【作者】 王磊；

【导师】 许东； 张光辉；

【作者基本信息】 山东大学 ， 材料学， 2014， 博士

【摘要】 L-精氨酸磷酸盐(LAP)晶体是一种性能优良的非线性光学材料,相比同类晶体,其非一般的高激光损伤阈值受到众多关注。此外,在受激布里渊散射(SBS)性能方面,LAP晶体具有远高于石英玻璃的SBS反射率以及较低的SBS阈值。最近,本课题组发现,LAP晶体在变温过程中出现了特殊的可逆相变,在质子固态核磁中表现出了超长的自旋晶格弛豫时间。物质的特殊性来源于其结构及分子变化特征,而LAP晶体在受到光、热或磁场等能量作用时表现出来的这些唯一性,目前还没有得到合理解释。前期文献调研中我们发现了与LAP晶体分子组成类似的磷酸精氨酸(PA)分子。PA作为无脊椎动物体内重要的磷源,在ADP转化为ATP的过程中,承担着生物能量存储和传递的功能。生物化学研究表明,PA分子之所以具有能量存储和传递的功能,主要在于①能量能够存储于其分子内磷酸与胍基间极强的相互作用中(即N-P键),②分子上精氨酸部分在伸展型与弯曲型构象之间的转变。同时,在更多的生物化学研究中发现,磷酸与精氨酸胍基间的特殊静电或氢键作用,在蛋白质分子间作用及其生物化学功能中扮演重要角色。并且,晶体结构中的磷酸与胍基间作用,也曾被作为生物分子间作用模型而研究。LAP晶体中存在与PA分子的类似的磷酸和精氨酸胍基基团,并且磷酸与胍基间作用在PA以及其他分子功能中具有重要意义。因此,对非生物环境中,磷酸与精氨酸分子间作用的研究,不仅有助于我们揭示LAP晶体的特殊性,进-步探讨LAP晶体能量作用过程与PA生物能存储过程的相关性,同时也有利于阐释分子的生物化学功能。晶体结构中原子、分子位置稳定,基团间作用较难观测。而在水溶液中,分子相对比较自由,溶质分子间存在各种氢键、范德华力和静电引力等,分子相互影响和作用,从而形成分子聚集或对溶液性质产生影响。并且,水溶液环境类似于生物环境。同时,溶液中的分子聚集是晶体生长的重要步骤,聚集体的结构和性质与晶体结构有很大关联。因此,分析其中的聚集体结构或性质变化,不仅能够探讨其中与生物分子功能相关的分子间作用,而且有助于理解晶体生长过程。本论文采用L-精氨酸盐晶体生长溶液为首要研究对象,分析了溶液中分子聚集及其性质,通过探讨LAP溶液性质的特殊性,发现了其中的LAP分子内特殊基团间作用。同时,开展了相关新晶体的制备和表征以及晶体结构中分子间作用的探讨。论文的主要研究内容包括：(一)L-精氨酸和LAP溶液中的分子聚集通过溶液’H-NMR,研究了室温下不同浓度L-精氨酸和LAP溶液中L-精氨酸分子的结构形式；采用Pseudo-Phase理论模型,对质子化学位移随浓度变化进行了分析,获得了L-精氨酸和LAP溶液中L-精氨酸分子相同的临界聚集浓度(0.05mol·L-1左右),表明两种溶液中存在类似的L-精氨酸分子聚集。室温下采用稳态荧光光谱研究了饱和浓度下L-精氨酸和LAP溶液的荧光光谱,发现了溶液中L-精氨酸分子聚集体的两个主要荧光发射(415和380nm)；结合溶液的发射和激发光谱,分析了溶液中分子聚集体的结构形式；通过对不同浓度下L-精氨酸和LAP溶液荧光发射的研究,发现荧光发射性质转变浓度与临界聚集浓度一致,表明溶液特征荧光发射来自于溶液中的分子聚集体。通过动态光散射粒度仪,以超纯水为溶剂,采用0.22μm注射膜过滤,对L-精氨酸和LAP饱和溶液中溶质聚集体的粒径分布进行了测试,结果表明两种溶液中粒径分布相同,都在0.8-1.5nm。通过原位的液相AFM观测室温下的LAP晶体生长界面,发现了生长台阶的推进与消退同时存在,表明生长界面层中溶质分子在晶体表面上同时有附着和脱附。实验获得实时生长时表面台阶高度为1.5和3.0nm两种,且大部分在1.3-1.6nm,该台阶高度能够反映界面层溶液中溶质分子聚集体的大小,因而,饱和溶液中聚集体粒径通常为1.5nm左右。(二)LAP溶液中的特殊分子间作用LAP溶液中存在L-精氨酸与磷酸基团,两者之间的作用是研究LAP分子与生物分子功能相关性的关键。在溶液荧光光谱实验中发现,LAP溶液中的L-精氨酸分子聚集体特征发射波长位于380nm,相对于L-精氨酸溶液415nm的特征发射,发生了明显的蓝移。磷酸基团不具有荧光结构,因此,溶液中的L-精氨酸分子解离形式和分子间作用,可能是影响其中分子聚集体荧光发射的原因。制备了几种L-精氨酸盐溶液,通过分析L-精氨酸离子形式随pH值分布情况,表明其中存在与LAP溶液中相同的L-精氨酸离子态。采用与L-精氨酸溶液荧光光谱相同实验条件,发现几种饱和L-精氨酸盐与L-精氨酸溶液的特征荧光发射波长都在415nm,表明L-精氨酸离子形式对荧光发射没有影响。因此,LAP溶液特殊的荧光发射与其中L-精氨酸和磷酸的分子间作用有关。L-赖氨酸分子含有与L-精氨酸相同的α-氨基、羧基和类似碳链结构。通过对等浓度下L-赖氨酸和几种L-赖氨酸盐溶液的荧光发射研究,发现了其特征荧光发射波长都在500nm,再次表明溶质分子的离子形式对溶液荧光性质没有影响。同时发现,L-赖氨酸磷酸与其他L-赖氨酸盐溶液的荧光发射相同,其发射波长也在500nm,表明溶液中L-赖氨酸与磷酸之间不存在影响荧光发射的分子间作用,即磷酸与羧基或α-氨基间不存在特殊作用。因此,LAP溶液的特殊荧光发射只能归功于磷酸与L-精氨酸上胍基之间的作用。通过磷酸滴定同一浓度L-精氨酸溶液的荧光及紫外吸收光谱实验,发现溶液中磷酸与L-精氨酸等摩尔量时,L-精氨酸溶液的荧光发射波长和296nm的紫外吸收强度同时发生突变。由于296nm弱的吸收产生于胍基,因而表明,溶液中LAP分子形成时,胍基参与的分子内基团间作用改变了L-精氨酸分子聚集体的特征荧光发射。因此,溶液中的LAP分子内存在特殊的磷酸与胍基间作用。通过对LAP溶液升温时的1H-NMR谱研究,发现溶液中L-精氨酸分子质子化学位移随温度升高而向高场偏移,且分子两端质子的变化程度不同。表明当温度升高时,LAP溶液中的L-精氨酸分子两端不同程度的弯曲趋向。采用原位红外光谱对饱和LAP溶液在变温下的分子基团振动变化进行了研究,发现当温度升高时,C=N伸缩振动开始表现明显,表明胍基基团上的电荷分布出现变化,同时发现磷酸四面体结构发生了畸变；降温时,磷酸基团的振动发生了恢复变化,而L-精氨酸分子基团振动变化不大。通过二维相关分析方法研究,发现在变温时,LAP溶液中磷酸基团振动变化与L-精氨酸分子基团振动变化有很强的相关性,但没有明显证据表明基团间的相关性仅在胍基上。(三)新的氨基酸盐和胍基衍生物晶体的制备和研究为了进一步研究胍基以及磷酸-胍基间作用对晶体中分子结构及性质的影响,设计、合成并生长了L-精氨酸对硝基苯甲酸(LANB)、L-赖氨酸对硝基苯酚(LLNP)以及L-赖氨酸对甲苯磺酸(LLTS)新晶体。其中LANB晶体属于单斜晶系,P21空间群；LLNP和LLTS晶体属于正交晶系,P212121空间群。采用降温法获得了体块单晶。通过对晶体的分子振动光谱、核磁共振谱、光学和热学性能等研究,三种晶体均具有较好的光透过性能,并且其中LANB和LLNP晶体在1064nm下的粉末二阶倍频强度约为KDP晶体的4倍,LLTS晶体具有高达259℃的熔点。采用NMR技术,分析了系列氨基酸盐晶体中氨基酸分子(L-精氨酸和L-赖氨酸)的构象变化规律,发现晶体中L-赖氨酸分子两端变化趋势一致,而在L-精氨酸盐晶体中,胍基使L-精氨酸分子构象两端变化不同,并发现LAP晶体中的胍基端更加特殊的L-精氨酸分子构象。同时,制备了胍基衍生物,肌酸酐三氟乙酸(CTF)和磷酸双乙酸胍(PBGA)新晶体,其中CTF晶体属于六方晶系,R3c空间群；PBGA晶体属于三斜晶系,P1空间群,并对晶体结构中的基团间氢键作用和分子振动光谱进行了研究。在PBGA晶体结构研究中发现,其中的磷酸-胍基氢键是晶体结构的重要链接。晶体的分子振动光谱发现,乙酸胍的C=N振动吸收位于1653cm-1,相较其他乙酸胍盐晶体,其向低波数发生了较大的偏移。本论文研究发现,LAP分子内形成的特殊磷酸-胍基作用,是导致LAP溶液中L-精氨酸分子聚集体荧光发射波长蓝移的原因。同时,在LAP和PBGA晶体中,分别发现了L-精氨酸分子胍基端构象的特异性和乙酸胍分子上胍基振动吸收的异常偏移,其结构中的磷酸与胍基间作用可能是导致该现象的主要原因。非生物环境下,磷酸与胍基相互作用的发现,使LAP分子与PA及其他生物分子具有了更多的共性,是进一步探讨LAP晶体特异性与分子能量存储功能相关性的重要突破。对于能量作用下,磷酸-胍基作用与分子性质变化的相关性研究将是下一步工作的主要内容。更多还原

【Abstract】 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.更多还原