Fig. 2: Electron donation for O₂ photoreduction in early spring thylakoid membranes.

a ¹⁶O₂ yields b ^16,18O₂ (note that O₂ consumption in the mixed labeled O₂ was small due to the extremely low natural abundance of ^16,18O₂ compared to ¹⁶O₂) in S and ES pine thylakoid membranes supplemented with 25 µM DCMU measured by TR-MIMS under partially degassed conditions in the presence of 250 µM PPBQ and 500 µM FeCy. Gray-shaded regions indicate dark periods before and after illumination of the thylakoid membranes with 1200 µmol of photons m⁻² s⁻¹ for 60 s. For TR-MIMS (10% H₂¹⁸O enrichment), one representative spectrum out of 3–5 independent measurements is shown. c Intersystem e-/P700 pool measured as a ratio between area under MT and area under ST flash, applied under 200 µmol of photons m⁻² s⁻¹ constant FR-light (720 nm) illumination (n = 3) from S and ES intact needles. d Chlorophyll fluorescent signal recorded from S (n = 2) and ES (n = 3) intact needles under constant actinic red light (320 µmol of photons m⁻² s⁻¹) illumination for 5–6 min followed by 120–180 s of dark relaxation (Time scale is set to ‘0’ upon actinic light off). c, d n = number of measurements where needles were pooled from 5 trees. e Redox state of the PQ pool presented as the ratio of plastoquinol (PQH2) to plastoquinone (PQ) (n = 9) from dark-adapted S and ES thylakoids where n defines independent measurement replication (n = 3) of individual biological replicates (n = 3). Statistically significant mean differences were calculated t test (p < 0.05) (Supplementary Table 5). f O₂ yields in S and ES thylakoid membranes in air-saturated buffer measured by a Clark-electrode with 1 mg ml⁻¹ HgCl₂ supplementation in the absence of PPBQ and µM FeCy (-A). The colored shaded regions around O₂ yield curve (mean) indicates ±SEM (n = 3) where n = biological replicates. Data represents O₂ exchange corresponding to the same chlorophyll content (50 µg) for both Clark-type electrode and MIMS measurements. O₂ exchange rates from Clarke-electrode are given in Supplementary Fig. 11. c, e The box bounds indicate ±SEM, minima/maxima indicates ±SD, middle line in the box indicates mean and the dots indicate data points. Source data are provided as a Source Data file.