Application of magnetic immobilized microorganisms

Magnetic calcium alginate yeast beads, made by incorporation of magnetite or colloidal magnetic liquid, Ferrofluidℳ, exhibited catalytic behavior similar to that of their nonmagnetic counterparts. The magnetic immobilized preparations’ shortterm performance, long-term operational stability, and capacity forin situ activation were unaffected by the inclusion of magnetic material. The magnetic quality of the alginate beads provides manipulatory advantages.     

Application of factorial design to study of heavy metals biosorption by waste biomass from beverage distillery

A full factorial design leading to 20 sets of sorption runs was conducted to study the influence of four variables (bleaching earth and biomass concentrations, pH, and sorption time) on the iron, nickel, and chromium removal from stainless steel effluent using waste biomass from a beverage industry. Similar factor effects and interactions were found for each metalinvolved in this biosorption study, and the main factors were pH (positive effect) and biomass concentration (negative effect). Response surface methodology was adopted and an empirical linear polynomial model constructed on the basis of the specific uptake (mg of metal/g of biomass as dry weight) for each metal species. Under optimized process conditions (pH 4.0, biomass concentration of 2.0 g/L, absence of Celite), uptake values of 155 mg of Fe/g, 38 mg of Cr/g, and 0.4 mg of Ni/g were achieved after 3 h. This corresponded to a reduction in heavy metals concentration of approx 94% for Cr, 57% for Fe, and 25% for Ni.     

Use of bioluminescent bacterial sensors as an alternative method for measuring heavy metals in soil extracts

The luminescence based bacterial sensor strains Pseudomonas fluorescens OS8 (pTPT11) for mercury detection and Pseudomonas fluorescens OS8 (pTPT31) for arsenite detection were used in testing their application in detecting heavy metals in soil extracts. Three different soil types (humus, mineral and clay) were spiked with 1, 100 or 500 μg g⁻¹ Hg²⁺ or As³⁺. Samples were taken 1, 14 and 30 days and extracted with water, ammonium acetate, hydrogen peroxide and nitric acid to represent water soluble, bioavailable, organic matter bound and residual fractions, respectively. The lowest mercury-concentration measured using biosensor (0.003 μg kg⁻¹) was considerably lower than by chemical method (0.05 μg kg⁻¹). The sensor strain with pTPT31 appeared to have a useful detection range similar to that of chemical methods. Concentration results with chemical and biosensor analysis were very similar in the case of mercury-spiked samples. Although some of the arsenite samples showed higher variation between methods, it is concluded that the bacteria can be used as an alternative traditional methods for different types of samples.     

Continuous production of butanol with immobilized cells ofClostridium acetobutylicum

Spores ofClostridium acetobutylicum were immobilized in calcium alginate. An active gel preparation was obtained after outgrowth of the spores to vegetative cells within the gel matrix. A 100 mL column containing the immobilized cells was used for continuous production. At steady-state conditions the productivity of butanol was 67 g/L reactor volume/day.     

Heavy metal removal by biosorption using Phanerochaete chrysosporium

Biosorption using microbial cells as adsorbents is being seen as a cost-effective method for the removal of heavy metals from wastewaters. Biosorption studies with Phanerochaete chrysosporium were performed for copper (II), lead (II), and cadmium (II) to evaluate the effectiveness and to optimize the operational parameters using response surface methodology. The operational parameters chosen were initial metal ion concentration, pH, and biosorbent dosage. Using this method, the metal removal could be correlated to the operational parameters, and their values were optimized. The results showed fairly high adsorptive capacities for all the metals within the settings of the operational parameters. The removal efficiencies followed the order Pb>Cu>Cd. As a general trend, metal removal efficiency decreased as the initial metal ion concentration increased, and the results fitted the Langmuir and Freundlich isotherms well.     

Biosorption of heavy metal ions by immobilized zoogloea and zooglan

ImmobilizedZoogloea and zooglan in calcium alginate-silica matrix was shown to have a high adsorption capacity for Cu and Cd ions. Our results showed that Cu-ion uptake in the presence of Ca and Mg ions can be enhanced using immobilizedZoogloea and zooglan. Heavy metal ion adsorption efficiency decreased in the following order: Cu > Cd > Zn > Cr. The adsorbed metal ions were desorbed completely using sulfuric acid. ImmobilizedZoogloea and zooglan which was repetitively regenerated adsorbed heavy metal ions without a loss of adsorption capacity.     

Reasons for the high stability of fumarase activity ofBrevibacterium flavum cells immobilized with k-carrageenan gel

Whole cells ofBrevibacterium flavum having high fumarase activity were immobilized using K-carrageenan. The reason for the high stability of fumarase activity of immobilized cells was investigated. One of main reasons for stabilizing fumarase activity by immobilization using K-carrageenan against organic solvents such as ethanol and acetone was the lower concentration of these solvents in the carrageenan gel compared with that in outer bulk solution. The stabilization of fumarase activity in the immobilized cells against protein-denaturing reagents was found to be related to rheological properties of K-carrageenan gel. Another reason for stabilizing fumarase activity by immobilization with K-carrageenan was to protect the cells from lysis. When immobilized cells were freeze-thawed, their fumarase activity increased and operation stability decreased. Therefore, one reason for the high decay of fumarase activity caused by the freeze-thawing may be a change in the pore size of the K-carrageenan gel. Fumarase activity and the operational stability of immobilized cells was found to depend on gelling conditions. Therefore, the steric structure of the K-carrageenan gel may be related to the decay of fumarase activity.     

Immobilized cells in microbial nitrate reduction

The microorganismPseudomonas denitrificans was immobilized in alginate. These immobilized cells were capable of reducing 0.8 mg NO ₃ ^- /min/g wet weight of cells.     

Determination of phenol and catechol concentrations with oxygen probes coated with immobilized enzymes or immobilized cells

The enzyme phenol 2-hydroxylase was immobilized on Sepharose and used in conjunction with an O₂ electrode for quantitating phenol. Similarly, catechol 1,2-oxygenase was used for quantitating catechol. A third probe was prepared by immobilization ofTrichosporon cutaneum cells rather than purified phenol 2-hydroxylase for phenol quantitation. The whole cell system gave results comparable to the immobilized enzyme system.     

Affinity chromatographic purification of lentil lectin using immobilized yeast cells

A model system for evaluating macroporous supports containing immobilized whole cells in affinity Chromatographic applications is described. Whole cells were immobilized in a polyacrylamide network in a two-step polymerization process. The affinity system discussed consists of immobilized cells ofSaccharomyces cervisiae in the purification of lentil lectin fromLens culinaris.     

Protection of immobilized sulfhydryl groups against autooxidation by alterations in their microenvironment

Immobilized sulfhydryl groups were prepared by partial thiolation of NH₂-glass beads. The microenvironment of the immobilized SH groups was varied by different chemical modifications of neighboring NH₂ groups. Introduction of a strong charge in the surroundings of immobilized sulfhydryls results in their dramatic stabilization against autooxidation. This effect is due to the salting of O₂ from the surface microlayer of the thiolated beads.     

Stabilization of fumarase activity ofBrevibacterium flavum cells by immobilization with k-carrageenan

Whole cells ofBrevibacterium flavum having fumarase activity were immobilized using K-carrageenan. The stabilities of fumarase activity in the immobilized cells against external factors, including heat, pH, organic solvents, and protein denaturing reagents, were compared with those of free cells and native enzyme. The stabilities of fumarase activity in immobillized cells against external factors were highest, and those of native enzyme were lowest. In the “gel-state,” K-carrageenan-immobilized cells showed a much higher stabilization effect for external factors than “sol-state” immobilized cells.     

The influence of products and by-products obtained by drinking water electrolysis on microorganisms

It is the aim of this work to underline the chemical character of disinfection processes and to present the results of treatment experiments for artificially and electrochemically added chemicals focusing on chlorine components and hydrogen peroxide obtained on IrO₂/RuO₂ anodes. A discontinuous cell with rotating anode placed 4 mm above an IrO₂ expanded mesh cathode of same diameter was used for the generation of electrolysis products. The thermostated experiments were carried out at a temperature of 20 °C and in constant current mode. Artificial solutions prepared using sodium salts and deionised water were electrolysed. Samples were analysed using the DPD (N, N-diethyl-p-phenylendiamine) method. Escherichia coli K-12, Bacillus subtilis DSM 2277 and Saccharomyces cerevisiae Kolin were used as test-microorganisms. Morphological changes were studied using transmission electron microscopy. The results show that electrolysed water had a higher lethal efficiency than Ca(OCl)₂ of the same measured active chlorine concentration. During the killing period mainly inner cell parts of the microorganisms react chemically with the disinfectants. In sufficiently high concentrations and reaction times, the cell material continues to react. Cell walls or membranes can break-through as electron microscopy pictures show. Electrolysed water had a higher lethal efficiency on microorganisms. This behaviour demonstrates the existence of additional by-products.     

Citric acid production with alginate bead entrappedAspergillus niger ATCC 9142

Aspergillus niger ATCC 9142 mycelium was entrapped in calcium alginate beads and employed in an air-lift completely stirred reactor for continuous production of citric acid. Maximum yield obtained from 10% (w/v) sucrose was 12 g dm^-3 with about 40% fermentation efficiency. Maximum rate of production 70 mg g^-1 h^-1 was about five times that obtained in classical batch fermentation.     

Pilot-plant for NOx, SO2 and HC removal from flue-gas of municipal waste incinerator by electron beam irradiation

According to the basic research performed using a small size reactor at TRCRE of JAERI, the electron beam irradiation process was proved to be very effective for NO_x, SO₂ and HC removals from flue-gas of municipal waste incinerators. Based on this result, a pilot-plant was constructed for the demonstration of NO_x, SO₂ and HC removal performance using electron accelerator of maximum energy 0.95 MeV and maximum power 15kW.The pilot-plant was constructed at Matsudo City waste Disposal Center. The flue-gas of 1,000 Nm³/hr is guided from the waste incinerator flue-gas line of 30,000Nm³/hr to the pilot-plant to be processed by spraying Ca(OH)₂ slurry or powder and irradiation with high-energy electron beam of the accelerator. NO_x, SO₂ and HC are removed simultaneously from the flue-gas by the enhanced reaction with Ca (OH)₂ under irradiation.A brief explanation of the pilot-plant and preliminary results of the experiments are introduced in this paper.     

Entrapment of living microbial cells in covalent polymeric networks

The kinetics of oxidative phenol degradation with microbial cellsCandida tropicalis, immobilized in a polyacrylamide and polymethacrylamide matrix, were mathematically simulated assuming zero-order and Michaelis-Menten rate equations. For zero-order kinetics an expanded equation for catalytic effectiveness as a function of the Thiele modulus, Biot number, and partition coefficients was derived and compared with numerical solutions for Michaelis-Menten kinetics. Errors with regard to the zero-order approximation become negligible ifc _o/K _M >2. Experimentally determined catalyst activities as a function of particle size and cell concentration were compared to calculated ones. Additional experiments to determine the diffusion and oxygen consumption ratios have been carried out in an effort to resolve the physical parameters to be used in the above mentioned calculations. Furthermore, experiments on cell growth during reincubation with nutrients and oxygen are reported; an increase in activity up to a factor of ten was observed. These experiments demonstrate that the microbial cells are entrapped in the polymer matrix in the living state.     

Influence of the support on the reaction course of tributyrin hydrolysis catalyzed by soluble and immobilized lipases

Lipases from different origins have been immobilized in supports chosen by its different aquaphilicity and used as biocatalysts for the hydrolysis of tributyrin. The changes of the concentration of tri-, di-, monobutyrin, glycerol, and butyric acid during the reactions catalyzed by soluble, as well as immobilized, lipases were evaluated by gas chromatography. The experimental data were fitted to a simple kinetic model for the sequential reaction of tributyrin hydrolysis. The calculated apparent rate constants were different for the biocatalysts used and were apparently related to diffusional effects and aquaphilicity of the supports. Maximal yields of dibutyrin were found with the soluble Candida lipase, whereas the highest yield of monobutyrin (90%) was obtained with the least aquaphylic derivative (Candida-Celite).     

Immobilization of isolated and cellular hydrogenase ofD. desulfuricans in radiation polymerized polyacrylamides

Purified hydrogenase fromDesulfovibrio desulfuricans was immobilized either by entrapment or absorption onto porous neutral and charged acrylamide beads. Surface absorption and crosslinking on the beads resulted in a high hydrogenase activity and a good immobilization coefficient compared to the enzyme and whole cells entrapped in the same matrix. Maximum enzyme activity (citrate-phosphate buffer) was shifted to pH 6.5 upon immobilization in contrast to 6.0 for the free enzyme and the range of 6–7 for whole cells. Both the purified enzyme and whole cells were most active when held in neutral matrices. Immobilization improved the temperature stability (65‡C) and long term storage (4‡C) of the hydrogenase activity of both the purified enzyme and whole cells.     

Removal of chemical oxygen demand,nitrogen, and heavy metals using a sequenced anaerobic-aerobic treatment of landfill leachates at 10-30 degrees C

As a first step of treatment of landfill leachates (total chemical oxygen demand [COD]: 1.43–3.81 g/L; total nitrogen: 90–162 mg/L), performance of laboratory upflow anaerobic sludge bed reactors was investigated under mesophilic (30°C), submesophilic (20°C), and psychrophilic (10°C) conditions. Under hydraulic retention times (HRTs) of about 0.3 d, when the average organic loading rates (OLRs) were about 5 g of COD/(L·d), the total COD removal accounted for 81% (on average) with the effluent concentrations close to the anaerobic biodegradability limit (0.25 g of COD/L) for mesophilic and submesophilic regimes. The psychrophilic treatment conducted under an average HRT of 0.34 d and an average OLR of 4.22 g of ducted under an average HRT of 0.34 d and an average OLR of 4.22 g of COD/(L·d) showed a total COD removal of 47%, giving effluents (0.75 g of COD/L) more suitable for subsequent biologic nitrogen removal. All three anaerobic regimes used for leachate treatment were quite efficient for elimination of heavy metals (Fe, Zn, Cu, Pb, Cd) by concomitant precipitation in the form of insoluble sulfides inside the sludge bed. The application of aerobic/anoxic biofilter as a sole polishing step for psychrophilic anaerobic effluents was acceptable for elimination of biodegradable COD and nitrogen approaching the current standards for direct discharge of treated waste-water.     

A kinetic study on the bioremediation of sodium cyanide and acetonitrile by free and immobilized cells ofPseudomonas putida

Pseudomonas putida capable of utilizing organic nitrile (acetonitrile) and inorganic cyanide (sodium cyanide) as the sole source of carbon and nitrogen was isolated from contaminated industrial sites and waste water. The bacterium possesses nitrile aminohydrolase (EC 3.5.5.1) and amidase (EC 3.5.1.4), which are involved in the transformation of cyanides and nitriles into ammonia and CO₂ through the formation of amide as an intermediate. Both of the enzymes have a high selectivity and affinity toward the⁻Cn group. The rate of degradation of aceotnitrile and sodium cyanide to ammonia and CO₂ by the calcium-alginate immobilized cells ofP. putida was studied. The rate of reaction during the biodegradation of acetonitrile and sodium cyanide, and the substrate- and product-dependent kinetics of these toxic compounds were studied using free and immobilized cells ofP. putida and modeled using a simple Michaelis-Menten equation.