Effect of Fe3+ on the growth and lipid content of Isochrysis galbana

Inducing lipid accumulation in microalgae cells without suppressing cell growth is vital to the economical production of biodiesel from microalgae. In two experiments, we demonstrate that the cell concentration and lipid content of marine microalgae Isochrysis galbana depend upon the iron concentration in the growth media. In Experiment I, adding chelated FeCl 3 to the medium at the late exponential growth phase prolonged this phase and increased the lipid content in I. galbana cells. The fi nal cell density and lipid content of I. galbana supplemented with chelated FeCl 3 was approximately 2 and 1.65 times higher than that of non-supplemented cultures, respectively. In Experiment II, I. galbana cells in the late exponential phase were collected and re-inoculated into new media containing Fe 3+ at various concentrations. The fi nal cell concentration and lipid content were maximized at the highest iron concentration(38% biomass by dry weight at 1.2×10-5 mol/L Fe 3+). In this study, intracellular neutral lipid storage was evaluated by fl uorescent spectrophotometry using fl uorochrome Nile red, and the measurement conditions were optimized.     

Removal of K^＋, Na^＋, Ca^2＋, and M^g2＋ from saline-alkaline water using the microalga Scenedesmus obliquus

The capability ofScenedesmus obliquus to remove cations （K^＋, Na^＋, Ca^2＋, Mg^2＋） from saline- alkaline water was investigated at different salinities （0, 5, 10, 15, 20, 25） and carbonate alkalinities （0, 5, 10, 15, 20, 25, 30, 35 mmol/L）. K^＋, Na^＋, Ca^2＋, and Mg^2＋ in saline-alkaline water were efficiently removed by S. obliquus. The maximum removal of the cations （29.37 mg for K^＋, 185.85 mg for Na^＋, 23.07 mg for Ca^2＋, 66.14 mg for Mg^2＋） occurred at salinity 25. The maximum removal of K^＋ （2.28 mg）, Na＋ （6.62 mg）, Ca^2＋ （1.01 mg）, and Mg2＋ （0.62 mg） occurred at carbonate alkalinities of 25 mmol/L for K＊, 35 mmol/L for Na＋, 20 mmol/L for Ca2＋, and 25 mmol/L for Mf＋, respectively. Under a salinity stress, the concentration of Na＇ in S. obliquus increased significantly, while that of K~ decreased significantly. The concentrations of Ca^2＋ and Mg2＋ decreased as well. The ratios of K＋/Na~, Ca2＋/Na^＋, and Mg^2＋/Na^＋ were significantly lower in all salinity treatments than those of the control. Under alkaline stress, the concentrations of Nan and K＋ in S. obliquus decreased significantly and the ratios of K^＋/Na^＋, Ca2＋/Na^＋, and Mg^2＋/Na^＋ were significantly higher in all treatments than in the control. Moreover, the concentrations of Ca2＋ and Mg2＋ in S. obliquus at alkalinities of 5-10 mmol/L were significantly higher than those of the other treatments. The removal of Na＋ by S. obliquus mainly occurs through biosorption, and Mg^2＋ and Ca^2 ＋ were removed through both biosorption and bioaccumulation.     

Production of dimethylsulfide and acrylic acid from dimethylsulfoniopropionate during growth of three marine microalgae

We measured the concentrations of dimethylsulfide （DMS）, acrylic acid （AA）, and dimethylsulfoniopropionate （DMSP） during growth of three microalgae： Prorocentrum micans, Gephyrocapsa oceanica, and Platymonas subcordiformis. The DMSP, AA, and DMS concentrations in culture media varied significantly among algal growth stages, with the highest concentrations in the late stationary growth stage or the senescent stage. In the stationary growth stage, the average DMSP concentration per cell in P. micans （0.066 5 pmol/cell） was 1.3 times that in G. oceanica （0.049 5 pmol/ cell） and 20.2 times that in P. subcordiformis （0.003 29 pmol/cell）. The average concentrations of AA were 0.044 6, 0.026 9, and 0.003 05 pmol/cell in P. micans, G. oceanica, and P. subcordiformis, respectively, higher than the concentrations of DMS （0.272, 0.497, and 0.086 2 fmol/cell, respectively）. There were significant positive correlations between cell density and AA, DMSP, and DMS concentrations. The ratios of DMS/AA and AA/（DMSP＋AA） in the three algae differed significantly over the growth cycle. In all three microalgae, the DMS/AA ratios were less than 25% during the growth period, suggesting that the enzymatic cleavage pathway, which generates DMS, was not the main DMSP degradation pathway. The changes in the DMS/AA ratio indicated that there was a higher rate of enzymatic breakdown of DMSP in the early growth period and a lower rate during senescence. In all three microalgae, the AA/（DMSP＋AA） ratio （degradation ratio of DMSP） decreased during the exponential growth phase, and then increased. The variations in these ratios can approximately indicate the cleavage mechanism of DMSP at different stages of algal growth.     

EFFECT OF ZINC ON THE GROWTH AND DEVELOPMENT OF LARVAE OF BAY SCALLOP ARGOPECTEN IRRADIANS

The 48-h straight-hinge larvae of Argopecten irradians were exposed for 9 days to various concentrations of zinc in seawater. The growth rate of larvae in the 50 ppb Zn mediums was 77% that of the controls, and nearly zero in the 200 ppb Zn mediums. A progressive decrease in growth and increase in larval deformity and mortality was observed with increasing zinc concentrations from 50 to 200 ppb. 50 and 100 ppb Zn resulted in delayed development of eye spot and metamorphosis and reduced numbers of larvae at both stages. All larvae subjected to higher zinc concentrations died before attaining the eye-spot stage. The zinc concentrations which caused 50% reduction in growth (G C50)and 50% mortality (LC 50)were 109 and 120 ppb respectively. Growing in the medium with 100 ppb Zn, the larvae fed with Isochysis galbana cultured in a medium with 100 ppb Zn showed more suppressed growth and development than those fed with nonnal food. It is suggested that zinc contaminated food has strong influence on the bay sca     

Effects of waterborne Fe(II) on juvenile turbot Scophthalmus maximus: analysis of respiratory rate, hematology and gill histology

The concentration of Fe(Ⅱ) is high in some groundwater supplies used in turbot culture,and the toxicity of waterborne Fe(Ⅱ) is unknown.We investigated the stress responses of juvenile turbot,Scophthalmus maximus,exposed to Fe(Ⅱ) of different concentrations (0.01,0.05,0.1,0.5,1,and 2 mg/L) for 1,7,14,and 28 d,under the same ambient conditions of other parameters.Changes in respiratory rate,hematological parameters,and gill structure were determined.The results show that waterborne Fe(Ⅱ) did not cause severe hematological perturbation to turbot.A low-medium Fe(Ⅱ) concentration (lower than 0.1 mg/L) could boost the respiratory rate,and caused no or very limited damage to fish.A high Fe(Ⅱ) concentration (0.1 mg/L or higher),however,caused gill damage,such as vacuoles in branchial lamellae,epithelial necrosis,and hypertrophy of epithelial cells,and even death after extended exposure time.Therefore,excess waterborne Fe(Ⅱ) and long-term exposure to Fe(Ⅱ) could be responsible for poor growth and high mortality of turbot in culture.The concentration of waterborne Fe(Ⅱ) in turbot culture should be kept below 0.1 mg/L.     

ESTIMATION OF ATMOSPHERIC INPUT OF MERCURY TO SOUTH LAKE AND JINGYUE POOL

Taking South Lake and Jingyue Pool in Changchun as examples,we determined particulate Hg in ambient air and Hg concentration in precipitation.Results indicated that particulate Hg in air and Hg concentration in precipita-tion in both sites in nonheating period were lower than that in heating period,which is influenced by coal combustion.The annual particulate Hg concentrations in precipitation were 0.268μg/L(South Lake)and 0.108μg/L(Jingyue Pool).The high-er particulate Hg concentrations in precipitation were 0.268μg/L(South Lake) and 0.108μg/L(Jingyue Pool).The high-er particulate Hg concentration in air and Hg concentration in precipitation in South Lake than that of Jingyue Pool indicat-ed that mercury deposition was influenced by anthropogenic sources(especially from coas combustion).Hg concentration in precipitation is related with particulate Hg in the atmosphere.Dry deposition fluxes were estimated by using dry deposi-tion model on the base of particulate Hg concentrations in the atmosphere,they were estimated by using dry deposi-tion model on the base of particulate Hg concentrations in the atmosphere,they were 35.5μg/(m^2.a) for South lake and 15.3μg/(m^2.a)for Jingyue Pool.Combined with precipitation amount,wet deposition fluxes were estimated in these two sites,152.2μg(m^2.a)for South Lake and 61.2μg/(m^2.a)for Jingyue Pool.Atmospheric Hg deposition fluxes were 160.2g/a for South Lake and 328.62g/a for Jingyue Pool.e     

Effect of Dietary Potassium on Growth,Nitrogen Metabolism,Osmoregulation and Immunity of Pacific White Shrimp （Litopenaeus vannamel） Reared in Low Salinity Seawater

An 8 weeks feeding experiment was conducted to determine the effect of dietary potassium on the growth and physio-logical acclimation of Pacific white shrimp （Litopenaeus vannamei） reared in diluted seawater （salinity 4）. Six semi-purified practical diets containing 0.59, 0.96, 1.26, 1.48, 1.74, and 2.17 g potassium K＋ per 100 g diet were formulated, respectively. The survival and feed conversion rate did not show significant difference among groups of shrimps given these diets （P〉0.05）. The shrimps fed the diets containing 0.96-1.48 g K＋ per 100g diet gained the highest weight, specific growth rate, and protein efficiency ratio. Their ammonium-N excretion rate as well as hemolymph concentration of Na＋ and C1- were significantly lower than those of the control （P〈0.05）, but a reverse trend was observed for their gill Na＋/K＋-ATPase. Moreover, the shrimps fed with 1.48 g K＋ per 100 g diet were the highest in hemolymph urea level, and the phenoloxidase and lysozyme activities were significantly higher than those of the control （P〈0.05）. The growth and physiological response of the test shrimps suggested that diet containing 1.48 g K＋ per 100 g diet improved the growth of L. vannamei in low-salinity seawater, and enhanced the physiological acclimation of the organism.     

The UV-absorbing compounds in gametophytic blades of Porphyra haitanensis and the effect on formation of sporangial branchlets of conchocelis

Endogenous levels of UVAC in different parts of matured gametophytic blades of Porphyra haitanensis and the effect of exogenous UVAC on sporangial branchlet formation are studied. The UVAC value in vegetative (36. 78 mg/g) and reproductive (29.25 mg/g) parts of female thallus is higher than their counterparts in male thallus (34.68 mg/g and 20.46 mg/g). The UVAC value of reproductive parts is higher than vegetative parts of same sexual thallus. The optimal UVAC concentration for sporangial branchlet formation is 0.1 mg/g.     

Photosynthetic responses of thalli and isolated protoplasts of Bryopsis hypnoides (Bryopsidales, Chlorophyta) during dehydration

Bryopsis hypnoides Lamouroux is a unique intertidal siphonous green alga whose extruded protoplasm can aggregate spontaneously in seawater to form numerous new cells that can develop into mature algal thalli.In this study,the photosynthetic responses during dehydration of both the thalli and protoplasts isolated from B.hypnoides were measured using a Dual-PAM(pulse amplitude modulation)-100 fluorometer.The results show that the photosynthetic rates of B.hypnoides thalli were maintained for an initial period,beyond which continued desiccation resulted in reduced rates of PSI and PSII.However,the photosynthetic performances of the isolated protoplasts dehydrated in air(CO2 concentration 600±700 mg/L) showed a slight increase of Y(II) at 20% water loss,but the rates decreased thereafter with declining water content.When protoplasts were dehydrated in CO2 deficient conditions(CO2 concentration 40±80 mg/L),the values of Y(II) declined steadily with increased dehydration without an initial rise.These results indicated that the thalli and isolated protoplasts of this alga can utilize CO2 in ambient air effectively,and the photosynthetic performances of the isolated protoplasts were significantly different from that of the thalli during dehydration.Thus the protoplasts may be an excellent system for the study of stress tolerance.     

Effects of lithium on the survival,growth,and reproduction of Daphniopsis tibetana Sars(Crustacea:Cladocera)

We evaluated the effects of lithium on the survival,growth,and reproduction of D.tibetana in the laboratory.The safe concentration value was 69.3 mg/L.The time to first brood was significant lower for individuals reared in 5 and 10 mg/L(24.4 and 24.0 d,respectively) compared with individuals reared in20,40,or 60 mg/L.Females reared in 5 mg/L lithium produced a mean of 16.5 neonates/brood and had the highest number of broods(3.0±1.95).The rate of egg production(a),the intrinsic rate of increase(r_m),net reproduction rate(R_0),and finite rate of increase(λ) were highest for D.tibetana reared at 5 and 10 mg/L lithium,and the duration of development was shorter than for the remaining groups.The results indicated that rearing in 5-10 mg/L lithium can accelerate the growth and reproduction of D.tibetana.     

The Influences of Mg^2 ,Ca^2 ,and Mg^2 /Ca^2 Ratio in Mixed Seawater on the Emergence Rate of Penaeus Japonicus Postlarva

This paper reports the approprite ranges of Mg^2 ,Ca^2 and their ratio Mg^2 /Ca^2 in mixed seawater for rearing of Penaeus japonicus larvae,The ranges for the above three indices are 1150-1450mg/L,360-440mg/L and 2.8-3.4,respectively,the proper salinity range of mixed seawater is 22.1-33.9 obtained by mixing estuarine water and concentrated seawater.     

Effects of heavy metals （Pb^2＋ and Cd^2＋） on the ultrastructure, growth and pigment contents of the unicellular cyanobacterium Synechocystis sp. PCC 6803

The unicellular cyanobacterium Synechocystis sp. PCC 6803, a model organism known for its unique combination of highly desirable molecular genetic, physiological and morphological characteristics, was employed in the present study. The species was cultured in BG11 liquid medium contained various initial concentrations of Pb²⁺ and Cd²⁺ (0, 0.5, 1, 2, 4, 6 and 8 mg/L). The experiment was conducted for six days and the metal induced alterations in the ultrastructure, growth and pigment contents were assessed. Alterations in the ultrastructure of the Synechocystis sp. PCC 6803 cells became evident with the increased (>4 mg/L Pb²⁺) metal concentration. The photosynthetic apparatus (thylakoid membranes) were found to be the worst affected. Deteriorated or completely destroyed thylakoid membranes have made large empty spaces in the cell interior. In addition, at the highest concentration (8 mg/L Pb²⁺), the polyphosphate granules became more prominent both in size and number. Despite the initial slight stimulations (0.2, 3.8 and 6.5% respectively at 0.5, 1 and 2 mg/L Pb²⁺), both metals inhibited the growth in a dose-dependent manner as incubation progressed. Pigment contents (chlorophyll α, β carotene and phycocyanin) were also decreased with increasing metal concentration. Cells exposed to 6 mg/L Pb²⁺, resulted in 36.56, 37.39 and 29.34% reductions of chlorophyll α, β carotene and phycocyanin respectively over the control. Corresponding reductions for the same Cd²⁺concentrations were 57.83, 48.94 and 56.90%. Lethal concentration (96 h LC₅₀) values (3.47 mg/L Cd²⁺ and 12.11 mg/L Pb²⁺) indicated that Synechocystis sp. PCC 6803 is more vulnerable to Cd²⁺ than Pb²⁺. Supported by the Chinese Scholarship Council     

Bioaccumulation of Pb<Superscript>2+</Superscript> and its effects on growth,morphology and pigment contents of <Emphasis Type="Italic">Spirulina</Emphasis> (<Emphasis Type="Italic">Arthrospira</Emphasis>) <Emphasis Type="Italic">platensis</Emphasis>

A laboratory experiment was conducted to assess the bioaccumulation of Pb²⁺ and its effects on growth, morphology and pigment contents of Spirulina (Arthrospira) platensis. The specimen cultured in Zarrouk liquid medium was treated with various initial metal concentrations (0, 5, 10, 30, 50 and 100 μg mL⁻¹). The growth of S. platensis was adversely affected by Pb²⁺ at high concentrations (30, 50 and 100 μg mL⁻¹). However, at low concentrations (5 μg mL⁻¹), Pb²⁺ could stimulate its growth slightly. The pigment contents (chlorophyll α and β carotene) were decreased in a dose-dependent manner. The highest reductions (67% and 53% respectively in chlorophyll α and β carotene) were observed in 100 μg mL⁻¹ treatment group. The LC₅₀ (96 h) of Pb²⁺ was measured as 75.34 μg mL⁻¹. Apart from a few cases of filament breakages at elevated concentrations (50 and 100 μg mL⁻¹), morphological abnormalities are not specific. Metal bioaccumulation increased with Pb²⁺ concentrations, but decreased with exposure time. The maximum accumulated amount was 188 mg g⁻¹ dry weight. The bioconcentration factor (BCF) reached to a peak at day 2, followed by a gradual reduction for all the exposure concentrations. S. platensis is able to tolerate considerably high Pb²⁺ concentrations. Consequently it can be used as a potential species to remove heavy metal from contaminated waters.     

The effects of nitrogen,phosphorus, vitamins and trace metals on the growth of the red tide organismProrocentrum Micans

Laboratory culture experiments showed that <100μ mol/L nitrate, amonium or mixture of amino acids promote the growth of the red tide organismProrocentrum micans Ehrenb, but that >100μmol/L of ammonium, or mixture of glycine and glutamate was harmful to growth, and that orthophosphate wasP. micans’ main phosphorous source in the ocean. Presence of 80μ mol/L EDTA, 0.5 to 1 μmol/L Fe³⁺, 1.0 to 20.0 μ mol/L Mn²⁺ 0.1 to 0.4 μmol/L Co²⁺ in the culture medium could improve the growth ofP. micans. Vitamin B₁ promoted growth, but vitamin B₁₂ and biotin did not. The estimated minimum cell quotas (q _o) for nitrogen and phosphorus being 0.74 pmole/cell and 0.045 pmole/cell show that phosphorus (more than nitrogen) limits the growth ofP. micans in the study area. This project was supported by the Natural Science Foundation of Zhejiang Province.     

Effects of iron electrovalence and species on growth and astaxanthin production of Haematococcus pluvialis

To increase the cell concentration and the accumulation of astaxanthin in the cultivation of Haematococcus pluvialis, effects of different iron electrovalencies (Fe²⁺-EDTA and Fe³⁺-EDTA) and species (Fe-EDTA, Fe(OH)_x^32x and FeC₆H₅O₇) addition on cell growth and accumulation of astaxanthin were studied. Results show that different iron electrovalencies have various effects on cell growth and astaxanthin accumulation of H. pluvialis. Compared with Fe³⁺-EDTA, Fe²⁺-EDTA stimulate more effectively the formation of astaxanthin. The maximum astaxanthin content (30.70 mg/g biomass cell) was obtained under conditions of 18 μmol/L Fe²⁺-EDTA, despite the lower cell density (2.3×10⁵ cell/ml) in such condition. Fe³⁺-EDTA is more effective than Fe²⁺-EDTA in improving the cell growth. Especially, the maximal steady-state cell density, 2.9×10⁵ cell/ml was obtained at 18 μmol/L Fe³⁺-EDTA addition. On the other hand, all the various species of iron (EDTA-Fe, Fe(OH)_x^32x, FeC₆H₅O₇) are capable to improve the growth of the algae and astaxanthin production. Among the three iron species, FeC₆H₅O₇ performed the best. Under the condition of a higher concentration (36 μmol/L) of FeC₆H₅O₇, the cell density and astaxanthin production is 2 and 7 times higher than those of iron-limited group, respectively. The present study demonstrates that the effects of the stimulation with different iron species increased in the order of FeC₆H₅O₇, Fe(OH)_x^32x and EDTA-Fe. Supported by Xiamen Scientific and Technologic Projects (No. 3052Z20031086), Xiamen University Alumni Association Foundation in Singapore, and the First National College Students Innovative Experimental Project     

Isolation of protoplasts from <Emphasis Type="Italic">undaria pinnatifida</Emphasis> by alginate lyase digestion

The aim of this study is to isolate protoplasts from Undaria pinnatifida. Protoplasts of the alga were isolated enzymatically by using alginate lyase, which was prepared by fermenting culture of a strain Vibrio sp. 510. Monofacterial method was applied for optimizing digestion condition. The optimum condition for protoplast preparation is enzymatic digestion at 28°C for 2h using alginate lyase at the concentration of 213.36 U (8 mL) every 0.5g fresh thalline with NaCl 50 and at the shaking speed of 150 r min⁻¹ during digestion. The protoplast yield can reach 2.62±0.09 million per 0.5 g fresh leave under the optimum condition. The enzyme activity is inhibited by Ca²⁺ and slightly enhanced by Fe²⁺ and Mn²⁺ at concentrations of 0.05, 0.08 and 0.10 mol L⁻¹.     

Tolerance of ciliated protozoan Paramecium bursaria （Protozoa,Ciliophora） to ammonia and nitrites

The tolerance to ammonia and nitrites in freshwater ciliate Paramecium bursaria was measured in a conventional open system. The ciliate was exposed to different concentrations of ammonia and nitrites for 2h and 12h in order to determine the lethal concentrations. Linear regression analysis revealed that the 2h-LC50 value for ammonia was 95.94 mg/L and for nitrite 27.35 mg/L using probit scale method (with 95% confidence intervals). There was a linear correlation between the mortality probit scale and logarithmic concentration of ammonia which fit by a regression equation y=7.32x-9.51 (R^2=0.98; y, mortality probit scale; x, logarithmic concentration of ammonia), by which 2 h-LC50 value for ammonia was found to be 95.50 mg/L. A linear correlation between mortality probit scales and logarithmic concentration of nitrite is also followed the regression equation y=2.86x 0.89 (R^2=0.95; y, mortality probit scale; x, logarithmic concentration of nitrite). The regression analysis of toxicity curves showed that the linear correlation between exposed time of ammonia-N LC50 valueand ammonia-N LC50 value followed the regression equation y=2 862.85e-0.08x (R^2=0.95; y, duration of exposure to LC50 value; x, LC50 value), and that between exposed time of nitrite-N LC50 value and nitrite-N LC50 value followed the regression equation y = 127.15e-0.13x (R^2=0.91; y, exposed time of LC50 value; x, LC50 value). The results demonstrate that the tolerance to ammonia in P. bursaria is considerably higher than that of the larvae or juveniles of some metozoa, e.g. cultured prawns and oysters. In addition, ciliates, as bacterial predators, are likely to play a positive role in maintaining and improving water quality in aquatic environments with high-level ammonium, such as sewage treatment systems.     

Effects of Mg2+, NaCl, citric acid, and other factors on synthesis and accumulation of β-carotene inDunaliella salina

The effect of Mg²⁺, NaCl and citric acid on the accumulation of β-carotene inDunaliella Salina was studied. The experimental results showed that 10.5 mmol/L Mg²⁺, 5 mol/L NaCl, 3 μmol/L citric acid, and CO₂ are favorable forDunaliella Salina cell growth and β-carotene accumulation. After 144 h culture under the above conditions, theDunaliella Salina biomass increased by 7.18 times; β-carotene reached 9.61%. Project 89023990 supported by the Shandong Natural Science Fund.     

Impacts of CO<Subscript>2</Subscript> enrichment on growth and photosynthesis in freshwater and marine diatoms

The physiological responses of Nitzschia palea Kvtzing, a freshwater diatom, to elevated CO2 were investigated and compared with those of a marine diatom, Chaetoceros muelleri Lemmermann previously reported. Elevated CO2 concentration to 700 μl/L increased the dissolved inorganic carbon （D!C） and lowered the pH in the cultures of N. palea, thus enhancing the growth by 4%-20% during the whole growth period. High CO2-grown N. palea cells showed lower levels of dark respiration rates and higher Ik values. Light-saturated photosynthetic rates and photosynthetic efficiencies decreased in N. palea with the doubling CO2 concentration in airflow to the bottom of cultures, although the doubling CO2 concentration in airflow to the surface cultures had few effects on these two photosynthetic parameters. N. palea cells were found to be capable of using HCO5 in addition to gaseous CO2, and the CO2 enrichment decreased their affinity for HCO5 and CO2. Although doubled CO2 level would enhance the biomass of N. patea and C. muelleri to different extents, compared with the marine diatom, it had a significant effect on the specific growth rates of N. palea. In addition, the responses of photosynthetic parameters of IV. palea to doubled CO2 concentration were almost opposite to those of C. muelleri.     

Effects of Na+/K+ ratio of groundwaters on the gill ion-transport enzyme activity, plasma osmolality and growth of Cynoglossus semilaevis juveniles

The effects of environmental Na⁺/K⁺ ratio on the gill ion-transport enzyme activity, plasma osmolality and growth of Cynoglossus semilaevis juveniles were investigated. The results showed that, plasma osmolality was similar among flounder adapted to different Na⁺/K⁺ ratios of saline groundwaters (P>0.05), while the growth, gill Na⁺, K⁺-ATPase and HCO₃ ⁻-ATPase activities were affected by Na⁺/K⁺ ratio significantly (P<0.05). The gill Na⁺, K⁺-ATPase activity reached its maximum on day 3, then decreased gradually from day 3 to day 9 and remained constant from day 9 to day 15. The peak values of gill Na⁺, K⁺-ATPase activity were detected on day 3 for all Na⁺/K⁺ ratios of saline groundwaters, then the enzyme activities descended, and on day 9 the enzyme activities achieved steady state, and the gill HCO₃ ⁻-ATPase activity increased rapidly and achieved steady state after one day. During steady state, the gill Na⁺, K⁺-ATPase and HCO₃ ⁻-ATPase activity of Na⁺/K⁺ ratios 20 and 40 treatments were significantly higher than those in the control group (Na⁺/K⁺ ratio 27.5), while there were no significant differences between the Na⁺/K⁺ ratio 30 treatment and the control group; the gill Na⁺, K⁺-ATPase activity of Na⁺/K⁺ ratio 20 and 40 treatments were significantly higher than that for ratio 30 treatment, but there were no significant differences of gill HCO₃ ⁻-ATPase activity among these treatments. At the end of the 15-day experiment, the weight gain (%) and specific growth rate (SGR) of flounders maintained in seawater were significantly higher than those in groundwaters; significant differences also occurred among the treatments; Na⁺/K⁺ ratio 30 treatment had the highest values (33.7% and 1.94 respectively), which were significantly higher than those under Na⁺/K⁺ ratios 20 and 40 treatments. Therefore, for the saline groundwater used in this experiment, it is suggested that the Na⁺/K⁺ ratio be adjusted to approximately 30, i.e., as close to that of natural seawater as possible in the culture of flounder.