Bioresource Technology 161 (2014) 402–409

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NaCl as an effective inducer for lipid accumulation in freshwater microalgae Desmodesmus abundans Ling Xia a,b, Junfeng Rong c, Haijian Yang a,b, Qiaoning He a,b, Delu Zhang d, Chunxiang Hu a,⇑ a

Key Laboratory of Algal Biology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, China University of Chinese Academy of Sciences, Beijing 100049, China c SINOPEC Research Institute of Petroleum Processing, Beijing 100083, China d Department of Biological Science and Biotechnology, Wuhan University of Technology, Wuhan 430070, China b

h i g h l i g h t s  0.25 g L

1

urea is the optima for biomass accumulation in D. abundans.

 NaCl was outstanding of different salts (NaCl, NaHCO3, NaAc and NaS2O3) on lipid induction.  Higher lipid productivity was obtained during the optimized cultivation process.  1.79 g L

1

of biomass concentration was the time point for NaCl addition.

 Strategy of NaCl addition for cultivating D. abundans was cost-efficient and net energy-positive.

a r t i c l e

i n f o

Article history: Received 25 November 2013 Received in revised form 11 March 2014 Accepted 14 March 2014 Available online 24 March 2014 Keywords: Desmodesmus abundans NaCl Lipid induction Net energy ratio Cost feasibility

a b s t r a c t In order to evaluate the efficiency and potential of salt addition-based two-stage cultivation technology, on the basis of urea as nitrogen source, we compared four types of salts (NaCl, NaHCO3, NaS2O3 and NaAc) as inducers for lipid production in Desmodesmus abundans. The maximum biomass productivity (270.08 mg L1 d1) was obtained by using 0.25 g L1 urea. The highest lipid productivity (67.08 mg L1 d1) and better biodiesel quality were realized by addition of 20 g L1 NaCl, and the optimal time point for salt addition was determined at 1.79 g L1 of biomass density. Further cost analysis demonstrated this cultivation process was relatively economical. Above results suggest that NaCl addition is an economical and applicable strategy for lipid enhancement and can be extended for microalgae-based biodiesel production. Ó 2014 Elsevier Ltd. All rights reserved.

1. Introduction Biodiesel, the mixture of fatty acid methyl esters (FAMEs), has recently received a great deal of attentions due to its environmental benefits and the fact that it is made from renewable resources. Microalgae are regarded as the feedstock of future for sustainable biodiesel production because they have higher photosynthetic efficiencies, enhanced lipid production and faster growth rates compare to conventional terrestrial plants without compromising landmass (Chisti, 2007). Nevertheless, the microalgal biodiesel has not been widely commercialized mainly due to its higher production costs (Chisti, 2007; Sun et al., 2011). In order to reduce the total cost of microalgal biodiesel production, increase of the

⇑ Corresponding author. Tel./fax: +86 27 68780866. E-mail address: [email protected] (C. Hu). http://dx.doi.org/10.1016/j.biortech.2014.03.063 0960-8524/Ó 2014 Elsevier Ltd. All rights reserved.

productivity of biomass and lipids is one of the efficient methods (Delrue et al., 2012). The lipid content of microalgae can be increased when the cells are subjected to nutrient imbalances or culturing stress imposed by chemical or physical stimuli (Mus et al., 2013). However, high lipid contents produced under such conditions usually associated with relatively low biomass productivity, and thus low overall lipid productivity. Hence, in order to overcome the contradiction between biomass yield and lipid production, a culture mode that allows an optimum growth rate and permits a lipid enhancement is necessary. Thus, two-stage strategies were envisaged, which separate biomass and lipid boost by applying appropriate conditions in each phase of culture to improve overall lipid productivity (Su et al., 2011; Mujtaba et al., 2012; Xia et al., 2013). Accordingly, in the first stage optimal conditions best suited for highly concentrated biomass production are provided, followed by necessitating medium with stressful conditions to induce lipid biosynthesis.

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Our previous study has developed a two-stage cultivation process, which composed of optimization culture in stage I and followed by directly adding NaCl into the medium for lipid induction in stage II (Xia et al., 2013). In microalgal biomass production of stage I, nitrogen was regarded as the most important limiting factor (Richmond, 2004). Typically, algae are capable of utilizing nitrite, ammonia and urea. Among these nitrogen sources, urea gained important favor generally in large-scale algal cultivation, because of its universal availability and relatively low cost (Hsieh and Wu, 2009). Moreover, urea also exhibited favorable effect on algal growth in green algae (Hsieh and Wu, 2009; Arumugam et al., 2013). Therefore, urea is used for culturing Desmodesmus abundans in this study. As to the stage II, the use of other sodium salts has also been reported previously to enhance lipid accumulation of green algae such as sodium bicarbonate (NaHCO3) (Gardner et al., 2012), sodium acetate (NaAc) (Heredia-Arroyo et al., 2011), sodium thiosulphate (NaS2O3) (Mandal and Mallick, 2009) and so on, but the comparisons of the effects on lipid accumulation between these salts on certain species has not been reported yet. We contrasted salt-induced lipid accumulation in D. abundans with addition of NaCl, NaHCO3, NaAc and NaS2O3, and chose the best stimuli for rapid lipid accumulation in stage II in this study. To further enhance the feasibility and practicality of this salt addition-based two-stage cultivation process, it is important to undertake the energy analysis and cost estimation. In this study, we first evaluated the cost and energy feasibility of this two-stage process for D. abundans in a large-scale photobioreactor. The aim of this investigation was to select the best stimuli with low cost and high efficiency on lipid accumulation for the salt addition-based two-stage culture mode. In pre-experiments the optimum urea concentration in terms of fast growth was determined. Following this, in a second set of experiments the most suitable stimuli in terms of lipid induction was selected. And then the optimized culture conditions were used in the large-scale bioreactor for cultivating D. abundans. Finally, the improvement of the fatty acids composition in terms of biodiesel quality and the economic feasibility were also studied as critical factors for the entire process evaluation. 2. Methods

(mixing at 150 rpm) at the middle of the reactor. 2% CO2 in air was supplied to each bioreactor during the daytime. In order to realize field conditions, no particular effort was made to maintain an axenic culture for algal cultivation. The medium was thoroughly compounded with tap water. For urea concentration optimization, the urea concentration settings were 0.10, 0.15, 0.20, 0.25 g L1. For lipid induction tests, different concentrations of sodium salts were added into the culture separately when cultures reached the late exponential phase at a biomass concentration of about 1.80 g L1. For initial concentration optimization, the pre-cultured biomass from the late exponential phase was centrifugally harvested and the supernatants were saved. All reactors were inoculated simultaneously with biomass concentration between 1.19 and 2.51 g L1 using the gathered supernatants as medium and at the same time adding 20 g L1 NaCl for each test. The experiments were conducted during May to August in 2012; the air temperature was 31.44 ± 5.80 °C during day time from 6:00 AM to 18:00 PM and 24.08 ± 2.79 °C during night from 18:00 PM to 6:00 AM. 2.3. Analytical procedures 2.3.1. Biomass measurement The cells were harvested by centrifugation and lyophilized using a vacuum freeze dryer (Alpha 1-2 LD plus, Christ). Generally, the dry cell weight (DCW) of microalgae is correlated to the optical density (OD) at certain wavelength from 450 to 680 nm (Ji et al., 2013). In this study, the biomass concentration (BC, mg L1) of D. abundans was determined by measuring the optical density of 680 nm (OD680) via an ultraviolet photospectrometer. The result was converted to DCW concentration using the calibration curve relating OD680 as following Eq. (1):

BC ¼ 320  OD680

ðR2 ¼ 0:996Þ

ð1Þ 1

The biomass productivity (BP, mg L according to Eq. (2):

BP ¼ ðB2  B1 Þ=T

d

1

) was calculated

ð2Þ

where B2 and B1 represents the dry weight biomass density at the time T (days) and at the start of the experiment, respectively.

2.1. Microalgal strain and culture medium The freshwater algae species used in this study was pure culture of D. abundans provided by Institute of Hydrobiology, the Chinese Academy of Sciences. The stock culture was maintained indoors in a sterilized BG11 medium containing 1.5 g NaNO3, 40 mg K2HPO4, 75 mg MgSO47H2O, 20 mg Na2CO3, 36 mg CaCl22H2O, 6 mg ammonium citrate monohydrate, 6 mg ammonium ferric citrate, 1 mg EDTA, 2.86 lg H3BO3, 1.81 lg MnCl24H2O, 0.222 lg ZnSO47H2O, 0.39 lg Na2MoO42H2O, 0.079 lg CuSO45H2O, 0.050 lg CoCl26H2O in 1 L water. 2.2. Experimental setup All experiments were conducted in bioreactors (5-L flask bioreactor or 140-L airlift bag columns) in a green house in Beijing, China (40°220 N, 116°200 E) in summer. The photobioreactors setup was previously described in details in Xia et al. (2013). The 140-L bioreactor composed with two connected 70-L hanging bags (1.80 m height  0.22 m in diameter), with an aeration rate of 18 L min1 in each column for cell mixing. The 5-L bioreactor had the same bottom diameter of 0.22 m and a height of 0.37 m. Aeration in 5-L flask was at a flow rate of 4 L min1, and the reactor was stirred using a 5 cm magnetic stir bar

2.3.2. Lipid analysis The total lipid was gravimetrically quantified after extraction using a Soxhlet’s extractor with chloroform/methanol (2/1 v/v) as solvent and incubated at 90 °C for 4 h. The lipid productivity (LP, mg L1 d1) was determined according to Eq. (3):

LP ¼ BP  LC

ð3Þ

where LC denotes the total lipid content (w/dry weight biomass, %) at time T. Biodiesel was determined as fatty acid methyl esters (FAMEs) after acidic transesterification of lipids. Lipid sample was suspended in 1 M H2SO4–methanol (2 mL) in a vial. The vial was flushed with nitrogen to ensure an inert atmosphere before sealing and heated at 100 °C for 1 h in a water bathe. After methylation, purified water (0.20 mL) and n-hexane (0.60 mL) were added, the mixture centrifuged and the top hexane layer contained FAMEs collected for gas chromatograph mass spectrometry (GC–MS; Thermo Scientific ITQ 700™, USA) analysis (Xia et al., 2013). For analysis of the biodiesel property, the degree of unsaturation (DU), iodine value (IV), cetane number (CN) were determined by empirical equations based on fatty acids (FAs) composition as

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described by Song et al. (2013), while cold filter plugging point (CFPP) by Nascimento et al. (2013). 2.3.3. Net energy ratio The net energy ratio (NER) is defined as the ratio of the ‘‘Energy produced’’ over ‘‘Primary energy input’’ as the following Eq. (4): X X NER ¼ Energy producedðlipid or biomassÞ= Energy requirements ð4Þ

NER is estimated using the method detailed in Jorquera et al. (2010) and based on the data obtained in 140-L bioreactor for cultivating D. abundans for 1 year. 2.3.4. Production cost estimation The algal oil production cost is based on 1 year production in 140-L bioreactor for cultivating D. abundans and the price level in Beijing, 2013. The total cost is the sum of the fertilizer, electricity, fresh water, building and infrastructure for producing 1 kg of algal oil in the greenhouse.

Table 1 Biomass and lipid productivities of D.abundans cultivated under different urea concentrations. Urea concentration (g L1)

LC (%)

BC (g L1)

BP (mg L1 d1)

LP (mg L1 d1)

0.10 0.15 0.20 0.25 0.30

20.18 16.64 16.25 18.25 19.24

2.85 2.92 3.35 3.40 3.06

223.49 230.19 265.84 270.08 241.28

45.10 38.30 43.20 49.29 46.42

BP, biomass productivity; BC, biomass concentration; LC, lipid content; LP, lipid productivity; the same below.

influence cell growth and lipid production and choosing suitable nitrogen concentration for certain alga is important. Taken together, 0.25 g L1 urea was the optimal nitrogen concentration for D. abundans growth, and this optimal concentration was used for the subsequent tests.

3. Results and discussion

3.2. Salts supplementation-based lipid enhancement in stage II

3.1. Influence of initial urea concentration on biomass and lipid content in stage I

3.2.1. Influence of different salts on the lipid enhancement Sodium salts like chloride, thiosulphate, bicarbonate, acetate and so on, have been used to induce lipid accumulation in microalgae (Mandal and Mallick, 2009; Heredia-Arroyo et al., 2011; Gardner et al., 2012; Mus et al., 2013) and the present data is the first report based on a comparison of lipid accumulation capability between these salts. To choose the best salt as stimuli for lipid accumulation for the freshwater green algae D. abundans, lipid contents in cultures of different concentrations of NaCl, NaHCO3, NaAc and NaS2O3 addition were analyzed. For all reported comparisons, D. abundans grew on modified BG-11 medium with 0.25 g L1 urea without any salt addition were considered the control culture. The lipid contents listed are based on the assay of 3 or 6 days cultivation after different salts addition. As summarized in Table 2, in all cultivations the lipid content increased with incubation time. An increasing in NaCl concentrations led to an increase in the lipid content. And after 6 days treatment, the lipid content increased from 23.76% to 34.70% with the increasing concentrations of NaCl from 0 to 20 g L1, further increasing NaCl concentration reduced the lipid accumulation. Analogous to the NaCl supplement cultures, the Na2S2O3 cultures demonstrated a similar trend with a peak lipid content of 31.30% in the culture of 1.00 g L1 salt supplement at day 6. This lipid content was higher than that obtained in Scenedesmus obliquus, with a lipid content of 30.40% with 0.60 g L1 Na2S2O3 supplement (Mandal and Mallick, 2009). However, lipid accumulation associated with NaHCO3 was concentration dependent, with an increase in NaHCO3 concentration increasing the lipid content. Further increase in concentration of NaHCO3 resulted in extreme high alkaline pH value of above 11 in the medium, which is too high for the cells to tolerate and they mostly died shortly after NaHCO3 addition (Table 2). Thus, the maximum specific lipid content was obtained in the culture of 25 g L1 NaHCO3 supplement, with a content of 34.98%. This lipid content was 1.50 times higher against 24.00% obtained in Tetraselmis suecica and Chlorella sp. with inorganic carbon addition (Moheimani, 2012). Unfortunately, the lipid accumulation in cultures of NaAc addition was not as achievable as expected. And no obvious increase in lipid accumulation was observed relative to the control culture. It is worth noticing that of all the cultures, the highest lipid content was obtained in the culture amended with 25 g L1 NaHCO3 or 20 g L1 NaCl, with a content of almost 35% after 6 days treatment. This maximum lipid content was comparable and even higher than that obtained from

Urea was tested in five different concentrations viz., 0.10, 0.15, 0.20, 0.25 and 0.30 g L1 in order to standardize the optimum concentration for D. abundans growth. As shown in Fig. 1, after 12 days of cultivation, an increase in urea concentrations from 0.10 to 0.25 g L1 led to an increase in the biomass concentration, from 2.85 to 3.40 g L1, and biomass productivity from 223.49 to 270.08 mg L1 d1 (Table 1). Further increase in urea concentration of 0.30 g L1 led to a significant decline in the biomass concentration and biomass productivity (Table 1). Some literature reports have also suggested that nitrogen concentration might be inhibitive at high concentrations (Xu et al., 2001; Li et al., 2008; Arumugam et al., 2013). This may be due to the deleterious effect of nitrogen at higher concentrations (Arumugam et al., 2013). The lipid content of D. abundans varied with the level of urea concentration in the culture and showed no obvious trend as summarized in Table 1. However, the highest lipid content was observed in the culture with the lowest urea concentration. The result was consistent with most reports that nitrogen limitation could largely enhance the lipid synthesis in microalgae (Hu et al., 2008; Li et al., 2008; Hsieh and Wu, 2009). Nevertheless, the highest lipid productivity was obtained at the initial urea feed of 0.25 g L1 because of the highest algal biomass productivity (270.08 mg L1 d1) (Table 1). The results imply that different urea concentrations

Fig. 1. Growth curves of D. abundans at different initial urea concentrations.

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L. Xia et al. / Bioresource Technology 161 (2014) 402–409 Table 2 Lipid production of salts supplementation-based cultures in stage II. Salts

Concentration 1

(g L

)

Lipid content (%, w/w)

pH

(M)

3d

6d

Sodium chloride

0 10 20 30

0 0.17 0.34 0.51

21.67 ± 2.36 26.13 ± 0.75 28.79 ± 1.12 23.08 ± 1.58

23.76 ± 1.10 30.21 ± 0.34 34.59 ± 0.58 28.71 ± 3.83

8.67 ± 0.33 8.47 ± 0.45 8.28 ± 0.56 8.11 ± 0.61

Sodium thiosulphate

0 0.50 1.00 1.50

0 0.003 0.006 0.009

21.67 ± 2.36 23.32 ± 0.74 28.37 ± 0.52 25.59 ± 0.13

23.76 ± 1.10 27.58 ± 0.76 31.30 ± 1.12 27.75 ± 0.44

8.67 ± 0.33 8.41 ± 0.59 8.27 ± 0.73 8.24 ± 0.72

Sodium bicarbonate

0 8 15 25

0 0.10 0.18 0.30

19.34 ± 0.79 23.71 ± 0.41 25.59 ± 0.83 28.37 ± 2.31

22.15 ± 0.71 27.11 ± 0.31 32.72 ± 2.43 34.98 ± 0.03

8.61 ± 0.53 9.13 ± 0.67 9.43 ± 0.26 10.06 ± 0.38

Sodium acetate

0 2 4 6

0 0.024 0.049 0.073

19.34 ± 0.79 20.97 ± 1.46 24.89 ± 1.57 20.45 ± 3.46

22.15 ± 0.71 19.87 ± 2.64 19.04 ± 2.77 20.93 ± 1.32

8.61 ± 0.53 8.89 ± 0.32 8.92 ± 0.27 9.02 ± 0.13

Values are the means and standard deviation of two independent experiments.

Fig. 2. Time courses of biomass and lipid content of D. abundans during single- and two-stage cultivation using 20 g L1 NaCl or 25 g L1 NaHCO3. During single-stage cultivation, the microalgae were cultured in the optimal medium of 0.25 g L1 urea. In the two-stage cultivation process, microalgae grew under the same culture conditions as in the single-stage process, but were subjected to NaCl or NaHCO3 addition in stage II.

other Desmodesmus species under nitrogen-starvation or high saline conditions in indoor cultures (Pan et al., 2011). D. abundans depicted a profound rise in lipid yield under conditions of sodium chloride, sodium thiosulphate and sodium bicarbonate addition. The increase in lipid content in terms of the addition of NaCl, might relate to the enhancing production of

neutral and polar lipids, or particular triacyglycerols and glycerol, which could help the organisms to tolerate high salinity (Xia et al., 2013). While the mechanisms of the lipid-enhancing in terms of bicarbonate addition are based on a combination of alkaline pH stress and elevated dissolved inorganic carbon (DIC) (Table 2) (Mus et al., 2013). The possible reason for the rise of lipid pool in terms of the addition of thiosulphate could be that a reducing environment leads to increased pool size of NADH which favors lipid synthesis (Mandal and Mallick, 2009). However, acetate addition had no effect on lipid boost unlike the aforementioned salts. This may be due to severe contamination of bacteria in the varying environments in the greenhouse. On the other hand, D. abundans may also have difficulty in directly utilizing organic carbon source. Other salts such as ferric, chromic, cupric and zinc salts, have also been found to increase the lipid content in some microalgae (Salama et al., 2013), but they are impossible to be widely used due to the severe environmental hazard. Given the aforementioned results, lipid contents at 20 g L1 NaCl and 25 g L1 NaHCO3 were the highest and had no significant differences. So these two salts were selected for further study because they provided the highest lipid content. 3.2.2. Comparisons of lipid production capability and biodiesel quality between NaCl and NaHCO3 supplemented cultures To further determine the best stimuli for the lipid enhancement of D. abundans, time courses of D. abundans growth and lipid content under conditions of 25 g L1 NaHCO3 and 20 g L1 NaCl were analyzed and presented in Fig. 2; the growth and lipid parameters obtained are summarized in Table 3. The salts were added into the cultures when cells achieved a biomass concentration of 1.75 g L1 after 8 days cultivation when the cells stepped into the late exponential phase. After salts addition, a small part of cells died and

Table 3 Comparisons of biomass and lipid productivities between growth-associated lipid production and two-stage production of D. abundans. Salts

Time (d) LC (%, w/w) BP (mg L1 d1) LP (mg L1 d1)

Stage I

Stage I + Stage II

Control

Control

NaCl

NaHCO3

Control

NaCl

NaHCO3

8 19.82 ± 1.67 205.52 ± 7.81 40.73 ± 1.88

11 20.51 ± 1.37 189.74 ± 14.51 38.91 ± 0.37

11 32.37 ± 1.03 156.68 ± 18.84 50.72 ± 7.71

11 30.44 ± 0.35 164.70 ± 6.65 50.13 ± 1.44

14 23.02 ± 1.53 194.61 ± 6.52 44.80 ± 1.47

14 35.50 ± 1.06 163.38 ± 2.29 58.00 ± 2.55

14 33.44 ± 0.52 162.29 ± 3.83 54.27 ± 0.44

Values are the means and standard deviation of two independent experiments.

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resulted in a decrease in biomass concentration. While 3 days later (day 11), the cells recovered and the biomass concentration increased. In addition, both NaCl and NaHCO3 supplement cultures resulted in a decrease in biomass production but increase in lipid content compared to the control without any salt supplement. And after 6 days treatment (day 14), the lipid content achieved the highest values (Fig. 2). Nevertheless, the lipid productivity was much higher in the cultures of NaCl and NaHCO3 addition than that in the control without any salt supplement (Table 3). Relatively, the culture supplemented with NaCl accumulated more lipid than that with NaHCO3. During the whole experiment, the culture supplemented with NaCl attained the maximum lipid productivity compared with the control and the culture supplemented with NaHCO3, reaching the value of 58.00 mg L1 d1 at the end of the experiment (Table 3). This maximum lipid productivity obtained was 1.30 times higher against 44.80 mg L1 d1 in culture without salt supplement. The results highlight the importance of high salinity stress on lipid accumulation in freshwater green algae and further prove the feasibility of the two-phase cultivation mode for microalgal biodiesel production. Besides lipid productivity, FAME profile in terms of biodiesel quality is another significant factor determining the feasibility of this two-stage cultivation process using salt addition. Cells growing in the photobioreactor analyzed for their FA composition both under high concentration of NaCl and NaHCO3 at the end of the experiments. As shown in Table 4, the most commonly synthesized FAMEs were FAs with C16-C18, which are the main components of biodiesel. Palmitic acid (16:0), Oletic acid (18:1) and linolenic acid (18:3) were the most abundant FAs found in D. abundans. A remarkable increase of the relative content of monounsaturated FAs (C16:1 and C18:1) were found both under high salinity and high alkaline compared to the control without any salt addition. The relative content of polyunsaturated FAs (PUFA), especially linolenic acid (18:3), on the other hand decreased substantially. These results was consistent with the previous report in freshwater Chlamydomonas mexicana and S. obtusus XJ-15 under high salinity (Salama et al., 2013; Xia et al., 2013), and Neochloris oleoabundans under high alkaline (Santos et al., 2012). The changes in the FA profile in response to high salinity and high alkaline are inevitable to keep the membrane fluid and prevent its destruction. On the other hand, stress conditions alter fatty acid synthesis to produce more

Table 4 Fatty acid profile and estimated biodiesel properties from D. abundans oils during single- and two-stage cultivation subjected to NaCl or NaHCO3. Fatty acids Myristic acid (14:0) Palmitic acid (16:0) Palmitoleic acid (16:1) Stearic acid (18:0) Oletic acid (18:1) Linoleic acid (18:2) Linolenic acid (18:3) Eicosenoic (20:0) Behenic (22:0) Others SFA MUFA PUFA UFA Biodiesel properties DU Viscosity (mm2 S1, at 40 °C) CN IV (g I2 100 g1) LCSF (wt.%) CFPP (°C)

Control

NaCl

of the monounsaturated FAs (MUFA) that mainly make up neutral lipids which in turn favors biodiesel production (Hu et al., 2008). As to the biodiesel quality, the most important characteristics of the biodiesel potentially produced from D. abundans oils were empirically estimated and summarized in Table 4. The biodiesel obtained from D. abundans grown in medium supplemented with either NaCl or NaHCO3 had lower viscosity value (4.36 and 4.30) than that of the maximum value (1, Table 6). Additionally, because of the increase in lipid content and lipid productivity, using this two-stage culture with lipid induction phase could help reduce the total cost by 14.40% compared to that

Fig. 5. Time courses of biomass, lipid content and lipid productivity of D. abundans during single- and two-stage cultivation in a 140-L bioreactor.

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Table 6 Comparative energy and economic analyses for biomass or bio-oil production between growth-associated lipid production and two-stage production based on 1 year of cultivating D. abundans in 140-L bubbled columns.

Energy analysis for producing 100,000 kg of algal biomass

Economic analysis for producing 1 kg of algal oil

a b c d e f g h i j k

Variable

Stage I + Stage II

Stage I

Annual biomass production (kg year1) Volumetric productivity (g L1 d1) or (kg m3 d1)a Illuminated areal productivity (kg m2 d1)a Biomass concentration (kg m3)a Reactor volume (m3)b Lipid content (%) a Energy consumption (W m3)c

100,000 0.065 0.017 1.48 4257.68 39.10 40

100,000 0.099 0.026 1.68 2779.19 20.52 40

Total energy for air pumping (kWh months1)d Total energy for biomass drying (kWh year1)e Total energy for oil recovery (kWh year1)e Total energy consumption for producing biomass (GJ year1) Total energy consumption for producing oil (GJ year1) Energy produced as oil (GJ year1)f Energy produced as 100,000 kg biomass (GJ year1)g NER for oil productionh NER for biomass production

51,092.16 9200.00 17,186.70 2240.29

33,350.28 9200.00 17,186.70 1473.85

2302.17

1535.72

1526.34 3155.30 0.66 1.41

801.04 3155.30 0.54 2.14

Input category

Stage I + Stage II (US $/kg algal oil)

Stage I (US $/kg algal oil)

Fertilizers Electricityj Fresh water Buildingk Infrastructurek Total

1.06i 7.21 0.0016 7.94 4.77 20.98

0.40 7.73 0.0034 10.23 6.14 24.51

Data were based on the trial in 140-L bioreactor. Determined by dividing the illuminated area actual by production the volume of each unit. Sierra et al. (2008). Includes 10 h of daily pumping. Stepan et al. (2002). Energy content of net oil yield (assumed value of 39.04 MJ kg1); Jorquera et al. (2010). Energy content of net biomass yield (assumed value of 31.55 MJ kg1); Jorquera et al. (2010). NER would be above 1 if including coproduct allocation (Sander and Murthy, 2010). Including the cost for additional NaCl. Sum of the energy input of air pumping, biomass harvesting, biomass dryer and oil recovery. Based on a 10-year depreciation schedule.

without lipid induction phase for producing 1 kg of algal oil (Table 6). So, NaCl addition is a promising strategy to reduce the cost of algal oil production and increase the energy recovery of the whole process. 4. Conclusions The performances of D. abundans for biodiesel production were evaluated via a two-stage system in a greenhouse. This microalgae flourished at 0.25 g L1 urea at stage I, and yielded the highest lipid productivity of 67.08 mg L1 d1 and better biodiesel quality using 20 g L1 NaCl as lipid inducer at the biomass concentration of 1.79 g L1 at stage II. The optimized conditions were used for cultivating D. abundans in 140-L column bioreactor and obtained concentrated biomass with a high lipid content accompanied by high lipid productivity. This study provided a cost-efficient way for microalgae-based biodiesel production. Acknowledgements This work was funded by National 863 program (2013AA065804) and Program of Sinopec, international partner program of innovation team (Chinese Academy of Sciences), Platform construction of oleaginous microalgae (Institute of Hydrobiology, CAS of China). We are indebted to Prof. Xu for providing us the organism. We also thank Prof. Peng and Ms. Fang for their help in the analytical work.

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NaCl as an effective inducer for lipid accumulation in freshwater microalgae Desmodesmus abundans.

In order to evaluate the efficiency and potential of salt addition-based two-stage cultivation technology, on the basis of urea as nitrogen source, we...
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