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Photosynthetic Oxygen Evolution

University of Turku (Finland)

Involved scientists: Vesa Havurinne and Esa Tyystjärvi

Photosynthesis produces oxygen and therefore plant scientists often measure the rate of oxygen evolution. The traditional methods of oxygen measurement require extensive maintenance and the methods are prone to electrical interference. We have found the optical oxygen meter FireStingO2 from Pyro Science reliable and fast and feel that this is the future of oxygen measurement in photosynthesis research.

In order to measure photosynthetic oxygen evolution in liquid medium, we used a calibration method based on hydrogen peroxide and catalase to ensure the precise reading in oxygen concentrations above the level of air-saturated water. The calibration method is based on the stoichiometric coefficients of the catalase reaction H2O2 à 2H2O + O2. Further comparison between the results obtained using different calibration methods (hydrogen peroxide, single-point and 2-point calibration with oxygen-free water) showed no clear deviation from each other, and therefore it is obvious that even the very simple single-point calibration in water is enough for oxygen evolution experiments.

By 3D-printing we were able to create a perfect fit cuvette system for a type OXB430-OI oxygen minisensor, an external temperature sensor and the FireStingO2 central module. Temperature control is provided by water circulation and the sample is mixed with a magnetic stirrer (Figs. 1 & 2). Oxygen measurements from isolated thylakoid membranes using dimethylbenzoquinone as electron acceptor (Fig. 3) showed the same results as obtained with an oxygen electrode using the same sample.

For further information, please contact Esa Tyystjärvi (esatyyutu.fi).

Fig. 1: 3D-printed cuvette system on a magnetic stirrer. A 1 ml thylakoid sample in the cuvette is covered by a quartz window to prevent equilibration with air. The white temperature sensor and the brown OXB430-OI oxygen sensor from Pyro Science fit tightly into the holes on the cuvette wall and just reach the sample. Water circulation keeps the temperature constant.

Fig. 2: The complete system at work measuring oxygen evolution in pumpkin thylakoids under high light. The optical oxygen meter FireStingO2 from Pyro Science in the front. A slide projector is used here as a source of light for the photosynthetic reactions.

Fig. 3: Logger software from Pyro Science showing the rising oxygen level (yellow) caused by water splitting by the oxygen evolving complex of photosystem II under illumination. The disturbance at ~1315 s is caused by filling the cuvette with the sample, and the rise of oxygen concentration occurs when the light is switched on. The blue line shows the temperature.