Genetically engineered peptide fusions for improved protein partitioning in aqueous two-phase systems: Effect of fusion localization on endoglucanase I of Trichoderma reesei

Genetic engineering has been used for fusion of the peptide tag, Trp–Pro–Trp–Pro, on a protein to study the effect on partitioning in aqueous two-phase systems. As target protein for the fusions the cellulase, endoglucanase I (endo-1,4-β- -glucan-4-glucanohydrolase, EC 3.2.1.4, EGI, Cel7B) of Trichoderma reesei was used. For the first time a glycosylated two-domain enzyme has been utilized for addition of peptide tags to change partitioning in aqueous two-phase systems. The aim was to find an optimal fusion localization for EGI. The peptide was (1) attached to the C-terminus end of the cellulose binding domain (CBD), (2) inserted in the glycosylated linker region, (3) added after a truncated form of EGI lacking the CBD and a small part of the linker. The different constructs were expressed in the filamentous fungus T. reesei under the gpdA promoter from Aspergillus nidulans. The expression levels were between 60 and 100 mg/l. The partitioning behavior of the fusion proteins was studied in an aqueous two-phase model system composed of the thermoseparating ethylene oxide (EO)–propylene oxide (PO) random copolymer EO–PO (50:50) (EO₅₀PO₅₀) and dextran. The Trp–Pro–Trp–Pro tag was found to direct the fusion protein to the top EO₅₀PO₅₀ phase. The partition coefficient of a fusion protein can be predicted with an empirical correlation based on independent contributions from partitioning of unmodified protein and peptide tag in this model system. The fusion position at the end of the CBD, with the spacer Pro–Gly, was shown to be optimal with respect to partitioning and tag efficiency factor (TEF) was 0.87, where a fully exposed tag would have a TEF of 1.0. Hence, this position can further be utilized for fusion with longer tags. For the other constructs the TEF was only 0.43 and 0.10, for the tag fused to the truncated EGI and in the linker region of the full length EGI, respectively.     

Solvent properties governing protein partitioning in polymer/polymer aqueous two-phase systems

Distribution coefficients of various proteins were measured in aqueous Dextran-Ficoll, Dextran-PES, and Ficoll-PES two-phase systems, containing 0.15M NaCl in 0.01 M phosphate buffer, pH 7.4. The acquired data were combined with data for the same proteins in different systems reported previously and known solvatochromic solvent properties of the systems to characterize the protein-solvent interactions. The relative susceptibilities of proteins to solvent dipolarity/polarizability, solvent hydrogen bond acidity, solvent hydrogen bond basicity, and solvent ability to participate in ion-ion and ion-dipole interactions were characterized. These parameters, which are representative of solute-solvent interactions, adequately described the partitioning of the proteins in each system. It was found that the relative susceptibilities of proteins to solvent dipolarity/polarizability are interrelated with their relative susceptibilities to solvent hydrogen bond acidity and solvent hydrogen bond basicity similarly to those established previously for small nonionic organic compounds.     

Extraction of endoglucanase I (Ce17B) fusion proteins from Trichoderma reesei culture filtrate in a poly(ethylene glycol)-phosphate aqueous two-phase system

Endoglucanases (EGI) (endo-1,4-beta-D-glucan-4-glucanohydrolase, EC 3.2.1.4, Ce17B) of Trichoderma reesei are industrially important enzymes. Thus, there is a great need for development of a primary recovery method suitable for large-scale utilization. In this study we present a concept applicable for large-scale purification of an EGI fusion protein by one-step extraction in a poly(ethylene glycol) PEG-sodium/potassium phosphate aqueous two-phase system. EGI is a two-module enzyme composed of an N-terminal catalytic module and a C-terminal cellulose binding module (CBM) separated by a glycosylated linker region. Partitioning of six different EGI constructs, containing the C-terminal extensions (WP)2, (WP)4 or the amphiphilic protein hydrophobin I (HFB) of T. reesei instead of the CBM were studied to evaluate if any of the fusions could improve the partition coefficient sufficiently to be suitable for large-scale production. All constructs showed improved partitioning in comparison to full length EGI. The (WP)4 extensions resulted in 26- to 60-fold improvement of partition coefficient. Consequently, a relative minor change in amino acid sequence on the two-module protein EGI improved the partition coefficient significantly in the PEG 4000-sodium/potassium phosphate system. The addition of HFBI to EGI clearly enhanced the partition coefficient (K=1.2) in comparison to full-length EGI (K=0.035). Partitioning of the construct with (WP)4 fused to the catalytic module and a short sequence of the linker [EGI(core-P5)(WP)4] resulted in the highest partition coefficient (K=54) and a yield of 98% in the PEG phase. Gel electrophoresis showed that the construct with the (WP)4 tag attached after a penta-proline linker could be purified from the other bulk proteins by only a single-step separation in the PEG 4000-sodium/potassium phosphate system. This is a major improvement in comparison with the previously studied model (ethylene oxide-propylene oxide)-dextran system. Hence, this construct will be suitable for further optimization of the extraction of the enzyme in a PEG 4000-sodium/potassium phosphate system from culture filtrate.     

Cutinase-peptide fusions in thermoseparating aqueous two-phase systems. Prediction of partitioning and enhanced tag efficiency by detergent addition

It is of increasing importance to develop efficient purification methods for recombinant proteins where the number of steps can be minimised. The aim has been to establish a method for predicting the partitioning of the wild-type target protein in an aqueous two-phase system, and with this as basis, develop fusion tags and optimise the phase system for enhanced partitioning of the target protein. The surface of the lipolytic enzyme cutinase from Fusarium solani pisi was investigated with a computer program, Graphical Representation and Analysis of Surface Properties (GRASP). The accessible surface areas for the different amino acid residues were used together with peptide partitioning data to calculate the partition coefficient for the protein. The separation system was composed of a thermoseparating random copolymer of ethylene oxide and propylene oxide. Breox PAG 50A 1000, as top phase forming polymer and a hydroxypropyl starch polymer, Reppal PES 200, as bottom phase polymer. The calculated partition coefficient for the wild-type protein (K= 1.0) agreed reasonably well with the experimentally determined value (K=0.85). Genetic engineering was used to construct fusion proteins expressed in Saccharomyces cerevisiae based on cutinase and peptide tags containing tryptophan, to enhance the partitioning in aqueous two-phase systems. The partitioning of the cutinase constructs could qualitatively be predicted from peptide partitioning data, i.e. the trends in partitioning could be predicted. A spacer peptide introduced between protein and tag increased the partitioning of the protein towards the ethylene oxide-propylene oxide (EOPO) copolymer top phase. The aqueous two-phase system was modified by addition of detergent to increase the partitioning of the cutinase variants towards the EOPO copolymer phase. Triton and a series of C12En detergents selectively increased the partitioning of cutinase constructs with (WP)4-based tags up to 14 times compared to wild-type cutinase. The protein partition could almost quantitatively be predicted from the peptide partition data.     

Membrane protein isolation by in situ solubilization, partitioning and affinity adsorption in aqueous two-phase systems. Purification of the human type 1 11beta-hydroxysteroid dehydrogenase

Recently developed aqueous two-phase systems based on non-ionic detergents and polymers are suitable for the separation of membrane proteins. Moreover, within this relatively membrane protein "friendly" environment, changes in temperature can be controlled and stabilizing agents may be added to ensure integrity of the target protein during isolation. Here, we use aqueous two-phase partitioning for the isolation of membrane bound 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1). Different detergents were used to find optimal conditions regarding solubilization and retaining target protein activity. We explored in situ solubilization by adding detergent directly to the aqueous two-phase system, as well as a batch metal affinity capture step of 6xHis tagged 11beta-HSD1 in the two-phase system. The use of detergent/polymer two-phase systems resulted in a specific enzyme activity of 3840 nmol mg(-1) min(-1) of the target membrane protein compared to a conventional purification protocol where a specific enzyme activity of 1440 nmol mg(-1) min(-1) was achieved.     

Affinity partitioning of proteins in aqueous two-phase systems containing polymer-bound fatty acids. I. Effect of polyethylene glycol palmitate on the partition of human serum albumin and alpha-lactalbumin

The partition of serum albumin between the two aqueous phases of a polyethylene glycol-dextran-water two-phase system is strongly influenced by replacing a small fraction of the polyethylene glycol by its palmitate ester. The partition of albumin is compared with the partition of polymer-bound palmitate using polyethylene glycol [1-14C]palmitate. The data indicate that a maximum ("saturation") effect on the partition of albumin corresponds to a binding of fewer than two palmitate groups per albumin molecule. The effect on alpha-lactalbumin corresponds to the maximum binding of 0.5 palmitate group per protein molecule. These data do not fit the most favoured available model for affinity partitioning. The deviation can be attributed to the observation that the palmitate groups probably form micelle-type aggregates in the polyethylene glycol-rich phase at the concentrations used for albumin extraction.     

Electrostatically self-assembled azides on zinc sulfide nanoparticles as multifunctional nanoprobes for peptide and protein analysis in MALDI-TOF MS

A simple method to synthesize electrostatically self-assembled azides on zinc sulfide nanoparticles (ZnS-N₃ NPs) was described and then it was further applied as a multifunctional nanoprobe such as enriching, desalting, accelerating and separation-/washing free nanoprobes for rapid analysis of peptides and proteins and microwave assisted tryptic digested proteins in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The ZnS-N₃ NPs were characterized by UV-vis, FT-IR, SEM and TEM spectroscopy. The ZnS-N₃ NPs can effectively enrich signal intensities for 2-10 times for various peptides and proteins including HW6, insulin, ubiquitin, cytochrome c, lysozyme, myoglobin and bovine serum albumin (BSA) in MALDI-TOF MS. Furthermore, we also demonstrated that the ZnS-N₃ NPs can serve as accelerating probes for microwave assisted tryptic digestion of proteins in MALDI-TOF MS. The applicability of the present method on complex sample analysis such as milk proteins from cow milk and ubiquitin and ubiquitin like proteins from oyster mushroom were also demonstrated.     

Design of functional guanidinium ionic liquid aqueous two-phase systems for the efficient purification of protein

A series of novel cationic functional hexaalkylguanidinium ionic liquids and anionic functional tetraalkylguanidinium ionic liquids have been devised and synthesized based on 1,1,3,3-tetramethylguanidine. The structures of the ionic liquids (ILs) were confirmed by ¹H nuclear magnetic resonance (¹H NMR) and 13C nuclear magnetic resonance (13C NMR) and the production yields were all above 90%. Functional guanidinium ionic liquid aqueous two-phase systems (FGIL-ATPSs) have been first designed with these functional guanidinium ILs and phosphate solution for the purification of protein. After phase separation, proteins had transferred into the IL-rich phase and the concentrations of proteins were determined by measuring the absorbance at 278 nm using an ultra violet visible (UV–vis) spectrophotometer. The advantages of FGIL-ATPSs were compared with ordinary ionic liquid aqueous two-phase systems (IL-ATPSs). The proposed FGIL-ATPS has been applied to purify lysozyme, trypsin, ovalbumin and bovine serum albumin. Single factor experiments were used to research the effects of the process, such as the amount of ionic liquid (IL), the concentration of salt solution, temperature and the amount of protein. The purification efficiency reaches to 97.05%. The secondary structure of protein during the experimental process was observed upon investigation using UV–vis spectrophotometer, Fourier-transform infrared spectroscopy (FT-IR) and circular dichroism spectrum (CD spectrum). The precision, stability and repeatability of the process were investigated. The mechanisms of purification were researched by dynamic light scattering (DLS), determination of the conductivity and transmission electron microscopy (TEM). It was suggested that aggregation and embrace phenomenon play a significant role in the purification of proteins. All the results show that FGIL-ATPSs have huge potential to offer new possibility in the purification of proteins.     

Effect of nucleic acid contamination on partitioning of proteins in two-phase PEG-Dextran system

Downstream processing of bioproducts results in considerable losses of compounds of interest in a large number of cases. For the intracellular enzyme tartrate dehydrogenase, an analysis of the laboratory process for enzyme recovery revealed that maximum losses occur in the initial stages of purification when the enzyme is separated from nucleic acids and other undesirable enzymes. Hence, aqueous twophase extraction was studied to investigate the separation of several enzymes from nucleic acids. Single-component and binary equilibria for three commercially available enzymes (bovine serum albumin, trypsin, chymotrypsin) and yeast RNA were studied in a two-phase system consisting of dextran and polyethylene glycol (PEG). The effects of pH and concentrations of the components and salts (NaC1) were investigated.     

On the influence of some strong electrolytes on the partitioning of acetic acid to aqueous/organic two-phase systems in the presence of tri-n-octylamine: Part I: Methyl isobutyl ketone as organic solvent

The recovery of carboxylic acids from aqueous phases is often achieved by reactive extraction with water-insoluble amines which are dissolved in an organic solvent. The basic design of such downstream processes requires a thermodynamic framework for the encountered liquid–liquid equilibrium. The thermodynamic framework should be able to describe the rather uncommon and surprising effects that comparatively small amounts of strong electrolytes might have. Such strong electrolytes can either reduce or increase the affinity of a carboxylic acid for the organic phase in particular at low aqueous phase concentrations of the carboxylic acids. That behavior was explained in previous investigations with citric acid as a model compound for a carboxylic acid and modeled by combining the dissociation/protonation equilibrium in the aqueous phase with the formation of organic phase complexes of (amine + acid(s) + water). In the present investigation this work is extended to acetic acid as another example for a carboxylic acid. New experimental results are reported for the influence of sodium chloride, sodium nitrate, sodium sulfate, sodium citrate and hydrochloric acid on the partitioning of acetic acid to coexisting aqueous/organic liquid phases of the system (water + methyl isobutyl ketone (organic solvent) + tri-n-octylamine (chemical extractant)) at 25 °C. The phase behavior is described by an extension of the previously published thermodynamic framework that is able to describe/predict the influence of a strong electrolyte on the partitioning of acetic acid.     

Hg (II) extraction in a PEG-based aqueous two-phase system in the presence of halide ions. I. Liquid phase analysis

The partition behaviour of Hg (II) was studied in an aqueous polyethylene glycol (PEG) — (NH₄)₂SO₄ two-phase system as a function of halide, halide concentration, and pH. For a system prepared by mixing equal volumes of 40 % (w/w) PEG (1550) with 40 % (w/w) (NH₄)₂SO₄, Hg(II) remains almost exclusively in the salt-rich phase. The addition of NaX (X = Cl⁻, Br⁻, I⁻) enhances Hg (II) partition into the PEG-rich phase due to the formation of halide complexes. The efficiency of halide extractants increases in the order: Cl⁻ < Br⁻ < I⁻. Mercury extraction is improved at lower halide ion concentration by higher stock salt solution acidity. From the distribution coefficients determined as a function of halide ion concentration, the extracted species were identified. The Hg (II) extractability is determined by the type and stability of the Hg (II) halide species, and depends on the stock salt solution acidity. The observed behaviour is discussed and a possible extraction mechanism is proposed.     

Nitroprusside-PEO enthalpic interaction as a driving force for partitioning of the [Fe(CN)(5)NO](2-) anion in aqueous two-phase systems formed by poly(ethylene oxide) and sulfate salts

Ions are known to concentrate in the salt-enriched phase of aqueous two-phase systems, with the only known exception being the pertechnetate anion, TcO(4)(-). We have discovered a second ion, nitroprusside anion (NP), which is markedly transferred from the salt phase to the polymer phase. The partitioning behavior of [Fe(CN)(5)NO](2-) anion was investigated in ATPS formed by poly(ethylene oxide) of molar mass 3350 and 35000 g mol(-1), and different sulfate salts (Na(2)SO(4), Li(2)SO(4), and MgSO(4)). On the basis of a model, the nitroprusside high affinity for the macromolecular phase was attributed to an enthalpic specific interaction between the anion and ethylene oxide unit. Partition coefficients increased exponentially with tie-line length increase, reaching values as high as 1000 and showing a relationship very dependent on the salt nature, but independent of the polymer molar mass.     

On the influence of some strong electrolytes on the partitioning of acetic acid to aqueous/organic two-phase systems in the presence of tri-n-octylamine: Part II: Toluene as organic solvent

Water-insoluble amines (dissolved in an organic solvent/organic solvent mixture) are often used for the extractive recovery of carboxylic acids from aqueous phases. The basic design of the extraction process requires a thermodynamic framework that should be able to describe the liquid–liquid phase equilibrium not only in the phase forming systems (water + carboxylic acid + organic solvent + reactive extractant), but also when the aqueous feed phase contains additionally small amounts of strong electrolytes. Even small amounts of strong electrolytes might considerably reduce the recovery rate. In part I of this series such a model was presented and discussed for methyl isobutyl ketone as organic solvent and tri-n-octylamine (TnOA) as the chemical extractant. The present part II is to demonstrate that the procedures/methods described for methyl isobutyl ketone as organic solvent can be applied also for other organic solvents. By way of example, here toluene is that organic solvent. New experimental results are reported for the influence of sodium chloride, sodium nitrate, sodium sulfate, sodium acetate and hydrochloric acid on the partitioning of acetic acid to coexisting aqueous/organic liquid phases of the system (water + toluene + tri-n-octylamine) at 25 °C. An extension/adaptation of the previously published thermodynamic framework is successfully applied to describe/predict the new experimental liquid–liquid phase equilibrium data.     

Structure‐enantioselectivity relationships for the study of chiral recognition in peptide enantiomer separation on cinchona alkaloid‐based chiral stationary phases by HPLC: Influence of the N‐terminal protecting group

Eleven different N‐terminal protecting groups (acetyl, benzoyl, FMOC, etc.) were employed for the HPLC separation of oligoalanine peptide enantiomers containing up to six amino acids. Isocratic HPLC separations were performed using a hydro‐organic buffered mobile phase and 4 mm ID columns containing three different chiral anion exchange stationary phases based on cinchona alkaloid‐derived chiral selectors. For most peptides successful separations could be obtained with all protecting groups, although those comprising aromatic moieties were found to yield higher enantioselectivities than those with aliphatic residues, since they are capable of undergoing favourable π‐π interactions with the selector. Systematic investigations concerning the presence or absence of structural features of related protecting groups showed that the use of protecting groups that are optimally adjusted to the binding pocket of the chiral selector effects a significant gain in enantioselectivity. At the same time these studies provided new insights into the chiral recognition mechanism.     

Effect of sucrose on molecular and interaction parameters of sodium caseinate in aqueous solution: relationship to protein gelation

The effect of sucrose on molecular and interaction parameters of sodium caseinate in aqueous medium has been investigated using static and dynamic multi-angle laser light scattering over a wide range of sucrose concentration (from 10 to 78 w/v%) and pH values (from 7.0 to 3.5). Measurements have been made of the molar mass, the radius of gyration, the hydrodynamic radius, and the second virial coefficient of sodium caseinate in aqueous solution. Pronounced dissociation of sodium caseinate sub-micelles¹ was found in the presence of sucrose at a pH above the protein's isoelectric point. The effect of sucrose at a pH near the isoelectric point is very different. This is reflected in the pronounced increase in molar mass, radius of gyration, and the difference between the radius of gyration and the hydrodynamic radius. It was found that the extent of the protein association, caused by the presence of sucrose, is a key factor contributing to the hydrophobic–hydrophilic balance of the protein surface, and hence to the thermodynamic affinity of the caseinate sub-micelles for the aqueous medium and for each other. Analysis of light-scattering data using structure-sensitive plots shows a clear transition from Gaussian to wormlike chain/rod behaviour for sodium caseinate on pH lowering. Apparent relationships between the effects of sucrose on the self-association of sodium caseinate and a marked enhancement of the viscoelasticity of acid-induced casein gels have been revealed. Moreover, the dissociation of sodium caseinate sub-micelles is in excellent agreement with the more homogeneous microstructure of acid-induced protein gels in the presence of sucrose as detected by confocal laser scanning microscopy. We discuss likely molecular mechanisms underlying the observed effects of sucrose on the interactions and rheology in acidified caseinate systems.     

Amino acids in aqueous solution. Effect of molecular structure and temperature on thermodynamics of dissolution

The enthalpies of dissolution of glycine (Gly) and L-α-alanine (Ala) in water at 288.15–318.15 K were measured. The results were compared with the earlier obtained data for L-α-phenylalanine (Phe) and L-α-histidine (His). The standard enthalpies of dissolution (Δ_soln H ₀) and differences (ΔC _p ⁰ ) between the limiting partial molar heat capacity of the amino acids in solution and the heat capacity of the amino acids in the crystalline state were calculated in the temperature interval 273–373 K. Changes in the entropy of dissolution (ΔΔ_soln S ₀) and reduced Gibbs energy [Δ (Δ_soln G ₀/T)] in the temperature interval from 273 to 373 K were determined from the known thermodynamic relationships. The ΔC _p ⁰ value is negative for hydrophilic glycine and positive for other amino acids. The ΔΔ_soln S ⁰ values increase with an increase in the hydrophobicity of the amino acids. The Δ(Δ_soln G ₀/T) values become more negative in the order Ala, Phe, Gly, His. Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 4, pp. 711–714, April, 2007.     

An improved and clean procedure for the synthesis of one-donor poly-acceptors systems containing 2,6-dicyanoamine moiety in aqueous media catalyzed by TEBAC in the presence and absence of K2CO3

A clean and simple synthesis of one-donor poly-acceptors systems containing 2,6-dicyanoamine moiety was accomplished via the reaction of 1-arylethylidenemalonodinitriles with arylidenemalonodinitriles in aqueous media catalyzed by TEBAC in the presence of K₂CO₃. The important intermediates were obtained successfully to confirm the mechanism in the absence of base under the same reaction conditions. The structures of 3m and 4b were confirmed by X-ray diffraction studies.     

Effect of small molecule surfactants on molecular parameters and thermodynamic properties of legumin in a bulk and at the air–water interface depending on a protein structure in an aqueous medium

This paper presents the effect of fatty acid salts, namely, Na-caprate and Na-palmitate on the legumin (11S globulin of Vicia Faba broad beans) molecular and thermodynamic properties in the bulk aqueous medium and at the air–water interface under different molecular states of the protein. That are the native state of the protein globule (pH 7.2, ionic strength of 0.05 mol dm⁻³), as well as the acidic denatured (pH 3.0, ionic strength of 0.01 mol dm⁻³) and the heat denatured ones (after heating at 90°C for 30 min, pH 7.2, ionic strength of 0.05 mol dm⁻³). In turn, an importance of the state of the small molecule surfactants in a solution in reference to the critical concentrations of micelle formation (CMC), for their effect on the protein properties, was also under our studying. The peculiarities of the legumin structure in the aqueous medium appeared in the different nature of the interactions between the protein and the fatty acid salts, as was indicated by the mixing calorimetry data. So, the hydrophobic contacts provided a basis for interactions between both the native and heat denatured legumin with the fatty acid salts. At the same time, the electrostatic interactions between the oppositely charged functional groups of the fatty acid salts and the acidic denatured protein formed principally a basis of their interactions in an aqueous medium. In response to interactions of the fatty acid salts with legumin the essential changes in the protein conformational stability, depending on both the protein molecular state and concentration of the fatty acid salts, were found using differential scanning calorimetry (DSC). The rather high level of the protein association was observed by light scattering in the bulk aqueous medium in the presence of the fatty acid salts. As this takes place, the surface hydrophilicity of the protein increased under the formation of the associates. The combined data of mixing calorimetry, differential scanning calorimetry and light scattering suggested the complex formation between legumin and the fatty acid salts. The interactions of the fatty acid salts with the protein produced a change in the surface activity for the mixture of the protein with the fatty acid salts. That is a decrease in the protein surface tension at the air–water interface for the mixed solutions in comparison with ones for both the protein and small molecule surfactant alone in the case of Na-caprate, and those are the intermediate values of the surface tension in the case of Na-palmitate. These results were observed independently of the protein state (native or acidic/heat denatured) in an aqueous medium. As this took place, the most dramatic increase in the surface activity was found for the mixtures of the acidic denatured protein with Na-caprate as if the most hydrophobic species were formed in this case. The combined data of mixing calorimetry, DSC, light scattering and tensiometry showed that the effect of the fatty acid salts on the legumin thermodynamic properties in a bulk and at interfaces is governed by a number of the key factors such as: a structure of both the protein and fatty acid salt (a length of the hydrocarbon chain); a degree of the protein association in the bulk aqueous phase (as a result of the interactions with the small molecule surfactants); a change in the protein conformational stability (flexibility) under the influence of the small molecule surfactants; as well as by the nature (hydrophobic, electrostatic) of the protein–small molecule surfactant interactions, determining ultimately the hydrophilic–lipophilic balance of the protein surface.     

Aggregation behavior and catalytic activity of systems based on calix[4]resorcinarene derivatives and surfactants. 1. Mixed micellization of aminomethylated calix[4]resorcinarenes with cetyltrimethylammonium bromide in aqueous dimethylformamide

Mixed micellization of amphiphilic aminomethylated calix[4]resorcinarenes and phenols, which are their structural units, with the cationic surfactant cetyltrimethylammonium bromide (CTAB) in aqueous 10—70 vol % DMF decreases the critical micelle concentration; the resulting aggregates are larger than those in the CTAB—DMF—water systems. The micellization of CTAB with aminomethylated calix[4]resorcinarenes proceeds in two steps, while its micellization with phenols is a single-step process. The micellization characteristics depend on the structure and hydrophobicity of the amphiphilic compound and the concentration of DMF.