Interaction Between Phosphate And Guanidine In L-arginine Salt Solutions And Preparation Of Novel Crystals

L-arginine phosphate monohydrate (LAP) crystal is an excellent nonlinear optical material, and compared with other similar crystal, whose unusual high laser damage threshold has been widely concerned. Further, LAP crystal has been discovered to have low SBS threshold and much higher SBS reflection than fused silica. Recently, our group found that, LAP crystal exhibited a special reversible phase-change in the variable temperature process, and showed a long spin-lattice relaxation time at solid-state NMR. The particularity of material derives from its structure and molecular variation character. LAP crystal has shown its uniqueness when subjected to the action of energy such as light, heat and magnetic field. However, for these special phenomena, there is no reasonable explanation.Recently, Phosphate arginine (PA) having the similar chemical composition with LAP crystal was found. As an important phosphorus source in invertebrates, PA is responsible for the biological energy storage and transfer in the process of ADP and ATP transformation. Biological studies have shown that, the energy storage and transfer functions of PA, which mainly include two processes:A. the variation of phosphate-guanidine special interaction. B. the conversion of arginine conformation between extended and folded. Meanwhile, more biochemical research found that, the special electrostatic or hydrogen bonding interaction between guanidine and phosphate, which plays an important role in protein molecule interactions and their biochemistry functions. Moreover, the interaction between phosphoate and guanidine group in crystal structure had also been studied as model of biomolecular interaction.LAP crystal has the similar phosphoate and arginine as PA molecular, and the interaction between phosphoate and guanidine group of arginine plays an important role in PA and other biomolecule functions. Thus, the study of special interaction between phosphoric acid and arginine molecules in abiotic environment, not only can helps us to reveal the particularity of LAP crystal, to further explore the correlation of LAP crystal and PA molecular under energy action, but also conducive to interpret molecular biochemical function. The positions of atom and molecule in crystal structure are relatively stable, whose interaction is more difficult to be observed. While, molecule is relatively freely in solution, and there are hydrogen bonds, van der Waals forces and electrostatic attractive forces between the molecules in solution. Due to the interaction between molecules, molecules form aggregates and the properties of solution will be changed. In crystal growth solution, the solute molecule usually exists in the form of aggregate, which have a strong associated with crystal structure. Moreover, the aqueous solution environment is more similar to the biological environment. The studies on structure and properties of aggregate, which can not only investigate the intermolecular interaction related with biomolecules molecular function, but also help us further understand the crystal growth process of amino acid salts. Therefore, in this dissertation, the crystal growth solution L-arginine salts has been used as the primary study object. The special interaction between the groups in L-arginine salt would be explored by studying the structure and properties of molecular aggregates in solution. Meanwhile, a series of new amino acid salt and guanidine derivative crystals have been prepared and characterized, and the group interaction of crystal structure has been preliminary analysised. The dissertation mainly comprises the following aspects:(I) Existing form of molecule in L-arginine and LAP solutionsBy measuring solution1H-NMR spectra at room temperature, the solute structures of L-arginine and LAP solutions at different concentrations have been analyzed; Using Pseudo-Phase model, the chemical shift changes of a-H from L-arginine with the solute concentration is analysised. We have obtained the critical aggregation concentrations (0.05mol·L-1) of L-arginine molecule in L-arginine and LAP solutions, and the similar L-arginine molecule aggregation has been found in two kinds of solutions.Using steady-state fluorescence spectra at room temperature, the characteristic fluorescence properties of L-arginine and LAP saturated solution have been discovered and researched. The two major characteristics fluorescence emission wavelength of L-arginine aggregate were found at415and380nm, respectively. The structure of molecule aggregate has also been discussed through investigating emission and excitation spectra. Through analyzing the fluorescence emission properties of L-arginine and LAP solutions at different concentrations, the critical aggregation concentration was obtained again, which is consistent with the result of^-NMR analysis. The results have indicated that molecule aggregate is fluorescent chromophore.L-arginine and LAP saturated solutions with ultra-pure water as solvent, which were filtered by using0.22μm membrane. By dynamic light scattering particle size tests, the hydrodynamic diameter of molecule aggregates in two solutions have been found as the same as from0.8to1.5nm.The LAP crystal growth interface was real-time observed by in situ liquid AFM. The subsiding and promoting of growth steps were found at the same time. The two kinds of step heights have been found at1.5and3.0nm, respectively, and mostly at1.3to1.6nm. Therefore, L-arginine molecules generally exist in dimers form in solution.(II) The special intermolecular interaction in LAP solution.L-arginine and phosphate groups are present in the LAP solution, whose interaction will be the key to study on the relationship between LAP and biomolecular function. The special fluorescent emission at380nm of LAP solution has been found in solution fluorescence spectral experiments, relatived to the emission of L-arginine solution at415nm, which has an obvious blue shift. Since, phosphoric acid has no fluorescent structure in LAP solution, the L-arginine ion states or molecule interactions should be the reason for the changes in the fluorescence emission of molecule aggregates.According to the distribution of L-arginine ionic state at different pH values, the same L-arginine ionic form has been found in LAP and other L-arginine salt solutions. With the experimental conditions of the same fluorescence spectra, the fluorescence emissions of L-arginine and several L-arginine salt solutions have been found to be same. The effect of L-arginine ionic form on the fluorescence properties of solution has been excluded. Therefore, the specific fluorescence emission of LAP solution should been related to the interaction between L-arginine and phosphoate.L-lysine molecule has the same α-amino group, carboxyl group and carbon chain as L-arginine. Several L-lysine salts were synthesisd and their solutions were prepared. By comparing the fluorescence emission properties of L-lysine and its salts solutions, there no interaction was found in L-lysine phosphate solution which affected the fluorescence properties. The results have indicated that there is no specific interaction between phosphoate and carboxyl or α-amino groups. Thus, the special fluorescence emission of LAP solution should be attributed to the interaction between phosphoate and guanidino group of L-arginine.L-arginine solutions containing different concentrations of phosphoric acid were perepared, whose ultraviolet absorption and fluorescence emission spectra have been analyzed. When LAP molecule formed in solution, the characteristic fluorescence emission wavelength and the UV absorption intensity at296nm of L-arginine solutions were changed. The weak absorption at296nm is likely to come from guanidine group. Therefore, the group interaction involved by guanidine had changed the fluorescence properties of L-arginine molecule dimer in LAP solution. Thus, the specific interaction between phosphoate and guanidine exists in LAP molecule.By measuring and analyzing1H-NMR spectra of LAP solution at varying temperatures, the proton chemical shifts of L-arginine were found to shift to high field with the temperature rises, and the both ends of molecule existed different degrees of change. It shown that, when the temperature rises, both ends of L-arginine tend to bend with varying degrees in LAP solution.The changes of group vibrations in LAP solution at variable temperatures has been investigated by the in situ infrared spectroscopy. When the temperature rises, C=N stretching vibration became more obvious, shown that the charge distribution of guandine group had been changed. Meanwhile, the structure of phosphate tetrahedron distorted. When the temperature decreases, the vibration of the phosphate group restored, and the group vibrations of L-arginine were no significant changes. Two-dimensional correlation analysis of infrared spectroscopy has showed that, there was a strong correlation between changes of phosphate and L-arginine group vibrations. However, there was no obvious evidence that the correlation only exists between phosphate and guanidine group.(Ⅲ) Several novel amino acid salts and guanidine derivative crystals have been prepared and characterized.In order to explore the influence of guanidine and phosphate-guanidine interaction on crystal structure and properties, we have designed and prepared L-arginine p-nitrobenzoate monohydrate (LANB), L-lysine p-nitrophenolate (LLNP) and L-lysine p-toluenesulfonate (LLTS) crystals. LANB belongs to monoclinic system, space group P21; Both LLNP and LLTS belong to orthorhombic system, space group P212121. The bulk single crystals have been obtained by the cooling method. Crystal structure, molecular spectroscopy, NMR spectra, optical and thermal properties have been investigated and analyzed. All the three amino acid salt crystals all have good transparency properties. The SHG efficiencies of LANB and LLNP are both about four times that of KDP at1064nm laser. The melting point of LLTS crystal is259°C.The different conformations of amino acids molecular (L-arginine and L-lysine) in a series amino acid salts crystals have also been analyzed by NMR. The change trend of both ends of L-lysine has been found to be consistent, while the change trend of both ends of L-arginine is different in L-arginine salt crystals. The guanidine group plays an important role in conformation varied of L-arginine molecular, and an unusual guanidine group of L-arginine has been found in LAP crystal.Meanwhile, guanidine derivative crystals, named creatinine trifluoroacetate (CTF) and phosphate bis(guanidinoacetate)(PBGA) have been designed and prepared. CTF belongs to hexagonal system, space group R3c; PBGA belongs to triclinic system, space group P1. The single crystals have been obtained by the solvent evaporation method. Crystal structure and molecular spectroscopic have been investigated and analyzed. The important hydrogen bond between phosphate and guanidine has been found in PBGA crystal. The C=N stretching vibration absorption of PBGA crystal has been found at1653cm-1, as compared with other guanidine acetate salt crystals, which has a big shift to the lower wavenumbers.In this dissertation, the special phosphate-guanidine interaction has been found to form in LAP molecular, which is the reason for fluorescence emission blue shift of L-arginine aggregates in LAP solution. Meanwhile, an abnormal migration of vibration absorption and a specific L-arginine conformation have been found in PBGA and LAP crystal, respectively. The interaction between phosphate and guanidine in the two crystal structures should be the main reasons for the phenomenon. Under abiotic environment, the discovery of interaction between phosphate and guanidine, makes LAP has more in common with PA. Therefore, this research is an important breakthrough to further explore the correlation between specificity of LAP crystal and biomolecular function. In the future work, the correlations of phosphate-guanidine interaction and molecular properties under the action of energy would be the main research content.