Immobilization of polyphenol oxidase on carboxymethylcellulose hydrogel beads: preparation and characterization

Carboxymethylcellulose (CMC) beads were prepared by a liquid curing method in the presence of trivalent ferric ions, and epicholorohydrin was covalently attached to the CMC beads. Polyphenol oxidase (PPO) was then covalently immobilized onto CMC beads. The enzyme loading was 603 µg g⁻¹ bead and the retained activity of the immobilized enzyme was found to be 44%. The K_m values were 0.65 and 0.87 mM for the free and the immobilized enzyme, and the V_max values were found to be 1890 and 760 U mg⁻¹ for the free and the immobilized enzyme, respectively. The optimum pH was 6.5 for the free and 7.0 for the immobilized enzyme. The optimum reaction temperature for the free enzyme was 40 °C and for the immobilized enzyme was 45 °C. Immobilization onto CMC hydrogel beads made PPO more stable to heat and storage, implying that the covalent immobilization imparted higher conformational stability to the enzyme. © 2000 Society of Chemical Industry     

Invertase immobilized on spacer‐arm attached poly(hydroxyethyl methacrylate) membrane: Preparation and properties

Microporous poly(2‐hydroxyethyl methacrylate) (pHEMA) membrane was prepared by UV‐initiated photopolymerization. The spacer arm (i.e., hexamethylene diamine) was attached covalently and then invertase was immobilized by the condensation reaction of the amino groups of the spacer arm with carboxyl groups of the enzyme in the presence of carbodiimides. The values of the Michael's constant K_m of invertase were significantly larger (ca. 2.5 times) upon immobilization, indicating decreased affinity by the enzyme for its substrate, whereas V_max was smaller for the immobilized invertase. Immobilization improved the pH stability of the enzyme as well as its temperature stability. Thermal stability was found to increase with immobilization and at 70°C the half times for the activity decay were 12 min for the free enzyme and 41 min for the immobilized enzyme. The immobilized enzyme activity was found to be quite stable in repeated experiments. © 2000 John Wiley & Sons, Inc. J Appl Polym Sci 75: 1685–1692, 2000     

In vitro cadmium removal from human serum by Cibacron Blue F3GA–thionein‐complex conjugated affinity membranes

A new membrane affinity biosorbent carrying thionein has been developed for selective removal of cadmium ions from human serum. Microporous poly(2‐hydroxyethyl methacrylate) (pHEMA) membranes were prepared by photopolymerization of HEMA. The pseudo dye ligand Cibacron Blue F3GA (CB) was covalently immobilized on the pHEMA membranes. Then, the cysteine‐rich metallopeptide thionein was conjugated onto the CB‐immobilized membrane. The maximum amounts of CB immobilized and thionein conjugated on the membranes were 1.07 µmol cm⁻² and 0.92 µmol cm⁻², respectively. The hydrophilic pHEMA membrane had a swelling ratio of 58% (w/w) with a contact angle of 45.8 °. CB‐immobilized and CB‐immobilized–thionein‐conjugated membranes were used in the Cd(II) removal studies. Cd(II) ion adsorption appeared to reach equilibrium within 30 min and to follow a typical Langmuir adsorption isotherm. The maximum capacity (q _m) of the CB‐immobilized membranes was 0.203 (µmol Cd(II)) cm⁻² membrane and increased to 1.48 (µmol Cd(II)) cm⁻² upon CB–thionein‐complex conjugation. The pHEMA membranes retained their cadmium adsorption capacity even after 10 cycles of repeated use. © 2000 Society of Chemical Industry     

Dye derived and metal incorporated affinity poly(2-hydroxyethyl methacrylate) membranes for use in enzyme immobilization

Microporous poly(2-hydroxyethyl methacrylate) (PHEMA) membranes were prepared by UV-initiated photopolymerization of HEMA in the presence of an initiator (α,α′-azobisisobutyronitrile, AIBN). An affinity dye Cibacron Blue F3GA (CB) was attached covalently and then Fe³⁺ ions incorporated. The PHEMA-CB and PHEMA-CB-Fe³⁺ membranes derived were used for adsorption of glucose oxidase (GOD). The adsorption capacities of these membranes were determined under conditions of different pH and with different concentrations of the adsorbate in the medium. The adsorption phenomena appeared to follow a typical Langmuir isotherm. The glucose oxidase adsorption capacity of the Fe³⁺ incorporated membrane (87μgcm^-2) was greater than that of the dye-derived membrane (66μgcm^-2). Non-specific adsorption of the glucose oxidase on the PHEMA membranes was negligible. The K_m values for both immobilized glucose oxidase PHEMA-CB-GOD (8·3) and PHEMA-CB-Fe³⁺-GOD (7·6) were higher than that of the free enzyme (6·2mM). Optimum reaction pH was 5·5 for the free and 6·0 for both immobilized preparations. The optimum reaction temperature of the adsorbed enzymes was 5°C higher than that of the free enzyme and was significantly broader. After 15 successive uses the retained activity of the adsorbed enzyme was 87%. It was observed that enzymes could be repeatedly adsorbed and desorbed on the derived PHEMA membranes without significant loss in adsorption capacity or enzymic activity. © 1998 SCI.     

Universal sequential outlier hypothesis testing

Universal outlier hypothesis testing is studied in a sequential setting. Multiple observation sequences are collected, a small subset of which are outliers. A sequence is considered an outlier if the observations in that sequence are generated by an “outlier” distribution, distinct from a common “typical” distribution governing the majority of the sequences. Apart from being distinct, the outlier and typical distributions can be arbitrarily close. The goal is to design a universal test to best discern all the outlier sequences. A universal test with the flavor of the repeated significance test is proposed and its asymptotic performance, as the error probability goes to zero, is characterized under various universal settings. The proposed test is shown to be universally consistent. For the model with at most one outlier, conditioned on the outlier being present, the test is shown to be asymptotically optimal universally when the typical distribution is known and as the number of sequences goes to infinity when neither the outlier nor the typical distribution is known. With multiple identical outliers, the test is shown to be asymptotically optimal universally when the number of outliers is the largest possible and with the typical distribution being known, and its asymptotic performance with neither the outlier nor the typical distribution being known is also characterized. Extensions of the findings to models with multiple distinct outliers are also discussed. In all cases, it is shown that the asymptotic performance guarantees for the proposed test when neither the outlier nor the typical distribution is known converge to those when the typical distribution is known as the number of sequences goes to infinity.     

Evaluation of performance characteristics of the heavy vehicle simulator in Florida

The Florida Department of Transportation (FDOT) Materials Office has recently acquired a heavy vehicle simulator (HVS) and constructed an accelerated pavement testing (APT) facility which uses this HVS. An investigation was conducted to evaluate the operational performance of the HVS, and to determine its most effective test configurations for use in evaluating the rutting performance of pavement materials and/or designs under typical Florida traffic and climate conditions. Five trial runs with the HVS used a super single tire with a load of 4082 kg, tire pressure of 793 kPa and a wheel traveling speed of 12.9 km/h. These five trial runs used different combinations of wheel traveling direction (uni-directional or bi-directional), total wheel wander and wander increments. The uni-directional loading was found to be a more efficient mode for evaluation of rutting performance using the HVS. As compared with the bi-directional loading mode, the uni-directional mode produced substantially higher rut depths for the same number of wheel passes and also for the same testing time duration. When the bi-directional loading with no wander was used, imprints of the tire treads were observed on the wheel track. It was found that using a loading mode with wander smoothened out the imprints of the tire treads considerably. The uni-directional loading mode with 10 cm wander using 2.5 cm increments was selected to be used for evaluation of rutting performance based on consideration of testing efficiency and realistic rutting results.     

The investigation of coal-pyrite/lignite concentration and their separation in the artificial mixture by oil agglomeration

In this study, the concentration of coal-pyrite and lignite taken from Yozgat-Ayridam (Turkey) Coal Management was investigated by oil agglomeration.In the previous studies, the agglomeration of coal-pyrite was investigated using different bridging liquids (fuel oil, diesel oil and kerosene) and the combination of reagent (KEX, Acorga M5397)+kerosene. When using only bridging liquids, the agglomeration recovery of pyrite was very low. To increase the hydrophobicity of pyrite, KEX was used. However, the pyrite was not agglomerated with an acceptable recovery. But when using Acorga M5397, which is a chelating reagent, the agglomeration recovery of pyrite was increased. The pyrite was agglomerated with a recovery of 76.70 wt% by single-stage agglomeration.In the optimum conditions which were determined for coal-pyrite, the agglomeration recovery of lignite was investigated. It was found that the lignite could not be agglomerated with an acceptable recovery.To investigate the separation of coal-pyrite and lignite, the artificial mixture of coal-pyrite and lignite was prepared with the weight ratio of 1/4 of coal-pyrite and lignite. It was found that the pyrite could be agglomerated at a recovery of 96.54% with three-stage agglomeration process. The lignite concentrate was produced with a recovery of 73.96 wt% and the pyrite content of 0.86 wt%.These findings showed that the coal-pyrite and lignite could be separated by oil agglomeration using appropriate reagent and bridging liquid.     

Feature extraction for ECG heartbeats using higher order statistics of WPD coefficients

This paper describes feature extraction methods using higher order statistics (HOS) of wavelet packet decomposition (WPD) coefficients for the purpose of automatic heartbeat recognition. The method consists of three stages. First, the wavelet package coefficients (WPC) are calculated for each different type of ECG beat. Then, higher order statistics of WPC are derived. Finally, the obtained feature set is used as input to a classifier, which is based on k-NN algorithm. The MIT-BIH arrhythmia database is used to obtain the ECG records used in this study. All heartbeats in the arrhythmia database are grouped into five main heartbeat classes. The classification accuracy of the proposed system is measured by average sensitivity of 90%, average selectivity of 92% and average specificity of 98%. The results show that HOS of WPC as features are highly discriminative for the classification of different arrhythmic ECG beats.     

Enzymatic removal of phenol and p-chlorophenol in enzyme reactor: horseradish peroxidase immobilized on magnetic beads

Horseradish peroxidase was immobilized on the magnetic poly(glycidylmethacrylate-co-methylmethacrylate) (poly(GMA-MMA)), via covalent bonding and used for the treatment of phenolic wastewater in continuous systems. For this purposes, horseradish peroxidase (HRP) was covalently immobilized onto magnetic poly(GMA-MMA) beds using glutaraldehyde (GA) as a coupling agent. The maximum HRP immobilization capacity of the magnetic poly(GMA-MMA)-GA beads was 3.35 mg g(-1). The immobilized HRP retained 79% of the activity of the free HRP used for immobilization. The immobilized HRP was used for the removal of phenol and p-chlorophenol via polymerization of dissolved phenols in the presence of hydrogen peroxide (H(2)O(2)). The effect of pH and temperature on the phenol oxidation rate was investigated. The results were compared with the free HRP, which showed that the optimum pH value for the immobilized HRP is similar to that for the free HRP. The optimum pH value for free and immobilized HRP was observed at pH 7.0. The optimum temperature for phenols oxidation with immobilized HRP was between 25 and 35 degrees C and the immobilized HRP has more resistance to temperature inactivation than that of the free form. Finally, the immobilized HRP was operated in a magnetically stabilized fluidized bed reactor, and phenols were successfully removed in the enzyme reactor.