Penicillin acylase catalysis in the presence of ionic liquids

Several ionic liquids were used as reaction media for penicillin G acylase catalysis. In all the assayed ionic liquids, [bmim]PF6 proved good media for PGA-catalyzed hydrolysis. A novel [bmim]PF6/water two-phase system is provided for 6-aminopenicillanic acid (APA) production, which will be more benefical than aquous batch systems used widely in industrial production of APA.     

Sequence-specific destabilization of azurin by tetramethylguanidinium-dipeptide ionic liquids

The thermal unfolding of the copper redox protein azurin was studied in the presence of four different dipeptide-based ionic liquids (ILs) utilizing tetramethylguanidinium as the cation. The four dipeptides have different sequences including the amino acids Ser and Asp: TMG-AspAsp, TMG-SerSer, TMG-SerAsp, and TMG-AspSer. Thermal unfolding curves generated from temperature-dependent fluorescence spectroscopy experiments showed that TMG-AspAsp and TMG-SerSer have minor destabilizing effects on the protein while TMG-AspSer and TMG-SerAsp strongly destabilize azurin. Red-shifted fluorescence signatures in the 25 °C correlate with the observed protein destabilization in the solutions with TMG-AspSer and TMG-SerAsp. These signals could correspond to interactions between the Asp residue in the dipeptide and the azurin Trp residue in the unfolded state. These results, supported by appropriate control experiments, suggest that dipeptide sequence-specific interactions lead to selective protein destabilization and motivate further studies of TMG-dipeptide ILs.     

A comparative study of myoglobin stability in the presence of Hofmeister anions of ionic liquids and ionic salts

To reveal the impact of ionic liquids (ILs) on the stability of proteins, a series of ILs possessing same 1-butyl-3-methylimidazolium cation [Bmim]⁺ with a set of Hofmeister anions such as SCN⁻, HSO₄⁻, Cl⁻, Br⁻, CH₃COO⁻ and I⁻ were used and their effects on the myoglobin (Mb) structure and stability were studied. For the sake of comparison and also to explore the extent of the stabilization behavior of ILs toward Mb stability, we have chosen a set of ionic salts (I_s) of a fixed sodium cation (Na⁺) with the same series of anions such as SCN⁻, SO₄⁻², Cl⁻, Br⁻, CH₃COO⁻ and I⁻. UV–vis, fluorescence and circular dichroism (CD) spectroscopic techniques were used in order to investigate the stability behavior of Mb in ionic species (I_s and ILs). The results reveal that both I_s and ILs had a negative influence on the stability of Mb. Apparently, the flexibility in the native structure of Mb gradually increases with the increase in the concentration of I_s and ILs at pH 7.0. Therefore, a sharp decrease in the transition temperature (T_m) of the native Mb is observed in the presence of I_s and ILs.     

Horseradish peroxidase in ionic liquids: Reactions with water insoluble phenolic substrates

The reactivity of horseradish peroxidase (HRP) with water insoluble phenolic compounds has been studied in 1-butyl-3-methylimidazolium tetrafluoroborate ([BMIM][BF₄])/water mixtures. The enzyme retained some catalytic activity up to 90% ionic liquid in water at 25 °C only at pH values higher than 9.0. Activity steadily decreased towards neutral and acidic conditions, as judged by 4-aminoantypirin/phenol activity tests. Inhibition of horseradish peroxidase under neutral acidic condition was due to the binding of fluoride anions released from tetrafluoroborate anion to the heme iron as demonstrated by the sharp UV–visible absorption transition diagnostic of the conversion from a five coordinated to a six coordinated high spin ferric heme iron. Thus, reactions with water insoluble phenols were carried out under alkaline reaction conditions and 75% [BMIM][BF₄]/water mixture. Under these conditions, the distribution of the reaction products was much narrower with respect to that observed in aqueous buffers or water/dimethylformamide or water/dimethylsulfoxide mixtures, and polymeric species other than dimers were not observed. Technical scale preparations of a novel 4-phenylphenol ortho dimer [2,2′-bi-(4-phenylphenol)] with a high yield of the desired product were obtained.     

Tyrosinase activity in ionic liquids

Activity of mushroom tyrosinase was studied in three ionic liquids, 1-butyl-3-methylimidazolium hexafluorophosphate ([BMIm][PF₆]), 1-butyl-3-methylimidazolium tetrafluoroborate ([BMIm][BF₄]) and 1-butyl-3-methylimidazolium methylsulfate ([BMIm][MeSO₄]), and was compared to that in chloroform. Kinetic parameters of the enzyme were determined and the results indicate that the enzyme in ionic liquids basically follows the same catalytic mechanism as in water, and that the ionic liquids may affect the enzyme activity by direct interacting with the enzyme and thus hindering the E–S binding due to their high hydrophilicity and polarity.     

Green microwave-assisted synthesis of cellulose/calcium silicate nanocomposites in ionic liquids and recycled ionic liquids

Fabrication of biomass materials by a microwave-assisted method in ionic liquids allows the high value-added applications of biomass by combining three major green chemistry principles: using environmentally preferable solvents, using an environmentally friendly method, and making use of renewable biomass materials. Herein, we report a rapid and green microwave-assisted method for the synthesis of the cellulose/calcium silicate nanocomposites in ionic liquids and recycled ionic liquids. These calcium silicate nanoparticles or nanosheets as prepared were homogeneously dispersed in the cellulose matrix. The experimental results confirm that the ionic liquids can be used repeatedly. Of course, the slight differences were also observed using ionic liquids and recycled ionic liquids. Compared with other conventional methods, the rapid, green, and environmentally friendly microwave-assisted method in ionic liquids opens a new window to the high value-added applications of biomass.     

Structural modifications of nucleosides in ionic liquids

Nucleoside chemistry represents an important research area for drug discovery, as many nucleoside analogs are prominent drugs and have been widely applied for cancer and viral chemotherapy. However, the synthesis of modified nucleosides presents a major challenge, which is further aggravated by poor solubility of these compounds in common organic solvents. Most of the currently available methods for nucleoside modification employ toxic high boiling solvents; require long reaction time and tedious workup methods. As such, there is constant effort to develop process chemistry in alternative medium to limit the use of organic solvents that are hazardous to the environment and can be deleterious to human health. One such approach is to use ionic liquids, which are ‘designer materials’ with unique and tunable physico-chemical properties. Studies have shown that methodologies using ionic liquids are highly efficient and convenient for the synthesis of nucleoside analogs, as demonstrated by the preparation of pharmaceutically important anti-viral drugs. This article summarizes recent efforts on nucleoside modification using ionic liquids.     

Efficient hydrolysis of chitosan in ionic liquids

Effective hydrolysis of chitosan, the N-deacetylated product of chitin, remains challenging. Here, we report acid-promoted hydrolysis of chitosan in imidazolium based ionic liquids with good total reducing sugars (TRS) yield under mild conditions. TRS yield reached over 60% in the presence of about 6.0 wt% concentrated hydrochloric acid at 100 °C within 7 h. Kinetic modeling of a typical experimental data set suggested that the hydrolysis most likely followed a consecutive first-order reaction sequence, where k₁ and k₂, the rate constants for TRS formation and degradation, were determined to be 0.01372 and 0.00015 min⁻¹, respectively. Our method may be useful to explore new applications of natural chitin resources.     

Chemoenzymatic dynamic kinetic resolution of rac-1-phenylethanol in ionic liquids and ionic liquids/supercritical carbon dioxide systems

Continuous dynamic kinetic resolution processes in different ionic liquid/supercritical carbon dioxide biphasic systems were carried out by simultaneously using both immobilized Candida antarctica lipase B (Novozym 435) and silica modified with benzenosulfonic acid (SCX) catalysts at 40°C and 10 MPa. SCX was seen to act as an efficient heterogeneous chemical catalyst for the racemization of (S)-1-phenylethanol in different ionic liquid media ([emim][NTf₂], [btma][NTf₂] and [bmim][PF₆]). Coating both chemical and enzymatic catalysts with ILs greatly improved the efficiency of the process, providing a good yield (76%) of (R)-1-phenylethyl propionate product with excellent enantioselectivity (ee = 91–98%) in continuous operation.     

NMR evidence for the participation of triflated ionic liquids in glycosylation reaction mechanisms

A systematic low-temperature NMR study of a glycosylation reaction was performed in the presence of different ionic liquids and acidic catalysts. The influence of the triflate anion derived from [emim][OTf] on the stereochemistry of the glycosylation products was evaluated.     

Covalent attachment of horseradish peroxidase to the outer surface of liposomes

We describe a method by which horseradish peroxidase may be attached covalently to the surface of liposomes under conditions which permit minimal non-covalent association of the enzyme with the lipids. The coupling method adopted does not allow the formation of homopolymers of liposomes or peroxidase. For phosphatidylethanolamine/phosphatidylcholine and stearylamine/phosphatidylcholine vesicles, minimal disruption of vesicular structure is observed, whilst for phosphatidylserine vesicles, the lipid-protein complex appears to form structures much smaller than 25 nm in diameter. Stearylamine/phosphatidylcholine vesicles have been shown to retain entrapped inulin, and activity measurements for the peroxidase suggest that it is located exclusively on the external surface of the liposome membrane. Peroxidase can be localized histochemically which has permitted the morphological study of the coated liposomes and their interactions with cells.     

Lipase-catalyzed glycerolysis in ionic liquids directed towards diglyceride synthesis

This work examined the lipase-catalyzed glycerolysis of triglycerides (TG) in a list of commercially available ionic liquids (ILs) with varied cations and anions for the purpose of developing an efficient reaction protocol for diglyceride (DG) production and to understand whether ILs could assist the reaction systems. The reaction performances (reaction rate, TG conversion and DG yield) were found to be greatly dependent on the structure and property of ILs. The reactions in [TOMA·Tf₂N] and Ammoeng 120 produced comparable yield of DG to those most efficient conventional systems but with less by-products. Temperature enhancement generally yields positive effect on the conversion of TG, which was much more significant for the ILs with high viscosity. Unusually, increasing substrate concentration in many types of ILs led to enhanced conversion and yield; whereas the increase of glycerol/TG ratio resulted in a dramatic improvement of the reactions in the ILs with strong acidic anions. This work also sorted out some promising IL candidates, namely the ILs with good DG formation selectivity and the ones being able to achieve high TG conversion, which offered possibility to design binary IL systems. Overall, this study presented the first attempt concerning evaluation and characterization of lipase-catalyzed glycerolysis of TG for DG production in IL-based systems.     

Crystallisation of inorganic salts containing 18-crown-6 from ionic liquids

Reaction of the zwitterionic imidazolium salt [(CH₂COOH)(CH₂COO)im] with K₂CO₃ or BaO in the presence of 18-crown-6 affords the salts [(CH₂COO)₂im][K(18-crown-6)] and [(CH₂COO)₂im]₂[Ba(18-crown-6)], respectively. Recrystallisation of these crown complexes from the ionic liquid 1-ethyl-3-methylimidazolium bis(trifluoromethylsulfonyl)amide, [emim][Tf₂N], at a water interface, results in the formation of new salts in which the original anion is replaced by Tf₂N⁻. Single crystal X-ray diffraction has been performed on two of the salts. Notably, the potassium structure containing 18-crown-6 and Tf₂N⁻ forms a linear chain coordination polymer that can be regarded as metal organic frameworks (MOFs). Moreover, this study provides insights into the separation of group I and II metal ions using crown ethers in combination with ionic liquids.     

Scope and limitations of imidazolium-based ionic liquids as room temperature glycosylation promoters

The scope and limitations of imidazolium-based ionic liquids as room temperature glycosylation promoters have been studied. Herein, we report the effects of modifying the structure of the imidazolium cation and how important the choice of counter ion becomes on model glycosylation reactions of thioglycosides at room temperature in the presence of N-iodosuccinimide (NIS).     

Facile ring opening of 2,3-epoxy-steroids with aromatic amines in ionic liquids

Efficient ring opening of steroidal 2,3-epoxides with stoichiometric amount of aromatic amines has been carried out using an ionic liquid ([bmim](+)[BF(4)](-)) both as solvent and catalyst. The reactions were completely regio- and stereoselective in each case. The aminoalcohol products have chair conformations in ring A. The ionic liquid-mediated ring opening can efficiently be carried out with aliphatic amines like morpholine as well.     

Pine nut peroxidase immobilized on chitosan crosslinked with citrate and ionic liquid used in the construction of a biosensor

A biosensor based on the ionic liquid 1-butyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide (BMI·Tf₂N) and a novel source of peroxidase (tissue from the pine nuts of Araucaria angustifolia) was constructed. This enzyme was immobilized on chitosan crosslinked with citrate and the biosensor used for the determination of rosmarinic acid by square-wave voltammetry. The peroxidase in the presence of hydrogen peroxide catalyzes the oxidation of rosmarinic acid to quinone and the electrochemical reduction of the product was obtained at a potential of +0.15 V vs. Ag/AgCl. Different analytical parameters influencing the biosensor response, that is, peroxidase units, pH, hydrogen peroxide concentration and parameters for the square-wave voltammetry (frequency, pulse amplitude and scan increment), were investigated. The best performance was observed for the biosensor under the following conditions: 1000 units mL⁻¹ peroxidase, pH 7.0 and 8.3 × 10⁻⁴ mol L⁻¹ hydrogen peroxide with a frequency of 30 Hz, pulse amplitude of 100 mV and scan increment of 5.0 mV. The biosensor gave a linear response to rosmarinic acid over the concentration range of 9.07 × 10⁻⁷ to 4.46 × 10⁻⁶ mol L⁻¹ with a detection limit of 7.25 × 10⁻⁸ mol L⁻¹. The recovery of rosmarinic acid in plant extracts ranged from 97.0% to 109.6% and the determination of this substance in these samples using the biosensor compared favorably with that using the capillary electrophoresis method.     

Spectrophotometric assay for protease activity in ionic liquids using chromogenic substrates

A new spectrophotometric assay has been developed to evaluate protease activity in ionic liquids (ILs). The assay consists of two strategies to enable real-time spectrometric analysis of enzymatic reaction in ILs. First, enzymes are modified with a comb-shaped poly(ethylene glycol), PM₁₃, to obtain a transparent enzyme solution in IL. Second, a chromogenic substrate is used to follow the enzymatic reaction in IL. p-Nitroaniline-derivatized substrates are subjected to protease-catalyzed alcoholysis to release chromogenic p-nitroaniline that can be quantitatively detected by a UV-Vis spectrophotometer. By using this method, we can evaluate protease activity in ILs quite easily without separation of products from the reaction mixture. The availability of the novel assay system was demonstrated in a kinetic analysis of subtilisin-catalyzed reaction in the IL 1-ethyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide ([Emim][Tf₂N]) under different reaction conditions. Because two different serine proteases, subtilisin and α-chymotrypsin, substantially retained its original substrate specificity in the IL, the assay can be extended to other enzymes by using suitable chromogenic substrates.     

Surface structure of ionic liquids under an external electric field

Abstract

Surface structure and properties play an important role in many applications of ionic liquids (ILs). ILs can form unique surface structures that are very different from the bulk. In imidazolium-based ILs, for example, polar groups form ordered layer structure, while cationic alkyl chains are bundled together and point out from the surface. In many applications, ILs work under an external electric field. However, the effect of external electric field on the surface structure of ILs is still not clear. Here by using coarse-grained molecular dynamics simulation, we found that an electric field as strong as 1 V/nm is required to alter the surface structure of 1-dodecyl-3-methylimidazolium nitrate. Under a strong external electric field, layered structure disappears, and instead a sparser, more homogeneous and less orientationally ordered surface develops.     

Environmentally friendly, efficient resolution of racemic secondary alcohols by lipase-catalyzed enantioselective transesterification in ionic liquids in the presence of organic bases

The lipase-catalyzed enantioselective transesterification of racemic secondary alcohols was studied using vinyl acetate as acyl donor in two imidazolium-based ionic liquids vs. hexane (Scheme), both in the absence and presence of catalytic amounts of organic bases such as triethylamine (Et(3)N) or pyridine. The organic bases generally enhanced both the rate and enantioselectivity of the reaction. Further, the system 1-butyl-3-methyl-1H-imidazolium hexafluorophosphate/Candida antarctica lipase B ([bmim][PF(6)]/CALB) could be readily recycled four times without significant loss in activity or enantioselectivity.     

Magnetic nanoparticles supported ionic liquids for lipase immobilization: Enzyme activity in catalyzing esterification

Candida rugosa lipase was immobilized on magnetic nanoparticles supported ionic liquids having different cation chain length (C₁, C₄ and C₈) and anions (Cl⁻, BF₄⁻ and PF₆⁻). Magnetic nanoparticles supported ionic liquids were obtained by covalent bonding of ionic liquids–silane on magnetic silica nanoparticles. The particles are superparamagnetic with diameter of about 55 nm. Large amount of lipase (63.89 mg/(100 mg carrier)) was loaded on the support through ionic adsorption. Activity of the immobilized lipase was examined by the catalysis of esterification between oleic acid and butanol. The activity of bound lipase was 118.3% compared to that of the native lipase. Immobilized lipase maintained 60% of its initial activity even when the temperature was up to 80 °C. In addition, immobilized lipase retained 60% of its initial activity after 8 repeated batches reaction, while no activity was detected after 6 cycles for the free enzyme.