Simultaneous biogas upgrading and centrate treatment in an outdoors pilot scale high rate algal pond

1 Simultaneous biogas upgrading and centrate treatment in an outdoors 1 pilot scale high rate algal pond 2 Esther Posadas1, David Marín1,3, Saúl Blanco2, Raquel Lebrero1, Raúl Muñoz*1 3 1 Department of Chemical Engineering and Environmental Technology, Valladolid University, Dr. 4 Mergelina, s/n, 47011, Valladolid, Spain. 5 2 Department of Biodiversity and Environmental Management, University of León, 24071 León, Spain. 6 3 Universidad Pedagógica Nacional Francisco Morazán, Boulevard Centroamérica, Tegucigalpa, 7 Honduras. 8 9 * Corresponding author: [email protected] 10 11 ABSTRACT 12 The bioconversion of biogas to biomethane coupled to centrate treatment was evaluated 13 in an outdoors pilot scale high rate algal pond interconnected to an external CO2-H2S 14 absorption column (AC) via settled broth recirculation. CO2-removal efficiencies ranged 15 from 50 to 95% depending on the alkalinity of the cultivation broth and environmental 16 conditions, while a complete H2S removal was achieved regardless of the operational 17 conditions. A maximum CH4 concentration of 94% with a limited O2 and N2 stripping 18 was recorded in the upgraded biogas at recycling liquid/biogas ratios in the AC of 1 and 19 2. Process operation at a constant biomass productivity of 15 g m-2 d-1 and the 20 minimization of effluent generation supported high carbon and nutrient recoveries in the 21 harvested biomass (C = 66±8%, N= 54±18%, P≈100% and S =16±3%). Finally, a low 22 diversity in the structure of the microalgae population was promoted by the 23 environmental and operational conditions imposed. 24 Keywords: algal-bacterial symbiosis, biogas upgrading, biomethane, microalgae, 25 outdoors conditions, wastewater treatment. 26 *Manuscript Click here to view linked References 2 1. Introduction 27 Biogas from the anaerobic digestion of organic solid waste and wastewater represents a 28 renewable energy source with a significant potential to reduce the current world´s fossil 29 fuel dependence (Hermann et al., 2016). Biogas can be used as a fuel for the on-site 30 generation of domestic heat or steam and electricity in industry, as a substrate in fuel 31 cells or as a substitute of natural gas prior upgrading (Andriani et al., 2014; Muñoz et 32 al., 2015). For instance, the use of this biofuel in the European Union during 2014 33 supported a production of electricity and heat of 63.4 and 32.2 TWh, respectively (EBA, 34 2016). Biogas conversion to biomethane is highly recommended due to the high 35 concentration of impurities present in the raw biogas: CO2 (25-60%), CO (<0.6%), H2S 36 (0.005-2%), N2 (0-2%), NH3 (<1%), H2O (5-10%), O2 (0-1%), siloxanes (0-0.02%) and 37 halogenated hydrocarbons (VOC <0.6%) (Ryckebosch et al., 2011). In fact, biogas 38 upgrading is a mandatory step required prior biomethane injection into natural gas grids 39 or use as a vehicle fuel, which must provide concentrations of CH4 ≥95%, CO2 ≤2%, 40 O2≤0.3% and negligible amounts of H2S according to most international regulations 41 (Muñoz et al., 2015). In this context, the removal of CO2 from raw biogas would 42 contribute to reduce the transportation costs and to increase the calorific value of 43 biomethane, while the removal of H2S would limit the corrosion in pipelines, boilers, 44 engines, etc. (Posadas et al., 2015a). 45 Several physical-chemical and biological technologies are nowadays available at 46 commercial scale to remove CO2 and H2S from biogas. Pressure swing adsorption, 47 amine/water/organic scrubbing or membrane separation are typically applied to remove 48 CO2, while activated carbon filtration, chemical precipitation or anoxic/aerobic 49 biotrickling filtration provide satisfactory levels of H2S removal (Mann et al., 2016; 50 Toledo-Cervantes et al., 2016; Muñoz et al., 2015). However, these H2S and CO2 51 3 removal technologies must be sequentially implemented to remove both biogas 52 contaminants, which makes physical-chemical biogas upgrading a costly and complex 53 two-stage process (Muñoz et al., 2015). The few technologies supporting a 54 simultaneous removal of CO2 and H2S from low S-strength biogas (i.e. chemical 55 scrubbing) exhibit high environmental impacts and operating costs (Tippayawong and 56 Thanompongchart, 2010). In this context, algal-bacterial photobioreactors have recently 57 emerged as an environmentally friendly and cost-efficient alternative to remove CO2 58 and H2S from raw biogas in a single step process (Bahr et al., 2014; Yan et al., 2016). 59 Photosynthetic biogas upgrading in algal-bacterial photobioreactors is based on the 60 simultaneous fixation of CO2 by microalgae and oxidation of H2S to SO42- by sulfur 61 oxidizing bacteria or chemical reactions, the latter supported by the high dissolved 62 oxygen (DO) concentrations present in the cultivation broth (Posadas et al., 2015a; 63 Toledo-Cervantes et al., 2016). The economic and environmental sustainability of this 64 process can be boosted via integration of biogas upgrading with the recovery of 65 nutrients from digestate in the form of a valuable algal-bacterial biomass (Serejo et al., 66 2015; Posadas et al., 2015a, 2016; Toledo-Cervantes et al., 2016; Yan et al., 2016). 67 Several investigations aiming at integrating photosynthetic biogas upgrading with 68 digestate treatment have been recently carried out in indoors high rate algal ponds 69 (HRAPs) interconnected to biogas absorption columns (AC) under artificial 70 illumination (Bahr et al. 2014; Alcántara et al., 2015; Posadas et al. 2015a, 2016; Serejo 71 et al. 2015; Meier et al. 2015; Toledo-Cervantes et al. 2016, 2017). Despite the rapid 72 optimization of this technology (Toledo-Cervantes et al., 2016, 2017), the constant 73 temperature (often in the optimum range) and irradiation (often too low compared to 74 solar irradiation) prevailing under laboratory conditions still hinder the complete 75 understanding of a process designed to be ultimately implemented outdoors under solar 76 4 irradiation. Therefore, the evaluation of the performance of photosynthetic biogas 77 upgrading under outdoors conditions is crucial to understand the influence of the diurnal 78 variations of light irradiance and temperature on the quality of the upgraded biogas. 79 Similarly, process operation to minimize the desorption of O2 and N2 from the 80 cultivation broth to the upgraded biogas, and to maximize nutrient recovery from 81 digestates, must be optimized to the particular conditions prevailing during outdoors 82 operation. 83 Despite the remarkable environmental advantages of using digestates as a nutrient 84 source during biogas upgrading, their high nutrients content results in high biomass 85 concentrations in the HRAPs (7-50 g L-1) and the need to operate the process at low 86 digestates flowrates. This severely decreases the photosynthetic efficiency of the system 87 as a result of mutual shading and entails a net consumption of water to compensate 88 evaporation losses (Posadas et al., 2016). In this context, all studies carried out to date 89 set the make-up water input to maintain similar effluent and influent flowrates in order 90 to guarantee a constant biomass output, which resulted in the generation of effluents 91 with residual nutrient concentrations (Toledo-Cervantes et al., 2016; Posadas et al., 92 2016). On this basis, there is an urgent need to develop novel photobioreactor designs 93 and operational strategies to minimize effluent generation while maintaining high 94 microalgae productivities using digestates as a nutrient source. 95 This work aimed at evaluating the potential of a novel pilot scale HRAP interconnected 96 to an AC via recirculation of the settled cultivation broth under outdoors conditions 97 during the simultaneous upgrading of biogas and treatment of centrate. Process 98 performance was evaluated under pseudo-steady state conditions at different alkalinity 99 levels and make-up water supply regimes from June to October. Under each operational 100 stage, process performance was also assessed during one diurnal cycle of temperature 101 5 and irradiance. A novel strategy decoupling biomass productivity from the effluent 102 flowrate via control of the biomass wastage from the settler was applied to maximize 103 the recovery of carbon and nutrients from biogas and centrate in the form of harvested 104 biomass. Finally, the influence of the recycling liquid/biogas (L/G) ratio on the 105 efficiency of biogas upgrading was also evaluated during a 24 h diurnal cycle. 106 2. Materials and methods 107 2.1 Biogas and centrate 108 A synthetic biogas mixture, composed of CO2 (29.5%), H2S (0.5%) and CH4 (70%), 109 was used as a model biogas (Abello Linde; Spain). Centrate was obtained from the 110 centrifuges dehydrating the anaerobically digested sludge of Valladolid wastewater 111 treatment plant and stored at 4 ºC prior to use. Centrate composition along the 112 experimental period was subjected to the typical variations of real wastewaters: total 113 organic carbon (TOC) = 70±8 mg L-1, inorganic carbon (IC) = 522±40 mg L-1, total 114 nitrogen (TN) = 580±102 mg L-1, N-NH4+ = 553±67 mg L-, P-PO43- = 34±7 mg L-1 and 115 SO42- = 9±9 mg L-1. 116 2.2 Experimental set-up 117 The pilot plant was located outdoors at the Department of Chemical Engineering and 118 Environmental Technology of Valladolid University (41.39º N, 4.44º W). The 119 experimental set-up consisted of a 180 L HRAP with an illuminated surface of 1.20 m2 120 (length = 170 cm; width = 82 cm; depth =15 cm) and two water channels divided by a 121 central wall and baffles in each side of the curvature. The HRAP was interconnected to 122 an external 2.5 L bubble absorption column (internal diameter = 4.4 cm; height = 165 123 cm) provided with a metallic gas diffuser (2 µm pore size) located at the bottom of the 124 column. The HRAP and AC were interconnected via external liquid recirculation of the 125 supernatant of the algal-bacterial cultivation broth from an 8 L settler located at the 126 6 outlet of the HRAP (Fig. 1). The internal recirculation velocity of the cultivation broth 127 in the HRAP was ≈ 20 cm s-1, which was provided by the continuous rotation of a 6-128 blade paddlewheel. 129 ˂Figure 1> 130 2.3 Operational conditions and sampling procedures 131 Process operation was carried out from June 29th to October the 4th 2016. Based on a 132 previous study conducted by Norvill et al. (2017) in a similar HRAP treating urban 133 wastewater at 4 days of hydraulic retention time (HRT) in the same location, a constant 134 biomass productivity of 15 g m-2 d-1 was set throughout the 92 days of operation. The 135 required C, N and P input to maintain this biomass productivity was 9.7 g C d-1, 1.9 g N 136 d-1 and 0.2 g P d-1, assuming a C, N and P biomass content of 45, 9 and 1%, respectively 137 (Posadas et al., 2015b). This required a centrate flow rate of 3.2 L d-1 (considering an IC 138 and N-NH4+ stripping of 20%, and the absence of P removal by precipitation; Posadas et 139 al. (2013)) and a biogas flow rate of 74.9 L d-1 (assuming an average CO2 removal 140 efficiency in the AC of 80% based on Posadas et al. (2015a)). The recycling 141 liquid/biogas (L/G) ratio in the AC was fixed at 0.5 according to Toledo-Cervantes et al. 142 (2016). The liquid and biogas residence time in the AC under these operational 143 conditions were 96 and 48 min, respectively. The settled biomass in the settler was 144 continuously recirculated to the HRAP at a flow rate of 7.2 L d-1. This, together with the 145 external recycling, resulted in a HRT in the settler of 4.4 h. This process configuration 146 has been shown to increase the settleability of the algal-bacterial biomass, while 147 avoiding biomass degradation in the settler (Valigore et al., 2012; Park et al., 2011, 148 2013). Biomass harvesting was performed by daily removing the required settled 149 biomass volume according to its total suspended solids (TSS) concentration in order to 150 maintain the above mentioned biomass productivity. 151 7 The HRAP was initially filled with tap water (IC = 550 mg L-1) and inoculated to an 152 initial concentration of 210 mg TSS L-1 with Chlorella sp. from a HRAP treating 153 centrate at the Department of Chemical Engineering and Environmental Technology of 154 Valladolid University (Spain). The system was inoculated on June 29th, and after 5 d of 155 inoculum acclimation batchwise, three different operational conditions were tested 156 (corresponding to stages I, II and III) to optimize the simultaneous outdoors biogas 157 upgrading and centrate treatment from a technical and environmental view point (Table 158 1). 159 ˂Table 1> 160 Stage I (reference state) was conducted at a centrate IC concentration of 522 ± 40 mg C 161 L-1. During stages II and III, the IC concentration of the centrate was increased up to 162 2024±124 mg C L-1 by addition of NaHCO3 and Na2CO3, which increased the pH of the 163 centrate from 8.38±0.33 in stage I to 9.94±0.09 and 10.06±0.13 in stages II and III, 164 respectively (Table 1). Tap water was fed to the HRAP in stages I and II to compensate 165 evaporation losses and maintain an effluent flowrate of 0.6±0.4 and 0.8±0.4 L d-1, 166 respectively, thus minimizing the loss of carbon, nutrients and fresh water. The effluent 167 from the system was returned to the HRAP in stage III to minimize the supply of 168 NaHCO3 and Na2CO3, with a subsequent decrease in the supply of make-up water. Each 169 operational stage was maintained for approximately one month, where temperature, 170 solar irradiation and number of sun hours remained approximately constant (Table 1). 171 The results obtained for the liquid phase throughout the three operational stages were 172 provided as average values along with their corresponding standard deviation from 173 measurements recorded for four consecutive days during each steady state. 174 The ambient and cultivation broth temperatures, influent and effluent flowrates, DO and 175 pH in the cultivation broth, and the photosynthetic active irradiation (PAR) were daily 176 8 monitored. Gas samples of 100 µL of the raw and upgraded biogas were drawn twice a 177 week to monitor the concentrations of CO2, H2S, CH4, O2 and N2. The inlet and outlet 178 biogas flowrates in the AC were also measured to accurately determine both CO2 and 179 H2S removals, and CH4 losses by absorption. Liquid samples of 100 mL from the 180 centrate and the treated effluent after settling were withdrawn twice a week to monitor 181 the pH, TSS concentration, and concentrations of dissolved TOC, IC, TN, N-NH4+, N-182 NO2-, N-NO3-, P-PO43- and SO42- following sample filtration through 0.20 µm nylon 183 filters. Likewise, liquid samples of 25 mL were drawn from the cultivation broth and 184 from the bottom of the settler twice a week to monitor the algal-bacterial TSS 185 concentration. The algal-bacterial biomass harvested from the settler under steady state 186 was washed three times with distilled water and dried for 24 hours at 105 ºC to 187 determine its elemental composition (C, N, P and S). Process monitoring and biomass 188 harvesting were always conducted at 9:00 a.m. along the entire experimental period. 189 At the end of each operational stage, the outdoors temperature and PAR, along with the 190 temperature, DO concentration and pH in the HRAP, settler and AC were measured 191 every 30 minutes during one entire diurnal cycle from one hour prior to dawn to one 192 hour after sunset. The composition and flowrate of the upgraded biogas were recorded 193 every hour, and the concentrations of TOC, IC and TN in the HRAP, settler and AC 194 were analyzed every 2 hours. 195 2.4 Influence of the L / G ratio on the quality of the upgraded biogas 196 L/G ratios ranging from 0.5 to 5 were tested at the end of stage III (4th - 7th October) to 197 optimize the quality of the upgraded biogas. A biogas flowrate of 74.9 L d-1 was 198 maintained while the liquid flowrates were set at 37.5, 74.9, 149.8 and 374.5 L d-1 199 (providing L/ G ratios of 0.5, 1, 2 and 5, respectively). Each L/G ratio was maintained 200 for 12 h during one-day diurnal cycle. The ambient temperature and PAR, along with 201 9 the temperature, DO and pH in the HRAP, settler and AC, and the composition and 202 flowrate of the upgraded biogas, were measured every two hours from one hour prior to 203 dawn to one hour after sunset. 204 2.5 Analytical procedures 205 The monthly average ambient temperatures, PARs and number of sun hours were 206 provided by the official AEMET meteorological station located at the University of 207 Valladolid. CO2, H2S, CH4, O2 and N2 gas concentrations were determined using a 208 Varian CP-3800 GC-TCD (Palo Alto, USA) according to Posadas et al. (2015a). 209 Temperature and DO concentration were determined using an OXI 330i oximeter 210 (WTW, Germany). An Eutech Cyberscan pH 510 (Eutech instruments, The 211 Netherlands) was used for pH determination. The PAR was measured with a LI-250A 212 light meter (LI-COR Biosciences, Germany). The concentrations of dissolved TOC, IC 213 and TN were measured using a Shimadzu TOC-VCSH analyzer (Japan) coupled with a 214 TNM-1 chemiluminescence module. N-NH4+ concentration was determined with an 215 ammonium specific electrode Orion Dual Star (Thermo Scientific, The Netherlands). 216 The concentrations of N-NO3-, N-NO2- , P-PO43- and SO42- were quantified by HPLC-IC 217 according to Posadas et al. (2013). All analyses were carried out according to Standard 218 Methods (APHA, 2005). 219 The determination of the C, N and S content of the algal-bacterial biomass was 220 conducted in a LECO CHNS-932 analyzer, while phosphorus content was determined 221 spectrophotometrically after acid digestion in a microwave according to Standard 222 Methods (APHA, 2005). The identification, quantification and biometry measurements 223 of the microalgae assemblage under steady state were performed by microscopic 224 examination (OLYMPUS IX70, USA) of biomass samples (fixed with lugol acid at 5% 225 and stored at 4 ºC prior to analysis) according to Sournia (1978). 226 16 2003), the removals of TOC estimated by mass balance calculations ranged from 376 59±7% (stage III) to 74±7% (stage I) (Table 2) (Fig. A.3). 377 ˂Table 2> 378 The TIC-REs in stage I were higher than those recorded in stages II and III as a result of 379 the higher inorganic carbon feeding and C-CO2 REs in the AC during these latter stages 380 (Table 2). Therefore, only 65±6 and 66±8% of the total carbon removed in stages II and 381 III was recovered in the harvested biomass, while a 97±1% carbon recovery was 382 observed during stage I (Table 3). Despite the higher pH values should have promoted 383 lower IC removals by stripping based on the limited CO2 aqueous equilibrium 384 concentration, the lower IC loading during stage I resulted in a lower fraction of C 385 removed by stripping (Table 3) (Posadas et al., 2013) (Fig. 5b). 386 Similar TN-REs of 86±4, 87±4 and 80±4% were recorded during stages I, II and III, 387 respectively, while a complete N-NH4+ removal occurred during the entire experimental 388 period (Table 2; Fig. 5c, 5d). Nitrification was not inhibited by the high pH values 389 prevailing during stages II and III or the low DO concentrations (<1 mg O2 L-1) present 390 in the first hours in the morning (Fig. A.3). N-NO2- concentrations were low compared 391 to N-NO3- despite temperatures higher than 28ºC were always recorded close to midday, 392 which are known to promote the partial oxidation of N-NH4+ (Fig. 5e; Figs. A.2-A.3) 393 (Metcalf and Eddy, 2003). The oxidation ratios (referred to [N-NO3-+ N-NO2-] mass 394 outputs compared to TN mass input, Posadas et al. (2015a)) were 11±2, 13±4 and 395 19±8% during stages I, II and III, respectively. The high nitrification activity, together 396 with the high evaporation rates, induced an increase in N-NO3- concentration in the 397 cultivation broth up to 148 mg L-1 in stage I, 198 mg L-1 in stage II and 293 mg L-1 in 398 stage III, this latter increase mediated by the absence of effluent from the HRAP (Fig. 399 5f). The nitrogen recovered in the harvested biomass accounted for 65±3, 54±18 and 400 17 76±19% of the total nitrogen removed during stages I, II and III, respectively (Table 3). 401 These values were considerably higher than those recorded by Posadas et al. (2015a) 402 (45±7%) and Toledo-Cervantes et al. (2017) (19±13% and 36±18%) in a similar indoors 403 experimental set-up during the simultaneous treatment of biogas and digestates as a 404 result of the lower microalgae productivities in those studies. 405 ˂Table 3> 406 High P-PO43- REs of 92±2, 84±5 and 85±5% were recorded during stages I, II and III, 407 respectively (Table 2). The higher P-RE in stage I was likely mediated by the higher P 408 content of the harvested biomass (Table 3). In this regard, P-PO43- concentration in the 409 cultivation broth increased up to 6 mg L-1 in stage I, 15 mg L-1 in stage II and 17 mg L-1 410 in stage III. These increasing P-PO43- concentration were also supported by the 411 evaporation rate and the low or negligible effluent flowrates (Fig. 5g). A P mass balance 412 revealed than approximately 100% of the P removed was recovered in the harvested 413 biomass, despite high pH values are known to promote PO43- precipitation (Cai et al., 414 2013) (Table 3). 415 Finally, H2S oxidation supported an increase in SO42- concentration in the cultivation 416 broth of the HRAP from 60 to 495 mg L-1 through the 92 operational days, also 417 triggered by the high evaporation rates and low effluent flowrates (Fig. 5h). The fraction 418 of H2S not fully oxidized to sulphate would have remained as S-intermediates in the 419 liquid phase (Sº, thiosulfate or sulfite) (Toledo-Cervantes et al., 2016). This was 420 confirmed by the observation of Sº accumulation on the walls and diffuser of the AC 421 during stage I (Photograph 1, appendix), while a S mass balance revealed that only 422 26±5, 17±3 and 16±3% of the S removed was recovered in the harvested biomass 423 during stages I, II and III, respectively (Table 3). Further analyses to determine the 424 actual sulfur compounds present in the cultivation broth are required. 425 18 3. 5 Concentration and composition of the algal-bacterial biomass 426 The steady state biomass concentrations in the HRAP during stages I, II and III 427 averaged 660±17, 1078±84 and 665±79 mg TSS L-1 (Fig. A. 14). The operational 428 strategy here evaluated based on the control of biomass productivity via regulation of 429 the settled biomass wastage rate successfully maintained the concentration of algal-430 bacterial biomass below light limiting values. At this point it should be stressed that the 431 theoretical biomass concentration generated based on the centrate composition would be 432 ≈2000 mg TSS L-1 (with P as the limiting nutrient). The good settling characteristics of 433 the algal-bacterial (supporting TSS-REs in the settler of 80±9%) were likely promoted 434 by the short HRT in the settler and the continuous recirculation of the settled biomass, 435 which boosted the enrichment of rapidly settling algal-bacterial flocs (Valligore et al., 436 2011; Park et al., 2011). 437 The elemental composition of the harvested biomass remained within the typical range 438 reported in literature, regardless of the operational stage (Posadas et al., 2016; Bi et al., 439 2013). C, N and P content in the biomass decreased from stage I to stage II and slightly 440 increased in stage III (Table 3). The different C/N/P (g/g/g) ratios present in the 441 cultivation broth of the HRAP (100/39/2, 100/6/1 and 100/12/1 during stages I, II and 442 III, respectively) could have influenced this final biomass composition, despite the C/N 443 ratio in the harvested biomass remained always at the optimum value of 6 regardless of 444 the operational conditions (Serejo et al., 2015). The main differences were recorded in 445 the S content, which decreased from 0.4% in stage I to 0.2% in stages II and III (Table 446 3). The higher S content in the biomass was recorded concomitantly with the occurrence 447 of S precipitation (Photograph 1, appendix), and was attributed to the likely S 448 absorption into the biomass. 449 19 The inoculated Chlorella sp. was gradually replaced by Chloroidium saccharophilums 450 (Chlorella saccharophila) during stage I. Chloroidium saccharophilum was the 451 dominant microalga species during stage I (94%) and stage III (100 %), while 452 Pseudanabaena sp. accounted for 6% and 54% of the total number of microalgae cells 453 in stages I and II, respectively (Fig. 6). Pseudanabaena sp. has been consistently found 454 in a similar indoors experimental set-up during the simultaneous upgrading of biogas 455 and digested vinasse treatment (Posadas et al. 2015a; Serejo et al. 2015). The lower 456 microalgae diversity recorded outdoors compared to that observed under laboratory 457 conditions in a similar experimental set-up was likely due to i) the recirculation of the 458 settled biomass and ii) the high alkalinity in the cultivation broth in stages II and III 459 (Serejo et al., 2015; Posadas et al., 2015a; Toledo-Cervantes et al., 2016, 2017; Park et 460 al., 2011). 461 ˂Figure 6> 462 4. Conclusions 463 This work constitutes the first proof-of-concept study of photosynthetic biogas 464 upgrading coupled with centrate treatment at pilot scale under outdoors conditions. The 465 feasibility of a zero-effluent process operation was also demonstrated. Temperature 466 played a key role on the efficiency of biogas upgrading at low-to-medium alkalinities, 467 while high alkalinities enhanced process robustness against daily temperature 468 variations. Process operation at L/G ratios of 1-2 provided a biomethane complying 469 with most international regulations. A consistent centrate treatment was achieved 470 regardless of the operational conditions, while the decoupling of biomass productivity 471 from the HRT allowed high recoveries of C, N and P. 472 ACKNOWLEGMENTS 473 20 This research was supported by MINECO and the European Union through the FEDER 474 program (CTM2015-70442-R and Red Novedar), the Regional Government of Castilla 475 y León (Project VA024U14 and UIC 71) and INIA (RTA2013-00056-C03-02). The 476 authors wish to thanks Julia Bilbao and Argimiro de Miguel from the Atmosphere and 477 Energy Laboratory at Valladolid University for kindly providing the temperature, 478 radiation data and number of sun hours. Valladolid University is also acknowledged for 479 funding the research contract of Esther Posadas. 480 REFERENCES 481 (1) Alcántara C., García-Encina P., Muñoz R., 2015. Evaluation of simultaneous 482 biogas upgrading and treatment of centrates in a HRAP through C, N and P mass 483 balances. Water Sci. 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The effects of various LED (light 573 emittion diode) lighting strategies on simultaneous biogas upgrading and biogas slurry 574 nutrient reduction by using of microalgae Chlorella sp. Energy 106, 554-561.575 25 FIGURE CAPTIONS 576 Figure 1. Schematic diagram of the outdoors experimental set-up used for the 577 continuous upgrading of biogas. 578 Figure 2. Time course of the concentration of (a) CH4 (■), (b) CO2 (♦) and H2S (▲), 579 and (c) O2 (●) and N2 (○) in the upgraded biogas. The removal efficiencies of CO2 (◊) 580 and H2S (∆) are also displayed in figure 2b. 581 Figure 3. Time course of the concentration of (a) CH4, (b) CO2, (c) O2 and (d) N2 in the 582 upgraded biogas during the one-day cycle evaluated in stages I (♦), II (■) and III (▲). 583 Figure 4. Time course of the concentration of (a) CH4, (b) CO2, (c) O2 and (d) N2 in the 584 upgraded biogas at L / G ratios of 0.5 (♦), 1 (□), 2 (▲) and 5 (○ ). 585 Figure 5. Time course of the influent (♦) and effluent (◊) concentrations of (a) TOC, (b) 586 IC, (c) TN, (d) N-NH4+, (e) N-NO2-, (f) N-NO3-, (g) P-PO43- and (h) SO42- throughout 587 the three operational stages. 588 Figure 6. Time course of the structure of microalgae population in the HRAP: ( ) 589 Chlorella sp., ( ) Pseudanabaena sp. and ( ) Chloroidium saccharophilum. 590 Table 1. Environmental and operational parameters during the three operational stages. STAGE PARAMETER I II III Date 05/07 - 08/08 09/08 – 06/09 07/09 – 04/10 Average temperature (ºC) 23.8 ± 6.7 23.5 ± 6.4 20.0 ± 6.7 Average PAR (µmol m-2 s-1) 1427 ± 65 1258 ± 140 946 ± 174 Number of sun hours (h) 12 ± 1 11 ± 1 9 ± 1 ICinfluent (mg L-1) 522 ± 40 2009 ± 135 2040 ± 120 Effluent from the settler (L d-1) 0.6 0.8 No effluent Table Table 2. Steady state removal efficiencies of total organic carbon, total inorganic carbon, total nitrogen, ammonium and phosphorus during the three operational stages. STAGE Removal efficiencies (%) TOC TIC TN N-NH4+ P-PO43- I 74±7 95±1 86±4 100±0 92±2 II 57±6 72±8 87±4 100±0 84±5 III 59±7 75±7 80±8 99±1 85±5 Table Table 3. Carbon and nutrient recovery via biomass assimilation estimated from the carbon and nutrients removal, and the biomass elemental composition of the harvested biomass during stages I, II and III. STAGE Carbon and nutrient recovery as biomass (%) Biomass elemental composition (%) C N P S C N P S I 97±1 65±3 100±0 26±5 41.1 6.7 1.1 0.4 II 65±6 54±18 91±9 17±3 35.8 5.7 0.7 0.2 III 66±8 76±19 99±1 16±3 37.8 6.5 0.8 0.2 Table Electronic Annex Click here to download Electronic Annex: Appendix.docx