crystals.
The purpose to exploit principles determined for inorganic solids to organic systems was based on the findings that crystallization of inorganic minerals and biomacromolecules from solution can be described by the same fundamental laws13. Furthermore, recent studies imply that kinetics of incorporation of growth units into a crystal surface is diffusion limited for the solution-solid phase transition of common small-molecule ionic substances and for organic macromolecules14. Diffusion in aqueous solution is determined by the water structure and dynamics in solvation shells15. This supported the possibility to apply the approach that considers impact of additives on crystal behavior in terms of the structural properties of the aqueous solvent to understand and control the effect of ILs on crystallization of inorganic solids and as well as protein crystals. Implementation of ILs in industrial crystallization processes could have potential beneficial environmental impact if these liquid salts were used as alternative to commonly employed volatile organic solvents. It is worth noting that this project intended to use ILs in order to manipulate water structure and water is the greenest and the most universal solvent on the Earth.
According to the objectives of the project, novel mechanisms underlying the response of inorganic minerals and protein crystals to the presence of ionic liquid (IL) additives have been established. The revealed apparent similarities between the effect of ILs on the crystallization process of small molecule ionic substances and of biomacromolecules have been explained by considering the general feedback mechanism between water structure dynamics (modified by ILs) and crystal growth. The response of the crystallizing systems to the presence of ILs enabled also to extract some information about particular properties of ILs. Therefore, the model systems studied gave more insight on the action of ILs on the liquid-solid phase transition processes and on the intrinsic features of ILs in aqueous solutions. The unraveled relationships can help in designing new IL-based processes and crystallization strategies.
As a first step, a systematic dependence between the size of the precipitating crystalline particles and the conductivity of ionic liquids (ILs) has been found. The unraveled correlation has been interpreted in the light of the effect of ILs on water structure dynamics and respective consequences for crystal nucleation. The response of the crystal formation process to the presence of specific ILs has been used to extract information about some particular properties of ILs (ion association and hydration) that modify the crystal hydration environment.
Our model system; precipitation of barium sulfate in IL containing solutions; provided further insights into the fundamentals of the nucleation phenomena in aqueous ionic solutions on one hand, and served as a relatively simple probe of hydration and association characteristics of ionic liquids on the other. To the best of our knowledge, this is the first study showing a systematic dependence of a nucleation rate on a given measurable property (conductivity) of IL solutions and providing a theoretical explanation for the observed correlation. The revealed relationship defines the effect of specific properties of ILs (hydration and association) on the hydration characteristics of other ionic species present in solution. The established correlation should be treated as a model that bears qualitative information and it is important to realize that the response of the system to changes in hydration induced by ILs depends on the properties of the particular system.
The principles developed for inorganic system have been applied to control crystallization of model proteins in ILs solutions and verify/determine relationship between composition of IL and precipitation of organic crystals. The influence of complex ionic liquid ions on protein crystallization has been systematized and rationalized by considering the hydration-dependent affinity of the IL ions to the protein surface and the effect of ILs on water structure dynamics in macromolecular solvation shell and in a solution bulk. The recognized distinct effects of IL-surface and IL-bulk water interactions (e.g. initial protein solubilization due to binding and its subsequent salting-out by increasing water surface tension) could be beneficial for example in those systems where crystallization of biomacromolecules is restricted due to their limited solubility. To the best of authors knowledge, this work (published as invited contribution in a special issue of CrystEngComm and selected as Highlight from RSC themed issues on ionic liquids) reports also for the first time hydration modification (caused by induction, via the presence of an additive) of protein charged compartments, with this mechanism being partially responsible for the success of crystallization, in the same way as previously unraveled for inorganic ionic crystals. The general feedback mechanism between ion hydration-water structure dynamics-and crystallization (self assembly) proved to be valid from inorganic to biomacromolecular systems and opens new horizons to control order formation in complex systems.
Using the potential of ionic liquids to shape water in crystallization processes, inorganic crystals with potential functionality have been synthesized. These refer to hollow core calcite rhombohedra with composite (eventually multishell) walls [9], fluorescent vaterite microcapsules with permeable walls and composite calcium carbonate crystals with fluorescent vaterite cores encapsulated inside calcite shells (unique chemistry, mixed structure) [10]. Rhombohedral crystals with internal cavities capable of occupying more that 70 % of crystal volume have been grown encompassing ionic-liquid stabilized gas bubbles in solution. Stabilization of bubbles and bubble aggregates is determined by the effect of ions on water structure dynamics. Composite walls of those crystals could grow in gel-like environment (environment with restricted water mobility or in other words structured water) generated by ionic liquids. Loadable vaterite capsules and mixed-structure composite crystals have been grown in a system where inorganic ions (crystal building units) induce spontaneous emulsification of the water-immiscible ionic liquid and water and build a porous crystal around an emulsion droplet. Unstable (emulsifying) ionic liquid-water interfaces were used to template formation of porous films (COIL). The designed crystalline structures can find practical applications as delivery carriers for the controlled release of compounds, photonic crystals, lightweight fillers, or chemical storage devices for active species. Cavities enable loading and potential use in smart delivery systems. Compositional zoning can be especially advantageous for the affinity-based loading with different target materials and for solubility-controlled release. Fluorescence can be potentially employed for tracking the load or for imaging purposes.
It is important to note, that in all crystallization experiments the concentration of ILs in aqueous solution was kept relatively low, so that water could be considered the main solvent (according to the objectives of the project) and its local structure modification by IL additives was affecting process of crystal nucleation and grow. Although the importance of the water mediated interactions in crystalline order formation has been appreciated (though mainly with respect to protein crystals), there has been no comprehensive theory explaining the effect of other solutes on hydration of crystal building units and respective consequences for crystal nucleation and growth. In CRYSTILS project water has been always considered as a crucial player. And it was shown that character of interaction of ILs with water molecules, that defines partitioning of IL ions to the interfaces (e.g. air/water) and IL ions´ mutual affinity in aqueous solution, determines also effect of those liquid salts on hydration of other solutes (here crystal building units) and therefore ability of the latter to form crystalline order in solution. Therefore outcome of this project creates some common framework connecting chemistry and molecular structure of multicomponent aqueous solution and crystallization processes. Recognition of the relationship between IL-water interactions and crystal formation opens possibility to employ those designer chemicals in crystallization strategies for potential scientific and engineering applications.
As a result of this project knowledge from the field of ILs is transferred to the field of biomacromolecular crystallization in the form of a joint project between Laboratory of Molecular Thermodynamics at ITQB (the Host laboratory of the ERG grant beneficiary) and Laboratory of X-ray Crystallography of Proteins at Faculdade de Ciencias e Tecnologia. The Marie Curie ERG beneficiary is a Principal Investigator in this project.