Yonamine Yusuke, Hiramatsu Kotaro, Ideguchi Takuro, Ito Takuro, Fujiwara Tomomi, Miura Yoshiko, Goda Keisuke, Hoshino Yu
Research Institute for Electronic Science, Hokkaido University Kita 21, Nishi 10, Kita-ku Sapporo 001-0021 Japan
Department of Chemistry, The University of Tokyo 7-3-1 Hongo, Bunkyo-ku Tokyo 113-0033 Japan.
RSC Adv. 2020 Apr 28;10(28):16679-16686. doi: 10.1039/d0ra02803g. eCollection 2020 Apr 23.
Cellular metabolites are valuable in a diverse range of applications. For example, the unicellular green alga produces as a secondary metabolite the carotenoid pigment astaxanthin (AXT), which is widely used in nutraceutical, cosmetic, and food industries due to its strong antioxidant activity. In order to enhance the productivity of , spatial and temporal understanding of its metabolic dynamics is essential. Here we show spatiotemporal monitoring of AXT production in cells by resonance Raman microscopy combined with stable isotope labeling. Specifically, we incorporated carbon dioxide (CO) labeled with a stable isotope (C) into cells through carbon fixation and traced its conversion to C-AXT using our resonance Raman microscope. We incubated cells under various conditions by switching, pulsing, and replacing CO and CO. By measurement of these cells we determined the fixation time of C-carbon, visualized the intracellular localization of C- and C-AXTs, and revealed the dynamic consumption-production equilibrium of the accumulated AXT. This work is a valuable step in the development of effective screening criteria for high AXT-producing cells.
细胞代谢物在多种应用中具有重要价值。例如，单细胞绿藻作为次生代谢产物产生类胡萝卜素虾青素（AXT），由于其强大的抗氧化活性，虾青素在营养保健品、化妆品和食品工业中被广泛使用。为了提高其产量，对其代谢动力学进行时空理解至关重要。在这里，我们展示了通过共振拉曼显微镜结合稳定同位素标记对绿藻细胞中虾青素产生的时空监测。具体而言，我们通过碳固定将用稳定同位素（¹³C）标记的二氧化碳（¹³CO₂）引入绿藻细胞，并使用我们的共振拉曼显微镜追踪其转化为¹³C-AXT的过程。我们通过切换、脉冲和替换¹²CO₂和¹³CO₂在各种条件下培养绿藻细胞。通过对这些细胞的测量，我们确定了¹³C-碳的固定时间，可视化了¹³C-和¹²C-AXTs的细胞内定位，并揭示了积累的虾青素的动态消耗-产生平衡。这项工作是为高产虾青素绿藻细胞制定有效筛选标准发展中的重要一步。
