Bioreactor Design Research Articles

Mechanical Effect on Gene Transfection Based on Dielectric Elastomer Actuator.

Gene transfection has been widely applied in genome function and gene therapy. Although many efforts have been focused on designing carrier materials and transfection methods, the influence of mechanical stimulation on gene transfection efficiency has rarely been studied. Herein, dielectric elastomer actuator (DEA)-based stimulation bioreactors are designed to generate tensile and contractile stress on cells simultaneously. With the example of the EGFP transfection, cells with high membrane tension in the stretching stimulation regions had lower transfection efficiency, while the transfection efficiency of cells in the compressing regions tended to increase. Besides, the duty cycle and loading frequency of the applied stress on cells were also important factors that affect gene transfection efficiency. Furthermore, the pathways of cell endocytosis with the effect of mechanical stimulation were explored on the mechanism for the change of EGFP transfection efficiency. This design of the DEA-based bioreactor, as a strategy to study gene transfection efficiency, could be helpful for developing efficient transfection methods.

Advanced Bioreactor Treatments of Hydrocarbon-Containing Wastewater

This review discusses bioreactor-based methods for industrial hydrocarbon-containing wastewater treatment using different (e.g., stirred-tank, membrane, packed-bed and fluidized-bed) constructions. Aerobic, anaerobic and hybrid bioreactors are becoming increasingly popular in the field of oily wastewater treatment, while high concentrations of petroleum hydrocarbons usually require physico-chemical pre-treatments. Most efficient bioreactor techniques employ immobilized cultures of hydrocarbon-oxidizing microorganisms, either defined consortia or mixed natural populations. Some advantages of fluidized-bed bioreactors over other types of reactors are shown, such as large biofilm–liquid interfacial area, high immobilized biomass concentration and improved mass transfer characteristics. Several limitations, including low nutrient content and the presence of heavy metals or toxicants, as well as fouling and contamination with nuisance microorganisms, can be overcome using effective inocula and advanced bioreactor designs. The examples of laboratory studies and few successful pilot/full-scale applications are given relating to the biotreatment of oilfield wastewater, fuel-contaminated water and refinery effluents.

An additive manufacturing approach to bioreactor design for mesenchymal stem cell culture

Bioreactor design is a challenging endeavour that aims to provide the most ideal environment in which cells can grow and biological reactions can occur. The emergence of regenerative medicine and stem cell therapies has led to the need for more diverse environmental requirements in the bioreactor design space.The study presented uses an additive manufacturing approach for the initial design phase of a packed/fluidized bed bioreactor for mesenchymal stem cell expansion. Combining 3D-printing with CFD for precision control over the bioreactor flow dynamics. Novel flow distributors were developed to engender swirling particle fluidization. The design was simulated and optimised using CFD, demonstrating an increase from 0.01 m/s to 0.02 m/s in the radial velocity of 3.0 mm macrocarriers (1080 kg/m3) at the minimum fluidization velocity. An autoclavable prototype was constructed to illustrate proof-of-concept in the use of swirling flow distribution to enhance cell attachment efficiency (compared to static culture system). Commercial Cytodex 1 carriers were tested: an improvement in attachment efficiency after 24 h from 50 % to 95 % was induced by the swirling flow distributor, with subsequent expansion of 2.4-fold after 6 days of culture.The computational design, modelling and 3D-printing of complex geometric architecture that control the flow dynamics within a bioreactor, provides a novel approach to bioprocess unit operation development for manufacturing novel ATMPs.

3D-Printed Bioreactor Enhances Potential for Tendon Tissue Engineering

Bioreactors have immense value within tissue engineering by enabling important control over design inputs within a microenvironment; however, they are often complex and expensive. Herein, we demonstrate a strategy where a bioreactor was 3D-printed as a means to provide an accessible route to culture scaffolds under physiological inputs and to determine an ideal tendon scaffold structure based on remodeling and degradation events. Furthermore, real-time monitoring of tissue remodeling is introduced through the bioreactor design. Following in vitro uniaxial stretch conditioning, scaffolds were positively altered to reflect improved biomechanical function, enhanced extracellular matrix (ECM) content, and increased mechanical properties of elastic moduli. A bioreactor was constructed to meet the following criteria: real-time readout of resistance, ability to be easily cleaned and sterilized, control of strain, speed, and time parameters, and capability to withstand 3 weeks of continuous load with minimal maintenance. The scaffolds cultured in a dynamic setting showed improved mechanics and promising ECM and collagen content generated. Overall, the bioreactor was effective as a tool to provide mechanical culture conditions that promote the tendon cell niche. The bioreactor supports the scaffolds as a possible therapeutic venture in tendon tissue engineering. Additionally, the 3D-printed bioreactor is a significant contribution to the tissue engineering field by making dynamic culturing more accessible for researchers. Bioreactors present physiological and dynamic conditions for tissue testing and are crucial in tissue engineering for product development, standardization, quality control, and large-scale synthesis. The low-cost, 3D-printed, programmable bioreactor provides cyclic stretch to scaffolds with real-time resistance readout and demonstrates tissue changes over time. The bioreactor enhances scaffold biomechanics and biofunctionality towards natural tendon by promoting cell alignment through mechanical force. The design components are culture chamber for tendon constructs, motor control of uniaxial stretching, and a base for stabilization. The results from conditioning scaffolds in the bioreactor lend to a better understanding of how specific inputs affect tissue outcomes.

MBR treatment of leachates originating from waste management facilities: A reference study of the design parameters for efficient treatment

The main objective of the study was to define the interaction between the solid retention time (SRT) and the contaminant loading rate on a membrane bioreactor's efficacy in removing contaminants frequently detected (chemical oxygen demand (COD), NH4+, total phosphorus and metals) above the discharge criteria in waste-originating leachates. The rates and coefficient calculated from this study's experimental data can be used for the design of membrane bioreactor treating wastewaters, even beyond the scope of this experiment. Over a period of 152 days, SRTs of 28 and 47 days and HRTs of 13, 25, 36 and 52 h were studied using a real leachate with a constant composition. Results showed that membrane bioreactors can efficiently treat >1850 mg COD L−1 d−1 of highly to moderately biodegradable COD, with the SRT having no significant impact on the removal of recalcitrant COD. Overall ammonium removal rates of >740 mg NH4–N L−1 d−1 can be achieved as long as a residual alkalinity of 200 mg CaCO3 L-1 and an adequate dissolved oxygen concentration (6–7 mg L−1) are both maintained. Overall phosphorus removal rates are independent of the phosphorus loading rate. However, the highest overall phosphorus removal rate (39 ± 2 mg P per g of total suspended solids) was obtained at the lowest SRT (28 days) due to an increased extracellular polymeric substance production. Finally, membrane bioreactor's metal removal capacity is mostly dependent on the metals' affinity to both the leachate's recalcitrant COD as well as sludge concentrations.

An Integrated Approach for Experimental Design, Control, and Optimization of Perfusion Bioreactors

This paper presents methods for experimental design, control, and optimization of perfusion bioreactors as well as the vision for their integration. After describing a generic model of perfusion bioreactors, the paper proposes a control scheme via rate estimation and feedback linearization with useful properties with respect to steady-state error, stability, and performance. The paper also introduces data-driven and hybrid procedures for experimental design that are tailored to the intended use of the model for steady-state optimization. Lastly, the methods are illustrated via a simulation example of a perfusion bioreactor.

Analysis of oxygen transport in microfluidic bioreactors for cell culture and organ‐on‐chip applications

In the recent decade, development of microfluidic bioreactors and organ‐on‐chip platforms for drug screening and disease modeling has been rising significantly. Prediction of oxygen level within the microfluidic bioreactors to create physiologically relevant oxygen tension for realistic cellular behavior is of critical importance. This article presented an analytical method to calculate oxygen tension in microchannel parallel plate bioreactors. Two‐dimensional convection‐diffusion equation was solved analytically with considering diffusion in two directions. Cellular oxygen consumption was assumed to follow Michaelis‐Menten kinetics. Effects of oxygenator design, gas permeability of the microfluidic channels, as well as flow rate and cell density on the oxygen distribution within the bioreactor were examined. In addition, a mathematical model was developed to predict oxygen tension in a fluidic circuit containing two interconnected bioreactors with and without recirculation of the media for the culture of hepatocytes and cardiomyocytes. The oxygenators were used to maintain normoxia in the bioreactors independently. The model allowed for the prediction and independent modulation of oxygen tension in the two bioreactors through changing the length of the oxygenators. In overall, the obtained results provide critical insights required for the design and operation of microfluidic bioreactors with desired levels of oxygen tension.

Validation of a novel perfusion bioreactor system in cancer research

Development of drugs is a complex, time- and cost-consuming process due to the lack of standardized and reliable characterization techniques and models. Traditionally, drug screening is based on in vitro analysis using two-dimensional (2D) cell cultures followed by in vivo animal testing. Unfortunately, application of the obtained results to humans in about 90 % of cases fails. Therefore, it is important to develop and improve cell-based systems that can mimic the in vivo-like conditions to provide more reliable results. In this paper, we present development and validation of a novel, user-friendly perfusion bioreactor system for single use aimed for cancer research, drug screening, anti-cancer drug response studies, biomaterial characterization, and tissue engineering. Simple design of the perfusion bioreactor provides direct medium flow at physiological velocities (100?250 ?m s-1) through samples of different sizes and shapes. Biocompatibility of the bioreactor was confirmed in short term cultivation studies of cervical carcinoma SiHa cells immobilized in alginate microfibers under continuous medium flow. The results have shown preserved cell viability indicating that the perfusion bioreactor in conjunction with alginate hydrogels as cell carriers could be potentially used as a tool for controlled anti-cancer drug screening in a 3D environment.

Effects of Physical, Chemical, and Biological Stimulus on h-MSC Expansion and Their Functional Characteristics.

Human adult mesenchymal stem or stromal cells (h-MSC) therapy has gained considerable attention due to the potential to treat or cure diseases given their immunosuppressive properties and tissue regeneration capabilities. Researchers have explored diverse strategies to promote high h-MSC production without losing functional characteristics or properties. Physical stimulus including stiffness, geometry, and topography, chemical stimulus, like varying the surface chemistry, and biochemical stimuli such as cytokines, hormones, small molecules, and herbal extracts have been studied but have yet to be translated to industrial manufacturing practice. In this review, we describe the role of those stimuli on h-MSC manufacturing, and how these stimuli positively promote h-MSC properties, impacting the cell manufacturing field for cell-based therapies. In addition, we discuss other process considerations such as bioreactor design, good manufacturing practice, and the importance of the cell donor and ethics factors for manufacturing potent h-MSC.

Impact of immobilization technology in industrial and pharmaceutical applications.

The current demands of the world's biotechnological industries are enhancement in enzyme productivity and development of novel techniques for increasing their shelf life. Compared to free enzymes in solution, immobilized enzymes are more robust and more resistant to environmental changes. More importantly, the heterogeneity of the immobilized enzyme systems allows an easy recovery of both enzymes and products, multiple reuse of enzymes, continuous operation of enzymatic processes, rapid termination of reactions, and greater variety of bioreactor designs. This review summarizes immobilization definition, different immobilization methods, advantages and disadvantages of each method. In addition, it covers some food industries, protein purification, human nutrition, biodiesel production, and textile industry. In these industries, the use of enzymes has become an inevitable processing strategy when a perfect end product is desired. It also can be used in many other important industries including health care and pharmaceuticals applications. One of the best uses of enzymes in the modern life is their application in diagnose and treatment of many disease especially when used in drug delivery system or when used in nanoform.

Design of a Bioreactor to Assess the Effect of Passive Joint Loading in a Live Chick Embryo In Ovo.

There is increasing interest in understanding how mechanical cues (e.g., physical forces due to kicking and other movements) influence the embryological development of tissues and organs. For example, recent studies from our laboratory and others have used the chick embryo model to demonstrate that the compositional and mechanical properties of developing tendons are strongly regulated by embryo movement frequency. However, current research tools for manipulating embryological movements and in ovo (or in utero) mechanical forces are generally limited to chemical treatments that either paralyze or overstimulate muscles without allowing for precise control of physical cues. Thus, in this study, we introduce an instrument that enables application of passive, dynamic ankle flexion at prescribed amplitudes and frequencies in live, developing chick embryos. This device meets the design goals of allowing for precise (<1.5°) control of different waveforms of ankle motion at a physiologically relevant frequency (0.17 Hz) across a range of ankle angles (0-90° plantarflexion) with maintenance of embryo viability comparable to other methods. Impact Statement We describe the design and implementation of a novel bioreactor to precisely control ankle motion in a chick embryo within its physiological environment. The chick embryo has been used for decades to study mechanobiology of musculoskeletal tissue development and regeneration, but approaches have been limited to chemical treatments that either paralyze or overstimulate muscles without allowing for precise control of physical cues. Thus, this novel instrument is a major advancement over current research tools for manipulating chick embryological movements in ovo (or in utero).

Advances in multi-scale analysis and regulation for fermentation process

Industrial fermentation focuses on realizing the uniform of high titer, high yield, and high productivity. Multi-scale analysis and regulation, including molecule level, cell level, and bioreactor level, facilitate global optimization and dynamic balance of fermentation process, which determine high efficiency of biosynthesis, targeted directionality of bioconversion, process robustness, and well-organized system. In this review, we summariz and discuss advances in multi-scale analysis and regulation for fermentation process focusing on the following four aspects: 1) kinetic modeling of metabolic pathways, 2) characteristic of cell metabolism, 3) co-coupling fermentation and purification, and 4) bioreactor design. Integrating multi-scale analysis of fermentation process and integrating multi-scale regulation are expected as an important strategy for realizing highly efficient fermentation by industrial microorganisms.

Design of a 1000 L pilot-scale airlift bioreactor for nitrification with application of a three-phase hydrodynamic mathematical model and prediction of a low liquid circulation velocity

In this study, a 1000L pilot scale internal loop airlift bioreactor was operated and compared to a mathematical model to determine the best design for optimal supply of oxygen for nitrification and sufficient air for biomass fluidization. The design model is based on parameters such as geometry, carrier density, and airflow of the 1000L pilot scale bioreactor. The model predicts a range of superficial air velocities (0.009–0.013m/s) under which the airlift bioreactor was fluidized. Three superficial air velocities (0.009m/s, 0.011m/s and 0.013m/s) were experimentally tested in the pilot plant and the obtained circulation velocities were compared with the predicted design scenarios. The predicted velocity was in agreement with the measured velocity. The aim of the mathematical model and the calculations of different geometry scenarios was to define the optimal geometry design for the physical model. The results show that the ratio of the cross-sectional area between the riser and the downcomer of 1.33 resulted in the lowest superficial liquid velocity of 0.076m/s in the riser at a relative low superficial air velocity of 0.011m/s and a carrier density of 1030kg/m3. This bioreactor design enabled longest retention time of particles in the oxygenated riser.

A Comprehensive Review on Hydrothermal Carbonization of Biomass and its Applications

Hydrothermal carbonization (HTC) has gained a lot of interest in the last few years for the process production of hydrochar from the different kinds of biological materials such as agricultural waste. Therefore, a thorough review of the literature on HTC has been conducted, which is more energy-efficient and involves hydrochar preparation at a lower temperature without any specific requirements for pressure. This review article primarily differentiates between the hydrochar and biochar developed from various sources. Also, a comparative analysis has been done to evaluate the maximum efficiency of hydrochar production. The various parameters, which influence the hydrochar yield to include pH, temperature, the concentration of the modifying agent, duration of exposure, salt, and phenolic compounds, have also been highlighted in this review. The scale-up processes for industrial level hydrochar production, along with bioreactor designs, have also been discussed. We have also focused on various applications of hydrochar, such as adsorbent in wastewater treatment, carbon sequestration, gas adsorption and in the field of agriculture for soil amendment. The adsorption effects of hydrochar towards the partial and proportionate adsorption of heavy metals and hazardous chemicals have been depicted. This review of the literature clearly illustrates the role of HTC in environmental remediation of soil, air and water and its usage in various industries.

Bioparticles consisting of olive mill wastewater (OMW)-adapted bacteria and OMW-polluted soil as carrier– An application in an anaerobic fluidized bed bioreactor

The world olive oil production represents a very significant sector of the alimentary industry. Nonetheless, the spillages associated to the sector –olive mill wastewater (OMW)– imply serious environmental risks due to their difficult treatments. This study showcases the use of bioparticles from a soil that has repeatedly been contaminated by OMW, to treat water also polluted by OMW. The bacterial biomass that develops overtime within a OMW-polluted course becomes adapted to its conditions and proves effective in its treatment. Water and soil samples were taken from a watercourse that has been suffering from periodic OMW discharges during the last 25 years. Two main factors were identified as the causes of the efficiency of these soils as a means of biological treatment: high concentrations and extended adaptation of active biomass to the pollutants present in the watercourse. The start-up times of the biological processes are reduced since no adaptation period is necessary. This makes it possible to eschew pre-treatment procedures in biological processes. A guide for the design of a continuous flow anaerobic fluidized bed bioreactor (AnFBR) is provided. A stream of 300 m3 d−1 and 5000 mg L−1 COD generates an electrical and thermal energy of 1654 kW h d−1 and 2341 kW h d−1 respectively.

Design, construction and characterisation of a novel nanovibrational bioreactor and cultureware for osteogenesis

In regenerative medicine, techniques which control stem cell lineage commitment are a rapidly expanding field of interest. Recently, nanoscale mechanical stimulation of mesenchymal stem cells (MSCs) has been shown to activate mechanotransduction pathways stimulating osteogenesis in 2D and 3D culture. This has the potential to revolutionise bone graft procedures by creating cellular graft material from autologous or allogeneic sources of MSCs without using chemical induction. With the increased interest in mechanical stimulation of cells and huge potential for clinical use, it is apparent that researchers and clinicians require a scalable bioreactor system that provides consistently reproducible results with a simple turnkey approach. A novel bioreactor system is presented that consists of: a bioreactor vibration plate, calibrated and optimised for nanometre vibrations at 1 kHz, a power supply unit, which supplies a 1 kHz sine wave signal necessary to generate approximately 30 nm of vibration amplitude, and custom 6-well cultureware with toroidal shaped magnets incorporated in the base of each well for conformal attachment to the bioreactor’s magnetic vibration plate. The cultureware and vibration plate were designed using finite element analysis to determine the modal and harmonic responses, and validated by interferometric measurement. This helps ensure that the vibration plate and cultureware, and thus collagen and MSCs, all move as a rigid body, avoiding large deformations close to the resonant frequency of the vibration plate and vibration damping beyond the resonance. Assessment of osteogenic protein expression was performed to confirm differentiation of MSCs after initial biological experiments with the system, as well as atomic force microscopy of the 3D gel constructs during vibrational stimulation to verify that strain hardening of the gel did not occur. This shows that cell differentiation was the result of the nanovibrational stimulation provided by the bioreactor alone, and that other cell differentiating factors, such as stiffening of the collagen gel, did not contribute.

Improved conidiation from entomopathogenic fungi through 26% oxygen pulses in solid-state culture depends on a balance between headspace volume and substrate amounts.

Oxygen-enriched atmospheres applied as periodic pulses increased conidia production from entomopathogenic fungi in agar surface cultures. However, this advantage has not been obtained in solid-state cultures (SSC), probably as a result of different biomass production between both culture systems. In this work, the biomass formation from two Isaria strains was limited in SSC using 5, 2·5 and 1 initial grams of substrate (gds). In the system with 5gds, conidia production decreased in 26% oxygen-enriched pulses compared to the normal atmosphere. Conversely, 26% oxygen pulses increased conidiation up to one order magnitude in systems with 2·5 and 1 gds, respective to the normal atmosphere. These results were explained by oxygen depletion and high CO2 accumulation in the 5 gds system. Whereas in systems with 2·5 or 1 gds, oxygen levels remained high enough to stimulate conidiation. These results were attributed to the headspace volume:gds ratio, which is suggested to be ≥48ml per gds. This ratio is proposed as a scaling-up criterion for bioreactor design when oxygen-enriched pulses are used in SSC for improvement of conidia production. SIGNIFICANCE AND IMPACT OF THE STUDY: Oxygen-enriched atmospheres applied as periodic pulses increase conidiation in entomopathogenic fungi (EF). However, this remained restricted to agar surface cultures, since conidiation decreased when carried out in solid-state culture (SSC) which is used as large-scale production system. We identified that in SSC the ratio between the headspace volume containing 26% oxygen-enriched pulses and the grams of substrate determines the conidiation response to oxygen-enriched pulses. For the first time, oxygen-enriched pulses increased conidiation in SSC respective to the normal atmosphere in four EF. This ratio is proposed as a bioreactor criterion design for large-scale conidia production of EF using oxygen-enriched pulses.

Experiments and finite element modeling of hydrodynamics and mass transfer for continuous gas-to-liquid biocatalysis using a biocomposite falling film reactor

We investigated the hydrodynamics and mass transfer performance of falling liquid films over a rough, hydrophilic paper surface with experiments and finite element modeling. These results are critical for designing a novel gas-to-liquid continuous bioreactor with cells immobilized on the vertical surface of a paper biocomposite. The paper substrate allows investigations at very low Reynolds numbers while maintaining an unbroken liquid film. A finite element model was developed to give 10 fold faster simulation result for designing a prototype laboratory scale bioreactor. Excellent agreement was found in both the film properties and mass transfer performance between experiments and simulations. At Re < 100, mass transfer coefficients kL and kLa were ∼1E-4 m/s and ∼1000 h−1, respectively, at ∼10 W/m3. That power input is 10–1000 fold less than most gas stripping bioreactors. This work highlights the potential of this finite element method for falling film, gas absorbing, bioreactor design and analysis.

Vinegar Engineering: a Bioprocess Perspective

Food engineering is an important sub-field that requires special attention in the food industry. The application of biochemical process engineering principles in food production often leads to the optimization of certain features of the food production process; similarly, it results in rapid production, improved quality and reduced food losses. Consequently, to address each aspect of food processing including engineering adequately, researchers must have a multidisciplinary approach, using aspects from a number of fields such as microbiology, chemistry, food technology, process engineering and molecular biology. Accordingly, this review focuses on the engineering of various vinegars. Furthermore, cognizance is given to the gaps that need to be addressed in vinegar engineering, particularly to address limitations employed in traditional approaches during vinegar production. Food engineering assessments address numerous functions in integrated systems for which fermentation systems are the primary process. Mathematical models are used to describe the process, simulate future fermentations and describe process performance. Vinegar engineering also includes the use or design of bioreactors intended for improved product yield and rapid production, improved mass or energy transfer efficiencies and the reduction of detrimental hydrodynamics fermentor conditions on the microorganisms used. For vinegar fermentation, bioreactor selection which might include cell immobilization requires that appropriate process control and optimization be conducted using mathematical models, with rates of acetification being influenced by parameters such as the ratio of dissolved oxygen consumption in comparison to acetic acid yield.

Sulfide-driven denitrification: detecting active microorganisms in fed-batch enrichment cultures by DNA stable isotope probing.

A microbial community was enriched in the anoxic compartment of a pilot-scale bioreactor that was operated for 180days, fed with sewage and designed for organic matter, nitrogen and sulfide removal by coupling anaerobic digestion, nitrification and mixotrophic denitrification. Denitrification occurred with endogenous electron donors, mainly sulfide and residual organic matter, coming from the anaerobic compartment. The microorganisms involved in denitrification with sulfide as electron donor were identified by DNA-stable isotope probing with [U-13C]-labelled CO2 and NaHCO3. Complete denitrification occurred every two days, and the applied NO3-/S2- ratio was 1.6. Bacteria belonging to the Sulfurimonas denitrificans was identified as a chemoautotrophic denitrifier, and those related to Georgfuchisa toluolica, Geothrix fermentans and Ferritrophicum radicicola were most probably associated with heterotrophic denitrification using endogenous cells and/or intermediate metabolites. This study showed that DNA-SIP was a suitable technique to identify the active microbiota involved in sulfide-driven denitrification in a complex environment, which may contribute to improve design and operation of bioreactors aiming for carbon-nitrogen-sulfur removal.