Enzymatic pretreatment of microalgae using fungal broth from Trametes versicolor and commercial laccase for improved biogas production

Coupling microalgae production to wastewater treatment can reduce the costs of microalgae production for non-food bioproducts and energy consumption for wastewater treatment. Furthermore, microalgae anaerobic digestion can be enhanced by applying pretreatment techniques. The aim of this study is to improve the biogas production from microalgal biomass grown in urban wastewater treatment systems by applying an enzymatic pretreatment with crude fungal broth and commercial laccase. To this end, the fungus Trametes versicolor was cultured, and the enzymatic activity of the culture broth analysed by measuring laccase concentration. The results showed that both the fungal broth and commercial laccase pretreatment (100 U L- 1) over an exposure time of 20 min increased the methane yield in batch tests. Indeed, the fungal broth pretreatment increased the methane yield by 74%, while commercial laccase increased the methane yield by 20% as compared to non-pretreated microalgal biomass. In this manner, laccase addition enhanced microalgal biomass anaerobic biodegradability, and addition of T. versicolor broth further improved the results. This fact may be attributed to the presence of other molecules excreted by the fungus.Peer ReviewedPostprint (author's final draft

Improving biogas production from microalgae by enzymatic pretreatment

In this study, enzymatic pretreatment of microalgal biomass was investigated under different conditions and evaluated using biochemical methane potential (BMP) tests. Cellulase, glucohydrolase and an enzyme mix composed of cellulase, glucohydrolase and xylanase were selected based on the microalgae cell wall composition (cellulose, hemicellulose, pectin and glycoprotein). All of them increased organic matter solubilisation, obtaining high values already after 6 h of pretreatment with an enzyme dose of 1% for cellulase and the enzyme mix. BMP tests with pretreated microalgae showed a methane yield increase of 8 and 15% for cellulase and the enzyme mix, respectively. Prospective research should evaluate enzymatic pretreatments in continuous anaerobic reactors so as to estimate the energy balance and economic cost of the process.Peer ReviewedPostprint (author's final draft

Effect of cultivation conditions on β-estradiol removal in laboratory and pilot-plant photobioreactors by an algal-bacterial consortium treating urban wastewater

The use of microalgal consortia for urban wastewater treatment is an increasing trend, as it allows simultaneous nutrient removal and biomass production. Emerging contaminants proposed for the list of priority substances such as the hormone 17β-estradiol are commonly found in urban wastewater, and their removal using algal monocultures has been accomplished. Due to the inherent potential of algae-based systems, this study aimed to assess the capability of native photobioreactor biomass to remove 17β-estradiol under indoor and outdoor conditions. At the same time, the microbial community changes in regular and bioaugmented operations with Scenedesmus were assessed. The results show that almost complete removal (>93.75%) of the hormone 17β-estradiol can be attained in the system under favourable seasonal conditions, although these conditions greatly influence biomass concentrations and microbial diversity. Even under the harsh conditions of low temperatures and solar irradiation, the established consortium removed more than 50% of the pollutant in 24 h. While species from genus Chlorella were stable during the entire operation, the microbial diversity analysis revealed that assorted and evenly distributed populations stimulate the removal rates. Bioaugmentation assays proved that the input of additional biomass results in higher overall removal and decreases the yield per mg of biomass

Enzymatic pretreatment of microalgae using fungal broth from Trametes versicolor and commercial laccase for improved biogas production

Es un article del grup de recerca BioremUAB (2014SGR476)Coupling microalgae production to wastewater treatment can reduce the costs of microalgae production for non-food bioproducts and energy consumption for wastewater treatment. Furthermore, microalgae anaerobic digestion can be enhanced by applying pretreatment techniques.. The aim of this study is to improve the biogas production from microalgal biomass grown in urban wastewater treatment systems by applying an enzymatic pretreatment with crude fungal broth and commercial laccase. To this end, the fungus Trametes versicolor was cultured, and the enzymatic activity of the culture broth analysed by measuring laccase concentration. The results showed that both the fungal broth and commercial laccase pretreatment (100 U/L) over an exposure time of 20 min increased the methane yield in batch tests. Indeed, the fungal broth pretreatment increased the methane yield by 74%, while commercial laccase increased the methane yield by 20% as compared to non-pretreated microalgal biomass. In this manner, laccase addition enhanced microalgal biomass anaerobic biodegradability, and addition of T. versicolor broth further improved the results. This fact may be attributed to the presence of other molecules excreted by the fungus

Degradation of pharmaceutical compounds by microalgae : photobioreactor wastewater treatment, biomass harvesting and methanization /

Premi Extraordinari de Doctorat concedit pels programes de doctorat de la UAB per curs acadèmic 2017-2018Els contaminants emergents (ECs) són un ampli grup de compostos orgànics detectats en diversos compartiments ambientals que degut a la seva presència en el medi ambient i la seva difícil degradació han aixecat un gran interès en la comunitat científica. Tot i que la seva concentració normalment està compresa entre els ng/L i els μg/L, poden representar una amenaça per a la salut humana i el medi ambient ja que són excretats per la orina i els fems, ja sigui en forma de compost actiu o com a metabòlit, ja que no són completament assimilats. D'entre tots els emergents, els compostos disruptors endocrins (EDCs) i els principis actius dels fàrmacs (PhACs) generen especial preocupació. Està àmpliament acceptat que la seva principal font d'entrada al medi ambient és a través dels efluents de les plantes depuradores (WWTP), on els tractaments convencionals de llots actius no són capaços de degradar-ne la majoria, sent capaços d'arribar a les aigües superficials, subterrànies i, posteriorment, a l'aigua potable. Per tant, s'han de buscar tractaments alternatius. Un d'aquests tractaments podria ser l'ús de les microalgues aprofitant la seva capacitat pel tractament d'aigües residuals, eliminant els nutrients de l'aigua residual i la seva posterior conversió en biocombustibles. Aquesta tesi avalua diferents aspectes relacionats amb la degradació de contaminants emergents i tractament d'aigües amb microalgues. A més de l'estudi d'un dels principals colls d'ampolla dels sistemes de microalgues, la collita, que té l'avantatge de clarificar l'aigua residual tractada i concentrar la biomassa algal per a la seva conversió a biogàs. Tant, cultius purs com efluents de microalgues s'han considerat. Primerament, s'ha estudiat la degradació individual de dos compostos estrogènics. Aquests dos compostos estrogènics s'han introduït recentment a la llista de substàncies prioritàries de la directiva de l'aigua: 17α-etinileestradiol (EE2) i 17β- estradiol (E2). La seva degradació s'ha monitoritzat a partir dels cultius purs de Chlamydomonas reinhardtii i Pseudokirchneriella subcapitata a nivell de laboratori. Per tal d'obtenir més informació sobre els mecanismes de degradació s'ha fet la identificació dels productes de transformació. A més, la degradació d'una mescla de 10 fàrmacs (9 antibiòtics i un antidepressiu) s'ha avaluat en cultius purs de microalgues a escala laboratori. S'ha realitzat la identificació dels productes de transformació de tres PhACs seleccionats. Posteriorment, un dels antibiòtics, la ciprofloxacina, s'ha analitzat. Els seus mecanismes d'eliminació han estat estudiats en sistemes algals, tant en reactors a escala laboratori com en escala pilot. Per altra banda, s'ha dissenyat un fotobioreactor (PBR) d'algues tractant aigua residual urbana i en operació durant 6 mesos. S'ha monitoritzat el seu seguiment i s'han provat diferents condicions d'operació. A més, un cop a l'estat estacionari l'eliminació de PhACs s'ha estimat i s'ha caracteritzat la població microbiana. També s'ha estudiat l'eliminació del compost estrogènic E2 en el PBR. En aquesta tesi també es remarca la importància de la collita de microalgues. Tres tècniques de collita (i.e., sedimentació natural, coagulació-floculació i la tècnica innovadora de co-pel·letització utilitzant el fong Trametes versicolor) s'han aplicat en dos efluents reals de microalgues i a una suspensió pura de Chlamydomonas reinhardtii. Finalment, s'ha dut a terme la metanització de biomassa exhausta. Abans del procés de digestió anaeròbia, per tal de solubilitzar la paret cel·lular les algues han estat sotmeses a un pretractament enzimàtic. S'han provat enzims específics i no específics, així com l'efecte combinat utilitzant una mescla d'enzims. Aquest estudi es completa amb la valorització de biomassa fúngica que prové del tractament d'efluents.Emerging contaminants (ECs) are a wide range of organic compounds detected in many environmental compartments that have raised an increasing interest in the scientific community due to their ubiquitous presence in the environment and their difficult degradation. Even though their environmental concentration is usually in the range of ng/L to μg/L, they still represent a threat to human health and environment since they are excreted with urine and faeces either as active substances or metabolites, because they are not completely assimilated. Among emerging contaminants, endocrine disrupting compounds (EDCs) and pharmaceutical active compounds (PhACs) are of major concern. It is widely accepted that the main source to the environment are the effluents of wastewater treatment plants (WWTP), where conventional activated sludge treatments are not able to degrade most of them being able to reach surface, groundwater and subsequently, drinking water. Therefore, alternative treatments should be found. One of those alternatives might be the use of microalgae by taking advantage of their capacity for wastewater treatment removing the nutrients contained in the wastewater and further microalgal biomass conversion into biofuels. The present thesis assesses different factors related to microalgal degradation of emerging contaminants and wastewater treatment. As well as the study of one of the major bottlenecks on microalgal systems, the harvesting, which has the advantage to clarify the treated wastewater and concentrate the microalgal biomass for further biogas production. Pure microalgal cultures and real microalgal effluents have been considered. First of all, individual degradation of two estrogenic compounds has been studied. The two estrogenic compounds evaluated have been recently incorporated in the priority substances in the Water Framework Directive: 17α-ethinylestradiol (EE2) and 17β- estradiol (E2). Their degradation by Chlamydomonas reinhardtii and Pseudokirchneriella subcapitata pure cultures at laboratory scale conditions has been monitored. To obtain further insights in the mechanism of degradation the transformation products have been identified. Moreover, the removal of a mixture of 10 PhACs (9 antibiotics and an antidepressant) has also been assessed in pure microalgal cultures at laboratory scale conditions. Transformation products from three chosen PhACs have been identified. One of the antibiotics, ciprofloxacin, was further evaluated. The removal mechanisms have been studied in real algal ponds in both, laboratory and pilot scale reactors. On the other hand, an algal photobioreactor (PBR) treating urban wastewater has been designed and operated during 6 months. Its performance has been monitored and different operating conditions have been tested. Furthermore, PhACs removal has been evaluated during the steady state and microbial diversity has been identified. The estrogenic compound E2 removal in the PBR has been studied. The importance of microalgal harvesting has been highlighted in the thesis. Three different harvesting techniques (i.e., natural sedimentation, coagulation-flocculation and the novel technique of co-pelletization using Trametes versicolor fungus) have been evaluated using two real microalgal effluents and a pure Chlamydomonas reinhardtii suspension. Finally, methanization of exhausted biomass has been conducted. Prior the anaerobic digestion process microalgal biomass has been submitted to an enzymatic pretreatment for its cell wall solubilisation. Specific and non-specific enzymes have been tested as well as the synergistic effects between an enzymatic mixture. The study is completed valorising fungal biomass coming from the treatment of effluents

Microalgae cultivation on wastewater digestate: β-estradiol and 17α-ethynylestradiol degradation and transformation products identification

Selenastrum capricornutum and Chlamydomonas reinhardtii were tested for possible biodegradation of the hormones β-estradiol (E2) and 17α-ethinylestradiol (EE2) when cultured in anaerobic digester centrate (ADC). Neither ADC nor the hormones had a negative or toxic effect on the microalgae growth but enhanced it. E2 and EE2 biodegradation was evaluated under different culture conditions. After 7 days of treatment, between 88% and 100% of E2 was removed by S. capricornutum. Overall, 42 and 54% of the removal was attributed to biodegradation processes, while the rest of the removal was due to adsorption onto the algae biomass. For EE2, removals between 60 and 95%, depending on the culture conditions, were achieved, with biodegradation accounting for 20-54% of the removal. E2 and EE2 were completely removed in the experiments performed with C. reinhardtii, except for EE2 in the presence of ADC, which decreased to 76%. However, C. reinhardtii presented higher adsorption percentages: 86% and 71% after 7 days for E2 and EE2, respectively. Transformation products (TPs) of E2 and EE2 generated in each treatment were also monitored. Two TPs were tentatively proposed as degradation products of E2 and EE2 by the algae. In addition, the removal of 26 endocrine disruptors and related pollutants present in the centrate was also monitored: bisphenol A was completely removed, whereas tris(2-butoxyethyl)phosphate was only removed in the absence of hormones

Performance of microalgal photobioreactor treating toilet wastewater : pharmaceutically active compound removal and biomass harvesting

In this study, a 1200L outdoor pilot scale microalgal photobioreactor (PBR) was used for toilet wastewater (WW) treatment and evaluate its ability to remove pharmaceutically active compounds (PhACs). The PBR was operated at two different hydraulic retention times (HRTs), which were 8 and 12days, during Period I (September-October) and Period II (October-December), respectively. Algal biomass concentrations varied by operating period because of seasonal changes. Nutrients (ammonia, nitrogen and total phosphorous) and chemical oxygen demand (COD) were monitored and efficiently removed in both periods (>80%), attaining the legislation limits. At the theoretical hydraulic steady state in both periods, pharmaceutical removal reached high levels (>48%). Two harvesting techniques were applied to the PBR microalgae effluent. Gravity sedimentation was efficient for biomass removal (>99% in 7min) in Period I when large particles, flocs and aggregates were present. In contrast, a longer sedimentation time was required when biomass was mainly composed of single cells (88% clarification in a 24h in Period II). The second harvesting technique investigated was the co-pelletization of algal biomass with the ligninolytic fungus Trametes versicolor, attaining >98% clarification for Period II biomass once pellets were formed. The novel technology of co-pelletization enabled the complete harvesting of single algae cells from the liquid medium in a sustainable way, which benefits the subsequent use of both biomass and the clarified effluent

Improving biogas production from microalgae by enzymatic pretreatment

In this study, enzymatic pretreatment of microalgal biomass was investigated under different conditions and evaluated using biochemical methane potential (BMP) tests. Cellulase, glucohydrolase and an enzyme mix composed of cellulase, glucohydrolase and xylanase were selected based on the microalgae cell wall composition (cellulose, hemicellulose, pectin and glycoprotein). All of them increased organic matter solubilisation, obtaining high values already after 6 h of pretreatment with an enzyme dose of 1% for cellulase and the enzyme mix. BMP tests with pretreated microalgae showed a methane yield increase of 8 and 15% for cellulase and the enzyme mix, respectively. Prospective research should evaluate enzymatic pretreatments in continuous anaerobic reactors so as to estimate the energy balance and economic cost of the process.Peer Reviewe

Insights into removal of antibiotics by selected microalgae (Chlamydomonas reinhardtii, Chlorella sorokiniana, Dunaliella tertiolecta and Pseudokirchneriella subcapitata)

The increasing contamination of worldwide water with numerous antibiotics has become an emerging environmental concern due to the considerable ecotoxicity and health issues associated with some antibiotics. Microalgae-mediated remediation of pollutants is of growing scientific interest due to the ability of microalgae to take up organic/inorganic nutrients and produce biomass that can be further used for energy or biofuel generation. Elimination of pollutants from water in microalgae experiments includes various mechanisms, such as photodegradation, sorption in biomass and biodegradation. The sorption of pollutants by microalgae has been widely reported, but few studies have proven the biodegradation of pollutants by microalgae. In this work, the ability of four selected microalgae (Chlamydomonas reinhardtii, Chlorella sorokiniana, Dunaliella tertiolecta and Pseudokirchneriella subcapitata) to remove 9 antibiotics and the antidepressant venlafaxine was studied under different experimental conditions. The results show that the highest efficiency for antibiotic removal was obtained with microalgae exhibiting the highest growth rates. Promising results have been obtained from C. reinhardtii and D. tertiolecta cultures, which exhibited a rapid growth and good removal percentages for some of the antibiotics studied. Photodegradation was the main removal mechanism for the 3 fluoroquinolones studied (ciprofloxacin, ofloxacin, norfloxacin) and pipemidic acid (>78%), while for the 3 macrolides (azithromycin, clarithromycin, erythromycin), the efficiency of photodegradation was less than 5%. For the macrolides, the combination of abiotic phenomena, sorption onto the biomass and biodegradation were the mechanisms responsible for total removal. Trimethoprim and venlafaxine were shown to be very stable compounds with low removal percentages. Sulfapyridine appeared to be eliminated mainly by algae biodegradation since low sorption and photodegradation efficiencies were observed. The presence of products resulting from transformations of azithromycin, erythromycin and sulfapyridine in the liquid phase of live algae confirmed that biodegradation is one way by which microalgae eliminate these antibiotics.This work has been funded by the Spanish Ministry of Economy and Competitiveness (project CTQ2013-48545-C2) and supported by the Generalitat de Catalunya (Consolidated Research Groups 2017-SGR-0014, 2017-SGR-1404 and 2017 SGR 1124) and co-financed by the European Union through the European Regional Development Fund. Andrea Hom-Díaz acknowledges the predoctoral grant from AGAUR (2013FI_B 00302). Adrián Jaén-Gil acknowledges the predoctoral grant from AGAUR (2019FI_B2 00202). Sara Rodriguez-Mozaz acknowledges the Ramon y Cajal program (RYC-2014-16707).Peer reviewe

Enzymatic pretreatment of microalgae using fungal broth from Trametes versicolor and commercial laccase for improved biogas production

Es un article del grup de recerca BioremUAB (2014SGR476)Coupling microalgae production to wastewater treatment can reduce the costs of microalgae production for non-food bioproducts and energy consumption for wastewater treatment. Furthermore, microalgae anaerobic digestion can be enhanced by applying pretreatment techniques.. The aim of this study is to improve the biogas production from microalgal biomass grown in urban wastewater treatment systems by applying an enzymatic pretreatment with crude fungal broth and commercial laccase. To this end, the fungus Trametes versicolor was cultured, and the enzymatic activity of the culture broth analysed by measuring laccase concentration. The results showed that both the fungal broth and commercial laccase pretreatment (100 U/L) over an exposure time of 20 min increased the methane yield in batch tests. Indeed, the fungal broth pretreatment increased the methane yield by 74%, while commercial laccase increased the methane yield by 20% as compared to non-pretreated microalgal biomass. In this manner, laccase addition enhanced microalgal biomass anaerobic biodegradability, and addition of T. versicolor broth further improved the results. This fact may be attributed to the presence of other molecules excreted by the fungus