Carriers For Cell Immobilization Research Articles

Immobilization of Lactobacillus bulgaricus with gellan gum and its application in continuous fermentation of D-lactic acid from corn straw

Biorefinery of chemicals from straw is an effective approach to alleviate the environmental pollution caused by straw burning. In this paper, we prepared gellan gum immobilized Lactobacillus bulgaricus T15 gel beads (LA-GAGR-T15 gel beads), characterized their properties, and established a continuous cell recycle fermentation process for D-lactate (D-LA) production using the LA-GAGR-T15 gel beads. The fracture stress of LA-GAGR-T15 gel beads was (91.68±0.11) kPa, which was 125.12% higher than that of the calcium alginate immobilized T15 gel beads (calcium alginate-T15 gel beads). This indicated that the strength of LA-GAGR-T15 gel beads was stronger, and the strain was less likely to leak out. The average D-LA production was (72.90±2.79) g/L after fermentation for ten recycles (720 h) using LA-GAGR-T15 gel beads as the starting strain and glucose as the substrate, which was 33.85% higher than that of calcium alginate-T15 gel beads and 37.70% higher than that of free T15. Subsequently, glucose was replaced by enzymatically hydrolyzed corn straw and fermented for ten recycles (240 h) using LA-GAGR-T15 gel beads. The yield of D-LA reached (1.74±0.79) g/(L·h), which was much higher than that of using free bacteria. The wear rate of gel beads was less than 5% after ten recycles, which indicated that LA-GAGR is a good carrier for cell immobilization and can be widely used in industrial fermentation. This study provides basic data for the industrial production of D-LA using cell-recycled fermentation, and provides a new way for the biorefinery of D-LA from corn straw.

Repeated-Batch Ethanol Fermentation from Sweet Sorghum Stem Juice under a Very High Gravity Condition Using a Stirred Tank Bioreactor Coupled with a Column Bioreactor by Immobilized Saccharomyces cerevisiae

The ethanol fermentation efficiency of sweet sorghum stem juice (SSJ) under a very high gravity (VHG) condition (250 g/L of sugar) was improved by immobilized Saccharomyces cerevisiae SSJKKU01, using a stirred tank bioreactor (STR) coupled with a column bioreactor (CR). Dried rattan pieces (as carriers for cell immobilization) at 50% of the working volume of the CR were suitable for use in a batch ethanol fermentation. The average ethanol concentration (PE) and ethanol productivity (QP) of repeated-batch fermentation in the CR for eight successive cycles were 109.85 g/L and 1.88 g/L⋅h, respectively. Then an STR coupled with a CR was applied for repeated-batch ethanol fermentation in two systems. System I was an STR (1.8 L working volume), and System II was an STR (1 L) coupled with a CR, referred to as a CR-F (0.8 L). Both systems were connected to a new CR, called CR-I, containing sterile dried rattan pieces at 50% of its working volume. Active yeast cells were inoculated only into the STR, and the medium circulation rate between bioreactors was 5.2 mL/min. The results showed that at least eight successive cycles could be operated with an average PE of 108.51 g/L for System I and 109.44 g/L for System II. The average QP and SC values of both systems were also similar, with values of 1.87 to 1.88 g/L⋅h and 93 to 94%, respectively. The morphology of the carriers with and without immobilized cells before and after the fermentation was investigated. The obtained results demonstrated that a repeated-batch fermentation by immobilized cells on rattan pieces, using an STR coupled with a CR, was successfully used to produce high levels of ethanol from SSJ under a VHG condition.

An Alternative Approach to Improve the Butanol Production Efficiency from Sweet Sorghum Stem Juice Using Immobilized Cells Combined with an In Situ Gas Stripping System

The effects of the nitrogen source and buffers used in butanol production with Clostridium beijerinckii TISTR 1461 from sweet sorghum stem juice (SSJ) containing 60 g/L of total sugar were first studied in this paper. Among the various nitrogen sources (dried spent yeast, urea, ammonium acetate, ammonium sulfate), urea was found to be the most suitable for butanol production. SSJ supplemented with urea (0.64 g/L) and cocktail buffers (KH2PO4, 0.5 g/L; K2HPO4, 0.5 g/L; ammonium acetate, 2.2 g/L) gave the highest butanol concentration (PB, 10.13 g/L). Then, the capability of immobilized C. beijerinckii TISTR 1461 cells for butanol fermentation was investigated. Two residual waste materials were examined as immobilized cell carriers. Bamboo chopstick pieces were more appropriate as carriers for cell immobilization than cigarette filter tips. The PB value of the immobilized cells on the bamboo chopstick pieces was ~13% higher than that on the cigarette filter tips. Using the response surface methodology (RSM), 1.9 cm bamboo chopstick pieces with a carrier loading of 1:32 (w/v) were the optimum conditions for cell immobilization for butanol production. Under these conditions, the PB value was 11.62 g/L. To improve the butanol production efficiency, a gas stripping system (GS) was connected to the fermenter. It was found that the PB (14.02 g/L) and butanol productivity (QB, 0.29 g/L·h) values improved by ~21% compared to butanol fermentation using no gas stripping.

An overview on cell and enzyme immobilization for enhanced biohydrogen production from lignocellulosic biomass

Hydrogen gas, a carbon-free energy carrier, can be produced via fossil fuel reforming, coal gasification, water electrolysis, photocatalysis, or biological process. Biohydrogen production from lignocellulosic biomass (LCB) in this regard, appears as an environmental benign, sustainable, non-food competing second generation fuel. LCB serves as the largest potential carbon source for sustainable biohydrogen production with an enormous global annual capacity of more than one trillion tons. Enzymes in this case, are widely used to hydrolyse the LCB into fermentable sugars, and subsequent hydrogen production is carried out by dark fermentation. However, the untapped non-food competing LCB is currently impeded by several critical bottlenecks including sensitivity of cells and enzymes to numerous denaturing conditions, recyclability, and high cost of enzyme. Low productivity of hydrolysis and hydrogen production in this regard, lead to a larger bioreactor and capital expenditures (CAPEX) requirement, which in turn, making this approach to be less competitive in commercial application. These bottlenecks can be overcome by immobilization technique, which enables the recyclability, improves stability and productivity of the enzyme and cells. Current review accommodates for the important outlook and critical insights into the immobilization techniques, providing important guidelines for the operation of immobilization techniques to elevate commercial competitiveness of biohydrogen production from LCB in the future. The effect of geometry, surface charges, and wettability of different type of carriers for cell immobilization to enhance biohydrogen production are discussed. The critical aspects of the immobilization parameters, such as temperature, pH, and duration which could significantly affect the properties of immobilized enzymes are thoroughly examined in this review. Suggestions and future directions of this field are provided to assist the development of an efficient, economic, and sustainable hydrogen production process.

Valorization of sugar beet pulp through biotechnological approaches: recent developments.

Sugar beet pulp (SBP) is a valuable by-product of the sugar beet industry and is predominantly composed of cellulose, hemicellulose, and pectin. It is commonly used as livestock feed because of its palatability, good energy levels, and highly digestible fibers such as pectins and glucans. However, the utilization of SBP for the production of value-added products via biotechnological approaches is gaining significance in recent years owing to its potential as a cost-effective nutrient source and technological advancements in its processing. SBP can be used as a substrate for bio-production of microbial enzymes, single cell protein, alcohols (e.g., ethanol), methane/biogas, hydrogen, lactic acid, ferulic acid, and pectic oligosaccharides. SBP can also be used as a carrier for cell immobilization in fermentation processes. This review focused on recent developments in biotechnological valorization of SBP.

Capability of Immobilized Clostridium beijerinckii TISTR 1461 on Lotus Stalk Pieces to Produce Butanol from Sugarcane Molasses

Immobilized Clostridium beijerinckii TISTR 1461 was used to enhance the butanol production efficiency from sugarcane molasses. Lotus stalk (LS) pieces were used as carriers for cell immobilization. Sugarcane molasses containing 50 g/L of sugar supplemented with 1 g/L of yeast extract was found to be an appropriate medium for bacterial cell immobilization on the LS pieces. Carrier size (4, 12 and 20 mm in length) and carrier loading (1:15, 1:30 and 1:45, w/v) were optimized for high levels of butanol production using response surface methodology (RSM). The batch fermentation was carried out under anaerobic conditions in 1 L screw-capped bottles at 37 °C and an agitation rate of 150 rpm. It was found that the optimum conditions for the butanol production were the carrier size of 4 mm and carrier loading of 1:31 (w/v). Under these conditions, the butanol concentration (PB) was 12.89 g/L, corresponding to the butanol productivity (QB) of 0.36 g/L∙h and butanol yield (YB/S) of 0.36 g/g. These values were higher than those using free cells (PB, 10.20 g/L, QB, 0.28 g/L∙h and YB/S, 0.32 g/g). In addition, it was found that a 24 h incubation time for cell immobilization was appropriate for the immobilization process, which was confirmed by the results of the scanning electron microscope (SEM) and atomic force microscopy (AFM) images and specific surface area measurement. When the fermentation using the immobilized cells was carried out in a stirred-tank reactor (STR), column reactor (CR) and CR coupled with STR, the results showed that all reactors could be used to produce butanol production from the immobilized cells on LS pieces. However, the PB using CR and CR coupled with STR were only 75% and 45% of those using the screw-capped bottle and STR.

Lactic acid production on molasses enriched potato stillage by Lactobacillus paracasei immobilized onto agro-industrial waste supports

In this study, the production of lactic acid (LA) on sugar beet molasses enriched potato stillage by immobilized Lactobacillus paracasei NRRL B-4564 was examined. Three agro-industrial materials, such as sunflower seed hull (SSH), brewers’ spent grain (BSG), and sugar beet pulp (SBP) were studied as carriers for cell immobilization. The carriers were physically characterized in terms of water adsorption index (WAI), critical humidity point (CHP) and porosity. Further, the stability and efficiency of the immobilized biocatalysts were evaluated in repeated batch fermentation of molasses enriched potato stillage and compared with free cell system. A strong cell attachment onto agro-industrial supports allowed easy separation from the fermentation media and efficient biocatalyst reuse in five successive batch cycles. The highest cell number attached on the support surface during the fermentation was detected for material with higher WAI and lower CHP. Porosimetry measurements showed that the attachment of L. paracasei cells to support materials and overall LA productivities achieved with different immobilized biocatalysts were not determined by the support porosity and surface morphology, but with the material characteristics such as electrostatic charge, chemical composition and hydrophilicity. LA productivity of 1.48 g/L h, maximal LA concentration of 80.10 g/L and average yield coefficient of 0.97 g/g were achieved in fermentation of molasses enriched potato stillage using SBP as a support material, followed by BSG and SSH. The studied approach showed to be an interesting alternative strategy for increasing the LA productivities on low-cost and abundant substrate.

Production of butyric acid by Clostridium tyrobutyricum coated with carboxyl functioned magnetic nanoparticles

AbstractBACKGROUNDMagnetic nanomaterial has been considered a promising carrier for cell immobilization, because of its low mass transfer resistance, high load rate, high specific surface and facility for reusability. In this study, carboxyl functioned magnetic nanoparticles (CMNPs) were prepared by chemical co‐precipitation and oleate modification methods, and then immobilized with Clostridium tyrobutyricum for the production of butyric acid.RESULTSThe immobilization conditions were systematically optimized to achieve maximum efficiency and the highest cell relative activity, in which the mass ratio, immobilization temperature and time were determined as 0.72 g g‐1, 35 °C and 30 min, respectively. The endurance of CMNPs‐cells was also evaluated in a repeated‐batch mode of eight continuous batches. Stable and reliable production of butyric acid was obtained in each batch with an average butyrate titer and yield of ∼11.02 g L‐1 and ∼0.43 g g‐1 utilized sugar. Compared with the free‐cell fermentation, higher butyrate yield and concentration were obtained with immobilized cells of C. tyrobutyricum in repeated‐batch fermentation, suggesting that the coated cells did not experience a mass transfer problem.CONCLUSIONConsidering the convenient separation of CMNPs‐cells from fermentation broth via an external magnetic force, as well as its excellent long‐term performance overcoming the possible accumulation of dead cells and culture degeneration, the developed usage of CMNPs‐cells could be very benign for industrial application. © 2018 Society of Chemical Industry

Sugarcane Bagasse as a Carrier for the Immobilization of Saccharomyces cerevisiaein Bioethanol Production

Sugarcane bagasse was used as a carrier to immobilize Saccharomyces cerevisiae in bioethanol production. This research aims to study the potential use of sugarcane bagasse as an alternative carrier for cell immobilization and improvement in the production process of cell immobilization in bagasse. The results showed that the physical characteristics of sugarcane bagasse as a carrier were water content (7.77 ± 0.35%), water retention (4.80 ± 0.44 g/g), water absorption index (8.58 ± 0.22 g/g), and lignin content (24.40 ± 1.52 %). Determination of cell retention was performed in an inoculum volume of 50 mL yeast suspension with various carrier weights (2.5, 5, 10, and 20 g). The highest cell retention was obtained in ratio of 2.5 g carrier/50 mL cell suspension with cell retention of 5.41 ± 1.06 mg/g, or known as biocatalyst. Biocatalyst, as much as 1.5, 3, 4.5, and 6 g, were used as inoculum for a 24 hour bioethanol fermentation. The best concentration and productivity of bioethanol, obtained by using 3 g of biocatalyst, were 23.95 ± 0.28 g/L and 1.24 ± 0.01 g/L/hours. The average of bioethanol yield for a 24 hour fermentation by using immobilized cells was three times higher than the free cells system.

Polyethyleneimine-grafted collagen fiber as a carrier for cell immobilization.

Collagen fiber (CF), an abundant natural biopolymer, features many favorable properties that make it a potential carrier for cell immobilization. In the present investigation, CF was grafted with polyethyleneimine (PEI) using glutaraldehyde (GA) as the cross-linking agent, resulting in the formation of a novel CF based carrier (CF-PEI). The properties of CF-PEI as a carrier were evaluated by the immobilization of Microbacterium arborescens (CICC 20196), which has glucose isomerase (EC 5.3.1.5) activity. It was found that M. arborescens cells immobilized on CF-PEI exhibited higher glucose isomerization than those using activated carbon or anion exchange resin as the carriers. The Michaelis constant (K m) of the isomerization reaction for the CF-PEI-immobilized M. arborescens cells was 0.528 mol/L, which was slightly higher than that of free cells (0.473 mol/L). In addition, the apparent activation energies (E a) of free and immobilized cells on CF-PEI were almost the same at 60 kJ/mol. In an isomerization reaction of glucose to fructose in a fixed-bed reactor, CF-PEI-immobilized M. arborescens cells showed appreciable activity and operational stability. The corresponding isomerization ratio was as high as 41 % for 20 days, and the half-life was about 40 days.

Immobilization of saccharomyces cerevisiae in novel hydrogels based on hybrid networks of poly(ethylene oxide), alginate and chitosan for ethanol production

Hydrogel matrices were designed as hybrid networks of poly(ethylene oxide) (PEO) with natural polymers,alginate or chitosan by UV irradiation. The networks were formulated in the single-stage procedure inwhich the alginate or chitosan were added to the crosslinking reaction solution of PEO, and two-stage procedure,with additional chemical crosslinking of alginate or chitosan. Double-layer hydrogels composed of PEOhydrogel core with entrapped cells and outer natural hydrogel layer were also synthesized. The hydrogels werecharacterized by gel fraction yield and degree of equilibrium swelling as well as by rheological measurements.The production of ethanol by immobilized Saccharomyces cerevisiae was used to test the suitability of the synthesizedhybrid hydrogels to serve as carriers for cell immobilization. The presence of cells affected the mechanicalproperties and the structure of the polymer networks. The best system for immobilization was found to bethe PEO/alginate/Ca, which exhibited high mechanical strength (G′, 830; GF, 93; ESH2O, 15) without affectingthe metabolic functions of the cells. The maximum ethanol yield was 0.42 g/g corresponding to 82 % of thetheoretical yield.

Sol–gel synthesis and structure of silica hybrid materials

In this work the research results on the sol–gel synthesis and structure of silica nanocomposites, containing carrageenan and their application as carriers for cell immobilization were described. The samples were prepared at room temperature by replacing different quantity of the inorganic precursor with κ-carrageenan. For studying the structure of the synthesized hybrids the following methods were used: FT-IR, XRD, BET-Analysis, SEM, AFM and Roughness Analysis. The influence of the type of silicon precursors, nature and quantity of organic component on the structure, surface area, design and size of nanostructures was established. The possibility of application of the synthesized biocatalysts in an enzyme degradation process of the toxic, carcinogenic and mutagenic substances benzonitrile, fumaronitrile, o-, m-, and p-tolunitriles was investigated at batch experiments. A two-step biodegradation process in a column bioreactor of fumaronitrile was followed. After operation of the system for 8 h at a flow rate 45 mL h−1 and at 60 °C, the overall conversion was 89%, showing a good stability of the developed process.

The application of corrugated parallel bundle model to immobilized cells in porous microcapsule membranes

To describe immobilized cells in porous microcapsule membranes with straight pores, a novel model called corrugated parallel bundle model (CPBM) was utilized. In this model, a network was developed with 10 main pores each composing 10 pore elements. Cell growth kinetic in the network was examined using non-structural models. Effectiveness factor and pore plugging time were calculated by solving reaction-diffusion equation set via finite difference method. The findings revealed that diffusion coefficient for lower order reactions will create a lesser impact on the reduction of effectiveness factor. These findings also indicated that the use of such supporting carrier for cell immobilization could enhance the operational life of the system.

Poly(vinyl alcohol) cryogels employed as matrices for cell immobilization. 2. Entrapped cells resemble porous fillers in their effects on the properties of PVA-cryogel carrier

Poly(vinyl alcohol) cryogels which are prepared by freeze-thawing of concentrated aqueous solutions of the polymer are used as carriers for cell immobilization. Biomass entrapment causes a variation in the mechanical properties of the composites obtained. With rheological testing and scanning electron microscopy, it was shown that cells which manifested the reinforcing action in respect to the carrier resembled in their “physical behavior” simple model inorganic fillers of definite porosity. This porosity allows macromolecules of gel-forming polymer to penetrate into the inner regions of the particles and promote the formation of tight contacts between the matrix and dispersed phase, such as entrapped cells in particular. The immobilized cultures and model fillers were as follows: Citrobacter intermedius, Zymomonasas mobilis, and Pseudomonas sp. bacteria, Saccharomyces cerevisiae native and modified yeast cells, titanium dioxide, silica gels, and controlled pore glass.

Effect of ferrous iron concentration on the catalytic activity of immobilized cells of Thiobacillus ferrooxidans

The kinetics of continuous oxidation of ferrous iron by immobilized cells of Thiobacillus ferrooxidans was studied in a packed-bed bioreactor. Polyurethane foam biomass support particles were used as carriers for cell immobilization. Effects of ferrous iron concentration and its volumetric loading on the kinetics of the reaction were investigated. Media containing different concentrations of ferrous iron in the range 5–20 kg m-3 were tested. For each medium the kinetics of the reaction at different volumetric loadings of ferrous iron, at a constant temperature of 30°C, were determined. With media containing 5 kg m-3 and 10 kg m-3 Fe2+, the fastest oxidation rates of 34.25 kg m-3 h-1 and 32 kg m-3 h-1 were achieved at a dilution rate of around 6 h-1, which represents a residence time of 10 min. Employing a higher concentration of ferrous iron (20 kg m-3) in the medium resulted in lower oxidation rates, with a maximum value of 10 kg m-3 h-1, indicating an inhibitory effect of ferrous iron on growth and activity of T. ferrooxidans. The reliable performance of the bioreactor during the course of the experiments confirmed the suitability of polyurethane foam biomass support particles as carriers for T. ferrooxidans immobilization.

Loofa (Luffa cylindrica) sponge as a carrier for microbial cell immobilization

Abstract The feasibility of using loofa (Luffa cylindrica) sponge as a carrier for cell immobilization was investigated. In comparison with other carriers used for immobilization by cell adhesion, the density of loofa sponge is very low while the porosity and specific pore volume are very high. Furthermore, loofa sponge is stable over the whole range of pH and can be autoclaved many times without any visible change in the shape, structure and texture. By simply inserting loofa sponge in a flocculating yeast cell suspension, the flocs were entrapped within the lattice structure and the immobilization was completed in less than 20 min. The immobilized cells grew within the void volume, reaching a high cell concentration of over 4.4 g-cells/g-sponge. As an example, the immobilized cells were used for ethanol production from both sucrose and molasses media. More than 35 repeated batches on sucrose medium were performed without any loss in the activity of the immobilized cells. Also with both sucrose and molasses media, more than 500 h of stable continuous processing were possible. During the continuous processing, the cells remained firmly immobilized to the sponge and the free cell concentration in the effluent was less than 0.1 g/l throughout the fermentation period. The above results demonstrate that loofa sponge is an excellent carrier for immobilization of flocculating cells.

Aerobic fluidized-bed treatment of polychlorinated phenolic wood preservative constituents

Degradation of polychlorinated phenols was studied in continuous-flow fluidized-bed reactors using pure oxygen for aeration and celite carrier for cell immobilization. High dilution rates and chlorophenols as the only source of carbon and energy were used for maintenance of the mixed biofilm cultures. The chlorophenol degradation performance was monitored as release of inorganic chloride and removal of total organic carbon and by direct gas chromatographic analyses. Continuous polychlorophenol biodegradation activity was maintained in a fluidized-bed reactor for 315 days. Chloride release and chlorophenol removal efficiencies of over 99% were achieved at substrate loading rates of up to 430 g 2,4,6-trichlorophenol/m1/d and 400 g 2,3,4,6-tetrachlorophenol/m1/d at 3–5 h hydraulic retention times, respectively. Immobilized mixed cultures biodegraded 2,3,4,6-tetrachlorophenol with partial efficiency at feed concentrations as high as 157 mg/l as indicated by inorganic chloride release. Inorganic chloride release in experiments with practical grade 2,3,4,6-tetrachlorophenol exceeded that which is possible by dechlorination of tetrachlorophenol alone suggesting dechlorination of unidentified chlorinated impurities of the preparation as well. Fluidized-bed treatment of simulated wood preservative contaminated groundwater at a chlorophenol loading rate of 217 g/m1/d and 5 h hydraulic retention time resulted in the following removal efficiencies: 99.5% for 2,4,6-trichlorophenol, 99.6% for 2,3,4,6-tetrachlorophenol and 92.5% for pentachlorophenol.

Specialized bacterial strains for the removal of dichloromethane from industrial waste

A flow-through process for the mineralization of dichloromethane was developed. To select a bacterial strain with a high degradation capacity for dichloromethane the growth properties of ten isolates were compared in batch and continuous culture. Growth rates varied between 0.01 and 0.11 h−1 and degradation rates were in the range between 0.05 and 0.86 g/1·h−1. Several strains were unable to grow on media containing 120 or 170 mM dichloromethane due to the high concentration of salt arising from the neutralization of the hydrochloric acid generated by the dehalogenation reaction. For all organisms the maximum growth rate decreased with an increase in substrate concentration. A fluidized bed reactor with sand particles as a carrier for cell immobilization was tested. The system was inoculated with a mixture of dichloromethane-degrading organisms and operated under non-sterile conditions. It exhibited a maximum degradation rate of 1.6 g dichloromethane/1·h−1 and was stable during long-time operation.