Abstract
Although the importance of nonphotochemical quenching (NPQ) on photosynthetic biomass production and crop yields is well established, the in vivo operation of the individual mechanisms contributing to overall NPQ is still a matter of controversy. In order to investigate the timescale and activation dynamics of specific quenching mechanisms, we have developed a technique called snapshot transient absorption (TA) spectroscopy, which can monitor molecular species involved in the quenching response with a time resolution of 30 s. Using intact thylakoid membrane samples, we show how conventional TA kinetic and spectral analyses enable the determination of the appropriate wavelength and time delay for snapshot TA experiments. As an example, we show how the chlorophyll-carotenoid charge transfer and excitation energy transfer mechanisms can be monitored based on signals corresponding to the carotenoid (Car) radical cation and Car S1 excited state absorption, respectively. The use of snapshot TA spectroscopy together with the previously reported fluorescence lifetime snapshot technique (Sylak-Glassman et al. in Photosynth Res 127:69–76, 2016) provides valuable information such as the concurrent appearance of specific quenching species and overall quenching of excited Chl. Furthermore, we show that the snapshot TA technique can be successfully applied to completely intact photosynthetic organisms such as live cells of Nannochloropsis. This demonstrates that the snapshot TA technique is a valuable method for tracking the dynamics of intact samples that evolve over time, such as the photosynthetic system in response to high-light exposure.
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Acknowledgements
We thank Drs. Dagmar Lyska and Masakazu Iwai for helpful discussions and for providing Nannochloropsis oceanica samples. We also thank Lam Lam for helpful assistance during her visit to the University of California, Berkeley. This work was supported by the U.S. Department of Energy, Office of Science, Basic Energy Sciences, Chemical Sciences, Geosciences, and Biosciences Division under Field Work Proposal 449B. G. R. F. thanks Magdalen College, Oxford for a visiting Fellowship, and the Department of Physical and Theoretical Chemistry, Oxford for their hospitality during the writing of this paper.
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Park, S., Steen, C.J., Fischer, A.L. et al. Snapshot transient absorption spectroscopy: toward in vivo investigations of nonphotochemical quenching mechanisms. Photosynth Res 141, 367–376 (2019). https://doi.org/10.1007/s11120-019-00640-x
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DOI: https://doi.org/10.1007/s11120-019-00640-x