Abstract: A dark green dwarf mutant, TGM 167, was isolated from a gamma ray + sodium azide mutagenized population of cultivated groundnut breeding line, TFDRG 5. The mutant had a 45.8% reduction in height due to its shorter internodal length. Further, it was found to be insensitive towards exogenous GA(3) application, although it had nearly the same level of endogenous GA(3) as the parent. Genetic analysis revealed that the dwarfism is under the control of a single dominant gene. This dominant dwarfing gene was mapped with an SSR marker TC3H02 at a distance of 9.7 cM.
Abstract: Thiourea (TU) has been found to enhance the stress tolerance of plants in our earlier field trials. In the present study, the TU mediated effect on the redox and antioxidant responses were studied in response to salinity (NaCl) stress in Indian mustard (Brassica juncea (L.) Czern.) seedlings. Biochemical analyses of reactive oxygen species (ROS) and lipid peroxidation revealed that TU supplementation to NaCl brought down their levels to near control values as compared to that of NaCl stress. These positive effects could be correlated to the significant increases in the 1,1-diphenyl-2-picrylhydrazyl (DPPH)-radical scavenging activity, in the levels of reduced glutathione (GSH) and GSH/GSSG (reduced/oxidized glutathione) ratio and in the activities of superoxide dismutase (SOD; EC 1.1.5.1.1) and glutathione reductase (GR; EC 1.6.4.2) in NaCl+TU treatment as compared to that of NaCl treatment. Further, TU supplementation allowed plants to avoid an over-accumulation of pyridine nucleotides, to stimulate alternative pathways (through higher glycolate oxidase activity; EC 1.1.3.15) for channeling reducing equivalents and thus, to maintain the redox state to near control levels. These positive responses were also linked to an increased energy utilization (analyzed in terms of ATP/ADP ratio) and presumably to an early signaling of the stress through stimulated activity of ascorbate oxidase (EC 1.10.3.3), an important component of stress signaling. A significant reduction observed in the level of sodium ion (Na(+)) accumulation indicated that TU mediated tolerance is attributable to salt avoidance. Thus, the present study suggested that TU treatment regulated redox and antioxidant machinery to reduce the NaCl-induced oxidative stress.
Abstract: Whole cells of recombinant Escherichia coli were immobilized on the screen printed carbon electrode (SPCE) using glutaraldehyde. Recombinant E. coli was having high periplasmic expression of organophosphorus hydrolase enzyme, which hydrolyzes the methyl parathion into two products, p-nitrophenol and dimethyl thiophosphoric acid. Cells immobilized SPCE was studied under SEM. Cells immobilized SPCE was associated with cyclic voltammetry and cyclic voltammograms were recorded before and after hydrolysis of methyl parathion. Detection was calibrated based on the relationship between the changes in the current observed at +0.1 V potential, because of redox behavior of the hydrolyzed product p-nitrophenol. As concentration of methyl parathion was increased the oxidation current also increased. Only 20 μl volume of the sample was required for analysis. Detection range of biosensor was calibrated between 2 and 80 μM of methyl parathion from the linear range of calibration plot. A single immobilized SPCE was reused for 32 reactions with retention of 80% of its initial enzyme activity.
Abstract: Inner epidermis of onion bulb scales was used as a natural support for immobilization of microbial cells for biosensor application. A bacterium Sphingomonas sp. that hydrolyzes methyl parathion into a chromophoric product, p-nitrophenol (PNP), has been isolated and identified in our laboratory. PNP can be detected by electrochemical and colorimetric methods. Whole cells of Sphingomonas sp. were immobilized on inner epidermis of onion bulb scale by adsorption followed by cross-linking methods. Cells immobilized onion membrane was directly placed in the wells of microplate and associated with the optical transducer. Methyl parathion is an organophosphorus pesticide that has been widely used in the field of agriculture for insect pest control. This pesticide causes environmental pollution and ecological problem. A detection range 4-80 μM of methyl parathion was estimated from the linear range of calibration plot of enzymatic assay. A single membrane was reused for 52 reactions and was found to be stable for 32 days with 90% of its initial hydrolytic activity. The applicability of the cells immobilized onion membrane was also demonstrated with spiked samples.
Abstract: In the present study, Hydrilla verticillata plants were exposed to arsenate (AsV; 50 microM) and arsenite (AsIII; 5 microM) under variable S supply: deficient (2 microM S, -S), normal (1 mM S, +S) and excess (2 mM S, +HS). Arsenic accumulation (microg g(-1) dw) in +HS plants was about 2-fold higher upon exposure to both AsV (30) and AsIII (50) than that observed in +S (12 & 24) and -S (14 & 26) plants. Despite lower As accumulation, -S plants experienced the maximum oxidative stress owing to an inadequate response of enzymatic and molecular antioxidants and significant decline in total thiols and the ratio of reduced glutathione (GSH) to oxidized glutathione (GSSG). By contrast +HS plants had significant increase in total thiols and an improved redox status, did not demonstrate any negative impact to antioxidants except catalase and hence experienced the least increase in oxidative stress parameters. In conclusion, an increase in S supply to plants may improve their accumulation capacity for As through enhanced tolerance caused by a positive effect on thiol metabolism and antioxidant status of the plants.
Abstract: The uranium (U) accumulation potential and ensuing biochemical responses were studied in Hydrilla verticillata (L.f.) Royle upon exposure to U (0, 20 and 100 mg L(-1)). There was a concentration-duration dependent increase in U accumulation with the maximum being 78 mg g(-1) DW at 100 mg L(-1) U after 24 h. Plants experienced an initial phase of the maximum toxicity (within 30 min) followed by almost complete recovery after 24 h. The recovery was attributed to an integrated modulation in the level of both enzymatic and molecular antioxidants (viz., guaiacol peroxidase, ascorbate peroxidase, catalase, proline, total phenolics) and also the constituents of thiol metabolism (viz., cysteine and glutathione). Thus, plants were found to be able to accumulate significant amount of U in a short time and to tolerate it efficiently. Hence, they may find application in U phytoremediation considering there accumulation ability, fast growth due to weed-like habit and world-wide distribution.
Abstract: Comparative antioxidant profiling of tolerant (TPM-1) and sensitive (TM-4) variety of Brassica juncea L. was performed after exposure to arsenate [As(V)] and arsenite [As(III)]. TPM-1 demonstrated higher accumulation of As upon exposure to both 500 microM As(V) and 250 microM As(III) (49 and 37 microg g(-1) dw after 15 days) as compared with that observed in TM-4. The activities of various antioxidant enzymes and the level of glutathione and proline demonstrated, in general, a comparatively better response in TPM-1 than in TM-4 that presumably allowed TPM-1 to tolerate higher As concentrations as compared with that of TM-4.
Abstract: Morphological transformation during evaporation-induced self-assembly of a mixed colloidal suspension in micrometric droplets has been investigated. It has been demonstrated that a buckling-driven shape transition of drying droplets of mixed colloidal suspension takes place during evaporation-induced self-assembly. Further, it is also shown that the distortion modulations get significantly amplified with enhancement in volume fraction of anisotropic soft colloidal component of the mixed colloids. It has been argued that the reduction in elastic modulus of formed shell, at the boundary of a drying droplet, and the anisotropic nature of one of the colloidal components facilitate the deformation process. Hierarchical structures of these assembled colloidal grains have been probed using electron microscopy and scattering techniques.
Abstract: Phosphate-solubilizing bacteria (PSBs) were isolated from different plant rhizosphere soils of various agroecological regions of India. These isolates showed synthesis of pyrroloquinoline quinone (PQQ), production of gluconic acid, and release of phosphorus from insoluble tricalcium phosphate. The bacterial isolates synthesizing PQQ also showed higher tolerance to ultraviolet C radiation and mitomycin C as compared to Escherichia coli but were less tolerant than Deinococcus radiodurans. Unlike E. coli, PSB isolates showed higher tolerance to DNA damage when grown in the absence of inorganic phosphate. Higher tolerance to ultraviolet C radiation and oxidative stress in these PSBs grown under PQQ synthesis inducible conditions, namely phosphate starvation, might suggest the possible additional role of this redox cofactor in the survival of these isolates under extreme abiotic stress conditions.
Abstract: Abiotic stresses including salinity are the major constraints to crop production. In this regard, the use of thiourea (TU) in imparting salinity-stress tolerance to Indian mustard (Brassica juncea) has been demonstrated earlier. To gain an insight into the mechanism of TU action, various molecular and biochemical studies were conducted.
Abstract: Arsenic (As) is a potential hazard to plants' health, however the mechanisms of its toxicity are yet to be properly understood. To determine the impact of redox state and energetic in stress imposition, plants of Hydrilla verticillata (L.f.) Royle, which are known to be potential accumulator of As, were exposed to 100 and 500 μM arsenate (AsV) for 4 to 96 h. Plants demonstrated significant As accumulation with the maximum being at 500 μM after 96 h (568 μg g(-1) dry weight, dw). The accumulation of As led to a significant increase in the level of reactive oxygen species, nitric oxide, carbonyl, malondialdehyde, and percentage of DNA degradation. In addition, the activity of pro-oxidant enzymes like NADPH oxidase and ascorbate oxidase also showed significant increases. These parameters collectively indicated oxidative stress, which in turn caused an increase in percentage of cell death. These negative effects were seemingly linked to an altered energetic and redox equilibrium [analyzed in terms of ATP/ADP, NADH/NAD, NADPH/NADP, reduced glutathione/oxidized glutathione, and ascorbate/dehydroascobate ratios]. Although there was significant increase in the levels of phytochelatins, the As chelating ligands, a large amount of As was presumably present as free ion particularly at 500 μM AsV, which supposedly produced toxic responses. In conclusion, the study demonstrated that the magnitude of disturbance to redox and energetic equilibrium of plants upon AsV exposure determines the extent of toxicity to plants.
Abstract: Organophosphorus pesticides such as methyl parathion have been widely used in the field of agriculture for insect pest control. These pesticides and their degradation products cause environmental pollution and ecological problem. With a view to monitor these pesticides biosensors are being developed. A bacterium Sphingomonas sp. from field soil has been isolated and identified in our laboratory that hydrolyzes the methyl parathion upto a chromophoric product, p-nitrophenol (PNP). PNP can be detected by electrochemical and colorimetric methods, which can be exploited to develop a biosensor for detection of the organophosphate pesticide. Whole cells of Sphingomonas bacteria were immobilized directly onto the surface of the wells of polystyrene microplates (96 wells) using glutaraldehyde as the cross-linker. SEM study confirmed the immobilization of Sphingomonas sp. Immobilized bacterial microplate was associated directly with the optical transducer, microplate reader. The microplate-based biosensor is having advantages as it has 96 reaction vessels and therefore it provides a convenient system for detecting multiple numbers of samples in a single platform. Detection range of the biosensor from the linear range was determined to be 4-80 μM methyl parathion. Cells-immobilized microplates were having reusability upto 75 reactions. Present study reports an innovative concept where the microplate can be used as immobilizing support for development of reusable microbial biocomponent.
Abstract: Large areas of the globe are becoming saline due to evapotranspiration and poor irrigation practices, and sustainability of agriculture is being seriously affected. Thiourea (TU) has been identified as an effective bioregulator imparting stress tolerance to crops. The molecular mechanisms involved in the TU-mediated response are considered in this study.
Abstract: Inner epidermal membrane of the onion bulb scales was studied as a natural polymer support for immobilization of the glucose oxidase (GOD) enzyme for biosensor application. Onion epidermal membrane was used for immobilization of glucose oxidase and was associated with dissolved oxygen (DO) probe for biosensor reading. Glucose was detected on the basis of depletion of oxygen, when immobilized GOD oxidizes glucose into gluconolactone. A wide detection range between 22.5 and 450 mg/dl was estimated from calibration plot. A single membrane was reused for 127 reactions with retention of approximately 90% of its initial enzyme activity. Membrane was stable for 45 days ( approximately 90% activity) when stored in buffer at 4 degrees C. Surface structure studies of the immobilized membranes were carried under SEM. To our knowledge, this is the first report on employing inner epidermal membrane of onion bulb scales as the solid support for immobilization of enzyme.
Abstract: Potential of Chromolaena odorata plants for remediation of (137)Cs from solutions and low level nuclear waste was evaluated. When plants were exposed to solutions spiked with three different levels of (137)Cs, namely 1 x 10(3) kBqL(-1), 5 x 10(3) kBqL(-1) and 10 x 10(3) kBqL(-1), 89%, 81% and 51% of (137)Cs was found to be remediated in 15 d, respectively. At the lowest Cs activity (1 x 10(3) kBqL(-1)), accumulation of Cs was found to be higher in roots compared to shoots, while at higher Cs activities (5 x 10(3) kBqL(-1) and 10 x 10(3) kBqL(-1)), Cs accumulation was more in shoots than roots. When plants were incubated in low level nuclear waste, 79% of the activity was removed by plants at the end of 15 d. The present study suggests that C. odorata could be used as a potential candidate plant for phytoremediation of (137)Cs.
Abstract: Thorium biosorption by Aspergillus fumigatus was carried out in a batch reactor to study the effect of initial pH and metal ion concentration, contact time, biomass dose and kinetics and equilibrium Th uptake. Thorium(IV) uptake by A. fumigatus was pH dependent (pH range, 2.0-6.0) and maximum sorption was observed at pH 4.0. The uptake was rapid and the biosorption process reached equilibrium within 2h of contact times at pH 2-4 and initial Th concentration of 50 and 100mg/L. The kinetics data fitted well to Lagergren's pseudo-second-order rate equation (r(2)>0.99). A maximum initial sorption rate of 71.94 (mg/g min) and second-order rate constant of 7.82 x 10(-2) (g/mg min) were observed at pH 4.0, 50 mg Th/L. The observed maximum uptake of thorium was 370 mg Th/g at equilibrium. Biosorption process could be well described by Langmuir isotherm in comparison to Freundlich and Temkin isotherms. Sodium bicarbonate was the most efficient desorbing reagent with desorption efficiency of more than 99%. Environmental scanning electron micrograph (ESEM) showed that the surface of the biomass after desorption was intact.
Abstract: Paecilomyces marquandii, a phosphate-solubilizing, starch-utilizing filamentous fungus, was immobilized on polyurethane foam (PUF). The immobilized fungus was applied in a repeated batch (six batches) fermentation process to solubilize Hirapur rock phosphate. The fungus was immobilized on PUF cubes and was used for phosphate solubilization in shake flask repeated batch cultivations. The fungus was also immobilized on PUF sheet and utilized in an airlift bioreactor in a repeated batch process. Maximum soluble phosphate (370 microg/ml) was recorded after third batch with 8 g rock phosphate/l. After 12 days of fermentation, a total production of 1,643 microg phosphate/ml was achieved.
Abstract: An indigenous strain HD-550 of Bacillus thuringiensis subsp. kenyae was found to be toxic to lepidopteran as well as dipteran insects. The cry2Aa gene (classified as cry2Aa14) from this isolate was cloned and expressed in Escherichia coli. Only a little amount of the expressed Cry2Aa14 protein was observed in soluble fraction under normal induction condition. The inclusions were non-toxic to test insects, whereas solubilized Cry2Aa14 was highly toxic to lepidopteran and dipteran insects. Cry2Aa14 protein was expressed as thioredoxin (trx) fusion protein for improving the yield of active protein. An enhancement of nearly 15% was observed in the yield of active Cry2Aa14. The TrxA-Cry2Aa14 protein purified from the solubilized fraction also showed toxicity profile similar to the wild-type protein. The LC(50) values of Cry2Aa14 and TrxA-Cry2Aa14 protein against Spodoptera litura was 694 and 696 ng/cm(2), respectively, while for Culex quinquefasciatus the LC(50) values were 894 and 902 ng/ml, respectively. The broad spectrum toxicity of the Cry2Aa14 thus indicates that this protein could be an important component in integrated pest management. Further, the trx tag clearly led to higher yield, which facilitates protein purification for biophysical and biochemical characterization.
Abstract: The mechanisms of perception of arsenic (As)-induced stress and ensuing tolerance in plants remain unresolved. To obtain an insight into these mechanisms, biochemical and transcriptional profiling of two contrasting genotypes of Brassica juncea was performed. After screening 14 varieties for As tolerance, one tolerant (TPM-1) and one sensitive (TM-4) variety were selected and exposed to arsenate [As(V)] and arsenite [As(III)] for 7 d and 15 d for biochemical analyses. The tolerant variety (TPM-1) demonstrated higher accumulation of As upon exposure to both 500 microM As(V) and 250 microM As(III) [49 microg g(-1) and 37 microg g(-1) dry weight (dw) after 15 d] as well as a better response of thiol metabolism as compared with the responses observed in the sensitive variety (TM-4). Transcriptional profiling of selected genes that are known to be responsive to sulphur depletion and/or metal(loid) stress was conducted in 15-d-old seedlings after 3 h and 6 h exposure to 250 microM As(III). The results showed an up-regulation of sulphate transporters and auxin and jasmonate biosynthesis pathway genes, whereas there was a down-regulation of ethylene biosynthesis and cytokinin-responsive genes in TPM-1 within 6 h of exposure to As(III). This suggested that perception of As-induced stress was presumably mediated through an integrated modulation in hormonal functioning that led to both short- and long-term adaptations to combat the stress. Such a coordinated response of hormones was not seen in the sensitive variety. In conclusion, an early perception of As-induced stress followed by coordinated responses of various pathways was responsible for As tolerance in TPM-1.
Abstract: The lyophilized biomass of bacterium Brevibacterium ammoniagenes was immobilized in polystyrene sulphonate-polyaniline (PSS-PANI) conducting polymer on a Pt twin wire electrode by potentiostatic electropolymerization. The bacterial cells retained their viability as well as urease activity under entrapped state, as confirmed with bacterial live-dead fluorescent assay and enzymatic assays. The entrapped cells were visualized using scanning electron microscope. The immobilized cells were used as a source of unpurified urease to develop a conductometric urea biosensor. The catalytic action of urease in the sensor released ammonia, thereby causing an increase in the pH of the microenvironment. The pH dependant change in the resistivity of the polymer was used as the basis of sensing mechanism. The sensor response was linear over a range of 0-75 mM urea with a sensitivity of 0.125 mM(-1). The sensor could be reused for 12-15 independent measurements and was quite stable in dry as well as buffered storage condition at 4 degrees C for at least 7 days.
Abstract: The present study was aimed to analyze the effects of variable S supply on arsenic (As) accumulation potential of Hydrilla verticillata (Lf.) Royle. Plants were exposed to either arsenate (AsV; 50 microM) or arsenite (AsIII; 5 microM) for 4 h and 1 day while S supply was varied as deficient (2 microM, -S), normal (1 mM, +S) and excess (2 mM, +HS). The level of As accumulation (microg g(-1) dw) after 1 day was about 2-fold higher upon exposure to either AsV (30) or AsIII (50) in +HS plants than that being in +S (12 and 24) and -S (14 and 26) plants. The +HS plants showed a significant stimulation of the thiol metabolism upon As exposure. Besides, they did not experience significant toxicity, measured in terms of malondialdehyde accumulation; an indicator of oxidative stress. By contrast, -S plants suffered from oxidative stress probably due to negative impact to thiol metabolism. Variable S supply also modulated the activity of enzymes of glycine and serine biosynthesis indicating an interconnection between S and N metabolism. In conclusion, an improved supply of S to plants was found to augment their ability for As accumulation through stimulated thiol metabolism.
Abstract: Morphological transition of droplets during evaporation driven self-assembly of colloidal dispersion of alumina particles has been investigated. It was found that a sphere to doughnut-like transition of the droplet morphology takes place even when the rate of drying remains moderate and is not extremely fast. Further, it has been seen that such transition is strongly dependent on the volume fraction of the colloids in the droplets. The transition proceeds via buckling of the initial spherical droplets, which occurs when the capillary forces driving the deformation overcomes the interparticle electrostatic forces. However, the transition is hindered and the buckling probability is reduced due to the inherent spatial constraint when the colloid volume fraction is increased. Mesoscopic structures of the assembled grains have been investigated by scanning electron microscopy, small-angle neutron scattering, and dynamic light scattering techniques. Interestingly, it has been observed that the functionality of photoluminescence spectrum of the dried nanoporous grains depends somewhat on the grain morphology.
Abstract: Aseptically grown Vetiveria zizanoides were evaluated for their potential for phytoremediation of phenol from Murashige and Skoog's liquid medium. Phenol was found to be completely removed from incubation medium at the end of 4 days by V. zizanoides plantlets, when medium was supplemented with 50 and 100 mg L(-1) phenol, while with 200, 500, and 1000 mg L(-1) of phenol, 89%, 76% and 70%, respectively, were removed. Phenol removal was found to be associated with inherent production of peroxidase and hydrogen peroxide. Coupled with H(2)O(2) formation, the levels of antioxidant enzymes like superoxide dismutase and peroxidase showed an enhancement when plants were exposed to phenol, whereas catalase levels initially showed a decline due to the utilization of H(2)O(2) by peroxidase for phenol oxidation. However, when peroxidase levels declined, there was an enhancement in catalase levels to minimize the presence of H(2)O(2) in the medium. Having confirmed that the removal of phenol was by V. zizanoides plantlets, in the next phase, micropropagated plantlets and well-developed plants grown in hydroponics were used under in vivo conditions to study the effect of phenol (200 mg L(-1)) on plant growth and reuse. Although plant growth was reduced in presence of phenol, the results of the reuse study indicated the possibility of plants getting adapted to phenol without any decline in potential for phenol remediation.
Abstract: Removal of heavy metals present in wastewaters has been a major concern due to their non-biodegradability and toxicity. Removal of copper ion using NaOH treated Rhizopus oryzae biomass was investigated in a batch reactor. The copper uptake exhibited substantial enhancement both in terms of kinetics of uptake as well as the loading capacity. The copper biosorption by viable and pretreated fungal biomass fit well to a Lagergren's pseudo second order reaction in comparison to pseudo first order kinetics. Investigation on effect of pH indicated improved performance in the range of pH 4-6 in alkali treated biomass. Copper uptake exhibited by viable biomass was highest at 21 degrees C, unlike pretreated biomass that showed maximum uptake across the range of temperature 21-55 degrees C. The maximum copper loading capacity of the viable and pretreated biomass according to Langmuir isotherm was 19.4 and 43.7 mg/g, respectively. Distribution coefficient of pretreated biomass showed improvement at lower residual concentration, indicating a change in the nature of binding by the treated biomass. Copper uptake decreased with an increasing dose of biosorbent, although enhancement in the total metal ion removal was observed at higher dose.
Abstract: One of the major wastes generated by fertilizer, explosive, and nuclear industries are nitrate (as high as 1,000 ppm NO(3)N) whose removal before disposal has become a growing concern. In this study, an active denitrifying sludge was immobilized onto support materials like cloth and polyurethane foam and their denitrification efficiency on high nitrate wastes [1,000 ppm NO(3) (225 ppm NO(3)N), 5,000 ppm NO(3) (1,129 ppm NO(3)N), 7,500 ppm NO(3) (1,693 ppm NO(3) N)] was studied. Results showed complete degradation of the nitrate wastes (225 ppm NO(3)N, 1,129 ppm NO(3)N, and 1,693 ppm NO(3)N) without any accumulation of nitrite in a period of only 1, 4, and 10 h, respectively. Based on adhering and entrapment principle, an immobilization unit was developed using a combination of cloth and foam as well as both individually. This system used for treating such high nitrate wastes was found to be quite effective in waste water treatment, particularly in problems associated with solid-liquid separation. The batch column reactor was run in about 45 batches without any loss in activity or reactor stability.
Abstract: The biosorption characteristics of uranium(VI) onto Catenella repens (a red alga), were evaluated as a function of pH, biosorbent size, time, biomass dosage, initial uranium concentration and temperature. Within the pH range studied (1.5-7.5), 4.5 was the optimum pH for the uptake of uranium(VI) by C. repens. Reduction in particle size did not increase the biosorption capacity. The metal removal was rapid, with more than 90% of total biosorption taking place in 30 min, and equilibrium was attained in 45 min. The maximum metal loading capacity of the alga was 303 mg/g. Within the temperature range studied (15-55 degrees C), there was no significant change in biosorption, under optimal conditions. Adsorption process could be well defined by both the Langmuir and Freundlich isotherms with r(2) of 0.94 and 0.96, respectively. The kinetic data fitted the pseudo-second-order kinetic model with the r(2) value of 0.99. At a low pH of 2.5, where most of the biomasses show either no or less metal uptake, a good (>15%) metal loading capacity of 25% was achieved. Therefore biosorption characteristics were also evaluated at pH 2.5.
Abstract: An indigenously isolated strain of Bacillus thuringiensis subsp. kenyae exhibited toxicity against lepidopteran as well as dipteran insects. The lepidopteran active cry1Ac protoxin gene coding sequence of 3.5 kb from this strain was cloned into vector pET28a(+). However, it could not be expressed in commonly used Escherichia coli expression hosts, BL21(DE3) and BL21(DE3)pLysS. This gene is classified as cry1Ac17 in the B. thuringiensis toxic nomenclature database. The coding sequence of this gene revealed that it contains about 3% codons, which are not efficiently translated by these expression hosts. Hence, this gene was expressed in a modified expression host, Epicurian coli BL21-Codonplus (DE3)-RIL. The expression of gene yielded a 130-kDa Cry1Ac17 protein. The protein was purified and its toxicity was tested against economically important insect pests, viz., Helicoverpa armigera and Spodoptera litura. LC(50) values obtained against these insects were 0.1 ng/cm(3) and 1231 ng/cm(2), respectively. The higher toxicity of Cry1Ac17 protein, compared to other Cry1Ac proteins, toward these pests demonstrates the potential of this isolate as an important candidate in the integrated resistance management program in India.
Abstract: A membrane was prepared using polyvinyl alcohol (PVA) with low and high degree of polymerization (DOP), acetone, benzoic acid (BA) and was cross-linked by UV treatment. Membrane composition was optimized on the basis of swelling index. Membrane prepared with 12% low DOP and 8% high DOP of PVA, 2% BA, dissolved in buffer containing 20% acetone and cross-linked with UV treatment exhibited lower swelling index. Fourier transform infrared (FTIR) study of the membranes showed appearance of a strong band at approximately 2337 cm(-1) when UV was used for cross-linking in the presence of benzoic acid. Scanning electron microscope (SEM) study revealed that membrane cross-linked with UV treatment was smoother. Glucose oxidase (GOD)-PVA membrane was associated with the dissolved oxygen (DO) probe for biosensor reading. Glucose was detected on the basis of depletion of oxygen, when immobilized GOD oxidizes glucose to gluconolactone. A wide detection range, 0.9-225 mg/dl was estimated from the linear range of calibration plot of biosensor reading. Membranes were reused for 32 reactions without significant loss of activity and stored for 30 days (approximately 90% activity) at 4 degrees C. Membranes were also used with real blood samples.
Abstract: A urea biosensor was developed using the urease entrapped in polyvinyl alcohol (PVA) and polyacrylamide (PAA) composite polymer membrane. The membrane was prepared on the cheesecloth support by gamma-irradiation induced free radical polymerization. The performance of the biosensor was monitored using a flow-through cell, where the membrane was kept in conjugation with the ammonia selective electrode and urea was added as substrate in phosphate buffer medium. The ammonia produced as a result of enzymatic reaction was monitored potentiometrically. The potential of the system was amplified using an electronic circuit incorporating operational amplifiers. Automated data acquisition was carried by connecting the output to a 12-bit analog to digital converter card. The sensor working range was 1-1000 mM urea with a response time of 120 s. The enzyme membranes could be reused 8 times with more than 90% accuracy. The biosensor was tested for blood urea nitrogen (BUN) estimation in clinical serum samples. The biosensor showed good correlation with commercial Infinitytrade mark BUN reagent method using a clinical chemistry autoanalyzer. The membranes could be preserved in phosphate buffer containing dithiothreitol, beta-mercaptoethanol and glycerol for a period of two months without significant loss of enzyme activity.
Abstract: A phosphate solubilizing fungus, Paecilomyces marquandii AA1 was isolated from phosphate deficient soil on Pikovskaya's medium buffered with Tris-HCl pH 8. The organism was identified on the basis of morphological characterization and by sequencing of 18S rRNA gene. The organism could release phosphate from both buffered and unbuffered medium and solubilized rock phosphates from various places. The effect of concentration of ore, temperature, carbon and nitrogen sources on solubilization of rock phosphate was studied. Ammonium salts were the best nitrogen source, followed by asparagine, sodium nitrate, potassium nitrate, urea and calcium nitrate in that order.
Abstract: Denitrification of synthetic high nitrate waste containing 9032 ppm NO(3)-N (40,000 ppm NO(3)) in a time period of only 6h has been achieved in our previous study using activated sludge. The activated sludge culture was acclimatized by a stepwise increase in the nitrate concentration of synthetic waste. In the present work, studies were carried out on the changing microbial population of the sludge and the physiology of nitrate metabolism during the various stages of adaptation process to high strength synthetic nitrate waste. During the course of adaptation, with an increase in the nitrate concentration, a sharp increase in the number of denitrifiers was found with an equally rapid decrease in the nitrifying community. Two key enzymes involved in the first two steps of the denitrification process were also studied during this period. The results of the study suggest that specific enzyme levels increase as the activated sludge adapts itself to higher nitrate concentrations. Biological denitrification of high nitrate waste is a slow process and to increase the rate of denitrification, parameters such as pH, temperature, C:N and biomass concentration of the process were optimized using orthogonal array method. Optimized conditions increased the specific nitrate reduction rate by 54% and specific nitrite reduction rate by 45%.
Abstract: Phytoremediation-the use of plants for cleaning up of xenobiotic compounds-has received much attention in the last few years and development of transgenic plants tailored for remediation will further enhance their potential. Although plants have the inherent ability to detoxify some xenobiotic pollutants, they generally lack the catabolic pathway for complete degradation/mineralization of these compounds compared to microorganisms. Hence, transfer of genes involved in xenobiotic degradation from microbes/other eukaryotes to plants will further enhance their potential for remediation of these dangerous groups of compounds. Transgenic plants with enhanced potential for detoxification of xenobiotics such as trichloro ethylene, pentachlorophenol, trinitro toluene, glycerol trinitrate, atrazine, ethylene dibromide, metolachlor and hexahydro-1,3,5-trinitro-1,3,5-triazine are a few successful examples of utilization of transgenic technology. As more genes involved in xenobiotic metabolism in microorganisms/eukaryotes are discovered, it will lead to development of novel transgenic plants with improved potential for degradation of recalcitrant contaminants. Selection of suitable candidate plants, field testing and risk assessment are important considerations to be taken into account while developing transgenic plants for phytoremediation of this group of pollutants. Taking advantage of the advances in biotechnology and 'omic' technologies, development of novel transgenic plants for efficient phytoremediation of xenobiotic pollutants, field testing and commercialization will soon become a reality.
Abstract: An optical microbial biosensor was described for the detection of methyl parathion pesticide. Whole cells of Flavobacterium sp. were immobilized by trapping in glass fiber filter and were used as biocomponent along with optic fiber system. Flavobacterium sp. has the organophosphorus hydrolase enzyme, which hydrolyzes the methyl parathion into detectable product p-nitrophenol. The immobilized microbial biocomponent was disposable, cost-effective and showed high reproducibility and uniformity. The detection of methyl parathion by the use of disposable microbial biocomponent with optical biosensor was simple, single step and direct measurement of very low quantity of the sample. The home made reaction vessel was small and needed only 75 microl of sample. A lower detection limit 0.3 microM methyl parathion was estimated from the linear range (4-80 microM) of calibration plot of organophosphorus hydrolase enzymatic assay. The applicability to synthetic methyl parathion spiked samples was demonstrated.
Abstract: Bacopa monnieri L. plants exposed to 10, 50, 100 and 200 microM cadmium (Cd) for 48, 96 and 144 h were analysed with reference to the accumulation of metal and its influence on various enzymatic and non-enzymatic antioxidants, thiobarbituric acid reactive substances (TBARS), photosynthetic pigments and protein content. The accumulation of Cd was found to be increased in a concentration and duration dependent manner with more Cd being accumulated in the root. TBARS content of the treated roots and leaves increased with increase in Cd concentration and exposure periods, indicating the occurrence of oxidative stress. Induction in the activities of superoxide dismutase (SOD), ascorbate peroxidase (APX) and guiacol peroxidase (GPX) was recorded in metal treated roots and leaves of B. monnieri. In contrast, a significant reduction in catalase activity in Cd treated B. monnieri was observed. An increase was also noted in the levels of cysteine and non-protein thiol contents of the roots of B. monnieri followed by a decline. However, in leaves, cysteine and non-protein thiol contents were found to be enhanced at all the Cd concentrations and exposure periods. A significant reduction in the level of ascorbic acid was observed in a concentration and duration dependent manner. The total chlorophyll and protein content of B. monnieri decreased with increase in Cd concentration at all the exposure periods. Results suggest that toxic concentrations of Cd caused oxidative damage as evidenced by increased lipid peroxidation and decreased chlorophyll and protein contents. However, B. monnieri is able to combat metal induced oxidative injury involving a mechanism of activation of various enzymatic and non-enzymatic antioxidants.
Abstract: Removal of phenol, a major pollutant in aqueous effluents was studied using plant hairy root cultures. Among four different species of hairy roots tested, Brassica juncea showed the highest potential for phenol removal. The effect of phenol concentration and reuse in a batch system was studied using B. juncea hairy root cultures. Unlike most of the studies reported earlier, phenol removal by the hairy roots was seen to take place without the need for addition of external hydrogen peroxide (H(2)O(2)). To understand the mechanism of phenol removal, levels of peroxidase and phenol oxidase were monitored in the hairy roots. Peroxidase activity in the roots was enhanced when exposed to phenol, while phenol oxidase remained constant. Since peroxidase has a pre-requisite for H(2)O(2), the levels of H(2)O(2) were monitored for its in situ synthesis. H(2)O(2) levels were seen to increase in the presence of phenol. Thus, a mechanism wherein hairy roots also produce H(2)O(2) besides peroxidase, as a protection strategy of plant against xenobiotic stress is plausible.
Abstract: The low-grade rock phosphate of Jhabua, Madhya Pradesh (India), was investigated for its possible application in the removal of lead, copper, zinc and cobalt ions from aqueous solutions. Effects of contact time, amount of adsorbent and initial concentration of metal ions were studied. Adsorption of heavy metal ions was found to follow the order: Pb2+ > Cu2+ > Zn2+ > Co2+. The probable mechanism of metal ions removal by rock phosphate was found to be by its dissolutions followed by subsequent precipitation.
Abstract: Potential of plants to remove radionuclides/toxic elements from soils and solutions can be successfully applied for removal of important radionuclides such as strontium-90 ((90)Sr) and cesium-137 ((137)Cs). When uptake of (137)Cs and (90)Sr by Calotropis gigantea plants incubated in distilled water spiked with the radionuclides either alone or in combination was studied, it was found to have a high efficiency for the removal of (90)Sr, with 90% being removed from solutions (5 x 10(3)kBql(-1)) within 24h of incubation. However, in case of (137)Cs, about 44% could be removed from solutions (5 x 10(3)kBql(-1)) at the end of 168h of incubation. Accumulation of (90)Sr and (137)Cs was higher in roots compared to shoots. The plants could remediate both (90)Sr and (137)Cs when they were added together to the solution. When two months old plants were incubated in low level nuclear waste, 99% of activity disappeared at the end of 15 days. The present study suggests that C. gigantea could be used as a potential candidate plant for phytoremediation of (90)Sr and (137)Cs.
Abstract: A Pseudomonas strain identified as a potent biosorbent of uranium (U) and thorium was immobilized in radiation-induced polyacrylamide matrix for its application in radionuclide containing wastewater treatment. The immobilized biomass exhibited a high U sorption of 202 mg g(-1) dry wt. with its optimum at pH 5.0. A good fit of experimental data to the Freundlich model suggested multilayered uranium binding with an affinity distribution among biomass metal binding sites. Scanning electron microscopy revealed a highly porous nature of the radiation-polymerized beads with bacterial cells mostly entrapped on pore walls. Energy dispersive X-ray analysis (EDXA) coupled with SEM ascertained the accumulation of uranium by the immobilized biomass without any physical damage to the cells. A significant (90%) part of biosorbed uranium was recovered using sodium bicarbonate with the immobilized biomass maintaining their U resorption capacity for multiple sorption-desorption cycles. Uranium loading and elution behavior of immobilized biomass evaluated within a continuous up-flow packed bed columnar reactor showed its effectiveness in removing uranium from low concentration (50 mg U L(-1)) followed by its recovery resulting in a 4-5-fold waste volume reduction. The data suggested the suitability of radiation polymerization in obtaining bacterial beads for metal removal and also the potential of Pseudomonas biomass in treatment of radionuclide containing waste streams.
Abstract: Development of reliable and eco-friendly process for synthesis of metallic nanoparticles is an important step in the filed of application of nanotechnology. One of the options to achieve this objective is to use natural processes such as use of biological systems. In this work we have investigated extracellular biosynthesis of silver nanoparticles using Aspergillus fumigatus. The synthesis process was quite fast and silver nanoparticles were formed within minutes of silver ion coming in contact with the cell filtrate. UV-visible spectrum of the aqueous medium containing silver ion showed a peak at 420 nm corresponding to the plasmon absorbance of silver nanoparticles. Transmission electron microscopy (TEM) micrograph showed formation of well-dispersed silver nanoparticles in the range of 5-25 nm. X-ray diffraction (XRD)-spectrum of the silver nanoparticles exhibited 2theta values corresponding to the silver nanocrystal. The process of reduction being extracellular and fast may lead to the development of an easy bioprocess for synthesis of silver nanoparticles.
Abstract: Immobilized callus cultures of Tinospora cordifolia (Willd) Miers ex Hooks and Thoms were investigated to find out the combined effect of elicitation, cell permeabilization with chitosan and in situ product recovery by polymeric neutral resin-like Diaion HP 20. In this study, callus cultures of T. cordifolia were immobilized using sodium alginate and calcium chloride and the beads were cultured in Murashige and Skoog's basal medium along with benzyl adenine (BA), 2,4-dichlorophenoxy acetic acid (2,4-D) and 3% sucrose. The immobilized cultures, when subjected to elicitation and cell permeabilization with chitosan and in situ removal of the secondary metabolites by addition of resin, showed a 10-fold increase in production of arabinogalactan (0.490% dry weight) as compared to respective controls devoid of resin and chitosan. This indicates that in situ adsorption may have reduced the feedback inhibition caused by accumulation of secondary metabolites in the media, while the dual effect of elicitation and cell permeabilization by chitosan may have released the intracellular (secreted) berberine and the polysaccharide arabinogalactan, respectively.
Abstract: Bioremediation is gaining a lot of importance in recent times as an alternate technology for removal of elemental pollutants in soil and water, which require effective methods of decontamination. Phytoremediation--the use of green plants to remove, contain or render harmless environmental pollutants--may offer an effective, environmentally nondestructive and cheap remediation method. The use of genetic engineering to modify plants for metal uptake, transport and sequestration may open up new avenues for enhancing efficiency of phytoremediation. Metal chelator, metal transporter, metallothionein (MT), and phytochelatin (PC) genes have been transferred to plants for improved metal uptake and sequestration. Transgenic plants, which detoxify/accumulate cadmium, lead, mercury, arsenic and selenium have been developed. A better understanding of the mechanisms of rhizosphere interaction, uptake, transport and sequestration of metals in hyperaccumulator plants will lead to designing novel transgenic plants with improved remediation traits. As more genes related to metal metabolism are discovered, facilitated by the genome sequencing projects, new vistas will be opened up for development of efficient transgenic plants for phytoremediation.
Abstract: Composite polymer membrane of polyvinyl alcohol (PVA) and acrylamide was prepared on cheesecloth support by gamma-irradiation induced free radical polymerization. The enzyme urease was entrapped in the membrane during polymerization and was cross-linked within the matrix using glutaraldehyde. The membranes could be reused a number of times without significant loss of urease activity.
Abstract: Polysaccharides bound to bacteria or in isolated form have been shown to bind heavy metals. A limitation of this technology can be overcome by immobilization. In view of this Ocimum basilicum seeds which swell upon wetting could serve as natural immobilized source of agriculturally-based polysaccharides. The seeds consist of an inner hard core and a pectinous fibrillar outer layer. Pretreating the seeds with acid, alkali, periodate or boiling in water was found to alter the metal binding capacity. Of the various treatments given, seeds boiled in water were found to be superior in terms of mechanical stability and exhibited fairly optimal Cr(VI) uptake kinetics. The maximum adsorption capacity as calculated from the Langmuir isotherm was 205 mg Cr/g dry seeds. Biosorption of Cr(VI) was found to be pH dependent with maximum uptake at pH 1.5 wherein sorption was not affected by the presence of other metal ions such as Cd(2+), Cu(2+), Ca(2+) and Na(+). Seeds were used in a packed bed reactor for the continuous removal of Cr(VI). Thus O. basilicum seeds may have application as a potential bioresource in tropical countries such as India where they are widely available.
Abstract: An immobilized preparation of whole cell-based catalase was obtained by cross-linking the yeast cells permeabilized with toluene in hen egg white using glutaraldehyde. Optimal preparations were obtained when cross-linking was carried out for 2 h at 4 degrees C. Immobilized cells could be reused for the removal of H2O2 from milk.
Abstract: Glucoamylase was immobilized on polyethylenimine (PEI)-coated cotton cloth by adsorption followed by cross-linking with 0.2% glutaraldehyde in the presence of starch. Optimal adsorption of the enzyme was seen when cloth treated with 2% PEI was contacted with the enzyme for 50 min. pH and temperature optima profiles were not changed appreciably on immobilization. However, the bound enzyme exhibited a higher thermal stability. The enzyme-bound cloth strips were used in a specially designed bioreactor for the continuous hydrolysis of starch. The reactor could be operated for over 21 days retaining about 70% of the original activity. An operational temperature of 45 degrees C was found to be optimal.
Abstract: Washed and dried rice husk was coated with 2% polyethylenimine (PEI). Invertase was immobilized on this support through adsorption followed by cross-linking with 2% glutaraldehyde. Immobilized enzyme was reused for the hydrolysis of sucrose without loss in activity. This approach may serve as a simple technique in the future for the covalent immobilization of enzymes on lignocellulosic supports.
Abstract: A microbial biosensor consists of a transducer in conjunction with immobilised viable or non-viable microbial cells. Non-viable cells obtained after permeabilisation or whole cells containing periplasmic enzymes have mostly been used as an economical substitute for enzymes. Viable cells make use of the respiratory and metabolic functions of the cell, the analyte to be monitored being either a substrate or an inhibitor of these processes. Bioluminescence-based microbial biosensors have also been developed using genetically engineered microorganisms constructed by fusing the lux gene with an inducible gene promoter for toxicity and bioavailability testing. In this review, some of the recent trends in microbial biosensors with reference to the advantages and limitations are been discussed. Some of the recent applications of microbial biosensors in environmental monitoring and for use in food, fermentation and allied fields have been reviewed. Prospective future microbial biosensor designs have also been identified.
Abstract: Uranium uptake by dried roots of Eichhornia crassipes was rapid and the biomass could remove 54% of the initial uranium present within 4 min of contact time. The process was favored at pH 5-6 and was least influenced by temperature. Biosorption data fitted to both Langmuir and Freundlich isotherm. The maximum loading capacity obtained was 371 mg U/g dry biomass. Distribution coefficient of 9336 ml/g was observed at a residual concentration of 4.9 mg U/L. Uptake increased at higher dose of biomass and reached a plateau beyond the concentration of 6 g/L. The specific metal ion uptake decreased with increasing initial uranium concentration. Anions (0.1 M) inhibited the uptake and followed the trend acetate > sulphate > nitrate > chloride. However presence of carbonate had no effect on uranium biosorption.
Abstract: Biosensors are finding applications in a variety of analytical fields. A biosensor basically consists of a transducer in conjunction with a biologically active molecule that converts a biochemical signal into a quantifiable electric response. The specificity of the biosensor depends on the selection of the biomaterial. Enzymes, antibodies, DNA, receptors, organelles, microorganisms as well as animal and plant cells or tissues have been used as biologic sensing materials. Advances in biochemistry, molecular biology, and immunochemistry are expected to lead to a rapid expansion in the range of biologic recognition elements to be used in the field of biosensors. Biomaterials that are stable and function even in highly acidic, alkaline, hydrophobic, or oxidizing environments as well as stable to high temperature and immune to toxic substrates in the processing stream will play an important role. Techniques for immobilization of the biomaterials have played a significant role in the biosensor field. Immobilization not only brings about the intimate contact of the biologic catalysts with the transducer, but also helps in the stabilization of the biologic system, thus enhancing its operational and storage stability. A number of techniques have been developed in our laboratory for the immobilization of enzymes, multienzyme systems, cells, and enzyme-cell conjugates. Some of these aspects that are of significance in biosensor applications have been highlighted.
Abstract: Crude cell-free extract of yeast cells was mixed with sufficient amount of Jack bean meal extract so as to precipitate all the invertase. The precipitate was then cross-linked using 2% glutaraldehyde retaining over 60% of the activity. The immobilized invertase could be reused for over ten batches without loss in activity.
Abstract: Lysozyme naturally present in raw hen egg white was immobilized by cross-linking the egg white foam with glutaraldehyde. Inclusion of N-acetyl glucosamine, a competitive inhibitor of lysozyme, was found to enhance the yield of lysozyme activity by fivefold.
Abstract: A technique has been described for the stabilization of calcium alginate beads using radiation polymerized acrylamide. The technique involved dropping a mixture containing the cells (20%), sodium alginate (2%), acrylamide (2.5%) and N-N'-methylene-bis-acrylamide (0.1%) through a syringe needle into cold (-75 degrees C) toluene. The frozen beads obtained were exposed to 60Co gamma-rays (0.5 KGy) and were then thawed in 0.1 M CaCl2 solution. Unlike the calcium alginate beads the conjugate beads were not found to be dissolved when incubated in 3% trisodium citrate solution. Stabilized beads containing entrapped yeast cells could be reused for over 15 batches for the inversion of sucrose without loss in activity or chemical integrity of the beads.
Abstract: The cells of Haloarcula vallismortis, an extreme halophilic archaebacterium, were permeabilized by various chemical, physical, and biological treatments. Biological permeabilization by lysozyme and papain showed effective results as observed by studying the in situ activity of halophilic glyceraldehyde-3-phosphate dehydrogenase (hGAPDH) as the model enzyme. Detergents N-cetyl-N, N, N-trimethyl ammonium bromide (CTAB) and digitonin also showed significant results. Other strains of halobacteria could also be permeabilized by lysozyme. The cell morphology did not show any significant change after permeabilization as observed by phase contrast microscopy. The enzyme characteristics of hGAPDH were studied in situ using permeabilized H. vallismortis cells. The properties, like optimum pH, Km for GAP and NAD(+), inhibition by heavy metals, sulphydryl reagents, and other compounds, showed remarkable similarity with those studied in vitro.
Abstract: Halobacteria require 20-25% NaCl for optimal growth and lyse when the salt concentration falls below 10%. The response of Haloferax mediterranei cells to low concentrations of NaCl (< 20%) in the medium was studied. The cells adapted to and grew in concentrations of NaCl as low as 10% and survived in concentrations lower than 5%. The cells synthesised a red pigment, bacterioruberin, in response to stress caused by a low concentration of NaCl (< 20%).
Abstract: A novel technique has been described for the immobilization of lysozyme, naturally present in hen egg white by cross-linking the egg white foam with glutaraldehyde. This technique results in a mechanically stable and porous matrix exhibiting about 6-times the lytic activity against Micrococcus lysodeikticus cells, as compared to the unfoamed matrix. Foamed egg white matrix can be used for the continuous lysis of bacterial cells.
Abstract: Yeast invertase, a glycoprotein, was covalently coupled to Ocimum basilicum seeds either through its protein or carbohydrate moiety. Of the various methods investigated, binding of the enzyme through its carbohydrate moiety resulted in the retention of considerably higher amounts of enzyme activity. Immobilized invertase showed a shift in the pH optimum toward the alkaline side without appreciable change in temperature optimum. However, the immobilized preparation was more thermostable than the free enzyme. Invertase bound to the seeds could be used repeatedly for the hydrolysis of sucrose syrups in a batch process without appreciable loss in activity. The seeds could serve as an inexpensive, ready-to-use, natural pellicular polysaccharide support for immobilizing enzymes.
Abstract: The cells of Haloferax mediterannei were stabilized by cross-linking with 0.5% glutaraldehyde for 10 min. Such cells were found to be osmotically stable even when suspended in water. The stabilized cells could be permeabilized by treatment with chloroform without leakage of intracellular components. No significant difference in the properties of an intracellular enzyme aldolase was observed, using either cell-free extract or the osmotically stabilized and permeabilized cells. This novel technique can serve as a useful tool for studying in situ regulatory characteristics of intracellular functions in halobacteria and can also help in their re-use under more stabilized conditions for biotechnological applications.
Abstract: A method has been described for obtaining coimmobilizates by the simultaneous binding of glucose oxidase to the cell and the enzyme-bound cell to cotton thread through adhesion using polyethylenimine (PEI). Glucose oxidase was found to adsorb onto PEI-coated yeast cells from a water suspension. The desorption observed at higher ionic strength could be obviated by cross-linking with 2% glutaraldehyde for 2 min. The enzyme-bound yeast cells could then be immobilized by adhesion on cotton thread. The coimmobilizate could be reused for over 10 batches without appreciable loss in activity.
Abstract: Yeast invertase was immobilized on polyethyleneimine-coated cotton thread by adsorption followed by crosslinking with glutaraldehyde. The thread-bound invertase was used as an easily retrievable system for the hydrolysis of 80% w/v commercial sucrose syrups. The immobilized enzyme was stable for over 90 days to a temperature of 50 degrees C, only when stored in 80% sucrose solution. Above this temperature, inactivation of enzyme was observed. The cotton threads were used in a batch reactor for hydrolysis of sucrose in about 30 batches carried out over a period of 50 days without loss in activity. The threads could also be used in a packed bed reactor (1.51) for 97% hydrolysis of 80% sucrose syrups at 50 degrees C at a rate of about 360 kg per month for a period of 3 months.
Abstract: Immobilized beta-galactosidase was obtained by crosslinking the enzyme with hen egg white using 2% glutaraldehyde. The gel obtained could be lyophilized to give a dry enzyme powder. The pH optimum of both the soluble and immobilized enzyme was found to be 6.8. The immobilized enzyme showed a higher K(m) for the substrates. The extent of enzyme inhibition by galactose was reduced upon immobilization. The stability towards inactivation by heat, urea, gamma irradiation, and protease treatment were enhanced. The bound enzyme as tested in a batch reactor could be used repeatedly for the hydrolysis of milk lactose. The possible application of this system for small-scale domestic use has been suggested.
Abstract: A stable immobilized preparation of fumarase (EC 4.2.1.2) was obtained by entrapment of rat liver mitochondria in acrylamide polymerized by using gamma irradiation (100 kR). The enhanced stability and the efficiency of the entrapped enzyme have shown potential for repeated use for the production of L-malic acid from fumaric acid. The possible formation of succinic acid in the system could be controlled by incorporating malonate along with detergents such as sodium deoxycholate or sodium dodecylsulfate in the reactor system.
