Up-scaling the production of bacteria for self-healing concrete application

Self-healing concrete has become the main focus of several research programs that have been bringing together people from academia and industry. The remediation of the micro-cracks that are formed in the concrete structures has been the aim of these research programs. These cracks are a quite well known problem that can lead to corrosion of the steel reinforcement and, in the worst cases, to the failure of the entire concrete structure. The maintenance costs due to the repair of these cracks can be of the order of € 130 (direct costs) per m3 of concrete. Hence, a self-healing technology is of interest to the industrial partners. Scientific studies indicate that a Microbial Induced Carbonate Precipitation (MICP), using microbial spores as active agent, can be an alternative for the actual repair methods. The precipitation of this concrete compatible material allows the filling of the cracks restoring the water tightness of the concrete structure. By avoiding the water ingression one also avoids the corrosion of the reinforcement. However, the production of bacterial spores is yet imposing considerable costs. Industrial concrete producers and contractors are willing to pay extra € 15 to € 20 per m3 of concrete for a bio-based self-healing product if the increase in service life is ensured. There is an urgent need to develop the production of a bio-additive to make the biological self-healing industrial applicable. Furthermore, protective carriers or encapsulation materials have been used in microbial self-healing concrete studies due to the harsh environment of concrete, i.e. very high pH (~ 13), relatively small pore size (< 0.1 nm) and dry conditions. These protection methodologies improve the survival probability of the bacterial agents once incorporated in the concrete matrix. This study deals with the up-scaling of suitable bacterial spores for self-healing concrete applications and with the study of some protection methodologies. The first part of this work is directly related with the up-scaling of axenic bacterial spore cultures while a non-axenic bacterial culture is studied in a second part. In the third part of the study different protection methods were tested in order to evaluate concrete compatibility with industrially available materials. The fourth and last part of this study deals with the economic and ecological consequences of this technology and its industrial feasibility. Up-scaling of axenic bacterial spore cultures Both Bacillus sphaericus and Bacillus cohnii spores were first produced at lab scale in order to evaluate and select the best growth conditions. Different media formulations were used to produce the bacterial spores in order to evaluate the most optimal one. MBS (Minimal Basal Salts) and SM2 (Sporulation Medium 2) media were selected among the tested possibilities as sporulation media for Bacillus sphaericus and Bacillus cohnii respectively. Biological parameters such as microbial activity and calcium carbonate precipitation capabilities were evaluated for both Bacillus sphaericus and Bacillus cohnii vegetative cells resulting from the germination of their respective bacterial spores. Increasing the production volume showed to have no effect on either microbial activity or calcium carbonate precipitation capabilities of both axenic cultures. Despite of the good microbial activity observed, a decrease in the biomass yield from 3.5 to 2.5 g CDW/L was registered in the Bacillus sphaericus cultures while no difference was noticed in the Bacillus cohnii cultures. Nevertheless, the biomass yield of the Bacillus sphaericus was still a factor 6 higher than the one of Bacillus cohnii cultures. Moreover, the amount of calcium carbonate precipitated by the Bacillus sphaericus cells was 5 times higher than the one obtained with the Bacillus cohnii cells. Additionally, the sporulation time required for Bacillus sphaericus cultures at 5 L scale, defined as the time required to obtain 90 % of the biomass present in the form of spores, was a factor 4 lower when compared with the one for Bacillus cohnii spores. Both bacterial spores unfortunately provoked a decrease in compressive strength when incorporated in mortar cubes. Nevertheless, both bacterial cultures, produced at lab scale, were able to perform self-healing in mortar specimens when properly immobilized in diatomaceous earth. Both types of bacterial spores produced at lab scale were able to originate vegetative cells capable of closing cracks up to 0.4 mm. For the bacterial cultures produced at 5 L scale only Bacillus sphaericus was tested regarding self-healing capabilities, revealing the complete closure of cracks up to 0.4 mm. Yet, the survival capability of Bacillus cohnii spores produced at 5 L scale was tested upon incorporation in mortar specimens. These bacterial spores, when immobilized in diatomaceous earth, were able to survive the mortar incorporation maintaining their viability. Up-scaling of non-axenic bacterial spores culture In this part, the production of a non-axenic ureolytic bacterial culture able to sporulate was evaluated. A powderous material containing an efficient ureolytic microbial community (Cyclic EnRiched Ureolytic Powder or CERUP) was developed. Ureolytic activity, calcium carbonate precipitation capability and the effects in concrete were evaluated at production scales of 5 and 50 L. The non-axenic culture obtained following this new selective process, at both 5 L and 50 L scales, showed to be as good as the benchmark i.e. axenic vegetative cells resultant from the germination of Bacillus sphaericus spores, both in terms of urea hydrolysis (20 g urea/L in 24 hours) and calcium carbonate precipitation (0.5 g CaCO3/g CDW.h). Plain incorporation of CERUP in concrete was found to be efficient at levels of 0.5 and 1 % of the cement weight. The use of inexpensive materials to produce CERUP was beneficial for the concrete application since it decreases the total production cost. Protection methods and their compatibility with concrete The use of axenic cultures for self-healing concrete requires some sort of protection of the bacterial agents. On the one hand, biological compatibility is, naturally, an important characteristic that must be attained by the protection material. On the other hand, concrete compatibility and cost feasibility must also be strongly considered. In this part of the study the influence of different protection methodologies was evaluated. Commercially available possible protection approaches (diatomaceous earth, metakaolin, zeolite and air entrainment) were tested regarding their influence on mortar setting and compressive strength when plain incorporated or combined with Bacillus sphaericus spores and their respective nutrients. The influence of CERUP, salt protected/immobilized, was also investigated within the same scope. The most severe effect in setting time was observed in all samples containing nutrients for Bacillus sphaericus with a significantly undesirable increase of 340 minutes. Samples containing Bacillus sphaericus spores showed the most significant undesirable decreases in compressive strength up to 68 %. Yet, the use of the novel salt protected/immobilized CERUP had no significant effect on the setting and the compressive strength of mortar. These results indicate that the use of non-axenic and self-protected bacterial cultures must be considered as a valuable alternative for the currently proposed axenic cultures. Economical and ecological aspects of the microbial based self-healing concrete technology The last part of the present study deals with cost calculation regarding both axenic and non-axenic production of self-healing bacterial agents. Concrete is a very large market and thus, one must consider the need to produce several thousands of tons of biological agent to supply it. Therefore, an economical evaluation upon the different possibilities must be conducted. It was found that the production cost of axenic cultures is, at least, a factor 10 higher than the one of non-axenic cultures. Nevertheless, despite of the lower production cost calculated for the non-axenic cultures, these are still very expensive for concrete application. Hence, process optimization and new techniques must be further studied in order to bring the bacterial self-healing concrete technology closer to full scale application. Carbon dioxide (CO2) emissions caused by the production of the bacterial agents was also evaluated. It was found that both axenic cultures are responsible for a higher CO2 emission when compared with the non-axenic ones. However, the production of Bacillus sphaericus spores showed to be responsible for only 2.7 times more CO2 emitted when compared with CERUP while the production of Bacillus cohnii spores caused CO2 emissions 40 times higher than the ones caused by the production of CERUP. Overall, the non-axenic culture (CERUP) showed not only a more feasible production process as also a lower CO2 footprint. Therefore, the use of non-axenic bacterial cultures for self-healing concrete purposes is more industrially feasible than the use of axenic ones. Biological self-healing concrete requires further optimization in order to reach industrially feasible values. Nonetheless, the technology is of considerable potential and warrants to be further investigated. Future research must be conducted using axenic but particularly also with non-axenic bacterial cultures which can be more robust and hold the potential to be more cost effective.