Apparent Kinetic Parameters Research Articles

Adsorption of Myoglobin on Calixarenes and Biocatalysis in Organic Media

Derivatives of p-tert-butylcalix(4,6,8)arene car- boxylic acids were used for selective adsorption of myoglobin. A mixture of myoglobin, laccase and peroxidase was used for extraction with calixarenes and only myoglobin was selectively extracted to organic media. Myoglobin and Mb c-calixarene exhibited pseudoactivity of peroxidase in aqueous and organic media. This protein-calixarene complex exhibited the highest specific activity of 1.37 × 10 −1 U.mg protein −1 at initial pH 6.5 of myoglobin aqueous solution. Apparent kinetic parameters (V � max, K �, k � cat and k � cat/K � m) for the pseudoperoxidase activity were determined in organic media for different initial pH values of myoglobin aqueous solution by Michaelis-Menten plot. The stability of this complex was studied for different initial pH values and t1/2 values were obtained in the range of 3.5-5.2 days. The extracted Mb c in organic media was recovered into fresh aqueous solutions at alkaline pH with a recovery of pseudoperoxidase activity of over 100%.

Kinetic Modeling of Liquid Phase Photocatalysis on Supported TiO2 Nanoparticles in a Rectangular Flat-Plate Photoreactor

The photocatalytic degradation of C.I. Basic Blue 3 (BB3) in water on supported TiO2 nanoparticles was carried out in a rectangular flat-plate photoreactor. The photocatalytic degradation kinetic characteristics were experimentally investigated under different light intensities and initial dye concentrations. Calculating corresponding “apparent adsorption constants”, KS, and “apparent rate constants”, kr, of experimental results according to the Langmuir−Hinshelwood (L-H) model emphasized that the L-H constants varied with regard to the light intensity. A new kinetic model was developed on the basis of the intrinsic element reactions, which took into account the effect of light intensity, reaction intermediates, organic compound content, and first order reaction kinetics. The new kinetic model described the experimental results in a more accurate way and explained consistently the dependence of the apparent kinetic parameters kr and KS on the light intensity.

Kinetic study of pyrolysis of waste water treatment plant sludge

AbstractActivated sewage sludge samples obtained from two different waste water treatment plants were investigated by thermogravimetric analysis. Due to a very high content of water in the sludge samples, these had to be dried at 160°C in an electrical oven in order to remove all adsorbed water. To ensure pyrolysis conditions, nitrogen atmosphere was applied. The pyrolysis decomposition process was carried out in the temperature range from ambient temperature to 900°C at three different heating rates: 2 K min−1, 5 K min−1, 10 K min−1. TGA and DTG curves of the decomposition processes were obtained. Temperature of onset decomposition, final temperature of decomposition, maximum decomposition rate, and decomposition temperature were determined by thermogravimetric analysis for both sludge samples used. The main decomposition process takes place at temperatures in the range from 230°C to 500°C. Above this temperature, there are only small changes in the mass loss which are often attributed to the decomposition of carbonates present in the sewage sludge samples. To determine the apparent kinetic parameters such as the activation energy and the preexponential factor, the so called Friedman isoconversional method was used. Because of the requirements of this method, initial and final parts of the decomposition process, where crossings of the decomposition lines occurred, were cut off. Obtained dependencies of the apparent activation energies and preexponential factors as a function of conversion were used backwards to calculate the modeled decomposition process of sewage sludge and the experimental data were in good accordance with the data obtained by simulation.

Biomass pyrolysis in pulverized-fuel boilers: Derivation of apparent kinetic parameters for inclusion in CFD codes

Biomass combustion in pulverized-fuel boilers is a growing way to produce electricity from a renewable source of energy. Slagging and fouling limit however the reliability of the units that were initially designed for coal combustion. Computational Fluid Dynamics (CFD) codes aiming at studying those phenomena include simplified models of biomass particle pyrolysis, of which the pertinence has already been questioned for the typical conditions of interest. A comprehensive model has been developed to investigate pyrolysis of particles in pulverized-fuel boilers, with sizes ranging from 17 μm to 2.5 mm. The detailed model accounts for internal heat conduction, internal gaseous convection, moisture evaporation and particle shrinkage. It includes a competitive, multi-component kinetic scheme, improved for high temperatures. The discrepancy between the simplified models integrated in most CFD applications and the detailed simulations is highlighted. The simplified isothermal models underestimate pyrolysis time for the largest particles. Moreover, such models delay and shorten the volatiles release. The flame lengths, the local temperature fields and the pollutant emissions might be importantly impacted in global combustion simulations. Apparent kinetic parameters have been derived from the detailed simulations. Their use in existing simplified models improves the behavior of the biomass particles during pyrolysis, and offers therefore an efficient alternative to the integration of complex pyrolysis models in CFD codes.

Characterization and catalytic evaluation of aluminum oxides obtained by the atrane route

The atrane route (aluminum tri-sec butoxide as the metal source mixed with tri-ethanolamine to yield the alumatrane complex) was used to synthesize aluminas that showed specific surface areas in the 150–275 m 2/g range and average pore sizes in the 60–180 Å range after calcination. Different techniques were used to characterize the aluminas (IR, XRD, 27Al MAS-NMR, N 2 adsorption). The amount of organic matter trapped in the inorganic matrix in the uncalcined materials decreased as a function of the increasing molar relationship between water and the atrane complex in the synthesis, which ranged from 9 to 78. Crystalline micro domains of the γ-AlOOH boehmite-type were formed in the uncalcined samples that increased their size as a function of the proportion of water in the synthesis, because water favors the hydrolysis and condensation of the Al[N(CH 2–CH 2–O) 3] 2H 3 complex. The calcination treatment induced the crystalline restructuration from boehmite-type to γ-alumina-type structures. Thus, the calcined mesoporous aluminas showed an amorphous structure, with crystal micro domains of γ-alumina-type that also increased their size as a function of the amount of water in the starting mixtures. As compared to the properties of a sample prepared by the conventional sol–gel method, the moderate hydrothermal treatment in the atrane synthesis route favored the release of trapped organic matter and the formation of initial boehmite-type micro domains; specific surface areas were similar, but the pore size distributions were sharper in the materials prepared with the atrane route. The activities of the aluminas prepared by the atrane route, as indicated by the TIPB conversion at 500 °C and short reaction times from 12 to 30 s in a CREC Riser Simulator laboratory reactor, were somewhat smaller than those observed in conventional aluminas. The apparent kinetic parameters in a simple, first order model were similar, suggesting that accessibility limitations from the pore systems were not present. The properties shown by these aluminas synthesized by the atrane route indicated a preliminary appropriate condition for being used as FCC catalyst matrices.

Resolution of racemic sulfoxides with high productivity and enantioselectivity by a Rhodococcus sp. strain as an alternative to biooxidation of prochiral sulfides for efficient production of enantiopure sulfoxides

Whole cells of Rhodococcus sp. ECU0066 were used a catalyst for resolution of racemic sulfoxides, as an alternative to asymmetric oxidation of sulfides for efficient production of enantiopure sulfoxides. Racemic sulfoxides were excellent substrates for biotransformation because of their lower biotoxicity compared to sulfides. Determination of apparent kinetic parameters indicated that phenyl methyl sulfide (PMS), but not racemic phenyl methyl sulfoxide (rac-PMSO) caused substrate inhibition. (S)-PMSO was formed at a higher concentration and good enantiomeric excess (37.8 mM and 93.7% ee(S)) in a fed-batch reaction, than by an asymmetric oxidation of PMS (10 mM and 80% eeP (S)). The bacterium also displayed fairly good activity (yields, 22.7-43.2%; within 1-8 h) and enantioselectivity (ee(S)>99.0%) towards para-substituted (methyl and chloro) phenyl methyl sulfoxides and ethyl phenyl sulfoxide, indicating it could be a promising agent for synthetic applications.

First description of a laccase-like enzyme in soil algae

Laccases (EC 1.10.3.2) are versatile multi-copper oxidases so far found in higher plants, fungi, insects, prokaryotes and lichens. In the present study, the production of an extracellular laccase-like enzyme by the coccoid green soil alga Tetracystis aeria was investigated and the enzyme was partly characterized, thereby providing the first description of a laccase-like enzyme in soil algae. Enzyme production in algae cultures was considerably increased by addition of the fungal laccase inducer copper sulphate. Maximal enzyme production was observed during the stationary growth phase. Peroxidase or tyrosinase activity was not detected. The native enzyme exhibits an apparent molecular mass of about 212 kDa as observed with size exclusion chromatography and about 210-260 kDa as estimated by zymograms. The enzyme efficiently oxidizes 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid) (ABTS), 2,6-dimethoxyphenol (2,6-DMP), syringaldazine (SGZ) and the anthraquinone dye Acid Blue 62, while guaiacol and Remazol Brilliant Blue R are only poorly oxidized. The apparent kinetic parameters obtained for ABTS, 2,6-DMP and SGZ oxidation are within the range reported for fungal laccases. Oxidation of the phenolic substrate 2,6-DMP displays a remarkably high pH optimum (pH 8.0-8.5), which is interesting with respect to potential biotechnological applications.

Comparison of a-amylase activity in some sprouting Nigerian cereals

The yield and characteristics of Alpha-amylase obtained from some sprouting Nigeria cereals (maize, acha, rice and sorghum) were compared using standard procedures. The result indicated maximum a-amylase yield was attained from 72-120 hours of sprouting cereals). Of all the sprouted cereals, acha gave rise to the highest amylase yield (16.6 ± 0.02x102 and the least was exhibited by maize (11.0±0.04x102). The optimum temperature for a-amylase activity was 600C for maize and rice while that of acha and sorghum was 700C. The optimal pH for the activity of a-amylase from sprouting maize, rice, acha, and sorghum were exhibited at 6.5, 5.5, 6.5 and 5.8 respectively. The apparent kinetic parameters, Vmax and Km, were 9.0x10-2 and 0.23; 12.5x10-2 and 0.56; 7.0x10-2 and 0.29; 12.5x10-2 and 0.50 for a-amylase from sprouting maize, acha, rice and sorghum respectively. It is concluded that a-amylase obtained from sprouting Nigeria cereals manifest differences in quality and quantity, and may be useful in different starch processing industries.

The Effect of Ozone on the Removal Effectiveness of Photocatalysis on Indoor Gaseous Biogenic Volatile Organic Compounds

In this study, the degradation of d-limonene by photocatalytic oxidation (PCO) (titanium dioxide [TiO2]/ultraviolet [UV]) and by the combination of PCO and ozone (O3) (TiO2/UV/O3) was investigated to evaluate the enhancement effect of O3. The degradation of d-limonene by UV/O3 was also investigated for comparison. The experiments were conducted with a quartz photoreactor under various gas flow rates (600–1600 mL min-1), d-limonene concentrations (0.5–9 parts per million [ppm]), and relative humidity (RH) (20–80%). The d-limonene removal efficiency of TiO2/UV/O3, TiO2/UV, and UV/O3 ranged from 62 to 99%, from 49 to 99%, and from 46 to 75%, respectively. The addition of 120-ppb O3 can enhance the d-limonene removal efficiency of PCO up to 12%. The apparent kinetic parameters (apparent rate constants, k apparent and Langmuir adsorption constants, K apparent) of TiO2/UV and TiO2/UV/O3 reactions obtained from fitting Langmuir–Hinshelwood models are TiO2/UV: k apparent = 1.45 × 10−3 ppm-m sec−1, K apparent = 0.34 ppm−1; TiO2/UV/O3: k apparent = 1.83 × 10−3 ppm-m sec−1, and K apparent = 0.35 ppm−1. When RH was higher than 40%, the residual intermediates yield rates of d-limonene of TiO2/UV/O3, TiO2/UV, and UV/O3 reactions ranged from 0.39 to 0.51 μmol carbon m−2 sec−1, 0.56 to 1.96 μmol carbon m−2 sec−1, and 157 to 177 μmol carbon m−3 sec−1, respectively. In the photocatalytic reaction experiments, the addition of 120-parts per billion (ppb) O3 can reduce the residual intermediates yield rates of d-limonene by up to 1.46 μmol carbon m−2 sec−1. These experimental results showed that O3 can enhance the effectiveness of photo-catalysis on the removal of d-limonene.

Optimal seed recipe design for crystal size distribution control for batch cooling crystallisation processes

The paper presents a novel quality-by-design framework for the design of optimal seed recipes for batch cooling crystallisation systems with the aim to produce a desired target crystal size distribution (CSD) of the product. The approach is based on a population balance model-based optimal control framework, which optimises the compositions of seed blends, based on seed fractions that result from standard sieve analysis. The population balance model is solved using a combined quadrature method of moments and method of characteristics (QMOM-MOCH) approach for the generic case of apparent size-dependent growth. Seed mixtures are represented as a sum of Gaussian distributions, where each Gaussian corresponds to the seed distribution in a particular sieve size range. The proposed methods are exemplified for the model system of potassium dichromate in water, for which the apparent size-dependent growth kinetic parameters have been identified from laboratory experiments. The paper also illustrates the simultaneous application of in situ process analytical tools, such as focused beam reflectance measurement (FBRM) for nucleation detection, attenuated total reflection (ATR) UV/Vis spectroscopy for concentration monitoring, as well as the in-line use of laser diffraction particle sizing for real-time CSD measurement.

Assessment of Selected Apparent Kinetic Parameters of the HDM and HDS Reactions of Two Kuwaiti Residual Oils, Using Two Types of Commercial ARDS Catalysts

Selected apparent kinetic parameters such as rate constants and activation energies of the hydrodemetallation (HDM) and hydrodesulfurization (HDS) reactions were assessed for two Kuwaiti Residual oils using two types of commercial atmospheric residual desulfurization (ARDS) catalysts. The feeds investigated in the study were 360 °C+ cuts of Kuwait Export crude (KEC) and Kuwait Ratawi/Burgan crude (KHC). The catalysts used in the study were two different catalysts of a complete ARDS catalyst system (Cat-B) consisting of five different catalysts. Cat-BA is an HDM catalyst used in the front reactor of an industrial ARDS process to remove possible contaminants present in the feed, such as metals, sulfur, nitrogen, and to reduce the feed asphaltene content to lowest levels possible. Cat-BC is an HDS catalyst, usually employed in the middle reactors, mainly to reduce the feed sulfur content to the desired levels. Testing was performed using a bench scale unit that was equipped with three fixed-bed reactors work...

Compensation effect and volcano curve in toluene hydrogenation catalyzed by transition metal sulfides

Within the framework of volcano curves, a kinetic study of toluene hydrogenation catalyzed by transition metal sulfides highlights the variation of the apparent kinetic parameters as a function of the ab initio sulfur-metal bond energy descriptor and sulfo-reductive reaction conditions.

Analysis of Unstirred Water Layer in In Vitro Permeability Experiments

In vitro permeability experiments are used widely in drug discovery and other areas of pharmaceutical research. Much effort has been expended in developing novel epithelial models but generally much less attention has been paid to the hydrodynamic barrier in the actual experiments. The restricted liquid flow in the vicinity of solid surfaces leads to a zone where the diffusional movement of molecules exceeds the convection. This leads to formation of a concentration gradient between the bulk solution and the surface. The formed unstirred water layer (UWL) reduces the apparent permeability (Papp) of compounds that rapidly pass through the actual epithelial layer. This lowers the resolution of Papp versus fraction-absorbed assay, complicates the structure-permeability analysis and skews apparent kinetic parameters of transporter substrates. This review describes the techniques that can be used to determine the UWL thickness in permeability experiments and apparatuses described in the literature to control the in vitro hydrodynamics. © 2009 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 98:4469–4479, 2009

Activity modulation and reusability of β- d-galactosidase confined in sol–gel derived porous silicate glass

In the present work we applied the sol–gel method to obtain glass lentils entrapping β- d-galactosidase (β-Gal) (E β-Gal) within a silicate matrix. The effect of pH, temperature, polarity and salt concentration on the activity of E β-Gal was studied. Apparent kinetic parameters for ortho-nitro-phenyl-β- d-galactopyranoside hydrolysis catalyzed by E β-Gal ( V ′ max , K ′ M ) were lower compared to the soluble enzyme (S β-Gal), reflecting the solute diffusion restriction imposed by the matrix observed in the time curves, a partial protein inactivation upon encapsulation, and an improvement in the affinity of E β-Gal for the substrate as compared with S β-Gal. At pH < 4, E β-Gal stability was higher than that of S β-Gal. E β-Gal could be reused after storage at 4 °C for up to 90 days, and retained its activity profile within the range of pH = 2–10 and saline concentration 0–400 mM. Pre-incubation at 75 °C for 30 min fully inactivated S β-Gal while E β-Gal retained approximately 90% of its activity, even in the reused samples. Encapsulation did not introduce additional impairments to the reaction rate measured in heterogeneous dispersions, beyond those derived from their own particle-crowded environment. This reusable E β-Gal was resistant to typical technological conditions applied in milk processing that would lead to the unfolding and inactivation of S β-Gal. The results are discussed from the biophysical viewpoint.

The conserved Lysine69 residue plays a catalytic role in Mycobacterium tuberculosis shikimate dehydrogenase

BackgroundThe shikimate pathway is an attractive target for the development of antitubercular agents because it is essential in Mycobacterium tuberculosis, the causative agent of tuberculosis, but absent in humans. M. tuberculosis aroE-encoded shikimate dehydrogenase catalyzes the forth reaction in the shikimate pathway. Structural and functional studies indicate that Lysine69 may be involved in catalysis and/or substrate binding in M. tuberculosis shikimate dehydrogenase. Investigation of the kinetic properties of mutant enzymes can bring important insights about the role of amino acid residues for M. tuberculosis shikimate dehydrogenase.FindingsWe have performed site-directed mutagenesis, steady-state kinetics, equilibrium binding measurements and molecular modeling for both the wild-type M. tuberculosis shikimate dehydrogenase and the K69A mutant enzymes. The apparent steady-state kinetic parameters for the M. tuberculosis shikimate dehydrogenase were determined; the catalytic constant value for the wild-type enzyme (50 s-1) is 68-fold larger than that for the mutant K69A (0.73 s-1). There was a modest increase in the Michaelis-Menten constant for DHS (K69A = 76 μM; wild-type = 29 μM) and NADPH (K69A = 30 μM; wild-type = 11 μM). The equilibrium dissociation constants for wild-type and K69A mutant enzymes are 32 (± 4) μM and 134 (± 21), respectively.ConclusionOur results show that the residue Lysine69 plays a catalytic role and is not involved in substrate binding for the M. tuberculosis shikimate dehydrogenase. These efforts on M. tuberculosis shikimate dehydrogenase catalytic mechanism determination should help the rational design of specific inhibitors, aiming at the development of antitubercular drugs.

Extraction of hemoglobin with calixarenes and biocatalysis in organic media of the complex with pseudoactivity of peroxidase

The present work involves the use of p- tert-butylcalix[4,6,8]arene carboxylic acid derivatives ( tButyl[4,6,8]CH 2COOH) for selective extraction of hemoglobin. All three calixarenes extracted hemoglobin into the organic phase, exhibiting extraction parameters higher than 0.90. Evaluation of the solvent accessible positively charged amino acid side chains of hemoglobin (PDB entry 1XZ2) revealed that there are 8 arginine, 44 lysine and 30 histidine residues on the protein surface which may be involved in the interactions with the calixarene molecules. The hemoglobin– tButyl[6]CH 2COOH complex had pseudoperoxidase activity which catalysed the oxidation of syringaldazine in the presence of hydrogen peroxide in organic medium containing chloroform. The effect of pH, protein and substrate concentrations on biocatalysis was investigated using the hemoglobin– tButyl[6]CH 2COOH complex. This complex exhibited the highest specific activity of 9.92 × 10 −2 U mg protein −1 at an initial pH of 7.5 in organic medium. Apparent kinetic parameters ( V′ max, K′ m, k′ cat and k′ cat/ K′ m) for the pseudoperoxidase activity were determined in organic media for different pH values from a Michaelis–Menten plot. Furthermore, the stability of the protein–calixarene complex was investigated for different initial pH values and half-life ( t 1/2) values were obtained in the range of 1.96 and 2.64 days. Hemoglobin–calixarene complex present in organic medium was recovered in fresh aqueous solutions at alkaline pH, with a recovery of pseudoperoxidase activity of over 100%. These results strongly suggest that the use of calixarene derivatives is an alternative technique for protein extraction and solubilisation in organic media for biocatalysis.

Plasticity of laccase generated by homeologous recombination in yeast

Laccase-encoding sequences sharing 65-71% identity were shuffledin vivo by homeologous recombination. Yeast efficiently repaired linearized plasmids containing clac1, clac2 or clac5 Trametes sp. C30 cDNAs using a clac3 PCR fragment. From transformants secreting active variants, three chimeric laccases (LAC131, LAC232 and LAC535), each resulting from double crossovers, were purified, and their apparent kinetic parameters were determined using 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid) and syringaldazine (SGZ) as substrates. At acidic pH, the apparent kinetic parameters of the chimera were not distinguishable from each other or from those obtained for the LAC3 enzyme used as reference. On the other hand, the pH tolerance of the variants was visibly extended towards alkaline pH values. Compared to the parental LAC3, a 31-fold increase in apparent k(cat) was observed for LAC131 at pH 8. This factor is one of the highest ever observed for laccase in a single mutagenesis step.

Characterization of a novel meta-fission product hydrolase from Dyella ginsengisoli LA-4

A novel meta-fission product hydrolase gene ( mfphA) located in the bphX gene region of Dyella ginsengisoli LA-4 was successfully cloned and heterologously expressed in this study. The deduced amino acid sequence of MfphA showed 75% identity with the sequence of 2-hydroxymuconic semialdehyde hydrolase from Pseudomonas putida UCC22. The results suggested that MfphA belonged to the α/β hydrolase family, which could hydrolyze the meta-fission products (MFPs) during the biodegradation process of monocyclic compounds. The His-Tag MfphA was purified, and the subunit molecular mass of MfphA was about 35.3 kDa by SDS-PAGE analysis. According to the apparent kinetic parameters, the specificity of MfphA was determined in the following order: 6-methyl-HODA > HODA > 5-methyl-HODA > 6-phenyl-HODA > 5-chloro-HODA (HODA, 2-hydroxy-6-oxohexa-2,4-dienoate). The activity of MfphA for HODA was higher than that for 6-methyl-HODA. It was shown that the maximum activity was at 70 °C, and MfphA was stable for storage at low temperature for 30 days.

N-Heptane isomerization over Pt/WO 3-ZrO 2: A kinetic study

Kinetics of n-C 7 isomerization was investigated over Pt loaded WO 3-ZrO 2 using a fixed-bed reactor under hydrogen pressure and the effect of reaction conditions on the activity of the catalyst towards n-C 7 isomerization and product selectivity was studied. The study shows that the cracked products appear mainly from di-branched isomers of heptane. By controlling the reaction conditions, the octane number of the feed was increased from 0 to 32 with 93% fuel volume recovery. The major product among the mono-branched isomers was 3-methyhexane while 2,3-dimethylpentane was dominant among di-branched isomers. Craking products consisted mainly of propane and isobutane which were produced from dimethylpentanes. Simplified kinetic models were developed based on the reaction conditions. The apparent kinetic parameters for mono-isomerization (M), di-isomerization (D) and cracking (C) reactions were estimated using nonlinear regression analysis. The apparent activation energies were found to decrease as follows: E C > E M > E D.

Biological decolorization of dye solution containing malachite green by Pandoraea pulmonicola YC32 using a batch and continuous system

In our study, we have isolated a relatively newly identified bacteria species, Pandoraea pulmonicola YC32, and first assessed its capability to treat malachite green (MG). The effects of various factors on decolorization efficiency were investigated in a batch system. The decolorization efficiency was found to be optimal within a pH of 7–10 and it increased, with increasing initial MG concentration up to 100 mg/l. The relationship between the decolorization rate and MG concentration agreed with Lineweaver–Burk equation. The apparent kinetic parameters, R MG,max and K m, were 6.23 mg-MG/g-cell/h and 153.4 mg/l, respectively. The initial step in the biodegradation pathway of MG by P. pulmonicola YC32 was a reduction or N-demethylation reaction. We achieved a decolorization efficiency of 85.2% with 50 mg/l MG in the immobilized P. pulmonicola YC32 continuous column system. This is the first report on the application of a continuous column system to decolorize MG using a microorganism.