Optimització de la immobilització bacteriana per la bio-producció d’hidrogen amb alfa-proteobacteris

Fleitas García, Ana Rosa
Currently, we find ourselves in a period of energy transition based on the search of cleaner and more sustainable energy systems allowing the progressive disuse of fossil compounds’ combustion-based systems in order to reduce pollutant emissions and greenhouse gases mainly responsible of the climate change. Hydrogen (H2) is a carbon-free fuel representing an ecological and renewable energy source that can be produced from bio-electrochemical systems. These systems take advantage of the capacity of microorganisms to carry out oxidation-reduction reactions when an electric potential is applied, and they are presented as an alternative to achieve the aforementioned objectives. These systems can be exploited in the hydrogen’s production using bacteria that have electro-chemical activity and that, also, have the enzymes necessary for their production. These characteristics are fulfilled by the purple non-sulfur bacteria (PNS) and within this group, more specifically, the alpha-proteobacteria. Bacterial immobilization is useful for the optimization of bio-processes since it provides a series of advantages over working with cells in suspension: it can be operated during the growth phase of bacteria for a long time period, it takes less space and volume of growth medium phase due to the fact that the cells are concentrated and a greater catalytic stability and greater resistance to toxins or external enzymatic inhibitors can be achieved. In the case of the bio-electrochemical systems, bacterial immobilization is important because, in addition, it allows to reduce the start-up time that is, the necessary time between the inoculation of the system and the operation of the complete performance. This project aims to verify the effectiveness of bacterial immobilization protocols in the production of H2 by the alpha-proteobacteria Rhodobacter sp. DSM 5864 and the isolate C1S119.2 of Rhodopseudomonas palustris in a bio-electrochemical system using an H-type reactor. To achieve this purpose, bio-electrochemical systems have been implemented by providing these bacteria with electrons and CO2 as a source of carbon in a mineral medium. For Rhodobacter sp. DSM 5864 we have applied potentials of -0,4, -0,6 and -0,8V vs. SHE (standard hydrogen electrode) and in the case of the isolate C1S119.2 we have fixed a potential of -0,8V vs. SHE. In both cases maintaining a temperature of 30°C and in a constant agitation. Hydrogen concentrations achieved in the liquid phase of the cathode were measured using a hydrogen probe and, from these concentrations, we could determine the hydrogen productions. We had used agar for the immobilization. In addition, for each experiment, controls were carried out without immobilization of the cathode with agar and with immobilization of the cathode with agar without the bacteria. It was determined the redox activity of Rhodobacter sp. through a cyclic voltammetry and the viability of the isolate C1S119.2 through the addition of 1,5 mL of Na-acetate (2,5 mM) in the cathodic chamber at specific moments. It was concluded that the bacterial strains used are not the most adequate with the conditions of the system with which we have been working. In addition, it was corroborated that agar isn’t a suitable material to perform the immobilization because it prevents the diffusion to the medium. In addition, the immobilization protocol was optimized thanks to the fact of measuring the optical density of the cultures used for the immobilization, the use of agar concentrations of 10 g/L and the application of the mixture of cells and agar covering the cathode thanks to the pocket designed by the team ​
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