微藻為優質的碳循環材料，由於其具備良好之固碳能力，使微藻生長對於地球溫室氣體減量具有重要的貢獻。而且微藻富含許多高價值的產物，像是色素、不飽和脂肪酸與功能性藻蛋白等，使其具有高值化開發的潛力。然而商業化的最大挑戰是生產成本過高，因此建立低成本的藻體量產技術以及微藻生物質高值化技術有其必要性。以循環經濟概念利用微藻能使廢棄物轉換為食品、飼料、化肥、營養品、化妝品、化學品與能源等，使其成為一項極具有前瞻性的永續材料是近年的研究熱點。基於綠藻的蛋白質含量相當豐富並且可提供多元的必需胺基酸，是極佳的蛋白質主要補充來源，因此本研究即針對本土藻株之功能性蛋白質生產進行一系列之探討。在本研究中，選用了六株本土藻種Synechococcus elongatus PCC7002、Chlamydomonas reinhardtii CC-400、Chlorella variabilis、Chlorella vulgaris ESP-31、Chlorella vulgaris FSP-E與Chlorella sorokiniana，當中篩選出具高蛋白質生產力的小球藻Chlorella vulgaris FSP-E (CV) 與Chlorella sorokiniana (CS)，進行優化蛋白生產程序。結果顯示，調控兩種微藻的生長條件中，二氧化碳濃度在2% 到17%時，最佳二氧化碳濃度為5% (v/v)；氮源濃度在2 mM到72 mM時，CV與CS的最佳氮源濃度分別為12 mM硝酸鈉與氯化銨。調控微量鐵元素能有效刺激微藻生長，在微藻培養下添加12 mg/L檸檬酸鐵銨(III)可增加CV與CS的生物量 (4.08 g/L與4.24 g/L)、生物量產率 (583.0 mg/L/d與606.3 mg/L/d) 與蛋白質產量 (793.3 mg/L與812.8 mg/L)，並分別增加CV與CS之蛋白質產率4.86倍與2.77倍 (113.3 mg/L/d與116.1 mg/L/d)。本研究進而採用半批式培養策略，透過80% 培養基置換率連續增加藻體內蛋白含量，使CV與CS可獲得最高生物量 (4.56 g/L與4.75 g/L)、蛋白質產量 (794.2 mg/L與852.2 mg/L) 與蛋白含量 (394.8 mg/g DCW與424.4 mg/g DCW)。由本研究獲得之微藻蛋白經氨基酸分析含有40% 必需氨基酸，且扮演卡爾文循環中能量儲存的一重要角色；在活性特性分析方面，CV與CS蛋白具有益生元活性，分別使益生菌鼠李糖乳桿菌Lactobacillus rhamnosus ZY之生長速率提升48% 與74%。且CV與CS蛋白具有良好的抗菌功能，分別對六株病原菌Aeromonas hydrophila、Bacillus cereus、Escherichia coli、 Pseudomonas putida、Rhodococcus erythropolis與Staphylococcus aureus形成抑菌圈直徑範圍為1.30公分 ~ 1.60公分。CV與CS蛋白更分別具有51.7% 與54.9% 之高羥基自由基清除率，商業用IC50值高達10.3 g/L與6.6 g/L。

Microalgae are high-quality carbonic circular materials. Due to their great carbon-fixing capacity, growth of microalgae makes a considerable contribution to the global greenhouse gas reduction. Moreover, microalgae contain many high-value products, such as pigments, unsaturated fatty acids and functional algal proteins, which have the potential for high-value utilization. However, the biggest challenge for commercialization is the high production cost. Therefore, it is necessary to establish low-cost biomass production techniques of microalgae and high-value technology for microalgal biomass. Microalga is one of the most promising sustainable feedstock as it uptakes waste to convert to large range of sectors: food, feed, fertilizers, nutraceuticals, cosmetics, chemicals, energy, etc. via a circular economy concept, which becomes research hotspot in recent years. According to the abundance of protein content and a variety of essential amino acids in microalgae, they are considered as the major sources of protein supplement. Therefore, a series of discussions on the functional proteins production from native algal strains were examined in this study. In this study, six indigenous algal strains, Synechococcus elongatus PCC7002, Chlamydomonas reinhardtii CC-400, Chlorella variabilis, Chlorella vulgaris ESP-31, Chlorella vulgaris FSP-E and Chlorella sorokiniana were selected. Two kinds of protein-rich microalgae, Chlorella vulgaris FSP-E (CV) and Chlorella sorokiniana (CS), were screened as promising sources for algal proteins due to their high protein productivity. Procedure for optimization of functional proteins production was further carried out. As a result of growth regulation of CV and CS, the optimal CO2 concentration for the growth of both microalgae was 5% (v/v) by adjusting the CO2 concentration from 2% to 17%. The optimal nitrogen source for CV and CS were 12 mM of NaNO3 and NH4Cl by adjusting the nitrogen concentration from 2 mM to 72 mM, respectively. Regulating trace iron element can effectively stimulate the growth of microalgae. With the addition of 12 mg/L ammonium iron (III) citrate, the biomass concentration (4.08 g/L and 4.24 g/L), biomass productivity (583.0 mg/L/d and 606.3 mg/L/d) and protein concentration (793.3 mg/L and 812.8 mg/L) of CV and CS were obtained, representing a 4.86 fold and 2.77 fold increase, respectively, in protein productivity of CV and CS (113.3 mg/L/d and 116.1 mg/L/d). Next, semi-batch cultivation strategy was employed to successively increase protein content of CV and CS at the medium replacement ratio of 80%, achieving the biomass concentration of 4.56 g/L and 4.75 g/L, protein concentration of 794.2 mg/L and 852.2 mg/L and protein content of 394.8 mg/g DCW and 424.4 mg/g DCW for CV and CS, respectively. The obtained microalgal proteins consist of 40% essential amino acids, participating in energy conservation in Calvin cycle. In terms of activity characteristics analysis, the CV and CS proteins possess prebiotic activities as they enhanced the growth of Lactobacillus rhamnosus ZY by 48% and 74%, respectively. Moreover, the CV and CS proteins have high antibacterial activities against predominant pathogens. The diameter of inhibition zones for microalgal proteins with six kinds of pathogen strains, such as Aeromonas hydrophila, Bacillus cereus, Escherichia coli, Pseudomonas putida, Rhodococcus erythropolis and Staphylococcus aureus, varied from 1.30 cm to 1.60 cm. The CV and CS proteins, respectively, also possess high hydroxyl free radical scavenging rate of 51.7% and 54.9%, which have commercial IC50 values of 10.3 g/L and 6.6 g/L.

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