Optimal 1,3-propanediol production: Exploring the trade-off between process yield and feeding rate variation

This paper proposes a new optimal control model for the production of 1,3-propanediol (1,3-PD) viamicrobial fed-batch fermentation. The proposed model is governed by a nonlinear multi stage dynamic system with two modes: feeding mode, in which glycerol and alkali substrates are added continuously to the fermentor; and batch mode, in which no substrates are added to the fermentor. The non-standard objective function incorporates both the final 1,3-PD yield and the cost of changing the input feeding rate, which is the control variable for the fed-batch fermentation process. Continuous state inequality constraints are imposed to ensure that the concentrations of biomass, glycerol, and reaction products lie within specified limits. Using the constraint transcription method, we approximate the continuous state inequality constraints by a conventional inequality constraint to yield an approximate parameter optimization problem. We then develop a combined particle swarm and gradient-based optimization algorithm to solve this approximate problem. The paper concludes with simulation results

Optimisation of Microbial Fuel Cells (MFCs) through bacterial-robot interaction

For over 100 years, Microbial Fuel Cells (MFCs) have been developed as eco-friendly alternatives for generating electricity via the oxidation of organic matter by bacteria. In the early 2000s, collectives of MFCs were proven fea-sible energy providers for low-power robots such as Gastrobot and EcoBots. Even though individual MFC units are low in power, significant progress has been achieved in terms of MFC materials and configurations, enabling them to generate higher output levels. However, up to this date, MFCs are produced and matured using conventional laboratory methods that can take up to three months to bring the MFCs to their maximum power aptitudes. In this work, an approach to use a low-cost (£1.5k) RepRap liquid handling robot called EvoBot was employed with the aim to automate the maturing process of MFCs and allow them to reach their maximum power ability in a shorter time span. Initially, the work focused on establishing an interface and interconnection between the living cells (in the MFC) and the robotic platform, and investigating whether the MFC voltage can trigger a feedback loop feeding mechanism. It was shown that the robot successfully matured the MFCs in just 6 days and, they were also 1.4 times more powerful than conventionally matured MFCs (from 19.1 mW/m2 to 26.5 mW/m2). This work took a rounded approach in improving the overall MFC perfor-mance. 3D-printable materials that can emerge from EvoBot were investi-gated for fabricating MFCs. MFCs employing these materials improved their power output by almost 50% (from 66μW to 130 μW) compared to the ones based on conventional, fluorinated materials. Furthermore, EvoBot was able to improve the fuel supply frequency and composition using evolutionally algorithms. For the first time, this project has demonstrated that the fabrica-tion, maintenance and power generation of MFCs can be optimised via the interaction and support of a dedicated robotic system

Contribution to the development of methods and systems for the automatization during the early stages of bioprocess development

This thesis is framed within the field of red biotechnology and more specifically in the development of bioprocesses for cell species that feature some therapeutical interest, either for the production of vaccines and monoclonal antibodies or stem cell experimental research. The main objective was the development and application of different instrumental techniques for the control and online monitorization of cell cultures. Oxygen consumption OUR (Oxygen Uptake Rate) was chosen as the central theme since this parameter has often been referenced as the most straighforward indicator of metabolic activity in animal cell culture. This thesis was carried out in the context of a Spin-Off project (Hexascreen Culture Technologies) whose objective was the development of disposable Minibioreactors intended for biopharmaceutical research. Obviously, this has led to a number of important trade-offs, as well as the proposal of several imaginative solutions to solve various technological challenges. For this reason and in order to offer a better idea of the work's scope, it was decided to include in the thesis not only the description of the method and results related to the OUR estimation but a detailed description of the systems developed. Results demonstrate the feasibility of a simplified procedure for estimating the oxygen consumption. This is a review of the Stationary liquid phase mass balance method which allows reducing the implementation cost and unlike the Dynamic method (The most usual thechnique) prevents changes on the oxygen tension that could affect the cell's normal arctivity. The proposed method is based on the accurate control of the oxygen concentration by means of PWM driven electrovalves and using the control loop internal signals to estimate the OUR.Aquesta Tesi doctoral està enquadrada en l'àmbit de la Biotecnologia vermella i més concretament en el desenvolupament de Bioprocessos relacionats amb espècies cel·lulars d’interès terapèutic, bé sigui per a la producció de vacunes, anticossos monoclonals o bé per a la recerca experimental amb cèl·lules mare. L'objectiu general ha estat el desenvolupament i aplicació de diferents tècniques instrumentals per al control i monitorització en línia de cultius cel·lulars, tant mateix d'entre les diferents tècniques emprades es va escollir la monitorització de la demanda d'oxigen O.U.R. (Oxygen Uptake Rate) com a tema central de la tesi degut a que aquest paràmetre ha estat referenciat sovint com un dels millors indicadors de l'activitat metabòlica en cultius de cèl·lules animals. Cal mencionar que la Tesi ha estat duta a terme en el context d'un projecte empresarial (HexaScreen Culture Technologies) l'objectiu del qual ha estat el desenvolupament de Minibioreactors d'un sol ús orientats al mon de la recerca Biofarmacèutica. Òbviament això ha comportant un número important de compromisos a l'hora d'abordar les diferents tasques, així com el plantejament de solucions imaginatives per a la resolució dels diferents reptes tecnològic. Per aquest motiu i per tal de transmetre una millor idea de l'abast del treball realitzat, es va decidir incloure en la tesi no només la descripció del mètode i resultats relacionats amb l'estimació de la O.U.R. sinó amés una descripció prou detallada dels sistemes desenvolupats. Pel que fa al tema central de la tesi, es demostra la viabilitat d'un procediment simplificat per a l'estimació de la demanda d’oxigen. Es tracta d'una revisió del procediment d'estimació de la OUR en condicions de concentració estacionària en la fase líquida que permet reduir-ne el cost de implementació tot prescindint de l'ús de cabalímetres màssics, així com a diferència del mètode dinàmic (Tècnica més habitual) evitar cap mena de canvi en la tensió d’oxigen que pogués afectar l’activitat normal de les cèl·lules. El mètode proposat, es basa en el control de la concentració d’oxigen mitjançant actuació PWM de les vàlvules d'aereació i l’ús dels propis senyals del llaç de control per tal d'estimar la O.U.R.Postprint (published version

Small scale/large scale MFC stacks for improved power generation and implementation in robotic applications

Microbial Fuel Cells (MFCs) are biological electrical generators or batteries that have shown to be able to energise electronic devices solely from the breakdown of organic matter found in wastewater. The generated power from a single unit is currently insufficient to run standard electronics hence alternative strategies are needed for stepping-up their performance to functional levels. This line of work deals with MFC miniaturisation; their proliferation into large stacks; power improvement by using new electrode components and finally a novel method of energy harvesting that will enhance the operation of a self-sustainable robotic platform. A new-design small-MFC design was developed using 3D printing technology that outperformed a pre-existing MFC of the same volume (6.25 mL) highlighting the importance of reactor configuration and material selection. Furthermore, improvements were made by the use of a cathode electrode that facilitates a higher rate of oxygen reduction reaction (ORR) due to the high surface area carbon nanoparticles coated on the outer layer. Consequently, a 24-MFC stack was built to simulate a small-scale wastewater treatment system. The MFC units were connected in various arrangements, both fluidically as a series of cascades and electrically in-parallel or in-series, for identifying the best possible configuration for organic content reduction and power output. Results suggest that in-parallel connections allow for higher waste removal and the addition of extra units in a cascade is a possible way to ensure that the organic content of the feedstock is always reduced to below the set or permitted levels for environmental discharge. Finally, a new method of fault-proof energy harvesting in stacks was devised and developed to produce a unique energy autonomous energy harvester without any voltage boosting and efficiencies above 90%. This thesis concludes with the transferability of the above findings to a robotic test platform which demonstrates energy autonomous behaviour and highlights the synergy between the bacterial engine and the mechatronics

Waste and wastewater clean-up using microbial fuel cells

A sustainable energy portfolio should include a range of carbon-neutral and renewable energy technologies. Amongst the renewable energy technologies, MFCs can offer a solution for both sustainable energy and clean water demands. In order to take the MFC technology to commercial level, more effort has to be spent to improve the performance and treatment efficiency. The goal for this thesis was to improve anode performance and waste utilisation. To achieve this goal, the approach taken was system scale-up through multiples of relatively small sized MFC units. Two main aspects of the MFC anode, design and biofilm affecting parameters, were investigated in order to better understand and enhance the anode performance. Through a number of experiments, better performing material for each MFC component was chosen. For example, by replacing the previous electrode material with modified anode and cathode, a 2.2 and 4.9 fold increase in power output was achieved respectively. Investigations into biofilm affecting parameters such as temperature, external load and feedstock, yielded novel findings helping to understand the dynamic characteristics of MFC anode biofilms. For the final part of this thesis, these findings were used to implement the MFC technology for practical applications such as treating wastes and resource recovery as well as producing electrical energy. Two troublesome wastes, urine and uric scale showed great potential for being power sources of MFC electricity generation. Furthermore it was demonstrated that MFCs can contribute to recovery of resources such as nitrogen and phosphorus in the form of struvite. A commercial electronic appliance was run continuously, powered by a stack of 8 MFCs fed with neat human urine, which successfully demonstrated a great potential of the MFC technology for both electricity generation and waste treatment

Self sustainable cathodes for microbial fuel cells

The ultimate goal of this thesis was to investigate and produce an MFC with self-sustainable cathode so it could be implemented in real world applications. Using methods previously employed [polarisation curve experiments, power output measurements, chemical assays for determining COD in wastewater and other elements present in anolyte or catholyte, biomass assessments] and with a focus on the cathode, experiments were conducted to compare and contrast different designs, materials and nutrient input to microbial fuel cells with appropriate experimental control systems.Results from these experiments show that: Firstly, the choice of polymeric PEM membrane showed that the most effective materials in terms of power performance were cation exchange membranes. In terms of cost effectiveness the most promising was CM-I, which was the preferred separator for later experiments.Secondly, a completely biotic MFC with the algal cathode was shown to produce higher power output (7.00 mW/m2) than the abiotic control (1.52 mW/m2). At the scale of the experimental system, the reservoir of algal culture produced sufficient dissolved oxygen to serve the MFCs in light or dark conditions. To demonstrate usable power, 16 algal cathode-designed MFCs were used to power a dc pump as a practical application.It has been presented that the more power the MFC generates, the more algal biomass will be harvested in the connected photoreactor. The biomass grown was demonstrated to be a suitable carbon-energy resource for the same MFC units in a closed loop scenario, whereby the only energy into the system was light.In the open to air cathode configuration various modifications to the carbon electrode materials including Microporous Layer (MPL) and Activated Carbon (AC) showed catholyte synthesis directly on the surface of the electrode and elemental extraction such as Na, K, Mg, from wastewater in a power dependent manner. Cathode flooding has been identified as an important and beneficial factor for the first time in MFCs, and has been demonstrated as a carbon capture system through wet scrubbing of carbon dioxide from the atmosphere. The captures carbon dioxide was mineralised into carbonate and bicarbonate of soda (trona). The novel inverted, tubular MFC configuration integrates design and operational simplicity showing significantly improved performance rendering the MFC system feasible for electricity recovery from waste. The improved power (2.58 mW) from an individual MFC was increased by 5-fold compared to the control unit, and 2-fold to similar sized tubular systems reported in the literature; moreover it was able to continuously power a LED light, charge a mobile phone and run a windmill motor, which was not possible before

Optimisation of the production of cathepsin L1 from a recombinant saccharomyces cerevisiae

Cathepsin L1 is a cysteine protease that has been previously isolated and functionally expressed in Saccharomyces cerevisiae. It has the potential to be employed as a vaccine for liver-fluke disease in cattle and other ruminants. Production of this recombinant enzyme, which is secreted into the media from recombinant yeast, was studied initially in shake flask cultures and subsequently in 5L and 15L fermenters. In early studies, low productivity and especially variations in Cathepsin L1 production was a significant problem. A standard operating protocol (SOP) has been designed to consistently supply an optimum inoculum for large-scale fermentations. This SOP which involved 'blending' colonies for inoculum cultures in conjunction with sub-culturing starter flasks for two successive cycles of 48 hours, proved to be the most successful for consistently high levels of enzyme production during the ensuing fermentation. The pH and temperature optima are pH 6.5 and 30°C respectively for culturing the recombinant yeast to produce both both high biomass levels and high enzyme activity. Addition of casamino acids to the selective media or replacing it with complex YEPD resulted in poor plasmid stability and low Cathepsin L1 production. By supplementing the selective media with extra yeast nitrogen base, using a glucose concentration of 20g/L, enzyme activity increased by 3-4 fold and much higher levels of plasmid stability than observed in non-selective media were sustained. Enzyme activity of 0.74 units/mL were obtained in supplemented media compared to 0.19 units/mL in selective media. Investigations were performed on the constitutive behaviour of the ADH1 promoter, which controls the expression of Cathepsin L1 in this recombinant yeast strain. It revealed that enzyme production is repressed at high concentrations of glucose but gradually increases as glucose is utilised. Cathepsin L1 is still expressed during the ethanol consumption phase, albeit at a slower rate than during the latter stages of glucose consumption