Estela Tapia-Venegas’s research while affiliated with Playa Ancha University of Educational Sciences and other places

Baccharis macraei Hook. & Arn (Asteraceae), commonly known as Vautro, is found in the coastal areas of central-southern Chile, including the industrial zone of Quintero-Puchuncaví, known for the contamination of its soils with heavy metals, which together with other factors generate abiotic stress in plant species, against which they present defensive mechanisms. For this reason, the objective was to evaluate the effect of abiotic stress generated by the proximity of B. macraei to the industrial complex by assessing the physiological and metabolic states reported by the extracts and compounds isolated from the species, as well as the photosynthetic capacity, metal content and production, and antioxidant activity and cytotoxicity against tumorigenic cell lines of the phytoconstituents. To this end, B. macraei was collected at two different distances from the industrial complex, observing that the closer the species is, the greater the concentration of copper in the soil, generating a decrease in the rate of electron transport in situ, but an increase in antioxidant activity with low cytotoxicity. This activity could be due to the presence of flavonoids such as Hispidulin, Cirsimaritina, and Isokaempferida, as well as monoterpenes, oxygenated and non-oxygenated sesquiterpenes identified in this study.

Hydrogen (H2) as a clean fuel holds global potential and can be produced through bio-processes. To enhance bioH2 yields, integrated systems have been proposed, combining dark fermentation (DF) of wastewater with a subsequent photofermentation (PF) stage involving purple non-sulfur (PNS) bacteria. Mixed cultures of PNS bacteria and their microbial ecology have been relatively understudied despite the known benefits of mixed cultures in industrial applications. The aim of this study was to obtain various mixed cultures of PNS bacteria under different environmental conditions during the enrichment stage. Four different mixed cultures were obtained (A, B, C, and D). However, in the H2 production phase, only Consortium A, which had been enriched with malic acid as the carbon source, exposed to 32 W m−2 of irradiance, and subjected to intermittent agitation, produced H2 with a yield of 9.37 mmol H2 g−1 COD. The consortia enriched were a hybrid of PF and DF bacteria. Especially in Consortium A, Rhodopseudomonas palustris was the dominant organism, and various DF bacteria were positively associated with H2 production, with their dominance comparable to that of PNS bacteria. Despite the reported low yields, optimizing environmental conditions for this culture could potentially enhance hydrogen production from DF effluents.

Senecio fistulosus, an endemic plant in Chile, is highly regarded for its medicinal properties and is popular in alternative medicine. It thrives even in polluted areas, like Puchuncaví Valley, Chile. Therefore, the study aimed to assess the impact of industrial pollution in Puchuncaví Valley, Chile, on the phytoconstituent content, as well as the antioxidant and cytotoxic activities, of S. fistulosus. Phenols, flavonoids, and anthraquinones content were measured, alongside the assessment of antioxidant activities. Additionally, a GC-MS analysis was conducted to profile the phytoconstituents, while the cytotoxic potential was evaluated in HT-29 and MCF-7 and cell line non-tumorigenic MCF-10. The Wild sample exhibited a greater concentration of phytoconstituents (0 to 169.48 mg·L−1) compared to the Commercial control (0 to 95.38 mg·L−1), directly correlating with its antioxidant activity. While the Wild species showed cytotoxic activity, the Commercial control demonstrated cytotoxic effects on MCF-10 and MCF-7. Noteworthy compounds identified were hexadecanoic acid (12.76 to 19.57% relative area) and (Z,Z,Z)-9,12,15-octadecatrienoic acid (18.36% relative area), with anticancer properties. In conclusion, the abiotic stress experienced by S. fistulosus led to higher phytoconstituent content and improved antioxidant activity when contrasted with the Commercial control. The Commercial species showed increased cytotoxic activity against both tumorigenic and non-tumorigenic cell lines.

This book chapter aims to address the microbial diversity associated with implementation prospects of nonconventional technologies for nitrogen removal in wastewater treatment plants. Special focus will be placed on the environmental advantages offered by these processes in contrast to conventional ones as heterotrophic denitrification and anammox. In addition, the operational and environmental conditions that promote the development of these microbial populations for nitrogen removal in wastewater will be addressed. Nitrate removal by an autotrophic process using sulfur compounds forms is the better characterized process and has been extensively applied in the treatment of wastewater with low C/N ratios rich in nitrates and sulfides. Hydrogenotrophic denitrification has attracted attention for application to nitrate removal from groundwater and industrial wastewater with very low organic matter content and high nitrate concentrations. This is because the process is clean, with a low biomass production and no residual organic matter. Anaerobic methanotrophic microorganisms using nitrate or nitrite (N-damo) have attracted attention for application in cooperation with anaerobic ammonia-oxidizing bacteria (anammox), resulting in the simultaneous removal of nitrogen (ammonium, nitrite, and nitrate) and methane.

One of the bottlenecks of the hydrogen production by dark fermentation is the low yields obtained because of the homoacetogenesis persistence, a metabolic pathway where H2 and CO2 are consumed to produce acetate. The central reactions of H2 production and homoacetogenesis are catalyzed by enzyme hydrogenase and the formyltetrahydrofolate synthetase, respectively. In this work, genes encoding for the formyltetrahydrofolate synthetase (fthfs) and hydrogenase (hydA) were used to investigate the diversity of homoacetogens as well as their phylogenetic relationships through quantitative PCR (qPCR) and next-generation amplicon sequencing. A total of 70 samples from 19 different H2-producing bioreactors with different configurations and operating conditions were analyzed. Quantification through qPCR showed that the abundance of fthfs and hydA was strongly associated with the type of substrate, organic loading rate, and H2 production performance. In particular, fthfs sequencing revealed that homoacetogens diversity was low with one or two dominant homoacetogens in each sample. Clostridium carboxivorans was detected in the reactors fed with agave hydrolisates; Acetobacterium woodii dominated in systems fed with glucose; Blautia coccoides and unclassified Sporoanaerobacter species were present in reactors fed with cheese whey; finally, Eubacterium limosum and Selenomonas sp. were co-dominant in reactors fed with glycerol. Altogether, quantification and sequencing analysis revealed that the occurrence of homoacetogenesis could take place due to (1) metabolic changes of H2-producing bacteria towards homoacetogenesis or (2) the displacement of H2-producing bacteria by homoacetogens. Overall, it was demonstrated that the fthfs gene was a suitable marker to investigate homoacetogens in H2-producing reactors. KEY POINTS: • qPCR and sequencing analysis revealed two homoacetogenesis phenomena. • fthfs gene was a suitable marker to investigate homoacetogens in H2 reactors.

The oxidation-reduction potential (ORP) is an important factor in H2 production via dark fermentation however its effect over microbial diversity in an acidogenic sludge has not, been well studied. This work studies the effect of ORP controlled by hydrogen peroxide and potassium ferricyanide on continuous hydrogen production and microbial diversity in an acidogenic sludge fed (HRT 12 h and pH 5.5) with glycerol. Results show that the more oxidizing ORP control environment (−540 mV) improves H2 yield by 50–70% (0.31–0.51 molH2/mol glycerol) over non-ORP control conditions. Oxidizing ORP values were shown to enrich microorganisms of the genus Clostridium, which have been linked to high H2 yields. Therefore, controlling ORP in an acidogenic sludge was shown to directly modify microbial diversity at the genus level, and could likely to indirectly regulate metabolic function. Additionally, metabolic pathways were regulated by the kind of agent used.

Hydrogen (H2) production by dark fermentation can be performed from a wide variety of microbial inoculum sources, which are generally pre-treated to eliminate the activity of H2-consuming species and/or enrich the microbial community with H2-producing bacteria. This paper aims to study the impact of the microbial inoculum source on pre-treatment behavior, with a special focus on microbial community changes. Two inocula (aerobic and anaerobic sludge) and two pre-treatments (aeration and heat shock) were investigated using glycerol as substrate during a continuous operation. Our results show that the inoculum source significantly affected the pre-treatment efficiency. In aerobic sludge no pre-treatment is necessary, while in anaerobic sludge the heat pre-treatment increased H2 production but aeration caused unstable H2 production. In addition, biokinetic control was key in Clostridium selection as dominant species in all microbial communities. Lower and unstable H2 production were associated with a higher relative abundance of Enterobacteriaceae family members. Our results allow a better understanding of H2 production in continuous systems and how the microbial community is affected. This provides key information for efficient selection of operating conditions for future applications.

H2 production by dark fermentation using mixed cultures has been studied intensively during the last two decades, and its feasibility has been demonstrated. Different substrates, operational conditions, and reactor technologies have been widely studied and there is a general agreement that the use of non-sterile fermentable substrates is required to make the process feasible for scaling up. Nonetheless, stability problems during long term operation may hinder its application at large scale. This work, written by members of the Latin American Biohydrogen Network, analyse and discuss instability causes and possible solutions in the H2 production by dark fermentation. It is concluded that instability is mostly linked to the biotic aspects of the process (i.e., changes in the microbial community composition, presence of organisms that consume hydrogen and compete for the substrate, and accumulation of fermentation products); regardless of the reactor configuration. However, some problems like excessive growth of microorganisms and methanogens presence were mostly reported in fixed bed reactors and granular sludge reactors. The novelty of this work relies on the comprehensive revision of the main causes behind the unstable and low hydrogen production and how these causes are linked to the technology used. The strategies to overcome the problems as well as the potential implications are also analysed.

Fermentative hydrogen production requires a continuous products-removal and effective upgrading steps to improve its general performance. Therefore, implementation of new technologies capable of achieving both requirements is essential. We present the operationalbehavior of a new process concept based on integration of membranes for gas separation and fermentation technology. This process, which we term as hydrogen extractive membrane bioreactor consists of coupling two dense polymeric membranes to a hydrogen producing culture. The process automatization of this system was essential to maintain the proper operational pressures in the membrane module and in the bioreactorgas- phase. This system was able to extract and partially separate the hydrogen and carbondioxide generated. The hydrogen partial pressure was reduced from 55.5 to 49 KPa, which means an increase of hydrogen yield of 16.3% (1.1e1.28 mol-H2/mol-glucose). Simultaneously, the implemented system generated a final hydrogen stream 13% (v/v) more concentrated than a conventional process.

Hydrogen production by dark fermentation is an emerging technology of increasing interest due to its renewable feature. Recent scientific advances have well investigated the operational conditions to produce hydrogen through the valorization of several wastes or wastewaters. However, the development of standardized protocols to accurately assess the biohydrogen potential (BHP) is of crucial importance. This work is the first interlaboratory and international effort to validate a protocol estimating hydrogen potential using batch tests, using glucose as individual model substrate. The repeatability of the hydrogen potential (HP) increased with variations of the proposed protocol: reducing substrate concentration, increasing the buffer capacity, and using an automatic device. The interlaboratory variation of the HP was reduced from 32 to 12%, demonstrating the reproducibility and robustness of the proposed protocol. Recommendations to run BHP tests were formulated in terms of i) repeatability and reproducibility of results, ii) criteria for results validation and acceptance, iii) workload of the proposed protocols.