Evaluation of chlorophyll fluorescence, in vivo spectrophotometric pigment absorption and ion leakage as biomarkers of UV-B exposure in marine macroalgae

The photosynthetic fluorescence ratio F_v:F_m, in vivo absorption spectra and ion leakage were evaluated as biomarkers of ambient and elevated UV-B (280 to 320 nm) exposure of the intertidal alga Enteromorpha intestinalis (Chlorophyta) and the sublittoral alga Palmaria palmata (Rhodophyta). Measurements of thallus growth were also used to assess adverse biological effects. Ambient and elevated UV-B significantly inhibited photosynthesis in both species. It was shown that the F_v:F_m ratio is a sensitive, non-specific general biomarker of UV-B exposure in both species. Moreover, the in vivo absorption of what was tentatively identified as chlorophylls a and b as well as phycoerythrin and/or carotenoids, phycoerythrobilin and phycocyanin decreased in a dose-response dependent manner and was associated with a decrease in growth rate in P. palmata. The intertidal alga E. intestinalis showed a greater degree of tolerance to UV-B exposure. These results indicate that changes in the F_v:F_m ratio together with reductions in in vivo pigment absorption could provide an early quantitative warning of the detrimental effects of UV-B in marine macroalgae. Received: 16 May 1997 / Accepted: 16 July 1997     

Concentration of lodine and bromine by plants in the seas of Japan and Okhotsk

Iodine and bromine content were measured in 24 species of red (Rhodophyta), brown (Phaeophyta) and green (Chlorophyta) seaweeds and 2 species of higher water plants (Embryophyta) from the Sea of Japan, as well as in 12 species of the abovecited taxa and 1 species of flowering plant from the Sea of Okhotsk. Iodine was determined by photometric extraction with brilliant green, and bromine by neutron activation of samples. Phaeophyta and Rhodophyta were richest in iodine and bromine content. Representatives of the order Ceramiales (Rhodophyta) had high iodine and bromine contents. Thus, iodine concentrations in Ptilota filicina, Campylaephora hypnaeoides and Myriogramme yezoensis, a new iodine concentrator discovered by us, amount to 0.42, 0.094 and 0.75%, respectively. Bromine content in representatives of the family Rhodomelaceae was 3.36 and 3.74% in Japan Sea and Okhotsk Sea Rhodomela larix, respectively. Polysiphonia japonica (Rhodomelaceae) is a newly discovered concentrator of bromine (3.20%). Many species of the order Laminariales (Phaeophyta) were characterized by high iodine contents: Laminaria japonica, L. cichoriodes, L. inclinatorhiza, Cymathaere japonica and Alaria marginata. The Br:I ratio for all the species except those that concentrated iodine, was more than 1. Seaweeds that grow at greater depths showed increased iodine and bromine contents. A tendency toward increased iodine content was observed in species growing further to the North. Iodine and bromine were accumulated selectively by various organs of Sargassum pallidum.     

Waterborne chemical compounds in tropical macroalgae: positive and negative cues for larval settlement

Settlement sites of marine invertebrate larvae are frequently influenced by positive or negative cues, many of which are chemical in nature. Following from the observation that many shallow-water, Hawai'ian marine macroalgae are free of fouling by sessile invertebrates, we predicted that the algae are chemically protected and dependent on either surface-bound or continuously released soluble compounds to deter settling invertebrate larvae. To address the importance of waterborne algal compounds, we experimentally determined whether larvae of two of Hawai'i's dominant hard-surface fouling organisms, the polychaete tube worm Hydroides elegans and the bryozoan Bugula neritina, would settle in the presence of waters conditioned by 12 species of common Hawai'ian macroalgae (representing the Phaeophyta, Chlorophyta, Rhodophyta and Cyanophyta). The results included a full spectrum of biological responses by each larval species to waterborne algal compounds. Larval responses to conditioned water were consistent for each algal species, but the outcomes were not predictable based on the taxonomic relationships of the algae. For example, among the species of Phaeophyta examined, different conditioned waters were: (1) toxic, (2) inhibited settlement, (3) simulated settlement, or (4) had no effect, compared to larvae in control dishes containing filtered seawater. Additionally, larval responses to aged (24 h) conditioned waters could not be predicted from the results of assays run with conditioned waters utilized immediately after preparation. Finally, settlement by larvae of one species did not predict outcomes of tests for the other species. Four of 12 shallow-reef Hawai'ian macroalgae tested released compounds into surrounding waters that immediately killed or inhibited settlement by both H. elegans and B. neritina (toxic: Dictyota sandvicensis; inhibitory: Halimeda discoidea, Sphacelaria tribuloides, Ulva reticulata); the remaining 8 algal species prevented settlement by one of these fouling organisms but for the other had no effect or, in some cases, even stimulated settlement     

Fatty acid composition of marine macroalgae from the Black Sea and Dardanelles

The fatty acid (FA) composition was determined in 14 species of marine macroalgae belonging to the Chlorophyta, Phaeophyta and Rhodophyta, which were collected from ?ile in the Black Sea and Kepez in the Dardanelles. Generally, polyunsaturated FAs and monounsaturated FAs were major components (50–77%). Total saturated FAs ranged from 22% to 50%, with 16?:?0 as the most abundant saturate (32–38%). Two samples of Cystoseira barbata collected from a different station had some differences from each other in their contents of 18?:?2n-6 and 18?:?3n-3 and in the 18?:?2n-6/20?:?4n-6 ratios. Green algal species had a significantly higher proportion of unsaturated FAs and a significantly lower proportion of saturated FAs than the red and brown algae. The amount of n-3 FAs was significantly higher in Ulva rigita, Chaetomorpha linum, Enteromorpha linza and Gracilaria verrucosa (8.88, 6.44, 5.31 and 5.24, respectively).     

Assimilation efficiency of a temperate-zone intertidal fish (Cebidichthys violaceus) fed diets of macroalgae

We studied assimilation efficiencies of the temperate-zone intertidal fish Cebidichthys violaceus (Girard, 1854) fed in the laboratory on each of the following species of macroalgae: Spongomorpha coalita (Chlorophyta), Ulva lobata (Chlorophyta), Iridaea flaccida (Rhodophyta) and Porphyra perforata (Rhodophyta). Together, these 4 algae make up over 75% of the natural summer diet of C. violaceus. Assimilation efficiency was calculated by proximate organic analysis of food and feces; the amount of ash in food and feces was used as a standard. Depending on the algal species, the fish assimilated 43 to 81% of the protein, 21 to 44% of the lipid, 45 to 62% of the carbohydrate and 31 to 52% of all three classes of organic material combined. These data are the first results showing that a temperate-zone marine fish can assimilate macroalgal constituents. Protein, carbohydrate and total organic material were absorbed more efficiently from rhodophytes than from chlorophytes. Conversely, lipid was absorbed more efficiently from chlorophytes than from rhodophytes. These results are compared with previous work showing that C. violaceus in nature eats more chlorophytes than rhodophytes, but in laboratory preference tests prefers rhodophytes to chlorophytes.     

UV absorption by mycosporine-like amino acids in Phaeocystis antarctica Karsten induced by photosynthetically available radiation

Mycosporine-like amino acids (MAAs), which occur in diverse taxonomic groups, exhibit in vivo absorption maxima between 310 nm and 360 nm and may play a photoprotective role against ultraviolet (UV) exposure. Using cultures of colonial Phaeocystis antarctica, we examined the relationship between MAA concentration, in vivo UV absorption, photoprotective (carotenoid) and photosynthetic pigments, and photosynthetically available radiation (PAR, 350–700 nm). UV absorption was high; chlorophyll-specific absorption, a ^* _ph, at 330 nm ranged from 0.06 to 0.41 m²/mg chlorophyll a. Values of a ^* _ph (330) were 4–13 times greater than a ^* _ph (676). Mycosporine-glycine, shinorine, and mycosporine-glycine valine are responsible for the strong in vivo UV absorption. The sum of all MAAs increased with irradiance when normalized to chlorophyll a or carbon concentrations, whereas individual MAAs varied independently from each other. Mycosporine-glycine concentrations showed no statistically significant change over the range of light intensities, whereas mycosporine-glycine and shinorine concentrations increased at higher irradiances. The relative fluorescence yield for chlorophyll a was low in the UV region compared to the visible region, implying that absorbed UV radiation (<375 nm) is transferred inefficiently to chlorophyll a in the reaction center. Quantitative estimates of UV screening by MAAs are attributed to elevated MAA concentrations and increased diameter at high light. Received: 31 March 1999 / Accepted: 13 July 2000     

High molecular weight algal substances in the sea

Phytoplankton exudates into culture media and extracts from littoral algae are shown to contain significant quantities of extracellular materials having molecular weights in excess of 50,000. A recent development in experimental fluid mechanics (the reduction of turbulent friction coefficients of flowing solutions by dissolved macromolecules) has been utilized as a specific test for high molecular weight compounds. Phytoplankton species in all major algal groups could be found which exude high molecular weight compounds into their culture media; the red alga Porphyridium cruentum typically produced friction reductions as high as 60%. Seaweed samples in the Chlorophyta, Phaeophyta, and Rhodophyta showed major changes in turbulent friction coefficient; in some cases the friction was less than 1/2 that of pure seawater. All samples of the genus Porphyra and Gigartina produced substantial friction changes. From these experiments it is concluded that algal exudates can be a prominent source of high molecular weight compounds in the sea.     

Settlement of larval blacklip abalone, Haliotis rubra, in response to green and red macroalgae

Surfaces from the habitat of adult Haliotis rubra were tested as inducers of larval settlement to determine the cues that larvae may respond to in the field. Settlement was high on the green algal species Ulva australis and Ulva compressa (Chlorophyta), the articulated coralline algae Amphiroa anceps and Corallina officinalis, and encrusting coralline algae (Rhodophyta). Biofilmed abiotic surfaces such as rocks, sand and shells did not induce settlement. Ulvella lens was also included as a control. Treatment of U. australis, A. anceps and C. officinalis with antibiotics to reduce bacterial films on the surface did not reduce the settlement response of H. rubra larvae. Similarly, treatment of these species and encrusting coralline algae with germanium dioxide to reduce diatom growth did not significantly reduce larval settlement. These results suggest that macroalgae, particularly green algal species, may play an important role in the recruitment of H. rubra larvae in the field and can be used to induce larval settlement in hatchery culture.     

In situ characterization of phytoplankton from vertical profiles of fluorescence emission spectra

Vertical profiling of the upper ocean with a laser/fiber optic fluorometer enabled the determination of fluorescence emission spectra of photosynthetic pigments over small vertical scales. Simultaneous acquisition of phycoerythrin (PE) and chlorophyll (chl) emission spectra allowed in situ differentiation between PE-containing cells (cryptomonads and cyanobacteria) and other chl-containing autotrophs. Further, fluorescence spectral peak shifts associated with different species of PE-containing cells resulted in even finer scale in situ taxonomic differentiation. We found that the phycoerythrin fluorescence emission maxima shifted from 578 nm near the surface, to 585 m at the base of the shallow thermocline (30% light level), and to 590 nm below the thermocline at the base of the euphotic zone (1% light level). These shifts in peak emission coincided with a taxonomic change in the PE-containing cells (as determined from analysis of discrete bottle samples) from a greater proportion of Synechococcus spp. in the upper water column to a greater proportion of cryptomonads at the base of the euphotic zone. These results indicate that the composition of the phytoplankton assemblage may be assessed in situ without sample collection.     

Habitat-induced morphological variation influences photosynthesis and drag on the marine macroalga <Emphasis Type="Italic">Pachydictyon coriaceum</Emphasis>

Photosynthesis, growth, distribution, and persistence of macroalgae are determined in part by the physical environment in which they live. Therefore, discerning how macroalgae interact with their physical environment is necessary to better understand their physiological performance. The purpose of this study was to examine what photosynthetic and hydrodynamic costs and benefits the morphology of Pachydictyon coriaceum (Phaeophyta) confers on the thallus in a given environment. Principal components analysis of morphometric measurements of Pachydictyon coriaceum from different flow habitats and depths separated thalli into three distinct morphs: shallow wave-exposed, shallow wave-protected, and deep. To test the hypothesis that thallus morphology affects net photosynthesis (NP), thalli of three morphotypes of P. coriaceum were incubated in an enclosed recirculating flume under three simulated light/water flow environments representing conditions from which the three morphotypes were collected. The wave-protected and deep morphs had significantly higher rates of photosynthesis than the wave-exposed morph for all three simulated environments. The dense, compact shape of the wave-exposed morph readily streamlines with flow and in doing so, potentially shades many of its internal blades likely accounting for its lower biomass-specific NP. Drag coefficients (C _d) were estimated for the three morphotypes over a range of flow velocities between 0.08 and 0.47 m s⁻¹. At lower water flow velocities (0.08–0.21 m s⁻¹), wave-exposed morphs had the lowest C _d among the three morphotypes. But drag coefficients of the three morphotypes converged with increasing flow velocities, and at velocities >0.31 m s⁻¹ there were no differences in C _d among the three morphotypes. The results of this study indicate that the environmentally-shaped morphs influence photosynthesis and, to a lesser degree, hydrodynamic forces acting on P. coriaceum.     

Fluorescence induction in the macroalgae Chondrus crispus (Rhodophyceae) and Ulva sp. (Chlorophyceae)

The induction of in vivo chlorophyll a (chl a) fluorescence (change in fluorescence intensity during a time-scale of ms to s) was measured to determine the potential of this technique for assessing the physiological condition of the macroalgae Chondrus crispus and Ulva sp. A gradient in variable fluorescence (P-F ₀ =peak minus initial fluorescence, a measure of Photosystem II activity) was found along the frond of C. crispus, the values increasing with distance from the thallus apex. No gradient was observed for Ulva sp. thalli. Nitrogen- or phosphorus-depleted Ulva sp. required a longer dark-conditioning period and had lower values of P-F ₀ than did controls. In contrast, no differences were found in P-F ₀ of N- or P-depleted C. crispus unless values were normalized to chl a. The irradiance history of C. crispus strongly influenced P-F ₀ , even after dark-conditioning: P-F ₀ declined by about 70% as the mean daily natural irradiance increased between 2 February and 14 March 1986; a negative correlation was observed between P-F ₀ and the photon flux density 1 d prior to the measurement; P-F ₀ remained elevated when C. crispus was grown under a low photon flux density; and P-F ₀ decreased in thalli within 5 d of transfer from growth under natural irradiance to an incubator with artificial irradiance. Changes in variable fluorescence at different growth irradiances of C. crispus may be due to adaptive changes in the relative absorption cross-section of this alga. The influence of irradiance history on Ulva sp. was minimal in comparison.Issued as NRCC No. 28730Part of this study was carried out while employed by Focal Marine Ltd., Bedford, Nova Scotia, Canada     

Contents of heavy metals in marine algae from Egyptian Red Sea coast

Seaweeds belonging to 14 different genera of Chlorophyta, Phaeophyta and Rhodophyta were analyzed to determine the levels of heavy metals in two areas of the Egyptian Red Sea coast. Among the trace metals analyzed, Mn and Zn showed the highest mass concentrations in the surface seawaters of the two studied areas. However, algae obtained from Suez area had the highest concentrations of the investigated heavy metals than those collected from Mars Alam area. Nevertheless, a high variability of the metal levels occurs among the studied algae and also between the investigated areas. Moreover, Zn was the most abundant metal in the seaweeds of the Suez area, while Pb was predominant in Mars Alam area in red and brown algae. L. farinosa had the highest average concentration factor of Zn in Suez (29161 fold), while it was 20091 fold in E. intestinalis at Mars Alam. The highest value of metal pollution index (MPI) was recoded in L. farinosa (22.0) at Suez. It represents 4.6 fold of that value recorded in L. farinosa at Mars Alam. Among green, brown and red algae in Suez, the highest values of MPI were recorded in Cladophora (mixed sub-species) and H. comuto (18.2 and 18.3), P. pavonia (16.2) and L. farinosa (22.1), respectively; while at Mars Alam, they were recorded in Cladophora (mixed sub-species) (6.6), P. pavonia (3.4) and L. farinosa (4.8), respectively.     

Pigments of meso- and bathypelagic chaetognaths

Pigments of the meso- and bathypelagic chaetognaths Sagitta macrocephala and Eukrohnia fowleri were studied by chromatographic analysis. Supplementary histological studies were also performed. Fat-soluble properties and absorption spectra of the chaetognath pigments indicated that all pigments were carotenoid, independent of chaetognath species or habitat. The major carotenoid in chaetognaths was very soluble in nonpolar solvents such as carotenes, although its absorption spectrum formed a single broad peak at around 460 nm. The characteristics of the carotenoids in the chaetognaths were different from those of the pigments in the plankton which formed their diet. It is therefore inferred that carotenoids in chaetognaths are not formed by the deposition of food pigments in the intestinal tissue, but are synthesized by the chaetognaths themselves.     

Nitrogen- versus phosphorus-limited growth and sources of nutrients for coral reef macroalgae

Recent investigations of nutrient-limited productivity in coral reef macroalgae have led to the conclusion that phosphorus, rather than nitrogen, is the primary limiting nutrient. In this study, comparison of the dissolved inorganic nitrogen:phosphorus ratio in the water column of Kaneohe Bay, Hawaii, with tissue nitrogen:phosphorus ratios in macroalgae from Kaneohe Bay suggested that nitrogen, rather than phosphorus, generally limits productivity in this system. Results of nutrient-enrichment experiments in a flow-through culture system indicated that inorganic nitrogen limited the growth rates of 8 out of 9 macroalgae species tested. In 6 of the species tested, specific growth rates of thalli cultured in unenriched seawater from the Kaneohe Bay water column were zero or negative after 12 d. These results suggest that, in order to persist in low-nutrient coral reef systems, some macroalgae require high rates of nutrient advection or access to benthic nutrient sources in addition to nutrients in the overlying water column. Nutrient concentrations in water samples collected from the microenvironments inhabited or created by macroalgae were compared to nutrient concentrations in the overlying water column. On protected reef flats, inorganic nitrogen concentrations within dense mats of Gracilaria salicornia and Kappaphycus alvarezii, and inorganic nitrogen and phosphate concentrations in sediment porewater near the rhizophytic algae Caulerpa racemosa and C. sertularioides were significantly higher than in the water column. The sediments associated with these mat-forming and rhizophytic species appear to function as localized nutrient sources, making sustained growth possible despite the oligotrophic water column. In wave-exposed habitats such as the Kaneohe Bay Barrier Reef flat, water motion is higher than at protected sites, sediment nutrient concentrations are low, and zones of high nutrient concentrations do not develop near or beneath macroalgae, including dense Sargassum echinocarpum canopies. Under these conditions, macroalgae evidently depend on rapid advection of low-nutrient water from the water column, rather than benthic nutrient sources, to sustain growth. Received: 1 December 1997 / Accepted: 9 July 1998     

Assessment of AHS (Airborne Hyperspectral Scanner) sensor to map macroalgal communities on the Ría de vigo and Ría de Aldán coast (NW Spain)

Ría de Vigo and Ría de Aldán have high biological richness that is reflected in the number of environmental protection areas like the Atlantic Islands National Park and five places of community interest. Benthic algal communities play an important role in these ecosystems due to their ecological functions and support a great part of this biological richness. We tested by means of bio-optical modelling and Airborne Hyperspectral Scanner (AHS) images to what extent remote sensing could be used to map these communities in Ría de Vigo and Ría de Aldán (NW Spain). Reflectance spectra of dominating macroalgae groups were modelled for different water depths in order to estimate the separability of different bottom types based on their spectral signatures and the spectral characteristics of the AHS. Our results indicate that separation between three macroalgae groups (green, brown and red) as well as sand is possible when the bottoms are emerged during low tide. The spectra differences decrease rapidly with increasing water depth. Two types of classifications were carried out with the three AHS images: maximum likelihood and spectral angle mapper (SAM). Maximum likelihood showed positive results reaching overall accuracy percentages higher than 95?% and kappa coefficients higher than 0.90 for the bottom classes: shallow sand, deep sand, emerged rock, emerged macroalgae and submerged macroalgae. Sand and algae substrates were then separately analysed with SAM. These classifications showed positive results for differentiation between green and brown macroalgae until 5?m depth and high differences between all macroalgae and sandy substrate. However, differences between red and brown macroalgae are only detectable when the algae are emerged.     

Vertical heterogeneity in physiological characteristics of Ulva spp. mats

In eutrophic areas, green macroalgae are frequently and for long periods arranged in mats, resulting in a steep light gradient. This study investigates the effect of this gradient on physiological characteristics [tissue nitrogen content, maximal photosynthetic efficiency (F_v/F_m), glutathione levels and redox ratio, absorbance and absorption spectra] of the green macroalga Ulva spp. Mats were sampled during the build-up (June), stationary (July), and decomposing (September) phases of a macroalgal bloom in the Veerse Meer, a eutrophic brackish (salinity 15–20 psu) lake in the southwest Netherlands. Water samples were taken for nutrient analyses. At all three sampling dates, the mats were composed almost entirely of Ulva spp.; in September the mats were in decay and covered with silt and epiphytes. In June and July, total dissolved inorganic nitrogen concentration (DIN) of the water within the mat was significantly higher than outside the mat. Pronounced vertical differences were found in tissue N, F_v/F_m values, total glutathione levels, glutathione redox ratios, and absorbance. In June and July, tissue N decreased from over 2.2% dry weight (DW; N-sufficient) in the bottom layers to around 1% DW (minimum level for survival) in the top layers. Wide-band absorption increased with depth in the mat and throughout the season, probably due to higher Chl a and b and lutein contents. The shape of the absorption spectrum was similar for all layers. The absorption of the silt/epiphyte film on the top Ulva layer was highest; its absorption spectrum (high absorption in the 500–560 nm range) indicates that the film on the top layers of the macroalgal mats mainly consisted of diatoms. In June, F_v/F_m and the glutathione redox ratio of the algae increased with depth in the layer, while total glutathione decreased. Low F_v/F_m values in the bottom and middle layers in September reflect the bad condition of the algae; the mats were largely decaying. It is concluded that multiple growth-limiting gradients occur in macroalgal mats: upper layers suffer from nitrogen limitation and photoinhibition while bottom layers are light limited. The algae in the mat acclimatize to low light conditions by increasing their absorption through increased pigment contents and by higher photosynthetic efficiency during the build-up and stationary period. This study qualifies the glutathione redox ratio as a promising candidate for stress indicator in macroalgae and provides suggestions for its further development.Communicated by S.A. Poulet, Roscoff     

Quantum yield,relative specific absorption and fluorescence in nitrogen-limited Chaetoceros gracilis

Decreases in cell-nitrogen quota resulted in changes in the carbon-based quantum yield of photosynthesis, the chlorophyll a-specific absorption coefficient, and in vivo fluorescence in the marine diatom Chaetoceros gracilis in laboratory experiments performed in 1983 and 1984. The three parameters were independently determined for the two spectral regions dominated by either chlorophyll a or fucoxanthin absorption. As cell-nitrogen quota decreased, the quantum yield for both pigments decreased; the specific absorption coefficient for chlorophyll a and the in vivo chlorophyll a fluorescence excited by each pigment increased. The observed increase in the in vivo fluorescence per chlorophyll a could be partially attributed to the increased specific absorption coefficient for chlorophyll a; the remainder of the fluorescence increase was related to a decline in photosystem activity. Energy transfer efficiency between light-harvesting pigments appeared to be maintained as cell-nitrogen quota decreased. The decrease in a fluorescence index [(F _DCMU-F _O)/F _DCMU] with nitrogen starvation suggested a decrease in Photosystem II activity. These results imply that decreases in reaction center and/or electron-transport system activity were responsible for the decline in rates of photosynthesis under conditions of notrogen deficiency.     

Phycoerythrin and photosynthesis of the pelagic blue-green alga Trichodesmium thiebautii in the waters of Kuroshio,Japan

The photosynthetic function of phycoerythrin was investigated in the alga Trichodesmium thiebautii from the waters of Kuroshio, Japan. The spectroscopic characteristics of the in vivo and isolated T. thiebautii phycoerythrin pigments are identical, and have 3 absorption bands at 495, 547 and 562 nm. Light at the wavelengths corresponding to each absorption band of phycoerythrin is equally efficient in T. thiebautii photosynthesis, indicating that phycoerythrin is active in trapping light energy for photosynthesis. In the natural habitat, phycoerythrin is considered to be the main photosynthetic pigment in T. thiebautii photosynthesis.     

Light absorption by a marine diatom: experimental observations and theoretical calculations of the package effect in a small Thalassiosira species

The relationship between in vivo light absorption efficiency of whole cells and in vitro absorption efficiency of algal pigments has been examined experimentally in the marine diatom Thalassiosira sp. In vitro absorption spectra were obtained for cells disrupted by either ultrasonic treatment or high-pressure shearing stress in a low-temperature (-40°C) pressure cell. A dimensionless measure of the magnitude of the package effect (Q _a ^*), calculated from the ratio of whole-cell to disrupted-cell absorption, ranged from about 0.5 at the blue absorption peak of chlorophyll a (λ=435 nm) to 0.7 at the red chlorophyll a peak (λ=670 nm) to 1.0 at the absorption minimum (λ=600 nm). Cell diameter was found to be an inappropriate measure of size for assessing the magnitude of the package effect. Instead, the effective optical diameter for calculation of intracellular self-shading was found to be less than the cell diameter. This observation is consistent with the fact that most algal pigments are contained within chloroplasts, and that chloroplast volume is necessarily smaller than cell volume.     

Determination of photosynthetic pigment composition in an individual phytoplankton cell in seas and lakes using fluorescence microscopy; properties of the fluorescence emitted from picophytoplankton cells

To develop a method for the determination of photosynthetic pigment species in individual phytoplankton cells, especially natural picophytoplankton cells, the fluorescence spectra of intact cells were studied with cultured phytoplankton. The study was made mainly with phycoerythrin-containing picophytoplankton collected off Japan in 1982 with reference to diatomal species and phycoerythrin-free cyanophycean species. The spectra were measured for cell suspensions with an ordinary spectrofluorometer, and for individual cells with a microscope spectrofluorometer, paying special attention to the effect of cell-fixation. Results indicated that: (1) the cell-fixation with the glutaraldehyde and paraformaldehyde mixture modified phycoerythrin emission from picophytoplankton markedly in its wavelength location and intensity, but (2) the emission from phycocyanin was affected far less, and (3) the emission from chlorophyll a was not altered. However, the phycoerythrin emission modified by the fixation was found to be easily distinguished from other emissions, and kept its intensity high enough for detection with a fluorescence microscope. The fluorescence properties after the fixation were kept unaltered for a long period of time. Based on the results, we propose a simple method for the determination of photosynthetic pigments in individual phytoplankton cells in seas and lakes using fluorescence microscopy. Results of our tests with natural samples of phytoplankton are presented, and problems for further improvement are also discussed.