Sukekuni Mukataka

Factors affecting the composition of oligosaccharides produced in chitosan hydrolysis using immobilized chitosanases.

The hydrolysis reaction of chitosan using immobilized chitosanases with regard to the composition of its products and the yield of the intermediate target products, pentamer and hexamer of chitosan oligosaccharides, was investigated. Chitosanase was immobilized onto agar or agarose gel particles by the multipoint attachment method. In batch experiments, surface enzyme density, support particle size, temperature, agitator speed, and initial substrate concentration significantly affected the composition of the oligosaccharides produced. It was believed that these factors all related to the reaction rate and mass transfer rate at the surface of the support materials immobilizing the enzymes. These effects were summarized as a correlation with Damköhler number ( Da), defined as the ratio of the maximum reaction rate to the maximum mass transfer rate. The result showed that the reaction conditions that give a low value of Da provide a high yield of pentamer and hexamer oligosaccharides.

Gluconic acid production at high concentrations by Aspergillus niger immobilized on a nonwoven fabric

Abstract Batch production of gluconic acid in the presence of a high concentration of glucose was investigated using free and immobilized mycelia of Aspergillus niger IAM 2094 with the aim of achieving repeatable constant production. Accumulation of 300 g/l of gluconate with a productivity of 60 g/l·h was achievable by intermittent addition of powdered glucose using filamentous-form mycelia in the presence of 150 ppm dissolved oxygen. However, this productivity became unattainable after a few repetitions. The use of pellet-form mycelia, in place of filamentous ones, did not prove effective either. However, when the mycelia were immobilized on a nonwoven fabric, a sustained level (220 g/l) of gluconate production was reproducible. Immobilized mycelia grown in a gas phase (air or oxygen) had a much longer durability than mycelia grown in a liquid culture medium. The gluconate-producing activity of immobilized mycelia grown in the presence of oxygen was much higher than that of mycelia grown in air. At 150 ppm dissolved oxygen, 220 g/l of gluconate was repeatedly produced 14 times at a constant production rate in a period of about 1,000 h.

Immobilization and stabilization of chitosanase by multipoint attachment to agar gel support.

Highly stable chitosanase immobilized on an agar gel support was prepared by the multipoint attachment method. The optimum pH range was broadened to between 4 and 6, whereas for free chitosanase, the pH was only 5.6. The optimum temperature was also increased from 60 degrees C to 80 degrees C after the immobilization. The activity of immobilized chitosanase remained at 95% of its initial activity level after 225 h of incubation at 50 degrees C, whereas for free chitosanase, it decreased to 20% after 1 h of incubation. The immobilization markedly increased the thermostability of chitosanase. These changes in the reaction characteristics are favorable for the practical use of chitosanase in industrial processes. The effect of glycidol concentration in the activation of agar gel was also examined. The surface density of the aldehyde residue increased with increasing glycidol concentration. A maximal activity of 11.9 U/g-support was obtained when the glycidol concentration was 0.7 M. At concentrations higher than this, thermostability was almost the same. It was therefore proven that the optimal glycidol concentration in this system is 0.7 M. The effects of glycidol concentration on the activity and the thermostability of chitosanase are discussed in relation to the number of covalent bonds between the chitosanase and its support. Chitosan oligosaccharides were continuously produced using a column reactor packed with the immobilized chitosanase. The percentage of hydrolyzed chitosan after 28 reaction days was 44%. This was a slight decrease from the 48% observed on the first day. The total concentration of pentamer and hexamer ranged from 1.3 mg/ml to 1.5 mg/ml during the 28 reaction days. This was approximately 30% of the chitosan concentration in the supplied substrate solution.

On the intermolecular crosslinking of PVA chains in an aqueous solution by γ-ray irradiation

Abstract We studied how the polymer chains of poly(vinyl alcohol)(PVA) are crosslinked in its O 2 -free aqueous solution by γ -rays. The static–dynamic laser light scattering method was employed for determining the molecular weight ( M w ), hydrodynamic radius ( R h ) and radius of gyration ( R g ) for the original PVA ( M w =1.0×10 5 ) and its radiation products. The γ -ray irradiation from Co 60 to the PVA solutions with different concentrations was performed at different dose levels. It was found that both molar mass and size (i.e., R g and R h ) of the radiation products increased with a dose increase. The magnitude of these increases was enhanced by decreasing the polymer concentration ( C p ). In particular, at C p C p using the dose curves of M w . It became apparent that, regardless of C p , the intermolecular crosslinking reactions were dominant in the initial stage of irradiation. The radiation yield of intermolecular crosslinking was found to increase linearly when increasing C p , and at C p > the overlap concentration it asymptotically approached a limiting value estimated from the dimerization of pentane-2,4-diol as a model compound of PVA.

The influence of polymer concentration on the radiation-chemical yield of intermolecular crosslinking of poly(vinyl alcohol) by γ-rays in deoxygenated aqueous solution

Abstract The effect of polymer concentration on G value of intermolecular crosslinking of poly(vinyl alcohol) (PVA) during γ-ray irradiation is reported here. The G value, determined by the measurement of weight average molecular weight (Mw) from static light scattering, increased initially due to the polymer concentration and approached a maximum value of 0.5 × 10−7 mol J−1. It then markedly decreased for the more concentrated water-swollen PVA film (polymer concentration Cp>300 g dm−3). Our results suggest that polymer concentration plays an important role in the crosslinking and degradation reactions of PVA during γ-ray irradiation.

Enzymatic Hydrolysis of Fats at High Substrate Concentrations in Biphasic Organic-Aqueous Systems

Abstract The hydrolysis of palm oil and beef tallow by lipase has been studied for practical applications in a biphasis isooactane-aqueous system using a high substrate concentration. The effective lipase concentration for the hydrolysis was found to be about 120 IU per g of substrate. The addition of twenty percent isooctane brought about the most rapid reaction and produced the highest percentage of hydrolysis. For both palm oil and beef tallow, a percentage of hydrolysis higher than 98% was achieved in the 20% isooctane system at a higher concentration of 50%. However, when the substrate concentration was higher than 50%, the final value of hydrolysis decreased as the concentration of the substrate increased. Utilization of recycled lipase was attempted using an ultrafiltration membrane reactor. Approximately 60

Highly Productive Droplet Formation by Anisotropic Elongation of a Thread Flow in a Microchannel

% of the lipase activity was recoverable after each reaction.

Utilization of powdered pig bone as a support for immobilization of lipase

We developed a microfluidic device to form monodisperse droplets with high productivity by anisotropic elongation of a thread flow, defined as a threadlike flow of a dispersed liquid phase in a flow of an immiscible, continuous liquid phase. The thread flow was anisotropically elongated in the depth direction in a straight microchannel with a step, where the microchannel depth changed. Consequently, the elongated thread flow was given capillary instability (Rayleigh-Plateau instability) and was continuously transformed into monodisperse droplets at the downstream area of the step in the microchannel. We examined the effects of the flow rates of the dispersed phase and the continuous phase on the droplet formation behavior, including the droplet diameter and droplet formation frequency. The droplet diameter increased as the fraction of the dispersed-phase flow rate relative to the total flow rate increased and was independent of the total flow rate. The droplet formation frequency proportionally increased with the total flow rate at a constant dispersed-phase flow rate fraction. These results are explained in terms of a mechanism similar to that of droplet formation from a cylindrical liquid thread flow by Rayleigh-Plateau instability. The microfluidic device described was capable of forming monodisperse droplets with a 160-microm average diameter and 3-microm standard deviation at a droplet formation frequency of 350 droplets per second from a single thread flow. The highest total flow rate achieved was 6 mL/h using the present device composed of a straight microchannel with a step. We also demonstrated parallel droplet formation by anisotropic elongation of multiple thread flows; the process was applied to form W/O and O/W droplets. The highly productive droplet formation process presented in this study is expected to be useful for future industrial applications.

Viscometric, light scattering, and size‐exclusion chromatography studies on the structural changes of aqueous poly(vinyl alcohol) induced by γ‐ray irradiation

Abstract Pig bone was examined for its suitability as a support material for lipase immobilization. It was observed that pig bone (PB) particles dispersed readily in both polar and nonpolar solvents, and lipase was easily adsorbed. In particular lipase adsorbed on olive oil-soaked pig bone (OPB) particles exhibited a higher hydrolytic activity than that in lipase adsorbed on a selection of other representative supports, regardless of removing the presoaked olive oil from the particles after immobilization of lipase. The optimum pH and temperature for hydrolytic activity of OPB-adsorbed lipase were the same as those for free lipase, although thermal resistance was increased by immobilization. When OPB-adsorbed lipase was used for repeated batch reactions of olive oil hydrolysis, an activity of more than 80% of the initial activity of each run could he retained after 46 h reaction. The results suggest that PB is an excellent support material.

Transesterification reaction between medium‐ and long‐chain fatty acid triglycerides using surfactant‐modified lipase

The crosslinking processes of aqueous poly(vinyl alcohol) (PVA) by γ-ray irradiation were studied by viscometry, dynamic and static light scattering (DLS and SLS), as well as size exclusion chromatography (SEC). Increases in the intrinsic viscosity ([η]), molecular weight (Mw), hydrodynamic radius (Rh), and radius of gyration (Rg), and a decrease in second virial coefficient (A2) were observed after γ-ray irradiation. However, both the values of [η] and A2 for irradiated PVA fell below the data of unirradiated PVA solutions, suggesting a conformational change of PVA chains after γ-ray irradiation. This structural change of PVA as a result of γ-ray irradiation was also indicated by the decreases in Rg/Rh from 1.5 to 1.39 by SLS and DLS, and in Mark–Houwink exponent αη from 0.54 to 0.26 by SEC-Viscometry. The broadening of the Mw distribution (MWD) as indicated by the polydispersity index increased from 2.2 to 6.5 because of γ-ray irradiation.