Analysis of the electron transfer pathway in small laccase by EPR and UV-vis spectroscopy coupled with redox titration

Bacterial small laccases(SLAC) are promising industrial biocatalysts due to their ability to oxidize a broad range of substrates with exceptional thermostability and tolerance for alkaline p H. Electron transfer between substrate, copper centers, and O2is one of the key steps in the catalytic turnover of SLAC. However, limited research has been conducted on the electron transfer pathway of SLAC and SLAC-catalyzed reactions, hindering further engineering of SLAC to produce tunable biocatalysts for novel applications. Herein, the combinational use of electron paramagnetic resonance(EPR) and ultraviolet-visible(UV-vis) spectroscopic methods coupled with redox titration were employed to monitor the electron transfer processes and obtain further insights into the electron transfer pathway in SLAC. The reduction potentials for type 1 copper(T1Cu), type 2 copper(T2Cu) and type 3copper(T3Cu) were determined to be 367 ± 2 mV, 378 ± 5 m V and 403 ± 2 mV,respectively. Moreover, the reduction potential of a selected substrate of SLAC, hydroquinone(HQ), was determined to be 288 mV using cyclic voltammetry(CV). In this way, an electron transfer pathway was identified based on the reduction potentials. Specifically,electrons are transferred from HQ to T1Cu, then to T2Cu and T3Cu, and finally to O2.Furthermore, superhyperfine splitting observed via EPR during redox titration indicated a modification in the covalency of T2Cu upon electron uptake, suggesting a conformational alteration in the protein environment surrounding the copper sites, which could potentially influence the reduction potential of the copper sites during catalytic processes. The results presented here not only provide a comprehensive method for analyzing the electron transfer pathway in metalloenzymes through reduction potential measurements, but also offer valuable insights for further engineering and directed evolution studies of SLAC in the aim for biotechnological and industrial applications.

Determination of pKa values of alendronate sodium in aqueous solution by piecewise linear regression based on acid-base potentiometric titration

As a mono-sodium salt form of alendronic acid,alendronate sodium presents multi-level ionization for the dissociation of its four hydroxyl groups.The dissociation constants of alendronate sodium were determined in this work by studying the piecewise linear relationship between volume of titrant and p H value based on acidbase potentiometric titration reaction.The distribution curves of alendronate sodium were drawn according to the determined p Ka values.There were 4 dissociation constants(pKa_1=2.43,pKa_2=7.55,pKa_3=10.80,pKa_4=11.99,respectively) of alendronate sodium,and 12 existing forms,of which 4 could be ignored,existing in different p H environments.

Pitfalls and Uncertainties of Using Potentiometric Titration for Estimation of Plant Roots Surface Charge and Acid-Base Properties

Amount and properties of roots surface charge are important for nutrient uptake and balance in plants. Roots surface charge markedly varies at different rizosphere conditions (particularly pH and ionic strength), which can markedly alter during vegetation season. Among recently available measuring methods, surface charge-pH dependence of roots (as well as other biological objects) is most easily evaluated by potentiometric titration. Use of this method is also easy at different ionic strengths. Potentiometric titration also allows for estimation of the distribution of charge generating surface groups. However, many applications of this method seem to be based either on incorrect methodical or theoretical approaches. In this paper we discuss the methodical and theoretical backgrounds of the titration method. Basing on experimental titration curves of roots of barley grown in nutrient solution, we show inconsistency of surface charge results obtained at different measuring conditions. Limitations of theoretical interpretations of the results are outlined also.

Investigation of a Simple and Cheap Source of a Natural Indicator for Acid-Base Titration: Effects of System Conditions on Natural Indicators

This study investigated a natural indicator for acid-base titration which is extracted from guinea corn leaves popularly called “waakye leaves” in Ghana. Four types of acid-base titration were studied: strong acid versus (v/s) strong base, strong acid versus weak base, weak acid versus strong base, and weak acid versus weak base. The indicator color change, pH range and the average titre values were determined for each type of acid-base titration. These values were comparable to those obtained from three standard indicators: methyl orange, methyl red and phenolphthalein. Total flavonoids (TF) and condensed tannin (CT) from the crude leaves extract were determined which might be the major reasons for the activity of the extract as an indicator for simple acid-base titration. The authors suggest that the natural indicator is cheap, available, simple to extract, user and environmentally friendly and could be an excellent replacement for standard indicators.

Spectroscopic study on variations in illite surface properties after acid-base titration

FT IR, Raman microscopy, XRD, 29 Si and 27 Al MAS NMR, were used to investigate changes in surface properties of a natural illite sample after acid base potentiometric titration. The characteristic XRD lines indicated the presence of surface Al Si complexes, preferable to Al(OH) 3 precipitates. In the microscopic Raman spectra, the vibration peaks of Si O and Al O bonds diminished as a result of treatment with acid, then increased after hydroxide back titration. The varied ratio of signal intensity between IV Al and VI Al species in 27 Al MAS NMR spectra, together with the stable BET surface area after acidimetric titration, suggested that edge faces and basal planes in the layer structure of illite participated in dissolution of structural components. The combined spectroscopic evidence demonstrated that the reactions between illite surfaces and acid leaching silicic acid and aluminum ions should be considered in the model description of surface acid base properties of the aqueous illite.

Application of Amino Acid-Based Surfactant in Dishwasher Detergent

The application of amino acid-based surfactant in dishwasher detergent was studied in this paper. The foam and oil-removing performance of amino acid surfactants in alkaline condition were test to discuss feasibility of amino acid-based surfactant from natural origin as the alternative to traditional nonionic surfactants from petroleum origin in dishwasher detergent. Comparative study was conducted through using various amino acid-based surfactants and nonionic surfactants respectively, together with enzymes and peroxide to formulate dishwasher detergents, and their application properties such as foam performance and detergency were also evaluated when washing different tableware in automatic dishwasher. The results showed that among all tested amino acid surfactants, glutamate surfactants showed low foaming and high oil-removing ability in alkaline condition, and excellent decontamination ability in dishwashing tests.

Triphenylmethanol and Tris(2-(hydroxymethyl)phenol) Derivatives: Synthesis and Application as Indicators for Acid-Base Volumetric Titration

Polyphenols are naturally occurring compounds found largely in fruits, vegetables, cereals and beverages. Currently, there is much interest in the potential health benefits of dietary plant polyphenols as antioxidants. The effect of polyphenols on human cancer cells is most often protective and induces a reduction in the number of tumors or rate of growth. During our course of study on anticancer prodrugs, twelve triphenylmethanol and one tris(2-(hydroxymethyl) phenol derivatives were synthesized as a carrier of several drugs with optimized lipophilicity. Besides application of these compounds as a foundation for anticancer drug delivery systems, these compounds were evaluated as indicators for the acid-base volumetric titration of a standard solution of hydrochloric acid with a standard solution of sodium hydroxide. The experiments indicated a moderate-to-sharp color transition of the solutions near the neutralization point for most indicators. These indicators may have potential applications for acid-base titrations in a narrow range.

Determination of Carboxyl Group on Fabric by Acid-base Titration

Polycarboxylic acids have been widely researched as non-formaldehyde DP finishes since late 1980s. However, there has no satisfied wet chemical method for quantifi-cation the carboxylic groups or the esterlinkages on fab-ric, which is important in investigation some basic as-pects of the esterification between polycarboxylic acids and cellulose. In this paper, the wet chemical analysis method, Ca(AC)2 back - titration, had been established to determinate the COOH on fabric. The reliability of this method had also been proved.

Determination of aniline in environmental water samples by alternating-current oscillopolarographic titration

A new method for the determination of aniline in environmental water based on oscillopolarographic titration was presented in this paper. Several factors including the kind, concentration, and volume of acid, the dosage of potassium bromide, the temperature and concentration of concomitant substances were investigated in detail. The experimental results indicated that this method was simple, rapid, and sensitive. The linear range was 8.367 × 10(?4) to 2.789 × 10(?2) mol L(?1), the relative standard deviation (R.S.D.) was lower than 0.96%, and the spiked recoveries of aniline in environmental water samples were in the range of 99.4–106.9% under the optimal conditions. The results indicated that the present method could be used as an alternative method for aniline determination in realworld water samples.

Study of Complexes of Lanthanum with Amino Acids by Titration Calorimeter

The stability constants and thermodynamic functions for complexes of lanthanum with eight kind of amino acids according to 1∶1 and 1∶2 in proportion have been determined by titration calorimeter at 298.15 K. The enthalpy change makes a predominant contribution to the stability of these complexes. The ring in amino acid associated with lanthanum ion helps to enhance the stability of complexes. Steric effects between rings in complexes leads to that the equilibrium constants of reaction of the complexes (1∶2) is much less than that of the complexes (1∶1).

POTENTIOMETRIC TITRATION OF CALCIUM IN SEAWATER

By using calcium ion selective elatrode (Ca ISE) as indicating electrode, saturated calomel electrode (SCE) as reference electrode and EGTA as titrant, calcium in seawater was determined by potentiometric titration in borate buffer solution. This method can reduce observation errors in the determination of the endpoint, and thus provide better analytical precision(<0.08%) than present complexometric titration.

Conductometric Titration for the Prediction of Conductivity in Solutions with Polymeric Membranes

The analytical technique of conductometric titration is used to characterize polymeric materials.This technique allows obtaining the polymer dry weight capacity(DWC) and the extent of reaction and establishing the optimal number of water molecules per sulfonic groups.In this particular case the polymer material under study was Nafion?117.Two different reactions were made: Neutralization and exchange.Both of them allow obtaining the same result.Conductometric titrations could be considered as a good method to study the neutralization and exchange reactions for polymeric materials used in fuel cells or electrolyzers.The implementation of this analytical technique allows reducing operation time of those types of materials.The waiting time between determinations permits to say that the amount of reactant consumed and products obtained were the same after each addition of titrant.

Extraction Mechanism of Rare Earths with Sec-Octylphenoxy Acetic Acid by Two-Phase Titration Technique

The compositions of the extracted complexes of La, Gd, Er and Y with sec octyl phenoxy acetic acid in heptane and the related apparent extraction equilibrium constants K M were determined using two phase titration technique. The stoichiometric compounds for La, Gd, Er and Y should be LaA 3·2.5HA, GdA 3·3HA, ErA 3·3.1HA and YA 3·4.3HA respectively. And their p K M are 3.43, 3.46, 3.08 and 2.58 respectively.

Rapid determination of phenolic compounds in water samples by alternating-current oscillopolarographic titration

A rapid, simple and sensitive method was demonstrated for the determination of phenolic compounds in water samples by alternating-current oscillopolarographic titration. With the presence of sulfuric acid, phenol could be transferred into a nitroso-compound by reacting with NaNO2. The titration end-point was obtained by the formation of a sharp cut in the oscillopolarographic with infinitesimal NaNO2 on double platinum electrodes. The results showed that phenol had an excellent linear relationship over the range of 4.82×10^-6 -9.65×10^-3 mol/L, the RSD of the proposed method was lower than 1.5%, and the spiked recoveries of three real water samples were in the range of 95.6%-106.9%.

Two Dehydroabietic Acid-based Arylamines: Synthesis, Crystal Structure and Fluorescent Properties

Two dehydroabietic acid-based arylamines have been synthesized and characterized by FTIR, 1H NMR, 13C NMR, MS spectra and elemental analysis. Their spatial structures were determined by X-ray diffraction analysis. UV-Vis absorption and fluorescence spectral characteristics of these compounds in methanol were investigated. Their fluorescence emission spectra in different polarity solvents were further evaluated. Fluorescent properties and structural relationship of the compounds showed that fluorescence intensity and quantum yield inversely increase with the non-coplanar degree. In addition, the solvent polarity has different effects on the fluorescence emission spectra of two compounds.

A Multiparameter Colloidal Titrations for the Determination of Cationic Polyelectrolytes

In water treatment processes and conditioning drinking water, PEs are widely used;however, their environmental impact is still doubtful, since residual concentrations increase organic matter content and represents a potential health hazard. This paper demonstrates a multiparametric study of two colloidal titration methods: spectrophotometric and zeta potential end point detection. The first one was optimized to guarantee the accuracy of cationic commercial PE quantification. It includes the indicator dose optimization using analytical criteria for competing equilibria, a calibration curve for two ranges of CPE concentration (1 - 5 ppm and 5 - 100 ppm) and the interference study of flocculant and Sn in the CPE quantification. The second method provides a physicochemical validation of the electric surface phenomena occurring during the colloidal titration and the end point detection. As an additional contribution the zeta potential titration was discussed and proposed as an alternative method for quantifying CPE when the sample is metal free.

Prospects for nucleic acid-based therapeutics against hepatitis C virus

In this review,we discuss recent advances in nucleic acid-based therapeutic technologies that target hepatitis C virus(HCV)infection.Because the HCV genome is present exclusively in RNA form during replication,various nucleic acid-based therapeutic approaches targeting the HCV genome,such as ribozymes,aptamers,siRNAs,and antisense oligonucleotides,have been suggested as potential tools against HCV.Nucleic acids are potentially immunogenic and typically require a delivery tool to be utilized as therapeutics.These limitations have hampered the clinical development of nucleic acid-based therapeutics.However,despite these limitations,nucleic acid-based therapeutics has clinical value due to their great specificity,easy and large-scale synthesis with chemical methods,and pharmaceutical flexibility.Moreover,nucleic acid therapeutics are expected to broaden the range of targetable molecules essential for the HCV replication cycle,and therefore they may prove to be more effective than existing therapeutics,such as interferon-αand ribavirin combination therapy.This review focuses on the current status and future prospects of ribozymes,aptamers,siRNAs,and antisense oligonucleotides as therapeutic reagents against HCV.

Interaction of Ca^(2+) and soil humic acid characterized by a joint experimental platform of potentiometric titration, UV–visible spectroscopy, and fluorescence spectroscopy

Rocky desertification has become a major environmental issue in the karst region of southwestern China.Karst rocky desertification was more severe in regions of limestone soil than in adjacent regions of other soils,despite the relatively higher soil organic matter(SOM)content in limestone soil.The underlying mechanism remains ambiguous.We speculated that the geochemical characteristics of limestone soils in the karst region plays an essential role,especially the high calcium content of limestone soil.To test this hypothesis,we collected limestone soil samples from a limestone soil profile in the southwestern China karst region and extracted humic acid(HA)from these limestone soil samples.We investigated the interaction of Ca^(2+)and three HA samples on a joint experimental platform,which consists of an automatic potentiometric titrator,a UV–visible spectrometer,and a Fluorescence spectrometer.HA solutions were titrated by Ca^(2+)and optical spectra of the HA solutions were monitored during the titration experiments.The results indicated that:(1)the interaction of Ca^(2+)and HA is a combined process of adsorption and complexation.Adsorption dominated the overall distribution behavior of Ca^(2+),which could be fit by Langmuir and Freundlich isotherm models.Complexation was distinguished only when the concentration of Ca2+is low;(2)the changes of UV–visible spectroscopy and excitation–emission matrix fluorescencespectroscopy spectra of HA samples when they were binding with Ca^(2+)implied the apparent molecular size and structure of HA became larger and more complex;(3)the combination of Ca^(2+)and HA plays an important role in the SOM preservation of limestone soils but the stability of the Ca–HA association was relatively weak.The present study draws attention to maintaining the relatively higher Ca^(2+)concentration in limestone soils in ecologic restoration attempts in karst regions.

Preparation and Characterization of Cathode Materials La_(0.7)Sr_(0.3-x)Ca_xCo_(0.9)Fe_(0.1)O_(3-δ) by Reverse Titration Co-Precipitation Method for ITSOFC

The precursors of La0.7Sr0.3-xCaxCo0.9Fe0.1O3-δ(LSCCF, x=0.05, 0.10, 0.15, 0.20) as the cathode materials for intermediate temperature solid oxide fuel cell (ITSOFC) were prepared by reverse titration co-precipitation method with metal-nitrates as starting materials and mixed alkali (NaOH and Na2CO3) as a precipitating agent. The formation process of LSCCF from the precursors was monitored by TG-DSC, and the crystal structure and particles morphology of the precursors which were calcined at 600, 800, 1000 ℃ for 3 h were characterized using XRD, SEM technologies. Compared with the solid state reaction of constituent oxides, when the pH value of the precipitating solution was in the range of 9.1～9.5, the LSCCF powders from the precursors caclined at 800 ℃ for 3 h had high purity, homogeneous and single perovskite phase. The electrical conductivity of the LSCCF samples sintered at 1200 ℃ for 3 h, which was measured as a function of temperatures from 100 to 800 ℃ by DC four-probe method in air, decreased with x from 0.05 to 0.20. The value of electrical conductivity was almost equal because of Ca2+, Sr2+ co-dopant resulting in the 'mix effect' while x=0.10 or 0.15. The electrical conductivity of all doped samples was higher than 100 S·cm-1 at intermediate temperatures from 500 to 800 ℃, and there was good compatibility between the LSCCF cathode and Ce0.8Sm0.2O2 electrolyte.