Bioprocess and Biosystems Engineering

The enzymatic hydrolysis of olive oil using Chromobacterium viscosum lipase B encapsulated in reversed micelles of AOT in isooctane was carried out in a continous reversed micellar membrane bioreactor. A tubular ceramic membrane installed in an ultrafiltration module was used to retain the lipase and separate the products from the reaction media. Water filled micelles were supplemented to the reactor together with the substrate/solvent solution to compensate for the permeation of reversed micelles. The influence of substrate concentration, residence time and water content in the productivity and conversion of the system were investigated. A linear relationship between productivity and conversion degree was found for each substrate concentration tested. Operational stability of the bioreactor was tested in a long term operation confirming the high stability of this catalytic system.

This study considers two aspects of the implementation of a biomass growth observer and specific growth rate controller in scale-up from small- to pilot-scale bioreactors towards a feasible bulk production process for whole-cell vaccine against whooping cough. The first is the calculation of the oxygen uptake rate, the starting point for online monitoring and control of biomass growth, taking into account the dynamics in the gas-phase. Mixing effects and delays are caused by amongst others the headspace and tubing to the analyzer. These gas phase dynamics are modelled using knowledge of the system in order to reconstruct oxygen consumption. The second aspect is to evaluate performance of the monitoring and control system with the required modifications of the oxygen consumption calculation on pilot-scale. In pilot-scale fed-batch cultivation good monitoring and control performance is obtained enabling a doubled concentration of bulk vaccine compared to standard batch production.

Au nanoparticles (AuNPs) have shown the potential for a variety of applications due to their unique physical and chemical properties. In this study, a facile and affordable method for the synthesis of AuNPs via the liquefied mash of cassava starch has been described and the functionalized AuNPs by l-cysteine improved activity of recombinant xylanase was demonstrated. UV–Vis absorption spectroscopy, transmission electron microscopy, and zeta potential measurements were performed to characterize the AuNPs and monitor their synthesis. The presence of Au was confirmed by energy-dispersive X-ray spectroscopy (EDX) and the X-ray diffraction patterns showed that Au nanocrystals were face-centered cubic. The C=O stretching vibration in the Fourier transform infrared spectrum of AuNPs suggested that the hemiacetal C–OH of sugar molecules performed the reduction of Au3+ to Au0. The presence of C and O in the EDX spectrum and the negative zeta potential of AuNPs suggested that the biomolecules present in liquefied cassava mash were responsible for the stabilization of AuNPs. The surface of AuNPs was easily functionalized by l-cysteine, which improved the stability of AuNPs. Moreover, cysteine-functionalized AuNPs could significantly improve recombinant xylanase efficiency and stability.

Polyunsaturated fatty acid (PUFA) synthase is a special and effective enzyme for PUFA synthesis, and dehydratase (DH) domain played a crucial role in it. In this work, we compared four different DH domains from different strains (Schizochytrium sp. HX-308 and Shewanella sp. BR-2) and different gene clusters. First bioinformatics analysis showed that DH1, 2 and DH3 were similar to FabA and PKS-DH, respectively, and all of them got a hot-dog structure. Second, four DH domains were expressed in Escherichia coli that increased biomass. Especially, Schi-DH1,2 presented the highest dry cell weight of 2.3 g/L which was 1.62 times of that of control. Fatty acids profile analysis showed that DH1,2 could enhance the percentage of unsaturated fatty acids, especially DH1,2 from Schizochytrium sp., while DH3 benefited for the saturated fatty acid biosynthesis. Furthermore, five kinds of fatty acids were added to the medium to study the substrate preferences. Results revealed that DH1,2 domain preferred to acting on C16:0, while DH3 domain trended acting on C14:0 and C15:0, which illustrated DH from different clusters do have specific substrate preference. Besides, DH expression could save the cell growth inhibition by mid-chain fatty acids. This study provided more information about the catalysis mechanism of polyunsaturated fatty acid synthase and might promote the modification study based on this enzyme.

The cells of Bacillus circulans (ATCC 21783) immobilized in sodium alginate gel matrix were able to synthesize the extracellular enzyme, Cyclodextrin glycosyl transferase (CGTase, E.C. 2.4.1.19) which is industrially employed for the preparation of cyclodextrins. Optimization for the maximum production of enzyme was carried out by varying the cell density (3.3–53.5 kg/m3) in the gel and the incubation temperature (30°–42°C). The CGTase activity was found to be the highest (45 units/cm3) with maximum cell loading at 37°C. The reusability of immobilized cells was ascertained by repeated batch experiments. The enzyme activity exhibited was in the range of 50 to 55 units/cm3 in each batch. The continuous synthesis of CGTase by immobilized cells has been demonstrated by operating a fluidized bed reactor at a dilution rate 1.1 · 10−4 sec−1 for a period of 15 days. The enzyme activity has decreased to 42.5 units/cm3 from an initial value of 61 units/cm3 during continuous operation.

Filamentous microorganisms populations present in biofilms from three wastewater treatment plants with a rotating biological contactor system have been studied. Results showed that filaments are an integral component of the biofilms, and that they were made up by: Beggiatoa ssp., Haliscomenobacter hydrossis, Sphaerotilus natans, Thiothrix ssp., and the 021N, 0803, 0914, 0961, 1701, 1851, and 1863 Eikelboom's types. Beggiatoa ssp. was the most frequent microorganism followed by S. natans and type 021N. Microthrix parvicella, Gordona amarae (=Nocardia amarae), Nostocoida limicola (I, II, III), and 0041, 0092, 0211, 0411, 0581, 0675, 1702, and 1852 Eikelboom's types appear infrequently.

 A method is reported for the treatment of industrial fertilizer effluent rich in urea by a new coupling method to immobilize crude urease onto polyester which is having high flow through property in columns. Kinetics of the immobilized enzyme is established in small column. A typical treatment process with two larger columns in packed bed mode is discussed with and without recycling the treated effluent.

ZnO hollow spheres were firstly prepared. A new type of amperometric hydrogen peroxide biosensor was fabricated by entrapping Hemoglobin (Hb) through the ZnO hollow spheres (ZHS) nanoparticles. The composition morphology and size were studied by transmission electron microscopy. The surface topography of the prepared films was imaged by atomic force microscope (AFM). Several techniques, including UV–vis absorption spectroscopy, cyclic voltammetry, chronoamperometry were employed to characterize the performance of the biosensor. The results indicated that the ZHS nanoparticles had enhanced the performance of the hydrogen peroxide sensors. The electrochemical parameters of Hb in the ZHS were calculated by the results of the electron-transfer coefficient (α) and the apparent heterogeneous electron-transfer rate constant K s as 0.5 and 3.1 s−1, respectively. The resulting biosensors showed a wide linear range from 2.1 × 10−6 to 5.18 × 10−3 M, with a low detection limit of 7.0 × 10−7 M (S/N = 3) under optimized experimental conditions. The results demonstrated that the ZHS matrix may improve the protein loading with the retention of bioactivity and greatly promote the direct electron transfer, which can be attributed to its unique morphology, high specific surface area, and biocompatibility. The biosensor obtained from this study possesses high sensitivity, good reproducibility, and long-term stability.