利用鹼性培養基捕捉高濃度二氧化碳用以培養耐鹼微藻突變株

Abstract

碳是生物系統的基本元素，而二氧化碳(CO2)是微藻主要碳來源，先前的研究指出養殖微藻的過程中給予較高濃度的CO2，可以得到較高的生物質產率。儘管如此，在實際大規模養殖微藻的情形下，建置CO2管線來養殖微藻過於昂貴而不可行。微藻的CO2來源可以藉由工業廢氣來提供，此不僅可提高生物質產率與CO2生物利用率，並有利於碳的捕獲而減少溫室氣體的排放。另外，煙道廢氣中的CO2濃度過高(10~30％)，大量的CO2來不及被水吸收就逸散回環境中，因此CO2生物利用率極低。若能有一高效率吸收廢氣中CO2的方式，如此便不需埋設大量氣體管線，也能將吸附有CO2的培養液提供微藻生長，此將適用於大規模養殖微藻。這篇研究主要探討鹼性培養基是否能夠吸附足夠的CO2提供微藻生長並且提高生物質產率，以期能夠利用在往後大規模微藻養殖系統之中。 本實驗使用氫氧化鈉配製鹼性培養基以提升其吸附CO2的能力，而微藻可以直接使用溶於液體中的CO2或碳酸根提供生長所需。結果顯示濃度0 (pH 6)、0.005 (pH 7)、0.01 (pH 8)、0.015 (pH 9)、0.02 (pH 10)、0.025 (pH 11)及0.06 M (pH 12)的氫氧化鈉分別可以吸附 226、507、677、903、1100、1214及3195 ppm CO2。之後將微藻養於添加不同濃度氫氧化鈉0、0.005、0.01、0.015、0.02、0.025及0.06 M的培養基之中，結果發現氫氧化鈉濃度越高，越能提供二氧化碳給微藻生長而有較高的生物質產率(0.29、0.40、0.46、0.48、0.51、0.54、0.55 g L-1 day-1)。此外發現到使用間歇性通氣養殖方式時，微藻長時間處於高pH環境生長會受到抑制，為了進一步提高微藻生物質產率於鹼性培養基系統中，我們將Chlorella sp.進行NTG突變，篩選出一株耐鹼藻株Chlorella sp. HPT，此耐鹼藻可於通空氣條件下在pH 8~11培養基中生長良好，在pH 10培養基時與原始藻相比有較佳的耐鹼性，培養7天後，耐鹼藻的最大生物質產率相較於Chlorella sp.增加2倍以上。之後將突變微藻Chlorella sp. HPT用於鹼性培養基養殖微藻系統中，於Chlorella sp. HPT培養期間每3、6、12及24小時間隔通入30分鐘10% CO2，培養液中的pH會逐漸呈現上升至10，Chlorella sp. HPT仍能生長良好，並且生物質產率較原始藻Chlorella sp. wild type高。當連續通入10% CO2和每3小時間隔通入30分鐘10% CO2進行Chlorella sp. HPT室內養殖，每3天置換一半培養液為期21天，平均微藻生物質濃度為5.08和4.35 g L-1，平均微藻生物質產率為0.878和0.726 g L-1 day-1。接著於室外每7天置換一半培養液為期21天養殖，白天連續通入10% CO2和每3小時間隔通入30分鐘10% CO2進行微藻養殖，平均微藻生物質濃度和生物質產率分別為2.13和1.87 g L-1及0.195和0.163 g L-1 day-1。研究顯示耐鹼藻Chlorella sp. HPT可於室內1-L光生物反應器(PBR)和室外60-L PBR以半連續置換培養液和間歇通入CO2的養殖條件下，同樣可以建立穩定且長期的微藻養殖製程。

Carbon is the fundamental element of the living system, and carbon dioxide (CO2) is the source of carbon for microalgae. Previous studies indicate that the relatively high CO2 concentration with high microalgal biomass productivity. However, it is too expensive to construct gas line in large scale microalgae pond. The microalgae growth can be provided by exhaust gases (flue gases) from industrial plants, which significantly increases the biomass productivity and the CO2 bioavailability for algae, and it is conducive to carbon capture and reduce greenhouse gas emissions. In addition, the CO2 concentration of flue gases is very high (10~30%), a lot of CO2 is too late to be absorbed by water and easy escape into the environment, so the CO2 bioavailability is extremely low. Therefore, it is essential to develop a system that has high efficient absorb CO2 from exhaust gas, then there is no need to lay a large number of gas pipelines, and also can absorb CO2 with culture medium to provide microalgae growth, the system will be effectively applied to large-scale farming microalgae. This study investigates alkaline medium could absorption more CO2 concentration and provides the technique to promote microalgae biomass productivity in large-scale cultivation. Present study enhances the CO2 retaining capacity of microalgae alkaline culture medium by the addition of NaOH. Microalgae absorb carbon source directly from the dissolved CO2 or HCO3- in the culture medium. Our data showed that microalgae culture medium capture 226, 507, 677, 903, 1100, 1214, and 3195 ppm CO2 by addition of 0 (pH 6), 0.005 (pH 7), 0.01 (pH 8), 0.015 (pH 9), 0.02 (pH 10), 0.025 (pH 11), and 0.06 M (pH 12) M NaOH. Moreover, the microalgae were incubated in a different concentrations (0, 0.005, 0.01, 0.0125, 0.02, 0.025, and 0.06 M) NaOH medium, as a result, the higher the concentration of NaOH capture higher concentration of CO2 to improve the biomass productivity (0.29, 0.40, 0.46, 0.48, 0.51, 0.54, and 0.55 g L-1 day-1). In addition, the microalgae grows in a high pH environment for a long time will be inhibited with intermittent CO2 aeration culture in alkaline medium. In order to improve the microalgae biomass productivity in alkaline medium systems, an alkaline-tolerance Chlorella sp. HPT mutant strain was screened by wild-type Chlorella sp. of NTG mutagenesis. Chlorella sp. HPT could grow well in pH 8~11 medium with air aeration and the mutant microalgae had better alkali resistance compared to the original microalgae Chlorella sp. wild type at pH 10. When microalgal cells were cultured in pH 10 medium, the maximum biomass productivity of Chlorella sp. HPT was higher approximately 2-fold than wild-type Chlorella sp. for 7-days cultivation. Then the mutant microalgae Chlorella sp. HPT was used in alkaline culture medium system. At intermittent CO2 aeration for 30 min at 3, 6, 12, and 24 hours the pH of medium was gradually close to 10 and Chlorella sp. HPT also could grow well in these alkaline culture medium, and the biomass productivity was higher than the Chlorella sp. wild type. When Chlorella sp. HPT aerated with 10% CO2 continuously and 10% CO2 intermittently for 30 min at 3 hours interval indoor semi-continuous cultivation with 3-day replacement for a period of 21 days, the average biomass concentration and biomass productivity of Chlorella sp. HPT were 5.08 and 4.35 g L-1, and 0.878 and 0.726 g L-1 day-1, respectively. For a period of 21 days outdoor semi-continuous cultivation with 7-day replacement, the average biomass concentration and average biomass productivity of Chlorella sp. HPT aerated with CO2 continuously and intermittently during the daytime were 2.13 and 1.87 g L-1, and 0.195 and 0.163 g L-1 day-1, respectively. The results demonstrate that a stable growth performance of Chlorella sp. HPT could be achieved for long-term microalgal cultivation indoor 1-L PBR and outdoor 60-L PBR in semi-continuous cultivation.