Valorization of Spent Escherichia coli Media Using Green Microalgae Chlamydomonas reinhardtii and Feedstock Production.
ABSTRACT: The coupling of Chlamydomonas reinhardtii biomass production for nutrients removal of Escherichia coli anaerobic broth (EAB) is thought to be an economically feasible option for the cultivation of microalgae. The feasibility of growing microalgae in using EAB high in nutrients for the production of more biomass was examined. EAB comprised of nutrient-abundant e?uents, which can be used to produce microalgae biomass and remove environment pollutant simultaneously. In this study, C. reinhardtii 21gr (cc1690) was cultivated in different diluted E. coli anaerobic broth supplemented with trace elements under mixotrophic and heterotrophic conditions. The results showed that C. reinhardtii grown in 1×, 1/2×, 1/5× and 1/10×E. coli anaerobic broth under mixotrophic conditions exhibited specific growth rates of 2.71, 2.68, 1.45, and 1.13 day-1, and biomass production of 201.9, 184.2, 175.5, and 163.8 mg L-1, respectively. Under heterotrophic conditions, the specific growth rates were 1.80, 1.86, 1.75, and 1.02 day-1, and biomass production were 45.6, 29.4, 15.8, and 12.1 mg L-1, respectively. The removal efficiency of chemical oxygen demand, total-nitrogen and total-phosphorus from 1×E. coli anaerobic broth was 21.51, 22.41, and 15.53%. Moreover, the dry biomass had relatively high carbohydrate (44.3%) and lipid content (18.7%). Therefore, this study provides an environmentally sustainable as well economical method for biomass production in promising model microalgae and subsequently paves the way for industrial use.
Project description:In this work, we studied a novel algae cultivation strategy, mixotrophic microalgae biofilm, to improve the productivity and cost-efficiency of algal biofuel production. In contrast to previous methods, this improved approach can achieve high productivity at low cost by harnessing the benefits of mixotrophic growth's high efficiency, i.e., capable of subsisting on inorganic and organic carbons thus unaffected by limited light, and microalgae biofilm's low harvesting cost. Our results, as one of the first studies of this type, proved that microalgae biofilms under mixotrophic condition exhibited significantly higher productivity and quality of biofuel feedstock: 2-3 times higher of biomass yield, 2-10 times higher of lipid accumulation, and 40-60% lower of ash content when compared to microalgae biofilms under autotrophic condition. In addition, we investigated the impact of cell-surface properties (hydrophobicity and roughness) on the growth activities of microalgae biofilms and found that the productivity of mixotrophic biofilms was significantly correlated with the surface hydrophobicity. Finally, our work demonstrated the applicability of integrating this novel cultivation method with wastewater for maximum efficiency. This study opens a new possibility to solve the long-lasting challenges of algal biofuel feedstock production, i.e., low productivity and high cost of algal cultivation.
Project description:Background:Microalgae are an important feedstock in industries. Currently, efforts are being made in the non-phototrophic cultivation of microalgae for biomass production. Studies have shown that mixotrophy is a more efficient process for producing algal biomass in comparison to phototrophic and heterotrophic cultures. However, cultivation of microalgae in pilot-scale open ponds in the presence of organic carbon substrates has not yet been developed. The problems are heterotrophic bacterial contamination and inefficient conversion of organic carbon. Results:Laboratory investigation was combined with outdoor cultivation to find a culture condition that favors the growth of alga, but inhibits bacteria. A window period for mixotrophic cultivation of the alga Graesiella sp. WBG-1 was identified. Using this period, a new sequential phototrophic-mixotrophic cultivation (SPMC) method that enhances algal biomass productivity and limits bacteria contamination at the same time was established for microalgae cultivation in open raceway ponds. Graesiella sp. WBG-1 maximally produced 12.5 g biomass and 4.1 g lipids m-2 day-1 in SPMC in a 1000 m2 raceway pond, which was an over 50% increase compared to phototrophic cultivation. The bacterial number in SPMC (2.97 × 105 CFU ml-1) is comparable to that of the phototrophic cultivations. Conclusions:SPMC is an effective and feasible method to cultivate lipid-rich microalgae in open raceway ponds. Successful scale-up of SPMC in a commercial raceway pond (1000 m2 culture area) was demonstrated for the first time. This method is attractive for global producers of not only lipid-rich microalgae biomass, but also astaxanthin and β-carotene.
Project description:Microalgae are promising microorganisms for the production of numerous molecules of interest, such as pigments, proteins or triglycerides that can be turned into biofuels. Heterotrophic or mixotrophic growth on fermentative wastes represents an interesting approach to achieving higher biomass concentrations, while reducing cost and improving the environmental footprint. Fermentative wastes generally consist of a blend of diverse molecules and it is thus crucial to understand microalgal metabolism in such conditions, where switching between substrates might occur. Metabolic modeling has proven to be an efficient tool for understanding metabolism and guiding the optimization of biomass or target molecule production. Here, we focused on the metabolism of Chlorella sorokiniana growing heterotrophically and mixotrophically on acetate and butyrate. The metabolism was represented by 172 metabolic reactions. The DRUM modeling framework with a mildly relaxed quasi-steady-state assumption was used to account for the switching between substrates and the presence of light. Nine experiments were used to calibrate the model and nine experiments for the validation. The model efficiently predicted the experimental data, including the transient behavior during heterotrophic, autotrophic, mixotrophic and diauxic growth. It shows that an accurate model of metabolism can now be constructed, even in dynamic conditions, with the presence of several carbon substrates. It also opens new perspectives for the heterotrophic and mixotrophic use of microalgae, especially for biofuel production from wastes.
Project description:Microalgae are fast-growing photosynthetic organisms which have the potential to be exploited as an alternative source of liquid fuels to meet growing global energy demand. The cultivation of microalgae, however, still needs to be improved in order to reduce the cost of the biomass produced. Among the major costs encountered for algal cultivation are the costs for nutrients such as CO?, nitrogen and phosphorous. In this work, therefore, different microalgal strains were cultivated using as nutrient sources three different anaerobic digestates deriving from municipal wastewater, sewage sludge or agro-waste treatment plants. In particular, anaerobic digestates deriving from agro-waste or sewage sludge treatment induced a more than 300% increase in lipid production per volume in <i>Chlorella vulgaris</i> cultures grown in a closed photobioreactor, and a strong increase in carotenoid accumulation in different microalgae species. Conversely, a digestate originating from a pilot scale anaerobic upflow sludge blanket (UASB) was used to increase biomass production when added to an artificial nutrient-supplemented medium. The results herein demonstrate the possibility of improving biomass accumulation or lipid production using different anaerobic digestates.
Project description:Background:The yield of microalgae biomass is the key to affect the accumulation of fatty acids. A few microalgae can assimilate organic carbon to improve biomass yield. In mixotrophic cultivation, microalgae can use organic carbon source and light energy simultaneously. The preference of the main energy source by microalgae determines the biomass yield. Auxenochlorella protothecoides is an oleaginous mixotrophic microalga that can efficiently assimilate glucose and accumulate a large amount of biomass and fatty acids. The current study focused on the effect of light on the growth and glucose assimilation of A. protothecoides. Results:In this study, we found that the uptake and metabolism of glucose in A. protothecoides could be inhibited by light, resulting in a reduction of biomass growth and lipid accumulation. We employed comparative proteomics to study the influence of light on the regulation of glucose assimilation in A. protothecoides. Proteomics revealed that proteins involving in gene translation and photosynthesis system were up-regulated in the light, such as ribulose-phosphate 3-epimerase and phosphoribulokinase. Calvin cycle-related proteins were also up-regulated, suggesting that light may inhibit glucose metabolism by enhancing the production of glyceraldehyde-3-phosphate (G3P) in the Calvin cycle. In addition, the redox homeostasis-related proteins such as thioredoxin reductase were up-regulated in the light, indicating that light may regulate glucose uptake by changing the redox balance. Moreover, the increase of NADH levels and redox potential of the medium under illumination might inhibit the activity of the glucose transport system and subsequently reduce glucose uptake. Conclusions:A theoretical model of how glucose assimilation in A. protothecoides is negatively influenced by light was proposed, which will facilitate further studies on the complex mechanisms underlying the transition from autotrophy to heterotrophy for improving biomass accumulation.
Project description:The stressed cultivations are widely used in microalgae R&D for the biofuel production with the repress on growth to a certain degree, which limits the overall productivity. The balance between the growth and energy storage compounds accumulation is a target needing the combination of both strain selection or construction and culture optimization. Here, an engineered strain of <i>Chlamydomonas reinhardtii</i>, in which the chloroplast type glyceraldehyde-3-phosphate dehydrogenase (cGAPDH) was overexpressed and named as P3-GAPDH, was cultured on the Algal Station platform. Compared with wild type (WT), <i>C. reinhardtii</i> CC137c, in Tris-acetate-phosphate (TAP) medium, the highest density of WT and P3-GAPDH were 1.23 ± 0.13 and 1.74 ± 0.09 g L<sup>-1</sup> within 96 h, and the maximum biomass productivity was 24.30 ± 1.65 and 28.54 ± 1.43 mg L<sup>-1</sup> h<sup>-1</sup>, respectively. In terms of the energy storage compounds, both carbohydrate and fatty acids content doubled in P3-GAPDH, from 0.13 ± 0.02 to 0.26 ± 0.04 g L<sup>-1</sup> for carbohydrate and from 0.08 ± 0.01 to 0.16 ± 0.01 g L<sup>-1</sup> for fatty acids, among which poly unsaturated fatty acids increased by 65.8%. Together with the continuous monitor of the chlorophyll fluorescence dynamics parameters <i>F</i> <sub>v</sub>/<i>F</i> <sub>m</sub> and <i>F</i> <sub>v</sub>'/<i>F</i> <sub>m</sub>' and pH of culture, enhanced Calvin cycle by overexpressed cGAPDH promoted the carbon conversion and subsequent energy storage compounds accumulation. <i>C. reinhardtii</i> P3-GAPDH strain showed the potential as a good chassis with high carbon conversion ability.
Project description:Microalgae are a promising biomass feedstock for biofuels production. The use of wastewater effluent as a nutrient medium would improve the economics of microalgal biofuels production. Bacterial communities in aquatic environments may either stimulate or inhibit microalgal growth. Microalgal productivity could be enhanced if the positive effects of indigenous bacteria could be exploited. However, much is unknown about the effects of indigenous bacteria on microalgal growth and the characteristics of bacterial communities associated with microalgae in microalgae-effluent culture. To assess the effects of the indigenous bacteria in wastewater effluent on microalgal growth, three microalgae, Chlamydomonas reinhardtii, Chlorella vulgaris, and Euglena gracilis, were cultured in two municipal wastewater effluents and one swine wastewater effluent with and without indigenous bacteria for 7 days.All microalgae grew better in all effluents with indigenous bacteria than without bacteria. Biomass production of C. reinhardtii, C. vulgaris, and E. gracilis increased?>?1.5, 1.8-2.8, and >?2.1-fold, respectively, compared to the axenic cultures of each microalga. The in situ indigenous bacterial communities in the effluents therefore promoted the growth of the three microalgae during 7-day cultures. Furthermore, the total numbers of bacterial 16S rRNA genes in the 7-day microalgae-effluent cultures were 109?793 times the initial numbers. These results suggest that the three microalgae produced and supplied organic carbon that supported bacterial growth in the effluent. At the phylum and class levels, Proteobacteria (Alphaproteobacteria and Betaproteobacteria) and Bacteroidetes (Sphingobacteriia and Saprospirae) were selectively enriched in all microalgae-effluent cultures. The enriched core bacterial families and genera were functions of the microalgal species and effluents. These results suggest that certain members of the bacterial community promote the growth of their "host" microalgal species.To enhance their own growth, microalgae may be able to selectively stimulate specific bacterial groups from among the in situ indigenous bacterial community found in wastewater effluent (i.e., microalgae growth-promoting bacteria: MGPB). The MGPB from effluent cultures could be used as "probiotics" to enhance microalgal growth in effluent culture. Wastewater effluent may therefore be a valuable resource, not only of nutrients, but also of MGPB to enable more efficient microalgal biomass production.
Project description:Microalgae are gaining commercial interests in the areas food, feed and biofuel sector. They have intrinsic ability to harness energy from sunlight and photosynthetically valorize CO2 into various bio-based products viz., triacylglycerols (TAGs), mono/poly-unsaturated fatty acids (MUFA, PUFA), pigments etc. Microalgae have adapted to grow in various nutritional environments due to their metabolic versatility and resilience. Strategic evaluation of newly isolated strain Chlorella sp. from a residential lake was performed. The strain was investigated by varying the nutritional modes to gain insights into biomass and fatty acids production. Maximum biomass (3.59 g/L) was observed in mixotrophic condition followed by heterotrophic (1.58 g/L) and autotrophic condition (0.59 g/L). The maximum lipid yield (670 mg/g DCW) was observed in mixotrophic condition whereas maximum total lipid content (36%) was observed in heterotrophic condition. Significant correlation was noticed between fluorescence parameters measured by OJIP and non-photochemical quenching (NPQ) with the function of nutritional mode variations. Autotrophic condition showed higher photosynthetic activity which was well correlated with high fluorescence intensity as represented by OJIP, NPQ1, and NPQ2 curves. Good balance of saturated fatty acids (SFA) and unsaturated fatty acids was observed in autotrophic mode, whereas polyunsaturated fatty acids (PUFA) and mono unsaturated fatty acid (MUFA) content were relatively higher in mixotrophic and heterotrophic conditions.
Project description:In this study, we propose a novel sensitive solid-based immunosensor developed on a plasmonic nanopaper platform for the detection of Escherichia coli (E. coli) bacteria. This plasmonic nanopaper that comprises of carboxylated bacterial cellulose (CBC) impregnated with gold nanoparticles (AuNP-CBC), employed as a quencher and a sustainable functionalized platform to be conjugated with protein A. Thus, the conjugated protein A allows the aligned linkage of EAb-QD (anti-E. coli conjugated quantum dot) and EAb-AF (anti-E. coli conjugated Alexa Fluor 488). Interestingly, once E. coli was captured by the AuNP-CBC/EAb-QD or AuNP-CBC/EAb-AF, the energy transfer from the QD or Alexa Fluor fluorophores is triggered due to the conformational change in the antibody structure and this, in turn, causes a decrease in the distance between fluorophores and the quencher nanopaper and, therefore diminishing their photoluminescence. The immunosensors performed successfully to recognize E. coli at concentrations as low as 50 CFU mL<sup>-1</sup> in the standard buffer. The examined functionality of the immunosensors in a real matrix such as chicken extract and lettuce juice demonstrated a highly efficient response while QD is the main fluorophore with a limit of detection around 100 CFU mL<sup>-1</sup>.
Project description:<h4>Background</h4>The production of microalgal biofuels, despite their sustainable and renowned potential, is not yet cost-effective compared to current conventional fuel technologies. However, the biorefinery concept increases the prospects of microalgal biomass as an economically viable feedstock suitable for the co-production of multiple biofuels along with value-added chemicals. To integrate biofuels production within the framework of a microalgae biorefinery, it is not only necessary to exploit multi-product platforms, but also to identify optimal microalgal cultivation strategies maximising the microalgal metabolites from which biofuels are obtained: starch and lipids. Whilst nutrient limitation is widely known for increasing starch and lipid formation, this cultivation strategy can greatly reduce microalgal growth. This work presents an optimisation framework combining predictive modelling and experimental methodologies to effectively simulate and predict microalgal growth dynamics and identify optimal cultivation strategies.<h4>Results</h4>Microalgal cultivation strategies for maximised starch and lipid formation were successfully established by developing a multi-parametric kinetic model suitable for the prediction of mixotrophic microalgal growth dynamics co-limited by nitrogen and phosphorus. The model's high predictive capacity was experimentally validated against various datasets obtained from laboratory-scale cultures of Chlamydomonas reinhardtii CCAP 11/32C subject to different initial nutrient regimes. The identified model-based optimal cultivation strategies were further validated experimentally and yielded significant increases in starch (+?270%) and lipid (+?74%) production against a non-optimised strategy.<h4>Conclusions</h4>The optimised microalgal cultivation scenarios for maximised starch and lipids, as identified by the kinetic model presented here, highlight the benefits of exploiting modelling frameworks as optimisation tools that facilitate the development and commercialisation of microalgae-to-fuel technologies.