abstract 1. introduction 2. methods

Short-Period photophysiological responses of Thalassiosira pseudonana during photoacclimation to near surface irrradiance Robert E. Reed Dept. of Marine, Earth, and Atmospheric Sciences North Carolina State University, Raleigh, N.C. 27695 Gary J. Kirkpatrick Mote Marine Laboratory 1600 Ken Thompson Parkway, Sarasota, FL 34236 Dan Kamykowski Dept. of Marine, Earth, and Atmospheric Sciences North Carolina State University, Raleigh, N.C. 27695

ABSTRACT A two day deployment of the Self-Contained Underwater Photosynthetic Apparatus (SUPA) was conducted to examine the effects of high and variable natural irradiance on the optical properties and primary productivity of the diatom Thalassiosira pseudonana (clone 3H). Study objectives included the determination of short time responses in the cycling of diadinoxanthin(DD) and diatoxanthin(DT), and associated changes in the rates of primary production, and quantum yield, and in cell absorption characteristics over a two day period. A nutrient replete, low light acclimated diatom culture was placed in SUPA and in a collocated culture reservoir fitted with a quartz top. The instrument was deployed in shallow water in Sarasota Bay under fluctuating, high irradiance typical for June. A pigment sampling series reveals a correlation of DD to DT cycling with exposure to high irradiance. Net quantum yields, determined by cell absorption spectra and minute-to-minute SUPA net primary productivity values, exhibit high values initially which decay to lower values upon exposure to high light. Short time scale changes in quantum yield are observed due to changes in DD/DT cycling and fluctuating irradiance.

1. INTRODUCTION The ability of a phytoplankter to acclimate to high and variable irradiance levels is an important factor in determining growth rates and overall success.10'16 Short time scale responses due to fluctuating light are not easily determined by traditional methods (e.g. radioactive carbon uptake) because the sample handling time exceeds the time scale of response. The biophysical instrument SUPA simultaneously can determine minute-to-minute changes in carbon uptake rates and oxygen evolution rates by a unialgal culture in situ2. Since, the light (oxygenic photosynthesis) and dark (Calvin cycle) reactions of photosynthesis are not directly coupled, divergences can occur in the rates of the two photosynthetic reactions. This research attempts to accurately determine changes in the T. pseudonana photosynthetic rates as measured by SUPA on a per minute basis and pigment complement as measured on periodic samples for two consecutive diurnal cycles under high and variable irradiance.

2. METHODS Unialgal batch cultures of T. pseudonana (clone 3H) were grown on a 12:12 hour light-dark cycle in F/2 growth media. ' A mixture of cool-white, and Vita-lite fluorescent lamps provided approximately 135 uEin m": s"1 of photosynthetically active radiation (PAR). All PAR measurements were made with a Biospherical Instruments, Inc. model QSL-100 quantum scalar irradiance sensor. The cultures were diluted to F/20 prior to loading into SUPA and the 18 liter reservoir. The SUPA dome and top of the enclosure was constructed of quartz glass to allow the penetration full spectrum irradiance. Changes in ambient irradiance, culture temperature, pH and dissolved oxygen are recorded by SUPA on a oneminute basis. All rates of change were calculated by algorithms developed for the specific parameters.' In vivo OD spectra collected on glass fiber filters were determined using a Varian model DMS-80 UV-VIS spectrophotometer. The resulting

spectra were corrected for multiple scattering using the method of Cleveland and Weidemann3. The spectral fourth derivative was then computed for each spectrum to resolve the position and any shifts in absorption peaks.4 Pigment analysis methods follow Mantoura and Llewellyn5 with detection on a Shimadzu SPD-M6A photo-diode array detector and quantification at 440nm. Cell numbers were enumerated using both Coulter Counter and hemacytometer methods. The SUPA and associated enclosure were deployed in Sarasota Bay on June 25, 1995 off the Mote Marine Laboratory dock. The sample chamber was approximately at 1 meter depth at high tide. The array was sampled at deployment and approximately every two hours thereafter via a darkened tube connected to a peristaltic pump. A 3m x 3m neutral density screen was constructed to provide some protection to the shade adapted culture if deemed necessary. Surface irradiance was recorded by the Mote Marine Laboratory weather station. In order to determine the fine scale changes in pigment complement due to short term irradiance fluctuations, the HPLC and cell absorption samples received the highest priorty for processing. Both were immediately filtered with low vacuum and frozen in liquid nitrogen.

3. RESULTS 3.1 Photosynthetic rate response to irradiance fluctuations A two day, minute-to-minute record of oxygen evolution and carbon uptake by T. pseudonana was collected by SUPA. If one takes the light field experienced as the input and the rate of carbon uptake or oxygen evolution in the context of variable and high irradiance exposure as the output of the system, the rates either mimic the light field or spikes and noise seem to predominate. For example, the carbon uptake rate (Fig. 1A & IB) after 1720 min. elapsed time, closely resembles the light input signal. Prior to this time, large swings in uptake occur on a regular basis that appear independent of the light. Also, the oxygen evolution data (Fig. 1C & ID) exhibit noise during the first day; however, the second day seems more stable and rate features more closely match the light signal. These gross features will be examined later in the context of cellular photoadaptative responses. Day One Carbon Uptake Rate

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