Plankton Biol. Ecol. 48 (2): 96-103, 2001 plankton biology & ecology

£ The Plankton Society of Japan 2001

Relationship between light absorption and the -cycle pigments in marine diatoms

Tetsuichi Fujiki & Satoru Taguchi

Laboratory of Biological Oceanography, Department of Bioengineering, Faculty of Engineering. Soka University, 1-236 Tangi-

cho, Hachioji 192-8577, Japan

Received 6 March 2001; accepted 13 May 2001

Abstract: The relative contribution of the xanthophyll-cycle pigments, plus diatoxan thin, to total in vivo pigment absorption was examined in 3 species of marine diatoms grown at 6 ir- radiances between 90 and 750^mol m"2s"1. These diatoms included Phaeodactylum tricornutum, Chaetoceros gracilis and Thalassiosira weissflogii. The pigment ratios of diadinoxanthin plus diatox- anthin to chlorophyll a (Chi a) for all 3 species increased with increasing growth irradiance. For P. tri cornutum, the ratio was 5.5 times higher in cells grown under 750jt/mol m~2s~1 compared to those grown at 90^mol m~2s"1. As the growth irradiance varied from 90 to 750^mol m~2s~\ the relative contribution of absorption by diadinoxanthin plus to total in vivo pigment absorption (for wavelengths from 400 to 700 nm [PAR]) increased from 4.5 to 17% for P. tricornutum, 5.8 to 19% for C. graciiis and 13 to 30% for T. weissflogii. The high contribution of absorption by diadinoxanthin plus diatoxanthin to total in vivo pigment absorption was observed between 450 and 500 nm, in which the contribution of Chl-a absorption was minimal. The maximum contribution of absorption by diadinoxanthin plus diatoxanthin to total in vivo pigment absorption was 57% at 462 nm in T. weiss flogii grown under 750//mol m~2s~1. For all species, a significant negative relationship was obtained between absorption of diadinoxanthin plus diatoxanthin and quantum yield for growth. This relation ship may suggest that the xanthophyll cycle dissipates the excess light energy and helps to maintain growth rate under high light conditions.

Key words: absorption spectra, quantum yield, spectral reconstruction technique, xanthophyll cycle

photoprotective systems of . This cycle can dis Introduction sipate the excess light energy by mutual transformations Phytoplankton can adapt lo changes in light quality (in between epoxy-containing (oxy-derivatives of tensity and/or spectrum) by varying the total amount of cel ) and epoxy-free xanthophylls (Hager 1975). In lular pigments, the ratio of different pigments, or both. The higher plants and chlorophytes, this cycle contains the 3 light energy absorbed by pigments is either used in photo components of , and synthesis, re-emitted as fluorescence or dissipated as heat (Hager 1975), and the formation of zeaxanthin helps in the (Falkowski & Raven 1997). In stressful high light condi process of dissipation of excess energy (Demmig et al. tions, if phytoplankton cells are unable to utilize the high 1987). The conversions are mediated by a reversible light level of energy absorbed by pigments, the excess energy epoxidizing enzyme in the thylakoid membranes that uses a will cause damage to intracellular materials or metabolic pH gradient across the lumen membrane (Gilmore & Ya- processes, e.g. destruction of membranes or in- mamoto 1993). The xanthophyll pigments, diadinoxanthin activation of enzymes. Some carotenoids can serve as pho- and diatoxanthin, found in chromophyte algae such a di toprotection against such photoinhibition by quenching the atoms, dinoflagellates and prymnesiophytes have recently excess energy (Vincent et al. 1984; Cogdell & Frank 1987; been reported to possess a similar photoprotective system Demmig-Adams 1990). The xanthophyll cycle is one of the to the xanthophyll cycles of higher plants and chlorophytes (Demers et al. 1991; Arsalane et al. 1994; Olaizola & Ya-

Corresponding author: Tetsuichi Fujiki; e-mail, [email protected] mamoto 1994; Moisan et al. 1998). When the algae are ex- Light Absorption and Pigments in Diatoms 97

posed to stressful high irradiance, some diadinoxanthin is lar to that described by Head & Home (1993). Integrated de-epoxidized to diatoxanthin within 5min. Light energy peak areas were quantified with external standards obtained absorbed by xanthophyll cycle pigments is not transferred from the International Agency for I4C Determination. to the reaction centers for photosynthesis, but is dissipated by mutual transformations between diadinoxanthin and dia Particulate carbon contents toxanthin. What is not well understood at present is how the xanthophyll cycle pigments contribute to cellular light ab Triplicate samples were filtered onto precombusted sorption. It is imperative to study quantitatively the photo- Whatman GF/F filters, dried at 60°C for 24 h, and analyzed protection process by diadinoxanthin and diatoxanthin. with an elemental analyzer (FISON Instrument NA1500). The spectral reconstruction technique for phytoplankton Acetanilide was used as an external standard. absorption can provide information on the relative contribu tion of individual pigments to total in vivo pigment absorp Light absorption tion (Bidigare et al. 1989; Hoepflfner & Sathyendranath Phytoplankton absorption spectra were measured with 1991; Babin et al. 1996). In this study, we have attempted to the glass fiber filter technique (e.g. Truper & Yentsch 1967; address this question by examining the contribution of ab Mitchell & Kiefer 1988; Roesler et al. 1989). Triplicate sorption by diadinoxanthin plus diatoxanthin to total in vivo samples were collected on Whatman GF/F filters at <50 pigment absorption for diatoms grown under high light mmHg. Optical density spectra were recorded from 400 to conditions using the spectral reconstruction technique (as 750 nm with a Beckman DU640 spectrometer using a wet suming that all diadinoxanthin and diatoxanthin is related Whatman GF/F filter as a blank. Opticaldensity spectra to the xanthophyll cycle). were converted to absorption coefficients ap(X) by subtract ing the optical density at 750 nm from all wavelengths, di viding by the geometrical path length (ratio of filtered vol Materials and Methods ume to the filtered clearance area of the filter) and adjusted Cultures for path length amplification. The path length amplification factor described by Cleveland & Weidemann (1993) was Cultures of the diatoms Thalassiosira weissfiogii employed for all absorption spectra. Pigments were then ex [Grunow] G. Fryxell & Hasle (NEPCC741), Chaetoceros tracted from filtered samples using cold methanol (Kishino gmcilis Schutt (NEPCC645) and Phaeodactylum thcornu- et al. 1985), and the absorption of the remaining particles tum Bohlin (CCMP1327) were grown semicontinuously in was measured (defined as the nonpigment absorption coef f/2 medium (Guillard & Ryther 1962) at 25°C. T. weiss ficient, od(A)). The phytoplankton absorption coefficient, fiogii and C. gracilis were obtained from The North East aph(A), was calculated as the difference between ap(A) and Pacific Culture Collection (NEPCC) at the University of ad(X). Finally, aph(X) was converted to the Chl-a-specific British Columbia, Canada. P. tricornutum was obtained absorption coefficient, a*(A), by dividing by the Ch\-a con from the Provasoli-Guillard National Center for Culture of centration. Marine Phytoplankton (CCMP), Bigelow Laboratory for Marine Sciences, Maine, USA. Cultures were exposed Spectral reconstruction of absorption coefficients under cool fluorescent lights ranging from 90 to 750 fimo\ m~2s~' on a 12:12 h light/dark cycle. Growth irradiance, /,„ Absorption spectra of pure pigment standards were was measured with a scalar quantum sensor (QSL-100, wavelength-shifted to match their in vivo absorption peaks Biospherical Instruments Inc.). The relative spectral quan and shoulders using the approach described by Bidigare et tum irradiance E'(X) was measured with a spectroradiome- al. (1990). Then, absorption spectra of the 6 pigments were ter (MSR-7000, OPTO Research Corporation). Cell densi scaled to their respective weight-specific absorption coeffi ties of the three species were determined using a micro cients to give in vivo weight-specific absorption coefficients scope and a hematocytometer. Specific growth rate (ft) was (Fig. 1). Reconstructed absorption coefficients were esti calculated over the exponential growth phase. The cells mated as the product of the volume-based pigment concen grown in light/dark cycles were harvested at the end of the trations and in vivo weight-specific absorption coefficients. photoperiod to compare photoadaptation responses due to The spectral reconstruction technique for phytoplankton ab differences in growth irradiance. sorption does not take package effectinto consideration. Moreover, there arealso some problems regarding spectral Pigment concentrations shape and molarity when compared with in vivo absorption (Bidigare et al. 1989; Hoepffner & Sathyendranath 1991; Triplicate samples were filtered onto Whatman GF/F Moisan & Mitchell 1999). However, if the spectral recon glass fiber filters and immediately immersed in 90% ace struction technique is used in combination with the glass tone. Extraction of pigments was conducted for 24 h in the fiber filter technique described above, one can obtain the dark at 4°C after grinding and sonicating the cells harvested contribution of absorption by individual pigments to totalin by filtration. Subsamples were analyzed onan HPLC (Sys vivo pigment absorption and this overcomes these problems tem Gold, Beckman) usinga solvent gradient system simi 98 T. Fujiki & S. Taguchi

(Babinetal. 1996). following equation, The relative contribution of absorption by diadinoxanthin JnE'(X) plus diatoxanthin to total in vivo pigment absorption for (2) J»/uu wavelengths from 400 to 700 nm (PAR), cDD+DT, was calcu E'(X)dX lated by the following equation, 400

J»700 where E'(X) is the relative spectral quantum irradiance fl*(A)[flDD+DT (X)/aT[,(X)]E0{X)dX measured with a spectroradiometer. _ 400 (1) J»/uo a*(X)El)(X)dX Results

400 Specific growth rate, fi, ranged from 0.64 to 0.98 d"1 for where aDD+DT(X) is the reconstructed absorption coefficient P. tricornutum, 0.85 to 1.33 d"1 for C. gracilis and 0.66 to of diadinoxanthin plus diatoxanthin, aTP(X) is the total re 1.14d~' for T. weissflogii (Table 1). The optimal irradiance constructed absorption coefficient of the six pigments providing the maximum specific growth rate differed found in marine diatoms (cf. Fig. 1). E0(X) is the spectral among the 3 species. Cellular carbon contents (CccII) ranged quantum scalar irradiance. E0(X) was determined by the from 7.3 to 10.3 pg cell"1 for P tricornutum, 20.1 to 26.6 pg cell"1 for C. gracilis and 122 to 155 pg cell"1 for T. weiss flogii (Table 1). The Ccdl of P. tricornutum was inversely re Chi c lated to irradiance (/?<0.05). The Cccl, of the other two species did not show such a relationship. Cellular Chl-o content (Chi acell) of all 3 species decreased exponentially with increasing irradiance (p<0.0\) (Table 1). For all three species, the ratio of cellular Chl-a content to carbon content (Chi a: C) decreased exponentially with increasing irradi ance (p<0.05) (Table 1). For all 3 species, Chi c, , diadinoxanthin, dia 400 toxanthin and j3- were measured as accessory pig ments. All three species showed no change in the pigment Wavelength (nm) ratios of Chi c, fucoxanthin and j8-carotene to Chi a in re

Fig. 1. In vivo weight-specific absorption coefficients (a-*) for sponse to irradiance, except for the ratio of Chi c to Chi a the major pigments found in marine diatoms using the approach which decreased with increasing irradiance for T. weiss described by Bidigare et al. (1990). Chi a=chlorophyll a; Chi flogii (p<0.05) (Table 2). The ratios of diadinoxanthin plus c=chlorophyll c; Fuco=fucoxanthin; j3-caro=/3-carotene; DD= diatoxanthin to Chi a of all species increased with increas diadinoxanthin; DT=diatoxanthin. The mean absorption coeffi ing irradiance (p<0.05). For example, this ratio for P. tri cient of DD and DT was used because the weight-specific absorp cornutum was 5.5 times higher in the cells grown under tion coefficients of DD and DT were similar. 750jUmolm~2s"1 compared to the cells grown under

Table 1. Variations in growth rate (jU), cellular carbon content (CccI|) and Chl-a content (Chi aceM) and the ratio of cellular Chl-o content to carbon content (Chi a: Q for 3 species of diatom grown under 6 irradiances. fi in d"1, Ccel, and Chi «„„ in pg cell"1.

Irradiance (//mol m 2s ') Species Variahles 90 190 330 450 600 750

Phaeodactylum tricornutum fi 0.67 0.64 0.98 0.87 0.90 0.90 Cci 10.3 10.0 8.78 8.10 8.74 7.27 Chi «ccll 0.36 0.30 0.22 0.21 0.17 0.14 Chi a: C 0.035 0.030 0.025 0.026 0.020 0.019 Chaetoceros gracilis 0.85 0.97 0.91 1.35 1.28 1.27 Qell 21.3 20.1 26.4 25.3 26.6 25.5 Chi acd, 0.98 0.67 0.65 0.63 0.49 0.36 Chi a: C 0.046 0.033 0.025 0.025 0.018 0.014 Thaiassiosira weissflogii 0.66 1.13 1.10 1.10 1.14 1.04 Cecil 138 122 146 155 143 139 Chi

Table 2. Ratios (w: w) of various accessory pigments to cellular Chl-o for 3 diatom species grown under different irradiances. Chi c= chlorophyll c; Fuco=fucoxanthin; /3-caro=j3-carotene; DD+DT=diadinoxanthin+diatoxanthin.

Irradiance (/jmolm~2s~')

uuvvivj

90 190 330 450 600 750

Phaeodactylum tricorn Mum Chic 0.139 0.150 0.162 0.159 0.145 0.165 Fuco 0.617 0.714 0.814 0.722 0.649 0.944 j3-caro 0.007 0.007 0.007 0.006 0.006 0.007 DD+DT 0.077 0.116 0.223 0.277 0.287 0.424 Chaetoceros gracilis Chic 0.171 0.143 0.125 0.148 0.152 0.102 Fuco 0.512 0.469 0.454 0.388 0.369 0.422 /3-caro 0.031 0.036 0.037 0.040 0.042 0.035 DD+DT 0.096 0.138 0.191 0.222 0.257 0.279 Thalassiosira weissflogii Chic 0.088 0.079 0.073 0.064 0.060 0.066 Fuco 0.413 0.439 0.452 0.321 0.295 0.410 /J-caro 0.026 0.030 0.032 0.045 0.045 0.036 DD+DT 0.206 0.278 0.300 0.293 0.325 0.543

90/imolm 2s '. This was primarily due to the increase in 0.5-i P. tricornutum the ratio of diatoxanthin to Chi a with increasing irradiance (Fig. 2). 0.4- The mean Chl-a-specific absorption coefficient from 400 0.3- to 700 nm (PAR), a*, increased with increasing irradiance (p<0.0\) (Table 3). Maximum variation in a* was observed 0.2- for C. gracilis, where a* varied about 1.6 times. The char acteristic absorption peaks by accessory pigments occurred 0.1- around 440, 462, 490, 636 and 675 nm. The absorption 0 peaks relating to diadinoxanthin plus diatoxanthin were lo cated at 440, 462 and 490 nm (cf. Fig. 1). The contribution 0.3 n C. gracilis of absorption by diadinoxanthin plus diatoxanthin to total in vivo pigment absorption («*dd+dt>.'Tp(^)) was determined by the ratio of the reconstructed absorption coefficient of 0.2- diadinoxanthin plus diatoxanthin (aDD^D1(X)) t0 tota' re" constructed absorption coefficient (#TP(A)) (Fig. 3). The

contribution of absorption by diadinoxanthin plus diatoxan Q thin to total in vivo pigment absorption at 440, 462 and 490 nm increased logarithmically with increasing irradiance (p<0.05) (Table 3). For example, the maximum contribu

tion of absorption by diadinoxanthin plus diatoxanthin to 0.6-1 T. weissflogii total in vivo pigment absorption was 57% at 462 nm in T. 0.5- weissflogii grown under 750 ^mol m"2 s~'. The cDD+DT calculated from equation I increased loga 0.4-

rithmically with increasing irradiance (p<0.01) (Fig. 4). As 0.3- the growth irradiance varied from 90 to 750^molm~2s"', 0.2- the cDD_DT increased 3.8 times ranging from 4.5 to 17% for

P. tricormttum, 3.3 times ranging from 5.8 to 19% for C. 0.1- gracilis, and 2.3 times ranging from 13 to 30% for T. weiss- 0 fiogii. 90 190 330 450 600 750

Discussion Irradiance (u.mol m"~ s )

A marked decrease in fj. due to photoinhibition was not Fig. 2. Ratios (w:w) of cellular diadinoxanthin (DD; hatched) observed for the 3 species studied, and for the range of irra plusdiatoxanthin (DT: open) to cellular Chia for 3 diatom species grown under 6 irradiances ranging from 90 to 750 j/mol irT2s~'. diances used in the present study (Table 1). This result may 100 T. Fujiki & S. Taguchi

Table 3. Variations in the mean specific absorption coefficient (a*) and the contribution of absorption by diadinoxanthin plus diatoxan thin to total in vivo pigment absorption («*dd+Dtvtp) at 440' 462 an(1490 nm. a* in m2 [mgChl a]~\

Irradiance (fjmo\ m 2 s ') Wavelength Species (nm) 90 190 330 450 600 750

Phaeodactvlum tricornutum 400-700 [a*] 0.014 0.016 0.017 0.018 0.019 0.021 440 0.066 0.092 0.156 0.193 0.206 0.251 462 0.110 0.144 0.225 0.281 0.308 0.334 490 0.114 0.146 0.224 0.287 0.316 0.323 Chaetoceros gracilis 400-700 [a*] 0.008 0.009 0.010 0.011 0.012 0.013 440 0.080 0.115 0.156 0.177 0.200 0.220 462 0.131 0.193 0.264 0.291 0.323 0.370 490 0.154 0.218 0.285 0.342 0.384 0.385 Thalassiosira weissflogii 400-700 [a*] 0.009 0.010 0.011 0.011 0.011 0.012 440 0.177 0.224 0.237 0.243 0.267 0.365 462 0.321 0.387 0.406 0.446 0.486 0.570 490 0.327 0.381 0.392 0.449 0.492 0.557

0.06 P. tricornutum imply that the photoprotection system acted well against 0.05 total excess light energy for the range of irradiances provided. The pigment ratio (w: w) of diadinoxanthin plus diatoxan 0.04 thin to Chi a, which showed the pool size of photoprotec- 0.03 tive pigments, increased linearly as the growth irradiance 0.02 increased (Table 2). Sakshaug et al. (1991) showed that

jT- 0.01 high-light (400^molm~2s~') adapted cells of the diatoms Thalassiosira nordenskioeldii and Chaetoceros furcellatus 2 0 contained up to 2 times more cellular diadinoxanthin E 0.04 plus diatoxanthin relative to Chi a than low-light (25 C. gracilis /imolm~2s"') adapted cells. Using the freshwater diatom 0.03- Nitzschia palea, Willemoes & Monas (1991) suggested that £ the conversion from diadinoxanthin to diatoxanthin took 0.02- place only as growth irradiance was saturated because dia toxanthin was only found in the cultures grown at the satu 0.01 rated irradiance levels over 200/anolirT2s~1. For all 3 S species used in this study, however, diatoxanthin was found a 0 in cultures where the growth irradiance was not saturated M 0.03 (Table l,Fig. 2). a. T. weissflogii As the growth irradiance varied from90 to 750

2 /imolm~2s~', the ratio (w:w) of diatoxanthin to diadinox U 0.02 anthin plus diatoxanthin, which indicated the activity of the xanthophyll cycle, increased 4 times ranging from 17 to 67% for P. tricornutum, 5 times ranging from 11 to 59% for 0.01- C. gracilis, and 7 times ranging from 10 to 69% for T. weissflogii respectively (Fig. 2). Cell size of the 3 species was determined from cellular carbon content, which indi 400 450 500 550 600 650 700 cated that T. weissflogii had the largest cell size, followed by C. gracilis and P. tricornutum (data not shown). The re Wavelength (nm) sults could suggest that the activity of the xanthophyll cycle Fig. 3. Partitioning of a*(X) between total pigments (thin line) is species dependent and is related to cell size. Willemoes and diadinoxanthin plus diatoxanthin (solid line) for three species & Monas (1991) pointed out that unavailable diadinoxan of diatoms grown under an irradiance level of 750Jumolm~2s~l, thinfor the xanthophyll cycle existed in the thylakoid mem as determinedby the ratio of the reconstructed absorption coeffi branes. The development of new methodology is required cient of diadinoxanthin plus diatoxanthin to the total reconstructed to quantify accurately the unavailable diadinoxanthin con absorption coefficient. tent, and to understand how this relates to total cellular di- Light Absorption and Pigments in Diatoms 101

0.4 -1 mained under 0.05 for all species, irrespective of growth ir radiance. Brown (1988) observed that /3-carotene was pref erentially found in pigment-protein complexes of photosyn- 0.3- thetic reaction centers compared to antenna pigment-pro tein complexes for 5 species of diatoms including, P. tricor nutum, C. gracilis and Thalassiosira sp.. These results may suggest that diadinoxanthin and diatoxanthin play a more 0.2- significant photoprotective role than /3-carotene in diatoms. Chi a specific absorption coefficient, a*(l), is used widely asthe fundamental index of light absorption by phy- 0.1- toplankton pigments (Morel & Bricaud 1981; Sosik & Mitchell 1994; Stuart et al. 1998). The a*(k) varies due to changes in package effect and the relative proportion of Chi a and accessory pigments (Sathyendranath et al. 1987; i Bricaud et al. 1988). The package effect describes the de 200 400 600 800 creased absorption of pigments in a cell compared to the absorption potential for the same amount of pigment in so Irradiance (|nmol m"~ s ) lution (Duysens 1956; Kirk 1976, 1994; Geider & Osborne 1987). An increase in package effect occurs as cell size in Fig. 4. The cDD+DT of Phaeodactyhtm tricornutum (D), Chaeto- creases or the cellular pigment concentration increases ceros gracilis (O) and Thalassiosira weissflogii (A) grown under (Kirk 1976; Morel & Bricaud 1981; Sosik & Mitchell 6 irradiances ranging from 90 to 750^mol m"2 s"'. 1994). In this study, the increase in a* with increasing irra diance may result from the combined increase in the contri bution of absorption by diadinoxanthin plus diatoxanthin to 0.04- total in vivo pigment absorption aswell as a decrease in the package effect due to a decrease in cellular pigment con centration (Table 3). Increase in the contribution of absorp § 0.03 tion by diadinoxanthin plus diatoxanthin to total in vivo o sz pigment absorption with increasing growth irradiance was most apparent between 450 and 500 nm, where the contri 1 0.02- bution of Chl-a absorption was minimal (Table 3). As the light energy absorbed by diadinoxanthin and dia toxanthin is not transferred to the photosynthetic reaction centers, the increase in cDD+DT would reduce the amount of 0.01- energy available for carbon fixation. To determine the molar ratio of carbon fixed for growth to the light energy absorbed by pigments, the quantum yield for growth 0p was calculated according to the model of Kiefer & Mitchell 0.1 0.2 0.3 0.4 (1983) for each species,

J»7(KI (3) Chla:C Fig. 5. Changes in M associated with cDD_DT for Phaeodaay- 4(X) lum tricornutum (D), Chaetoceros gracilis (O)and Thalassiosira where fj is the specific growth rate and Chi a: C is the ratio weissflogii (A) grown under 6 irradiances ranging from 90 to 750 of cellular Chl-o content to carbon content. As the growth irradiance varied from 90 to 750^molm~2s~', the 0/( de creased from 0.018 to 0.0038 molC(mol photon)"1 for P. adinoxanthin. tricornutum, 0.031 to 0.012 molC(mol photon)"1 forC. The ratios of Chi c to Chi a and fucoxanthin to Chi a var gracilis, and 0.034 to 0.0072 molC(mol photon)"1 for T. ied little with growth irradiance, but species-specific differ weissflogii. For the range of irradiances studied, the mini ences were observed (Table 2). Both Chi c and fucoxanthin mum 0^ were always observed at the highest irradiance of are able to transfer absorbed light energy to Chia (Fried 750Jumolm~2s~1. The fy, of all 3 species decreased loga man & Alberte 1984; Owens & Wold 1986). 0-carotene is rithmically with increasing cDD+DT (p<0.05) (Fig. 5). The thought to play an important photoprotective role in chloro- relationships were species dependent. Variations in the en phytes (Loeblich 1982). The ratio of/3-carotene to Chia re ergy transfer efficiency, the turnover time of reaction cen- 102 T. Fujiki & S. Taguchi ters, and the ratio of pigment in photosystem 1 to photosys- ties of diverse phytoplanktonic species: experimental results tem 2 could all contribute to the change in 0^ (Kolber et al. and theoretical interpretation. J. Plankton Res. 10: 851-873. 1988; Sosik & Mitchell 1991; Babin et al. 1996). These Brown, J. S. 1988. Photosynthetic pigment organization in di atoms (Bacillariophyceae). J. Phycol. 24: 96-102. may be some of the reasons why the relationship between Cleveland, J. S. & A. D. Weidemann 1993. Quantifying absorption cdd+dt and 0// was species dependent. The relationship be by aquatic particles: A multiple scattering correction for glass- tween cDD+DT and 0/( suggests that the increase in the ab fiber filters. Limnol. Oceanogr. 38: 1321-1327. sorption by diadinoxanthin plus diatoxanthin due to photo Cogdell, R. J. & H. A. Frank 1987. How carotenoids function in protection is closely related to the decrease in (j>t, at high ir photosynthetic bacteria. Biochim. Biophys. Acta 895: 63-79. radiances. Demers, S., S. Roy, R. Gagnon & C. Vignault 1991. Rapid light- induced changes in cell fluorescence and in xanthophyll-cycle Conclusion pigments of Alexandrium excavation (Dinophyceae) and Tha- lassiosira pseudonana (Bacillariophyceae): a photo-protection In this study, the spectral reconstruction technique was mechanism. Mar. Ecol. Prog. Ser. 76: 185-193. utilized to quantify absorption by the xanthophyll cycle Demmig, B., K. Winter, A. Kriiger & F. C. Czygan 1987. Photoin- pigments in relation to total in vivo pigment absorption. hibition and zeaxanthin formation in intact leaves. Plant Phys- The diatoms T. weissflogii, C. gmcUis and P. tricornutum /o/. 84:218-224. demonstrated a two-component xanthophyll cycle between Demmig-Adams, B. 1990. Carotenoids and photoprotection in diadinoxanthin and diatoxanthin at non-saturating growth plants: a rolefor the xanthophyll zeaxanthin. Biochim. Biophys. irradiances. The contribution of absorption by diadinoxan Acta 1020: 1-24. thin plus diatoxanthin to total in vivo pigment absorption Duysens, L. N. M. 1956. The flattening of the absorption spec (a(DD+DT>Tp(^)) was significant at high irradiances because trumof suspensions, as compared to that of solutions. Biochim. the pigment ratio of diadinoxanthin plus diatoxanthin to Biophys. Acta 19: 1-12. Chi a increased substantially with increasing irradiance. Falkowski, P. G. & J. A. Raven 1997. Aquatic Photosynthesis. 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