Potentialités des bioélectrodes et des bioréacteurs à biomasse fixée pour l'estimation rapide de la DBO

La méthode standard de détermination de la Demande Biochimique en Oxygène (DBO-5 jours par dilutions) présente certains inconvénients bien connus dont celui de ne pas être exploitable pour la gestion en temps réel d'un procédé d'épuration. De ce fait, divers types d'appareils d'estimation rapide de la DBO, basés sur des méthodes respirométriques, ont été imaginés dans le passé et même commercialisés pour certains.Dans cette étude, des appareils d'estimation rapide de la DBO de types bioélectrode et réacteurs à biomasse fixée (écoulement piston et parfaitement mélangé) ont été mis en oeuvre, après avoir été développés ou modifiés sur base de concepts déjà existants. Une attention particulière a été portée sur la validité de ces appareils, la crédibilité de leurs mesures et la définition de leur champ d'action.De par leur principe de fonctionnement (injection en flux), la bioélectrode et le bioréacteur piston sont des appareils qui doivent être calibrés et dont la biomasse immobilisée doit être préalablement adaptée à l'échantillon à analyser. La solution de calibration est essentielle et doit être qualitativement la plus proche possible de l'échantillon à analyser.Le bioréacteur parfaitement mélangé se distingue des deux premières méthodes car il travaille théoriquement en consommation totale du substrat. La méthode ne nécessite pas de calibration mais implique, avant toute mesure de respirogramme, la détermination d'un paramètre supplémentaire, le coefficient de transfert de l'oxygène.De manière générale, la principale conclusion de l'étude réside dans la difficulté de ces appareils à fournir une DO (Demande en oxygène) corrélable à la DBO5 conventionnelle. Leur champ d'application reste en effet limité aux substrats rapidement biodégradables.Leurs potentialités restent cependant conformes aux besoins de contrôle rapide des charges organiques polluantes en entrée et sortie des stations d'épuration.BOD (Biochemical Oxygen Demand) is an important parameter to characterize organic pollution in aquatic environments and sewage. The five-day period required by the classical dilution method (BOD5) is incompatible with real-time control of a sewage treatment plant. Moreover, the assay procedure (closed respirometer, very diluted samples) is not only far from real growing conditions but also far from conditions in sewage treatment plants.Several devices for rapid BOD estimation, all based on respirometric methods, have been developed and tested. These devices can be grouped into 2 categories:- an immobilized biomass plug-flow reactor and a bioelectrode, both based on a flow injection principle; - an immobilized biomass perfectly mixed reactor, based on an open respirometer principle. We have focused mainly on validating the principles, checking the measurement reliability, and defining more precisely the scope of the various devices.The bioelectrodeThe BOD bioelectrode that we developed relies on a classical configuration that uses a yeast strain (Trichosporon cutaneum) as the biological receptor and a Clark probe as the transducer. The main changes made in this system are as follows:- a second biomembraneless Clark probe was added to the 16-ml measuring cell to serve as a reference probe in order to remove experimental disturbances (temperature, oxygen transfer coefficient, dissolved oxygen concentration, etc.). - the second change was to include the respirogram area among the data available for processing. The signal utilized in this set-up is the difference between the signals provided by the two probes. Our conclusions are as follows:- Concerning the signal processing, manufacturers are currently guided by the desire to develop devices able to estimate the BOD of a large range of substrates or effluents in a very short time (a few minutes for the newest devices). However, it seems useful, even necessary in many cases (complex mixtures of components that are oxidized at variable rates), to use information provided by the respirogram shape and area. This approach allows one to maximize the BOD bioelectrode's range for a given immobilized strain, although the trade-off is a longer total run time. - Concerning the adaptation period for microorganisms, it is impossible to correctly estimate BOD from various effluents without first adapting the biomaterial to the type of substrate to be analyzed. Therefore it is dangerous to consider a BOD bioelectrode as an analytical instrument, because an adaptation period is required after any change in the composition or, even more so, type of effluent. - Concerning the correspondence between rapid BOD and BOD5, the BOD sensor can detect only the BOD of soluble compounds that can diffuse through the biomembrane and that will be metabolized during the time of analysis. The difference observed between BOD sensor and BOD5 depends on the calibration solution but also and even more on the structure and size of the molecules constituting the sample to be analyzed. - Concerning the choice of calibration solution, choosing the right calibration solution is crucial. The calibration solution should therefore be qualitatively as close as possible to the test sample. The immobilized biomass bioreactorsThe plug-flow reactorThe plug-flow reactor design was validated for simple substrates; its working principle is similar to the bioelectrodes, since it relies on flow-injection analysis (FIA). In the case of the plug-flow reactor, the only usable information for BOD estimation is the respirogram area, as the peak height quickly reaches a rather constant value due to saturation of the immobilized microorganisms. However, the importance of the many physical and biological processes that occur concomitantly in the system (transfer, adsorption, substrate consumption, substrate saturation phenomena, dilution rate, etc.) makes a theoretical mathematical model of the reactor more difficult to establish. A long-term, more fundamental study of various natural or artificial substrates might ultimately enable us to reach such a goal.On the other hand, a variant of this reactor that recirculates the partially-degraded effluent until it is completely consumed yielded a linear correlation between system response (respirogram area) and substrate amount. In this system, oxidation of the rapidly biodegradable substrates is total under our operating conditions. This alternative reactor seems to have some very interesting possibilities, especially with regard to the automation of the system.The perfectly mixed reactorThe utilization of the perfectly mixed reactor for rapid BOD estimation is based on two sequential experiments - although this has the disadvantage of increasing the total run time - to obtain the respirogram area (S) and oxygen transfer coefficient (KL.a), as the units of the product (KL.a x S) of these parameters are equivalent to those of oxygen demand and their product is the variable that best correlates with the substrate injection volume. This correlation was observed for a large range of substrates. In the case of the perfectly mixed reactor, unlike bioelectrodes,- diffusion processes have no effect on measurement, as the substrate is consumed completely during the experiment; - the substrate consumption rate does not affect the measurements, thereby freeing the method from the influence of various experimental parameters (air flow-rate, the quantity of biomass, the liquid volume in the reactor, etc.), as our tests have shown. For a given substrate, the linearity of the correlation between the product of KL.a x S and the amount of substrate is generally excellent. The attempt to correlate the system's response with the BOD5 measurements for a variety of substrates proved to be difficult, however. Although the bioreactor's analytical range is wider than that of bioelectrodes, the system still fails to give any measurable responses for complex substrates such as starch, cellulose, proteins, etc., and the KL.a x S values estimated by the bioreactor remained much lower than the corresponding BOD5 values. KL.a x S is probably characteristic of the portion of the substrate that is degraded rapidly by the microorganisms to meet their immediate energy needs, whereas the remaining substrate is probably kept for reserve and biomass synthesis