Alginate-entrapped Cells Research Articles

Manganese Removal by Biofiltration Using Activated Carbon-barium Alginate-entrapped Cells: Morphology, Durability, Settling Velocity, and Treatment Efficiency

Occurrence of manganese in water supplies causes colored water and pipe rusting in water treatment and distribution systems. Moreover, consumption of manganese-contaminated water may lead to neurotoxicity in humans. Biofilters have the potential to alleviate the manganese issue through bio-oxidation, particle separation, and adsorption processes. Biofiltration performance can be enhanced by augmentation of the manganese-oxidizing bacterium entrapped in polymeric materials. This study aimed to investigate the potential of barium alginate-entrapped cells supplemented with powdered activated carbon (PAC) for manganese removal. Streptomyces violarus strain SBP1, an effective manganese-oxidizing bacterium, was selected. The experiments were divided into 2 parts, including 1) characterization of barium alginate bead and 2) manganese removal testing. Effect of PAC content (1, 5, and 10% w/v) in the entrapment material on bead morphology, bead durability, and settling velocity (relative to a filtration medium) was investigated. Micro-structural observation using a scanning electron microscope showed that the PAC was distributed through intra-porous structure of the beads. The PAC-supplemented barium alginate beads improved durability (up to three-time higher Young’s modulus values). The PAC-supplemented barium alginate beads gave similar settling velocity compared to a filtration medium. The manganese removal efficiencies by the PAC-supplemented beads (no cells) ranged from 48 to 53%. Based on barium alginate bead characterization and manganese removal performance, the bead with 5% PAC content was selected for cell entrapment. The investigation revealed that the entrapped cells achieved faster manganese removal rate than free cell system. The findings from this study indicated high potential of the entrapped cells for use in future biofilter applications.

Whole cell immobilization of thermostable ?-amylase from phosphate solubilizing Bacillus thuringiensis A5-BRSC

Whole cell immobilization of thermostable ?-amylase from phosphate solubilizing Bacillus thuringiensis A5-BRSC - IJMR- Print ISSN No: - 2394-546X Online ISSN No:- 2394-5478 Article DOI No:- 10.18231/2394-5478.2018.0055, Indian Journal of Microbiology Research-Indian J Microbiol Res

Ethanol production from coffee mucilage fermentation by S. cerevisiae immobilized in calcium-alginate beads

Soluble sugars in coffee mucilage were directly fermented into ethanol using immobilized Saccharomyces cerevisiae cells in calcium alginate beads. Several tests were performed via alginate-entrapped cells in different bead sizes (3 or 7 mm) combined to alginate concentrations (2–4%). The highest yield of 0.33 g ethanol/g sugar was achieved at 18 h using alginate-entrapped cells in 3 mm diameter and 2% (w/v) alginate, which corresponded to 64% of the theoretically achievable yield of ethanol. Tests with 7 mm beads took 6 h longer to reach maximum ethanol productions. The reduction of bead size increased mass transfer of substrates from the liquid to the immobilized cells, accelerating sugar consumption and ethanol production. Entrapment enabled reuse of the cells for up to 3 consecutive batches with stable ethanol production (up to 72 h). This work confirms the reuse of alginate-entrapped cells is feasible for ethanol fermentation of mucilage without supplement of nutrients.

Mitigation of bactericidal effect of carbon nanotubes by cell entrapment

This study investigated the effects of the alginate and polyvinyl alcohol (PVA) entrapment on the viability of Escherichia coli cells exposed to single wall carbon nanotubes (SWCNTs) with a diameter of 1–2nm. Viability was examined using a galactosidase enzyme assay, LIVE/DEAD BacLight assay, and total ribonucleic acid quantity. Variables studied included SWCNT concentration (5, 10, 20, 50, 100, 200, 500, and 1000μg/ml), SWCNT length (0.5–2μm for short SWCNTs and 5–30μm for long SWCNTs), and initial bacterial concentration (6.5 log10 CFU and 9 log10 CFU per test). Results showed that both alginate and PVA entrapments mitigate the bactericidal effect of SWCNTs. At the highest SWCNT concentration tested (1000μg/ml), the viability of the cells relative to controls (systems with only E. coli, no SWCNTs), was 0–60% for free cells and 60–90% for alginate and PVA entrapped cells. The bactericidal effect depended on SWCNT type and concentration, and bacterial concentration. In general, long SWCNTs (5–30μm) caused significantly greater reductions in the viability of entrapped cells than the short SWCNTs except for the two highest SWCNT concentrations studied, 500 and 1000μg/ml. Microscopy showed that the entrapment matrices prevented SWCNTs from entering the beads. This study shows that bacterial entrapment is effective at limiting the bactericidal effect of SWCNTs.

Biodegradation of dimethyl phthalate ester using free cells, entrapped cells of Variovorax sp. BS1 and cell free enzyme extracts: A comparative study

Whole cells of Variovorax sp. BS1 had the ability to biodegrade up to 600 mg L−1 (3.08 mM) of dimethyl phthalate (DMP) at the end of 30 h of incubation at 30 °C. The total organic carbon analyses suggested mineralization of DMP using isolated bacterial strain. The cell-free extracts of Variovorax sp. BS1 contained mixture of enzymes required for the complete biodegradation of DMP, diethyl phthalate (DEP) and dibutyl phthalate (DBP). Transient intermediates or end products were not detected by HPLC and LC-MS analyses during or after biodegradation of DMP by Variovorax sp. BS1. Addition of Zn+2 to assay mixture containing DMP and cell free extract resulted in the accumulation of detectable quantity of metabolite, ortho-phthalate thereby suggesting that DMP biodegradation proceeded via de-esterification reactions. Cells of Variovorax sp. BS1 entrapped in calcium alginate beads could be re-used for atleast ten sequential cycles in plain water with diminutive loss of DMP biodegradation activity. Results suggested that whole cells, alginate entrapped cells of Variovorax sp BS1 and enzymes present in the cell free extracts may be used for efficient biodegradation of dimethyl phthalate.

Packed bed reactor for degradation of simulated cyanide-containing wastewater.

The discharge of cyanide-containing effluents into the environment contaminates water bodies and soil. Effective methods of treatment which can detoxify cyanide are the need of the hour. The aim of the present study is to develop a bioreactor for complete degradation of cyanide using immobilized cells of Serratia marcescens RL2b. Alginate-entrapped cells of S. marcescens RL2b were used for complete degradation of cyanide in a packed bed reactor (PBR). Cells grown in minimal salt medium (pH 6.0) were harvested after 20 h and exhibited 0.4 U mg−1 dcw activity and 99 % cyanide degradation in 10 h. These resting cells were entrapped using 3 % alginate beads and packed in a column reactor (20 × 1.7 cm). Simulated cyanide (12 mmol l−1)-containing wastewater was loaded and fractions were collected after different time intervals at various flow rates. Complete degradation of 12 m mmol l−1 (780 mg l−1) cyanide in 10 h was observed at a flow rate of 1.5 ml h−1. The degradation of cyanide in PBR showed direct dependence on retention time. The retention time of cyanide in the reactor was 9.27 h. The PBR can degrade 1.2 g of cyanide completely in 1 day.

Bioreactor study employing bacteria with enhanced activity toward cyanobacterial toxins microcystins.

An important aim of white (grey) biotechnology is bioremediation, where microbes are employed to remove unwanted chemicals. Microcystins (MCs) and other cyanobacterial toxins are not industrial or agricultural pollutants; however, their occurrence as a consequence of human activity and water reservoir eutrophication is regarded as anthropogenic. Microbial degradation of microcystins is suggested as an alternative to chemical and physical methods of their elimination. This paper describes a possible technique of the practical application of the biodegradation process. The idea relies on the utilization of bacteria with a significantly enhanced MC-degradation ability (in comparison with wild strains). The cells of an Escherichia coli laboratory strain expressing microcystinase (MlrA) responsible for the detoxification of MCs were immobilized in alginate beads. The degradation potency of the tested bioreactors was monitored by HPLC detection of linear microcystin LR (MC-LR) as the MlrA degradation product. An open system based on a column filled with alginate-entrapped cells was shown to operate more efficiently than a closed system (alginate beads shaken in a glass container). The maximal degradation rate calculated per one liter of carrier was 219.9 µg h−1 of degraded MC-LR. A comparison of the efficiency of the described system with other biological and chemo-physical proposals suggests that this new idea presents several advantages and is worth investigating in future studies.

Degradation of cationic surfactants using Pseudomonas putida A ATCC 12633 immobilized in calcium alginate beads

In this study, the degradation of tetradecyltrimethylammonium bromide (TTAB) by freely suspended and alginate-entrapped cells from the bacteria Pseudomonas putida (P. putida) A ATCC 12633 was investigated in batch cultures. The optimal conditions to prepare beads for achieving a higher TTAB degradation rate were investigated by changing the concentration of sodium alginate, pH, temperature, agitation rate and initial concentration of TTAB. The results show that the optimal embedding conditions of calcium alginate beads are 4% w/v of sodium alginate content and 2×10(8) cfuml(-1) of P. putida A ATCC 12633 cells that had been previously grown in rich medium. The optimal degradation process was carried out in pH 7.4 buffered medium at 30°C on a rotary shaker at 100rpm. After 48h of incubation, the free cells degraded 26mgl(-1) of TTAB from an initial concentration of 50mgl(-1) TTAB. When the initial TTAB concentration was increased to 100mgl(-1), the free cells lost their degrading activity and were no longer viable. In contrast, when the cells were immobilized on alginate, they degraded 75% of the TTAB after 24h of incubation from an initial concentration of 330mgl(-1) of TTAB. The immobilized cells can be stored at 4°C for 25days without loss of viability and can be reused without losing degrading capacity for three cycles.

Effects of entrapment on nucleic acid content, cell morphology, cell surface property, and stress of pure cultures commonly found in biological wastewater treatment

The effects of cell entrapment on nucleic acid content, cell morphology, cell surface property, and stress of major groups of bacteria (betaproteobacteria and gammaproteobacteria) in biological municipal wastewater treatment were investigated. Three different entrapment media (alginate, carrageenan, and polyvinyl alcohol) were examined. Results indicated that the entrapment and type of entrapment media affected nucleic acid content, cell morphology, cell surface property, and stress of the three representative species (Alcaligenes faecalis, Comamonas testosteroni, and Pseudomonas putida) studied. The highest deoxyribonucleic acid and ribonucleic acid increases were observed with the alginate and polyvinyl alcohol (PVA) entrapment, respectively. A cell morphological change from bacilli to coccoidal was observed in the case of alginate entrapment while the PVA-entrapped cells had a slim morphology when compared to non-entrapped cells and formed putative nanowires. The entrapment increased or decreased the surface roughness of cells depending on the type of entrapment media. Expression of a nitrosative stress gene, which is linked to oxygen deprivation, was observed more in the alginate-entrapped cells. These research findings advance the fundamental understanding of the entrapped cell physiology which can lead to more efficient entrapped cell-based wastewater treatment.

Significant enhancement of ( R)-mandelic acid production by relieving substrate inhibition of recombinant nitrilase in toluene–water biphasic system

The enantioselective hydrolysis of mandelonitrile with whole cells of a recombinant Escherichia coli expressing nitrilase activity was severely inhibited by the substrate at high concentrations (>300 mM), which resulted in a low yield of the target product ( R)-(−)-mandelic acid. To relieve the substrate inhibition and to enhance the ( R)-(−)-mandelic acid productivity, eight water–organic solvent biphasic systems were attempted in this work. Toluene was found to be the most suitable solvent as the organic phase among the solvents tested. Various parameters were systematically examined and optimized in shake flasks. The phase volume ratio, buffer pH and reaction temperature were shown to be sensitive parameters affecting both the yield and the enantiopurity of product in the biphasic system. Under the optimized conditions, significant enhancement of substrate tolerance from 200 mM to 500 mM and average productivity from 179.6 g l −1 d −1 to 352.6 g l −1 d −1 were achieved. Subsequently, the biocatalytic hydrolysis of mandelonitrile was successfully carried out in a stirred reactor (2-l scale) by repeated use of the calcium alginate entrapped cells for 5 batches, affording 110.7 g ( R)-(−)-mandelic acid in 98.0% ee (enantiomeric excess) and a specific production of 13.8 g (mandelic acid) g −1 (cell), respectively.

Ethanol production from sugar beet molasses by S. cerevisiae entrapped in an alginate–maize stem ground tissue matrix

A new alginate–maize stem ground tissue matrix was developed as a Saccharomyces cerevisiae carrier for ethanol fermentation from sugar beet molasses. There were several fermentation procedures in the present study: using free cells and alginate-entrapped cells with and without maize stem ground tissue supplementation (F; F + C; AB; AB + C), and using a new combined alginate–maize stem ground tissue carrier (ABC). It was found that addition of maize stem ground tissue meal (C), with honeycomb configuration, provided high surface areas for cell attachment and biofilm growth, and also increased alginate matrix porosity, enabling better mass transfer characteristic, better physical strength and stability of beads. The highest values of process parameters were obtained in the case of new carrier (ABC): the ethanol concentration of 60.36 g/l, percentage of the theoretical ethanol yield of 96.56%, ethanol yield of 0.493 g/g and the volumetric ethanol productivity of 2.51 g/l h. The medium supplementation with maize stem ground tissue significantly decreased acetaldehyde and acetic acid content, did not affect fusel alcohol and ethylacetate content of the distillate.

Effects of cell entrapment on nucleic acid content and microbial diversity of mixed cultures in biological wastewater treatment

The effects of entrapment on nucleic acid content and microbial diversity of mixed cultures in biological municipal wastewater treatment were investigated. Deoxyribonucleic acid content increased 1.6-5.5 times more in alginate entrapped cells than in free and polyvinyl alcohol (PVA) entrapped cells. PVA entrapment resulted in 1.1- to 5.9-fold more increases in ribonucleic acid content compared to that experienced by free and alginate entrapped cells. Entrapment in carrageenan changed the bacterial community structure more than the alginate and PVA entrapments (35-80% versus 0-35%) as determined by single-strand conformation polymorphism analyses. The change in the bacterial community structure of alginate entrapped cells was less time dependent than that of PVA entrapped cells. This study enhances understandings on the physiology of entrapped cells and their community evolution in wastewater treatment environments.

Nitrate Removal from Agricultural Infiltrate by Bioaugmented Free and Alginate Entrapped Cells

A bench-scale sand column experiment was conducted to investigate nitrate removal from synthetic agricultural infiltrate by denitrifying bacterial cells entrapped in calcium alginate compared to free cells. The effects of methanol as a carbon source and cell loading were examined. Low (0 to 50%) nitrate removal was observed in both entrapped and free-cell columns without methanol supplement. In the presence of methanol, nitrate removals of 90 to 99% and 56 to 75% were obtained for entrapped and free-cell columns, respectively. Nitrate removal followed first-order kinetics. The entrapped-cell columns achieved higher nitrate removal than the free-cell columns because of less bacterial losses. Scanning electron microscopic images supported the nitrate removal results that the denitrifying bacteria proliferated in the entrapment matrix, and several nitrogen gas voids were produced from denitrification.

Immobilization of Bacillus circulans MTCC 7906 for enhanced production of alkaline protease under batch and packed bed fermentation conditions

Alkaline protease production by immobilized cells of Bacillus circulans MTCC 7906 was studied using agar-agar and calcium alginate as entrapment materials. Under batch conditions using agar agar, 2% agar concentration, 6.25mm bead size and 200 beads/ flask inoculum size were the optimal conditions for a maximum alkaline protease production of 19.9 IU/ml and while for alginate beads, 3% alginate concentration, 5.2mm bead size and 200 beads/ flask inoculum size were the optimal conditions for a maximum alkaline protease production of 19.7 IU/ml. The alginate and agar agar beads were reusable for four and nine successive batch cycles respectively. A maximum enzyme production of 28 and 27 IU/ml was obtained with incubation time of 36 h, column length of 30cm and flow rate of 10ml/h with agar-agar and alginate entrapped cells respectively under packed bed fermentation conditions. Scanning electron microscopy (SEM) of the agar agar entrapped cells showed a random distribution of cells on the surface with pockets of high cell density.

Biodegradation of Naphthalene by Pseudomonas stutzeri in Marine Environments: Testing Cells Entrapment in Calcium Alginate for Use in Water Detoxification

ABSTRACT The biodegradation of naphthalene in sea water by freely suspended and alginate-entrapped cells of Pseudomonas stutzeri 19SMN4 has been investigated in batch cultures. The results showed that immobilized cells can be stored at 4°C for 1 month without loss of viability. The biodegradation was highly affected by the availability of nitrogen and phosphorous, so at 30°C a naphthalene concentration of 25 mM was almost completely degraded (93%) by free cells in 6 days in samples supplemented with these nutrients, whereas only 42% naphthalene was consumed in the nonsupplemented samples. Biodegradation was much slower at 16°C than at 30°C; after 6 days of culture at 30°C, almost all naphthalene was degradated by free and immobilized cells, whereas only 22% and 34% at 16°C, respectively. The degradation rate remained unaffected when the naphthalene concentration was reduced from 25 to 10 mM. Alginate of three different viscosities was used for immobilization of cells. After 7 days of culture, beads formed with 31.4 cP alginate were fragmented, whereas beads formed with 240 and 3600 cP did not display structural changes and afforded the same degradation rate. Beads formed with high-viscosity alginate retained cells more efficiently.

Biosurfactant production by free and alginate entrapped cells of Pseudomonas fluorescens

Production of biosurfactant by free and alginate-entrapped cells of Pseudomonas fluorescens Migula 1895-DSMZ was investigated using olive oil as the sole carbon and energy source. Biosurfactant synthesis was followed by measuring surface tension and emulsifying index E24 over 5 days at ambient temperature and at neutral pH. Diffusional limitations in alginate beads affected the kinetics of biosurfactant production when compared to that obtained with free cells culture. Nevertheless, the emulsion stability was improved and fewer by-products interfered with the biosurfactant activity. A decrease in pH down to 5 in the case of immobilized cells was observed during the first 3 days, after which it returned to its initial value. The minimum values of surface tension were 30 and 35 dynes cm(-1) achieved after 40 and 72 h with free and immobilized cells, respectively, while the corresponding maximum E24 values were 67 and 62%, respectively. After separation by acetone precipitation, the biosurfactant showed a rhamnolipid-type in nature, and had a good foaming and emulsifying activities. The critical micellar concentration was found to be 290 mg l(-1). The biosurfactant also showed good stability during exposure to high temperatures (up to 120 degrees C for 15 min), to high salinity (10% NaCl) and to a wide range of pH (4-9).

Effects of cell entrapment on growth rate and metabolic activity of mixed cultures in biological wastewater treatment

The effects of cell entrapment on the growth rate and metabolic activity of mixed cultures in biological municipal wastewater treatment were investigated. Three different cell entrapment media (alginate, carrageenan and polyvinyl alcohol) and three cell-to-matrix ratios (2.5%, 5% and 15%, w/v) were examined. Free (non-entrapped) mixed culture cells were included in the study for comparative purpose. Results indicated that the entrapment, type of entrapment media, and cell-to-matrix ratio had significant effects on the growth and metabolic activity of the mixed culture cells. In most cases, polyvinyl alcohol entrapped cells had the highest specific growth and specific substrate utilization rates while alginate entrapped cells provided the lowest or second lowest rates. The specific growth and specific substrate utilization rates decreased as cell-to-matrix ratio increased from 2.5% to 5% or 15%. This study enhances the fundamental understanding about the physiological behaviors of entrapped cells in wastewater treatment environment. It provides results that are useful for increasing the performances of entrapped cells for wastewater treatment.

Cell immobilization technique for the enhanced production of α-galactosidase by Streptomyces griseoloalbus

Streptomyces griseoloalbus was immobilized in calcium alginate gel and the optimal immobilization parameters (concentrations of sodium alginate and calcium chloride, initial biomass and curing time) for the enhanced production of α-galactosidase were determined. The immobilization was most effective with 3% sodium alginate and 0.1 M calcium chloride. The optimal initial biomass for immobilization was approximately 2.2 g (wet wt.). The alginate-entrapped cells were advantageous because there was a twofold increase in the enzyme yield (55 U/ml) compared to the highest yield obtained with free cells (23.6 U/ml). Moreover, with immobilized cells the maximum yield was reached after 72 h of incubation in batch fermentation under optimal conditions, whereas in the case of free cells the maximum enzyme yield was obtained only after 96 h of incubation. The alginate beads had good stability and also retained 75% ability of enzyme production even after eight cycles of repeated batch fermentation. It is significant that this is the first report on whole-cell immobilization for α-galactosidase production.

Studies On Free And Immobilised Cells Of Bacillus Species On The Production Of Alpha-Amylase

Studies On Free And Immobilised Cells Of Bacillus Species On The Production Of Alpha-Amylase

Degradation of 4-aminobenzenesulfonate by alginate encapsulated cells of Agrobacterium sp. PNS-1

Studies were carried out on 4-aminobenzenesulfonate (4-ABS) degradation by free and alginate entrapped cells of Agrobacterium sp. PNS-1. Degradation rate in batch reactors with free cells was marginally higher than Ca-encapsulated cells. Comparison of Ca 2+ and Ba 2+ as gelling agents showed that 4-ABS removal rate was significantly less with Ba-alginate entrapped cells. Specific degradation rates, using linear regression analysis and based on the initial biomass in the beads, varied from 49.7 mg/mg biomass/h to 92.0 mg/mg biomass/h for Ca-alginate encapsulated cells for different initial 4-ABS concentrations ranging from 200 to 800 mg/L. UV spectra of the aliquots drawn at different time intervals from batch reactors did not show accumulation of any intermediate during degradation. Ca-alginate immobilized cells could be repeatedly reused upto five cycles without any loss of activity. Studies with packed bed reactors, operated in a semi-continuous mode, showed that this could be used for 4-ABS degradation.