Preparation of ACE-inhibitory peptides from milk protein in continuous enzyme membrane reactor with gradient dilution feeding substrate

In this study, a novel method of gradient dilution feeding substrate (GDFS) was established to improve the yield of angiotensin-converting enzyme (ACE) inhibitory peptides from milk protein. The hydrolysis process stability, enzymatic efficiency and kinetics of the method were studied and compared with traditional feeding modes, viz., adding water after feeding substrate or constant concentration feeding substrate. Results showed that the GDFS mode achieved the highest membrane flux and lowest fluctuation of protein concentration in the reactor. Moreover, the GDFS maximized protein conversion rate, yield of peptides, and ACE-inhibitory activity, with their values of 67.58 %, 138.51 g/(g*Neutrase), and 0.74 mg/mL (IC₅₀), respectively. In further study, the kinetic model of GDFS mode was successfully established with K_M of 69.481 g/L and V_max of 0.752 g·L⁻¹ min⁻¹. Based on the optimum condition of the kinetic model, the practical longest runtime was 720 min. Obtained results suggested that GDFS mode could be used as an alternative method in the preparation of high-yield bioactive peptides.     

Mass cultivation of Nannochloropsis sp. in annular reactors

A study was made on the mass cultivation of Nannochloropsis sp. in newly designed annular reactors operated under natural, artificial or combined illumination. The annular reactor consists of two 2-m-high Plexiglas cylinders of different diameter placed vertically one inside the other so as to form an annular culture chamber. Artificial illumination is supplied by lamps placed inside the inner cylinder. Two annular reactors of different diameter (50 and 91 cm), light path (4.5 and 3.0 cm) and illuminated surface area (5.3 and 9.3 m²) were experimented with. The effect of two different artificial light sources (fluorescent tubes and metal halide lamps) on culture productivity was investigated in both systems. The highest productivity on a per reactor basis (about 34 g (d. wt) reactor^–1 24 h^–1) was achieved with the larger reactor illuminated by a 400-W metal halide lamp. From February to May a 91-cm reactor illuminated only with natural light was operated in parallel with a 91-cm reactor subjected to combined illumination. Under natural illumination productivity increased from 16.6 g (d. wt) reactor^–1 d^–1 in February to 34.1 g (d. wt) reactor^–1 d^–1 in May. Under combined illumination productivity was 41.3 g (d. wt) reactor^–1 d^–1 in February and increased up to 48.3 g (d. wt) reactor^–1 d^–1 in May. Although the culture exposed to combined illumination always attained higher yields, the productivity gap between the two cultures decreased gradually along the season as solar radiation and minimum night temperatures increased. A 1200-L plant made of ten 50-cm annular reactors was set up and operated for two years with combined illumination yielding an average of 270 g of dry Nannochloropsis sp. biomass per day. More than 2000 L of concentrate suspension (50 g (d. wt) L^–1) of Nannochloropsis sp. were produced and successfully used by fish hatcheries as live feed for rotifers and for rearing seabream larvae with the green-water technique. This study indicates that the annular reactor can be profitably used for long-term cultivation of Nannochloropsis in temperate climates. Besides reliability and ease of operation, the main advantage of the system is that it can be used under natural illumination, yet artificial light can be also supplied to maintain high productivity levels in winter or on cloudy days.     

Plasmid dynamics in a model soil column

Continuous-flow column reactors were used to study the dynamics of plasmid exchange in a structured, thermodynamically open system containing either Enterobacter cloacae or Pseudomonas cepacia , both carrying the transmissible recombinant plasmid R388::Tn1721. Plasmid transfer rates were higher in vermiculite and sterile soil columns supplied with nutrient solution than those in sterile and non-sterile soil columns without input of nutrient solution. For both species, donor and recipient strains took about 5 days to reach their maximum densities in effluents from the columns supplied with nutrient solution. After about 8 day s the donor and transconjugant populations of P. cepacia in the effluent solution decreased exponentially, whereas E. cloacae donor, recipient and transconjugant strains maintained steady-state concentrations. The difference between plasmid stability in the two species may have significant consequences in terms of releasing plasmid-bearing genetically modified microorganisms into the natural environment. The plasmid is persistent in E. cloacae in non-sterile soil even though its transfer to the marked recipient in non-sterile soil was minimal.     

Theoretical investigations of the experimentally observed selectivity of a cobalt imprinted polymer

A cobalt imprinted polymer synthesised, for reducing the volume of radioactive waste generated during nuclear reactor decontaminations, using vinylbenzyl iminodiacetate (VbIDA) as the functional ligand, has been found to be selective for cobaltous ions over excess ferrous ions. The selectivity of the polymer has been investigated through theoretical calculation of the formation energies of complexes involved by using the ab-initio density functional theory (DFT) code SIESTA (Spanish Initiative for Electronic Simulations with Thousands of Atoms). The formation energies of complexes of Fe²⁺, Co²⁺, Cu²⁺ and Ni²⁺ with the free functional ligands as well as with ligands attached to the crosslinkers have been calculated. The calculations revealed that the ferrous forms an unstable complex with the ligands attached to the crosslinkers. The formation energy calculation results were found to corroborate the experimentally observed selectivity order.     

Performance and kinetic evaluation of anaerobic moving bed biofilm reactor for treating milk permeate from dairy industry

High strength milk permeate derived from ultra-filtration based cheese making process was treated in an anaerobic moving bed biofilm reactor (AMBBR) under mesophilic (35 °C) condition. Total chemical oxygen demand (TCOD) removal efficiencies of 86.3–73.2% were achieved at organic loading rates (OLR) of 2.0–20.0 g TCOD L⁻¹ d⁻¹. A mass balance model gave values of methane yield coefficient (Y_G/S) and cell maintenance coefficient (k_m) of 0.341 L CH₄ g⁻¹ TCOD_removed and 0.1808 g TCOD_removed g⁻¹ VSS d⁻¹, respectively. The maximum substrate utilization rate U_max was determined as 89.3 g TCOD L⁻¹ d⁻¹ by a modified Stover–Kincannon model. Volumetric methane production rates (VMPR) were shown to correlate with the biodegradable TCOD concentration through a Michaelis–Menten type equation. Moreover, based on VMPR and OLR removed from the reactor, the sludge production yield was determined as 0.0794 g VSS g⁻¹ TCOD_removed.     

Quantitative detection of selenate-reducing bacteria by real-time PCR targeting the selenate reductase gene

We designed a primer set to target selenate reductase (SerA) for detecting selenate reducing bacteria (SeRB). Our serA gene-based PCR primer set has high specificity in that it and positively amplified some SeRB, but not denitrifying bacteria (DB). Phylogenetic analysis of serA clone sequences of environmental samples from selenate-reducing membrane biofilm reactor (MBfR) biofilms showed that these sequences were closely grouped and had high similarity to selenate reductase gene sequences from SeRB Thauera selenatis and DB Dechloromonas; however, they were distant to other genes from dimethylsulfoxide (DMSO) enzyme family. Constructing a standard curve targeting the serA gene, we found that the good linearity for the qPCR assay when applied it to quantify SeRB in MBfR biofilms, and the gene copies of SeRB correlated well to the selenate removal percentages. Our results demonstrated the feasibility of using the serA gene-based PCR primer set to detect and quantify SeRB in environmental samples.     

Enzymatic synthesis of a series of alkyl esters using novozyme 435 in a packed-bed,miniaturized, continuous flow reactor

Using lipase catalysed, enzymatic esterification as a model reaction, we successfully demonstrate the use of miniaturized technology for biocatalytic reactions. Benchmarked against batch reactions, nine alkyl esters have been synthesized effectively, using Novozyme 435 in a pressure driven, packed-bed, miniaturized, continuous flow reactor. In some cases close to 100% ester conversions were obtained. The paper also demonstrates the ability to screen the enzyme for substrate specificity.     

Specific cephalosporin C production of Acremonium chrysogenum is independent of the culture density

Specific cephalosporin C production of Acremonium chrysogenum grown on a glucose-based minimal medium using conventional batch and dialysis membrane reactor systems was independent of the cell density in the range of 0.4 to 40 g biomass l^–1.     

Comparison of the relative and k0 methods for the standardization of NAA with stable low-flux reactors

Thek ₀ standardization method was Actapted for NAA with stable low-flux reactors where flux monitors are not needed. The modifiedk ₀ method offers the convenience of the use of libraries of sensitivity constants. It was compared to the relative method for 52 elements using a SLOWPOKE reactor and 6 counting geometries. The sensitivity constants determined fromk ₀ values were found to be as accurate or more accurate than those measured with standards. NAA with this modifiedk ₀ method should be accurate to 3% for light elements and 5% for heavy elements.     

Immobilisation of pig liver esterase in hollow fibre membranes

Different methods were evaluated to immobilise Pig Liver Esterase (PLE) in hollow fibre membranes. Four covalent bonding techniques (using epoxy, imidazol, amino and carboxylic acid terminal groups) were tested to link the enzyme to microporous nylon membranes. Physical immobilisation was also studied, by entrapment of the enzyme inside the microporous structure of a polysulfone asymmetric ultrafiltration membrane. The entrapment method lead to a higher retention of enzymatic activity for a longer period of time. This technique was selected to be used in a biphasic membrane bioreactor where the microporous hydrophilic membrane, containing the enzyme, is used to separate an aqueous from an organic phase, in which the substrate is dissolved. Different enzyme loading procedures were studied in the biphasic reactor and the resulting axial and radial enzyme distribution in the hollow fibre module were related to the global enzymatic activity.