Fourth Workshop

Fourth Workshop

LUMINESCENCE & PHOTOSYNTHESIS
OF MARINE PHYTOPLANKTON

Sopot - Mogilany

22-26 September 2003

1. Basic information 2. List of lectures 3. Some abstracts 4. Authors and co-authors of lectures

Bogdan Woźniak - Institute of Oceanology, Polish Academy of Science, Sopot, Poland

Jerzy Dera - Institute of Oceanology, Polish Academy of Science, Sopot, Poland

Andrei B. Rubin - M.V. Lomonosov Moscow State University (Depart. of Biophysics), Moscow, Russia

Vadim N. Pelevin - P.P. Shirshov Institute of Oceanology RAS, Moscow, Russia

Mirosława Ostrowska - Institute of Oceanology, Polish Academy of Science, Sopot, Poland

Ryszard Hapter - Institute of Oceanology, Polish Academy of Science, Sopot, Poland

Mirosław Darecki - Institute of Oceanology, Polish Academy of Science, Sopot, Poland

Sławomir Kaczmarek - Institute of Oceanology, Polish Academy of Science, Sopot, Poland

Maciej Sokólski - Institute of Oceanology, Polish Academy of Science, Sopot, Poland

A.B. Rubin (invited) Regulation of the primary photosynthetic processes of algae

Yu.V. Kazimirko, S.I. Pogosyan Principles of fluorometric methods and equipment for the assessment of the state of photosynthetic apparatus in algae

M. Ostrowska, B. Woźniak Some fluorometric methods of marine photosynthesis study

S.I. Pogosyan, D.N. Matorin, E.N. Volkova, T.K. Antal, Yu.V. Kazimirko, S.V. Vostokov,
A.B. Rubin Fluorescence methods for estimation of phytoplankton condition in the Black Sea

E.N. Voronova, S.I. Pogosyan, E.V. Volkova, Yu.V. Kazimirko Response of Photosynthetic Apparatus of Diatom Thallassiosira weisflogii to High Light Illumination

D.N. Matorin, L.V. Ilyash, B. Woźniak The use fluorescence methods for biological monitoring and biotesting of the effects of environmental pollution

L.V. Ilyash, D.N. Motorin Investigation of distribution and photosynthetic activity of natural phytoplankton in the bay of Nhatrang (South China Sea) with the use of a submersible two-flash fluorometer

M. Koblizek, P.G. Falkowski, Z.S. Kolber Marine aerobic photosynthetic bacteria new metabolic strategy in the ocean

R.M. Siegoczynski (invited) Molecular interactions, fluorescence and radiationless energy transfer of electronic excitation in organic compounds

B. Woźniak, M. Darecki, M. Ostrowska, A. Pacuszka, K. Tyszka (invited) Quantum yield of phytoplankton chlorophyll a fluorescence in marine environment; some questions of determination, modelling and applications

D. Ficek, B. Woźniak New results of the study of photosynthesis quantum yield in the Baltic

R. Majchrowski, B. Woźniak A simplified polynomial methods of determination of phytoplankton light-adaptations and light-absorption capacity, in sea surface layers

T. Król Modeling of absorption and attenuation of light by green algae cells

D.V. Khlebnikov, V.A. Matushenko, V.N. Pelevin Measurements of brightness coefficient of Atlantic waters onboard r/v "Ak. Ioffe" in 2002

V.N. Pelevin, V.V. Rostovtseva Modelling of bio-optical parameters of Atlantic Ocean waters using the completed data of phytoplankton pigments absorption

V.N. Pelevin, O.I. Abramov, G.G. Karlsen, V.V. Pelevin, D.V. Khlebnikov Laser probing of Atlantic Ocean waters from board r/v "Ak. Ioffe" from Kaliningrad to Ushuaia during the three voyages in 2002-2003

A.B. Rubin (invited) Regulation of the primary photosynthetic processes of algae

The regulation of the photosystem II activity in microalgae under the influence of environmental factors, mechanisms of photochemical and non-photochemical quenching of chlorophyll fluorescence and methods for assessment of these processes by measuring kinetics of variable fluorescence of chlorophyll are discussed.

Yu.V. Kazimirko, S.I. Pogosyan Principles of fluorometric methods and equipment for the assessment of the state of photosynthetic apparatus in algae

The using chlorophyll fluorescence parameters such as F₀, F_m, F_v/F_m, qP, qN etc is widely accepted for assessments of functional state and productivity of phytoplankton. The application of this technique requires both adequate instrumentation and algorithms. In this report we present technical characteristics of the fluorometric complex developed in this laboratory consisting of a submersible fluorometer, on-board fluorometer and microfluorometer. The possible applications of the complex in field studies of marine phytoplankton are discussed.

M. Ostrowska, B. Woźniak Some fluorometric methods of marine photosynthesis study

The traditional, direct methods to determine the main photosynthetic characteristic are inefficient and the results widely and randomly scattered in time and space. The poster presents convenient and physically justified methods of determining chlorophyll a concentration and primary production using contact measurements carried out in situ with submersible fluorometers (especially using PumpProbe fluorometer) and some other simple measurements.

The chlorophyll a concentration can be determined from the fluorometric equation using the initial phytoplankton fluorescence F₀, and the optical depth, which is measured with a quantometer. The fluorometric measurements can be carried out with both classical and Pump Probe fluorometers.

The magnitudes of primary production can be determined using the phytoplankton fluorescences: initial F₀(z) and maximal F_m(z), which can be measured only with a Pump Probe fluorometer. Also required are measurements of the temperature temp (with a thermometer connected to the fluorometer) and measurements of the irradiance PAR(z) performed with a quantometer calibrated in absolute units. The determination of the primary production during longer periods of time will also require additional pyranometric measurements.

The methods presented here are far more accurate than the previous ones, which were based on the statistical correlations between photosynthetic characteristics and measured fluorescences.

The results of empirical verifications showed that accuracy of elaborated methods is similar to accuracy of traditional methods of chlorophyll and primary production determination.

Hence the necessity for the introduction of fluorometric techniques to oceanological practice, especially as they are much simpler and more efficient than traditional measurements.

S.I. Pogosyan, D.N. Matorin, E.V. Volkova, T.K. Antal, Yu.V. Kazimirko, S.V. Vostokov,
A.B. Rubin Fluorescence methods for estimation of phytoplankton condition in the Black Sea

The functional state of phytoplankton community was studied in coastal and open regions of the north-eastern Black Sea by means of fluorometric methods. Numerous data on phytoplankton fluorescence properties determination in euphotic layer of the sea were obtained. The phytoplankton pigment content, photosynthesis efficiency, water temperature and underwater irradiance (PAR) were plotted. Fluorescent characteristics of photosynthesis efficiency were measured for individual sells of the most abundant algae species in the Blue and Gelendzhik Bays. Dependence of nonphotochemical fluorescence quenching of phytoplankton chlorophyll on depth was also explored. Application of different fluorometric method for the Black Sea ecosystem modern state analysis and future changes forecasting was discussed.

E.N. Voronova, S.I. Pogosyan, E.V. Volkova, Yu.V. Kazimirko Response of Photosynthetic Apparatus of Diatom Thallassiosira weisflogii to High Light Illumination.

The response of the photosynthetic apparatus to high light illumination (440-2200 W^.m^-2) was studied on diatom Th. weisflogii algae by measuring characteristics of chlorophyll fluorescence. Changes in photosynthetic apparatus were revealed by measuring chlorophyll fluorescence parameters F₀, F_m, F_v/F_m and qN for several hours after illumination of the alga with high light. Incubation of the alga with an inhibitor of the deepoxydase of diadinoxanthin cycle carotenoids (2 mM dithiothreitol -DTT) led to a decrease in the non-photochemical quenching of chlorophyll fluorescence and a drop in the F_v/F_m ratio, that reflects the quantum efficiency of the primary light energy conversion in photosynthesis. Light-induced absorption changes associated with transformations of carotenoids of diadinoxanthin cycle were recorded in vivo in algal suspension in the absence and in the presence of DTT. Using microfluorometric method, we measured cell distribution over the efficiency of the primary processes of photosynthesis (F_v/F_m) after illumination. We found cells with a high tolerance of their photosynthetic apparatus to photooxidative damage. The relatively high tolerance of a portion of the cell population to high light illumination can be related to the light-induced transformation of carotenoids and to functioning of other protective systems of the photosynthetic apparatus in diatoms.

D.N. Matorin, L.V. Ilyash, B. Woźniak The use fluorescence methods for biological monitoring and biotesting of the effects of environmental pollution

Luminescence methods, developed at the Dep. of Biophysics, Moscow State University and IO PAN, and designed for diagnostics of the physiological state of algae are considered. Series of the field studies with the use of a submersible fluorometer, devices for measuring characteristics of delayed fluorescence and thermoluminescence were performed to assess phytoplankton state in diverse regions of the world ocean, Baikal and Issyk-kul Lake. The application of these methods and instruments in natural water bodies, especially for the testing of the impact of anthropogenic pollution, as well as for bioassay of water quality is discussed.

L.V. Ilyash, D.N. Matorin Investigation of distribution and photosynthetic activity of natural phytoplankton in the bay of Nhatrang (South China Sea) with the use of a submersible two-flash fluorometer

The species composition, abundance of phytoplankton (B), chlorophyll concentration (Chl), the vertical distribution of fluorescence parameters (Fo, Fv/Fm), temperature and irradiance were evaluated in bay of Nhatrang. A according B and Chl the Bay is mezotrophic. Diatoms dominated in species diversity and abundance. B and Chl in bottom layer exceeded those in upper layer. The diel dynamic of Fv/Fm showed that surface phytoplankton was photoinhibited during the most part of light period, while at the bottom photosynthetic activity was high. Water column B changed in order of magnitude during diel cycle mainly because of change B in the bottom layer.

M. Koblizek, P.G. Falkowski, Z.S. Kolber Marine aerobic photosynthetic bacteria new metabolic strategy in the ocean

Recently, the presence of bacterial photosynthesis was registered in the open ocean by means of infra-red fast repetition rate fluorometry (Kolber et al. Nature 407, 177, 2000, Kolber et al. Science 292, 2492, 2001). We isolated about 30 strains of strictly aerobic, anoxygenic, bacteriochlorophyll-containing bacteria isolated from surface waters of Atlantic, Pacific and Indian Oceans, the Black and the Mediterranean Seas. We characterized their pigment and lipid composition, spectroscopic properties, substrate preferences, as well as 16S rDNA and pufM gene sequences. The phylogenetic analyses place all the isolates within the alpha 4 and alpha 3 subclass of Proteobacteria. The functionality of the photosynthetic apparatus was proven by kinetic bacteriochlorophyll fluorescence measurements and CO2 fixation assays. However, these bacteria are not capable of truly autotrophic growth, as they require a supply of organic carbon. The presence of the photosynthetic apparatus seems to represent an ecological advantage in nutrient poor marine environments serving as an alternative source of ATP for basal metabolism. The mixed photoheterothophic metabolism represent a novel component in the marine carbon cycle.

R.M. Siegoczynski (invited) Molecular interactions, fluorescence and radiationless energy transfer of electronic excitation in organic compounds

The electronic transport properties of organic compounds especially of aromatic hydrocarbons have been studied extensively in recent years, but a satisfactory understanding of the basic mechanism of transport is still lacking. In molecular crystals and films the electron exchange interactions are much weaker and the electron mobility is correspondingly much lower than in inorganic semiconductors. The electron-phonon interactions, on the order hand, are expected to be much the same in both types of materials. The question therefore arises whether in molecular crystal or films the electron-phonon interaction can be still treated as a small perturbation to the quasi-free-electron wave function. The proper zero-order solution often leads to a self-trapped or quasi-localized electron, which in the next order of approximation makes random jumps to neighbouring molecules, a mechanism known as hopping.

Transfer of electronic excitation energy in organic molecular crystals (aromatic hydrocarbons) is well-known phenomenon. Energy transfer takes place also in amorphous organic polymers especially in polyvinyl carbazole (PVCz), both in solution and the solid state. Through monitoring the fluorescence properties in PVCz as a function of different types of fluorescence quenching substances as benzanthrone (Bt), it was shown the dominant mechanism for energy transfer is exciton diffusion.

The transfer of electronic excitation energy from the monomer chromophore sites where absorption occurs to the dimmer, excimer, exciplex and charge-transfer sites where emission occurs has been the subject of our investigation. We propose here the hopping model of exciton diffusion which explains the dependence of monomer fluorescence intensity on the concentration of excimer sites and charge-transfer fluorescence intensity in presence of guest molecules in strong electric fields.

D. Ficek, B. Woźniak New results of the study of photosynthesis quantum yield in the Baltic

R. Majchrowski, B. Woźniak A simplified polynomial methods of determination of phytoplankton light-adaptations and light-absorption capacity, in sea surface layers

The aim of this work was to find simple methods of determination of phytoplankton light absorption coefficient for total phytoplankton pigments, for photosynthetic pigments and for photoprotecting pigments, in sea surface layers.

On the basis of complicated model of the in vivo spectral absorption of algal pigments presented in Oceanologia 42(2) (Woźniak et al., 2000; Majchrowski et al., 2000) phytoplankton spectral absorption coefficients in the surface water were calculated. These coefficients were calculated for given total chlorophyll concentrations C_a(0) and irradiance of photosynthetically available radiation just below the water surface PAR(0⁺).

To make the calculation easier we derived polynomial methods of determining light spectral absorption coefficients of: phytoplankton a_pl, photosynthetic pigments a_pl,PSP and photoprotecting pigments a_pl,PPP, in sea surface layers.

In all cases the errors are relatively small (for example, the statistical error in all cases was < 4 %).

T. Król Modeling of absorption and attenuation of light by green algae cells

Based on the pigments absorption spectra and the estimated pigment composition in phytoplankton cells the distribution of the values of the complex refraction index of light by the phytoplankton cell or its inner structures were determined.

The microscopic measurements allowed to estimate the cell and inner structures size distribution of the examined phytoplankton population.

Then the Mie solution gives the light attenuation and absorption cross-section distribution for spherical-shaped targets that model the cells and their inner structures. Such spectra averaged according to cell-size distribution result in model light absorption and attenuation spectra of phytoplankton culture and its inner structure. This modeling was realized for two similar spherical cells of green algae Chlorella vulgaris and Chlorella kesleri differing in size from each other as well.

D.V. Khlebnikov, V.A. Matushenko, V.N. Pelevin Measurements of brightness coefficient of Atlantic waters onboard r/v "Ak. Ioffe" in 2002

V.N. Pelevin, V.V. Rostovtseva Modelling of bio-optical parameters of Atlantic Ocean waters using the completed data of phytoplankton pigments absorption

The data obtained from board a research vehicle “Akademik Ioffe» during its voyage across the Atlantic Ocean in 2002 are used to estimate the concentrations of optical-biological admixtures. The optical properties of the Atlantic waters were studied by three different passive methods. The spectra of the radiance coefficient of water surface were obtained with the special three-channeled spectrophotometer. The spectra of the water extinction coefficient were measured in the ship laboratory for numerous water samples and their filtrates. The spectra of water absorption coefficient were obtained for the filtrates with the comparative spectrophotometer.

The whole data set was used to create “the optical portraits” of the different areas of the Atlantic. Three basic areas are discussed: the central Atlantic, the La-Plata mouth area, the Baltic Sea.

The new completed model of phytoplankton pigment absorption developed in IOPAS is used for interpretation of these “optical portraits” and for estimation of the three main natural admixtures concentrations: phytoplankton pigment, “yellow substance” and suspended matter.

The results can be used for developing the detailed optical sea-water models. They are also useful for improving satellite measurements.

M.V. Lomonosov Moscow State University (Depart. of Biophysics), Moscow, Russia
L.V. Ilyash
Yu.V. Kazimirko
D.N. Matorin
S.I. Pogosyan
A.B. Rubin
E.V. Volkova
E.N. Voronova
S.V. Vostkov

P.P. Shirshov Institute of Oceanology RAS, Moscow, Russia
O.I. Abramov
D.V. Khlebnikov
V.A. Matushenko
V.N. Pelevin
V.V. Pelevin
V.V. Rostovtseva

Russian Elektrotechnic Institute, Moscow, Russia
G.G. Karlsen

Institute of Microbiology (Lab. of photosynthesis), Opatovicky Mlyn, Trebon, Czech Republic
M. Koblizek

Rutgers University (Inst. of Marine and Coastal Sciences), New Brunswick, USA
P.G. Falkowski
Z.S. Kolber

Institute of Oceanology PAS, Sopot, Poland
M. Darecki
J. Dera
T. Król
M. Ostrowska
B. Woźniak
A. Pacuszka
K. Tyszka

Pomeranian Pedagogical Academy (Depart. of Environment. Physics), Słupsk, Poland
D. Ficek
R. Majchrowski

Warsaw Technical Univ. (Faculty of Physics), Warsaw, Poland
R.M. Siegoczyński