Efficient proteolysis using a regenerable metal-ion chelate immobilized enzyme reactor supported on organic-inorganic hybrid silica monolith

A metal‐ion chelate immobilized enzyme reactor (IMER) supported on organic–inorganic hybrid silica monolith was developed for rapid digestion of proteins. The monolithic support was in situ prepared in a fused silica capillary via the polycondensation between tetraethoxysilane hydrolytic sol and iminodiacetic acid conjugated glycidoxypropyltrimethoxysilane. After activated by Cu²⁺, trypsin was immobilized onto the monolithic support via metal chelation. Proteolytic capability of such an IMER was evaluated by the digestion of myoglobin and BSA, and the digests were further analyzed by microflow reversed‐phase liquid chromatography with ESI‐MS/MS. Similar sequence coverages of myoglobin and BSA were obtained by IMER, in comparison to those obtained by in‐solution digestion (91 versus 92% for 200 ng myoglobin, and 26 versus 26% for 200 ng BSA). However, the digestion time was shortened from 12 h to 50 s. When the enzymatic activity was decreased after seven runs, the IMER could be easily regenerated by removing Cu²⁺ via EDTA followed by trypsin immobilization with fresh Cu²⁺ introduced, yielding the equal sequence coverage (26% for 200 ng BSA). For ～5 μg rat liver extract, even more proteins were identified with the immobilized trypsin digestion within 150 s in comparison to the in‐solution digestion for 24 h (541 versus 483), demonstrating that the IMER could be a promising tool for efficient and high‐throughput proteome profiling.     

Biodiesel production using Aspergillus niger as a whole‐cell biocatalyst in a packed‐bed reactor

Enzymatic lipase transesterification of palm oil to biodiesel in a packed‐bed reactor (PBR) using a novel strain of the fungus Aspergillus niger, immobilized within polyurethane biomass support particles (BSPs), was investigated. A three‐step addition of methanol was used to reduce lipase inhibition by immiscible methanol. The influence of water content and PBR flow rate was investigated. FAME yield was enhanced with an increase of PBR flow rate in the range of 0.15–30 L h^?1, where inefficient mixing of the reaction mixture at lower flow rates resulted in low conversion rates i.e. 69% after 72‐h reaction. Adding the third mole equivalent of methanol resulted in lipase inhibition due to methanol migration into the accumulated glycerol layer. Glutaraldehyde (GA) solution (0.5 vol.%) was used to stabilize lipase activity, which led to a high FAME yield (>90%) in the PBR after 72‐h of reaction time at a flow rate of 15 L h^?1, and a water content of 15%. Moreover, a high conversion rate (>85%) was maintained after four palm oil batch conversion cycles in the PBR. In contrast, lipase activity of non‐GA‐treated cells decreased with each PBR batch cycle, where only 70% FAME was produced after the forth PBR cycle. Transesterification of palm oil in a PBR system using BSPs‐immobilized A. niger as a whole‐cell biocatalyst is a viable process for enzymatic biodiesel production.     

Immobilized cells cultivated in semi-continuous mode in a fluidized bed reactor for xylitol production from sugarcane bagasse

Summary Xylitol production from sugarcane bagasse hemicellulosic hydrolyzate was evaluated in a fluidized bed reactor operated in semi-continuous mode, using cells immobilized on porous glass. The fermentative process was performed during five successive cycles of 72 h each one. The lowest xylitol production occurred in the first cycle, where a high cell concentration (12 g l⁻¹) was observed. In the subsequent cycles the xylitol concentration was ever increasing due to the cells adaptation to the medium. In the last one, 18 g xylitol l⁻¹ was obtained with a yield factor of 0.44 g g⁻¹ and volumetric productivity of 0.32 g l⁻¹ h⁻¹.     

A comparative evaluation of oxygen mass transfer and broth viscosity using Cephalosporin-C production as a case strategy

The production of Cephalosporin-C (CPC) a secondary metabolite, using a mold Acremonium chrysogenum was studied in a lab scale Internal loop air lift reactor. Cephalosporin-C production process is a highly aerobic fermentation process. Volumetric gas–liquid mass transfer coefficient (k_La) and viscosity (η) were evaluated, during the growth and production phases of the microbial physiology. An attempt has been made to correlate the broth viscosity, η and volumetric oxygen transfer coefficient, k_La during the Cephalosporin-C production in an air lift reactor. The impact of biomass concentration and mycelial morphology on broth viscosity has been also evaluated. The broth exhibits a typical non-Newtonian fermentation broth. Rheology parameters like consistency index and fluidity index are also studied.     

Hydrodynamics and mass transfer coefficient in activated sludge aerated stirred column reactor: experimental analysis and modeling

The aerated stirred reactor (ASR) has been widely used in biochemical and wastewater treatment processes. The information describing how the activated sludge properties and operation conditions affect the hydrodynamics and mass transfer coefficient is missing in the literature. The aim of this study was to investigate the influence of flow regime, superficial gas velocity (U(G)), power consumption unit (P/V(L)), sludge loading, and apparent viscosity (mu(ap)) of activated sludge fluid on the mixing time (t(m)), gas hold-up (epsilon), and volumetric mass transfer coefficient (k(L)a) in an activated sludge aerated stirred column reactor (ASCR). The activated sludge fluid performed a non-Newtonian rheological behavior. The sludge loading significantly affected the fluid hydrodynamics and mass transfer. With an increase in the U(G) and P/V(L), the epsilon and k(L)a increased, and the t(m), decreased. The epsilon, k(L)a, and t(m), were influenced dramatically as the flow regime changed from homogeneous to heterogeneous patterns. The proposed mathematical models predicted the experimental results well under experimental conditions, indicating that the U(G), P/V(L), and mu(ap) had significant impact on the t(m), epsilon, and k(L)a. These models were able to give the t(m), epsilon, and k(L)a values with an error around +/-8%, and always less than +/-10%.     

High rate treatment of terephthalic acid production wastewater in a two-stage anaerobic bioreactor

The feasibility was studied of anaerobic treatment of wastewater generated during purified terephthalic acid (PTA) production in two-stage upflow anaerobic sludge blanket (UASB) reactor system. The artificial influent of the system contained the main organic substrates of PTA-wastewater: acetate, benzoate, and terephthalate. Three parallel operated reactors were used for the second stage, and seeded with a suspended terephthalate degrading culture, with and without additional methanogenic granular sludge (two different types). The first stage UASB-reactor was seeded with methanogenic granular sludge. Reactors were operated at 37 degrees C and pH 7. During the first 300 days of operation a clear distinction between the biomass grown in both reactor stages was obtained. In the first stage, acetate and benzoate were degraded at a volumetric loading rate of 40 g-COD/L . day at a COD-removal efficiency of 95% within the first 25 days of operation. No degradation of terephthalate was obtained in the first stage during the first 300 days of operation despite operation of the reactor at a decreased volumetric loading rate with acetate and benzoate of 9 g-COD/L . day from day 150. Batch incubation of biomass from the reactor with terephthalate showed that the lag-phase prior to terephthalate degradation remained largely unchanged, indicating that no net growth of terephthalate degrading biomass occurred in the first stage reactor. From day 300, however, terephthalate degradation was observed in the first stage, and the biomass in this reactor could successfully be enriched with terephthalate degrading biomass, resulting in terephthalate removal capacities of 15 g-COD/L . day. Even though no single reason could be identified why (suddenly) terephthalate degradation was obtained after such a long period of operation, it is suggested that the solid retention time as well the prevailing reactor concentrations acetate and benzoate may have played an important role. From day 1 of operation, terephthalate was degraded in the second stage. In presence of methanogenic granular biomass, high terephthalate removal capacities were obtained in these reactors (15 g-COD/L . day) after approximately 125 days of operation. From the results obtained it is concluded that terephthalate degradation is the bottleneck during anaerobic treatment of PTA-wastewater. Pre-removal of acetate and benzoate in staged bioreactor reduces the lag-phase prior to terephthalate degradation in latter stages, and enables high rate treatment of PTA-wastewater.     

Anaerobic Processes as the Core Technology for Sustainable Domestic Wastewater Treatment: Consolidated Applications, New Trends, Perspectives, and Challenges

Anaerobic digesters have been responsible for the removal of large fraction of organic matter (mineralization of waste sludge) in conventional aerobic sewage treatment plants since the early years of domestic sewage treatment (DST). Attention on the anaerobic technology for improving the sustainability of sewage treatment has been paid mainly after the energy crisis in the 1970s. The successful use of anaerobic reactors (especially up-flow anaerobic sludge blanket (UASB) reactors) for the treatment of raw domestic sewage in tropical and sub-tropical regions (where ambient temperatures are not restrictive for anaerobic digestion) opened the opportunity to substitute the aerobic processes for the anaerobic technology in removal of the influent organic matter. Despite the success, effluents from anaerobic reactors treating domestic sewage require post-treatment in order to achieve the emission standards prevailing in most countries. Initially, the composition of this effluent rich in reduced compounds has required the adoption of post-treatment (mainly aerobic) systems able to remove the undesirable constituents. Currently, however, a wealth of information obtained on biological and physical-chemical processes related to the recovery or removal of nitrogen, phosphorus and sulfur compounds creates the opportunity for new treatment systems. The design of DST plant with the anaerobic reactor as core unit coupled to the pre- and post-treatment systems in order to promote the recovery of resources and the polishing of effluent quality can improve the sustainability of treatment systems. This paper presents a broader view on the possible applications of anaerobic treatment systems not only for organic matter removal but also for resources recovery aiming at the improvement of the sustainability of DST.     

Direct rhizogenesis and establishment of fast growing normal root organ culture of Withania somnifera Dunal

Direct rooting from leaf explants of Withania somnifera was achieved on half strength Murashige and Skoog’s medium supplemented with 15 g l⁻¹ sucrose, and different concentrations of growth regulators. Basal medium supplemented with 2.85 μM indoleacetic acid and 9.85 μM indolebutyric acid achieved maximum number of roots with 100% response. The roots were cultured on MS liquid medium for the establishment of root-organ culture with the same plant growth regulators and incubated on an orbital shaker at 80 rpm at 25 ± 2 °C. A root biomass of 6.15 ± 0.17 g was obtained after 5 weeks. When 1 g roots were inoculated to 2.5 l bubble column reactor, 47 g roots were obtained after 6 weeks. The concentration of alkaloids was increased as compared to field grown roots. The maximum concentration of withanolides (10 mg g⁻¹ dry weight) was obtained in the bioreactor.     

Estimating biofilm reaction kinetics using hybrid mechanistic-neural network rate function model

This work describes an alternative method for estimation of reaction rate of a biofilm process without using a model equation. A first principles model of the biofilm process is integrated with artificial neural networks to derive a hybrid mechanistic-neural network rate function model (HMNNRFM), and this combined model structure is used to estimate the complex kinetics of the biofilm process as a consequence of the validation of its steady state solution. The performance of the proposed methodology is studied with the aid of the experimental data of an anaerobic fixed bed biofilm reactor. The statistical significance of the method is also analyzed by means of the coefficient of determination (R²) and model efficiency (ME). The results demonstrate the effectiveness of HMNNRFM for estimating the complex kinetics of the biofilm process involved in the treatment of industry wastewater.     

Anthracene Removal using Activated Soil Reactors

The aim of this study was to evaluate anthracene removal using activated soil reactors, previously inoculated, under both aerobic and anaerobic conditions. In the reactors, the soil was maintained at 60% moisture (weight basis), room temperature, in the dark, and under constant agitation at 100 rpm. Two experiments were run during and after acclimatization to evaluate anthracene removal under both aerobic and anaerobic conditions. The first one took place during inoculum acclimatization using three different concentrations of anthracene (50, 100, and 500 mg anthracene/L per day) during 90 days. The second experiment took place after acclimatization (during 132 days). The results of anthracene removal were compared with controls in which no additional inoculum was added. During the two experiments, the behavior of pH, chemical oxygen demand (COD), and biogas production was evaluated. Results indicate that the bacterial community adapted for removal of anthracene became enriched through the acclimatization process. Anthracene biodegradation occurred in the soil model with both types of reactors (aerobic and anaerobic), but the rates and extent of biodegradation in the aerobic reactor were higher (95%) than those in anaerobic conditions (74%). Microbial activity also contributed to enhancing bioremediation in the soil by reducing anthracene sorption.