Establishment of cluster of differentiation 20 immobilized cell membrane chromatography for the screening of active antitumor components in traditional Chinese medicine.

Non-Hodgkin lymphoma (NHL) is a heterogeneous group of malignant tumors occurring in B or T lymphocytes, and no small molecule-positive drugs to treat NHL have been marketed. Cluster of differentiation 20 (CD20) is an important molecule regulating signaling for the life and differentiation of B lymphocytes and possesses the characteristics of a drug target for treating NHL. 2-Methoxyestradiol induces apoptosis in lymphoma Raji cells and CD20 protein is highly expressed by Raji lymphoma cells. Therefore, in this study, a CD20-SNAP-tag/CMC model was developed to validate the interaction of 2-methoxyestradiol with CD20. 2-Methoxyestradiol was used as a small molecule control compound, and the system was validated for good applicability. The cell membrane chromatography model was combined with high-performance liquid chromatography ion trap time-of-flight mass spectroscopy (HPLC-IT-TOF-MS) in a two-dimensional system to successfully identify, analyze, and characterize the potential active compounds of Schisandra chinensis (Turcz.) Baill. extract and Lysionotus pauciflorus Maxim. extract, including Schisandrin A, Schizandrol A, Schizandrol B, Schisantherin B, and Nevadensin, which can act on CD20 receptors. The five potential active compounds were analyzed by non-linear chromatography. The thermodynamic and kinetic parameters of their interaction with CD20 were also analyzed, and the mode of interaction was simulated by molecular docking. Their inhibitory effects on lymphoma cell growth were assessed using a Cell Counting Kit-8 (CCK-8). Nevadensin and Schizandrin A were able to induce apoptosis in Raji cells within a certain concentration range. In conclusion, the present experiments provide some bases for improving NHL treatment and developing small molecule lead compounds targeting CD20 with low toxicity and high specificity.

Uranium biomineralization by immobilized Chryseobacterium sp. strain PMSZPI cells for efficient uranium removal.

Uranium (U) contamination is hazardous to human health and the environment owing to its radiotoxicity and chemical toxicity and needs immediate attention. In this study, the immobilized biomass of Chryseobacterium sp. strain PMSZPI isolated from U enriched site, was investigated for U(VI) biomineralization in batch and column set-up. Under batch mode, the fresh or lyophilized cells successfully entrapped in calcium alginate beads demonstrated effectual U precipitation under acid and alkaline conditions. The maximum removal was detected at pH 7 wherein ∼98-99% of uranium was precipitated from 1 mM uranyl carbonate solution loading ∼350 mg U/g of biomass within 24 h in the presence of organic phosphate substrate. The resulting uranyl phosphate precipitates within immobilized biomass loaded beads were observed by SEM-EDX and TEM while the formation of U biomineral was confirmed by FTIR and XRD. Retention of phosphatase activity without any loss of uranium precipitation ability was observed for alginate beads with lyophilized biomass stored for 90 d at 4 °C. Continuous flow through experiment with PMSZPI biomass immobilized in polyacrylamide gel exhibited U loading of 0.8 g U/g of biomass at pH 7 using 1 l of 1 mM uranyl solution. This investigation established the feasibility for the application of immobilized PMSZPI biomass for field studies. ENVIRONMENTAL IMPLICATION: Uranium contamination is currently a serious environmental concern owing to anthropogenic activities and needs immediate attention. We have developed here a biotechnological method for successful uranium removal using immobilized cells of a uranium tolerant environmental bacterium, Chryseobacterium sp. strain PMSZPI isolated from U ore deposit via phosphatase enzyme mediated uranium precipitation. The ability of immobilized PMSZPI cells to precipitate U(VI) as long-term stable U phosphates under environmental conditions relevant for contaminated waters containing high concentrations of U that exerts toxicity for biological systems is explored here. The long term stability of the immobilized biomass without compromising its U removal capacity shows the relevance of the bioremediation strategy for uranium contamination proposed in this work.

Cyclic extraction of phosphate from soybean meal using immobilized Aspergillus oryzae SBS50 phytase.

Phytase enzyme found in plants, animals, and microorganisms is mainly involved in catalyzing the systematic removal of a phosphate group from phytic acid. Enzyme immobilization is one of the cost-effective methods for the wide usage of enzymes in the industrial sector. This paper reports the covalent immobilization of phytase on glutaraldehyde-activated aluminum oxide beads. The immobilization yield, efficiency, and activation energy were found to be 47.8%, 71.5%, and 15.78 J/mol, respectively. The bound enzyme displayed a shift in pH optima from 5.5 to 4.5, which is more beneficial to increase digestibility in comparison with the free enzyme. Immobilized phytase retained 42.60% of its activity after 1.0 h incubation at 80 °C, whereas free enzyme retained only 4.20% of its activity. Thermodynami increase in half-lives, D-values, enthalpy and free energy change after covalent immobilization could be credited to the enhanced stability. Immobilized phytase could be reused for five consecutive cycles retaining 51% of its initial activity with sodium phytate. The immobilized phytase was also found effective to hydrolyze the soybean meal, thus increasing the digestibility of poultry feed. The hydrolyzing reaction of soybean meal was carried out for six consecutive cycles and immobilized phytase retained nearly 50% of activity till the fifth cycle. The amount of phosphorus released after treatment with immobilized phytase was far higher than that from free phytase. Immobilization on this support is significant, as this support can sustain high mechanical resistance at high pH and temperature. This considerable stability and reusability of the bound enzyme may be advantageous for its industrial application.

Hydrodynamic cavitation-assisted acid pretreatment and fed-batch simultaneous saccharification and co-fermentation for ethanol production from sugarcane bagasse using immobilized cells of Scheffersomyces parashehatae.

A new alternative for hydrodynamic cavitation-assisted pretreatment of sugarcane bagasse was proposed, along with a simultaneous saccharification and co-fermentation (SSCF) process performed in interconnected columns. Influential variables in the pretreatment were evaluated using a statistical design, indicating that an ozone flow rate of 10 mg min-1 and a pH of 5.10 resulted in 86 % and 72 % glucan and xylan hydrolysis yields, respectively, in the subsequent enzymatic hydrolysis process. Under these optimized conditions, iron sulfate (15 mg L-1) was added to assess Fenton pretreatment, resulting in glucan and xylan hydrolysis yields of 92 % and 71 %, respectively, in a material pretreated for 10 min. In SSCF, ethanol volumetric productivities of 0.33 g L-1 h-1 and of 0.54 g L-1 h-1 were obtained in batch and fed-batch operation modes, achieving 26 g L-1 of ethanol in 48 h in the latter mode.

"3-in-1" Hybrid Biocatalysts: Association of Yeast Cells Immobilized in a Sol-Gel Matrix for Determining Sewage Pollution.

This study presents a novel ″3-in-1″ hybrid biocatalyst design that combines the individual efficiency of microorganisms while avoiding negative interactions between them. Yeast cells of Ogataea polymorpha VKM Y-2559, Blastobotrys adeninivorans VKM Y-2677, and Debaryomyces hansenii VKM Y-2482 were immobilized in an organosilicon material by using the sol-gel method, resulting in a hybrid biocatalyst. The catalytic activity of the immobilized microorganism mixture was evaluated by employing it as the bioreceptor element of a biosensor. Optical and scanning electron microscopies were used to examine the morphology of the biohybrid material. Elemental distribution analysis confirmed the encapsulation of yeast cells in a matrix composed of methyltriethoxysilane (MTES) and tetraethoxysilane (TEOS) (85 and 15 vol %, respectively). The resulting heterogeneous biocatalyst exhibited excellent performance in determining the biochemical oxygen demand (BOD) index in real surface water samples, with a sensitivity coefficient of 50 ± 3 × 10-3·min-1, a concentration range of 0.3-31 mg/L, long-term stability for 25 days, and a relative standard deviation of 3.8%. These findings demonstrate the potential of the developed hybrid biocatalyst for effective pollution monitoring and wastewater treatment applications.

A microscale system for in situ investigation of immobilized microalgal cell resistance against liquid flow in the early inoculation stage.

In attached microalgae cultivation systems, cell detachment due to fluid hydrodynamic flow is not a subject matter that is commonly looked into. However, this phenomenon is of great relevance to optimizing the operating parameters of algae cultivation and feasible reactor design. Hence, this current work miniaturizes traditional benchtop assays into a microfluidic platform to study the cell detachment of green microalgae, Chlorella vulgaris, from porous substrates during its early cultivation stage under precisely controlled conditions. As revealed by time lapse microscopy, an increase in bulk flow velocity facilitated nutrient transport but also triggered cell detachment events. At a flow rate of 1000 µL min-1 of growth medium for 120 min, the algal cell coverage was up to 5% lower than those at 5 µL min-1 and 50 µL min-1. In static seeding, the evolution of attached cell resistance toward liquid flows was dependent on hydrodynamic zones. The center zone of the microchannel was shown to be a "comfortable zone" of the attached cells to sequester nutrients effectively at lower medium flow rates but there was a profile transition where outlet zones favored cell attachment the most at higher flow rates (1.13 times higher than the center zone for 1000 µL min-1). Besides, computational fluid dynamics (CFD) simulations illustrated that the focusing band varied between cross-sections and depths, while the streamline was the least concentrated along the side walls and bottom plane of the microfluidic devices. It was intriguing to learn that cell detachment was not primarily happening along the symmetry streamline. Insight gained from this study could be further applied in the optimization of operating conditions of attached cultivation systems whilst preserving laminar flow conditions.

Continuous production of gamma aminobutyric acid by engineered and immobilized Escherichia coli whole-cells in a small-scale reactor system.

γ-Amino butyric acid (GABA) is a non-proteinogenic amino acid and a human neurotransmitter. Recently, increasing demand for food additives and biodegradable bioplastic monomers, such as nylon 4, has been reported. Consequently, considerable efforts have been made to produce GABA through fermentation and bioconversion. To realize bioconversion, wild-type or recombinant strains harboring glutamate decarboxylase were paired with the cheap starting material monosodium glutamate, resulting in less by-product formation and faster production compared to fermentation. To increase the reusability and stability of whole-cell production systems, this study used an immobilization and continuous production system with a small-scale continuous reactor for gram-scale production. The cation type, alginate concentration, barium concentration, and whole-cell concentration in the beads were optimized and this optimization resulted in more than 95 % conversion of 600 mM monosodium glutamate to GABA in 3 h and reuse of the immobilized cells 15 times, whereas free cells lost all activity after the ninth reaction. When a continuous production system was applied after optimizing the buffer concentration, substrate concentration, and flow rate, 165 g of GABA was produced after 96 h of continuous operation in a 14-mL scale reactor. Our work demonstrates the efficient and economical production of GABA by immobilization and continuous production in a small-scale reactor.

Biotreatment efficiency, degradation mechanism and bacterial community structure in an immobilized cell bioreactor treating triclosan-rich wastewater.

Biotreatment of triclosan is mainly performed in conventional activated sludge systems, which, however, are not capable of completely removing this antibacterial agent. As a consequence, triclosan ends up in surface and groundwater, constituting an environmental threat, due to its toxicity to aquatic life. However, little is known regarding the diversity and mechanism of action of microbiota capable of degrading triclosan. In this work, an immobilized cell bioreactor was setup to treat triclosan-rich wastewater. Bioreactor operation resulted in high triclosan removal efficiency, even greater than 99.5%. Nitrogen assimilation was mainly occurred in immobilized biomass, although nitrification was inhibited. Based on Illumina sequencing, Bradyrhizobiaceae, followed by Ferruginibacter, Thermomonas, Lysobacter and Gordonia, were the dominant genera in the bioreactor, representing 38.40 ± 0.62% of the total reads. However, a broad number of taxa (15 genera), mainly members of Xanthomonadaceae, Bradyrhizobiaceae and Chitinophagaceae, showed relative abundances between 1% and 3%. Liquid Chromatography coupled to Quadrupole Time-Of-Flight Mass Spectrometry (LC-QTOF-MS) resulted in the identification of catabolic routes of triclosan in the immobilized cell bioreactor. Seven intermediates of triclosan were detected, with 2,4-dichlorophenol, 4-chlorocatechol and 2-chlorohydroquinone being the key breakdown products of triclosan. Thus, the immobilized cell bioreactor accommodated a diverse bacterial community capable of degrading triclosan.

Reduction of metronidazole in municipal wastewater and protection of activated sludge system using a novel immobilized Aspergillus tabacinus LZ-M.

Metronidazole (MNZ) accumulation inhibits municipal wastewater treatment bio-systems, and an effective solution to augment anaerobic activated sludge (AAS) is required. This research discovered that Aspergillus tabacinus LZ-M could degrade 77.39% of MNZ at 5 mg/L. MNZ was metabolized into urea, and the enzymes involved in its degradation were aminotransferase, methyltransferase, monooxygenase, and CN cleavage hydrolase. The strain was immobilized in polyurethane foam and used in AAS for the treatment of MNZ-containing municipal wastewater. The results showed that, using immobilized LZ-M, MNZ was completely removed, and the degradation efficiency of wastewater's chemical oxygen demand (COD) was increased from 11.7% to 83.31%. The extracellular polymer and ROS levels indicated that MNZ's toxicity on AAS was reduced. Furthermore, bioaugmentation stabilized its microbial community, and decreased MNZ resistance genes. These observations confirm that the immobilized fungi are effective in protecting AAS against antibiotic contamination in the treatment process of municipal wastewater.

Enhanced biodegradation of thiamethoxam with a novel polyvinyl alcohol (PVA)/sodium alginate (SA)/biochar immobilized Chryseobacterium sp H5.

Long-term and extensive usage of thiamethoxam, the second-generation neonicotinoid insecticide, has caused a serious threat to non-target organisms and ecological security. Efficient immobilized microorganism techniques are a sustainable solution for bioremediation of thiamethoxam contamination. A Gram-negative aerobic bacterium Chryseobacterium sp H5 with high thiamethoxam-degrading efficiencies was isolated from activated sludge. Then we developed a novel polyvinyl alcohol (PVA)/sodium alginate (SA)/biochar bead with this functional microbe immobilization to enhance the biodegradation and removal of thiamethoxam. Results indicated that the total removal and biodegradation rate of thiamethoxam with PVA/SA/biochar (0.7 %) beads with Chryseobacterium sp H5 immobilization at 30 °C and pH of 7.0 within 7 d reached about 90.47 % and 68.03 %, respectively, much higher than that using PVA/SA immobilized microbes (75.06 %, 56.05 %) and free microbes (61.72 %). Moreover, the PVA/SA/biochar (0.7 %) immobilized microbes showed increased tolerance to extreme conditions. Biodegradation metabolites of thiamethoxam were identified and two intermediates were first reported. Based on the identified biodegradation intermediates, cleavage of C-N between the 2-chlorothiazole ring and oxadiazine, dichlorination, nitrate reduction and condensation reaction would be the major biodegradation routes of thiamethoxam. Results of this work suggested the novel PVA/SA/biochar beads with Chryseobacterium sp H5 immobilization would be helpful for the effective bioremediation of thiamethoxam contamination.

Improving mass transfer rates in microbial cell immobilization system for environmental applications: Synergistic interaction of cells on crude oil biodegradation.

Among the various techniques used to clean up polluted environments, bioremediation is the most cost-effective and eco-friendly option. The diversity of microbial communities in a consortium can significantly affect the biodegradability of hazardous organic pollutants, particularly for in situ bioremediation processes. This is largely attributed to interactions between members of a consortium. In this study, the effect of internal diffusion limitations in substrate model biodegradation was firstly examined by immobilized bacterial cells at different particle sizes produced by the electrospray technique. According to the obtained results, for particles with large size, the effectiveness factors (η) were about 0.58-0.67, and the resistance to diffusive on the biodegradation rate was significant, while with decreasing the particle size, η increases and approaches about 1. After selection of suitable bead size, heavy crude oil biodegradation was investigated using a consortium consisting of three oil-degrading bacterial strains at different treatment systems. The removal rate in the suspended co-culture system stands at minimum value of 38% with all three strains which is an indicator of negative interactions among consortium members. Independent immobilization of microorganisms minimizes the competition and antagonistic interactions between strains and leads to more crude oil removal, so that, the biodegradation rate reached 60%.

Development and operation of immobilized cell plug flow bioreactor (PFR) for treatment of textile industry effluent and evaluation of its working efficiency.

The release of untreated/partially treated effluent and solid waste from textile dyeing industries, having un-reacted dyes, their hydrolysed products and high total dissolved solids (TDS) over the period of time had led to the deterioration of ecological niches. In an endeavour to develop a sustainable and effective alternative to conventional approaches, a plug flow reactor (PFR) having immobilized cells of consortium of three indigenous bacterial isolates was developed. The reactor was fed with effluent collected from the equalization tank of a textile processing unit located near city of Amritsar, Punjab (India). The PFR over a period of 3 months achieved 97.98 %, 82.22 %, 87.36%, 77.71% and 68.75% lowering of colour, chemical oxygen demand (COD), biological oxygen demand (BOD), total dissolved solids (TDS) and total suspended solids (TSS) respectively. The comparison of the phytotoxicity and genotoxicity of untreated and PFR-treated output samples using plant and animal models indicated significant lowering of respective toxicity potential. This is a first report, as per best of our knowledge, regarding direct treatment of textile industry effluent without any pre-treatment and with minimal nutritional inputs, which can be easily integrated into already existing treatment plant. The successful implementation of this system will lower the cost of coagulants/flocculants and also lowering the sludge generation.

Continuous process technology for bottom-up synthesis of soluble cello-oligosaccharides by immobilized cells co-expressing three saccharide phosphorylases.

BACKGROUND: Continuous processing with enzyme reuse is a well-known engineering strategy to enhance the efficiency of biocatalytic transformations for chemical synthesis. In one-pot multistep reactions, continuous processing offers the additional benefit of ensuring constant product quality via control of the product composition. Bottom-up production of cello-oligosaccharides (COS) involves multistep iterative ß-1,4-glycosylation of glucose from sucrose catalyzed by sucrose phosphorylase from Bifidobacterium adeloscentis (BaScP), cellobiose phosphorylase from Cellulomonas uda (CuCbP) and cellodextrin phosphorylase from Clostridium cellulosi (CcCdP). Degree of polymerization (DP) control in the COS product is essential for soluble production and is implemented through balance of the oligosaccharide priming and elongation rates. A whole-cell E. coli catalyst co-expressing the phosphorylases in high yield and in the desired activity ratio, with CdP as the rate-limiting enzyme, was reported previously. RESULTS: Freeze-thaw permeabilized E. coli cells were immobilized in polyacrylamide (PAM) at 37-111 mg dry cells/g material. PAM particles (0.25-2.00 mm size) were characterized for COS production (~ 70 g/L) in mixed vessel with catalyst recycle and packed-bed reactor set-ups. The catalyst exhibited a dry mass-based overall activity (270 U/g; 37 mg cells/g material) lowered by ~ 40% compared to the corresponding free cells due to individual enzyme activity loss, CbP in particular, caused by the immobilization. Temperature studies revealed an operational optimum at 30 °C for stable continuous reaction (~ 1 month) in the packed bed (volume: 40 mL; height: 7.5 cm). The optimum reflects the limits of PAM catalyst structural and biological stability in combination with the requirement to control COS product solubility in order to prevent clogging of the packed bed. Using an axial flow rate of 0.75 cm- 1, the COS were produced at ~ 5.7 g/day and ≥ 95% substrate conversion (sucrose 300 mM). The product stream showed a stable composition of individual oligosaccharides up to cellohexaose, with cellobiose (48 mol%) and cellotriose (31 mol%) as the major components. CONCLUSIONS: Continuous process technology for bottom-up biocatalytic production of soluble COS is demonstrated based on PAM immobilized E. coli cells that co-express BaScP, CuCbP and CcCdP in suitable absolute and relative activities.

Use of yeast biocapsules as a fungal-based immobilized cell technology for Indian Pale Ale-type beer brewing.

Immobilized cell technologies (ICT) have been used in wort fermentation, beer maturation, or production of alcohol-free or low-alcohol beer. The purpose of ICT is to restrict intact cells to a specific location while allowing biological function. It improves cell stability, operational flexibility, and control in brewing, as well as ease in executing continuous operations. We investigated the use of yeast biocapsules for Indian Pale Ale (IPA) type beer wort fermentation, a novel ICT in brewing. Yeast biocapsules are a spherical yeast immobilization system in which yeast cells are encapsulated and connected to the hyphae of an inactivated hollow filamentous fungus pellet. Fermentations with yeast encapsulated in alginate beads, as the standard immobilization practice, and in free (non-immobilized) forms were carried out in parallel. We found that yeast biocapsules are a better option for cell reutilization than alginate beads, but worse for beer must clarity. Beer brewed with yeast biocapsules differed in concentration for five volatile compounds (acetaldehyde, diacetyl, ethyl acetate, 1,1-diethoxyethane, and isoamyl alcohol) and three sensory characters (persistency of the foam, malt, and yeast character). KEY POINTS: • Yeast biocapsules were investigated for beer wort fermentation • Biocapsules improve cell reutilization but are limited for beer clarification • Beer brewed with biocapsules is chemically different than conventional beer • Most sensory features did not differ between biocapsule and control beer.

High-sodium maltobionate production by immobilized Zymomonas mobilis cells in polyurethane.

The purpose of this study was the production of maltobionic acid, in the form of sodium maltobionate, by Z. mobilis cells immobilized in polyurethane. The in situ immobilized system (0.125-0.35 mm) was composed of 7 g polyol, 3.5 g isocyanate, 0.02 g silicone, and 7 g Z. mobilis cell, at the concentration of 210 g/L. The bioconversion of maltose to sodium maltobionate was performed with different cell concentrations (7.0-9.0 gimobilized/Lreaction_medium), temperature (30.54-47.46 °C), pH (5.55-7.25), and substrate concentration (0.7-1.3 mol/L). The stability of the immobilized system was evaluated for 24 h bioconversion cycles and storage of 6 months. The maximum concentration of sodium maltobionate was 648.61 mmol/L in 34.34 h process (8.5 gdry_cell/Lreaction_medium) at 39 °C and pH 6.30. The immobilized system showed stability for 19 successive operational cycles of 24 h bioconversion and 6 months of storage, at 4 °C or 22 °C.

Phenol biodegradation by immobilized Rhodococcus qingshengii isolated from coking effluent on Na-alginate and magnetic chitosan-alginate nanocomposite.

Phenol is a hazardous organic solvent to living organisms, even in its small amounts. In order to bioremediation of phenol from aqueous solution, a novel bacterial strain was isolated from coking wastewater, identified as Rhodococcus qingshengii based on 16S rRNA sequence analysis and named as strain Sahand110. The phenol-biodegrading capabilities of the free and immobilized cells of Sahand110 on the beads of Na-alginate (NA) and magnetic chitosan-alginate (MCA) nanocomposite were evaluated under different initial phenol concentrations (200, 400, 600, 800 and 1000 mg/L). Results illustrated that Sahand110 was able to grow and complete degrade phenol up to 600 mg/L, as the sole carbon and energy source. Immobilized cells of Sahand110 on NA and MCA were more competent than its free cells in degradation of high phenol concentrations, 100% of 1000 mg/L phenol within 96 h, indicating the improved tolerance and performance of the immobilized cells against phenol toxicity. Therefore, the immobilized Sahand110 on the studied beads, especially MCA bead regarding its suitable properties, has significant potential to enhanced bioremediation of phenol-rich wastewaters.

Single-Cellular Biological Effects of Cholesterol-Catabolic Bile Acid-Based Nano/Micro Capsules as Anti-Inflammatory Cell Protective Systems.

Recent studies in our laboratories have shown promising effects of bile acids in ➀ drug encapsulation for oral targeted delivery (via capsule stabilization) particularly when encapsulated with Eudragit NM30D® and ➁ viable-cell encapsulation and delivery (via supporting cell viability and biological activities, postencapsulation). Accordingly, this study aimed to investigate applications of bile acid-Eudragit NM30D® capsules in viable-cell encapsulation ready for delivery. Mouse-cloned pancreatic ß-cell line was cultured and cells encapsulated using bile acid-Eudragit NM30D® capsules, and capsules' images, viability, inflammation, and bioenergetics of encapsulated cells assessed. The capsules' thermal and chemical stability assays were also assessed to ascertain an association between capsules' stability and cellular biological activities. Bile acid-Eudragit NM30D® capsules showed improved cell viability (e.g., F1 < F2 & F8; p < 0.05), insulin, inflammatory profile, and bioenergetics as well as thermal and chemical stability, compared with control. These effects were formulation-dependent and suggest, overall, that changes in ratios of bile acids to Eudragit NM30D® can change the microenvironment of the capsules and subsequent cellular biological activities.

A critical review for advances on industrialization of immobilized cell Bioreactors: Economic evaluation on cellulose hydrolysis for PHB production.

Advances such as cell-on-cell immobilization, multi-stage fixed bed tower (MFBT) bioreactor, promotional effect on fermentation, extremely low temperature fermentation, freeze dried immobilized cells in two-layer fermentation, non-engineered cell factories, and those of recent papers are demonstrated. Studies for possible industrialization of ICB, considering production capacity, low temperatures fermentations, added value products and bulk chemical production are studied. Immobilized cell bioreactors (ICB) using cellulose nano-biotechnology and engineered cells are reported. The development of a novel ICB with recent advances on high added value products and conceptual research areas for industrialization of ICB is proposed. The isolation of engineered flocculant cells leads to a single tank ICB. The concept of cell factories without GMO is a new research area. The conceptual development of multi-stage fixed bed tower membrane (MFBTM) ICB is discussed. Finally, feasible process design and technoeconomic analysis of cellulose hydrolysis using ICB are studied for polyhydroxybutyrate (PHB) production.

Cyanide removal from aqueous environment by resting cells and PTFE immobilized cells of Sphingobacterium spp.

Microbial detoxification of cyanide offered an inexpensive, safe, and viable alternative to physiochemical processes for the treatment of cyanide in industrial effluents or contaminated sites. This study involved isolation of novel strain with high resistance against cyanide toxicity and able to degrade the cyanide radical. The strain was isolated from rocky area and identified as Sphingobacterium multivorium using 16S ribosomal RNA. Resting pregrown cells were used in simple reaction mixture to avoid the complication associated with the media. One-gram fresh weight of this bacteria was able to remove 98.5% from 1.5 g/L cyanide which is a unique result. Factor affecting the biochemical process such as pH, temperature, agitation, glucose concentration was examined. The optimum conditions were, pH 6-7, 30-40°C, and 100-150 rpm shaking speed and 0.25% glucose. Furthermore, the cells were used after immobilization in polytetrafluoroethylene (PTFE) polymer. The PTFE is very safe carrier and the cells withstand the entrapment process and were able to remove 92% (1 g/L cyanide). The immobilized cells were used for six successive cycles with about 50% removal efficiency. The storage life extended to 14 days. No previous work studied the cyanide removal by Sphingobacterium spp. The strain showed good applicable characters.

Bioremediation of oily seawater by Bacteria immobilization on a novel carrier material containing nutrients.

A novel carrier material was obtained by coating puffed rhubarb rice (PRR) with calcium alginate (CA) membrane. The carrier material was prepared to contain oil-degrading bacterial strains and inorganic nutrients through entrapping them in different locations. This formulation possessed floatability, biodegradability and nutrient slow-release properties. Therefore, it could be applied for oil biodegradation on seawater surfaces. For controlling the release rate of nutrients, the optimal preparation technique was established. The number of viable cells immobilized on the carrier material reached 2 × 109 CFU/g. This formulation could be stored at -20 °C for three months without a significant decrease in the number of viable immobilized cells (4 × 108 CFU/g). Scanning electron microscope (SEM) results showed that the cells were immobilized on the outer CA membrane, and the inorganic nutrients were entrapped in the inner PRR and CA membrane. The immobilized cells were able to remove 86% of the diesel oil at an initial diesel oil concentration of 1% (v/v), an incubation temperature of 37 °C, during three days of incubation. Gas chromatography-mass spectrometry (GC-MS) analysis results showed that most components of diesel oil were degraded by the formulations.