KINETICS OF SILICIC ACID UPTAKE AND RATES OF SILICA DISSOLUTION IN THE MARINE DIATOM THALASSIOSIRA PSEUDONANA1,2

Tracer techniques using the stable isotope ³⁰Si were used to measure rates of silicic acid uptake and silica dissolution in silicon replete and silicon depleted populations of 2 clones of the marine diatom Thalassiosira pseudonana Hasle & Heimdal. Uptake kinetics were describable using the Michaelis-Menten equation for enzyme kinetics, and no threshold concentration for uptake was evident. The maximum specific uptake rate of the estuarine clone 3H (0.062–0.092 · h^?1) was higher than that of the Sargasso Sea clone 13-1 (0.028–0.031 · h^?1), but half-saturation constants for uptake by the 2 clones were not measurably different (0.8–2.3 μM for 3H; 1.4–1.5 μM for 13-1). There was little or no light dependence of uptake in populations grown under optimal light conditions prior to the experiment. Exponentially growing populations released silicic acid to the medium by dissolution of cellular silica at rates ranging from 6.5 to 15% of the maximum uptake rate.     

Ammonium-limited growth and uptake by Oscillatoria agardhii in chemostat cultures

Growth of Oscillatoria agardhil was studied in ammonium-limited chemostat cultures, at various dilution rates (=growth rates, μ). The uptake kinetics for ammonium of nitrogen (ammonium or nitrate)-limited chemostat cultures also was investigated. The kinetics of ammonium-limited growth could be adequately described by both the Monod and Droop equations, and were closely similar to the nitrate-limited growth kinetics of this species. The uptake kinetics for ammonium showed similarities as well as differences with the uptake kinetics for nitrate. The similarities were apparent in the uptake capacity values for ammonium and nitrate , which were identical, high and independent of μ. The differences were to be found in the half-saturation constants for ammonium uptake and nitrate uptake , the former being hardly influenced by μ. A consitutive, high affinity, system is likely to operate in the uptake and assimilation of ammonium by nitrogen-limited O. agardhii. The use of ammonium uptake parameters in studies of growth-limiting factors in nature can provide information as to whether a nitrogen-limitation prevails in natural habitats of this species.     

LIGHT‐INDUCED RESPONSES OF OXYGEN PHOTOREDUCTION,REACTIVE OXYGEN SPECIES PRODUCTION AND SCAVENGING IN TWO DIATOM SPECIES1

Diatoms are frequently exposed to high light (HL) levels, which can result in photoinhibition and damage to PSII. Many microalgae can photoreduce oxygen using the Mehler reaction driven by PSI, which could protect PSII. The ability of Nitzschia epithemioides Grunow and Thalassiosira pseudonana Hasle et Heimdal grown at 50 and 300 μmol photons · m^?2 · s^?1 to photoreduce oxygen was examined by mass spectrometric measurements of ¹⁸O₂. Both species exhibited significant rates of oxygen photoreduction at saturating light levels, with cells grown in HL exhibiting higher rates. HL‐grown T. pseudonana had maximum rates of oxygen photoreduction five times greater than N. epithemoides, with 49% of electrons transported through PSII being used to reduce oxygen. Exposure to excess light (1,000 μmol photons · m^?2 · s^?1) produced similar decreases in the operating quantum efficiency of PSII (F_q′/F_m′) of low light (LL)‐ and HL‐grown N. epithemoides, whereas HL‐grown T. pseudonana exhibited much smaller decreases in F_q′/F_m′ than LL‐grown cells. HL‐grown T. pseudonana and N. epithemioides exhibited greater superoxide and hydrogen peroxide production, higher activities (in T. pseudonana) of superoxide dismutase (SOD) and ascorbate peroxidase (APX), and increased expression of three SOD‐ and one APX‐encoding genes after 60 min of excess light compared to LL‐grown cells. These responses provide a mechanism that contributes to the photoprotection of PSII against photodamage.     

Interactive Effects of CO2, Temperature,Irradiance, and Nutrient Limitation on the Growth and Physiology of the Marine Diatom Thalassiosira pseudonana (Coscinodiscophyceae)

The marine diatom Thalassiosira pseudonana was grown in continuous culture systems to study the interactive effects of temperature, irradiance, nutrient limitation, and the partial pressure of CO₂ (pCO₂) on its growth and physiological characteristics. The cells were able to grow at all combinations of low and high irradiance (50 and 300 μmol photons · m⁻² · s⁻¹, respectively, of visible light), low and high pCO₂ (400 and 1,000 μatm, respectively), nutrient limitation (nitrate-limited and nutrient-replete conditions), and temperatures of 10–32°C. Under nutrient-replete conditions, there was no adverse effect of high pCO₂ on growth rates at temperatures of 10–25°C. The response of the cells to high pCO₂ was similar at low and high irradiance. At supraoptimal temperatures of 30°C or higher, high pCO₂ depressed growth rates at both low and high irradiance. Under nitrate-limited conditions, cells were grown at 38 ± 2.4% of their nutrient-saturated rates at the same temperature, irradiance, and pCO₂. Dark respiration rates consistently removed a higher percentage of production under nitrate-limited versus nutrient-replete conditions. The percentages of production lost to dark respiration were positively correlated with temperature under nitrate-limited conditions, but there was no analogous correlation under nutrient-replete conditions. The results suggest that warmer temperatures and associated more intense thermal stratification of ocean surface waters could lower net photosynthetic rates if the stratification leads to a reduction in the relative growth rates of marine phytoplankton, and at truly supraoptimal temperatures there would likely be a synergistic interaction between the stresses from temperature and high pCO₂ (lower pH).     

COPPER‐UPTAKE KINETICS OF COASTAL AND OCEANIC DIATOMS1

We investigated copper (Cu) acquisition mechanisms and uptake kinetics of the marine diatoms Thalassiosira oceanica Hasle, an oceanic strain, and Thalassiosira pseudonana Hasle et Heimdal, a coastal strain, grown under replete and limiting iron (Fe) and Cu availabilities. The Cu‐uptake kinetics of these two diatoms followed classical Michaelis–Menten kinetics. Biphasic uptake kinetics as a function of Cu concentration were observed, suggesting the presence of both high‐ and low‐affinity Cu‐transport systems. The half‐saturation constants (K_m) and the maximum Cu‐uptake rates (V_max) of the high‐affinity Cu‐transport systems (～7–350 nM and 1.5–17 zmol · μm^?2 · h^?1, respectively) were significantly lower than those of the low‐affinity systems (>800 nM and 30–250 zmol · μm^?2 · h^?1, respectively). The two Cu‐transport systems were controlled differently by low Fe and/or Cu. The high‐affinity Cu‐transport system of both diatoms was down‐regulated under Fe limitation. Under optimal‐Fe and low‐Cu growth conditions, the K_m of the high‐affinity transport system of T. oceanica was lower (7.3 nM) than that of T. pseudonana (373 nM), indicating that T. oceanica had a better ability to acquire Cu at subsaturating concentrations. When Fe was sufficient, the low‐affinity Cu‐transport system of T. oceanica saturated at 2,000 nM Cu, while that of T. pseudonana did not saturate, indicating different Cu‐transport regulation by these two diatoms. Using CuEDTA as a model organic complex, our results also suggest that diatoms might be able to access Cu bound within organic Cu complexes.     

Effect of different CO2 concentrations on biomass,pigment content,and lipid production of the marine diatom Thalassiosira pseudonana

The marine diatom Thalassiosira pseudonana grown under air (0.04% CO₂) and 1 and 5% CO₂ concentrations was evaluated to determine its potential for CO₂ mitigation coupled with biodiesel production. Results indicated that the diatom cultures grown at 1 and 5% CO₂ showed higher growth rates (1.14 and 1.29 div day⁻¹, respectively) and biomass productivities (44 and 48 mg_AFDWL⁻¹ day⁻¹) than air grown cultures (with 1.13 div day⁻¹ and 26 mg_AFDWL⁻¹ day⁻¹). The increase of CO₂ resulted in higher cell volume and pigment content per cell of T. pseudonana. Interestingly, lipid content doubled when air was enriched with 1–5% CO₂. Moreover, the analysis of the fatty acid composition of T. pseudonana revealed the predominance of monounsaturated acids (palmitoleic-16:1 and oleic-18:1) and a decrease of the saturated myristic acid-14:0 and polyunsaturated fatty acids under high CO₂ levels. These results suggested that T. pseudonana seems to be an ideal candidate for biodiesel production using flue gases.

     

NITROGEN UTILIZATION AND METABOLISM RELATIVE TO PATTERNS OF N2 FIXATION IN CULTURES OF TRICHODESMIUM NIBB1067

We measured uptake kinetics for four combined N sources, ambient rates of N uptake and N₂ fixation, glutamine synthetase activity (transferase and biosynthetic), and concentrations of intracellular pools of glutamate (glu) and glutamine (gln) in cultures of Trichodesmium NIBB1067. N dynamics and metabolism were examined to assess the relative importance of N₂ fixation and N uptake to Trichodesmium nutrition. Comparisons were made between cultures grown on medium without added N, with excess NO, or with excess urea. Of the combined N sources tested, Trichodesmium NIBB1067 had the highest affinity for NH; high uptake capacities for NH, urea, and glu; and little capacity for NO uptake. In cultures grown on medium without added N, NH accumulated in the medium during growth, resulting in high NH uptake rates relative to rates of N₂ fixation. Glu uptake rates were low but consistent throughout the diel period. In cultures grown on excess NO or urea, uptake of these compounds supplied the majority of the daily N demand, although some N₂ fixation occurred during the light period. NO uptake rates were reduced when N₂-fixation rates were high. In all of the cultures, the highest gln/glu ratios and the lowest glutamine synthetase transferase/biosynthetic ratios were observed during the period when rates of total N uptake were highest. In cultures growing exponentially on medium without added N, N₂ fixation accounted for 14%– 16% of the total daily N uptake. Uptake of NH and glu, presumably regenerated within the culture vessels, represented 84%–86% of the daily N uptake. Because these systems were closed, net growth was constrained by the rate at which N₂ could be fixed into the system. However, total daily N turnover was greater than that necessary to accommodate the observed increase in culture biomass. The rapid N turnover rates observed in these cultures may support gross productivity and balance the high rates of C fixation observed in natural populations of Trichodesmium.     

NITROGEN ASSIMILATION OF AN OCEANIC DIATOM IN NITROGEN-LIMITED CONTINUOUS CULTURE1

The oceanic diatom Thalassiosira pseudonana Hasle and Heimdal (formerly Cyclotella nana) was grown with 12L:12D illumination cycles in nitrogen-limited continuous culture with a mixture of ammonium and nitrate as the N source. Measurements included, at 3 different growth rates (degrees of N limitation), cell concentration, cell carbon, nitrogen, and chlorophyll a contents, cell volume, photosynthetic carbon assimilation vs. irradiance, short-term uptake of ammonium and nitrate vs. their ambient concentrations, and in vitro activities of the assimilatory enzymes nitrate reductase and glutamic dehydrogenase. The various parameters showed either an increase (pattern a) or a decrease (pattern b) with increasing N limitation. Those following pattern a were nitrate reductase activity and the capacity to assimilate nitrate and ammonium. Those following pattern b were glutamic dehydrogenase activity, photosynthetic rate, nitrogen:carbon and chlorophyll a:carbon composition ratios. Results are discussed in terms of the interpretation such measurement for natural phytoplankton and effects of circadian periodicity.     

INTERACTIONS BETWEEN PULSED NUTRIENT SUPPLIES AND A PHOTOCYCLE AFFECT PHYTOPLANKTON COMPETITION FOR LIMITING NUTRIENTS IN LONG-TERM CULTURE1

The influence of periodic nutrient supplies and a photocycle on phytoplankton competition for limiting nutrients was examined using the diatoms Thalassiosira rotula Meunier (clone 411) and Chaetoceros sp. cf. vixvisibilis Schiller (clone 847). Chaetoceros sp. cf. vixvisibilis displaced T. rotula from ammonium-limited cultures under constant light irrespective of whether ammonium was supplied continuously, in 6 pulses.day^?1 or in a single daily pulse. In contrast, the species coexisted under the 14:10 h LD photocycle under either continuous or pulsed ammonium supplies with the relative abundance of C. sp. cf. vixvisibilis increasing as the interval between ammonium additions lengthened. Coexistence was not observed with either silicic acid or nitrate limitation. Chaetoceros sp. cf. vixvisibilis displaced T. rotula from both nitrate- and silicic acid-limited chemostat cultures and from semi-continuous cultures grown under the same photoperiod that produced coexistence with a daily pulse of ammonium. The presence of a photocycle was both necessary and sufficient to permit coexistence with ammonium limitation. Under continuous ammonium supply the photocycle may have induced a temporal separation of ammonium uptake between species, permitting sharing of the limiting nutrient and coexistence. In contrast, the species were shown to be in direct competition for the daily ammonium pulse. A competition model suggested that coexistence in this case arose from a balance between the species’ammonium uptake rates and their nitrogen demands for steady-state growth induced by the photocycle. The results indicate that variations in nutrient supply rates may contribute to the coexistence of phytoplankton species in the sea, but that the identity of the limiting nutrient and the influence of variations in other non-limiting resources play important roles in affecting the outcome of nutrient competition among planktonic algae.     

Nitrate assimilatory properties of barley grown under long-term N limitation: Effects of local nitrate supply in split-root cultures

The effect of nitrate availability on characteristics of the nitrate assimilatory system was investigated in N-limited barley (Hordeum valgare L. cv. Golf), grown with the seminal root system split into initially equal-sized halves. The cultures were continuously supplied with nitrate-N at a relative addition rate (RA) of 0.09 day^?1, which resulted in relative growth rates (RG) that were ca 85% of those observed under surplus nitrate nutrition. The total N addition was divided between the subroots in ratios of 100:0, 80:20, 70:30, 60:40, and 50:50. For comparison, standard cultures were grown at RAs ranging from 0.03 to 0.18 day^?1. Initially, biomass and N partitioning to the subroots responded strongly and proportionally to the nitrate distribution ratio. After 12-14 days no further effect was observed. The V_max for net nitrate uptake and in vitro nitrate reductase (NR) activity were measured in acclimated plants, i.e., after > 14 days under a certain nitrate regime. In subroots fed from 20 to 100% of the total N addition, V_max for net nitrate uptake increased slightly, whereas NR activity was unaffected. Uptake and NR activities were insignificant in the 0%-subroot. Uneven nitrate supply to individual subroots had negligible effect on the whole-plant ability for nitrate uptake, and the relative V_max (unit N taken up per unit N in whole plant tissue and time) remained about 7-fold in excess of the demand set by growth. Balancing nitrate concentrations (the resulting external nitrate concentrations at a certain RA) generally ranged between 2 and 10 μM at growth-limiting RA, both when predicted from uptake kinetics and when actually measured. When comparing split root and standard cultures when acclimated, it appears that uptake and NR activities in roots respond more strongly to over-all nitrate availability than to nitrate availability to individual subroots.     

Acclimation conditions modify physiological response of the diatom Thalassiosira pseudonana to elevated CO2 concentrations in a nitrate‐limited chemostat

Diatoms are responsible for a large proportion of global carbon fixation, with the possibility that they may fix more carbon under future levels of high CO₂. To determine how increased CO₂ concentrations impact the physiology of the diatom Thalassiosira pseudonana Hasle et Heimdal, nitrate‐limited chemostats were used to acclimate cells to a recent past (333 ± 6 μatm) and two projected future concentrations (476 ± 18 μatm, 816 ± 35 μatm) of CO₂. Samples were harvested under steady‐state growth conditions after either an abrupt (15–16 generations) or a longer acclimation process (33–57 generations) to increased CO₂ concentrations. The use of un‐bubbled chemostat cultures allowed us to calculate the uptake ratio of dissolved inorganic carbon relative to dissolved inorganic nitrogen (DIC:DIN), which was strongly correlated with fCO₂ in the shorter acclimations but not in the longer acclimations. Both CO₂ treatment and acclimation time significantly affected the DIC:DIN uptake ratio. Chlorophyll a per cell decreased under elevated CO₂ and the rates of photosynthesis and respiration decreased significantly under higher levels of CO₂. These results suggest that T. pseudonana shifts carbon and energy fluxes in response to high CO₂ and that acclimation time has a strong effect on the physiological response.     

EFFECTS OF COPPER TOXICITY ON SILICIC ACID UPTAKE AND GROWTH IN THALASSIOSIRA PSEUDONANA11

The toxicity of Cu to Thalassiosira pseudonana (Hustedt) Hasle and Heimdal was investigated by examining both short and long term effects of Cu on cellular processes. Toxic levels of Cu (pCu* < 13) were found to inhibit short term Si(OH)₄ uptake rates with kinetics characteristic of irreversible inhibition at a hypothetical Cu-sensitive Si(OH)₄ transport site. Residual Si(OH)₄ concentrations (those below which no uptake could occur) were found to increase with increasing levels of Cu, and the toxic effects of Cu could be reversed by increasing the concentrations of Si(OH)₄ in the medium. The actual uptake of Cu by the cells was found to vary inversely with the ambient Si(OH)₄ concentration. Copper did not inhibit the uptake of NO₃^? or PO₄^3-. The long term inhibition of growth rate by Cu in this species was shown not to be a result of Si deficiency caused by the inhibition of Si(OH)₄ uptake. Cu inhibited cells were found to have higher Si cell quotas (including a sizable soluble pool) than the control cultures. They were, however, observed to have aberrant frustules, significantly larger than the control cells, suggesting interference with the silification process as a possible mechanism for inhibition of growth by Cu. A conceptual model is proposed for the Cu-Si(OH)₄-growth relationship. It includes a Cu sensitive Si(OH)₄ transport site that may also serve to transport Cu into the cell, and growth inhibition mediated by intracellular Cu concentrations which may block cell division or cause general cellular disfunction.     

Regulation of nitrate uptake by amino acids in maize cell suspension culture and intact roots

The effect of amino acids on nitrate transport was studied in Zea mays cell suspension cultures and in Zea mays excised roots. The inclusion of aspartic acid, arginine, glutamine and glycine (15mM total amino acids) in a complete cell-culture media containing 1.0 mM NO₃ ^- strongly inhibited nitrate uptake and the induction of accelerated uptake rates. The nitrate uptake rate increased sharply once solution amino acid levels fell below detection limits. Glutamine alone inhibited induction in the cell suspension culture. Maize seedlings germinated and grown for 7 days in a 15 mM mixture of amino acids also had lower nitrate uptake rates than seedlings grown in 0.5 mM Ca(NO₃)₂ or 1 mM CaCl₂. As amino acids are the end product of nitrate assimilation, the results suggest an end-product feed-back mechanism for the regulation of nitrate uptake.     

STUDIES OF CARBON AND NITROGEN METABOLISM BY THREE MARINE PHYTOPLANKTON SPECIES IN NITRATE-LIMITED CONTINUOUS CULTURE1,2

Characteristics of carbon production, excretion and dark respiration, and nitrate uptake kinetics were studied using continuous culture techniques for Thalassiosira allenii Takano, Monorhrysis lutheri Droop and Dunaliella tertiolccta Butcher. Fur T. allenii. the ratio of dark C loss to daytime net C production varied between 0.1 and 0.2 over a growth rate range from ca. 0.005 to 0.06 h^-1. For M. lutheri and D. tertiolecta. this same ratio varied belween 0.2 and 0.3 between growth rates of ca. 0.005 and 0.025 h^-1, but declined at higher growth rates when the dark nitrate uptake capacity of the cells was exceeded by the pumping rate. Carbon excretion rates averaged less than 1.5% of daytime net C production rates. Productivity indices showed little correlation with growth rate, due to the significant poisitive correlation between chl a:C ratios and growth rate. Chlorophyll a:C ratios for T. allenii were less than 0.01 al growth rates less than 0.03 h^-1, and appoached zero at zero growth rate. Dark nitrate maximum uptake rates for M. lutheri, D. tertiolecta and T. allenii averaged 23, 64 and 120%, respectively, of light nitrate maximum uptake rates. Excretion of nitrite was observed during most nitrate uptake experiments. This excretion reduced net uptake of nitrate spikes in the dark for M. lutheri and D. tertiolecta by 79 and 23%, respectively.     

INTERACTION (ALLELOPATHY) BETWEEN MARINE DIATOMS: THALASSIOSIRA PSEUDONANA AND PHAEODACTYLUM TRICORNUTUM1

Two marine diatoms were studied singly and in mixed culture. Thalassiosira pseudonana (Hust.) Hade & Heimdal was capable of a higher growth rate (μ_max) than Phaeodactylum tricornutum Bohlin. In two-species batch cultures P. tricornutum took over in the latter portion of the exponential phase, possibly due to allelopathy. The filtrate from this species caused an initial lag phase and a reduced terminal population density for T. pseudonana. Two-species continuous cultures showed verification of these interactions. At high dilution rate (i.e., high growth rate) P. tricornutum washed out when added at low density, whereas T. pseudonana maintained constant cell density. However, when sufficient density of P. tricornutum was added as a contaminant, both species washed out. At a lower dilution rate P. tricornutum increased in density when added and eventually reached a stable population; T. pseudonana then washed out.     

COMPARATIVE KINETIC STUDIES OF NITRATE-LIMITED GROWTH AND NITRATE UPTAKE IN PHYTOPLANKTON IN CONTINUOUS CULTURE1

A comparative study of nitrate-limited growth and nitrate uptake was carried out in chemostat cultures of Ankistrodesmus falcatus (Corda) Ralfs., Asterionella formosa Hass., and Fragilaria crotonensis Kit. In each species growth rate (μ) was related to total cell nitrogen or cell quota (q) by the empirical Droop growth function. Nitrate uptake was a function of both external N concentration and q. The apparent maximum uptake rate (V_m') at a given μ was inversely related to q – q₀, where q₀ is the minimum quota. The apparent half-saturation constant for uptake, (K_m') appears to show a slight inverse trend with μ, although statistical analysis shows that this trend is inconclusive. When q approaches q₀, V_m' is several orders of magnitude greater than μq, the calculated steady-state uptake rate. As q increases, however, the difference between these two variables decreases sharply until q approaches q_m, the cell quota for nitrogen-rich cells. At this point the difference between μq and V_m' disappears. This behavior is explained by the feedback regulation of N uptake. The inverse relationship between V_m' and q – q₀ can be described by an empirical three-parameter equation.     

GROWTH OF VITAMIN B12-LIMITED CULTURES: THALASSIOSIRA PSEUDONANA,MONOCHRYSIS LUTHERI,AND ISOCHRYSIS GALBANA1,2

The growth rates of 3 species of phytoplankton were found to be dependent on the vitamin B₁₂ concentrations in the media. In batch cultures, the vitamin B₁₂ half-saturation constants and standard errors were 0.39 ± 0.042 μμg/ml for Thalassiosira pseudonana (clone 3H), 1.69 ± 0.24 μμg/ml for Isochrysis galbana, and 2.77 ± 1.65 μμg/ml for Monochrysis lutheri. A chemostat was used to grow T. pseudonana with vitamin B₁₂ as the controlling factor. In the chemostat the yield and standard deviation, 102 ± 21 × 10⁴ cells/μμg vitamin B₁₂, was the same as in the batch culture, 126 ± 13 ± 10⁴ cells/μμg. The chemostat half-saturation constant, 0.26 ± 0.068 μμg/ml vitamin B₁₂, and maximum growth rate were in agreement with those obtained in batch cultures. Vitamin concentrations for maximum growth, rates were greater than those calculated necessary from yield data to give observed population densities similar to those in natural waters. In the sea the effect of vitamin B₁₂ concentration on growth rates may be complicated by low concentrations of other nutrients or the presence of inhibitors.     

RELATIONSHIPS BETWEEN NITRATE REDUCTASE ACTIVITY AND RATES OF GROWTH AND NITRATE INCORPORATION UNDER STEADY-STATE LIGHT OR NITRATE LIMITATION IN THE MARINE DIATOM THALASSIOSIRA PSEUDONANA (BACILLARIOPHYCEAE)1

Although activity of the enzyme nitrate reductase (NR) can potentially be used to predict the rate of nitrate incorporation in field assemblages of marine phytoplankton, application of this index has met with little success because the relationship between the two rates is not well established under steady-state conditions. To provide a basis for using NR activity measurements, the relationships among NR activity, growth rate, cell composition, and nitrate incorporation rate were examined in cultures of Thalassiosira pseudonana (Hustedt)Hasle and Heimdal, growing a) under steady-state light limitation, b) during transitions between low and high irradiance (15 or 90 μmol quanta.m^?2.s^?1), and c) under steady-state nitrate limitation. Using a modified assay for NR involving additions of bovine serum albumin to stabilize enzyme activity, NR activity in light-limited cultures was positively and quantitatively related to calculated rates of nitrate incorporation, even in cultures that were apparently starved of selenium. During transitions in irradiance, growth rates acclimated to new conditions within 1 day; through the transition, the relationship between NR activity and nitrate incorporation rate remained quantitative. In nitrate-limited chemostat cultures, NR activity was positively correlated with growth rate and with nitrate incorporation rates, but the relationship was not quantitative. NR activity exceeded nitrate incorporation rates at lower growth rates (<25% of nutrient-replete growth rates), but chemostats operating at such low dilution rates may not represent ecologically relevant conditions for marine diatoms. The strong relationship between NR activity and nitrate incorporation provides support for the idea that NR is rate-limiting for nitrate incorporation or is closely coupled to the rate-limiting step. In an effort to determine a suitable variable for scaling NR activity, relationships between different cell components and growth rate were examined. These relationships differed depending on the limiting factor. For example, under light limitation, cell volume and cell carbon content increased significantly with increased growth rate, while under nitrate limitation cell volume and carbon content decreased as growth rates increased. Despite the differences found between cell composition and growth rate under light and nitrate limitation, the relationships between NR activity scaled to different compositional variables and growth rate did not differ between the limitations. In field situations where cell numbers are not easily determined, scaling NR activity to particulate nitrogen content may be the best alternative. These results establish a strong basis for pursuing NR activity measurements as indices of nitrate incorporation in the field.