Qayyum Husain

Potential applications of enzymes immobilized on/in nano materials: A review

Several new types of carriers and technologies have been implemented in the recent past to improve traditional enzyme immobilization which aimed to enhance enzyme loading, activity and stability to decrease the enzyme biocatalyst cost in industrial biotechnology. These include cross-linked enzyme aggregates, microwave-assisted immobilization, click chemistry technology, mesoporous supports and most recently nanoparticle-based immobilization of enzymes. The union of the specific physical, chemical, optical and electrical properties of nanoparticles with the specific recognition or catalytic properties of biomolecules has led to their appearance in myriad novel biotechnological applications. They have been applied time and again for immobilization of industrially important enzymes with improved characteristics. The high surface-to-volume ratio offered by nanoparticles resulted in the concentration of the immobilized entity being considerably higher than that afforded by experimental protocols based on immobilization on planar 2-D surfaces. Enzymes immobilized on nanoparticles showed a broader working pH and temperature range and higher thermal stability than the native enzymes. Compared with the conventional immobilization methods, nanoparticle based immobilization served three important features; (i) nano-enzyme particles are easy to synthesize in high solid content without using surfactants and toxic reagents, (ii) homogeneous and well defined core-shell nanoparticles with a thick enzyme shell can be obtained, and (iii) particle size can be conveniently tailored within utility limits. In addition, with the growing attention paid to cascade enzymatic reaction and in vitro synthetic biology, it is possible that co-immobilization of multi-enzymes could be achieved on these nanoparticles.

Potential Applications of the Oxidoreductive Enzymes in the Decolorization and Detoxification of Textile and Other Synthetic Dyes from Polluted Water: A Review

ABSTRACT Recently, the enzymatic approach has attracted much interest in the decolorization/degradation of textile and other industrially important dyes from wastewater as an alternative strategy to conventional chemical, physical and biological treatments, which pose serious limitations. Enzymatic treatment is very useful due to the action of enzymes on pollutants even when they are present in very dilute solutions and recalcitrant to the action of various microbes participating in the degradation of dyes. The potential of the enzymes (peroxidases, manganese peroxidases, lignin peroxidases, laccases, microperoxidase-11, polyphenol oxidases, and azoreductases) has been exploited in the decolorization and degradation of dyes. Some of the recalcitrant dyes were not degraded/decolorized in the presence of such enzymes. The addition of certain redox mediators enhanced the range of substrates and efficiency of degradation of the recalcitrant compounds. Several redox mediators have been reported in the literature, but very few of them are frequently used (e.g., 1-hydroxybenzotriazole, veratryl alcohol, violuric acid, 2-methoxy-phenothiazone). Soluble enzymes cannot be exploited at the large scale due to limitations such as stability and reusability. Therefore, the use of immobilized enzymes has significant advantages over soluble enzymes. In the near future, technology based on the enzymatic treatment of dyes present in the industrial effluents/wastewater will play a vital role. Treatment of wastewater on a large scale will also be possible by using reactors containing immobilized enzymes.

β Galactosidases and their potential applications: a review

β Galactosidases have been obtained from microorganisms such as fungi, bacteria and yeasts; plants, animals cells, and from recombinant sources. The enzyme has two main applications; the removal of lactose from milk products for lactose intolerant people and the production of galactosylated products. In order to increase their stability, reusability, and use in continuous reactors, these enzymes have been immobilized on both organic and inorganic support via adsorption, covalent attachment, chemical aggregation, microencapsulation, and entrapment. Free and immobilized preparations of β galactosidases have been exploited in various applications such as industrial, biotechnological, medical, analytical, and in different other applications. β galactosidase is widely used in food industry to improve sweetness, solubility, flavor, and digestibility of dairy products. Immobilized β galactosidases are employed for the continuous hydrolysis of lactose from whey and milk in a number of reactors such as hollow fiber reactors, tapered column reactors, packed bed reactors, fluidized bed reactors etc.

Applications of Redox Mediators in the Treatment of Organic Pollutants by Using Oxidoreductive Enzymes: A Review

An enzymatic approach has attracted much interest in the remediation/degradation of various organic pollutants present in the wastewater coming out of industries. The treatment of several pollutants sometimes causes problems due to the recalcitrant nature of the compounds. However, these recalcitrant compounds were degraded/transformed by enzymes in the presence of certain redox mediators. These redox mediators enhanced the range of substrates and efficiency of degradation of the recalcitrant compounds by severalfold. Enzymes whose potential has been exploited for this purpose are laccases, lignin peroxidases, manganese peroxidases, horseradish peroxidases, turnip peroxidases, bitter gourd peroxidases, and others. Several redox mediators have been reported in the literature but very few of them are frequently used, for example, 1-hydroxybenzotriazole, veratryl alcohol, violuric acid, 2-methoxy-phenothiazone, 3-hydroxyanthranilic acid, anthraquinone 2,6-disulfonic acid, 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), N-hydroxyacetanilide, phenol red, 3,3′,5,5′-tetramethyl benzidine; dichlorophenol red, 2,2′,6,6′-tetramethylpiperidine-N-oxyl radical, syringaldehyde and acetosyringone, and others. In the near future the enzyme–redox mediator combined treatment of aromatic compounds present in the industrial effluents/wastewater will play a vital role. Use of the enzyme–redox mediator system certainly will enhance the chances of remediation of a wide spectrum of aromatic compounds present in various industrial effluents.

Concanavalin A: A useful ligand for glycoenzyme immobilization—A review

Concanavalin A is finding increasing applications as a useful ligand in glycoenzyme immobilization. An attempt therefore, has been made to summarize the work available in the area. Glycoenzymes that are recalcitrant to immobilization procedures involving covalent coupling to solid supports can be immobilized in high yields by binding to matrices precoupled with concanavalin A. In addition, glycoenzymes associated with concanavalin A matrices usually exhibit high retention of activity and enhanced stability against various forms of inactivation. Binding of the glycoenzymes on the concanavalin A supports, being noncovalent, can be reversed by incubating the preparation with a high concentration of sugars/glycosides or at acidic pH. The association can be, however, rendered covalent by crosslinking the preparations with bifunctional reagents like glutaraldehyde. Crosslinking may be accompanied by further increase in stability, albeit at the expense of the loss of some enzyme activity. Several laboratory-size reactors containing concanavalin A matrix-bound glycoenzyme have been successfully operated for reasonably long durations with only small losses in catalytic activity. Insoluble glycoenzyme preparation can also be obtained by precipitating them from solution as concanavalin A complexes. Such complexes have small particle dimensions but can be successfully used in column reactors after a subsequent immobilization step. Insoluble concanavalin A-flocculates containing various microorganisms and glycoenzymes that successfully carry out multistep transformations have also been obtained by several investigators.

Immobilization of Aspergillus oryzae β galactosidase on zinc oxide nanoparticles via simple adsorption mechanism.

The present study demonstrates the immobilization of Aspergillus oryzae β galactosidase on native zinc oxide (ZnO) and zinc oxide nanoparticles (ZnO-NP) by simple adsorption mechanism. The binding of enzyme on ZnO-NP was confirmed by Fourier transform-infrared spectroscopy and atomic force microscopy. Native ZnO and ZnO-NP showed 60% and 85% immobilization yield, respectively. Soluble and immobilized enzyme preparations exhibited similar pH-optima at pH 4.5. ZnO-NP bound β galactosidase retained 73% activity at pH 7.0 while soluble and ZnO adsorbed enzyme lost 68% and 53% activity under similar experimental conditions, respectively. There was a marked broadening in temperature-activity profile for ZnO-NP adsorbed β galactosidase; it showed no difference in temperature-optima between 50°C and 60°C. Moreover, ZnO-NP adsorbed β galactosidase retained 53% activity after 1h incubation with 5% galactose while the native ZnO- and soluble β galactosidase exhibited 35% and 28% activity under similar exposure, respectively. Native ZnO and ZnO-NP adsorbed β galactosidase retained 61% and 75% of the initial activity after seventh repeated use, respectively. It was noticed that 54%, 63% and 71% milk lactose was hydrolyzed by soluble, ZnO adsorbed and ZnO-NP adsorbed β galactosidase in batch process after 9h while whey lactose was hydrolyzed to 61%, 68% and 81% under similar experimental conditions, respectively. In view of its easy production, improved stability against various denaturants and excellent reusability, ZnO-NP bound β galactosidase may find its applications in constructing enzyme-based analytical devices for clinical, environmental and food technology.

Remediation and treatment of organopollutants mediated by peroxidases: a review

In this paper an effort has been made to review the literature on the role of peroxidases in the remediation and treatment of a wide spectrum of aromatic pollutants. Peroxidases can catalyse degradation/transformation of polycyclic aromatic hydrocarbons, polychlorinated biphenyls, organochlorines, 2,4,6-trinitrotoluene, phenolic compounds and dyes. These enzymes are also capable of treating various types of recalcitrant aromatic compounds in the presence of redox mediators. Immobilised peroxidases from plant and fungal sources have been used for the remediation of such types of industrial pollutants on a large scale.In this paper an effort has been made to review the literature on the role of peroxidases in the remediation and treatment of a wide spectrum of aromatic pollutants. Peroxidases can catalyse degradation/transformation of polycyclic aromatic hydrocarbons, polychlorinated biphenyls, organochlorines, 2,4,6-trinitrotoluene, phenolic compounds and dyes. These enzymes are also capable of treating various types of recalcitrant aromatic compounds in the presence of redox mediators. Immobilised peroxidases from plant and fungal sources have been used for the remediation of such types of industrial pollutants on a large scale.

Concanavalin A layered calcium alginate–starch beads immobilized β galactosidase as a therapeutic agent for lactose intolerant patients

A novel therapeutic agent in the form of beta galactosidase immobilized on the surface of concanavalin A layered calcium alginate-starch beads has been developed. Immobilized beta galactosidase exhibited significantly very high stability against conditions of digestive system such as pH, salivary amylase, pepsin and trypsin. Soluble and immobilized beta galactosidase exhibited same pH-optima. However, the immobilized enzyme retained greater fraction of catalytic activity at higher and lower pH to pH-optima as compared to soluble enzyme. Immobilized enzyme preparation was quite stable under conditions present in mouth, stomach and intestine. Immobilized beta galactosidase retained 65% activity even after its sixth repeated use.

Relation of oxidant-antioxidant imbalance with disease progression in patients with asthma

CONTEXT: Asthma is a chronic airway disorder which is associated to the inflammatory cells. Inflammatory and immune cells generate more reactive oxygen species in patients suffering from asthma which leads to tissue injury. AIMS: To investigate the role of oxidant-antioxidant imbalance in disease progression of asthmatic patients. SETTINGS AND DESIGN: In this study, 130 asthmatic patients and 70 healthy controls were documented. METHODS: For this malondialdehyde level, total protein carbonyls, sulfhydryls, activity of superoxide dismutase (SOD), catalase, glutathione peroxidase (GPx), total blood glutathione, and total antioxidant capacity (FRAP) were measured. STATISTICAL ANALYSIS USED: Analysis of the data was done using unpaired student t test and one-way ANOVA analysis. P < 0.05 was considered significant. RESULTS: The present work showed that the systemic levels of MDA (4.19 ± 0.10 nmol/ml, P < 0.001) and protein carbonyls (1.13 ± 0.02 nmol/mg, P < 0.001) were found to be remarkably higher in asthmatic patients while protein sulfhydryls (0.55 ± 0.01 mmol/l, P < 0.05) decreased as compared to controls (2.84 ± 0.12 nmol/ml, 0.79 ± 0.02 nmol/mg and 0.60 ± 0.02 mmol/l, respectively). We also observed decrease in activities of SOD (2047 ± 50.34 U/g Hb, P < 0.05), catalase (4374 ± 67.98 U/g Hb, P < 0.01), and GPx (40.97 ± 1.05 U/g Hb, P < 0.01) in erythrocytes compared to control (2217 ± 60.11 U/g Hb, 4746 ± 89.94 U/g Hb, and 48.37 ± 2.47 U/g Hb, respectively). FRAP level (750.90 ± 21.22 μmol/l, P < 0.05) in plasma was decreased, whereas total blood glutathione increased (0.94 ± 0.02 mmol/l, P < 0.05) as seen in control (840.40 ± 28.39 μmol/l and 0.84 ± 0.04 mmol/l). CONCLUSIONS: This work supports and describes the hypothesis that an imbalance between oxidant-antioxidant is associated to the oxidative stress which plays a significant role in severity of the disease.

Designing and surface modification of zinc oxide nanoparticles for biomedical applications.

The present study aimed to work out a simple and high yield procedure for the immobilization of β galactosidase on bioaffinity support, concanavalin A (Con A) layered zinc oxide nanoparticles (ZnO-NP). Thermogravimetric analysis of bioaffinity support revealed 4% loss in weight at 600°C whereas its thermal decomposition was observed at 530°C by differential thermal analysis. No significant change was noticed in the band intensity of pUC19 plasmid after its treatment with Con A layered ZnO-NP. Comet assay further exhibited negligible change in tail length of comet after treating the lymphocytes by bioaffinity matrix. The bioaffinity matrix binds 89% of the enzyme activity. Atomic force microscopy analysis showed that the prepared matrix has an advantageous microenvironment and large surface area for binding significant amount of the enzyme. The functional groups present in native and parent compound were monitored by Fourier transform-infrared spectroscopy. Michaelis constant, K(m) was 2.38 and 5.88 mM for free and immobilized β galactosidase, respectively. V(max) for the soluble and immobilized enzyme was 0.520 mM/min and 0.460 mM/min, respectively. Concanavalin A layered ZnO-NP bound β galactosidase exhibited a shift in the temperature-optima and retained nearly 86% activity even after its 6th repeated use.