Lactic acid extraction with trioctyl amine

Lactic acid extraction was carried out with trioctyl amine (TOA) in three diluents. The effect of initial lactic acid concentrations on the extraction efficiency was investigated. It was observed that although the percentage extraction remained constant or decreased but the loading ratio was increased in all the cases. The overloading was observed in the case of TOA in methyl isobutyl ketone (MIBK). The extraction of lactic acid was favored at a lower aqueous pH?in all the diluents. The improvement of the extraction efficiency at a higher aqueous pH?(=?6) was achieved by using the modified TOA (treated with HCl) in MIBK. However, the recovery of lactic was very poor in the case of modified TOA in MIBK, although the complete recovery was obtained for untreated TOA.     

Reactive extraction of lactic acid with mixed tertiary amine extractants

The mixture of tripropylamine (TPA) and trioctylamine (TOA) dissolved in 1-octanol/n-heptane was used in the reactive extraction of (L+)lactic acid in aqueous solution. Maximum distribution coefficients were obtained in the range from 6:4 to 8:2 weight ratio of TPA/TOA at 5% (w/w) lactic acid in aqueous phase and their extraction efficiencies were above 90%. By introducing TPA into TOA, the third phase formation could be overcome, thereby, the settling time is shorter than in the case of TOA only.     

Kinetics of the extraction of succinic acid with tri-n-octylamine in 1-octanol solution

Kinetic studies for the extraction of succinic acid from aqueous solution with 1-octanol solutions of tri-n-octylamine (TOA) were carried out using a stirred cell with a microporous hydrophobic membrane. The interfacial concentrations of species were correlated and thus the intrinsic kinetics was obtained. The overall extraction process was controlled by the chemical reaction at or near the interface between the aqueous and organic phases. The formation reaction of succinic acid-TOA complex was found to be first order with respect to the concentration of succinic acid in the aqueous phase and the order of 0.5 with respect to that of TOA in the organic phase with a rate constant of (3.14 +/- 0.6) x 10(-8) m(2.5) x mol(-0.5) x s(-1). The dissociation reaction of succinic acid-TOA complex was found to be the second-order with respect to that of succinic acid-TOA complex in the organic phase and the order of -2 with respect to that of TOA in the organic phase with a rate constant of (1.44 +/- 1.4) x 10(-4) mol x m(-2) x s(-1).     

Modulation of Hydra attenuata rhythmic activity: phase response curve.

We investigated the effect of photic stimulation on the frequency of Hydra attenuata column contractions. We used positive or negative abrupt light transitions, single or repetitive light or darkness pulses, and alternation of light and darkness periods. The main results are: (a) The frequency of the contraction pulse trains (CPTs) varies transiently in response to an abrupt variation of the light intensity. (b) CPTs in progress can be inhibited by different types of photic stimuli. (c) The response time to a single photic stimulus varies during the inter-CPT interval and depends also on the polarity of the stimulus. (d) The CPTs are entrainable with repetitive light stimulation of various frequencies. (e) Long-lasting variations of the frequency of CPTs occur after the end of a repetitive light stimulation. We suggest that the mechanism responsible for the rhythym of column contractions is quite similar to that on which other biological rhythmic phenomena are based.     

Investigations of biocompatible systems for reactive extraction of propionic acid using aminic extractants (TOA and Aliquat 336)

This paper presents the reactive extraction of propionic acid from aqueous solution by amine based extractants such as tri-n-octylamine and Aliquat 336, dissolved in a mixture of n-dodecane and 1-decanol. Equilibrium experiments were carried out to investigate the effects of various parameters such as modifier (1-decanol) concentration, extractant type, extractant composition, diluent composition, and initial acid concentration on the extraction efficiency. The extraction efficiency was found to be increased with an increase in modifier composition and extractant composition, and decreased with increases in initial acid concentration. Different biocompatible extractant/diluent systems such as (1) 20% TOA, 20% 1-decanol and 60% n-dodecane, (2) 20% TOA, 30% 1-decanol and 50% n-dodecane, (3) 30% TOA, 20% 1-decanol and 50% n-dodecane and (4) 25% Aliquat 336, 25% 1-decanol and 50% n-dodecane are developed and used in this study. A mathematical model based on mass action law and a population-based search algorithm (differential evolution, DE) is proposed, and is used to estimate the extraction equilibrium constant (K _E) and stoichiometry of reactive extraction. Individual equilibrium constants for the simultaneous formation of (1:1) and (2:1) acid:amine complexes are also determined. The extraction system comprised of 20% TOA, 30% 1-decanol, and 50% n-dodecane was found to be the best among the four biocompatible extractant/diluent systems studied. The loading ratios found in the range of 0.113 ～ 1.05 indicated the simultaneous formation of 1:1 and 2:1 complexes between acid and TOA.     

Extraction of succinic acid with 1-octanol/n-heptane solutions of mixed tertiary amine

The reactive extraction of succinic acid was carried out by mixed tertiary amine which consisted of tripropylamine (TPA) and trioctylamine (TOA) as the extraction agent in 1-octanol/n-heptane diluent. Maximum distribution coefficient was obtained at 8:2 weight ratio of TPA/TOA. At this ratio, its extraction efficiency is above 90% at the 3.9 wt.% of succinic acid in aqueous solution. Furthermore, the prevention of the third phase formation made the phase separation between organic phase and aqueous phase easy.     

Predispersed solvent extraction of succinic acid aqueous solution by colloidal liquid aphrons in column

A study of the PDSE (predispersed solvent extraction) for succinic acid by colloidal liquid aphrons was conducted. The organic phase contaning TOA (tri-n-octylamine) and 1-octanol permits a selective extraction of succinic acid from its aqueous solution. There was no difference of the extractability of PDSE and that of conventional mixer-settler type extraction. Taking into account the no mechanical mixing in PDSE, it was concluded that the PDSE process is more adaptive than the conventional mixer-settler type extraction process. From mass transfer analysis at the various concentration of TOA in counter-current continuous operation, the concentration of TOA had no influence on the mass transfer coefficient. The loading values in continuous PDSE were almost same as those in batch operation.     

Effect of pH on the extraction characteristics of succinic and formic acids with Tri-n-octylamine dissolved in 1-octanol

A study was made on the extraction equilibria of succinic and formic acids from aqueous solutions using tri-n-octylamine (TOA) in 1-octanol. It was shown that the loading values of TOA decreased with increasing pH values. The apparent equilibrium constants for each acid-amine complex were determined by an equilibrium model. In the case of succinic acid, the formation of a bisuccinate anion played an important role in the stoichiometry of the acid-amine complex.     

Cloud-point temperatures for lysozyme in electrolyte solutions: effect of salt type, salt concentration and pH

Liquid-liquid phase-separation data were obtained for aqueous saline solutions of hen egg-white lysozyme at a fixed protein concentration (87 g/l). The cloud-point temperature (CPT) was measured as a function of salt type and salt concentration to 3 M, at pH 4.0 and 7.0. Salts used included those from mono and divalent cations and anions. For the monovalent cations studied, as salt concentration increases, the CPT increases. For divalent cations, as salt concentration rises, a maximum in the CPT is observed and attributed to ion binding to the protein surface and subsequent water structuring. Trends for sulfate salts were dramatically different from those for other salts because sulfate ion is strongly hydrated and excluded from the lysozyme surface. For anions at fixed salt concentration, the CPT decreases with rising anion kosmotropic character. Comparison of CPTs for pH 4.0 and 7.0 revealed two trends. At low ionic strength for a given salt, differences in CPT can be explained in terms of repulsive electrostatic interactions between protein molecules, while at higher ionic strength, differences can be attributed to hydration forces. A model is proposed for the correlation and prediction of the CPT as a function of salt type and salt concentration. NaCl was chosen as a reference salt, and CPT deviations from that of NaCl were attributed to hydration forces. The Random Phase Approximation, in conjunction with a square-well potential, was used to calculate the strength of protein-protein interactions as a function of solution conditions for all salts studied.     

cis-Prenyltransferase AtCPT6 produces a family of very short-chain polyisoprenoids in planta

cis-Prenyltransferases (CPTs) comprise numerous enzymes synthesizing isoprenoid hydrocarbon skeleton with isoprenoid units in the cis (Z) configuration. The chain-length specificity of a particular plant CPT is in most cases unknown despite the composition of the accumulated isoprenoids in the tissue of interest being well established. In this report AtCPT6, one of the nine Arabidopsis thaliana CPTs, is shown to catalyze the synthesis of a family of very short-chain polyisoprenoid alcohols of six, seven, and eight isoprenoid units, those of seven units dominating. The product specificity of AtCPT6 was established in vivo following its expression in the heterologous system of the yeast Saccharomyces cerevisiae and was confirmed by the absence of specific products in AtCPT6 T-DNA insertion mutants and their overaccumulation in AtCPT6-overexpressing plants. These observations are additionally validated in silico using an AtCPT6 model obtained by homology modeling. AtCPT6 only partially complements the function of the yeast homologue of CPT-Rer2 since it restores the growth but not protein glycosylation in rer2Δ yeast. This is the first in planta characterization of specific products of a plant CPT producing polyisoprenoids. Their distribution suggests that a joint activity of several CPTs is required to produce the complex mixture of polyisoprenoid alcohols found in Arabidopsis roots.     

Recovery of pyruvic acid from biotransformation solutions

The aim of this investigation was to separate pyruvic acid of biotransformation solutions from lactic acid through complex extraction. For this purpose, complex extraction was investigated from model solutions. Tri-n-octanylamine (TOA) was used as the extractant. The effects of various diluents, the stoichiometry of pyruvic acid to TOA, and the initial pH of the aqueous phase on the extraction process were investigated in this study. The effects of sodium hydroxide (NaOH) and trimethylamine (TMA) on the back extraction process were also studied, respectively. The optimal conditions attained from the model solutions proved efficient on the biotransformation solutions of different concentrations. A total recovery of 71–82% of pyruvic acid was obtained, whereas 89–92% of lactic acid was removed. The purity of pyruvic acid reached 97% after the removal of TMA by a simple distillation.     

Iodination of proteins by IPy2BF4, a new tool in protein chemistry

Iodination is a very useful method for protein characterization and labeling. However, derivatization chemistries used in most conventional iodination procedures may cause substantial alterations in protein structure and function. The IPy(2)BF(4) reagent [bis(pyridine)iodonium (I) tetrafluoroborate] has been shown to be an effective iodinating reagent for peptides. Herein we report the first application of IPy(2)BF(4) in protein iodination in an aqueous medium using three representative substrates: insulin, lysozyme, and the enzyme 1,3-1,4-beta-d-4-glucanohydrolase. Our results show that IPy(2)BF(4) has clear advantages over existing methods in that the reaction is quantitative, fast, and selective for the most accessible Tyr residues of a protein, and it preserves the functional integrity of the protein when moderate Tyr labeling levels are pursued.     

Secondary structures of proteins adsorbed onto aluminum hydroxide: infrared spectroscopic analysis of proteins from low solution concentrations

Comparative studies of the secondary structures of six model proteins, adsorbed onto aluminum hydroxide gel (Alhydrogel) or in aqueous solution, were carried out by Fourier transform infrared (FTIR) spectroscopy. The analysis of high-quality spectra of all six model proteins, with a broad range of secondary structure compositions, obtained at 15 mg/ml by the conventional method and at 0.5 and 1.0 mg/ml adsorbed to Alhydrogel revealed that adsorption onto hydrophilic surfaces of aluminum hydroxide particles did not alter the secondary structures of the proteins. The results of this study suggest that adsorbing proteins to Alhydrogel provides a means of obtaining FTIR spectra to study secondary structure and conformational changes of proteins in aqueous solution at very low concentrations. The new procedure effectively lowers the concentration requirement for FTIR studies of proteins in aqueous solutions by at least 40-fold, as compared with the conventional FTIR method. It permits FTIR study of proteins to be carried out in the same concentration range as is used for circular dichroism and fluorescence, thereby making it possible to compare structural information obtained by three commonly used techniques in protein biophysical characterization.     

Structure and properties of camptothecin derivatives, their protonated forms, and model interaction with the topoisomerase I-DNA complex

The structure and properties of the 11 Camptothecin derivatives (CPTs) and their different mono-, di-, and triprotonated forms, depending on the number of proton accepting centers in the molecules are studied both theoretically and experimentally by quantum chemical approaches, electronic absorption, and CD spectroscopy. The study of the protonated forms of the CPTs and search of the electron-withdrawing groups is crucial of the water-solubility of the novel medications. Thus, the model interaction of the different protonated molecular species with the Topoisomerase I-DNA complex are elucidated and discussed with a view to understand the mode of binding of the CPTs depending on the type of the substituents and pH of the medium.     

Economic comparison of multiple techniques for recovering leaf protein in biomass processing

Leaf protein concentrates (LPC) can be used as a valuable co-product to cellulosic biofuel production and can also mitigate the food versus fuel controversy. Two major approaches have been considered for LPC production: a well-characterized mechanical pressing method and a less studied method involving aqueous extraction with recovery using ultrafiltration. Experimental results with switchgrass extracts show low protein recovery after filtration, particularly if protein is recovered after cellulose hydrolysis. Economic modeling suggests that aqueous extraction costs less than mechanical pressing, but due to lower protein yields and lower quality, overall profit is higher for mechanical pressing versus aqueous extraction ($26/Mg feedstock vs. $14/Mg). If modest improvements can be made in extraction yields, filtration recovery, and protein quality, then the profitability of the aqueous extraction approach can be increased to $37/Mg feedstock. This study suggests that aqueous extraction is a viable alternative for LPC co-production in a biorefinery if key improvements can be made in the process.     

Molecular dynamics method for proteins with ionization-conformation coupling and equilibrium titration

A new realization of the constant pH molecular dynamics simulation is proposed in mean force potential of proton reservoir in constant titration conditions. The MD-pH-ET method is used for a protein in the most probable ionization microstate, which is the state with minimal energy of the current conformation taking into account the correcting ionization potential of mean force considering an equilibrium ensemble of ionization states. The new MD-pH-ET method allows one to carry out an optimization of protein structure and the total free energy of a protein in the aqueous solution at constant pH and to calculate the pH-dependent properties. The MD-pH-ET method possesses the following unique features: (1) it uses the most precise and computationally effective realization of calculation of electrostatic energy of a protein in aqueous solution, the model of continuous dielectric media with Poisson equation, and the generalized Born method with “ideal” Born atomic radii; (2) it uses the same model of the potential energy surface in the ionization-conformation phase space both for calculating the potential energy of the protein and atomic forces and for determining the most probable ionization states; (3) it calculates the total free energy of the protein in the aqueous solution in a proton reservoir under the conditions of equilibrium titration. The workability of the new method MD-pH-ET is demonstrated for a molecule of the BPTI protein.     

Effect of operating variables on back-extraction characteristics of succinic acid from organic phase

Tri-n-octylamine (TOA) is an effective extractant for the recovery of succinic acid from fermentation broth. The recovery of succinic acid from organic phase depends on the operating variables such as temperature, pH, volume of aqueous phase, and use of displacer. In thermal recovery of succinic acid, 34% of succinic acid was recovered at 90°C. More than 90% of succinic acid was back-extracted by pH-swing. Efficiency of back-extraction was increased 23% as increasing volume of aqueous phase. Use of oleic acid as a displacer increased efficiency of back-extraction to 76%. Aqueous phase volume and concentration of oleic acid were controlled simultaneously to increase the efficiency of back-extraction and percent recovery of succinic acid reached to 90.     

Purification of recombinant protein A by aqueous two-phase extraction integrated with affinity precipitation

Aqueous two-phase extraction incorporated affinity precipitation was examined as a technique for protein purification. An enteric coating polymer, Eudragit S100, was employed as a ligand carrier. Eudragit was specifically partitioned to the top phase in the aqueous two-phase systems. For application of this method to purification of recombinant protein A using human IgG coupled to Eudragit in an aqueous two-phase system, 80% of protein A added was recovered with 81% purity. The purity was enhanced 26-fold by thid method. The IgG-Eudragit could be used repeatedly for the purification process. This seperation method should be applicable to industrial-scale purification as a new purification procedure combining the advantages and compensating for the disadvantages of the aqueous two-phase method and affinity precipitation method. (c) 1992 John Wiley & Sons, Inc.     

Protein extraction using the sodium bis(2-ethylhexyl) phosphate (NaDEHP) reverse micellar system

The reverse micellar system of sodium bis(2-ethylhexyl) phosphate (NaDEHP)/isooctane/brine was used for liquid-liquid extraction of proteins. We investigated the solubilization of cytochrome-c and alpha-chymotrypsin into the NaDEHP reverse micellar phase by varying the pH and NaCl concentration in the aqueous phase. At neutral pH and relatively low ionic strength, the proteins are extracted into the micellar phase with high yield. By contacting the micellar phase with a divalent cation (e.g., Ca(2+)) aqueous solution, the reverse micelles are destabilized and release the protein molecules back into an aqueous solution for recovery. This method separates the proteins from the surfactant with very high overall efficiencies. (c) 1996 John Wiley & Sons, Inc.     

Relationship between preferential interaction of a protein in an aqueous mixed solvent and its solubility

The present paper is devoted to the derivation of a relation between the preferential solvation of a protein in a binary aqueous solution and its solubility. The preferential binding parameter, which is a measure of the preferential solvation (or preferential hydration) is expressed in terms of the derivative of the protein activity coefficient with respect to the water mole fraction, the partial molar volume of protein at infinite dilution and some characteristics of the protein-free mixed solvent. This expression is used as the starting point in the derivation of a relationship between the preferential binding parameter and the solubility of a protein in a binary aqueous solution. The obtained expression is used in two different ways: (1) to produce a simple criterion for the salting-in or salting-out by various cosolvents on the protein solubility in water, (2) to derive equations which predict the solubility of a protein in a binary aqueous solution in terms of the preferential binding parameter. The solubilities of lysozyme in aqueous sodium chloride solutions (pH=4.5 and 7.0), in aqueous sodium acetate (pH=8.3) and in aqueous magnesium chloride (pH=4.1) solutions are predicted in terms of the preferential binding parameter without any adjustable parameter. The results are compared with experiment, and for aqueous sodium chloride mixtures the agreement is excellent, for aqueous sodium acetate and magnesium chloride mixtures the agreement is only satisfactory.