利用纤维床反应器驯化原壳小球藻提高对蔗渣水解液耐受性研究

Domestication of Chlorella protothecoides for High Tolerance to Sugarcane Bagasse Hydrolysate Using a Fibrous-bed Bioreactor

原壳小球藻可快速利用蔗渣水解液中的可发酵糖,但水解液中副产物对细胞生长有抑制作用。为了提高其在高浓度水解液中的异养生长能力,本研究利用纤维床反应器(FBB)驯化细胞,系统研究了蔗渣水解液的制备及其组成、分批补料培养种子液、FBB中的细胞固定化,并在FBB中利用水解液为培养基进行细胞驯化。结果表明,蔗渣经酸解酶解后,其水解液的主要成分为葡萄糖、木糖、乙酸、纤维二糖和阿拉伯糖,浓度分别为18.40g/L、16.17g/L、6.13g/L、5.10g/L和2.29g/L;在发酵罐中采用Basal培养基补料分批培养细胞,117h后细胞密度可达到12.37g/L;将发酵罐与FBB连接并循环培养基33h后形成了固定化细胞床;随后以水解液培养基代替Basal培养基,通过逐级提高水解液培养基浓度来驯化培养固定化细胞,最终从纤维床上分离获得了能在含有35g/L葡萄糖的水解液中异养生长的高耐受性藻株,而野生型藻株不能生长。

Chlorella protothecoides can rapidly utilize fermentable sugars in sugarcane bagassehydrolysate（SCBH）, but its cell growth is inhibited by by-products in thehydrolysate. In this study, to enhance theheterotrophic growth capability inhigh-concentration SCBH, C. protothecoides was cultured in a fibrous-bed bioreactor（FBB）. The preparation and components of SCBH, seed culture by batch-fed fermentation, cell immobilization in FBB, and cell domestication in SCBH medium in an FBB were systemically studied. The results showed that after acidhydrolysis and enzymolysis, the main components of SCBH were glucose, xylose, acetic acid, cellobiose, and arabinose with concentrations of 18.40, 16.17, 6.13, 5.10, and 2.29g/L, respectively. The cell density was up to 12.37g/L for batch-fed cultures in Basal medium after 117hours in a fermenter. The immobilized cell bed formed when the medium was recycled for 33hours after connecting the fermenter with the FBB. Then, the Basal medium was replaced with SCBH medium, and the concentration of the SCBH medium was gradually increased to domesticate the immobilized cells. Ahighly tolerant strain, which grewheterotrophically in SCBH medium containing 35g/L glucose, was isolated from the fibrous bed, while the wild-type strain was unable to grow.