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Phosphorus removal using a microalgal biofilm in a new biofilm photobioreactor for tertiary wastewater treatment
w a t e r r e s e a r c h 7 1 ( 2 0 1 5 ) 5 5 e6 3
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Phosphorus removal using a microalgal biofilm in a new biofilm photobioreactor for tertiary wastewater treatment a,*, Martin Trtı´lek b, Toma s Rataj a,1 Katerina Sukacova Global Change Research Centre, Academy of Sciences of the Czech Republic, Belidla 986/4a, Brno 603 00, Czech Republic b sov 470, Czech Republic Photon Systems Instruments, 664 24 Dra a
article info
abstract
Article history:
Eutrophication of surface water has been an important environmental issue for nearly half
Received 13 June 2014
a century. High concentrations of phosphorus contribute to the process of eutrophication,
Received in revised form
resulting in the demand for effective and economic methods of phosphorus removal from
4 November 2014
treated water. The aim of this study was to evaluate the capacity for phosphorus removal
Accepted 29 December 2014
of a microalgal biofilm during different light regimes. The photobioreactor was operated for
Available online 7 January 2015
nine months each year over a two-year period without interruption and without any need of re-inoculation. The algal biofilm was able to remove 97 ± 1% of total phosphorus from
Keywords:
wastewater during 24 h of continuous artificial illumination. The average TP uptake rate in
Microalgal biofilm
our experiments was 0.16 ± 0.008 g m2 d1. Phosphorus removal values ranged from 36 to
Phosphorus removal
41% when the algal biofilm was illuminated by natural light (12 h sunlighte12 h night). The
Wastewater treatment
biomass production rate was 12.21 ± 10 g dry weight m2 d1 in experiments with continuous artificial light and 5.6 ± 1 g dry weight (DW) m2 d1 in experiments with natural light. These results indicate the great potential of microalgal biofilms in the tertiary treatment of wastewater. © 2015 Elsevier Ltd. All rights reserved.
1.
Introduction
Degradation of water ecosystems is a major problem in most countries of the world. Industrial and agricultural manufacturing is responsible for a huge production of wastewater, which must be treated to prevent pollution of water ecosystems (Omernik, 1977; Schlesinger, 1991; Vitousek et al., 1997). High concentration of phosphorus starts a process of eutrophication in the water environment. Dodds et al.
* Corresponding author. Tel.: þ420 511 440 038. ova ). E-mail address: [email protected] (K. Sukac 1 Tel.: þ420 511 440 038. http://dx.doi.org/10.1016/j.watres.2014.12.049 0043-1354/© 2015 Elsevier Ltd. All rights reserved.
(1997) concluded that maintenance of stream water total phosphorus concentration at <30 mg L1 would be necessary to keep benthic algal biomass below nuisance levels of 100 mg m2. Higher concentration of phosphorus leads to degradation of freshwater ecosystems and reduces the possibility of water reclamation. As a result, there is an urgent need for new, effective and economic methods of phosphorus removal. Currently, chemical methods are used most often for phosphorus removal, including co-precipitation with iron, alum or lime (Penetra et al., 1999).
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Biological methods of nutrient removal are considered the most environmentally favorable and least expensive of wastewater treatment methods, as there are no need of chemical precipitants (Mantzavinos and Kalogerakis, 2005). Biological removal of phosphorus is performed by different groups of microorganisms. Biological methods involve enhanced biological phosphorus removal (EBPR) which are based on polyphosphate accumulating organisms (PAOs). These microorganisms are able to store phosphate as intracellular polyphosphate in the specific conditions during wastewater treatment (De-Bashan and Bashan, 2004). The EBPR process is capable of efficient phosphorus removal performance. However disturbances and long periods of insufficient P removal have been observed at full-scale plants on numerous occasions (Oehmen et al., 2007). Other biological methods involve the use of algae and cyanobacteria for phosphorus removal (Rawat et al., 2011). Numerous studies have shown the ability of microalgae and cyanobacteria to grow in and reduce the nutrient content of wastewater (Chevalier et al., 2000; Doria et al., 2012; Renuka, 2013). This method of remediation is more environmentally sustainable because it does not generate additional waste such as sludge, it provides an opportunity for efficient recycling of nutrients, and it has the potential for sustainable productions of algal biofuels or bioactive compounds (Singh et al., 2005; Pittman et al., 2011). Microalgae and cyanobacteria are cultivated mainly in suspension (Christenson and Sims, 2011). However, algae are able to grow as biofilms on solid surfaces. Phototrophic biofilms are multi-layered community of photoautotrophs (cyanobacteria and microalgae) and heterotrophs (bacteria, fungi and protozoa) which play a key role in the self-purification of aquatic ecosystems (Sabater et al., 2002). In the case of biological phosphorus removal there have been numerous studies using algal biofilms successfully for the phosphorus removal under laboratory conditions (Guzzon et al., 2008; Johnson and Wen, 2010; Boelee et al., 2011; Liu et al., 2013; Shi et al., 2014). However, little data have been published concerning the capacity of microalgal biofilms for phosphorus removal in the large scale cultivation. Previously, the attached benthic algae were used in algal turf scrubber units (ATS) for the removal of nitrogen and phosphorus from different type of wastewater such as dairy manure (Kebede-Westhead et al., 2003) or domestic sewage (Craggs et al., 1996). ATS technology has been tested mainly in the southern regions of the USA, especially in California and Florida (Adey et al., 2011). The use of ATS for water quality improvement is regarded as an established practise (Adey and Loveland, 2007). As compared with ATS technology the application of phototrophic biofilms in tertiary wastewater treatment is still uncommon (Guzzon et al., 2008) and needs further research to develop effective technology for wastewater treatment. The current study has two aims: (a) to evaluate the capacity of an algal biofilm for phosphorus removal under light conditions consistent with the temperate climate of Central Europe; and (b) to estimate the comprehensive benefit of the presented technology, including biomass production and the assessment of harvested biomass as a source for biogas.
2.
Materials and methods
2.1.
Horizontal Flat Panel photobioreactor
A new large scale experimental photobioreactor was designed and built for the study of biofilms (Fig. 1). The Horizontal Flat Panel (HFP) photobioreactor for algal biofilm cultivation was developed by Photon Systems Instruments (Brno, Czech Republic) and consisted of two modules. One module was a concrete slab (1 2 0.02 m) supported by a metal frame which formed a 2 m2 cultivation area for biofilm growth. Wastewater was pumped from a reservoir (0.2 m3, with operational volume of 0.1 m3) by a magnetically powered centrifugal pump (IDRA 231 720, Sicce S.p.A., Italy) to the upper part of the cultivation area. Wastewater flowed over the cultivation area in a thin layer and then back to the reservoir. The flow rate of water pumped from the reservoir to the cultivation area was 8.8 L min1, with a laminar flow velocity of about 0.25 m s1 over the cultivation area. The circulating volume of wastewater was 100 L, with a retention time in the reservoir of approximately 7 min. The HFP photobioreactor (two modules) had a total cultivation area of 4 m2, and was placed in a greenhouse. Photosynthetically active radiation (PAR) was measured with a Quantum Sensor (EMS 12, EMS Brno, Czech Republic), inside the greenhouse. The average daily sum of irradiance was calculated from actual PAR values and expressed in MJ m2 d1. The temperature of the circulating wastewater was recorded regularly and varied between 19 and 24 C.
2.2.
Microalgal biofilm sampling and cultivation
An algal biofilm assemblage was sampled from the pools of a rendering plant (Glogow, Poland, 51 370 39.45600 N, 16 90 47.83600 E) to obtain a community with tolerance to high nutrient concentrations. The algal samples were taken from the mud surface of sludge pools and put into plastic bottles and stored in a cool box for transportation to the laboratory. Algal biofilm sheets were put into Bold's Basal medium (Bischoff and Bold, 1963) in several Erlenmeyer flasks to obtain sufficient volume of algal biomass for algal biofilm cultivation. The algal assemblage was cultivated under 30 mmol photons m2 s1 at 25 C for two weeks, then the species composition of algal assemblage in each Erlenmeyer flask was determined. The identification of microalgae and cyanobacteria was performed
Fig. 1 e Scheme of the experimental setup.
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Table 1 e Taxonomical analyses of the species present in biofilm in the season 2012 and 2013. Species
Inoculum used for
Start of experiments
2012
2012
2013
2013
HFP-photobioreactor
25 April 2012
16 August
29 October
25 April
14 August
Rare Dominant Rare
Dominant Dominant x
Dominant Dominant Rare
Dominant Dominant Rare
Dominant Dominant Rare
Dominant Dominant Rare
Dominant Dominant
Dominant x
Dominant x
Dominant x
Dominant x
Dominant x
Rare
Rare
Rare
Rare
Rare
Rare
Cyanobacteria Phormidium autumnale Pseudanabaena sp. Chrococcus sp. Coccal green algae Scenedesmus acutus Monoraphidium contortum Diatoms Cymbella minuta
rek and Anagnostidis (2005) and Ettl and according to Koma € rtner (1995) using an Olympus CX 31 microscope. The Ga microalgal assemblage with filamentous cyanobacteria and non-motile unicellular green algae was selected for following cultivation. The inoculum for the HFP-photobioreactor was prepared in a laboratory using the same design planned for a large scale system. A concrete slab (0.2 0.3 m) was placed in a plastic box with cultivation medium, and the medium was pumped to the cultivation area where it flowed over the surface of the slab. The selected microalgal assemblage was inoculated onto the surface of the slab where it formed a uniform film. The species composition of microalgal assemblage used for inoculation of the HFP-photobioreactor is given in Table 1. Inoculation of the HFP-photobioreactor was performed in a similar manner. An algal suspension with a biomass density of about 25 g DW L1 was applied to the cultivation area before starting the photobioreactor operation and pumping wastewater across the slab. The Algal biofilm was cultivated under sunlight (4e8 MJ m2 d1). The period of sunshine was approximately 14 h per day. The firm algal biofilm covered the whole cultivation area after 3 weeks.
2.3.
Experimental setup
Experiments were started when the algal biofilm covered the whole cultivation area. Performed experiments and experimental conditions are described in Table 2. Initially, batch
experiments were performed to identify TP removal during a five day period. Hydraulic retention time for continuous experiments was determined according to the results of batch experiments.
2.3.1.
Batch experiments
Batch experiments were conducted in two periods with markedly different light conditions. Daily values of irradiance ranged between 7 and 11 MJ m2 d1 in summer. The range of daily irradiance was between 2 and 10 MJ m2 d1 in autumn and spring. Batch experiments lasted five days and involved artificial wastewater.
2.3.2.
Continuous experiments
Continuous experiments were performed over four days repeatedly using two regimes: (a) During continuous experiments A and B, the algal biofilms were illuminated continuously by high pressure sodium lamps (Osram NAV-T Super 4Y e 600W) at an intensity of 90 ± 15 mmol photons m2 s1 and with a daily sum of irradiance of 1.7 MJ m2 d1. Artificial wastewater was used during experiments. (b) Continuous experiments C and D were performed under solar irradiance using real wastewater. Solar irradiance and real wastewater were used to imitate real conditions in the wastewater treatment plant. Average daily sums of solar irradiance were 1.57 ± 0.88 MJ m2 d1
Table 2 e Summary of performed experiments and experimental conditions. Experiment
Type of light
Batch experiment Exp. with high values Sunlight of solar radiation Exp. with low values Sunlight of solar radiation Continual experiment Experiment A Artificial light Experiment B Artificial light Experiment C Sunlight Experiment D Sunlight Experiments over 24-h time periods Experiment 1 Artificial light Experiment 2 Sunlight
Light regime
Light intensity (mmol m2 s1)
DayeNight
0e1650
7e11
Artificial
DayeNight
0e970
2e10
Artificial
Continual Continual DayeNight DayeNight
90 ± 15 90 ± 15 0e833 0e904
1.7 1.7 1.57 ± 0.88 3.1 ± 0.64
Artificial Artificial Real Real
Continual DayeNight
90 ± 15 0e815
1.7 1.73 ± 0.47
Artificial Real
Aver. daily sum of irradiance (MJ m2 d1)
Wastewater type
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(exp. C) and 3.1 ± 0.64 MJ m2 d1 (exp. D). The period of sunshine was approximately 12 h per day.
2.3.3.
Experiments over 24-h time periods
Experiments were carried out in the continual light regime and also in the dayenight light regime using both artificial and real wastewater. Experiments were run using a batch set-up. TP concentration and pH were measured every two hours to evaluate the influence of a diurnal light cycle on phosphorus removal.
2.3.4.
Wastewater characteristics
Artificial wastewater was constituted with total phosphorus (TP) concentration of 3 mg L1, typical of wastewater effluent from a municipal wastewater treatment plant that does not practice enhanced phosphorus removal (Water Environment Federation, 2010). The artificial wastewater was enriched by major and trace elements: 120 mg L1 NaNO3; 6 mg L1 MgSO4$7H2O; 2 mg L1 CaCl2$2H2O; 2 mg L1 NaCl; 6 mg L1 K2HPO4; 14 mg L1 KH2PO4; 4 mg L1 EDTA$2H20; 0.4 mg L1 FeSO4$7H2O; 0.9136 mg L1 H3BO3; 0.7056 mg L1 ZnSO4$7H2O; 0.1152 mg L1 MnCl2$4H2O; 0.1936 mg L1 Na2MoO4$2H2O; 0.1256 mg L1 CuSO4$5H2O; 0.0392 mg L1 Co(NO3)2$6H2O. Real wastewater was collected in the effluent from a sov (3500 Population wastewater treatment plant in Dra Equivalent), Czech Republic. Chemical parameters of real wastewater are given in Table 3.
2.4.
Analytical methods
Samples of wastewater were taken every day during batch and continuous experiments. The samples for phosphorus analysis were filtered through a 0.45 mm membrane filter (ME 25 e mixed cellulose ester, Whatman). TP concentration was measured using an ammonium molybdate spectrophotometric method (ISO 6878: 2004). Algal biomass was sampled from a defined area (9 cm2). Biofilm biomass was dried on pre-weighed filters (Whatmann, GFC filters) to a constant weight at 90 C, then the filters were weighed. Biomass production was expressed as g DW m2. Most of the biomass was harvested by squeegee after finishing the experiments. Harvested biomass was dried and used for anaerobic digestion tests. The rest of the algal biomass left on the cultivation area served for renewal of the algal biofilm. New experiments were conducted immediately after harvesting. The analysis of biomass distribution over the cultivation area was carried out at the end of the experimental season. One hundred samples of biofilm biomass were taken from the cultivation area. Each sample had an area of 9 cm2. The sampled areas were equally spread over the cultivation area.
Table 3 e Chemical characteristic of real wastewater. Chemical parameter Total phosphorus Total nitrogen NO3eN NO2eN NH4eN
Concentration (mg L1) 2.9 ± 0.1 49.4 ± 3 48.6 ± 2 0.23 ± 0.02 0.33 ± 0.03
2.5.
Anaerobic digestion processes
Anaerobic digestion tests were performed as described in Samson and LeDuy (1986). Adapted sludge was taken from the large anaerobic digester (second stage of fermentation pro c , Czech Republic). The adapted cess) in the biogas station (Cej sludge was placed in laboratory digesters. 15 g of dried biofilm biomass was added to the sludge. Tests were carried out in triplicate for two week periods in mesophilic temperature conditions (41 C). The total amount of produced gas and methane content were measured daily by gas analysis with €ger X-am® 7 000, infrared and electrochemical sensor (Dra € ger Safety AG & Co. KGaA, Germany). Dra
3.
Results
3.1.
Microalgal biofilm cultivation
The HFP photobioreactor was operated in Brno in the southern Czech Republic over a 9 month period for two years (AprileDecember 2012, AprileDecember 2013). After inoculation the algal biofilm grew across the entire slab surface within 3 weeks. Biomass ranged from 30 to 70 g DW m2 on the cultivation area. Nearly 50% of the cultivation area was covered with 40e50 g DW m2, with 30e40 g DW m2 over 26% of the area and 50e60 g DW m2 over 20% of the area. 5% of the area was covered with 60e70 g DW m2 of biofilm. The cyanobacterialemicroalgal assemblage constituted mostly of a mat created by the filamentous cyanobacteria Phormidium autumnale and Pseudanabaena sp., as well as the unicellular green alga Scenedesmus acutus which was distributed among cyanobacterial filaments. These dominant species were accompanied by Cymbella minuta (diatom) and Chroococcus sp. (unicellular cyanobacteria). The species composition did not change substantially (Table 1).
3.2.
Phosphorus removal and uptake
3.2.1.
Batch experiments
Fig. 2 shows the reduction of TP concentration during batch experiments. In the high solar radiation period (7e11 MJ m2 d1) the algal biofilm removed 92 ± 7% of TP with an average value of TP uptake rate of 0.14 ± 0.011 g m2 d1. This value was achieved by 24 h. In the low solar irradiance period, 50 ± 16% of TP removal was obtained after 24 h. The percentage gradually increased, attaining 81 ± 20% after 72 h. At the end of the experiments (96 h), values of TP removal were 74 ± 15%. The maximum TP uptake rate was 0.069 ± 0.024 g m2 d1 by 24 h during the period of low solar radiation. Maximum TP uptake rate was observed after 24 h of the experiment starting. During the following two days, the minimum values of TP uptake rate were 0.022 ± 0.007 g m2 d1 (48 h) and 0.025 ± 0.035 g m2 d1 (72 h).
3.2.2.
Continuous experiments
Under continual illumination the algal biofilm removed 97 ± 1% of TP from wastewater within 24 h. The average influent value 3.25 ± 0.16 mg TP L1 dropped to 0.082 ± 0.032 mg TP L1 after 24 h in the case of experiments
w a t e r r e s e a r c h 7 1 ( 2 0 1 5 ) 5 5 e6 3
59
concentration dropped only slightly. An increase in pH was detected with reduction of TP concentration, the highest changes in pH level coinciding with maximum reduction rate of TP. A maximal pH value of 9.98 ± 0.6 was measured after 24 h. The process of phosphorus removal evaluated during experiments with dayenight light regime (Figs. 4 and 5) was markedly different than in other experiments. During the diurnal cycle there were two phases of TP removal. Nearly 99% of TP was removed during the light phase (time period 8 h), during which pH values rose to a maximum value 10.17. Declining pH values were measured during the dark phase, during which TP concentration increased from 0.04 mg L1 to a maximum value of 1.48 mg L1 over a time period 12 h. Fig. 2 e Reduction of total phosphorus concentration during batch experiments.
with continuous light. The average TP uptake rate was 0.16 ± 0.008 g m2 d1. Phosphorus removal values were lower in the experiments in which solar irradiance was used for illumination of the algal biofilm. The influent value of 2.96 ± 0.022 dropped to only 1.76 ± 0.67 mg TP L1, and only 41 ± 11% of TP was removed during experiment C. In experiment D, the average influent concentration of 2.3 ± 0.06 decreased to 1.6 ± 0.3 and the algal biofilm removed 36 ± 9% of TP after 24 h. The average values of TP uptake rate were 0.06 ± 0.016 g m2 d1 (exp. C) and 0.042 ± 0.01 g m2 d1 (exp. D).
3.2.3.
Experiments over 24-h
Fig. 3 illustrates results of experiments over a 24-h time period using continuous light. After 18 h the TP concentration decreased from 3.0 ± 0.17 mg L1 to 0.09 ± 0.014 mg L1, the algal biofilm removing 97% of TP from the wastewater. The maximum reduction rate of TP was reached during the 8-h period at the start of the experiment, after which the TP
Fig. 3 e Diurnal course of pH values and total phosphorus concentration during experiments over 24-h time period with continual illumination.
3.3.
Biomass production and anaerobic digestion
Measured values of biomass production rate varied enormously during experiments. Markedly higher biomass production was measured in experiments with continuous illumination. The average value was 12.21 ± 10 g DW m2 d1. Lower values of 5.6 ± 1.4 g DW m2 d1 were measured in the experiments using solar irradiance with a dayenight light regime. A biomass production rate of 9.8 ± 5 g DW m2 d1 was measured during batch experiments. Harvested biomass was assessed for potential biogas production. During anaerobic digestion of dried biomass, the biogas yield reached 0.33 ± 0.03 m3 kg1 VS (Volatile Solids) added. Methane production was measured at 0.153 ± 0.015 m3 kg1 VS. Average methane content was measured at 46% in the yield biogas, with a maximum content measured at 66% after 10 days of anaerobic digestion. Maximum TP uptake capacity was used to estimate the required area of an algal biofilm for tertiary wastewater treatment in a plant with a capacity from 2000 to 6000 population equivalent (PE). The estimated area was calculated to be in range from 0.3 ha (2000 PE) to 1 ha (6000 PE), assuming a reduction of TP concentration from 3 mg TP L1 to 0.09 mg TP L1. Table 4 shows the production potentials associated with algal biofilm biomass yield per treatment of 1 m3 of
Fig. 4 e Diurnal course of pH values and total phosphorus concentration during experiments over 24-h time period with dayenight light regime.
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Fig. 5 e Diurnal course of total phosphorus concentration and the light intensity in the experiment C.
wastewater. Dry biomass production of 0.13 kg DW would be obtained from 1 m3 treated wastewater. Biogas and biomethane yield would be 0.043 and 0.03 m3 per m3 of wastewater, respectively. It was assumed that biogas contains 66% of methane. A power yield 1.13 MJ could be obtained during treatment of 1 m3 of wastewater, assuming energy content of CH4 to be 37.8 MJ m3 (Prajapati et al., 2013). Annual dry biomass production was estimated at 44.4 t ha1 y1.
4.
Discussion
A new type of photobioreactor for algal biofilm cultivation was tested and validated for technical design and functionality. An algal biofilm was cultivated and weekly harvested for nearly two years after initial inoculation without interruption and without the need for re-inoculation. Similarly Sandefur et al. (2011) described a nine-month operating period for an algal turf scrubber system, and Mulbry et al. (2008) reported on algal turf scrubber raceways that were operated for approximately 270 days each year between 2003 and 2006 in central Maryland. This study showed that biofilm activity decreased TP concentrations in all the experiments, with particularly high efficiency of phosphorus removal achieved during experiments with a continuous light regime. We measured an average TP 2 1 d uptake rate about 20% higher than the 0.13 g PO3 4 eP m reported by Boelee et al. (2011) during laboratory experiments
under continual illumination 230 mE m2 s1. Although a lower intensity of illumination (90 ± 15 mE m2 s1) was used in the current study, a higher efficiency of phosphorus removal was attained. We assume these results were due to using a different design of the algal biofilm system or because there was a different species composition in the algal biofilm. The Boelee et al. (2011) biofilm consisted mainly of pennate diatoms and green filamentous algae, whereas in our study the microalgal assemblage consisted of a net of filamentous cyanobacteria with unicellular green algae interspersed. The phosphorus uptake difference could be due to the presence of more cyanobacteria and coccal green algae. In both cyanobacteria and coccal green algae, phosphate can be accumulated as polyphosphate granules (Jansson, 1988; Guzzon et al., 2008). The efficiency of phosphorus removal was less in experiments with a dayenight light regime and using solar radiation. Light is one of the most important factor influencing the uptake of phosphate by algae, because photosynthesis is et al., 1997). Phosphate uptake by depended on light (Laliberte cyanobacteria is usually greater in the light than in the darkness (Whitton, 1992). During this study TP removal values varied between 36 and 41%. Sandefur et al. (2011) reported an average value of TP removal of 48% for the algal turf scrubber system under the warm climate of Arkansas, USA. Boelee et al. (2014a) reported an average removal efficiency of PO3 4 eP of 14% for an outdoor phototrophic biofilm reactor operated in the cooler, lower light, conditions in the Netherlands. Placement of our bioreactor in the greenhouse attenuated the impact of low temperature and other weather events in the Czech Republic, and allowed TP removal only 7e12% lower than measured in the warmer, sunnier climate of Arkansas. Diurnal changes of phosphorus concentration and pH values were observed during experiments over a 24 h period. Values of pH rose significantly during the light period. Similar pH shifts have been reported during phosphorus uptake by et al., 1997; numerous authors (Chevalier et al., 2000; Laliberte Sandefur et al., 2011). The pH shift corresponds with the absorption of CO2 during high photosynthetic activity (Small and Adey, 2001). Algae remove phosphorus from wastewater by processes of adsorption, assimilation and precipitation (Craggs et al., 1996). The precipitation of phosphorus with cations (such as Ca2þ and Mg2þ) at high pH is known to begin between pH 8.9e9.5 (Belsare and Belsare, 1987). Our results indicate that
Table 4 e Estimated production potential of algal biofilm wastewater treatment process. Process
Value
Dry biomass production (kg DW m3 treated wastewater) Biogas yield (m3 biogas m3 treated wastewater) Biomethane yield (m3 biomethane m3 treated wastewater) Estimated power yield (MJ m3 treated wastewater) Dry biomass production (kg dry weight ha1 y1) Biogas yield (m3 biogas ha1 y1) Biomethane yield (m3 biomethane ha1 y1) Estimated power yield (GJ ha1 y1)
0.13
Expressed to 1 m3 treated wastewater
Remarks
0.043 0.03
Expressed to 1 m3 treated wastewater 66% CH4 in biogas, Expressed to 1 m3 treated wastewater
1.13 44 408 14 655 9770 0.37
CH4 energy content 37.9 MJ m3, Expressed to 1 m3 treated wastewater Estimated annual production 66% CH4 in biogas, Estimated annual production Estimated annual production CH4 energy content 37.9 MJ m3, Estimated annual production
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TP removal depends on both absorption and precipitation, as indicated by the decrease of pH values, and simultaneous increase of TP concentration, during the night, indicating dissolution of precipitated phosphorus. This process was also reported by Craggs et al. (1996) for the algal turf scrubber system. We assumed that the 58% of TP returned to the water during night corresponded with the proportion of phosphorus which had been removed by precipitation. The rest of the TP (41%) would be the proportion removed by assimilation (include luxury uptake) of the algal biofilm. Boelee et al. (2014b) reported increased content of phosphorus (P), magnesium (Mg) and calcium (Ca) in the algal biofilm during the P-removal process, and assumed that this indicated precipitation of Ca and Mg phosphates in the biofilm. A reduction of precipitated phosphorus dissolution during the night would result in a more effective process of phosphorus removal. One option is the application of a continuous light regime. Exposure to sunlight during day and to artificial light during the night would increase the efficiency of the whole process. In our study, observed values of arial biomass production rate varied considerably during experiments. Biomass over the cultivation area ranged from 30 to 70 g DW m2. This high variability could have been caused by differences in the dispersion of biomass over the large cultivation area (4 m2). The degree of variability was also affected by cultivation conditions, since homogenous cultivation conditions are almost impossible to maintain over such a large cultivation area (4 m2). Boelee et al. (2014) also reported excessively variable values of arial biomass production during cultivation of the algal biofilm. Measured values of arial biomass production were similar to results of Boelee et al. (2011) who reported a maximum biomass production of 7.7 g DW m2 d1. However, our values are higher than the biomass production of 2.57 g DW m2 d1 reported for an attached microalgal growth system using polystyrene foam as a supporting material (Johnson and Wen, 2010). On the other hand, algal biomass production reported for an algal turf scrubber system at 25 g DW m2 d1 (Mulbry et al., 2008) were more than double the values measured during this study. Sandefur et al. (2011) reported 31 ± 19 g DW m2 d1. However, both studies were conducted under warmer climate conditions. Harvested biomass was assessed for potential of biogas production. Measured biogas production was similar to that reported in other studies. Prajapati et al. (2014) measured biogas production in a range from 0.34 to 0.464 m3 kg1 VS for different Chlorella species, which is similar to our values of 0.33 m3 kg1 VS. Mussgnug et al. (2010) investigated anaerobic digestion for different algal species and measured maximal biogas yield of 0.587 m3 kg1 VS for Chlamydomonas rheinhardtii, and minimal yield of 0.287 for Scenedesmus obliquus. In our study, the average methane content was low compared with other studies (e.g. Prajapati et al., 2013), although the maximal achieved methane content of 66% corresponds with the finding of other researchers (e.g. Sialve et al., 2009). The low average methane content in the biogas resulted from the short time period of the digestion test, lasting only 15 days. Maximal methane content was observed after 10 days of the test. We
61
would expect a higher methane yield following a longer anaerobic digestion process. The annual calculated dry biomass production in our study is about 30% more than that reported by Prajapati et al. (2014) for Chlorella pyrenoidosa, and that reported by Chinnasamy et al. (2010) for algal assemblage cultivated in carpet industry wastewaters. These data indicate that cyanobacteriaealgae assemblages immobilized in a biofilm can produce similar amount of biomass to algae cultivated in suspension. Moreover, biofilm cultivation allows for simple and low cost harvesting protocols. The annual potential energy generation of the harvested biofilm is estimated about 369 GJ ha1 y1, enough for warming the greenhouse and operation of the bioreactor. Anaerobic digestion plays an important role in the disposal of activated waste sludge, reducing treatment costs of wastewater plants, and their environmental footprint. The process is considered a major and essential part of a modern wastewater treatment plant (Appels et al., 2008). Combining wastewater treatment facilities with biogas production processes is an exciting possibility for wastewater management.
5.
Conclusions
This study demonstrates the ability of an algal biofilm to remove phosphorus from municipal wastewater. An algal biofilm was able to remove 97 ± 1% of total phosphorus from wastewater after 24 h under artificial continuous illumination. Phosphorus removal values ranged from 36 to 41% when the algal biofilm was illuminated by solar irradiance in a 12 h lighte12 h dark regime. pH values in wastewater decreased in the dark phase of the experiments, associated with CO2 release and dissolution of precipitated phosphorus. The results of experiments indicated that efficiency of P removal could be increased by imposing a regime of sunlight during the day and artificial light during the night. Measured biogas yield showed the economic potential to integrate algal biofilm wastewater treatment with further anaerobic digestion in small and medium wastewater treatment plants.
Acknowledgments This publication is an output of the CzechGlobe Centre that is being developed within the OP RDI and co-financed from EU funds and the State Budget of the Czech Republic (Project: CzechGlobe e Centre for Global Climate Change Impacts Studies), Reg. No. CZ.1.05/1.1.00/02.0073. The authors like to thank Dr. Tomas Vitez for anaerobic digestion of algal rkova for critically reading the biomass, Dr. Jaroslava Koma manuscript and Dr. Stephen Hunt (Qubit Systems Inc., Kingston, Canada) for reviewing the manuscript.
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Adey, W.H., Kangas, P.C., Mulbry, W., 2011. Algal Turf Scrubbing: cleaning surface waters with solar energy while producing a biofuel. Bioscience 61 (6), 434e441. Appels, L., Bayens, J., Degreve, J., Dewil, R., 2008. Principles and potential of the anaerobic digestion of waste-activated sludge. Prog. Energy Combust. Sci. 34, 755e781. Belsare, D.K., Belsare, S.D., 1987. High-rate oxidation pond for water and nutrient recovery from domestic sewage in urban areas in the tropics. Arch. Hydrobiol. Beih. Ergebn. Limnol. 28, 123e128. Boelee, N.C., Temmink, H., Janssen, M., Buisman, C.J.N., Wijffels, R.H., 2011. Nitrogen and phosphorus removal from municipal wastewater effluent using microalgal biofilms. Water Res. 45, 5925e5933. Boelee, N.C., Janssen, M., Temmink, H., Shrestha, R., Buisman, C.J.N., Wijffels, R.H., 2014a. Nutrient removal and biomass production in an outdoor pilot-scale phototrophic biofilm reactor for effluent polishing. Appl. Biochem. Biotechnol. 172, 405e422. iu te, L., Boelee, N.C., Janssen, M., Temmink, H., Taparavic Buisman, C.J.N., Wijffels, R.H., noska, A., Khiewwijit, R., Ja 2014b. The effect of harvesting on biomass production and nutrient removal in phototrophic biofilm reactors for effluent polishing. J. Appl. Phycol. 26, 1439e1452. Chevalier, P., Proulx, D., Lessard, P., Vincent, W.F., Nou¨e, J., 2000. Nitrogen and phosphorus removal by high latitude matforming cyanobacteria for potential use in tertiary wastewater treatment. J. Appl. Phycol. 12, 105e112. Chinnasamy, S., Bhatnagar, A., Hunt, R.W., Das, K.C., 2010. Microalgae cultivation in a wastewater dominated by carpet mill effluents for biofuel applications. Bioresour. Technol. 101, 3097e3105. Christenson, L., Sims, R., 2011. Production and harvesting of microalgae for wastewater treatment, biofuels and bioproducts. Biotechnol. Adv. 29, 686e702. Craggs, R.J., Adey, W.H., Jessup, B.K., Oswald, W.J., 1996. A controlled stream mesocosm for tertiary treatment of sewage. Ecol. Eng. 6, 149e169. De-Bashan, L.E., Bashan, Y., 2004. Recent advances in removing phosphorus from wastewater and its future use as fertilizer. Water Res. 38, 4222e4246. Dodds, W.K., Smith, V.H., Zander, B., 1997. Developing nutrient targets to control benthic chlorophyll levels in streams: a case study of the Clark Fork River. Water Res. 31, 1738e1750. Doria, E., Longoni, P., Scibilia, L., Iazzi, N., Cella, R., Nielsen, E., 2012. Isolation and characterization of a Scenedesmus acutus strain to be used for bioremediation of urban wastewater. J. Appl. Phycol. 24, 375e383. € rtner, G., 1995. Syllabus der Boden-, Luft- und Ettl, H., Ga Flechtenalgen. Gustav Fischer Verlag, Stuttgart, Germany, 721 pp. Guzzon, A., Bohn, A., Diociaiuti, M., 2008. Cultured phototrophic biofilms for phosphorus removal in wastewater treatment. Water Res. 42 (16), 4357e4367. Jansson, M., 1988. Phosphate uptake and utilization by bacteria and algae. Hydrobiologia 170, 177e189. Johnson, M., Wen, Z., 2010. Development of an attached microalgal growth system for biofuel production. Appl. Microbiol. Biotechnol. 85, 525e534. Kebede-Westhead, E., Pizzaro, C., Mulbry, W.W., 2003. Production and nutrient removal by periphyton grown under different loading rates of anaerobically manure digested flushed dairy. J. Phycol. 39, 1275e1282. rek, J., Anagnostidis, K., 2005. Cyanoprokaryota, 2. Teil: Koma € rtner, G., Krienitz, L., Oscillatoriales. In: Bu¨del, B., Ga Schagerl, M. (Eds.), Su¨sswasserflora von Mitteleuropa, vol. 18/ 2. Elsevier, Mu¨nchen, p. 759.
, G., Lessard, P., de la Nou¨e, J., Sylvestre, S., 1997. Effect of Laliberte phosphorus addition on nutrient removal from wastewater with the cyanobacterium Phormidium bohneri. Bioresour. Technol. 59, 227e233. Liu, T., Wang, J., Hu, Q., Cheng, P., Ji, B., Liu, J., Chen, Y., Zhang, W., Chen, X., Chen, L., Gao, L., Ji, Ch., Wang, H., 2013. Attached cultivation technology of microalgae for efficient biomass feedstock production. Bioresour. Technol. 127, 216e222. Mantzavinos, D., Kalogerakis, N., 2005. Treatment of olive mill effluents: part I. Organic matter degradation by chemical and biological processes e an overview. Environ. Int. 31, 289e295. Mulbry, W., Kondrad, S., Pizarro, C., Kebede-Westhead, E., 2008. Treatment of dairy manure effluent using freshwater algae: algal productivity and recovery of manure nutrients using pilot-scale algal turf scrubbers. Bioresour. Technol. 99, 8137e8142. Mussgnug, J.H., Klassen, V., Schlu¨ter, A., Kruse, O., 2010. Microalgae as substrates for fermentative biogas production in a combined biorefinery concept. J. Biotechnol. 150, 51e56. Oehmen, A., Lemon, P.C., Carvalho, G., Yuan, Z., Keller, J., Blackall, L.L., Reis, M.A.M., 2007. Advances in enhanced biological phosphorus removal: from micro to macro scale. Water Res. 41, 2271e2300. Omernik, J., 1977. Nonpoint Source-stream Nutrient Level Relationships: a Nationwide Study (US EPA e 600/3-77-105). Special Studies Branch. Corvalis Environmental Research Laboratory, Office of Research and Development, US Environmental Protection Agency, Washington D.C. Penetra, R.G., Reali, M.A.P., Foresti, E., Campos, J.R., 1999. Posttreatment of effluents from anaerobic reactor treating domestic sewage by dissolved-air flotation. Water Sci. Technol. 40, 137e143. Pittman, J.K., Dean, A.P., Osundeko, O., 2011. The potential of sustainable algal biofuel production using wastewater resources. Bioresour. Technol. 102, 17e25. Prajapati, S.K., Kaushik, P., Malik, A., Vijay, V.K., 2013. Phycoremediation and biogas potential of native algal isolates from soil and wastewater. Bioresour. Technol. 135, 232e238. Prajapati, S.K., Malik, A., Vijay, V.K., 2014. Comparative evaluation of biomass production and bioenergy generation potential of Chlorella spp. through anaerobic digestion. Appl. Energy 114, 790e797. Rawat, I., Kumar, R.R., Mutanda, T., Bux, F., 2011. Dual role of microalgae: phycoremediation of domestic wastewater and biomass production for sustainable biofuels production. Appl. Energy 88, 3411e3424. Renuka, N., Sood, A., Ratha, S.K., Prasanna, R., Ahluwalia, A.S., 2013. Evaluation of microalgal consortia for treatment of primary treated sewage effluent and biomass production. J. Appl. Phycol. 25 (5), 1529e1537. ~ oz, I., 2002. The effect of Sabater, S., Guasch, H., Romanı´, A., Mun biological factors on the efficiency of river biofilms in improving water quality. Hydrobiologia 469, 149e156. Samson, R., LeDuy, A., 1986. Detailed study of anaerobic digestion of Spirulina maxima algal biomass. Biotechnol. Bioeng. 28, 1014e1023. Sandefur, H.N., Matlock, M.D., Costello, T.A., 2011. Seasonal productivity of a periphytic algal community for biofuel feedstock generation and nutrient treatment. Ecol. Eng. 37, 1476e1480. Schlesinger, W.H., 1991. Biogeochemistry: an Analysis of Global Change. Academic Press, San Diego. Shi, J., Podola, B., Melkonian, M., 2014. Application of a prototypescale Twin-Layer photobioreactor for effective N and P removal from different process stages of municipal wastewater by immobilized microalgae. Bioresour. Technol. 154, 260e266.
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Sialve, B., Bernet, N., Bernard, O., 2009. Anaerobic digestion of microalgae as a necessary step to make microalgal biodiesel sustainable. Biotechnol. Adv. 27 (4), 409e416. Sing, S., Kate, B.N., Banerjee, U.C., 2005. Bioactive compounds from cyanobacteria and microalgae: an overview. Crit. Rev. Biotechnol. 25 (3), 73e95. Small, A.M., Adey, W.H., 2001. Reef corals, zooxanthellae and free-living algae: a microcosm study that demonstrates synergy between calcification and primary production. Ecol. Eng. 16 (4), 443e457.
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Phosphorus removal using a microalgal biofilm in a new biofilm photobioreactor for tertiary wastewater treatment a,*, Martin Trtı´lek b, Toma s Rataj a,1 Katerina Sukacova Global Change Research Centre, Academy of Sciences of the Czech Republic, Belidla 986/4a, Brno 603 00, Czech Republic b sov 470, Czech Republic Photon Systems Instruments, 664 24 Dra a
article info
abstract
Article history:
Eutrophication of surface water has been an important environmental issue for nearly half
Received 13 June 2014
a century. High concentrations of phosphorus contribute to the process of eutrophication,
Received in revised form
resulting in the demand for effective and economic methods of phosphorus removal from
4 November 2014
treated water. The aim of this study was to evaluate the capacity for phosphorus removal
Accepted 29 December 2014
of a microalgal biofilm during different light regimes. The photobioreactor was operated for
Available online 7 January 2015
nine months each year over a two-year period without interruption and without any need of re-inoculation. The algal biofilm was able to remove 97 ± 1% of total phosphorus from
Keywords:
wastewater during 24 h of continuous artificial illumination. The average TP uptake rate in
Microalgal biofilm
our experiments was 0.16 ± 0.008 g m2 d1. Phosphorus removal values ranged from 36 to
Phosphorus removal
41% when the algal biofilm was illuminated by natural light (12 h sunlighte12 h night). The
Wastewater treatment
biomass production rate was 12.21 ± 10 g dry weight m2 d1 in experiments with continuous artificial light and 5.6 ± 1 g dry weight (DW) m2 d1 in experiments with natural light. These results indicate the great potential of microalgal biofilms in the tertiary treatment of wastewater. © 2015 Elsevier Ltd. All rights reserved.
1.
Introduction
Degradation of water ecosystems is a major problem in most countries of the world. Industrial and agricultural manufacturing is responsible for a huge production of wastewater, which must be treated to prevent pollution of water ecosystems (Omernik, 1977; Schlesinger, 1991; Vitousek et al., 1997). High concentration of phosphorus starts a process of eutrophication in the water environment. Dodds et al.
* Corresponding author. Tel.: þ420 511 440 038. ova ). E-mail address: [email protected] (K. Sukac 1 Tel.: þ420 511 440 038. http://dx.doi.org/10.1016/j.watres.2014.12.049 0043-1354/© 2015 Elsevier Ltd. All rights reserved.
(1997) concluded that maintenance of stream water total phosphorus concentration at <30 mg L1 would be necessary to keep benthic algal biomass below nuisance levels of 100 mg m2. Higher concentration of phosphorus leads to degradation of freshwater ecosystems and reduces the possibility of water reclamation. As a result, there is an urgent need for new, effective and economic methods of phosphorus removal. Currently, chemical methods are used most often for phosphorus removal, including co-precipitation with iron, alum or lime (Penetra et al., 1999).
56
w a t e r r e s e a r c h 7 1 ( 2 0 1 5 ) 5 5 e6 3
Biological methods of nutrient removal are considered the most environmentally favorable and least expensive of wastewater treatment methods, as there are no need of chemical precipitants (Mantzavinos and Kalogerakis, 2005). Biological removal of phosphorus is performed by different groups of microorganisms. Biological methods involve enhanced biological phosphorus removal (EBPR) which are based on polyphosphate accumulating organisms (PAOs). These microorganisms are able to store phosphate as intracellular polyphosphate in the specific conditions during wastewater treatment (De-Bashan and Bashan, 2004). The EBPR process is capable of efficient phosphorus removal performance. However disturbances and long periods of insufficient P removal have been observed at full-scale plants on numerous occasions (Oehmen et al., 2007). Other biological methods involve the use of algae and cyanobacteria for phosphorus removal (Rawat et al., 2011). Numerous studies have shown the ability of microalgae and cyanobacteria to grow in and reduce the nutrient content of wastewater (Chevalier et al., 2000; Doria et al., 2012; Renuka, 2013). This method of remediation is more environmentally sustainable because it does not generate additional waste such as sludge, it provides an opportunity for efficient recycling of nutrients, and it has the potential for sustainable productions of algal biofuels or bioactive compounds (Singh et al., 2005; Pittman et al., 2011). Microalgae and cyanobacteria are cultivated mainly in suspension (Christenson and Sims, 2011). However, algae are able to grow as biofilms on solid surfaces. Phototrophic biofilms are multi-layered community of photoautotrophs (cyanobacteria and microalgae) and heterotrophs (bacteria, fungi and protozoa) which play a key role in the self-purification of aquatic ecosystems (Sabater et al., 2002). In the case of biological phosphorus removal there have been numerous studies using algal biofilms successfully for the phosphorus removal under laboratory conditions (Guzzon et al., 2008; Johnson and Wen, 2010; Boelee et al., 2011; Liu et al., 2013; Shi et al., 2014). However, little data have been published concerning the capacity of microalgal biofilms for phosphorus removal in the large scale cultivation. Previously, the attached benthic algae were used in algal turf scrubber units (ATS) for the removal of nitrogen and phosphorus from different type of wastewater such as dairy manure (Kebede-Westhead et al., 2003) or domestic sewage (Craggs et al., 1996). ATS technology has been tested mainly in the southern regions of the USA, especially in California and Florida (Adey et al., 2011). The use of ATS for water quality improvement is regarded as an established practise (Adey and Loveland, 2007). As compared with ATS technology the application of phototrophic biofilms in tertiary wastewater treatment is still uncommon (Guzzon et al., 2008) and needs further research to develop effective technology for wastewater treatment. The current study has two aims: (a) to evaluate the capacity of an algal biofilm for phosphorus removal under light conditions consistent with the temperate climate of Central Europe; and (b) to estimate the comprehensive benefit of the presented technology, including biomass production and the assessment of harvested biomass as a source for biogas.
2.
Materials and methods
2.1.
Horizontal Flat Panel photobioreactor
A new large scale experimental photobioreactor was designed and built for the study of biofilms (Fig. 1). The Horizontal Flat Panel (HFP) photobioreactor for algal biofilm cultivation was developed by Photon Systems Instruments (Brno, Czech Republic) and consisted of two modules. One module was a concrete slab (1 2 0.02 m) supported by a metal frame which formed a 2 m2 cultivation area for biofilm growth. Wastewater was pumped from a reservoir (0.2 m3, with operational volume of 0.1 m3) by a magnetically powered centrifugal pump (IDRA 231 720, Sicce S.p.A., Italy) to the upper part of the cultivation area. Wastewater flowed over the cultivation area in a thin layer and then back to the reservoir. The flow rate of water pumped from the reservoir to the cultivation area was 8.8 L min1, with a laminar flow velocity of about 0.25 m s1 over the cultivation area. The circulating volume of wastewater was 100 L, with a retention time in the reservoir of approximately 7 min. The HFP photobioreactor (two modules) had a total cultivation area of 4 m2, and was placed in a greenhouse. Photosynthetically active radiation (PAR) was measured with a Quantum Sensor (EMS 12, EMS Brno, Czech Republic), inside the greenhouse. The average daily sum of irradiance was calculated from actual PAR values and expressed in MJ m2 d1. The temperature of the circulating wastewater was recorded regularly and varied between 19 and 24 C.
2.2.
Microalgal biofilm sampling and cultivation
An algal biofilm assemblage was sampled from the pools of a rendering plant (Glogow, Poland, 51 370 39.45600 N, 16 90 47.83600 E) to obtain a community with tolerance to high nutrient concentrations. The algal samples were taken from the mud surface of sludge pools and put into plastic bottles and stored in a cool box for transportation to the laboratory. Algal biofilm sheets were put into Bold's Basal medium (Bischoff and Bold, 1963) in several Erlenmeyer flasks to obtain sufficient volume of algal biomass for algal biofilm cultivation. The algal assemblage was cultivated under 30 mmol photons m2 s1 at 25 C for two weeks, then the species composition of algal assemblage in each Erlenmeyer flask was determined. The identification of microalgae and cyanobacteria was performed
Fig. 1 e Scheme of the experimental setup.
57
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Table 1 e Taxonomical analyses of the species present in biofilm in the season 2012 and 2013. Species
Inoculum used for
Start of experiments
2012
2012
2013
2013
HFP-photobioreactor
25 April 2012
16 August
29 October
25 April
14 August
Rare Dominant Rare
Dominant Dominant x
Dominant Dominant Rare
Dominant Dominant Rare
Dominant Dominant Rare
Dominant Dominant Rare
Dominant Dominant
Dominant x
Dominant x
Dominant x
Dominant x
Dominant x
Rare
Rare
Rare
Rare
Rare
Rare
Cyanobacteria Phormidium autumnale Pseudanabaena sp. Chrococcus sp. Coccal green algae Scenedesmus acutus Monoraphidium contortum Diatoms Cymbella minuta
rek and Anagnostidis (2005) and Ettl and according to Koma € rtner (1995) using an Olympus CX 31 microscope. The Ga microalgal assemblage with filamentous cyanobacteria and non-motile unicellular green algae was selected for following cultivation. The inoculum for the HFP-photobioreactor was prepared in a laboratory using the same design planned for a large scale system. A concrete slab (0.2 0.3 m) was placed in a plastic box with cultivation medium, and the medium was pumped to the cultivation area where it flowed over the surface of the slab. The selected microalgal assemblage was inoculated onto the surface of the slab where it formed a uniform film. The species composition of microalgal assemblage used for inoculation of the HFP-photobioreactor is given in Table 1. Inoculation of the HFP-photobioreactor was performed in a similar manner. An algal suspension with a biomass density of about 25 g DW L1 was applied to the cultivation area before starting the photobioreactor operation and pumping wastewater across the slab. The Algal biofilm was cultivated under sunlight (4e8 MJ m2 d1). The period of sunshine was approximately 14 h per day. The firm algal biofilm covered the whole cultivation area after 3 weeks.
2.3.
Experimental setup
Experiments were started when the algal biofilm covered the whole cultivation area. Performed experiments and experimental conditions are described in Table 2. Initially, batch
experiments were performed to identify TP removal during a five day period. Hydraulic retention time for continuous experiments was determined according to the results of batch experiments.
2.3.1.
Batch experiments
Batch experiments were conducted in two periods with markedly different light conditions. Daily values of irradiance ranged between 7 and 11 MJ m2 d1 in summer. The range of daily irradiance was between 2 and 10 MJ m2 d1 in autumn and spring. Batch experiments lasted five days and involved artificial wastewater.
2.3.2.
Continuous experiments
Continuous experiments were performed over four days repeatedly using two regimes: (a) During continuous experiments A and B, the algal biofilms were illuminated continuously by high pressure sodium lamps (Osram NAV-T Super 4Y e 600W) at an intensity of 90 ± 15 mmol photons m2 s1 and with a daily sum of irradiance of 1.7 MJ m2 d1. Artificial wastewater was used during experiments. (b) Continuous experiments C and D were performed under solar irradiance using real wastewater. Solar irradiance and real wastewater were used to imitate real conditions in the wastewater treatment plant. Average daily sums of solar irradiance were 1.57 ± 0.88 MJ m2 d1
Table 2 e Summary of performed experiments and experimental conditions. Experiment
Type of light
Batch experiment Exp. with high values Sunlight of solar radiation Exp. with low values Sunlight of solar radiation Continual experiment Experiment A Artificial light Experiment B Artificial light Experiment C Sunlight Experiment D Sunlight Experiments over 24-h time periods Experiment 1 Artificial light Experiment 2 Sunlight
Light regime
Light intensity (mmol m2 s1)
DayeNight
0e1650
7e11
Artificial
DayeNight
0e970
2e10
Artificial
Continual Continual DayeNight DayeNight
90 ± 15 90 ± 15 0e833 0e904
1.7 1.7 1.57 ± 0.88 3.1 ± 0.64
Artificial Artificial Real Real
Continual DayeNight
90 ± 15 0e815
1.7 1.73 ± 0.47
Artificial Real
Aver. daily sum of irradiance (MJ m2 d1)
Wastewater type
58
w a t e r r e s e a r c h 7 1 ( 2 0 1 5 ) 5 5 e6 3
(exp. C) and 3.1 ± 0.64 MJ m2 d1 (exp. D). The period of sunshine was approximately 12 h per day.
2.3.3.
Experiments over 24-h time periods
Experiments were carried out in the continual light regime and also in the dayenight light regime using both artificial and real wastewater. Experiments were run using a batch set-up. TP concentration and pH were measured every two hours to evaluate the influence of a diurnal light cycle on phosphorus removal.
2.3.4.
Wastewater characteristics
Artificial wastewater was constituted with total phosphorus (TP) concentration of 3 mg L1, typical of wastewater effluent from a municipal wastewater treatment plant that does not practice enhanced phosphorus removal (Water Environment Federation, 2010). The artificial wastewater was enriched by major and trace elements: 120 mg L1 NaNO3; 6 mg L1 MgSO4$7H2O; 2 mg L1 CaCl2$2H2O; 2 mg L1 NaCl; 6 mg L1 K2HPO4; 14 mg L1 KH2PO4; 4 mg L1 EDTA$2H20; 0.4 mg L1 FeSO4$7H2O; 0.9136 mg L1 H3BO3; 0.7056 mg L1 ZnSO4$7H2O; 0.1152 mg L1 MnCl2$4H2O; 0.1936 mg L1 Na2MoO4$2H2O; 0.1256 mg L1 CuSO4$5H2O; 0.0392 mg L1 Co(NO3)2$6H2O. Real wastewater was collected in the effluent from a sov (3500 Population wastewater treatment plant in Dra Equivalent), Czech Republic. Chemical parameters of real wastewater are given in Table 3.
2.4.
Analytical methods
Samples of wastewater were taken every day during batch and continuous experiments. The samples for phosphorus analysis were filtered through a 0.45 mm membrane filter (ME 25 e mixed cellulose ester, Whatman). TP concentration was measured using an ammonium molybdate spectrophotometric method (ISO 6878: 2004). Algal biomass was sampled from a defined area (9 cm2). Biofilm biomass was dried on pre-weighed filters (Whatmann, GFC filters) to a constant weight at 90 C, then the filters were weighed. Biomass production was expressed as g DW m2. Most of the biomass was harvested by squeegee after finishing the experiments. Harvested biomass was dried and used for anaerobic digestion tests. The rest of the algal biomass left on the cultivation area served for renewal of the algal biofilm. New experiments were conducted immediately after harvesting. The analysis of biomass distribution over the cultivation area was carried out at the end of the experimental season. One hundred samples of biofilm biomass were taken from the cultivation area. Each sample had an area of 9 cm2. The sampled areas were equally spread over the cultivation area.
Table 3 e Chemical characteristic of real wastewater. Chemical parameter Total phosphorus Total nitrogen NO3eN NO2eN NH4eN
Concentration (mg L1) 2.9 ± 0.1 49.4 ± 3 48.6 ± 2 0.23 ± 0.02 0.33 ± 0.03
2.5.
Anaerobic digestion processes
Anaerobic digestion tests were performed as described in Samson and LeDuy (1986). Adapted sludge was taken from the large anaerobic digester (second stage of fermentation pro c , Czech Republic). The adapted cess) in the biogas station (Cej sludge was placed in laboratory digesters. 15 g of dried biofilm biomass was added to the sludge. Tests were carried out in triplicate for two week periods in mesophilic temperature conditions (41 C). The total amount of produced gas and methane content were measured daily by gas analysis with €ger X-am® 7 000, infrared and electrochemical sensor (Dra € ger Safety AG & Co. KGaA, Germany). Dra
3.
Results
3.1.
Microalgal biofilm cultivation
The HFP photobioreactor was operated in Brno in the southern Czech Republic over a 9 month period for two years (AprileDecember 2012, AprileDecember 2013). After inoculation the algal biofilm grew across the entire slab surface within 3 weeks. Biomass ranged from 30 to 70 g DW m2 on the cultivation area. Nearly 50% of the cultivation area was covered with 40e50 g DW m2, with 30e40 g DW m2 over 26% of the area and 50e60 g DW m2 over 20% of the area. 5% of the area was covered with 60e70 g DW m2 of biofilm. The cyanobacterialemicroalgal assemblage constituted mostly of a mat created by the filamentous cyanobacteria Phormidium autumnale and Pseudanabaena sp., as well as the unicellular green alga Scenedesmus acutus which was distributed among cyanobacterial filaments. These dominant species were accompanied by Cymbella minuta (diatom) and Chroococcus sp. (unicellular cyanobacteria). The species composition did not change substantially (Table 1).
3.2.
Phosphorus removal and uptake
3.2.1.
Batch experiments
Fig. 2 shows the reduction of TP concentration during batch experiments. In the high solar radiation period (7e11 MJ m2 d1) the algal biofilm removed 92 ± 7% of TP with an average value of TP uptake rate of 0.14 ± 0.011 g m2 d1. This value was achieved by 24 h. In the low solar irradiance period, 50 ± 16% of TP removal was obtained after 24 h. The percentage gradually increased, attaining 81 ± 20% after 72 h. At the end of the experiments (96 h), values of TP removal were 74 ± 15%. The maximum TP uptake rate was 0.069 ± 0.024 g m2 d1 by 24 h during the period of low solar radiation. Maximum TP uptake rate was observed after 24 h of the experiment starting. During the following two days, the minimum values of TP uptake rate were 0.022 ± 0.007 g m2 d1 (48 h) and 0.025 ± 0.035 g m2 d1 (72 h).
3.2.2.
Continuous experiments
Under continual illumination the algal biofilm removed 97 ± 1% of TP from wastewater within 24 h. The average influent value 3.25 ± 0.16 mg TP L1 dropped to 0.082 ± 0.032 mg TP L1 after 24 h in the case of experiments
w a t e r r e s e a r c h 7 1 ( 2 0 1 5 ) 5 5 e6 3
59
concentration dropped only slightly. An increase in pH was detected with reduction of TP concentration, the highest changes in pH level coinciding with maximum reduction rate of TP. A maximal pH value of 9.98 ± 0.6 was measured after 24 h. The process of phosphorus removal evaluated during experiments with dayenight light regime (Figs. 4 and 5) was markedly different than in other experiments. During the diurnal cycle there were two phases of TP removal. Nearly 99% of TP was removed during the light phase (time period 8 h), during which pH values rose to a maximum value 10.17. Declining pH values were measured during the dark phase, during which TP concentration increased from 0.04 mg L1 to a maximum value of 1.48 mg L1 over a time period 12 h. Fig. 2 e Reduction of total phosphorus concentration during batch experiments.
with continuous light. The average TP uptake rate was 0.16 ± 0.008 g m2 d1. Phosphorus removal values were lower in the experiments in which solar irradiance was used for illumination of the algal biofilm. The influent value of 2.96 ± 0.022 dropped to only 1.76 ± 0.67 mg TP L1, and only 41 ± 11% of TP was removed during experiment C. In experiment D, the average influent concentration of 2.3 ± 0.06 decreased to 1.6 ± 0.3 and the algal biofilm removed 36 ± 9% of TP after 24 h. The average values of TP uptake rate were 0.06 ± 0.016 g m2 d1 (exp. C) and 0.042 ± 0.01 g m2 d1 (exp. D).
3.2.3.
Experiments over 24-h
Fig. 3 illustrates results of experiments over a 24-h time period using continuous light. After 18 h the TP concentration decreased from 3.0 ± 0.17 mg L1 to 0.09 ± 0.014 mg L1, the algal biofilm removing 97% of TP from the wastewater. The maximum reduction rate of TP was reached during the 8-h period at the start of the experiment, after which the TP
Fig. 3 e Diurnal course of pH values and total phosphorus concentration during experiments over 24-h time period with continual illumination.
3.3.
Biomass production and anaerobic digestion
Measured values of biomass production rate varied enormously during experiments. Markedly higher biomass production was measured in experiments with continuous illumination. The average value was 12.21 ± 10 g DW m2 d1. Lower values of 5.6 ± 1.4 g DW m2 d1 were measured in the experiments using solar irradiance with a dayenight light regime. A biomass production rate of 9.8 ± 5 g DW m2 d1 was measured during batch experiments. Harvested biomass was assessed for potential biogas production. During anaerobic digestion of dried biomass, the biogas yield reached 0.33 ± 0.03 m3 kg1 VS (Volatile Solids) added. Methane production was measured at 0.153 ± 0.015 m3 kg1 VS. Average methane content was measured at 46% in the yield biogas, with a maximum content measured at 66% after 10 days of anaerobic digestion. Maximum TP uptake capacity was used to estimate the required area of an algal biofilm for tertiary wastewater treatment in a plant with a capacity from 2000 to 6000 population equivalent (PE). The estimated area was calculated to be in range from 0.3 ha (2000 PE) to 1 ha (6000 PE), assuming a reduction of TP concentration from 3 mg TP L1 to 0.09 mg TP L1. Table 4 shows the production potentials associated with algal biofilm biomass yield per treatment of 1 m3 of
Fig. 4 e Diurnal course of pH values and total phosphorus concentration during experiments over 24-h time period with dayenight light regime.
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Fig. 5 e Diurnal course of total phosphorus concentration and the light intensity in the experiment C.
wastewater. Dry biomass production of 0.13 kg DW would be obtained from 1 m3 treated wastewater. Biogas and biomethane yield would be 0.043 and 0.03 m3 per m3 of wastewater, respectively. It was assumed that biogas contains 66% of methane. A power yield 1.13 MJ could be obtained during treatment of 1 m3 of wastewater, assuming energy content of CH4 to be 37.8 MJ m3 (Prajapati et al., 2013). Annual dry biomass production was estimated at 44.4 t ha1 y1.
4.
Discussion
A new type of photobioreactor for algal biofilm cultivation was tested and validated for technical design and functionality. An algal biofilm was cultivated and weekly harvested for nearly two years after initial inoculation without interruption and without the need for re-inoculation. Similarly Sandefur et al. (2011) described a nine-month operating period for an algal turf scrubber system, and Mulbry et al. (2008) reported on algal turf scrubber raceways that were operated for approximately 270 days each year between 2003 and 2006 in central Maryland. This study showed that biofilm activity decreased TP concentrations in all the experiments, with particularly high efficiency of phosphorus removal achieved during experiments with a continuous light regime. We measured an average TP 2 1 d uptake rate about 20% higher than the 0.13 g PO3 4 eP m reported by Boelee et al. (2011) during laboratory experiments
under continual illumination 230 mE m2 s1. Although a lower intensity of illumination (90 ± 15 mE m2 s1) was used in the current study, a higher efficiency of phosphorus removal was attained. We assume these results were due to using a different design of the algal biofilm system or because there was a different species composition in the algal biofilm. The Boelee et al. (2011) biofilm consisted mainly of pennate diatoms and green filamentous algae, whereas in our study the microalgal assemblage consisted of a net of filamentous cyanobacteria with unicellular green algae interspersed. The phosphorus uptake difference could be due to the presence of more cyanobacteria and coccal green algae. In both cyanobacteria and coccal green algae, phosphate can be accumulated as polyphosphate granules (Jansson, 1988; Guzzon et al., 2008). The efficiency of phosphorus removal was less in experiments with a dayenight light regime and using solar radiation. Light is one of the most important factor influencing the uptake of phosphate by algae, because photosynthesis is et al., 1997). Phosphate uptake by depended on light (Laliberte cyanobacteria is usually greater in the light than in the darkness (Whitton, 1992). During this study TP removal values varied between 36 and 41%. Sandefur et al. (2011) reported an average value of TP removal of 48% for the algal turf scrubber system under the warm climate of Arkansas, USA. Boelee et al. (2014a) reported an average removal efficiency of PO3 4 eP of 14% for an outdoor phototrophic biofilm reactor operated in the cooler, lower light, conditions in the Netherlands. Placement of our bioreactor in the greenhouse attenuated the impact of low temperature and other weather events in the Czech Republic, and allowed TP removal only 7e12% lower than measured in the warmer, sunnier climate of Arkansas. Diurnal changes of phosphorus concentration and pH values were observed during experiments over a 24 h period. Values of pH rose significantly during the light period. Similar pH shifts have been reported during phosphorus uptake by et al., 1997; numerous authors (Chevalier et al., 2000; Laliberte Sandefur et al., 2011). The pH shift corresponds with the absorption of CO2 during high photosynthetic activity (Small and Adey, 2001). Algae remove phosphorus from wastewater by processes of adsorption, assimilation and precipitation (Craggs et al., 1996). The precipitation of phosphorus with cations (such as Ca2þ and Mg2þ) at high pH is known to begin between pH 8.9e9.5 (Belsare and Belsare, 1987). Our results indicate that
Table 4 e Estimated production potential of algal biofilm wastewater treatment process. Process
Value
Dry biomass production (kg DW m3 treated wastewater) Biogas yield (m3 biogas m3 treated wastewater) Biomethane yield (m3 biomethane m3 treated wastewater) Estimated power yield (MJ m3 treated wastewater) Dry biomass production (kg dry weight ha1 y1) Biogas yield (m3 biogas ha1 y1) Biomethane yield (m3 biomethane ha1 y1) Estimated power yield (GJ ha1 y1)
0.13
Expressed to 1 m3 treated wastewater
Remarks
0.043 0.03
Expressed to 1 m3 treated wastewater 66% CH4 in biogas, Expressed to 1 m3 treated wastewater
1.13 44 408 14 655 9770 0.37
CH4 energy content 37.9 MJ m3, Expressed to 1 m3 treated wastewater Estimated annual production 66% CH4 in biogas, Estimated annual production Estimated annual production CH4 energy content 37.9 MJ m3, Estimated annual production
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TP removal depends on both absorption and precipitation, as indicated by the decrease of pH values, and simultaneous increase of TP concentration, during the night, indicating dissolution of precipitated phosphorus. This process was also reported by Craggs et al. (1996) for the algal turf scrubber system. We assumed that the 58% of TP returned to the water during night corresponded with the proportion of phosphorus which had been removed by precipitation. The rest of the TP (41%) would be the proportion removed by assimilation (include luxury uptake) of the algal biofilm. Boelee et al. (2014b) reported increased content of phosphorus (P), magnesium (Mg) and calcium (Ca) in the algal biofilm during the P-removal process, and assumed that this indicated precipitation of Ca and Mg phosphates in the biofilm. A reduction of precipitated phosphorus dissolution during the night would result in a more effective process of phosphorus removal. One option is the application of a continuous light regime. Exposure to sunlight during day and to artificial light during the night would increase the efficiency of the whole process. In our study, observed values of arial biomass production rate varied considerably during experiments. Biomass over the cultivation area ranged from 30 to 70 g DW m2. This high variability could have been caused by differences in the dispersion of biomass over the large cultivation area (4 m2). The degree of variability was also affected by cultivation conditions, since homogenous cultivation conditions are almost impossible to maintain over such a large cultivation area (4 m2). Boelee et al. (2014) also reported excessively variable values of arial biomass production during cultivation of the algal biofilm. Measured values of arial biomass production were similar to results of Boelee et al. (2011) who reported a maximum biomass production of 7.7 g DW m2 d1. However, our values are higher than the biomass production of 2.57 g DW m2 d1 reported for an attached microalgal growth system using polystyrene foam as a supporting material (Johnson and Wen, 2010). On the other hand, algal biomass production reported for an algal turf scrubber system at 25 g DW m2 d1 (Mulbry et al., 2008) were more than double the values measured during this study. Sandefur et al. (2011) reported 31 ± 19 g DW m2 d1. However, both studies were conducted under warmer climate conditions. Harvested biomass was assessed for potential of biogas production. Measured biogas production was similar to that reported in other studies. Prajapati et al. (2014) measured biogas production in a range from 0.34 to 0.464 m3 kg1 VS for different Chlorella species, which is similar to our values of 0.33 m3 kg1 VS. Mussgnug et al. (2010) investigated anaerobic digestion for different algal species and measured maximal biogas yield of 0.587 m3 kg1 VS for Chlamydomonas rheinhardtii, and minimal yield of 0.287 for Scenedesmus obliquus. In our study, the average methane content was low compared with other studies (e.g. Prajapati et al., 2013), although the maximal achieved methane content of 66% corresponds with the finding of other researchers (e.g. Sialve et al., 2009). The low average methane content in the biogas resulted from the short time period of the digestion test, lasting only 15 days. Maximal methane content was observed after 10 days of the test. We
61
would expect a higher methane yield following a longer anaerobic digestion process. The annual calculated dry biomass production in our study is about 30% more than that reported by Prajapati et al. (2014) for Chlorella pyrenoidosa, and that reported by Chinnasamy et al. (2010) for algal assemblage cultivated in carpet industry wastewaters. These data indicate that cyanobacteriaealgae assemblages immobilized in a biofilm can produce similar amount of biomass to algae cultivated in suspension. Moreover, biofilm cultivation allows for simple and low cost harvesting protocols. The annual potential energy generation of the harvested biofilm is estimated about 369 GJ ha1 y1, enough for warming the greenhouse and operation of the bioreactor. Anaerobic digestion plays an important role in the disposal of activated waste sludge, reducing treatment costs of wastewater plants, and their environmental footprint. The process is considered a major and essential part of a modern wastewater treatment plant (Appels et al., 2008). Combining wastewater treatment facilities with biogas production processes is an exciting possibility for wastewater management.
5.
Conclusions
This study demonstrates the ability of an algal biofilm to remove phosphorus from municipal wastewater. An algal biofilm was able to remove 97 ± 1% of total phosphorus from wastewater after 24 h under artificial continuous illumination. Phosphorus removal values ranged from 36 to 41% when the algal biofilm was illuminated by solar irradiance in a 12 h lighte12 h dark regime. pH values in wastewater decreased in the dark phase of the experiments, associated with CO2 release and dissolution of precipitated phosphorus. The results of experiments indicated that efficiency of P removal could be increased by imposing a regime of sunlight during the day and artificial light during the night. Measured biogas yield showed the economic potential to integrate algal biofilm wastewater treatment with further anaerobic digestion in small and medium wastewater treatment plants.
Acknowledgments This publication is an output of the CzechGlobe Centre that is being developed within the OP RDI and co-financed from EU funds and the State Budget of the Czech Republic (Project: CzechGlobe e Centre for Global Climate Change Impacts Studies), Reg. No. CZ.1.05/1.1.00/02.0073. The authors like to thank Dr. Tomas Vitez for anaerobic digestion of algal rkova for critically reading the biomass, Dr. Jaroslava Koma manuscript and Dr. Stephen Hunt (Qubit Systems Inc., Kingston, Canada) for reviewing the manuscript.
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