Spatiotemporal monitoring of intracellular metabolic dynamics by resonance Raman microscopy with isotope labeling

Cellular metabolites are valuable in a diverse range of applications. For example, the unicellular green algaHaematococcus lacustrisproduces 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 ofH. lacustris, spatial and temporal understanding of its metabolic dynamics is essential. Here we show spatiotemporal monitoring of AXT production inH. lacustriscells by resonance Raman microscopy combined with stable isotope labeling. Specifically, we incorporated carbon dioxide (¹³CO₂) labeled with a stable isotope (¹³C) intoH. lacustriscells through carbon fixation and traced its conversion to¹³C-AXT using our resonance Raman microscope. We incubatedH. lacustriscells under various conditions by switching, pulsing, and replacing¹³CO₂and¹²CO₂. By measurement of these cells we determined the fixation time of¹³C-carbon, visualized the intracellular localization of¹³C- and¹²C-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-producingH. lacustriscells.