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Scalable single-use adherent cell-based biomanufacturing platforms are essential for unlocking the full potential of cell and gene therapies. The primary objective of this study is to design and develop a novel fixed bed bioreactor platform tailored specifically for scaling up adherent cell culture. The bioreactor comprises a packed bed of vertically stacked woven polyethylene terephthalate mesh discs, sandwiched between two-fluid guide plates. Leveraging computational fluid dynamics modeling, we optimized bioreactor design to achieve uniform flow with minimal shear stress. Residence time distribution measurements demonstrated excellent flow uniformity with plug flow characteristics. Periodic media sampling coupled with offline analysis revealed minimal gradients of crucial metabolites (glucose, glutamine, lactate, and ammonia) across the bioreactor during cell growth. Furthermore, the bioreactor platform demonstrated high performance in automated cell harvesting, with ≈96% efficiency and ≈98% viability. It also exhibited linear scalability in both operational parameters and performance for cell culture and adeno-associated virus vector production. We developed mathematical models based on oxygen uptake rates to accurately predict cell growth curves and estimate biomass in real-time. This study demonstrates the effectiveness of the developed fixed-bed bioreactor platform in enabling scalable adherent cell-based biomanufacturing with high productivity and process control.
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Biomassa , Reatores Biológicos , Técnicas de Cultura de Células , Técnicas de Cultura de Células/métodos , Técnicas de Cultura de Células/instrumentação , Animais , Glucose/metabolismo , Adesão Celular , Proliferação de Células , Hidrodinâmica , Células CHO , Cricetulus , Humanos , Desenho de EquipamentoRESUMO
Quasi-perfusion culture was employed to intensify lentiviral vector (LV) manufacturing using a continuous stable producer cell line in an 8-day process. Initial studies aimed to identify a scalable seeding density, with 3, 4, and 5 × 104 cells cm-2 providing similar specific productivities of infectious LV. Seeding at 3 × 104 cells cm-2 was selected, and the quasi-perfusion was modulated to minimize inhibitory metabolite accumulation and vector exposure at 37°C. Similar specific productivities of infectious LV and physical LV were achieved at 1, 2, and 3 vessel volumes per day (VVD), with 1 VVD selected to minimize downstream processing volumes. The optimized process was scaled 50-fold to 1,264 cm2 flasks, achieving similar LV titers. However, scaling up beyond this to a 6,320 cm2 multilayer flask reduced titers, possibly from suboptimal gas exchange. Across three independent processes in 25 cm2 to 6,320 cm2 flasks, reproducibility was high with a coefficient of variation of 7.7% ± 2.9% and 11.9% ± 3.0% for infectious and physical LV titers, respectively. The optimized flask process was successfully transferred to the iCELLis Nano (Cytiva) fixed-bed bioreactor, with quasi-perfusion at 1 VVD yielding 1.62 × 108 TU.
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Continuous manufacturing of lentiviral vectors (LVs) using stable producer cell lines could extend production periods, improve batch-to-batch reproducibility, and eliminate costly plasmid DNA and transfection reagents. A continuous process was established by expanding cells constitutively expressing third-generation LVs in the iCELLis Nano fixed-bed bioreactor. Fixed-bed bioreactors provide scalable expansion of adherent cells and enable a straightforward transition from traditional surface-based culture vessels. At 0.5 vessel volume per day (VVD), the short half-life of LVs resulted in a low total infectious titer at 1.36 × 104 TU cm-2. Higher perfusion rates increased titers, peaking at 7.87 × 104 TU cm-2 at 1.5 VVD. The supernatant at 0.5 VVD had a physical-to-infectious particle ratio of 659, whereas this was 166 ± 15 at 1, 1.5, and 2 VVD. Reducing the pH from 7.20 to 6.85 at 1.5 VVD improved the total infectious yield to 9.10 × 104 TU cm-2. Three independent runs at 1.5 VVD and a culture pH of 6.85 showed low batch-to-batch variability, with a coefficient of variation of 6.4% and 10.0% for total infectious and physical LV yield, respectively. This study demonstrated the manufacture of high-quality LV supernatant using a stable producer cell line that does not require induction.
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The COVID-19 pandemic has led to high global demand for vaccines to safeguard public health. To that end, our institute has developed a recombinant viral vector vaccine utilizing a modified vesicular stomatitis virus (VSV) construct, wherein the G protein of VSV is replaced with the spike protein of SARS-CoV-2 (rVSV-ΔG-spike). Previous studies have demonstrated the production of a VSV-based vaccine in Vero cells adsorbed on Cytodex 1 microcarriers or in suspension. However, the titers were limited by both the carrier surface area and shear forces. Here, we describe the development of a bioprocess for rVSV-ΔG-spike production in serum-free Vero cells using porous Fibra-Cel® macrocarriers in fixed-bed BioBLU®320 5p bioreactors, leading to high-end titers. We identified core factors that significantly improved virus production, such as the kinetics of virus production, the use of macrospargers for oxygen supply, and medium replenishment. Implementing these parameters, among others, in a series of GMP production processes improved the titer yields by at least two orders of magnitude (2e9 PFU/mL) over previously reported values. The developed process was highly effective, repeatable, and robust, creating potent and genetically stable vaccine viruses and introducing new opportunities for application in other viral vaccine platforms.
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Municipal water resource recovery facilities are not designed to eliminate micropollutants, leading to many pollutants entering the aquatic environment. Within this study, as part of the project MicroStop, the biological treatment of nanofiltration effluent (retentate) under pure aerobic (without nitrification) as well as nitrifying and denitrifying conditions has been investigated for micropollutant elimination. A potential of further biotransformation under increased hydraulic retention time (HRT) of 14 days was shown. Under both HRT of 7 and 14 days, eliminations below LOQ were achieved in the aerated bioreactor for gabapentin, iomeprol, and metoprolol, reaching > 95%, > 69 to > 92%, and > 72%, respectively. The reduction of diclofenac was positively influenced by longer HRT leading to an elimination of up to 67%. Sulfamethoxazole was reduced under denitrification, but accumulated under aeration, resulting in fluctuating results and an overall elimination of 78% under 14 days HRT. PRACTITIONER POINTS: The micropollutant elimination in fixed-bed bioreactors of highly concentrated nanofiltration retentate was studied. Pure aerobic (without nitrification), nitrifying, and denitrifying conditions were investigated under hydraulic retention times (HRT) of 7 and 14 days. Higher initial pollutant concentrations enhanced the biological degradability in attached growth for substances being moderately degradable in activated sludge systems. 4A potential of further biological micropollutant elimination was shown for gabapentin, iomeprol, metoprolol, and diclofenac.
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Diclofenaco , Águas Residuárias , Gabapentina , Metoprolol , Esgotos , Nitrificação , Reatores Biológicos , Eliminação de Resíduos Líquidos/métodosRESUMO
Lentiviral vectors (LVVs) play a critical role in gene delivery for ex vivo gene-modified cell therapies. However, the lack of scalable LVV production methods and the high cost associated with them may limit their use. In this work, we demonstrate the optimization and development of a scalable, chemically defined, animal component-free LVV production process using adherent human embryonic kidney 293T cells in a fixed-bed bioreactor. The initial studies focused on the optimization of the culture process in 2D static cultures. Process changes such as decreasing cell seeding density on day 0 from 2.5 × 104 to 5 × 103 cells/cm2, delaying the transient transfection from 24 to 120 h post-seeding, reducing plasmid DNA to 167 ng/cm2, and adding 5 mM sodium butyrate 6 h post-transfection improved functional LVV titers by 26.9-fold. The optimized animal component-free production process was then transferred to the iCELLis Nano bioreactor, a fixed-bed bioreactor, where titers of 1.2 × 106 TU/cm2 were achieved when it was operated in perfusion. In this work, comparable functional LVV titers were obtained with FreeStyle 293 Expression medium and the conventional Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum both at small and large scale.
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This work investigated the performance of the integrated system (i.e., a Photocatalytic reactor followed by a Fixed bed bioreactor (PC-FBR)) for the degradation of complex Acid Blue 113 from wastewater. Initially, a Photocatalytic reactor was employed to improve the biodegradability index (i.e., BOD/COD) of wastewater from 0.21 ± 0.0062 to 0.395 ± 0.0058. The preliminary photocatalytic oxidation study revealed a maximum of 86.42 ± 0.33 % dye removal at TiO2 loading of 1.5 g/L and an initial concentration of 50 mg/L of AB 113. An integrated reactor system significantly achieved a maximum of 92 ± 2.6 % of dye removal efficiency under a retention time of 120 hr. The stand-alone FBR dye shock loading study suggested that the reactor system was reasonably able to further restore its degradation efficiency. Langmuir-Hinshelwood kinetic model, Monod model, and Andrew-Haldane model were fitted. The bacterial toxicity assessment was carried out using the Pseudomonas fluorescens.
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Despite solid evidence of the success of rotavirus vaccines in saving children from fatal gastroenteritis, more than 82 million infants worldwide still lack access to a rotavirus vaccine. The main barriers to global rotavirus vaccine coverage include cost, manufacturing capacity and suboptimal efficacy in low- and lower-middle income countries. One vaccine candidate with the potential to address the latter is based on the novel, naturally attenuated RV3 strain of rotavirus, RV3-BB vaccine administered in a birth dose strategy had a vaccine efficacy against severe rotavirus gastroenteritis of 94% at 12 months of age in infants in Indonesia. To further develop this vaccine candidate, a well-documented and low-cost manufacturing process is required. A target fully loaded cost of goods (COGs) of ≤$3.50 per course of three doses was set based on predicted market requirements. COGs modelling was leveraged to develop a process using Vero cells in cell factories reaching high titers, reducing or replacing expensive reagents and shortening process time to maximise output. Stable candidate liquid formulations were developed allowing two-year storage at 2-8 °C. In addition, the formulation potentially renders needless the pretreatment of vaccinees with antacid to ensure adequate gastric acid neutralization for routine oral vaccination. As a result, the formulation allows small volume dosing and reduction of supply chain costs. A dose ranging study is currently underway in Malawi that will inform the final clinical dose required. At a clinical dose of ≤6.3 log10 FFU, the COGs target of ≤$3.50 per three dose course was met. At a clinical dose of 6.5 log10 FFU, the final manufacturing process resulted in a COGs that is substantially lower than the current average market price, 2.44 USD per dose. The manufacturing and formulation processes were transferred to BioFarma in Indonesia to enable future RV3-BB vaccine production.
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Infecções por Rotavirus , Vacinas contra Rotavirus , Rotavirus , Animais , Criança , Chlorocebus aethiops , Análise Custo-Benefício , Humanos , Indonésia , Lactente , Malaui , Infecções por Rotavirus/prevenção & controle , Vacinação , Vacinas Atenuadas , Células VeroRESUMO
The accelerating development of gene therapy from research towards clinical trials and beyond has elevated the demand for practical viral vector-manufacturing solutions. The use of disposable upstream technology is gaining traction in clinical manufacturing. Packed-bed or fixed-bed reactors, where column is packed with immobilized biocatalyst particles providing surface to constrain the cells in a particular region of the reactor, have been widely used in bioprocessing applications since mid-1900s. However, the world's first single-use, fully integrated, high cell density, fixed-bed bioreactor was launched only approximately a decade ago. By now, several single-use, fixed-bed technology solutions have been developed in a small scale. Scaling-up the manufacturing can be challenging and for commercial-scale manufacturing, there is practically only one single-use, good manufacturing practice-compliant option available. This study reviews the latest, fully disposable, fixed-bed bioreactors; compares the virus production in the different systems; and discusses important manufacturing cost-related topics. It is predicted that single-use, fixed-bed bioreactors will receive even more attention in the field of viral vector manufacturing and commercialization, especially with the need for higher virus titers and virus yields.
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Reatores Biológicos , Vetores Genéticos , Cultura de Vírus , Terapia GenéticaRESUMO
In this study, we describe the development of a hybrid bioreactor with integrated chlorinated polyethylene (CPE) fixed-bed and zeolite as a microorganism nutrition carrier (MNC), aiming at enhancing and sustaining biohydrogen production during the anaerobic digestion (AD) process. In the batch test, the hybrid bioreactor achieved a maximum biohydrogen production of 646.3 mL/L. Accordingly, the hybrid bioreactor significantly enhanced biohydrogen production and maintained a stable performance for 50 days of semi-continuous operation. This result should be attributed to the CPE providing roughness surface and high porosity for microorganism immobilization, resulting in the enhancement of microbial quantity, confirmed by our scanning electron microscope and immobilized biomass analyses. Moreover, the element ratio significantly decreased, indicating that zeolite could provide metal cations for stimulating microbial bioactivity and growth, as well as contributing to superior biohydrogen productivity during the 50-day operation. In order to further enhance and sustain long-term biohydrogen production, raw zeolite was modified with iron. The hybrid-Fe bioreactor (CPE with Fe-modified zeolite) operated mainly following the acetate pathway and exhibited higher sustainability in improving biohydrogen production with a peak value of 1893.0 mL/L during a 72-day-lasting operation. The synergistic mechanism of the Fe-modified zeolite and CPE fixed-bed revealed that it could effectively induce favorable pathways and contribute to the synthesis of essential enzymes, micronutrient supplementation, electoral conductivity, and microbial immobilization for biohydrogen production. Therefore, a hybrid-Fe bioreactor could provide a unique alternative for the enhancement of hydrogen production for practical applications.
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Zeolitas , Biomassa , Reatores Biológicos , Fermentação , HidrogênioRESUMO
Fixed-bed bioreactors packed with macrocarriers show great potential to be used for vaccine process development and large-scale production due to distinguishing features of low shear force, high cell adhering surface area, and easy replacement of culture media in situ. As an initial step of utilizing this type of bioreactors for Pseudorabies virus production (PRV) by African green monkey kidney (Vero) cells, we developed a tube-fixed-bed bioreactor in the previous study, which represents a scale-down model for further process optimization. By using this scale-down model, here we evaluated impacts of two strategies (use of serum-free medium and low cell inoculum density) on PRV production, which have benefits of simplifying downstream process and reducing risk of contamination. We first compared Vero cell cultures with different media, bioreactors and inoculum densities, and conclude that cell growth with serum-free medium is comparable to that with serum-containing medium in tube-fixed-bed bioreactor, and low inoculum density supports cell growth only in this bioreactor. Next, we applied serum-free medium and low inoculum cell density for PRV production. By optimization of time of infection (TOI), multiplicity of infection (MOI) and the harvesting strategy, we obtained total amount of virus particles ~ 9 log10 TCID50 at 5 days post-infection (dpi) in the tube-fixed-bed bioreactor. This process was then scaled up by 25-fold to a Xcell 1-L fixed-bed bioreactor, which yields totally virus particles of 10.5 log10 TCID50, corresponding to ~ 3 × 105 doses of vaccine. The process studied in this work holds promise to be developed as a generic platform for the production of vaccines for animal and human health.
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Reatores Biológicos , Contagem de Células , Herpesvirus Suídeo 1/genética , Células Vero/virologia , Animais , Chlorocebus aethiops/genética , Chlorocebus aethiops/crescimento & desenvolvimento , Meios de Cultura/química , Meios de Cultura/farmacologia , Herpesvirus Suídeo 1/crescimento & desenvolvimento , Cultura de Vírus/métodosRESUMO
The increasing importance of viral vaccine manufacturing has driven the need for high cell density process optimization that allows for higher production levels. Vero cells are one of the more popular adherent cell lines used for viral vaccine production. However, production is limited due to the logistical limitations surrounding adherent cell line processes, such as large equipment footprints, time and labor-intensive processes, and larger costs per dose. We have addressed this limitation with the establishment of a viral vaccine production system utilizing the novel single use scale-X™ carbo bioreactor. The unit is compact and is scalable and one of the novel features of the system is the continuous in-line downstream purification and concentration processes associated with the bioreactor vessel. We present the results from a campaign featuring a proprietary Vero cell line for production of a live recombinant Vesicular stomatitis virus vaccine that features the Lassa Fever virus glycoproteins. Metabolite analyses and viral yield comparison between traditional flasks, cell factories, and the scale-X carbo bioreactor were performed, and on average, the single use bioreactor produced 2-4 logs higher titers per surface area, approximately 5 × 1010 pfu/cm2, compared to classical flatstock, 2.67 × 106 pfu/cm2, and cell factories production, 5.77 × 108 pfu/cm2. Overall, we describe a novel bioreactor platform that allows for a cost-efficient and scalable process for viral vaccine production.
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Reatores Biológicos , Vacinas Virais , Animais , Linhagem Celular , Chlorocebus aethiops , Vacinas Atenuadas , Células Vero , Cultura de VírusRESUMO
This study described a successful application of the Quality by Design (QbD) approach to pseudorabies virus (PRV) production process development in a fixed-bed bioreactor using the serum-free medium (SFM). The innovated tube-fixed-bed bioreactor was used as a scale-down model of the fixed-bed bioreactor for process development. Risk analysis was performed using Ishikawa diagram combined with failure mode effects analysis (FMEA). The comparative experiment was performed to screen proper medium for adherent African green monkey kidney (Vero) cells from three commercially available SFMs (VP-SFM, ProVERO-1 and Vero-A). The Vero-A medium showed as an outstanding one for further study. The PRV titer in harvest medium was consider as Critical Quality Attribute (CQA) and the Critical Process Parameters (CPPs) [time of infection (TOI), multiplicity of infection (MOI) and initial inoculation cell density] ranked high with risk priority number (RPN) were taken into design of experiment (DoE) methodology. Then prediction model of PRV production process was established and a robust PRV production process was explored. Under the robust setpoint conditions, the Xcell 1 L laboratory-scale fixed-bed bioreactor yielded PRV titer up to 7.87 log10 TCID50/mL at 3 dpi, which was comparable with that in the tube-fixed-bed bioreactor. Combination of the tube-fixed-bed bioreactor and QbD approach could further accelerate the development of a robust virus production process.
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In this study, a novel tube-fixed-bed bioreactor which consists of a TubeSpin bioreactor 50 tube and 0.44 g macrocarriers was developed as the scale-down model of a fixed-bed bioreactor. The adherent Vero cell-based pseudorabies virus (PRV) production process was tested in this novel model. The Vero cells grew well in the tube-fixed-bed bioreactor, and the cell density reached 5.8 × 106 cells/mL after 7 days of culture. The PRV production parameters (time of infection, multiplicity of infection, and harvest process) were optimized in the tube-fixed-bed bioreactor. Then the optimized process (time of infection = 3 days, multiplicity of infection = 0.001 and multiple harvest process) was scaled up 25-fold to an Xcell 1-L laboratory-scale fixed-bed bioreactor and 125-fold to an Xcell 5-L fixed-bed bioreactor successfully. The total PRV harvest in the Xcell 1-L bioreactor at 5 days after infection (dpi) was 10.25 log10 TCID50 which corresponds to 177,827 doses of vaccine. The total PRV harvest in the Xcell 5-L bioreactor at 5 dpi was 11.13 log10 TCID50 which corresponded to 1,348,962 doses of vaccine. The comparable growth curve, metabolism, and PRV production profile of the scaled-up bioreactors confirmed the feasibility and scalability of the tube-fixed-bed bioreactor as a scale-down model of the fixed-bed bioreactor for virus production process development.
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Herpesvirus Suídeo 1 , Vacinas contra Pseudorraiva , Animais , Reatores Biológicos , Chlorocebus aethiops , Células VeroRESUMO
The quality of the bio-treated coking wastewater (BTCW) is difficult to meet increasingly stringent coking wastewater discharge standards and future wastewater recycling needs. In this study, the pre-treatment process of BTCW was installed including the two up-flow fixed-bed bioreactors (UFBRs) which were separately filled with alkali-pretreated or no alkali-pretreated corncobs used as solid carbon sources as well as biofilm carriers. Results showed that this pre-treatment process could significantly improve the biodegradability of BTCW and increase the C/N ratio. Thus, over 90% of residual nitrate in BTCW were removed stably. Furthermore, GC-MS analysis confirmed that the typical refractory organic matters decreased significantly after UFBRs pre-treatment. High-throughput sequencing analysis using 16S rRNA demonstrated that dominant denitrifiers, fermentative bacteria and refractory-organic-pollutants-degrading bacteria co-existed inside the UFBRs system. Compared with no alkali-pretreated corncobs, alkali-pretreated corncobs provided more porous structure and much stable release of carbon to guarantee the growth and the quantity of the functional bacteria such as denitrifiers. This study indicated that the UFBRs filled with alkali-pretreated corncobs could be utilized as an effective alternative for the enhanced treatment of the BTCW.
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Biodegradação Ambiental , Coque/análise , Nitratos/análise , Eliminação de Resíduos Líquidos/métodos , Águas Residuárias/química , Poluentes Químicos da Água/análise , Biofilmes , Reatores Biológicos , Carbono , Poluentes Ambientais , Nitratos/metabolismo , RNA Ribossômico 16S , Reciclagem , Poluentes Químicos da Água/metabolismo , Zea maysRESUMO
Recently, removal of arsenic from different industrial effluent discharged using simple, efficient and low-cost technique has been widely considered. In this study, removal of arsenic (As) from real wastewater has been studied employing modified bio-oxidation followed by adsorptive filtration method in a novel continuous flow through the reactor. This method includes biological oxidation of ferrous to ferric ions by immobilized Acidothiobacillus ferrooxidans bacteria on granulated activated carbon (GAC) in fixed bed bio-column reactor with the adsorptive filtration unit. Removal efficiency was optimized regarding the initial flow rate of media and ferrous ions concentration. Synthetic wastewater sample having different heavy metal ions such as Arsenic (As), Cobalt (Co), Chromium (Cr), Copper (Cu), Iron (Fe), Lead (Pb) and Manganese (Mn) were also used in the study. The structural and surface changes occurring after the treatment process were scrutinized using FT-IR and Scanning Electron Microscopy (SEM) analysis. The finding showed that not only arsenic can be removed considerably in the bioreactor system, but also removing efficiency was much more (<90%) for other heavy metals in real wastewater sample. The results from TCPL test confirms that solid spent media was non-hazardous and can be safely disposed of. This study verified that combination of bio-oxidation with adsorptive filtration method improves the removal efficiency of arsenic and other heavy metal ions in wastewater sample.
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Arsênio , Poluentes Químicos da Água , Adsorção , Filtração , Espectroscopia de Infravermelho com Transformada de Fourier , Águas ResiduáriasRESUMO
A single-use fixed-bed bioreactor (iCELLis nano) can be used for cultivating non adherent insect cells, which can be then recovered for scaling up or for harvesting a membrane-associated viral glycoprotein with high quality in terms of preserved protein structure and biological function. Here, we describe the procedures for establishing genetically modified Drosophila melanogaster Schneider 2 (S2) cell cultures in the iCELLis nano bioreactor and for quantifying by ELISA the recombinant rabies virus glycoprotein (rRVGP) synthesized. By using the described protocol of production, the following performance can be regularly achieved: 1.7 ± 0.6 × 1E10 total cells; 2.4 ± 0.8 × 1E7 cells/mL and 1.2 ± 0.9 µg of rRVGP/1E7 cells; 1.5 ± 0.8 mg of total rRVGP.
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Glicoproteínas/metabolismo , Insetos/metabolismo , Vírus da Raiva/metabolismo , Proteínas Recombinantes/metabolismo , Proteínas Virais/metabolismo , Animais , Reatores Biológicos , Linhagem Celular , Drosophila melanogaster/metabolismoRESUMO
A single-use fixed-bed bioreactor (iCELLis nano) can be used for cultivating non adherent insect cells, which can be then recovered for scaling up or for harvesting a membrane-associated viral glycoprotein with high quality in terms of preserved protein structure and biological function. Here, we describe the procedures for establishing genetically modified Drosophila melanogaster Schneider 2 (S2) cell cultures in the iCELLis nano bioreactor and for quantifying by ELISA the recombinant rabies virus glycoprotein (rRVGP) synthesized. By using the described protocol of production, the following performance can be regularly achieved: 1.7 ± 0.6 × 1E10 total cells; 2.4 ± 0.8 × 1E7 cells/mL and 1.2 ± 0.9 μg of rRVGP/1E7 cells; 1.5 ± 0.8 mg of total rRVGP.
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Intermittent illumination combined with bio-zeolite fixed-bed process was utilized to improve the efficiency of anaerobic digestion with ammonium-rich substrate. The batch experiments were carried out at NH4+-N concentration of 2211mg/L under intermittent illumination and dark (as control) conditions, respectively. The illuminated bioreactor achieved higher methane production (287mL/g-DOC) and ATP value (0.38µmol/L) than that under dark condition. Then the bio-zeolite fixed-bed bioreactor (NH4+-N concentration: 3000mg/L) was used to study the additional efficiency on the illuminated ammonium-rich anaerobic digestion process. The result showed that the illuminated fixed-bed bioreactor presented the greatest methane concentration (70%), methane yield (283mL/g-DOC) and quantity of methanogens comparing with no-bed bioreactor. Furthermore, the illuminated fixed-bed bioreactor achieved better performance during 118-day semi-continuous fermentation. The combination of the intermittent illumination and bio-zeolite fixed-bed strategy contributed to the higher efficiency and stability of the ammonium-rich anaerobic digestion process.
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Compostos de Amônio/análise , Reatores Biológicos , Biotecnologia/instrumentação , Biotecnologia/métodos , Resíduos , Zeolitas/química , Trifosfato de Adenosina/metabolismo , Anaerobiose , Bactérias/metabolismo , Bactérias/efeitos da radiação , Biomassa , Reatores Biológicos/microbiologia , Fermentação , Luz , Metano/análise , Esgotos/microbiologiaRESUMO
Textile effluents are highly polluting and have variable and complex compositions. They can be extremely complex, with high salt concentrations and alkaline pHs. A fixed-bed bioreactor was used in the present study to simulate a textile effluent treatment, where the white-rot fungus, Trametes versicolor, efficiently decolourised the azo dye Reactive Black 5 over 28 days. This occurred under high alkaline conditions, which is unusual, but advantageous, for successful decolourisation processes. Active dye decolourisation was maintained by operation in continuous culture. Colour was eliminated during the course of operation and maximum laccase (Lcc) activity (80.2 UâL(-1)) was detected after glycerol addition to the bioreactor. Lcc2 gene expression was evaluated with different carbon sources and pH values based on reverse transcriptase-PCR (polymerase chain reaction). Glycerol was shown to promote the highest lcc2 expression at pH 5.5, followed by sucrose and then glucose. The highest levels of expression occurred between three and four days, which corroborate the maximum Lcc activity observed for sucrose and glycerol on the bioreactor. These results give new insights into the use of T. versicolor in textile dye wastewater treatment with high pHs.