Modelling of Enzyme Properties in Organic Solvents

 In this article we review how molecular modeling techniques can be used to shed some light on the influence of organic solvents on the molecular characteristics of proteins and enzymes. Molecular dynamic simulations on bovine pancreas trypsin inhibitor, chymotrypsin, and subtilisin make it possible to get a deeper understanding into how increased intramolecular interactions improve conformational rigidity, thus explaining the lower reactivity and the higher thermostability of enzymes in non-aqueous media. The application of thermodynamics-based models allows first qualitative predictions on the selectivity of many reaction types; however, the application of quantum mechanical/molecular mechanical methods is required for the development of quantitative models of actual reactivity patterns.     

Modelling of Enzyme Properties in Organic Solvents

Summary.  In this article we review how molecular modeling techniques can be used to shed some light on the influence of organic solvents on the molecular characteristics of proteins and enzymes. Molecular dynamic simulations on bovine pancreas trypsin inhibitor, chymotrypsin, and subtilisin make it possible to get a deeper understanding into how increased intramolecular interactions improve conformational rigidity, thus explaining the lower reactivity and the higher thermostability of enzymes in non-aqueous media. The application of thermodynamics-based models allows first qualitative predictions on the selectivity of many reaction types; however, the application of quantum mechanical/molecular mechanical methods is required for the development of quantitative models of actual reactivity patterns. Received November 18, 1999. Accepted (revised) February 8, 2000     

Thermodynamic Study of the Solubility of Benzocaine in some Organic and Aqueous Solvents

The thermodynamic functions free energy, enthalpy, and entropy of solution, were evaluated from solubility data of benzocaine determined at several temperatures in octanol, water, and the mutually saturated solvents, in isopropyl myristate, water, and the mutually saturated solvents, and in cyclohexane. In aqueous media the solubility was determined at pH 7.4 and ionic strength 0.15 mol-L^–1. The excess free energy and the activity coefficients of the solutes also were determined. The solubility was higher in organic media, such as octanol and isopropyl myristate, than in aqueous media and cyclohexane.     

芦荟大黄素在水和非水溶剂中的电化学性质

在HAc-NaAc水溶液或二甲亚砜(DMSO)非水溶剂中,芦荟大黄素于铂电极上均显示一个准可逆的双电子转移过程,与HAc-NaAc溶液相比,芦荟大黄素在DMSO溶剂中的峰电位明显负移,且波形改变很大;HAc-NaAc溶液中,峰电流Ipa与芦荟大黄素的浓度在1.0×10-5~6.0×10-4mol/L范围内成线性关系,检测限为1.0×10-6mol/L.溶剂对芦荟大黄素的电化学性能有甚大的影响.     

有机溶剂性质及其水含量对酶催化合成含D－氨基酸残基的二肽衍生物的影响

有机溶剂性质及其水含量对酶催化合成保护的寡肽及其衍生物有显著影响。系 统地研究了有机溶剂性质及在不同有机溶剂中水含量对α－胰凝乳蛋白酶催化合成 含D－氨基酸残基的二肽衍生物产率的影响。用α－胰凝乳蛋白酶、枯草杆菌蛋白 酶和嗜热杆菌蛋白酶在有机溶剂中催化合成了一系列保护的二肽及其衍生物，并研 究了水含量对反应产率的影响，得到了十分有意义的结果。     

Temperature Dependence of Solubility for Ibuprofen in Some Organic and Aqueous Solvents

The thermodynamic functions—Gibbs energy, enthalpy, and entropy of solution—were evaluated from the solubilities of ibuprofen determined at several temperatures in the pure solvents: octanol, isopropyl myristate, chloroform, cyclohexane, and water. The organic solvent-saturated aqueous media and water-saturated organic solvents were also studied, except for cyclohexane. In aqueous media, the solubility was determined at pH = 7.4 and an ionic strength 0.15 mol-L^–1 (physiological values). The excess Gibbs energy and the activity coefficients of the solutes were also determined. The solubilities are higher in organic media such as chloroform and octanol than in aqueous media and cyclohexane.     

Properties and Synthetic Applications of Enzymes in Organic Solvents

Biotransformations already represent an effective and sometimes preferable alternative to chemical synthesis for the production of fine chemicals and optically active compounds. To further widen the versatility of the biological approach, the so-called "nonaqueous enzymology", which now represents an important area of research and biotechnological development, has emerged in the last ten years or so. This new methodology is especially suitable for the modification of precursors of pharmaceutical compounds and fine chemicals, which, in most cases, are insoluble or poorly soluble in water. Even though the idea of carrying out an enzymatic process in organic solvent was initially considered with scepticism, biocatalysis in such media is now investigated and exploited in numerous academic and industrial laboratories. One of the reasons that makes enzymatic catalysis in nonaqueous media so appealing, is the important new properties that enzymes exhibit in organic solvents. For example, they are often more stable and can catalyze reactions that are impossible or difficult in water. Furthermore, enzyme selectivity can also differ from that in water and can change, or even reverse, from one solvent to another. This phenomenon, which can be called "medium engineering", can be exploited as a valid alternative to protein engineering. The first part of this review examines the thermodynamic, kinetic, spectroscopic, and physical approaches that have been adopted to investigate the factors that affect activity, stability, structure, and selectivity of enzymes in organic solvents. These combined studies have brought the understanding of enzyme catalysis in organic solvents to a level almost comparable to that reached for biocatalysis in aqueous media. The second part surveys a number of the synthetic applications of enzymes in organic media, which span from the preparation of milligrams of specifically labeled compounds to the modification of fats on multiton scale and from the preparation of complex key intermediates for the pharmaceutical industry to the synthesis of polymers.     

Preferential Solvation in Aqueous–Organic Mixed Solvents Using Solvatochromic Indicators

Solvatochromism of the twisted intramolecular charge-transfer (TICT) fluorescence of 4-(N,N-dimethylamino)benzonitrile (DMABN) in pure water, methanol, ethanol, 1-propanol, 2-propanol, acetone, acetonitrile, and in the corresponding aqueous–organic binary mixed solvents was systematically studied and an empirical solvent polarity scale (F _B) based on the DMABN solvatochromism was defined. The F _B parameters of the explored binary mixed solvents as a function of solvent composition were analyzed by a stepwise solvent-exchange (SSE) model to clarify the preferential solvation (PS) of the probe dye in these binary mixed solvents. Solvation diagrams toward DMABN in the mixed solvents, i.e., the local solvent composition in the solvation shell of DMABN molecules was depicted as a function of bulk solvent composition to visualize the PS in these mixed solvents. For comparison, a similar PS analysis was applied to the solvatochromism of 2,6-diphenyl-4-(2,4,6-triphenylpyridinium-1- yl)phenolate (ET-30) and pyrene (Py) in these mixed solvents; the responsive PS pattern of the mixed solvents toward the specific indicator dye of DMABN, ET-30, and Py was then discussed in terms of the chemical properties of the probe dye, the properties of the mixed solvents, and the solute–solvent and solvent–solvent interactions.     

铝（Ⅲ）—桑色素—含氧有机溶剂荧光体系的研究

本文系统研究了醇、酮等7种含氧有机溶剂对铝-桑色素络合物荧光反应的溶剂化效应。研究发现,上述有机溶剂对荧光体系均有增敏作用,以乙醇的效果最佳,使络合物的荧光强度增加6倍多。据此,本文建立了测定微量铝的高灵敏度荧光分析方法,线性范围为0～1.0μg/10ml,检测限为1.6×10~(-9)g/ml。用所建立的方法对化学试剂中的微量杂质铝进行了测定,结果令人满意。并对有机溶剂化效应的机理作了探讨。     

On the High Viscosity of Aqueous Solution of Lysozyme Induced by Some Organic Solvents

A remarkably high viscosity has been induced in protein aqueous solutions by the addition of certain structurally related organic solvents. The effect has been observed for lysozyme aqueous solutions containing tetramethylurea (TMU), dimethylsulfoxide (DMSO), dimethylformamide, and hexamethylphosphortriamide. The effect has also been induced in ferrocytochrome c aqueous solutions by TMU. Critical concentrations for both the protein and organic solvent were verified for the onset of the viscosity increase. A common feature of the solvents which were able to induce the effect is a dipolar moiety (C=O, S → O and P → O) and a nonpolar region represented by the methyl groups. The resulting fluids show an extremely restricted flow and a typical non-Newtonian, pseudoplastic behavior. Use was made of¹H nuclear magnetic resonance (NMR) and Raman spectroscopy to characterize protein structural modifications and of¹³C NMR to investigate changes in relaxation times and chemical shifts in the solvent/water solutions. A systematic rheological characterization of the systems was undertaken for some of the solvents, and unusual patterns of viscous effects were identified for the solvent/water systems both with and without protein. The process was found to be at least partially reversible, as concluded from the recovered original solution rheological characteristics and the original protein¹H NMR spectrum, after eliminating the organic solvent by ultrafiltration. The whole process was characterized as consisting of two mutually independent stages. The first involves an extensive conformational transition of the polypeptide backbone, from a predominantly α-helical to increased random coiled and β-sheet structures, with the occurrence of nonorthodox protein secondary structures at regions above the solvent critical point. The second stage consists of short-lived interchain contacts leading to an entanglement of the macromolecular system as a whole. A microphase reversion in the organic solvent/water mixture, supported by¹³C NMR and rheological results, is proposed as the driving force causing the observed behavior.     

溶剂对镓Corrole光谱性质的影响

合成了一系列周边取代的Corrole(1, 2, 3, 4)及其镓配合物1-Ga(Py)、2-Ga(Py)、3-Ga(Py)、4-Ga(Py),通过核磁共振氢谱(¹H NMR), 紫外-可见光谱, 电喷雾离子质谱(ESI-MS)的方法对其进行了表征. 研究了不同溶剂对这一系列自由Corrole 及镓Corrole 的紫外-可见(UV-Vis)吸收光谱, 稳态荧光和时间分辨荧光光谱, 将获得的荧光衰减动力学曲线采用单指数拟合并进行解卷积处理获得荧光的寿命值. 非（弱)极性溶剂对镓Corrole 紫外光谱的影响服从Bayliss 方程, 且镓Corrole 非辐射能量损失hc(ν¹_A-ν_F)与F(n)呈线性相关.     

Effects of Organic Solvents on Immobilized Lipase in Pectin Microspheres

Lipase from Brevibacillus agri 52 was found stable up to 90% diethylenglycol (DEG), glycerol (GLY), and 1,2 propanediol (1,2 PRO) at 37 degrees C for 1 h and the stability was reduced only approximately 20% after 12 h incubation, but in 40% dimethylsulfoxide (DMSO), lipase activity was stable only for 1 h. Inhibition of the biocatalysts with dimethylformamide (DMF) was detected at 20% solvent concentration. In water immiscible systems, the stability of lipase in n-hexane, n-tetradecane and n-heptane resembles the water activity, but in the presence of isobutanol, 1-hexanol, and butylbutirate, the stability was significantly reduced. Lipase 52 precipitates in the presence of 50% acetone or ethanol/water mixtures, but enzymatic activity was partially recovered by adding 20% GLY, DEG, 1,2 PRO, or DMSO to the reaction mixture. Furthermore, by increasing DEG in 70% DMF/DEG mixtures, the lipase activity was protected. Encapsulation of lipase in pectin gels cross-linked with calcium ions brings three to four times more enzymatic activity in 70% water miscible organic solvents compared to aqueous systems.     

Studies on Systems of Salts and Mixed Solvents. XXXI. On the Solubility and Solvation Behaviour of Magnesium Chloride in Mixed Aqueous Organic Solvents

Continuing our investigations on the solvation of chlorides of selected main group elements in concentrated solutions of electrolytes this paper is intended to discuss the possibility of the qualitative interpretation of the solubility data by the example of regions of the 25°C solubility isotherm of 11 systems of the type MgCl₂ ? H₂O — Organic solvent. As to magnesium chloride a significant influence on the solubility can be noticed in a general analogy to aluminium(III)-chloride according to the solvent groups of the Kolthoff classification. The results obtained correlate with the results of spectroscopic investigations. Special significance is attached to solid phases so far unknown in literature.     

Molecular and Conformational Properties of Dendronized Polymethacrylate in Organic Solvent

The conformational and dynamo-optical properties of a homologous series of polymethacrylate modified by first-generation Fréchet-type dendrons in the molecular mass range of 70–8800 kDa in organic solvents were studied by viscometry, isothermal diffusion, dynamic light scattering, and flow birefringence. The Mark-Kuhn-Houwink equations for this dendronized polymer in toluene and chloroform together with hydrodynamic diameter and Kuhn segment length in toluene are defined as well as the value of intrinsic anisotropy of polarizability for the monomer unit.     

Preferential Solvation Study of Polyvinylpyrrolidone in Organic or Aqueous Mixed Solvents by GPC

Abstract

This paper deals with the use of gel permeation chromatography for the determination of the preferential solvation of polyvinylpyr-rolidone in binary solvent mixtures. It is the first time that this technique is applied to preferential solvation in both organic and aqueous systems. With this rapid and accurate method, we determined values of the preferential solvation coefficient λ₁, which are compared with those obtained by light scattering or dialysis equilibrium.     

Transference Numbers of NaCl in Aqueous Mixtures with Organic Solvents of Moderate to Low Permittivities

Electromotive forces (emf) were measured in the transference cells: AgAgCl- Nacl (m₂) in Z NaCl (m₁) in ZAgClAg and Na_xHg_1-xNaCl (m₁) in Z NaCl (m₂) in ZNa_xHg_1-x (where Na_xHg_1-x denotes a flowing Na–amalgam electrode and Z an aqueous-organic solvent mixture) at various molalities m ₂ > m₁ of NaCl in Z = (ethylene glycol + water), (acetonitrile + water) and (1,4-dioxane + water), with mass fractions of the organic components 0.8. The transference number of Na⁺ in (ethylene glycol + water) and (1,4-dioxane + water) varies little with respect to that in pure water medium, whereas in (acetonitrile + water) it increases remarkably with increasing proportion of acetonitrile so as to approach equitransference, behavior similar to that previously found in (methanol + water) and in (ethanol + water). At acetonitrile mass fraction 0.6 NaCl is sufficiently close to equitransference to emerge as a useful salt bridge, nearly at the same level as the popular aqueous KCl.     

Studies on Systems of Salts and Mixed Solvents. XXVI. Solubility and Solvation Behaviour of Aluminium(III) Chloride in Mixed Aqueous Organic Solvents

Using sections of 25°C solubility isotherms of 14 systems of the type AlCl₃? H₂O-organic solvent the investigations are intended to illustrate the possibility of qualitative interpretation of the solubility data on the basis of empirical solvent parameters. Within the group of solvents according to the Kolthoff classification a largely analogous influence on the solubility of aluminium chloride has been noticed. Similarities result from the typical influence on the solvation state of AlCl₃. The results obtained from the solubility investigations and parageneses of the solid phases correlate with the results of spectroscopic investigations. Solid phases so far not observed are reported.