Immobilization Of Dextranase Research Articles

The hydroxyapatite nanowires as high loading carrier for immobilization of dextranase by crosslinking method

The hydroxyapatite nanowires as high loading carrier for immobilization of dextranase by crosslinking method

Immobilization of Dextranase Obtained from the Marine Cellulosimicrobium sp. Y1 on Nanoparticles: Nano-TiO2 Improving Hydrolysate Properties and Enhancing Reuse.

Dextranase is widely used in sugar production, drug synthesis, material preparation, and biotechnology, among other fields. The immobilization of dextranase using nanomaterials in order to make it reusable, is a hot research topic. In this study, the immobilization of purified dextranase was performed using different nanomaterials. The best results were obtained when dextranase was immobilized on titanium dioxide (TiO2), and a particle size of 30 nm was achieved. The optimum immobilization conditions were pH 7.0, temperature 25 °C, time 1 h, and immobilization agent TiO2. The immobilized materials were characterized using Fourier-transform infrared spectroscopy, X-ray diffractometry, and field emission gun scanning electron microscopy. The optimum temperature and pH of the immobilized dextranase were 30 °C and 7.5, respectively. The activity of the immobilized dextranase was >50% even after 7 times of reuse, and 58% of the enzyme was active even after 7 days of storage at 25 °C, indicating the reproducibility of the immobilized enzyme. The adsorption of dextranase by TiO2 nanoparticles exhibited secondary reaction kinetics. Compared with free dextranase, the hydrolysates of the immobilized dextranase were significantly different, and consisted mainly of isomaltotriose and isomaltotetraose. The highly polymerized isomaltotetraose levels could reach >78.69% of the product after 30 min of enzymatic digestion.

Immobilization of Dextranase on Nano-Hydroxyapatite as a Recyclable Catalyst.

The immobilization technology provides a potential pathway for enzyme recycling. Here, we evaluated the potential of using dextranase immobilized onto hydroxyapatite nanoparticles as a promising inorganic material. The optimal immobilization temperature, reaction time, and pH were determined to be 25 °C, 120 min, and pH 5, respectively. Dextranase could be loaded at 359.7 U/g. The immobilized dextranase was characterized by field emission gun-scanning electron microscope (FEG-SEM), X-ray diffraction (XRD), and Fourier-transformed infrared spectroscopy (FT-IR). The hydrolysis capacity of the immobilized enzyme was maintained at 71% at the 30th time of use. According to the constant temperature acceleration experiment, it was estimated that the immobilized dextranase could be stored for 99 days at 20 °C, indicating that the immobilized enzyme had good storage properties. Sodium chloride and sodium acetic did not desorb the immobilized dextranase. In contrast, dextranase was desorbed by sodium fluoride and sodium citrate. The hydrolysates were 79% oligosaccharides. The immobilized dextranase could significantly and thoroughly remove the dental plaque biofilm. Thus, immobilized dextranase has broad potential application in diverse fields in the future.

A Comparative Study Among Different Protocols of Immobilization of Dextranase Using Chitin as a Matrix

In the current research, second most abundant renewable polysaccharide chitin is utilized for meaningful purpose. Owing to auspicious features of chitin it is used as a matrix for immobilization of valuable biocatalyst dextranase. Dextranase belongs to hydrolase family and has broad commercial application in different fields. Functionality of dextranase is further improve with the help of its immobilization on chitin by different protocols. Dextranase used in this study was isolated from a thermophilic bacteria Bacillus megaterium KIBGE-IB31 explored from hydrothermal spring. Isolated dextranase was immobilized on chitin by two different methods namely; adsorption and covalent binding. A comparative study was conducted between soluble, adsorbed and covalently cross linked dextranase. It was observed that although, there was slight deviation in characteristics of dextranase after immobilization but there was marked improvement in stability of enzyme after immobilization. The comparative analysis revealed that dextranase immobilized by adsorption was less stable and fails to retained enzyme stability for increase reusability. The adsorption method was improved after treatment of chitin with acid and showed improved stability as well as reusability. However, dextranase immobilized by covalent cross linking displayed highest stability at high temperature along with increase recycling efficiency as compare to enzyme immobilized by adsorption method. Hence, it can be concluded that though chitin is cheap, easily accessible matrix for immobilization of dextranase but still among different protocols of immobilization covalent cross linking was found to be more appropriate in improving enzyme stability and its reusability at industrial scale.

Immobilization of Dextranase Using Anionic Natural Polymer Alginate as a Matrix for the Degradation of a Long-Chain Biopolymer (Dextran)

Alginate is an inexpensive, nontoxic, valuable biopolymer utilized in the study for the immobilization of commercially applicable biocatalyst dextranase. Dextranase was immobilized by an entrapment method, and alginate hydrogel spheres were synthesized after optimizing several parameters. A sodium alginate concentration of 4.0% was noticed to be suitable along with a calcium chloride concentration of 0.2 molar after providing a curing time of 20 minutes. After comparing the characteristics of the entrapped enzyme with those of the soluble one, it was observed that the characteristics were more or less the same except for the change in reaction time which was noticed to be prolonged in the case of entrapped dextranase while the change in temperature and pH optima was not observed. The variation inVmaxandKmvalues of dextranase after entrapment was also noted. However, after extensive stability examination studies, it was found that dextranase became more stable after entrapment; as a result, it retained more than 50% of its original activity at elevated temperature even after exposure for about 2.0 hours. The reusability of dextranase was up to 7.0 cycles after performing catalytic activity under constant condition.

Hydrophilic polyethylenimine modified magnetic graphene oxide composite as an efficient support for dextranase immobilization with improved stability and recyclable performance

Development of appropriate support for enzyme immobilization with approving loading capacity, stability and recyclable performance is a significant topic in enzyme chemistry. In this study, the hydrophilic polyethylenimine (PEI) modified magnetic graphene oxide (GO) composite (Fe3O4@PDA@GO-PEI) was fabricated and acted as an efficient solid support for the immobilization of dextranase to improve the stability and recyclable performance. The GO nanosheet component not only provided abundant binding sites for decorating PEI molecules and further increasing the loading capacity of immobilized dextranase, but also acted as a soft matrix for enhancing the stability of immobilized dextranase. In addition, in combination of the unique magnetic separation property, the recyclable ability of immobilized dextranase was further improved. The loading amount of immobilized dextranase on as-developed Fe3O4@PDA@GO-PEI was determined to be 305.1 mg g−1, which was 1.61 times that on the Fe3O4@PDA-PEI without GO component. Moreover, the as-developed Fe3O4@PDA@GO-PEI-dextranase exhibited high immobilization efficiency (76.27%) and enzymatic activity (83.67%), improved stabilities in wide temperature conditions and pH solutions, and excellent reusability (85.78% of its initial activity after 20 cycles). These experimental results suggest that the immobilized dextranase is promising for various applications in biocatalytic engineering fields.

Chitosan hydrogel microspheres: an effective covalent matrix for crosslinking of soluble dextranase to increase stability and recycling efficiency.

Dextranase is a unique biocatalyst that has high specificity and stereo-selectivity towards a complex biopolymer known as dextran. Dextranase has wide industrial application, but most of the time harsh environmental conditions adversely affect the functionality and stability of the enzyme. To overcome this issue, a covalent cross-linking immobilization method was adapted in the current study utilizing a nontoxic and biocompatible matrix known as chitosan. Chitosan hydrogel microspheres were synthesized using chitosan which exhibited noteworthy physical and mechanical strength. After treatment with glutaraldehyde, chitosan hydrogel microspheres were used for immobilization of dextranase. The kinetic characteristics of immobilized dextranase were compared with that of the soluble enzyme. A shift in optimum pH and temperature from 7.0 to 7.5 and 50 to 60°C was observed after immobilization, respectively. Recycling efficiency, thermal stability, and activation energy distinctly improved after immobilization, whereas anchoring of substrate at the active site of the soluble dextranase exhibited an increase in K m with no change in V max after crosslinking. This technique involves the reduction in the size of carrier molecules (microspheres) that provide a larger surface area for improved immobilization efficiency. Therefore, it is concluded that increased stability and reusability of this immobilized biocatalyst makes it a promising aspirant for the utilization at commercial level.

Combined of ultrasound irradiation with high hydrostatic pressure (US/HHP) as a new method to improve immobilization of dextranase onto alginate gel

In this research work, dextranase was immobilized onto calcium alginate beads by the combination of ultrasonic irradiation and high hydrostatic pressure (US/HHP) treatments. Effects of US/HHP treatments on loading efficiency and immobilization yield of dextranase enzyme onto calcium alginate beads were investigated. Furthermore, the activities of immobilized enzymes prepared with and without US/HHP treatments and that prepared with ultrasonic irradiation (US) and high hydrostatic pressure (HHP), as a function of pH, temperature, recyclability and enzyme kinetic parameters, were compared with that for free enzyme. The maximum loading efficiency and the immobilization yield were observed when the immobilized dextranase was prepared with US (40W at 25kHz for 15min) combined with HHP (400MPa for 15min), under which the loading efficiency and the immobilization yield increased by 88.92% and 80.86%, respectively, compared to immobilized enzymes prepared without US/HHP treatment. On the other hand, immobilized enzyme prepared with US/HHP treatment showed Vmax, KM, catalytic and specificity constants values higher than that for the immobilized enzyme prepared with HHP treatment, indicated that, this new US/HHP method improved the catalytic kinetics activity of immobilized dextranase at all the reaction conditions studied. Compared to immobilized enzyme prepared either with US or HHP, the immobilized enzymes prepared with US/HHP method exhibited a higher: pH optimum, optimal reaction temperature, thermal stability and recyclability, and lower activation energy, which, illustrating the effectiveness of the US/HHP method. These results indicated that, the combination of US and HHP treatments could be an effective method for improving the immobilization of enzymes in polymers.

Ultrasound-assisted dextranase entrapment onto Ca-alginate gel beads

In this research work, dextranase has immobilized onto calcium alginate beads using a novel ultrasound method. The process of immobilization of the enzyme was carried out in a one-step ultrasound process. Effects of ultrasound conditions on loading efficiency and immobilization yield of the enzyme onto calcium alginate beads were investigated. Furthermore, the activity of the free and immobilized enzymes prepared with and without ultrasound treatment, as a function of pH, temperature, recyclability and enzyme kinetic parameters, was compared. The maximum loading efficiency and the immobilization yield were observed when the immobilized dextranase was prepared with an ultrasonic irradiation at 25kHz, 40W for 15min, under which the loading efficiency and the immobilization yield increased by 27.21% and 18.77%, respectively, compared with the immobilized enzymes prepared without ultrasonic irradiation. On the other hand, immobilized enzyme prepared with ultrasonic irradiation showed Vmax and KM value higher than that for the immobilized enzyme prepared without ultrasonic irradiation, likewise, both the catalytic and specificity constants of immobilized enzyme prepared with ultrasonic irradiation were higher than that for immobilized enzyme prepared without ultrasound, indicating that, this new ultrasonic method improved the catalytic kinetics activity of immobilized dextranase at all the reaction conditions studied. Compared with immobilized enzyme prepared without ultrasound treatment, the immobilized enzymes prepared with ultrasound irradiation exhibited: a higher pH optimum, optimal reaction temperature, activation energy, and thermal stability, as well as, a higher recyclability, which, illustrating the effectiveness of the sonochemical method. To the best of our knowledge, this is the first report on the effect of ultrasound treatments on the immobilization of dextranase.

Production, characterization and (co-)immobilization of dextranase from Penicillium aculeatum

Fermentation kinetics of Penicillium aculeatum ATCC 10409 demonstrated that fungal growth and dextranase release are decoupled. Inoculation by conidia or mycelia resulted in identical kinetics. Two new isoenzymes of the dextranase were characterized regarding their kinetic constants, pI, MW, activation energy and stabilities. The larger enzyme was 3-fold more active (turnover number: 2,230 +/- 97 s(-1)). Pre-treatment of bentonite with H(2)O(2) did not affect adsorption characteristics of dextranase. Enzyme to bentonite ratios above 0.5:1 (w/w) resulted in a high conservation of activity upon adsorption. Furthermore, dextranase could be used in co-immobilizates for the direct conversion of sucrose into isomalto-oligosaccharides (e.g. isomaltose). Yields of co-immobilizates were 2-20 times that of basic immobilizates, which consist of dextransucrase without dextranase.

Immobilization of dextranase from Chaetomium erraticum

In order to facilitate the Co-Immobilization of dextransucrase and dextranase, various techniques for the immobilization of industrial endo-dextranase from Chaetomium erraticum (Novozymes A/S) were researched. Adsorption isotherms at various pH-values have been determined for bentonite (Montmorillonite), hydroxyapatite and Streamline DEAE. Using bentonite and hydroxyapatite, highest activity loads (12,000 U g −1; 2900 U g −1, respectively) can be achieved without a significant change of the apparent Michaelis–Menten constant K M. For successful adsorption, enzyme to bentonite ratios greater than 0.4 (w/w) have to be used as lower ratios lead to 90% enzyme inactivation due to bentonite contact. In addition, covalent linkage using the activated oxiran carriers Eupergit C and Eupergit C250L as well as linkage with aminopropyl silica via metaperiodate activation of glycosyl moiety of dextranase are discussed. This is also the first report probing the structure of a matrix containing dextranase by use of substrate species with different molecular weights. From this we can observe a relationship between the porosity of Eupergit and dextran dependent activity. For the reactor concept using Co-Immobilisates, hydroxyapatite will be preferred to Eupergit because of its higher specific activity and dispersity.

Purification and immobilization of dextranase

AbstractAn enzymic characteristic of Novo dextranase was presented. In addition to a high dextranolytic activity (7,200 U/ml), the crude enzyme also contained small amounts of protease, glucoamylase, polygalacturonase, carboxymethylcellulase, laminarinase and chitinase. A highly purified dextranase was then simply separated from a commercial preparation by column chromatographies on DEAE‐Sepharose, CM‐Sepharose, and by chromatofocussing on Polybuffer Exchanger PBE‐94. The enzyme was recovered with an over 200‐fold increase in specific activity and a yield of 84%. The final preparation was homogeneous, as observed during high performance liquid chromatography (HPLC). Size‐exclusion HPLC indicated that dextranase had a molecular mass of 35 kDa and its isoelectric point, established by chromatofocussing, was 4.85. Analysis of the dextran break‐down products indicated that purified dextranase represents an endolytic mode of action, and isomaltose and isomaltotriose were identified as the main reducing sugars of dextran hydrolysis. The enzyme was then covalently coupled to the silanized porous glass beads modified by glutaraldehyde (Carrier I) or carbodiimide (Carrier II). It was shown that immobilization of dextranase gave optimum pH and temperature ranges from 5.4 to 5.7 and from 50°C to 60°C, respectively. The affinity of the enzyme to the substrate decreased by a factor of more than 13 for dextranase immobilized on Carrier I and increased slightly (about 1.4‐times) for the enzyme bound to Carrier II.

Immobilization of dextranase on bentonite

Dextranase (1,6-α- d-glucan 6-glucanohydrolase, EC 3.2.1.11) from Penicillium aculeatum culture has been immobilized on a bentonite support. The matrix-bound enzyme could be stored as acetone-dried powder or as a suspension in acetate buffer, pH 5.6, for about three weeks at 4°C without any loss of activity. There was no change in the specific activity of the enzyme on immobilization and the enzyme yield was 0.1–0.6 mg/g bentonite matrix. In the presence of sucrose, thermal stability of the immobilized enzyme was high and the bound enzyme could be used for about six cycles.