Paper title: Development of Algal Culture by Using Waste of Steel Industry

Transcription

1 Paper title: Development of Algal Culture by Using Waste of Steel Industry Author(s): Arunee Ewechareon 1, Natthawut Yodsuwan 2,3, Yothaka Puchcha 2 and Sarote Sirisansaneeyakul 2,3 Address: 1 Iron and Steel Institute of Thailand, Soi Trimitr, Rama IV Rd., Prakanong, Khlongtoey, Bangkok, THAILAND 2 Fermentation Technology Research Center, Department of Biotechnology, Kasetsart University, Chatuchak, Bangkok, 10900, Thailand 3 Center for Advanced Studies in Tropical Natural Resources, National Research University- Kasetsart University, Kasetsart (CASTNAR, NRU-KU, Thailand) University, Chatuchak, Bangkok, 10900, Thailand Abstract: The feasibility study of microalgal cultivation by using the nutrients from iron industrial wastewater was performed. Seven species of microalgae which are Chlorella sp. TISTR 8990 Chlorella sp. TISTR 8991 Chlorella sp. IFRPD 1162 Chlorella sarakiniana DMKU 522 Chlorella protothecoides UTEX 25 Ankistrodesmus sp. and Scenedesmus sp. were studied. The preliminary study was performed in mixed wastewater which used as algal growth medium. Unfortunately, no microalgae could grow on this defined medium. Thus, the acclimatization of these microalgae has been done in Horikoshi basal medium mixed with 5% v/v wastewater, ph 6.0 for 14 days. The result showed that there were 4 species including Chlorella sp. TISTR 8990 Chlorella sp. TISTR 8991 C. sarakiniana DMKU 522 and C. protothecoides UTEX 25 could be acclimatized satisfactorily. Thus, these 4 species would be potentially used for next step which were acclimatized in 150 ml Horikoshi basal medium mixed with 5% v/v wastewater, ph 6.0 for 10 days. There were used as algal starter for the 1st fed-batch cultivation which had the same conditions as described previously. The 2nd fedbatch cultivation was done after 10 days of the 1st fed-batch cultivation. The Horikoshi basal medium mixed with 5% v/v wastewater was added into the culture from the 1st fed-batch cultivation to provide the working volume at 150 ml. The 2nd fed-batch cultivation was performed for 10 days. The results showed that the C. protothecoides UTEX 25 which cultivated in the Horikoshi basal medium mixed with 5% v/v wastewater grew satisfactorily comparing with control condition (no mixed wastewater added). Moreover, it showed more potential than the other strains. However, the limitations of fed-batch growth that contains the Horikoshi basal medium mixed with wastewater were found during the cultivation. As compared with the acclimatized batch cultivation, the specific rate of growth which calculated from the fed-batch condition showed the low values and the precipitation of microalgae in the 2nd fed-batch cultivation was also found. Moreover, the specific rate of growth which calculated from the 2nd fed-batch cultivation was also found to be lower than obtained from the 1st fed-batch cultivation. In conclusion, the microalgae had the possibility

2 to cultivate in the medium which added with nutrients from iron industrial wastewater. However, the stage of acclimatization is needed, as well as, the consideration of the algal starter and the other factors which effect on promotion and inhibition the microalgal growth are necessary. Keywords: Algal Culture, Waste of Steel Industry Chemical & Equipment: Horikoshi basal medium Wastewater from Steel Industry Problem and Statement Global warming becomes more serious due to greater demands of consumers, which leads to additional resources consumption. With this reason, the United Nations has proposed the Kyoto Protocol under which the industrial countries are required to reduce their release of greenhouse gas by applying the best technology with the aim to solve the said problem. In addition, the 2008 alternative energy development strategy of the Department of Alternative Energy Development and Efficiency, Ministry of Energy includes the plan for using biodiesel of 4.5 million liters per day in Thus, sources of raw materials for manufacturing biodiesel have been sought. Most of these raw materials are agricultural products such as palm oil, coconut oil, soybean oil, sesame oil, and sunflower oil. However, the problem is that the prices of these agricultural products are based on consumption demand so their production costs are high and volatile, by which the biodiesel price is affected. Furthermore, these agricultural products also require a large storage area and a long harvesting period so they are not suitable for manufacturing biodiesel. Therefore, the sources of other short-harvesting period raw materials with high productivity are studied. It is reported that Halimeda sp. of Thailand can be used as a source of raw material for manufacturing biodiesel because it needs a small storage area and has a short-harvesting period. Additionally, it also helps reduce the release of greenhouse gas. This algal species is capable of absorbing up to 2,400 tons of carbon dioxide per year, 600 tons of which is used for photosynthesis and remaining 1,800 tons per year is used for generating calcium carbonate. Furthermore, some other algal species can produce oil (Beer, 2009) such as Oscilltoria sp. and Botryococcusbraunii. As algae become a source of raw materials for manufacturing alternative energy and its fermentation also generates biogas, they are interesting alternative energy plants. In cultivation of algae as a source of alternative energy, the problem is that the price of dehydrated medium is rather high so it is not worthy to do so. With this reason and from the study of preliminary information on components of steel industrial waste e.g. sediment arising from the steel production process, wastewater from the coating or plating process is conducive to the algal growth, and the steel industry has released greenhouse gas, CO 2, NO x and So x. The preceding study reveals that greenhouse gas can dissolve into water as a nutrient for algal cultivation which helps reduce the release of greenhouse gas of the industry sector.

3 Moreover, the steel industry needs reducing gas or hydrogen or methane gas for the direct reduction process which pulls out oxygen from the iron ore to ensure that it is suitable for being used as a raw material of the next process. This research thus studies the feasibility of using the steel industry waste as a nutrient and for fermentation of algal biomass to produce required gas for the steel industry, with an objective to establish the body of knowledge for the industrial sector. This will lead to the concrete and sustainable development. Experimental Method 1. Study of the acclimatization of microalgae growth in basal medium mixed with different concentration of wastewater 1.1 Preparation of microalgae starter Prepare the starter of 7 microalgae species in 50 ml Horikoshi basal medium, which were cultivated in 200 ml cultivation tubes, ph 6.0 at 30 C under the light intensity of 15 klilolux, contrast ratio of 8:16 hours and continuous feeding rate of carbon oxide at 0.67 vvm for 14 days. 1.2 Study of the acclimatization of microalgae growth after adding different concentration of wastewater Mix the wastewater from steel industry with ph 6.0 and culture each species of microalgae into test tubes then add the starter of microalgae deriving from 1.1 to the (sterilized) basal medium with the working volume of 10 ml as shown in Table 1. Table 1: The percentage of microalgae and wastewater Percentage (%) No. Wastewater of Steel microalgae industry Horikoshi basal medium 1 (Control) (Control) Cultivate the microalgae under the lighting condition (fluorescent lamp) at the room temperature (around 30 C) for 14 days and add 5% v/v wastewater to tube no. 2 and no. 6 and 10% v/v wastewater to tube no. 3 respectively, then cultivate the microalgae under the same condition for 14 days, compare the growth of microalgae with wastewater mixed in the control condition by measuring the turbidity at 680 nm. On the last day of cultivation and select the microalgae which can grow in the wastewater for the next step.

4 2. Cultivation of microalgae with nutrients from industrial wastewater 2.1 Preparation of the starter of 4 microalgae species in 50 ml Horikoshi basal medium, ph 60, which were cultivated in 200 ml cultivation tubes (three tubes of each species: control condition i.e. tube no. 1 and no. 2), at 30 C under the light intensity of 15 klilolux, the contrast ratio of 8:16 hours and continuous feeding rate of carbon oxide at 0.67 vvm for 7 days, then cultivate the same with the working volume at 150 ml in Horikoshi basal medium for 7 days. 2.2 Acclimatization of the growth of microalgae by adding 5% v/v wastewater in the fedbatch cultivation Dilute the starter of microalgae deriving from 2.1 to have the turbidity of 680 nm. at in Horikoshi basal medium then add 5% v/v wastewater, ph 6.0 to the starter (repeating the experiment as per 2 in tube 1 and no. 2) which were cultivated in 200 ml tubes to provide the working volume at 150 mm, at 30 C under the light intensity of 15 kilolux, the contrast ratio of 8:16 hours and the continuous feeding rate of carbon dioxide at 0.67 vvm for 10 days, and collect samples every day to measure the turbidity of 680 nm., ph and weight of dry cell st fed-batch cultivation with 5% v/v wastewater added Cultivate the microalgae deriving from 2.2 in Horikoshi basal medium mixed with 5% v/v wastewater, ph 6.0 (in tube no. 1 and no. 2), which were cultivated in 200 ml tubes to provide the working volume at 150 mm, at 30 C under the light intensity of 15 kilolux, the contrast ratio of 8:16 hours and the continuous feeding rate of carbon dioxide at 0.67 vvm for 10 days, and collect samples every day to measure the turbidity of 680 nm., ph and weight of dry cell nd fed-batch cultivation with 5% v/v wastewater added Cultivate the microalgae deriving from 2.2 in Horikoshi basal medium mixed with 5% v/v wastewater, ph 6.0 (in tube no. 1 and no. 2), which were cultivated in 200 ml cultivation tubes to provide the working volume at 150 mm, at 30 C under the light intensity of 15 kilolux, the contrast ratio of 8:16 hours and the continuous feeding rate of carbon dioxide at 0.67 vvm for 10 days, and collect samples every day to measure the turbidity of 680 nm., ph and weight of dry cell. Results 1. Study of the acclimatization of microalgae growth in basal medium mixed with different concentration of wastewater The cultivation of various microalgae species i.e. Chlorella sp. TISTR 8990, Chlorella sp. TISTR 8991, Chlorella sp. IFRPD 1162, Chlorella sarakiniana DMKU 522, Chlorella protothecoides UTEX 25, Ankistrodesmus sp. and Scenedesmus sp. in Horikoshi basal

5 medium mixed with 5% and 10% v/v wastewater, ph 6.15 for 14 days in the lighting condition (florescent lamp) at the room temperature shows that all 7 microalgae species grew satisfactory. For the first 7 days after the cultivation, there was no difference between testing set and control condition. After 14 days of cultivation, the following 4 microalgae species are qualified for cultivating in the wastewater of the steel industry: Chlorella sp. TISTR 8990, Chlorella sp. TISTR 8991, C. sarakiniana DMKU 522, C. protothecoides UTEX Cultivation of microalgae in nutrients from industrial wastewater The starter of microalgae species namely Chlorella sp. TISTR 8990, Chlorella sp. TISTR 8991, C. sarakiniana DMKU 522, and C. protothecoides UTEX 25, cultivated for 7 days were acclimatized in Horikoshi basal medium mixed with 5% v/v wastewater, ph 6.0 in tube no. 1 and no. 2 compared with the control condition (no wastewater added). Cultivation and sampling were conducted for 10 days. After acclimatization of the 4 microalgae species, their starters were transferred to Horikoshi basal medium mixed with 5% v/v wastewater, ph 6.0, and the 1 st fed-batch cultivation was conducted in 200 ml tubes in the lighting condition for 10 days compared to the control condition (no wastewater added). Cultivation and sampling were conducted for 10 days. Thereafter, Horikoshi basal medium mixed with 5% v/v wastewater, ph 6.0 was added to the microalgae sediment until it reached 150 ml and cultivation was made in 200 ml tubes as the 2 nd fed-batch cultivation under the lighting condition for 10 days compared to the control condition (no wastewater added). Cultivation and sampling were conducted for 10 days. The results of each species are as follows: Table 2 : The summary result Parameter Batch Batch Type of microalgae Chlorella sarakiniana Chlorella sp. TISTR 8990 Chlorella sp. TISTR 8991 DMKU 522 Fed Fed Fed Fed Fed Batch Batch Batch Batch Batch Batch 1 st 2 nd 1 st 2 nd 1 st Fed Batch 2 nd Batch Chlorella protothecoides UTEX 25 Fed Fed Batch Batch 1 st 2 nd Growth rate (mg/l.day) coefficient (per day) Batch cultivation : Comparison of the growth of the 4 microalgae species i.e. Chlorella sp. TISTR 8990, Chlorella sp. TISTR 8991, C. sarakiniana DMKU 522, and C. protothecoides UTEX 25, cultivated by ways of fed-batch cultivation in basal medium with 5% v/v wastewater added for acclimatization purpose was made. The parameter values are as shown in Table 24. It is found that C. protothecoides UTEX 25 generated the highest concentration of the last cell when compared to the remaining strains (Chlorella sp. TISTR 8990, Chlorella sp. TISTR 8991, C. sarakiniana DMKU 522, respectively). This is in line with the cell production rate in terms of volume, but the specific rates of growth are different. It is also found that Chlorella sp. TISTR 8990 had the highest specific rate of growth

6 followed by C. sarakiniana DMKU 522, C. protothecoides UTEX 25, and Chlorella sp. TISTR 8991, respectively. When compared to the control condition with no wastewater added, it is found that Chlorella sp. TISTR 8990 and Chlorella sp. TISTR 8991 had the declining specific rate of growths when they were cultivated in the basal medium mixed with 5% v/v wastewater (4.36% and 40.4%, respectively when compared to the specific rate of growth under the control condition). However, C. sarakiniana DMKU 522 and C. protothecoides UTEX 25 had the increasing specific rate of growth when adding 5% v/v wastewater (4.16% and 26.84%, respectively when compared to the specific rate of growth under the control condition). 1 st fed-batch cultivation : Comparison of growth of the 4 microalgae species from the 1 st fed-batch cultivation in basal medium with 5% v/v wastewater added was made. The parameter values are shown in Table 25, it is found that C. protothecoides UTEX 25 generated the highest concentration of the last cell when compared to the remaining strains (Chlorella sp. TISTR 8990, Chlorella sp. TISTR 8991, and C. sarakiniana DMKU 522, respectively). This is in line with the cell production rate of the batch cultivation in terms of volume, but the specific rates of growth are different. It is also found that Chlorella sp. TISTR 8990 had the highest specific rate of growth followed by C. protothecoides UTEX 25, Chlorella sp. TISTR 8991, and C. sarakiniana DMKU 522, respectively. When compared to the control condition with no wastewater added, it is found that Chlorella sp. TISTR 8990 had the increasing specific rate of growth after adding 5% v/v wastewater (15.06% when compared to the specific rate of growth under the control condition). However, C. Chlorella sp. TISTR 8991, C. sarakiniana DMKU 522 and C. protothecoides UTEX 25 had the declining specific rate of growth when being cultivated in the basal medium mixed with 5% v/v wastewater added (8.33%, 18.79%, and 10.40%, respectively when compared to the specific rate of growth under the control condition). 2 nd fed-batch cultivation : Comparison of growth of the 4 microalgae species from the 2 nd fed-batch cultivation in basal medium with 5% v/v wastewater added was made. The parameter values are shown in Table 26. It is found that C. protothecoides UTEX 25 generated the highest concentration of the last cell when compared to the remaining strains (Chlorella sp. TISTR 8991, C. sarakiniana DMKU 522, and Chlorella sp. TISTR 8990, respectively). This is in line with the cell production rate in terms of volume, but the specific rates of growth are different. It is found that C. sarakiniana DMKU 522 had the highest specific rate of growth followed by Chlorella sp. TISTR 8991, C. protothecoides UTEX 25, and Chlorella sp. TISTR 8990, respectively. When compared to the control condition with no wastewater added, it is also found that Chlorella sp. TISTR 8991 had the increasing specific rate of growths after adding 5% v/v wastewater (12.40% when compared to the specific rate of growth under the control condition). However, Chlorella sp. TISTR 8990, C. sarakiniana DMKU 522, and C. protothecoides UTEX 25 had the declining specific rate of growth when being cultivated in the basal medium mixed with 5% v/v wastewater added (36.19%, 20.56%, and 21.43%, respectively when compared to the specific rate of growth under the control condition).

7 This shows that the acclimatization of microalgae growth by ways of batch cultivation in Horikoshi basal medium with 5% v/v wastewater added affects their growth, increasing and declining, as same as the fed-batch cultivation. However, it is found that with the fed-batch cultivation, the growth rate tends to decrease more. This may be because with the fed-batch cultivation, the microalgae cells used as starter are livelong so when they were used as starter for the next fed-batch cultivation, these cells could not grow fast unlike the microalgae cells in the state of multiple growth. Furthermore, ore or metal in the mixed wastewater which was added in the 1 st and 2 nd fed-batch cultivation may cause the sediment of algal cells due to unsuitable condition. This affects the declination of the growth rate of microalgae cultivated under such condition. Conclusion The feasibility study of microalgae cultivation by using the nutrients from the steel industrial wastewater was performed. In this respect, the 7 microalgae species i.e. Chlorella sp. TISTR 8990 Chlorella sp. TISTR 8991, Chlorella sp. IFRPD 1162, Chlorella sarakiniana DMKU 522, Chlorella protothecoides UTEX 25, Ankistrodesmus sp. and Scenedesmus sp. in the mixed wastewater were studied. Unfortunately, no microalgae could grow on this defined medium. Thus, the acclimatization of these microalgae has been done in Horikoshi basal medium mixed with 5% v/v wastewater, ph 6.0 for 14 days. The result showed that there were 4 species including Chlorella sp. TISTR 8990, Chlorella sp. TISTR 8991, C. sarakiniana DMKU 522, and C. protothecoides UTEX 25 could be acclimatized satisfactorily. Thus, these 4 species would be potentially used for next step. These 4 microalgae species were cultivated by ways of batch cultivation to acclimatize in Horikoshi basal medium mixed with 5% v/v wastewater, ph 6.0 for 10 days in 200 ml tubes before there were used as algal starter for the 1 st fed-batch cultivation in 150 ml Horikoshi basal medium mixed with 5% v/v wastewater in 200 ml tubes for 10 days. The 2 nd fed-batch cultivation was done for 10 days. For this cultivation, Horikoshi basal medium mixed with 5% v/v wastewater was added to the culture from the 1 st fed-batch cultivation to provide the working volume at 150 ml. The results showed that C. protothecoides UTEX 25 grew satisfactorily when compared to control condition with no mixed wastewater added. Moreover, it showed more potential than the three remaining strains. However, the limitations of fed-batch cultivation are that when compared to the batch cultivation for acclimatization, the specific rate of growth declined dramatically and microalgae sediment arose from the 2 nd fed-batch cultivation with 5% v/v wastewater added in Horikoshi basal medium. In addition, the 2 nd fed-batch cultivation resulted that the specific rate of growth decreased from that of the 1 st fed-batch cultivation, which may be due to the age of microalgae used as starter. This decreased the algal growth capability. After considering the feasibility of the algal cultivation in the wastewater of the steel industrial, it is found that microalgae can be cultivated but acclimatization is needed before proceeding with the next step. This research reveals that the fed-batch cultivation could be made once. However, it depends on the strength of the starter used in the cultivation and limitations such as cultivation period, required concentration of the first and last algal cells,

8 quantify of nutrients, heavy metal in the mixed wastewater which affect the promotion and suspension of microalgae growth. In this regard, further study of other factors has to be conducted for an in-dept explanation. Reference Beer, L.L., et al., Engineering algae for biohydrogen and biofuel production. Current Opinion in Biotechnology, (3): p Berberoglu, H., Gomez, P.S. and Pilon, L., Radiation characteristics of Botryococcus braunii, Chlorococcum littorale, and Chlorella sp. used for CO2 fixation and biofuel production. Journal of Quantitative Spectroscopy and Radiative Transfer, (17): p Borowitzka, M.A., Marine and halophilic algae for the production of biofuels. Journal of Biotechnology, (Supplement 1): p. S7-S7. Lehr, F. and Posten, C., Closed photo-bioreactors as tools for biofuel production. Current Opinion in Biotechnology, (3): p Maeda, K., Owada, M., Kimura, N., Omata, K., Karube, I., CO2 fixation from the flue gas on coal-fired thermal power plant by microalgae. Energy Conservation Management, (6-9): p Nagase, H., et al, Improvement of Microalgal NOx Removal in Bubble Column and Airlift Reactors. Journal of fermentation and engineering, (4): p Posten, C. and Schaub, G., Microalgae and terrestrial biomass as source for fuels--a process view. Journal of Biotechnology, (1): p Yanagi, M., Watanabe, Y. and Saiki, H., CO2 Fixation by Chlorella sp. HA-1 and Its Utilization. Energy Conservation Management, (6-9): p Yun, Y.S., Lee, S.B., Park, J.M., Lee, C.I. and Yang, J.W., Carbon dioxide fixation by algal cultivation using wastewater nutrients. Journal of Chemical Technology and Biotechnology, : p