菱形藻的培养及积累岩藻黄素的调控机制的探讨

Fucoxanthin, a natural product in seaweeds, has unique molecular structure. It was recently revealed that fucoxanthin has significant anti-cancer activity and anti-obese effect without any side effects. Its high commercial value made it great exploitation foreground. However, the traditional process in preparation of fucoxanthin from brown algae shown disadvantages including complex purifying process and high cost. And fucoxanthin derived from microalgae gradually has got much attentions. However, the mechanism in metabolism and regulation of fucoxanthin for accumulation in algae has not been clear. Therefore, this algae, Nitzschia sp. MD1, which was proved to be poly-extremophilic and fucoxanthin-rich, will be used as researched object in this program. These key factors on growth and fucoxanthin accumulation of MD1 will be assessed by culture medium and culture conditions optimization, microalgal cell physiological and biochemical characteristics and fucoxanthin production. Then carotenoids metabolic and carotenoids synthetic key enzyme gene transcriptional level of MD1 induced by key factors will be studied to investigate the biosynthetic pathway of fucoxanthin in MD1 and the possible mechanism of induction of fucoxanthin. Research results of this program will provide the basis information of large-scale cultivation of micralgae for fucoxanthin prodution.

海藻天然产物岩藻黄素分子结构独特，具有显著的抗肿瘤和减肥等效果并无任何的毒副作用，商业附加值高，应用前景巨大。然而，传统工艺采用大型褐藻制备岩藻黄素，纯化工艺复杂且成本高，以微藻生产岩藻黄素逐渐成为备受关注的研究方向。然而，合成积累岩藻黄素的调控机制目前尚未清楚，因此，本项目在筛选出一株岩藻黄素含量高、适应性强的优良藻株菱形藻MD1的基础上，拟以该藻株为研究对象，通过对培养基和培养条件优化、藻细胞生理生化特性以及岩藻黄素的研究，确定影响MD1生长与岩藻黄素积累的关键因素，以此为线索，深入开展MD1类胡萝卜素代谢和类胡萝卜素合成关键酶基因转录水平的研究，初步探讨MD1岩藻黄素合成途径和积累诱导机理，可望为规模化生产微藻岩藻黄素奠定重要理论与技术基础。

微藻中岩藻黄素的含量可达到大型藻的数倍甚至数十倍，开发微藻作为天然岩藻黄素提取的优质原料这一课题开始吸引研究界和产业界的关注。微藻细胞中岩藻黄素的积累与藻种种质、培养条件等密切相关。.项目开展了富含岩藻黄素微藻的藻种选育，分离得到60多株硅藻，从中筛选出2株具有优良品质的藻种，包括菱形藻MD1（SCSIO788）、三角褐指藻(SCSIO433)。并以这2株硅藻为研究对象，开展了以下工作：.研究了培养基中NaHCO3、NaCl浓度及pH三种因素对菱形藻MD1生长和色素含量的影响；光质、番茄红素对三角褐指藻生长以及光合色素积累的影响；进一步从转录组水平上探讨了以红为主的红蓝组合光条件下诱导岩藻黄素合成积累的机制；此外，对三角褐指藻联合生产岩藻黄素和油脂进行了初步评估，建立了微藻岩藻黄素提取纯化工艺，体外评估了岩藻黄素抗氧化、降血糖活性。研究表明，最适宜菱形藻MD1生长的NaHCO3、NaCl浓度以及pH及分别为0.2mol L-1、10 g L-1和9.5，最适宜岩藻黄素积累分别为0.2 mol L-1、20 g L-1和9.5，优化条件后岩藻黄素含量最高达1.69%；单种光质红光和绿光不仅抑制藻细胞的生长，还抑制光合色素的合成积累；蓝光有利于藻细胞的生长，但抑制岩藻黄素的积累。红蓝光组合最有利于岩藻黄素的积累，含量达1.22%，显著高于白光对照组；转录组研究发现红蓝光处理提高了前番茄红素异构酶(CRTISO)、紫黄质脱环氧化酶（VDE）和玉米黄质环氧化酶(ZE）基因的表达。通过简单、污染小的岩藻黄素纯化技术，可以获得纯度为76%的岩藻黄素；岩藻黄素清除ABTS·+和DPPH的半数抑制浓度分别为0.50和0.85 mg/ml，1 mg/mL岩藻黄素对AGEs抑制率为67.72%。通过研究可得出以下结论：通过优化培养基、光质等条件可以促进岩藻黄素积累；菱形藻MD1和三角褐指藻可作为提取岩藻黄素的原料。本项目的研究为微藻岩藻黄素的工业化生产有指导性意义。