Nadine Schubert Instituto de Ciencias del Mar y Limnología de la UNAM

1. PHOTOBIOLOGY Nadine Schubert Instituto de Ciencias del Mar y Limnología de la UNAM Unidad de Sistemas Arrecifales, Puerto Morelos, México

2. WHAT DOES PHOTOBIOLOGY MEAN? Photosynthesis Photomorphogenesis Cirvadian Rhythm Ultraviolet Radiation

3. PHOTOBIOLOGY Part 1: Photosynthesis and Fluorescence Part 2: Photoacclimation/-adaptation Part 3: Photoprotection

4. Part 1: Photosynthesis and Fluorescence

5. PHOTOSYNTHESIS

6. LIGHT ABSORPTION

7. THE PHOTOSYNTHETIC APPARATUS

8. THE PHOTOSYNTHETIC APPARATUS ATPase PSII LHCII Cyt bf PSI LHCI

9. LIGHT ABSORPTION The absorbed light energy is funneled by excitation transfer into the RC’s, where energy conversion by charge separation takes place. Antenna pigments Antenna pigments PS II PS II Photochemistry Photochemistry

10. LIGHT ABSORPTION photon excited state molecule absorbs photon Increasing energy ground state

11. EXCITATION ENERGY TRANSFER Electron transfer Excitation transfer Acceptor Light e- Reaction Center e- Donor Antenna

12. ELECTRON TRANSFER ATP ADP + Pi NADPH NADP + H+ 2H+ Fd ATPase PQH2 PSII LHCII Cyt bf PSI LHCI PQ PC H+ 2H2O O2+ 4H+ 2H+

13. LIGHT ABSORPTION AND ENERGY TRANSFER

14. PHOTOSYNTHESIS AND FLUORESCENCE

15. PHOTOSYNTHESIS AND FLUORESCENCE photon excited state excited state Photochemistry molecule absorbs photon Fluorescence Heat ground state ground state

16. PHOTOSYNTHESIS AND FLUORESCENCE Antenna pigments Heat Fluorescence PS II Photochemistry

17. PHOTOSYNTHESIS AND FLUORESCENCE Antenna pigments Non-light -tress conditions Heat Fluorescence PS II Photochemistry

18. PHOTOSYNTHESIS AND FLUORESCENCE Photochemistry = 1 Fluorescence = 0 Photochemistry = 0 Fluorescence = 1 Whitmarsh & Govindjee (2002)

19. CHLOROPHYLL FLUORESCENCE MEASUREMENT PS = 0 NPQ = 0 Fv/Fm = (Fm-Fo)/Fm Fm = maximum fluorescence (RC’s closed) Fo = minimum fluorescence (RC’s open) (higher plants – 0.85, macroalgae usually lower) PS = 1 NPQ = 0

20. Fv/Fm – MAXIMUM QUANTUM YIELD Quantum yield: Probability that the energy of a photon absorbed will be used for photosynthesis (i.e. enters in the e- - transport chain)  Indicator of photosynthetic efficiency Maximum quantum yield:requires complete relaxation of the competing mechanisms with the photochemical energy conversion

21. Fv/Fm – Diurnal and spatial variation Chondrus crispus Depth (m) Colombo-Pallotta (2007) Macrocystis pyrifera Hanelt et al. (1992)

22. Fv/Fm – Comparison of stress responses between species Littoral Sublittoral Sublittoral Sublittoral Littoral Sublittoral van de Poll et al. (2001)

23. CHLOROPHYLL FLUORESCENCE MEASUREMENT Fv/Fm F/Fm’ PS = 0 1  NPQ  0 1  PS  0 1  NPQ  0

24. F/Fm’ – EFFECTIVE QUANTUM YIELD Used to describe the variation in the photochemical efficiency of PSII under illuminated conditions. Measurement of this parameter at certain irradiance value.  Indicator of the ability of an organism to move electrons beyond PSII (ETR) F/Fm’ = (Fm’-F)/Fm’

25. ELECTRON-TRANSPORT RATE (ETR)– CURVES ETR = Irradiance  F/Fm’  0,5  Absorptance (Genty et al. 1989) F/Fm’ = effective quantum yield (under light) 0,5 = Assumption that 50% of these quanta are absorbed by PSII Absorptance = fraction of incident light that is absorbed by the photosynthetic tissue. Not the same as absorbance (quantifies how much of the incident light is absorbed by an object).

26. ELECTRON-TRANSPORT RATE (ETR)– CURVES • ETR = Irradiance  F/Fm’  0,5  Absorptance • Relative ETR = Irradiance  F/Fm’  0,5(Ralph et al. 2002) • ETR: when absorption characteristics change between species, acclimations, seasons… • - rel. ETR: use only when it is sure that there are no differences in the absorption characteristics

27. ETR– CURVES AS AN ANALOGUE TO P-E- CURVES Macrocystis pyrifera Colombo-Pallotta et al. (2006)

28. CHLOROPHYLL FLUORESCENCE • EXTENSIVELY USE DUE TO: • NON-DESTRUCTIVE • NON-INVASIVE • RAPID • SENSITIVE • IN REAL-TIME • Since 1995 the number of articles published applying chlorophyll fluorescence on the analysis of the photosynthetic performance in macroalgae and seagrasses has increased more than five times.

29. FLUOROMETERS • The Chl fluorometer should be capable of measuring the fluorescence yield in a non-intrusive way: • very low measuring light (i.e. exciting light) intensity for assessment of the fluorescence yield of a dark-adapted sample • the detection system has to be very selective to distinguish between fluorescence excited by the measuring light and the much stronger signals caused by ambient and actinic light (full sun light, saturating light pulses for assessment of maximum fluorescence) • fast time response to resolve the rapid changes in fluorescence yield upon dark-light and light-dark transitions • PAM fluorometers: Pulse-Amplitude-Modulated fluorometers

30. Pulse-Amplitude-Modulated Fluorometers Distinguish between fluorescence and ambient light  Allows measurement of fluorescence in the presence of actinic light (light absorbed by the photosynthetic apparatus to drive photosynthesis) How? – Measuring light is modulated and the fluorescence amplifier is highly selective for the modulated signal (yield of chlorophyll fluorescence) - pulse-modulated measuring light can be generated either by a light-emitting diode (LED; most PAM fluorometers) or a flash discharge lamp (i.e. XE-PAM)

31. Pulse-Amplitude-Modulated Fluorometers DUAL-PAM IMAGE-PAM MINI-PAM XE-PAM DIVING-PAM