Development of a Process for an Efficient Exploitation of CO2

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Development of a process for an efficient exploitation of CO2 captured from flue gases as liquid carbonates for Chlorella protothecoides cultivation Barbara Gris, Eleonora Sforza, Luca Vecchiato, and Alberto Bertucco Ind. Eng. Chem. Res., Just Accepted Manuscript • DOI: 10.1021/ie502336d • Publication Date (Web): 03 Oct 2014 Downloaded from http://pubs.acs.org on October 15, 2014

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Industrial & Engineering Chemistry Research

Development of a process for an efficient exploitation of CO2 captured from flue gases as liquid carbonates for Chlorella protothecoides cultivation. Barbara Grisa, Eleonora Sforzaa, Luca Vecchiatob, Alberto Bertuccoa

a

Department of Industrial Engineering DII, University of Padova, Via Marzolo 9, 35131 Padova, Italy

b

Eco Management S.r.l. Via Emilia Monselice, Padova, Italy

Original Manuscript Submitted to I&EC research

June 2014 Revised August 2014 Revised October 2014

*

Corresponding author: Eleonora Sforza

e-mail: [email protected] Tel.: +39-0498275462; fax: +39-0498275461.

ACS Paragon Plus Environment


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ABSTRACT The use of bicarbonate as carbon supply in a large scale photobioreactor for industrial microalgae production could reduce the compression and transportation costs associated with the direct use of CO2 from flue gas. Some microalgae species are able to directly exploit bicarbonate as a carbon source, but the actual yield in view of large scale production is not clear, in particular for eukaryotic and lipid rich strains. In this work, the exploitation of bicarbonate as carbon source by Chlorella protothecoides was critically assessed. Both batch and continuous experiments were carried out with addition of NaHCO3 to the culture medium. The growth of this species was found strongly affected by the increased pH, resulting in a reduced yield if compared with air-CO2 bubbling supply. To overcome this issue, a pH control system was applied, so that the pH value was maintained below 8. In these conditions a specific growth rate was measured close to that obtained under CO2 bubbling, probably due to the equilibrium of carbonate, which is shifted to H2CO3 and free CO2. Thus, it appears that C. protothecoides is not able to directly use bicarbonate, but exploits the CO2 made available from the shifting of carbonate equilibrium. In addition, the presence of bicarbonate caused also a lipid accumulation in C. protothecoides, up to about 40% DW, compared to the constitutive value of 10-12% DW. Based on the results obtained, a process scheme was proposed, integrating microalgae cultivation with CO2 capture by absorption with a re-circulating carbonate solution. Process simulation was carried out to show the technical feasibility of this process. KEYWORDS pH control, bicarbonate, CO2 bubbling, carbonate equilibrium HIGHLIGHTS 

The increased pH due to the addition of bicarbonate to the culture medium strongly inhibits algal growth.

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The pH control is necessary to efficiently exploit bicarbonate.

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Experimental measurements suggested that C. protothecoides prefer to utilize CO2 as carbon source.

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The presence of bicarbonate stimulates a lipid accumulation in C. protothecoides.

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A new process concept is proposed to effectively exploit CO2 from flue gas at large scale.

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1. Introduction The global energy crisis has currently promoted the research to develop and optimize technology in biofuels as a sustainable alternative for fuel supply. In this regard, biodiesel from microalgae has shown a great potential that could impact the world's supply of transport fuel in the medium term. However, moving from laboratory to large scale, microalgal cultivation requires careful design and planning, and a full scale commercial application appears challenging for the time being1. Significant obstacles still need to be overcome before microalgae–based biofuel production becomes cost-effective2. In particular, a large scale microalgae cultivation is associated with high demands of nutrients, including carbon, nitrogen and phosphorus. Almost all pilot scale algal cultures depend on purchased carbon dioxide that contributes substantially (about 50%) to the cost of producing the biomass3. Algal culture for fuels is not feasible unless carbon dioxide and other nutrients are available for free4. Therefore, a multipurpose approach must be applied for large scale cultivation, for instance exploiting N and P from wastewaters, and CO2 from flue gas, which turns out to be also a promising carbon capture technique5–7. However, due to the very large surface area required for extensive microalgae production, CO2 has to be compressed and transported to algae photobioreactors (PBRs) some distance away. More energy is consumed for pumping air to provide sufficient mixing and optimum gas–liquid mass transfer8. These operations are energy intensive and enhance the unacceptability of production costs in a large scale PBR. Moreover, the transport of CO2 from gas to liquid is inefficient, requiring very high air flow rate which results in remarkable energy consumption when sufficient carbon source has to be ensured to microalgae during photosynthesis9. To address these challenges, a process where CO2 is captured as bicarbonate and used as feedstock for algae culture would significantly reduce the costs because the transport of the aqueous bicarbonate solution requires much less energy10. A number of studies have reported that bicarbonate ion (

) can serve as an alternative carbon

source to grow microalgae. In fact, several unicellular algae possess an inducible CO2-concentrating mechanism (CCM) which is able to elevate CO2 levels around the CO2-fixing enzyme ribulose-1,5-bisphospate carboxylase/oxygenase (Rubisco). This CCM is highly expressed under low CO2 concentrations (e.g. atmospheric concentration of CO2) compared with cells cultivated under high-CO2 conditions (e.g. 2–5% CO2)11. Generally, does not diffuse across the cell membrane, but the presence at outer membrane level of carbonic anhydrase (CA) enzyme (which is responsible for the conversion of carbonate to free CO2) facilitates CO2 assimilation12. Alternatively, an active transportation on the cell surface allows bicarbonate assimilation by the microalgal cell2. The direct uptake of bicarbonate by an active transport system has been found in several species12–14. Compared with cyanobacteria, less information is available on inorganic carbon (Ci) transport systems in eukaryotic algae10. However, it has been reported that, in addition to Ci transporters in the cell membrane, eukaryotic algae, such as Chlamydomonas reinhartdii, also have chloroplast Ci transporters, because photosynthesis in eukaryote microalgae occurs in the chloroplast10. Thus, some microalgae can directly fix carbon in the form of soluble carbonates for cell growth 15. On the other hand, according to the equilibrium relationship: (1) 2



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protons are consumed during the conversion of bicarbonate to

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.

Thus, the use of bicarbonate as the original carbon source for photosynthesis leaves OH– in the cell, which has to be neutralized by H+ uptake from the extracellular environment, unavoidably leading to an increased pH, with subsequent changes of the equilibrium between different Ci species and inhibitions of cell growth. The problem of achieving pH control in microalgal cultures has been addressed, among others, by Scherholz and Curtis16, who showed how an improved understanding of nutrient utilization can facilitate pH control during growth on excess CO2. Several authors6,10,17,18 recently proposed a new methodology to use efficiently flue gases as CO2 source in the production of photosynthetic microorganisms, where the carbon dioxide is absorbed in an aqueous phase that is then regenerated by microalgae. All of them, however, carried out experiments of microalgal growth with cyanobacteria, which are able to directly utilize bicarbonate19 but cannot accumulate significant amounts of lipids. In this context, literature data reported an increased lipid accumulation due to the addition of sodium bicarbonate in Chlorophytes20. Gardner et al.21 showed that, by manipulating the inorganic carbon concentration supply, it is possible to induce lipid and starch accumulation during nitrogen depletion in C. reinhardtii. However, to obtain a considerable biomass productivity, higher concentrations of CO2 are needed21. Thus, if the aim of microalgal cultivation is the production of liquid fuel, the feasibility of using bicarbonate as carbon source for eukaryotic algae has to be assessed, by critically evaluating the productivity actually achievable. Even though several studies reported growth curves of green algae in the presence of bicarbonates, the productivity is generally quite low, in particular if compared to that under CO2-air bubbling22,23. In addition, to our knowledge, only batch experiments have been carried out with eukaryotic algae and bicarbonate solutions in the open literature so far. In this work, the feasibility of using bicarbonate as carbon source for Chlorella protothecoides cultivation was tested, both in batch and continuous experiments, by comparing the productivity with that obtainable under CO 2 bubbling. Algal growth and carbonate ions consumption in the medium were measured, by taking into account the effects of pH, and eventually controlling its value, the cation associated to the bicarbonate anion and the presence of a simultaneous air bubbling. In addition, the effect of bicarbonate on lipid content in C. protothecoides was evaluated. Finally, based on experimental results, a simplified large scale process block flow diagram is proposed which is simultaneously an alternative for microalgae cultivation plant and a way to capture CO2 from flue gases and natural sources. Process simulation was performed by Aspen Plus® following this scheme. 2. Materials and Methods 2.1 Algal species and culture media Chlorella protothecoides 33.80 (from SAG Goettingen, Germany), was maintained in liquid BG11 medium24 for the inoculum. Experiments with bicarbonate were carried out in modified BG11, by adding different concentrations of NaHCO3 or KHCO3 (Sigma-Aldrich), as reported in the corresponding sections of results. 2.2

Experimental set-up and apparatus

3


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The experiments were performed in both batch and continuous processes. The temperature was controlled at 23°C in an incubator (Frigomeccanica Andreaus, Padova), and artificial light (white OSRAM neon lamps) was provided continuously at an intensity of 100 µE m-2 s-1 of PAR (Photosynthetic Active Radiation), measured by a photoradiometer (Model LI-189, LI-COR, USA). Batch experiments were performed in 250-mL-working-volume glass vertical tubes, with a 5-cm diameter, continuously mixed by a stirring magnet placed at the bottom of the bottle. A first series of runs was carried out without gas supply, by adding bicarbonate as the only carbon source. As a control, batch experiments were carried out in BG11 without inorganic carbon addition. A second series of experiments was carried out with air bubbling with a total gas flow rate of 1 L h-1. All batch experiments were compared to a CO2 air bubbling air (5% v/v of CO2, gas flow rate was 1 L h-1). The preinocula were also grown in this culture conditions. Each batch experiment started with an initial microalgae inoculation of OD750 = 0.5, corresponding to a cell concentration of about 15x106 cells mL-1. Continuous flow experiments were carried out in vertical glass tubes of 250 mL working volume with a 5-cm diameter. At the beginning C. protothecoides was inoculated into the reactor at significant concentration (order of 108 cells mL-1) and fresh medium was fed by a peristaltic pump (Sci-Q 400, Watson Marlow,USA). Part of reactor volume was pumped out at the same inlet flow-rate from the top, in order to ensure a constant reactor volume. Thus, the residence time results: (2) where

and

are the volume of the PBR and the flow rate respectively. It was directly controlled by the

peristaltic pump. After a transient period, the outlet biomass concentration was found stable and a steady state was reached. At steady state, the biomass productivity

was calculated as: (3)

where Cx is the average biomass concentration of at least 5 experimental measures in different days. Continuous experiments were carried out both with CO2 5%-air bubbling gas, as a control, and with the supply of bicarbonate solution, in the absence of bubbling. In the latter case, mixing was provided by magnetic bars. In the experiments with pH control, the pH of the culture was controlled continuously by a pH sensor (D.O. Apparecchiature Elettroniche) connected to a chromatographic pump (LC-20AT Prominence), which was activated when the pH in the reactor exceeded the set point, by using an HCl solution to adjust the pH value. The HCl concentration was chosen to avoid substantial volume changes in the reactor, due to the control. 2.3 Analytical methods The growth in both batch and continuous experiments was monitored daily by a spectrophotometric analysis of the optical density (measured at 750 nm by an UV-Visible UV 500 double beam spectrophotometer from Spectronic Unicam, UK) correlated to cell concentration, measured by a Bürker Counting Chamber (HBG, Germany). Specific growth rates in batch experiments were measured by linear regression of 6-10 experimental points of logarithmic phase of growth, from two or three independent biological replicates. At the end of the growth curve, the final biomass concentration of each experiment was measured as dry weight (DW) in terms of g L-1. DW was measured gravimetrically in cells previously harvested with a 0.22 µm filter: the filters were dried 4



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for 4 hr at 100 °C in a laboratory oven. In continuous experiments, biomass concentration in terms of cell mL-1 and DW were determined daily, and the steady state concentration was averaged on 5 to 10 experimental points, sampled once a day. The carbonate species concentrations were measured daily by using a common titration method, consisting in the use of 2 different dyes, phenolphthalein and bromcresol green (Sigma Aldrich), pHsensitive, allowing to determinate

-

and

concentrations in the solution.

Total lipids of biomass outlet at steady state were extracted overnight from dried cells by using chloroform:methanol (1:2 vol/vol) in accordance with the Bligh&Dyer method 25, in a Soxhlet apparatus. The lipid mass was measured gravimetrically after solvent removal by using rotary evaporator. 2.4 Statistical Analysis T-student tests were applied to ascertain significant differences in biomass concentration in terms of g L -1, cellular weights at different pH in the continuous experiments, as average values of steady state measures (5 to 10 experimental points). The level of statistical significance was assumed for P

Development of a Process for an Efficient Exploitation of CO2

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