Significance of cellular nitrate content in natural populations of marine phytoplankton growing in shipboard cultures

Phytoplankton intracellular nitrate concentrations have been monitored in a 56-h experiment on a shipboard culture of surface sea water from an upwelling region. These measurements were related to parameters of biomass (particulate nitrogen) and nitrate assimilation using the ¹⁵N isotope technique and the nitrate reducase (NR) assay. The procedure for measuring cellular nitrate concentrations is described. This parameter exhibited diurnal variations, ranging from 3.1 to 20.6 ng-at nitrate per g-at particulate nitrogen, and could be correlated positively with NR activity. Nitrogen budgets show that NR activity represents only 12% of nitrate incorporation in organic phytoplankton material when nitrate is available in the sea water. However, upon depletion of the environmental nitrate (zero uptake), NR activity can fully account for the decrease of internal nitrate. From the results, it seems that internal nitrate content is a better index of nitrate consumption by marine phytoplankton than the external concentration of nitrate-nitrogen.     

Nitrogenous nutrition in the euphotic zone of the Central North Pacific Gyre

The uptake rates of the three nitrogen compounds ammonium, nitrate, and urea were measured in the oligotrophic North Central Pacific Gyre in August–September 1985. The measurements were performed by using ¹⁵N-labelled substrates and incubating for short-time periods (3 to 4 h) under simulated in situ conditions. Ambient concentrations of the nitrogenous nutrients were generally below 0.10 mol l^-1. The average total daily nitrogen uptake rate, integrated over the euphotic zone, was 12.5 mmol N m^-2 d^-1. Diel studies in the upper water mass resulted in a calculated phytoplankton growth rate of 1.3 d^-1. Ammonium was the dominating nutrient, accounting for on the average 54% of the total nitrogen uptake, while urea uptake represented 32% and nitrate 14%. Ammonium uptake rates at a coastal station off the Hawaiian Islands were very close to the rates found at the oceanic station. Organisms <3 m dominated the nitrogen assimilation, being responsible for about 75% of the ammonium uptake. The nitrogen uptake rates in this study seem to be higher than those found by earlier investigations in the area, but correlated well with other productivity measurements performed during the same cruise.     

Nitrate storage by phytoplankton in a coastal upwelling environment

The storage of nitrate by phytoplankton cells during the early phases of upwelling was studied in coastal stations off northern Spain (southern Bay of Biscay) between 1990 and 1994. In this region, a persistent upwelling during summer is characterised by intermittent pulses of variable intensity, and increased nutrient concentrations in the surface layer. The main effect of an upwelling pulse on phytoplankton distribution is the shifting of the chlorophyll a and primary production maxima to near the surface. When the upwelling relaxes, thermal stratification of the water column occurs, and a distinct subsurface chlorophyll maximum develops below the production maximum. An accumulation of intracellular nitrate characterized the early phases of upwelling (mean = 2.73 μmol N m⁻³), maximum concentrations being attained at depths where biomass and production values were moderate. In contrast, phytoplankton cells from non-upwelling situations contained significantly lower concentrations of intracellular nitrate (mean = 0.17 μmol N m⁻³). The variations in the intracellular pool of nitrate may result from the differential allocation of resources within the cell as a result of variations in the energy available, since the uptake and assimilation of nitrate is a relatively expensive process involving several enzymatic systems. We hypothesize that nitrate storage by phytoplankton cells is characteristic of early phases of upwelling and is linked to patterns of carbon fixation. Average nitrogen budgets for upwelling and non-upwelling situations indicate that intracellular nitrate reserves are not responsible for maintaining high phytoplankton growth rates, since they only account for <2% of daily primary production during upwelling events. Received: 28 August 1996 / Accepted 3 December 1996     

Nitrogen regeneration by two surf-zone mysids,Mesopodopsis slabberi andGastrosaccus psammodytes

Nitrogen regeneration by two surf zone mysids,Mesopodopsis slabberi andGastrosaccus psammodytes, was determined under laboratory conditions. The mysids were collected from the lower Sundays River estuary, South Africa, from early spring 1984 to late summer 1985. The forms of nitrogen excreted and the effects of mass, temperature and feeding on excretion rate were determined for each species at three experimental temperatures. Comparison of the forms of nitrogen excreted revealed only slight differences between species, with ammonia the major form and urea and amino acids the secondary excretory products in both cases. Mass significantly influenced the rate of ammonia excretion at all experimental temperatures, with no significant difference in slope (common b=0.602) detected between species. During the day sediment deprivation resulted in a 15% and 20% increase in mean ammonia excretion rates of juvenile and adultG. psammodytes respectively, whereas no significant differences were found at night. The mean ammonia excretion rates of fedM. slabberi andG. psammodytes were 2 and 2.5 times higher than starved excretion rates, respectively.G. psammodytes andM. slabberi recycle 139 to 150 g N per running meter of surf zone per year and 1 007 to 1 208 g N m^-1 yr^-1, respectively. Togehter this constitutes 10% of total phytoplankton nitrogen requirements in the surf zone.     

Excretion of ammonia by zooplankton and its potential contribution to nitrogen requirements for primary production in the Catalan Sea (NW Mediterranean)

Excretion of ammonia by mesozooplankton (>200 m zooplankton) and its potential contribution to the nitrogen requirement for phytoplankton growth has been estimated for different hydrographical situations along a transect across the Catalan Sea (Northwestern Mediterranean). The nitrogen excreted as ammonia was estimated from mesozooplankton biomass and specific excretion rates. Nitrogen requirements of phytoplankton were estimated by means of carbon fixation rates and C:N ratios of <200 m particulate organic matter. Minimum C:N ratios and maximum primary production, zooplankton biomass, phytoplankton nitrogen requirements, and nitrogen excretion of zooplankton occurred near the Catalan density front. On average, the nitrogen regenerated by the mesozooplankton accounted for 43% of the nitrogen requirements of the phytoplankton. The specific excretion rates of ammonia and the percentage of phytoplanktonnitrogen requirements supplied by excreted nitrogen were higher at coastal stations. In some coastal and frontal stations, the ammonia excreted exceeded the phytoplanktonnitrogen demand. Bacteria competing for nutrient supply and the possible uncoupling between rate processes and standing stocks of phyto- and zooplankton could explain the apparent excess of regenerated ammonia.     

Inorganic nitrogen metabolism in marine bacteria: Nitrate uptake and reduction in a marine pseudomonad

Growth experiments in batch cultures indicated that the uptake of nitrate by the marine pseudomonad PL1 was inhibited in the presence of ammonia provided that the ammonia concentration was higher than 1 mM. At ammonia concentrations of less than about 1 mM, however, both nitrate and ammonia were utilised simultaneously. The saturation constants for nitrate and ammonia uptake were both 2.6x10^-4 M, and similar to the Michaelis constants of nitrate reductase for nitrate (2.9x10^-4 M) and glutamine synthetase for ammonia (2x10^-4 M). Nitrate reductase activity linked to NADH was detected in chemostat-grown cultures with nitrate as nitrogen source, and in cultures containing limiting concentrations of nitrate and ammonia, ammonia or glutamate. Enzyme synthesis appeared to be repressed in cultures containing an excess of ammonia or glutamate. Chemostat cultures utilised ammonia or glutamate in preference to nitrate, while there was no marked preference between ammonia and glutamate.     

Regulation of nitrate and ammonium uptake in the Greenland Sea

The uptake of nitrate and ammonium was investigated experimentally during early spring 1989 in the Greenland Sea, with particular attention placed on the roles of irradiance, nitrogen concentrations and nitrateammonium interactions. The phytoplankton assemblage was dominated by the colonial prymnesiophyte Phaeocystis pouchetii. Nitrate concentrations ranged from undetectable at the end of the cruise to greater than 10 M, and ammonium levels ranged from less than 0.1 to 1.9M. The uptake of both nitrate and ammonium as a function of irradiance was found to be a saturation response. Photoinhibition occurred and was found to be greater for ammonium uptake. Ammonium uptake also saturated at irradiance levels five times lower than those needed to saturate nitrate uptake. Nitrate and ammonium uptake as a function of nitrogen concentration also was characterized by a saturation response, with the estimated half-saturation constant (K _s) value for nitrate uptake being 0.29 M. Elevated ammonium concentrations inhibited nitrate uptake, and the response appeared to be one of exponential decrease with increasing concentrations of ammonium. The most important factor in the Greenland Sea influencing ammonium uptake during the spring was irradiace, while both irradiance and ammonium concentrations played major roles in regulating nitrate uptake and new production.     

Seasonal growth in Laminaria longicruris: Relations with dissolved inorganic nutrients and internal reserves of nitrogen

Observations have been made on seasonal fluctuations in dissolved inorganic nutrients, internal reserves of nitrogen and growth rates in Laminaria longicruris. The onset of winter growth in shallow-water stations (6 and 9 m) correlated well with improved dissolved nitrate conditions in the sea. During the winter, reserves of NO ₃ ^- were accumulated by the plants and reached maximum values of 150 moles per g fresh weight in March. This represents a concentration factor of approximately 28,000 over the ambient levels, or an internal nitrogen reserve of 2.1% of the dry weight of the tissue. Depletion of this nitrogen pool followed the disappearance of the external NO ₃ ^- with a lag period of up to 2 months. Rapid kelp growth was measured during this period. Reserves of organic nitrogen also reached maximum values in March and declined slowly throughout the summer into autumn. It is suggested that the combined inorganic and organic nitrogen reserves sustain the rapid growth rates into July and at reduced rate through the late summer. Fertilization of an experimental perimental kelp bed with NaNO₃ increased the internal plant reserves of NO ₃ ^- and produced a much improved summer growth rate. The enriched plants developed very small reserves of carbohydrate during the rapid summer growth phase.NRCC No. 15549.     

Nitrogen uptake and growth of the germlings and mature thalli of Fucus distichus

Fucus distichus L. was collected near Vancouver, Canada, in late fall and early winter, 1981. The effects of the forms of nitrogen (nitrate, ammonium or urea) and periodic exposure to air on growth, rhizoid development and nitrogen uptake in germlings was investigated. Gamete release, fertilization, germination and germling growth had no requirement for a specific form of nitrogen. Periodic exposure to air increased secondary rhizoid development twofold. Nitrate and ammonium uptake rates of the germlings were higher than for the mature thalli (20 to 40 times for nitrate and 8 times for ammonium), while the halfsaturation constant (K _s) values for nitrate were similar (1 to 5 M). The germlings showed saturable uptake kinetics but the mature thalli did not. When germlings were exposed to air it caused a 70% decrease in nitrate uptake, but not change in ammonium uptake. Ammonium uptake in the mature thalli was proportional to the ambient ammonium concentration. Nitrate uptake in the mature thalli appeared to follow saturation kinetics at low nitrate concentrations, but showed a non-saturable component at concentrations greater than 10 M. Presence of ammonium inhibited nitrate uptake by the mature plants but not by the germlings.     

Nitrogen regeneration by the surf zone penaeid prawn Macropetasma africanus

Nitrogen excretion of individual Macropetasma africanus (Balss) from an exposed beach/surf zone in Algoa Bay, South Africa was monitored under laboratory and field conditions in relation to body mass, temperature and feeding during 1985. Excretion rate experiments were performed on starved prawns at 15°, 18°, 20° and 25°C, as well as on individuals fed on four different diets (mussel, fish, shrimp and natural diet) at 15° and 20°C. The ratios of the excreted compounds to total nitrogen excreted were similar for the four diets despite differences in their nitrogen content and in the amount of food consumed. At 15° and 20°C, ammonia excretion rates of fed individuals were four to seven times higher than in starved prawns. the excretion rates were not correlated with nitrogen content of diets. M. africanus recycles 1 557 g NH₄–N per metre strip per year or 1 832 g total nitrogen m^-1 yr^-1, which constitute 12 and 14%, respectively, of total phytoplankton requirements of the surf zone. This study indicates that large motile crustaceans, when abundant, can play an important role in nutrient recycling in turbulent marine environments.     

Microphytoplankton in the Strait of Otranto (eastern Mediterranean)

The Strait of Otranto is the connection between the Adriatic and Ionian Seas. Low nutrient concentrations, high transparency, and low phytoplankton cell density and biomass reflect the oligotrophic character of the area. Enrichment of the euphotic layer with nutrients is mainly due to discharge of Albanian and Greek rivers, as well as mixing and upwelling in winter/early spring. Following phytoplankton bloom in April, a progressive decrease of phytoplankton cell density is due to the consumption of nutrients throughout the proceeding summer and autumn. Nitrogen was a strong limiting factor for phytoplankton growth in summer. Deep biomass maxima were detected in the 50 to 100 m layer and corresponded mostly to cells smaller than 20 m. The eastern part of the strait is mostly influenced by the northerly inflowing current from the Ionian Sea, and the western part by the southerly outflowing current from the Adriatic Sea. This typical circulation could be disturbed by inertial oscillations in the current field, generated by the strong oscillating winds and cyclonic eddies. The type of circulation determined the distribution of thermohaline characteristics, abundance, biomass, as well as taxonomic composition of phytoplankton, across the strait. Ecological characteristics of the water masses on two sides of the strait were significantly different during the formation of a longitudinal thermohaline front in May 1990.     

Urea as a nitrogen source for the phytoplankton in the Oslofjord

Ambient concentrations of urea in the inner Oslofjord, Norway, showed a pronounced yearly cycle in 1980, with values in the range 0.1 to 10.0 μg-at N l^-1; this cycle resemble that of ammonia although urea concentrations were usually lower. The uptake of urea by phytoplankton was investigated using ¹⁵N. Urea was usually a less important N source than NH ₄ ⁺ , and accounted for 0 to 53% (mean 19%) of summed NH ₄ ⁺ +NO ₃ ^- + urea uptake rates from April to October. Absolute as well as relative (specific) uptake rates of urea were higher in the summer (June–August) than at other times. Uptake of urea was inhibited by NH ₄ ⁺ concentrations higher than 1 to 2 μg-at N l^-1. The summed NH ₄ ⁺ +NO ₃ ^- + urea uptake rate was exponentially related to temperature.     

Nutrient uptake kinetics and growth of zooxanthellae maintained in laboratory culture

Growth characteristics and nutrient uptake kinetics were determined for zooxanthellae (Gymnodinium microadriaticum) in laboratory culture. The maximum specific growth rate (_max) was 0.35 d^-1 at 27 °C, 12 hL:12 hD cycle, 45 E m^-2 s^-1. Anmmonium and nitrate uptake by G. microadriaticum in distinct growth phases exhibited Michaelis-Menten kinetics. Ammonium half-saturation constants (K_s) ranged from 0.4 to 2.0 M; those for nitrate ranged from 0.5 to 0.8 M. Ammonium maximum specific uptake rates (V_max) (0.75 to 1.74 d^-1) exceeded those for nitrate (0.14 to 0.39 d^-1) and were much greater than the maximum specific growth rate (0.35 d^-1), suggesting that ammonium is the more significant N source for cultured zooxanthellae. Ammonium and nitrate V_max values compare with those reported from freshly isolated zooxanthellae. Light enhanced ammonium and nitrate uptake; ammonium inhibited nitrate uptake which was not reported for freshly isolated zooxanthellae, suggesting that physiological differences exist between the two. Knowledge of growth and nutrient uptake kinetics for cultured zooxanthellae can provide insight into the mechanisms whereby nutrients are taken up in coral-zooxanthelae symbioses.Contribution No. 1515 from the University of Maryland Center for Environmental and Estuarine Studies, Chesapeake Biological Laboratory, Solomons, Maryland 20688-0038, USA     

Characteristics of the uptake system for L-lysine and L-arginine inPhaeodactylum tricornutum

Cells ofPhaeodactylum tricornutum Bohlin develop the ability to take up L-lysine when they are deprived of nitrogen (illuminated in nitrogen-free medium), carbon (incubated in darkness) or both. Cells with a developed uptake system take up and accumulate lysine in an unchanged form. Uptake occurs under either aerobic or anaerobic conditions and is dependent on the presence of sodium⁺ ions (K _s ^{Na +}=,ca. 10 mM). Some potassium⁺ ions are necessary for uptake, presumably within the cells, but with potassium⁺-replete cells, increasing K⁺ concentration depresses lysine uptake. The lysine-uptake porter also transports L-arginine.K _s values are about 1.5 M for lysine and 0.5 M for arginine. It is, however, possible that the uptake system developed by incubating cells in darkness differs from that produced in light; it shows a pronounced pH optimum at pH 8.5, whereas the activity of the light-developed system declines from pH 6.5 to pH 9.0 and correlates well with the concentration of lysine⁺. The uptake system developed in darkness may also have a higher affinity for lysine. Lysine uptake is not inhibited by 1 mM concentrations of nitrate, nitrate, ammonium, or urea nor by similar concentrations of amphoteric or acidic amino acids.     

Enhancement of chlorophyll a production in Gulf Stream surface seawater by synthetic versus natural rain

Rainwater concentrations of either ammonium or nitrate were sufficient to stimulate chlorophyll a (chl a) production in bioassay experiments using Gulf Stream surface water collected off North Carolina during the summer of 1991. Previous studies primarily examined inshore waters and did not address the impact of rainwater ammonium. An increase in chl a occurred within 1 d of the addition of synthetic rainwater (2 or 5% rainwater, 98 or 95% seawater) containing up to 10 M ammonium; this increase was followed by a decrease in chl a the following day. A similar response to nitrate addition (5% addition of 20 M nitrate rain) was observed. In separate experiments, natural rainwater having nitrate and ammonium concentrations less than those in the experimental synthetic rain yielded a greater chl a response than synthetic rain when added at similar dilutions (0.5 to 5.0% rain). The maximum dissolved inorganic nitrogen concentration in the enriched seawater in these bioassays was 1.8 M; prior to enrichment the maximum was < 0.4 M. Bioassay experiments begun 2 d after a major storm event (sustained NE winds with gusts to 13 m s^-1 and ca. 390 mol m^-2 inorganic nitrogen deposition from rain) showed a chl a increase in response to addition of natural rainwater, but not to synthetic rainwater with similar dissolved inorganic nitrogen concentration. These results suggest that phytoplankton stimulants, in addition to nitrate and ammonium, exist in natural rain but not in the synthetic rain used in these experiments.     

Biological production of nitrite in seawater

The relative importance of 3 different sources for biological production of nitrite in seawater was studied. Decomposition of fecal pellets of the copepod Calanus helgolandicus (at a concentration of approximately 12 g-at N/l), in seawater medium, released small amounts of ammonia over a 6 week period. It nitrifying bacteria were added to the fecal pellets nitrite was barely detectable over the same period. Decomposition of phytoplankton (present at a concentration of about 8 g-at particulate plant N/l) with added heterotrophic bacteria, released moderate amounts of ammonia over a 12 week period. If the ammonia-oxidizing bacterium Nitrosocystis oceanus was added to the decomposing algae, nitrite was produced at a rate of 0.2 g-at N/l/week. Heterotrophic nitrification was not observed when 7 open-ocean bacteria were tested for their ability to oxidize ammonia. The diatom Skeletonema costatum, either non-starved or starved of nitrogen, produced nitrite when growing with 150 or 50 g-at NO ₂ ^- -N/l at a light intensity of about 0.01 ly/min. When nitrate in the medium was exhausted, S. costatum assimilated nitrite. If starved of vitamin B₁₂, both non-N-starved and N-starved cells of S. costatum produced nitrite in the medium with 150 g-at NO ₃ ^- -N/l. Nitrate was not exhausted and cell densities reached 2x10⁵/ml due to vitamin B₁₂ deficiency. If light intensity was reduced to 0.003 ly/min under otherwise similar conditions, cells did not grow due to insufficient light, and nitrite was not produced. In the sea, it appears that, in certain micro-environments, decomposition of particulate matter releases ammonia with its subsequent oxidation to nitrite. The amounts of these nutrients and the rate at which they are produced are dependent upon the nature of the materials undergoing decomposition and the associated bacteria. In certain other areas of the sea, where phytoplankton standing stock is high and nitrate is non-limiting, excretion by these organisms is a major source of nitrite.     

The common jellyfish Aurelia aurita: standing stock,excretion and nutrient regeneration in the Kiel Bight,Western Baltic

The population dynamics, ammonia and inorganic phosphate excretion, and nutrient regeneration of the common jellyfish Aurelia aurita was investigated from 1982 to 1984 in the Kiel Bight, western Baltic Sea. During summer 1982, medusae abundance ranged between 14 and 23 individuals 100 m^-3, biomass was estimated at about 5 g C 100 m^-3 and the mean final diameter of individuals was 22 cm. Abundance, based on numbers, in 1983 and 1984 was an order of magnitude lower; biomass was less than 2 g C 100 m^-3 and jellyfish grew to 30 cm. During the summers of 1983 and 1984, A. aurita biomass constituted roughly 40% of that of the total zooplankton>200 m. In 1982, for which zooplankton data were lacking, it was assumed that medusae biomass was greater than that of all other zooplankton groups. Total ammonia excretion ranged between 6.5 and 36 mol h^-1 individual^-1, whereas inorganic phosphate release was 1.4 to 5.7 mol h^-1 individual^-1. Allometric equations were calculated and exponents of 0.93 for NH₄–N release and 0.87 for PO₄–P excretion were determined. Nitrogen and phosphorus turnover rates were 5.4 and 14.6% d^-1, respectively. In 1982, the medusae population released 1 100 mol NH₄–N m^-2 d^-1, about 11% of the nitrogen requirements of the phytoplankton. The inorganic phosphate excretion (150 mol m^-2 d^-1) sustained 23% of the nutrient demands of the primary producers. In the other two years the nutrient cycling of the medusae was much less important, and satisfied only 3 to 6% of the nutrient demands. It is suggested that in some years A. aurita is the second most important source of regenerated nutrients in Kiel Bight, next to sediment.     

Partitioning of nitrogen and carbon in cultures of the marine diatom Thalassiosira fluviatilis supplied with nitrate,ammonium, or urea

Small or negligible differences in growth rates, average cell size, yields in cell numbers and total cell volumes were found in cultures of Thalassiosira fluviatilis inriched with nitrate, ammonium, or urea. Intracellular pools of unassimilated nitrate, nitrate, and ammonium were found in nutrient-rich conditions, but urea was not accumlated internally. Nitrogen assimilation into organic combination rather than nitrogen nutrient uptake was a critical rate-limiting step in nitrogen utilization. The free amino acid pool, protein, lipid-associated nitrogen, pigments, and total cell nitrogen were all highest in young or mature phase cells and decreased with age in senescent cells, whereas chitan, lipid, carbohydrate, and total cellular carbon all continued to increase during senescence. Dissolved organic nitrogen compounds accumulated in the medium only during senescence. C:N and lipid:protein were sensitive indicators of nitrogen depletion and age in T. fluviatilis.     

Effect of nitrogen supply on nitrogen uptake,accumulation and assimilation in Porphyra perforata (Rhodophyta)

Porphyra perforata J. Ag. was collected from a rocky land-fill site near Kitsilano Beach, Vancouver, British Columbia, Canada and was grown for 4 d in media with one of the following forms of inorganic nitrogen: NO ₃ ^- , NH ₄ ⁺ and NO ₃ ^- plus NH ₄ ⁺ and for 10 d in nitrogen-free media. Internal nitrogen accumulation (nitrate, ammonium, amino acids and soluble protein), nitrate and ammonium uptake rates, and nitrate reductase activity were measured daily. Short initial periods (10 to 20 min) of rapid ammonium uptake were common in nitrogen-deficient plants. In the case of nitrate uptake, initial uptake rates were low, increasing after 10 to 20 min. Ammonium inhibited nitrate uptake for only the first 10 to 20 min and then nitrate uptake rates were independent of ammonium concentration. Nitrogen starvation for 8 d overcame this initial suppression of nitrate uptake by ammonium. Nitrogen starvation also resulted in a decrease in soluble internal nitrate content and a transient increase in nitrate reductase activity. Little or no decrease was observed in internal ammonium, total amino acids and soluble protein. The cultures grown on nitrate only, maintained high ammonium uptake rates also. The rate of nitrate reduction may have limited the supply of nitrogen available for further assimilation. Internal nitrate concentrations were inversely correlated with nitrate uptake rates. Except for ammonium-grown cultures, internal total amino acids and soluble protein showed no correlation with uptake rates. Both internal pool concentrations and enzyme activities are required to interpret changes in uptake rate during growth.     

Assessment of simultaneous uptake of nitrogenous nutrients (15N) and inorganic carbon (13C) by natural phytoplankton populations

The simultaneous uptake of nitrogenous nutrients and inorganic carbon was measured in shipboard incubations of natural phytoplankton populations, using tracer additions of ¹³C-bicarbonate and ¹⁵N-labelled nitrogenous substrates. From March 1991 through March 1992, three stations on the Scotian Shelf (eastern Canada) were sampled monthly at ten depths in the euphotic zone. Additions of labelled nitrogen compounds ranged between 0.5 and 98% of ambient concentrations. Most of the C/N (at/at) uptake ratios were lower than the Redfield ratio, suggesting that nitrogen was not limiting. The fixation of carbon with and without addition of nitrate, ammonium or urea was generally similar. Some samples presented significant differences in carbon uptake rate between the four treatments, but these differences were not related to nitrogen enrichment (percent or nitrogen species). Given these results, the double-labelling method appears to be a reliable tool for measuring the simultaneous uptake of carbon and nitrogen by natural phytoplankton.