Growth dynamics and the proximate biochemical composition and fatty acid profile of the heterotrophically grown diatom <Emphasis Type="Italic">Cyclotella cryptica</Emphasis>

To investigate the nutritional value of the diatom Cyclotella cryptica as an alternative feed for aquaculture, its heterotrophic growth characteristics were studied. First, the proximate biochemical composition and fatty acid profiles were studied under a controlled heterotrophic growth condition. The approximate total ash, carbohydrate, lipid, and protein content were 245 mg g⁻¹ (dry weight), 360 mg g⁻¹, 165 mg g⁻¹ and 260 mg g⁻¹, respectively. Polyunsaturated fatty acids accounted for 24.5, 31.3, 45.1 and 17.3% of the total lipids in the phospholipid, sterol, free fatty acid and triglyceride classes. Secondly, the effect of aeration and agitation rates on the specific growth rate of C. cryptica under heterotrophic conditions was studied. The maximum specific growth rate was not significantly affected (P > 0.05) by the rate of agitation within the range of 100 to 160 rpm, but it was significantly affected (P > 0.05) by the rate of aeration. Optimal growth occurred when the aeration rate was within the range of 0.44 to 1.07 v/v/min. Viability measurements throughout the growth period showed that the C. cryptica cells remained viable in spite of the varied cultivation conditions. Hydrodynamic forces are an important parameter within biological systems, and optimisation is crucial for the successful scale-up of microalgal cultivation systems. Whilst the investigation was preliminary in nature, the information gained in this study will be useful for the continual development of an alternative and cost-effective feed for bivalve spat rations.     

The Effects of Changes in the Inorganic Nitrogen Supply on the Growth and Development of Marsilea in Aseptic Culture

The effects of changes in the inorganic nitrogen source of themedium on the growth and development of Marsilea in asepticculture are described. A change in nitrate concentration to0·2 or 5 x that of the normal medium had initially noeffect on growth or heteroblastic development, but in the 0·2nitrogen medium there was finally a reversion to juvenile stagesfollowed by early cessation of growth. Growth and heteroblastic development were stimulated by thesubstitution of the nitrate of the medium by ammonium saltsor urea. Nitrite was little inferior to nitrate as a nitrogensource, but hydroxylamine was toxic even at low concentrations.Addition of sodium chloride led to improved growth and developmentin nitrate media, an effect referred to beneficial effects ofthe chloride ion. Symptoms of toxicity and eventual death of the cultures whichoccurred after prolonged growth on ammonium media were due toan acid drift of the media rather than to a specific effectof the ammonium ion.     

EFFICIENCY OF GROWTH AND NUTRIENT UPTAKE FROM WASTEWATER BY HETEROTROPHIC,AUTOTROPHIC, AND MIXOTROPHIC CULTIVATION OF CHLORELLA VULGARIS IMMOBILIZED WITH AZOSPIRILLUM BRASILENSE1

Heterotrophic growth of microalgae presents significant economic advantages over the more common autotrophic cultivation. The efficiency of growth and nitrogen, phosphorus, and glucose uptake from synthetic wastewater was compared under heterotrophic, autotrophic, and mixotrophic regimes of Chlorella vulgaris Beij. immobilized in alginate beads, either alone or with the bacterium Azospirillum brasilense. Heterotrophic cultivation of C. vulgaris growing alone was superior to autotrophic cultivation. The added bacteria enhanced growth only under autotrophic and mixotrophic cultivations. Uptake of ammonium by the culture, yield of cells per ammonium unit, and total volumetric productivity of the culture were the highest under heterotrophic conditions when the microalga grew without the bacterium. Uptake of phosphate was higher under autotrophic conditions and similar under the other two regimes. Positive influence of the addition of A. brasilense was found only when light was supplied (autotrophic and mixotrophic), where affinity to phosphate and yield per phosphate unit were the highest under heterotrophic conditions. The pH of the culture was significantly reduced in all regimes where glucose was consumed, similarly in heterotrophic and mixotrophic cultures. It was concluded that the heterotrophic regime, using glucose, is superior to autotrophic and mixotrophic regimes for the uptake of ammonium and phosphate. Addition of A. brasilense positively affects the nutrient uptake only in the two regimes supplied with light.     

Prediction of proton exchange and bacterial growth on various substrates using constraint-based modeling approach

Proton exchange between cells and medium is one of the most important factors affecting culture pH, and hence its prediction is advantageous. In this research, proton exchange flux across the cell membrane was predicted using a genome-scale model. Calculated proton exchange flux was then exploited as a criterion to predict the trends and intensities of pH changes in cultures of Bacillus subtilis containing various C-sources, i.e. glucose, sucrose, glycerol, lactate, and citrate, as well as N-sources, i.e. ammonium chloride, sodium nitrate, urea, and histidine. The results showed that glucose, sucrose, and glycerol lowered culture pH as compared to citrate and lactate, which raised it. With regard to N-sources, the model predicted that ammonium chloride lowered culture pH while other sources raised pH. Furthermore, maximum theoretical biomass yield using the various C&N-sources was calculated, and sensitivity of microbial growth to proton exchange flux was investigated using robustness analysis to identify the effect of pH on growth of B. subtilis using different substrates. Finally, the effect of ammonium nitrate, a widely used nitrogen source, on growth of B. subtilis was studied. Experimental data obtained by cultivation of B. subtilis DSM 3256 on mineral salt media containing various C&N-sources were used to confirm model predictions. Model predictions were in good agreement with the experimental results for all of the examined C-sources as well as ammonium chloride and sodium nitrate as N-sources. However, the predictions for the N-sources urea and histidine showed deviations, possibly because these two compounds serve as both C&N-sources.     

Growth of and omega-3 fatty acid production by Phaeodactylum tricornutum under different culture conditions

Detailed studies were carried out on the effects of nitrogen source, phosphate, sodium chloride, growth factors, precursors, CO2, temperature, initial pH, and inoculum size on biomass and eicosapentaenoic acid (EPA) production by Phaeodactylum tricornutum. The EPA content of total fatty acids increased with increasing concentrations of nitrate and urea. Sodium chloride was not required for growth or EPA production. While vitamins B1 and B12 did not affect growth significantly, EPA yield was increased by 65% by B12 supplementation. Maximum EPA production occurred when the air gassing supply was supplemented with 1% CO2. Optimum culture temperature and initial pH for EPA production were 21.5 to 23 degrees C and 7.6, respectively. EPA yields of up to 133 mg/liter of culture were observed. EPA constituted up to 30 to 40% of total fatty acids.     

Nitrogen and sulphur requirements of Colletotrichum inamdarii Lal

Summary Nitrogen and sulphur requirements ofColletotrichum inamdarii Lal isolated from the leaves ofCarissa carandas L. have been studied. DL-serine, L-asparagine and L-phenylalanine have been found to be of good nitrogen source followed by potassium nitrate, calcium nitrate, magnesium nitrate, DL-alanine, ammonium nitrate, glutamic acid, ammonium sulphate, DL-valine, aspartic acid, ammonium chloride, ammonium hydrogencarbonate, L-histidine and potassium nitrite. There was no growth in the absence of nitrogen.Sporulation was excellent on calcium nitrate and sodium nitrate, Very good on DL-serine, potassium nitrate, and magnesium nitrate. Good on L-asparagine, L-phenylalanine and ammonium oxalate. Fair on DL-alanine, DL-leucine, ammonium sulphate, DL-valine, ammonium chloride and L-histidine whereas poor on glutamic acid, aspartic acid, ammonium tartarate and ammonium nitrate. Few spores were observed on ammonium hydrogencarbonate but potassium nitrite did not show any sporulation.Amongst the sulphur compounds sodium bisulphate gave the best growth and good sporulation, followed by sodium thiosulphate, magnesium sulphate, ammonium sulphate and potassium sulphate. Thiourea gave negligible growth whereas it failed to grow on zinc sulphate and potassium persulphate.     

Growth of and omega-3 fatty acid production by Phaeodactylum tricornutum under different culture conditions.

Detailed studies were carried out on the effects of nitrogen source, phosphate, sodium chloride, growth factors, precursors, CO2, temperature, initial pH, and inoculum size on biomass and eicosapentaenoic acid (EPA) production by Phaeodactylum tricornutum. The EPA content of total fatty acids increased with increasing concentrations of nitrate and urea. Sodium chloride was not required for growth or EPA production. While vitamins B1 and B12 did not affect growth significantly, EPA yield was increased by 65% by B12 supplementation. Maximum EPA production occurred when the air gassing supply was supplemented with 1% CO2. Optimum culture temperature and initial pH for EPA production were 21.5 to 23 degrees C and 7.6, respectively. EPA yields of up to 133 mg/liter of culture were observed. EPA constituted up to 30 to 40% of total fatty acids.     

Effects of nitrogen source and concentration on growth rate and fatty acid composition of Ellipsoidion sp. (Eustigmatophyta)

In order to study the effects of different nitrogen source and concentrationon the growth rate and fatty acid composition, a marine microalga Ellipsoidion sp. with a high content of eicosapentaenoic acid (EPA) wascultured in media with different nitrogen sources and concentrations.During the pre-logarithmic phase, the alga grew faster with ammoniumas N source than with nitrate, but the reverse applied during thepost-logarithmic phase. The alga grew poorly in N-free mediumor medium with urea as the sole N source. In the same growth phase,ammonium medium resulted in higher yield of total lipid, but the EPA yielddid not differ significantly different from that using nitrate medium. Themaximum growth rate occurred in medium containing 1.28 mmolL^-1 sodium nitrate, while maximum EPA and total lipid contents werereached at 1.92 mmol L^-1, when EPA accounted for 27.9% totalfatty acids. The growth rate kept stable when NH₄Cl ranged from0.64 to 2.56 mmol L^-1, and the maximum content of total lipidand EPA occurred in the medium with 2.56 mmol L^-1NH₄Cl. The EPA content was higher in the pre- thanpost-logarithmic phase, though the total lipid content was lower. Thehighest EPA content expressed as percent total fatty acid was 27.9% innitrate medium and and 39.0% in ammonium medium.     

Urea as sole source of nitrogen for plant growth

Summary Spirodela oligorrhiza grown in sterile culture was able to use urea as sole source of nitrogen but only when the pH of the culture medium was below 4.3. Plants inoculated into urea media at pH 6.4 initially made little growth and became nitrogen-deficient in appearance and composition although they contained about 100 grams of urea per gram fresh weight of tissue. After a period the pH of the medium usually fell below 4.3 and growth commenced. Growth with other compounds, e.g. ammonium, nitrate or allantoin, as sources of nitrogen was not similarly affected by the pH of the culture medium.Urease activity could always be detected in the tissues of Spirodela oligorrhiza growing on urea. Plants with little or no urease activity soon developed significant activity when inoculated into urea media at pH 4.0. When the pH of the medium was higher there was no increase in urease activity and no growth ensued. Plants growing on urea possessed an activity of about 50 milliunits per gram fresh weight of tissue, but if the pH of the medium fell to 3.5 or lower, the activity present rose to 10 times this level.Urease activity also appeared, in the absence of supplied urea, as plants became increasingly nitrogen-deficient.     

Growth and amino acid contents of<Emphasis Type="Italic">Spirulina platensis</Emphasis> with different nitrogen sources

The growth and amino acid contents of the cyanobacterium,Spirulina platensis strain NIES 46, were investigated using ammonium, nitrate, nitrite, or urea as the sole nitrogen source in a batch culture. Chlorophylla concentration was highest at 2,096 μg/L in the nitrate group after 10 days of cultivation, while the dry weight ofS. platensis was highest at 4.5 g/L in the ammonium group after 30 days of cultivation. The total amino acid content was highest at 174 mg/g dry weight ofS. platensis in the urea group at the end of the cultivation period, yet the amino acid patterns forS. platensis were similar for all the experimental groups. Therefore, it seemed that the growth and amino acid composition ofS. platensis varied depending on the type of nitrogen sources, while the amino acid patterns were not changed. Also, the most efficient harvesting time forS. platensis seemed to be approximately 10 days after cultivation.     

Effect of inorganic N on the population,in vitro colonization and morphology of Acetobacter diazotrophicus (syn. Gluconacetobacter diazotrophicus)

We investigated whether Acetobacter diazotrophicus (syn.Gluconacetobacter diazotrophicus) could be recovered only from sugarcane plants either with low or no application of fertiliser N. We report here the enrichment and enumeration of A. diazotrophicus from high N-fertilised samples where high heterotrophic populations reduce the numbers of A. diazotrophicus ultimately diminshing its isolation frequency as reported earlier. The growth medium of micropropagated sugarcane seedlings of the varieties Co 8021, Co 86249, Co 86010, Co 86032, and Co 87025 was amended with potassium nitrate, ammonium nitrate, ammonium chloride and urea. The colonisation and AR activity of A. diazotrophicus were affected in the presence of high levels (25 mM) of ammonium chloride and ammonium nitrate but remained unaffected in low levels of N (i.e 1/10th of MS liquid medium) and with high levels of potassium nitrate (25 mM) and urea (500 ppm). A. diazotrophicus was detected in the inoculated plants both at low and high levels of N based on the amplification of a specific 16S rRNA gene fragment using PCR based method targeting a stretch of 445 bp with primers AC and DI. High levels of N in the growth medium induced morphological changes on A. diazotrophicus cells resulting in long pleomorphic cells. The percentage of pleomorphic cells was in the decending order from NH₄NO₃, NH₄Cl, KNO₃, and urea. These changes were more prominent in ammonium chloride and ammonium nitrate than potassium nitrate, urea and N free medium. The morphological changes and the increased heterotrophic populations may play a role on the survival ofA. diazotrophicus in high N-fertilised samples/environments.     

Development of High‐Level Omega‐3 Eicosapentaenoic Acid (EPA) Production from Phaeodactylum tricornutum

Phaeodactylum tricornutum is a lipid‐rich marine diatom that contains a high level of omega‐3 polyunsaturated fatty acids, especially eicosapentaenoic acid (EPA). In an effort to reduce costs for large‐scale cultivation of this microalga, this study first established a New BBM medium (0.3 x strength BBM with only 3% of the initial phosphate level) to replace the traditional F/2 medium. Phaeodactylum tricornutum could grow in extremely low phosphate concentrations (25 µM), without compromising the EPA content. In the presence of sea salts, silicate addition was not necessary for high rate growth, high EPA content, or lipid accumulation in this species. Using urea as the sole nitrogen source tended to increase EPA contents per dry biomass (by 24.7%) while not affecting growth performance. The use of sea salts, rather than just sodium chloride, led to significantly improved biomass yields (20% increase) and EPA contents of total fatty acid (46–52% increase), most likely because it supplied sufficient essential elements such as magnesium. A salinity level of 35 led to significantly higher biomass yields compared with 20, but salinity had no significant influence on EPA content. EPA became the dominant fatty acid with average levels of 51.8% of total fatty acids during the exponential growth phase at 20 ppt in New BBM medium with sea salts.     

Importance of seed Zn content for wheat growth on Zn-deficient soil

Eggplants (Solanum melongena L. cv. Bonica) were grown in a glasshouse during summer under natural light with one unbranched shoot or one shoot with 3 to 4 branches and with or without fruit in quartz sand buffered and not buffered with 0.5% CaCO₃ (w : v), respectively. Nutrient solutions supplied contained nitrate or ammonium as the sole nitrogen source. Compared with nutrient solutions containing nitrate (10 mM), solutions containing ammonium (10 mM) caused a decrease in net photosynthesis of eggplants during early stages of vegetative growth when grown in quartz sand not buffered with CaCO₃. The decrease was not observed before leaves showed interveinal chlorosis. In contrast, net photosynthesis after bloom at first increased more rapidly in eggplants supplied with ammonium than with nitrate nitrogen. However, even in this case, net photosynthesis decreased four weeks later when ammonium nutrition was continued. The decrease was accompanied by epinasty and interveinal chlorosis on the lower leaves and later by severe wilting, leaf drop, stem lesions, and hampered growth of stems, roots, and fruits. These symptoms appeared later on plants not bearing fruits than on plants bearing fruits. If nutrient solutions containing increasing concentrations of ammonium (0.5–30 mM) were supplied after the time of first fruit ripening, shoot growth and set of later flowers and fruits were promoted. In contrast, vegetative growth and reproduction was only slightly affected by increasing the concentration of nitrate in the nutrient solutions. In quartz sand buffered with CaCO₃ ammonium nutrition caused deleterious effects only under low light conditions (shade) and on young plants during rapid fruit growth. If eggplants were supplied with ammonium nitrogen before bloom, vegetative growth was promoted, and set of flowers and fruit occurred earlier than on plants supplied with nitrate. Furthermore, the number of flowers and fruit yield increased. These effects of ammonium nutrition were more pronounced when plants were grown with branched shoots than with unbranched shoots. The results indicate that vegetative and reproductive growth of eggplants may be manipulated without causing injury to the plants by supplying ammonium nitrogen as long as the age of the plants, carbohydrate reserves of the roots, quantity of ammonium nitrogen supplied, and pH of the growth medium are favourable. T W Rufty Section editor     

真盐生植物盐角草对不同氮形态的响应

盐角草(Salicornia europaea L.)是一种喜盐植物,其最佳生长的实现需要200—400 mmol/L NaCl。为了解盐渍环境下盐角草氮素吸收利用特点,在水培添加200或400 mmol/L NaCl情况下,从生长指标,光合参数,根系体积和活力,硝酸还原酶与谷氨酰胺合成酶活力,蛋白、总氮、硝态氮及铵态氮含量等方面检测了硝态氮、铵态氮和尿素3种氮形态对盐角草生长的影响。研究发现以氮摩尔浓度(mmol/L)计,在0.1—400 mmol/L浓度范围进行测试,盐角草在0.1 mmol/L低氮条件下仍能维持生长,同时,抑制盐角草生长的氮浓度域值较高,其中铵态氮、尿素和硝态氮分别为50、50及400 mmol/L。研究结果表明盐角草吸收利用氮素的能力强,对氮素的浓度耐受范围宽,3种氮形态都可作为氮源满足其生长需要,但有效促进生长的效果存在差异,总体顺序从高到低依次为硝态氮、铵态氮和尿素。研究为揭示盐生植物氮吸收利用特点提供了基础数据,对提高盐生植物生产力,指导沿海滩涂生态建设具有一定的指导意义。     

Inorganic nitrogen source for autotrophic growth of Euglena mutabilis Schmitz

The effect of inorganic nitrogen source on population growth of Euglena mutabilis, an acidophillic benthic protozoa colonizing on the sediment of acid mine drainage, was investigated. Sodium nitrate, ammonium chloride, and ammonium sulfate were tested as nitrogen sources. The population density of E. mutabilis at equilibrium density cultivated in ammonium chloride‐ and ammonium sulfate‐containing media was 9–11 times higher than that in sodium nitrate‐containing medium at the optimal salt medium concentration. The population growth of E. mutabilis in ammonium sulfate‐containing medium was rapid and reached half of the equilibrium density after ca. 228 h, which was ca. 77 h earlier than that in ammonium chloride‐containing medium. Culture medium with ammonium sulfate as the nitrogen source achieved the highest maximum population density and the fastest growth rate among the three nitrogen salts used as nitrogen sources.     

冬小麦等4种作物对铵,硝态氮的吸收能力

采用水培试验探讨了冬小麦、大豆、油菜和莴笋４种作物对硝、铵态氮的相对吸收能力以及这两种氮源对它们生长发育的影响。试验表明：（１）不同氮源对供试作物的生长发育影响极大。供给硝态氮,这些作物生长发育良好,供给等量的ＮＯ＾－３和ＮＨ＾－４（１：１）时,蔬菜作物莴笋生长量下降幅度最大;供给铵态氨,莴笋和大豆极为敏感,供给ＮＯ＾－３时莴笋吸氮量显著高于供给等氮量ＮＯ＾－３和ＮＨ＾＋４,莴上麦供给等量ＮＯ＾－     

不同氮源及氮浓度对真眼点藻纲微藻生长及油脂积累的影响

以真眼点藻纲8株微藻(类波氏真眼点藻(Eustigmatos cf. polyphem)、大真眼点藻(Eustigmatos magnus)、波氏真眼点藻(Eustigmatos polyphem)、魏氏真眼点藻(Eustigmatos vischeri)、斧形魏氏藻(Vischeria helvetica)、点状魏氏藻(Vischeria punctata)、星形魏氏藻(Vischeria stellata)和眼点拟微绿球藻(Nan-nochloropsis oculata))为研究材料, 用3种氮源(硝酸钠、碳酸氢铵或尿素)和4种氮浓度(18、9、6和3 mmol) 在改良的BG-11培养基中对藻细胞进行培养。比较分析这8株微藻在不同培养条件下的藻液pH、生物量、油脂含量、脂肪酸组成的差异, 从而筛选出适合该类微藻生长和油脂积累的最适氮源与最佳氮浓度。结果表明, 这8株微藻均能在3种氮源中生长, 但是随着培养时间延长, 以碳酸氢铵和尿素为氮源时藻液pH逐渐降低, 其变化范围为5.0—6.0, 而以硝酸钠为氮源时藻液pH保持在7.0—8.0, 变化不大。当以尿素为氮源培养时, 能获得较高的生物量, 但是不同藻株在不同尿素浓度时达到最高生物量。最高生物量是波氏真眼点藻(E. polyphem)在9 mmol时达到, 为10.96 g/L。总脂含量分析发现, 在低氮浓度下均能促进8株微藻油脂的积累, 真眼点藻属中的魏氏真眼点藻(E. vischeri)在8株藻中获得最高油脂含量, 达到59.24%。进一步对脂肪酸分析发现, 8株微藻总脂肪酸含量为细胞干重的50%—58%, 主要脂肪酸组成为豆蔻酸(C14鲶0)、棕榈酸(C16鲶0)、棕榈油酸(C16鲶1)、油酸(C18鲶1)和二十碳五烯酸(C20鲶5), 其中拟微绿球藻(N. oculata)细胞中棕榈酸的含量最高占总脂肪酸50%左右; 其他7株微藻细胞中棕榈油酸的含量较高, 其占总脂肪酸含量范围在40%—60%。8株微藻均表现出较高的生物量与油脂积累能力, 以尿素为氮源, 氮浓度为6 mmol时更有利于该类微藻生物量和油脂的积累。总体来说, 真眼点藻纲的微藻是一类极具潜力适合于微藻生物燃料生产的微藻, 而真眼点藻属藻株表现更为明显的优势。     

Different nitrogen sources and growth responses of Spirulina platensis in microenvironments

Spirulina platensis was cultivated, in comparative studies, using several sources of nitrogen. The standard source used (sodium nitrate) was the same as that used in the synthetic medium Zarrouk, whereas the alternative nitrogen sources consisted of ammonium nitrate, urea, ammonium chloride, ammonium sulphate or acid ammonium phosphate. The initial nitrogen concentrations tested were 0.01, 0.03 and 0.05 M in an aerated photobioreactor at 30 °C, with an illuminance of 1900 lux, and 12 h-light/12 h-dark photoperiod over a period of 672 h. Maximum biomass was produced in medium containing sodium nitrate (0.01–0.03–0.05 M), followed by ammonium nitrate (0.01 M) and urea (0.01 M). The final biomass concentrations were 1.992 g l^–1 (0.03 M sodium nitrate), 1.628 g l^–1 (0.05 M sodium nitrate), 1.559 g l^–1 (0.01 M sodium nitrate), 0.993 g l^–1 (0.01 M ammonium nitrate) and 0.910 g l^–1 (0.01 M urea). This suggested that it is possible to utilize nitrogen sources other than sodium nitrate for growing S. platensis, in order to decrease the production costs of scaled up projects.     

Effects of fertilizer-based culture media on the production of exocellular polysaccharides and cellular superoxide dismutase by <Emphasis Type="Italic">Phaeodactylum tricornutum</Emphasis> (Bohlin)

Microalgae cultures are receiving attention because of increasing biotechnological and biomedical production of active biomolecules. We evaluated various fertilizer-based culture media to scale up production of the marine microalga Phaeodactylum tricornutum for production of exocellular polysaccharides (EPS), soluble proteins, and cellular superoxide dismutase (SOD). The standard source of sodium nitrate was the same as that used in the synthetic f/2 culture medium and ammonium nitrate, urea, ammonium sulfate, and calcium nitrate as alternative sources of nitrogen. The maximum production of EPS was achieved in microalgae cells grown in the culture media containing 63 and 23% nitrogen from ammonium sulfate, and also in microalgae cells grown in the culture media containing 3% nitrogen from ammonium nitrate. The maximum production of cellular SOD was achieved in microalgae cells grown in the culture media containing 35 and 26% nitrogen from ammonium sulfate, and in the culture media containing 17% nitrogen from urea. The results suggest that it is possible to use a source of nitrogen, other than sodium nitrate, to scale up growth of P. tricornutum for production of EPS and SOD at reduced costs.