The augmented lipid productivity in an emerging oleaginous model alga <Emphasis Type="Italic">Coccomyxa subellipsoidea</Emphasis> by nitrogen manipulation strategy

The lipid productivity controlled by both of biomass and lipid content was really crucial for economic-feasibility of microalgae-based biofuels production. This study attempted at augmenting lipid productivity in an emerging oleaginous model alga Coccomyxa subellipsoidea by different nitrogen manipulation including one-stage continuous N-sufficiency (OCNS), N-deprivation (OCND), N-limitation (OCNL), and also two-stage batch N-starvation (TBNS). Amongst four tested nitrogen manipulation strategies, OCNS performed remarkable promoting effect on cell metabolic growth and the maximum biomass was achieved by 7.39 g/L. Whereas TBNS regime induced the highest lipid content (over 50.5%). Only OCNL treatment augmented the lipid productivity by 232.37 mg/L/day, representing 1.25-fold more than TBNS and even as much as 5.06-fold more than that of OCND strategy. OCNL also strengthened the proportions of saturated (C16:0 and C18:0) and monounsaturated fatty acid (C18:1) which were inclined to high-quality biofuels-making. This might be due to that most part of energy and metabolic flux (e.g. acetyl-CoA) derived from TCA cycle and glycolysis flowed into fatty acids biosynthesis pathway (especially C18:1) response to OCNL manipulation. This study represented a pioneering work of utilizing OCNL for lipids production by C. subellipsoidea and clearly implied that OCNL might be a feasible way for algal lipid production on a commercial scale and also promoted the potential of C. subellipsoidea as an ideal biodiesel feedstock.     

Interspecific biodiversity enhances biomass and lipid productivity of microalgae as biofuel feedstock

Large improvements in biomass and lipid production are required to make massive scale algal biodiesel production an economic reality. The application of the biodiversity strategy to enhance algal biomass as biofuel feedstock is little. The algal diversity was manipulated in this study to investigate the effects of a combination of biodiversity complementarity and a new medium consisting of seawater and agricultural fertilizer on lipid productivity. The algae diverse community includes two strains of Dunaliella salina (Dunaliella salina 19/30 and 19/18) and three species of Nannochloropsis (Nannochloropsis oculata, Nannochloropsis salina, and Nannochloropsis gaditana). The results showed that the most diverse community (5 species) produced an average of sixfold more biomass in the new medium than did the standard f/2 culture medium. The most diverse polyculture had the highest growth rate (1.01 day^?1), biomass (1.2 g L^?1), and lipid productivity (0.31 g L^?1 day^?1). The assessment of algal polycultures relative to monocultures is particularly interesting and novel for this biofuel field, and the observations that these polycultures resulted in significant lipid productivity improvements are very useful addition to the biofuel research. The possible mechanism (resource diversity) to explain the synergy in mixed cultures warrants further investigation.     

Low-cost cultivation of <Emphasis Type="Italic">Scenedesmus</Emphasis> sp. with filtered anaerobically digested piggery wastewater: biofuel production and pollutant remediation

A laboratory study was conducted on biomass and lipid production by Scenedesmus sp. and the removal of total nitrogen (TN) and total phosphorus (TP) from filtered anaerobically digested piggery wastewater. The dry weight (DW), lipid content and productivity, total nitrogen, and total phosphorus removal rate were assessed in five media: modified soil extract (MSE) medium, 5 % anaerobic digested wastewater (ADWW), 10 % ADWW, 15 % ADWW, and 5 % ADWW supplemented with NaNO₃. The highest biomass productivity appeared in the 15 % ADWW group, which was 20.4 % higher than MSE group. The highest lipid content was found in the 5 % ADWW group (31.60 %), while the highest lipid productivity was in the 10 % ADWW group (27.01 mg L^?1 day^?1). Compared with the 5 % ADWW group, the 5 % ADWW group supplemented with NaNO₃ had a similar biomass amount but lower lipid content and productivity. The fatty acids percentage of Scenedesmus sp. showed a slight difference in different media, but with the four dominant fatty acids (C16:0, C18:1, C18:2, C18:3) accounting for 87 % of the total fatty acids, suggests that Scenedesmus sp. in ADWW medium was no different than MSE in terms of lipid composition and content. TN removal rates were 82.85, 82.51, 85.85, 91.28, and 78.71 % in groups 1 to 5, and TP removal rates were 53.05, 88.53, 87.77, 88.72, and 80.64 %. Our experiment also shows the feasibility of using ADWW as a substitute of all the elements of MSE medium except for carbon, which would significantly reduce the costs of microalgal culture.     

Comparison of Trophic Modes to Maximize Biomass and Lipid Productivity of <Emphasis Type="Italic">Micractinium inermum</Emphasis> NLP-F014

An optimum trophic mode condition was investigated to maximize biomass and lipid productivity of Micractinium inermum NLP-F014, which grown successfully in blended wastewater medium. In this study, four trophic modes were used, including photoautotrophic, photoheterotrophic, heterotrophic and mixotrophic modes. Mixotrophic mode showed the highest biomass and lipid productivity. However, a high concentration of organics resulted the negative effect on the growth of M. inermum NLP-F014. Mixotrophic cultivation using glucose below 500 mg/L was able to produce maximum biomass productivity up to 0.90 ± 0.03 g/L/day as well as maximum lipid productivity up to 129.31 ± 0.10 mg/L/day. From lipid analysis on mixotrophic mode using glucose, the major fatty acids are oleic acid (C18:1), linoleic acid (C18:2) and palmitic acid (C16:0). These results suggest that mixotrophic mode cultivation with wastewater containing chemical oxygen demand (COD) below 500 mg/L could be applicable for biodiesel production of M. inermum NLP-F014.     

Application of high-salinity stress for enhancing the lipid productivity of <Emphasis Type="Italic">Chlorella sorokiniana</Emphasis> HS1 in a two-phase process

Increased lipid accumulation of algal cells as a response to environmental stress factors attracted much attention of researchers to incorporate this stress response into industrial algal cultivation process with the aim of enhancing algal lipid productivity. This study applies high-salinity stress condition to a two-phase process in which microalgal cells are initially grown in freshwater medium until late exponential phase and subsequently subjected to high-salinity condition that induces excessive lipid accumulation. Our initial experiment revealed that the concentrated culture of Chlorella sorokiniana HS1 exhibited the intense fluorescence of Nile red at the NaCl concentration of 60 g/L along with 1 g/L of supplemental bicarbonate after 48 h of induction period without significantly compromising cultural integrity. These conditions were further verified with the algal culture grown for 7 days in a 1 L bottle reactor that reached late exponential phase; a 12% increment in the lipid content of harvested biomass was observed upon inducing high lipid accumulation in the concentrated algal culture at the density of 5.0 g DW/L. Although an increase in the sum of carbohydrate and lipid contents of harvested biomass indicated that the external carbon source supplemented during the induction period increased overall carbon assimilation, a decrease in carbohydrate content suggested the potential reallocation of cellular carbon that promoted lipid droplet formation under high-salinity stress. These results thus emphasize that the two-phase process can be successfully implemented to enhance algal lipid productivity by incorporating high-salinity stress conditions into the pre-concentrated sedimentation ponds of industrial algal production system.     

Improving of lipid productivity of the biodiesel promising green microalga <Emphasis Type="Italic">Chlorella pyrenoidosa</Emphasis> via low-energy ion implantation

High lipid content in microalgae is an essential parameter for adopting of microalgal biomass as a feedstock for biodiesel. Mutation is one approach to obtain desired algal strain with high lipid production. In this study, a mutant strain of Chlorella pyrenoidosa was isolated using 1.5?×?10¹⁵ ions cm^?2 s^?1 of N⁺ ion beam implantation technique, which has been widely used in mutagenesis of agricultural crops. N⁺ implantation slightly improved the growth of the mutant over the corresponding wild strain with significant increase in lipid content (32.4 % higher than the wild strain), which resulted in significant increase in lipid productivity by 35 %. In addition, ion implantation mutagenesis of C. pyrenoidosa resulted in 21.4 % decrease in total saturated fatty acids (SFAs) compared to the wild type, with a noticeable increase in polyunsaturated fatty acids (PUFAs). The increase in PUFAs was due mainly to stimulation of hexadecadienoic acid (C16:2) and octadecadienoic acid (C18:2) production. However, the SFA content of wild and mutant strains was 31.7 and 24.9 % of total fatty acids, respectively, highlighting the oxidative stability of biodiesel produced by both strains according to the European standards. Cultivation of C. pyrenoidosa mutant in selenite enrichment medium for five successive cultivation experiments showed insignificant changes in biomass productivity, lipid content, and lipid productivity alongside the study period, which confirms the genetic stability of the produced mutant. The present study confirmed the feasibility of generation of microalgae mutants with significant high lipid production using ion beam implantation.     

Fatty Acid Characterization and Biodiesel Production by the Marine Microalga <Emphasis Type="Italic">Asteromonas gracilis</Emphasis>: Statistical Optimization of Medium for Biomass and Lipid Enhancement

Lipid production is an important indicator for evaluating microalgal species for biodiesel production. In this study, a new green microalga was isolated from a salt lake in Egypt and identified as Asteromonas gracilis. The main parameters such as biomass productivity, lipid content, and lipid productivity were evaluated in A. gracilis, cultivated in nutrient-starved (nitrogen, phosphorous), and salinity stress as a one-factor-at-a-time method. These parameters in general did not vary significantly from the standard nutrient growth media when these factors were utilized separately. Hence, response surface methodology (RSM) was assessed to study the combinatorial effect of different concentrations of the abovementioned factor conditions and to maximize the biomass productivity, lipid content, and lipid productivity of A. gracilis by determining optimal concentrations. RSM optimized media, including 1.36 M NaCl, 1 g/L nitrogen, and 0.0 g/L phosphorus recorded maximum biomass productivity, lipid content, and lipid productivity (40.6 mg/L/day, 39.3%, and 15.9 mg/L/day, respectively) which agreed well with the predicted values (40.1 mg/L/day, 43.6%, and 14.6 mg/L/day, respectively). Fatty acid profile of A. gracilis was composed of C16:0, C16:1, C18:0, C18:3, C18:2, C18:1, and C20:5, and the properties of fuel were also in agreement with international standards. These results suggest that A. gracilis is a promising feedstock for biodiesel production.     

Cultivation of <Emphasis Type="Italic">Chlorella vulgaris</Emphasis> with swine wastewater and potential for algal biodiesel production

In this study, an alga-based simultaneous process of treating swine wastewater (SWW) and producing biodiesel was explored. Chlorella vulgaris (UTEX-265) was employed as a model species, and a SWW-based medium was prepared by dilution with tap water. Chlorella vulgaris grew well in the SWW-based medium, and at optimum dilution ratios, it exceeded the conventional culture medium in terms of biomass concentration and productivity. In eightfold diluted SWW, which supported the maximum growth, biomass productivity was 0.247 g L^?1 day^?1, while the productivity was merely 0.165 g L^?1 day^?1 in standard tris-acetate-phosphorous (TAP) algal medium. In addition, fatty acid methyl ester (FAME) productivity was greater in the SWW-based medium (0.067 versus 0.058 g L^?1 day^?1). This enhanced productivity resulted in more than 95 % removal of both nitrogen and phosphorous. All these show that C. vulgaris cultivation is indeed possible in a nutrient-rich wastewater with appropriate dilution, and in so doing, the wastewater can effectively be treated.     

Assessment of <Emphasis Type="Italic">Nannochloropsis gaditana</Emphasis> growth and lipid accumulation with increased inorganic carbon delivery

Algal biomass refineries for sustainable transportation fuels, in particular biodiesel, will benefit from algal strain enhancements to improve biomass and lipid productivity. Specifically, the supply of inorganic carbon to microalgal cultures represents an area of great interest due to the potential for improved growth of microalgae and the possibility for incorporation with CO₂ mitigation processes. Combinations of bicarbonate (HCO₃^?) salt addition and application of CO₂ to control pH have shown compelling increases in growth rate and lipid productivity of fresh water algae. Here, focus was placed on the marine organism, Nannochloropsis gaditana, to investigate growth and lipid accumulation under various strategies of enhanced inorganic carbon supply. Three gas application strategies were investigated: continuous sparging of atmospheric air, continuous sparging of 5% CO₂ during light hours until nitrogen depletion, and continuous sparging of atmospheric air supplemented with 5% CO₂ for pH control between 8.0 and 8.3. These gas sparging schemes were combined with addition of low concentrations (5 mM) of sodium bicarbonate at inoculation and high concentration (50 mM) of sodium bicarbonate amendments just prior to nitrogen depletion. The optimum scenario observed for growth of N. gaditana under these inorganic carbon conditions was controlling pH with 5% CO₂ on demand, which increased both growth rate and lipid accumulation. Fatty acid methyl esters were primarily comprised of C16:0 (palmitic) and C16:1 (palmitoleic) aliphatic chains. Additionally, the use of high concentration (50 mM) of bicarbonate amendments further improved lipid content (up to 48.6%) under nitrogen deplete conditions when paired with pH-controlled strategies.     

Effect of nitrate feeding strategies on lipid and biomass productivities in fed-batch cultures of Nannochloropsis gaditana

Controlled nitrate feeding strategies for fed-batch cultures of microalgae were applied for the enhancement of lipid production and microalgal growth rates. In particular, in this study, the effect of nitrate feeding rates on lipid and biomass productivities in fed-batch cultures of Nannochloropsis gaditana were investigated using three feeding modes (i.e., pulse, continuous, and staged) and under two light variations on both lipid productivity and fatty acid compositions. Higher nitrate levels negatively affected lipid production in the study. Increasing the light intensity increased the lipid contents of the microalgae in all three fed-batch feeding modes. A maximum of 58.3% lipid- to dry weight ratio was achieved when using pulse-fed cultures at an illumination of 200 μmol photons m⁻² s⁻¹ and 10 mg/day of nitrate feeding. This condition also resulted in the maximum lipid productivity of 44.6 mg L⁻¹ day⁻¹. The fatty acid compositions of the lipids consisted predominantly of long-chain fatty acids (C:16 and C:18) and accounted for 70% of the overall fatty acid methyl esters. Pulse feeding mode was found to significantly enhance the biomass and lipid production. The other two feeding modes (continuous and staged) were not ideal for lipid and biomass production. This study demonstrates the applicability of pulse feeding strategies in fed-batch cultures as an appropriate cultivation strategy that can increase both lipid accumulation and biomass production.     

Microalgal Biomass and Lipid Production on Dairy Effluent Using a Novel Microalga, <Emphasis Type="Italic">Chlorella</Emphasis> sp. Isolated from Dairy Wastewater

The present study focused on cost-effective production of microalgal biomass and lipid production on dairy effluent. The novel microalga, Chlorella sp. isolated from the dairy effluent showed high growth and lipid production on the undiluted and two-fold diluted dairy effluent which were four to five times higher than those of Chlorella vulgaris (control). The high growth of Chlorella sp. was thought to be possibly due to its heterotrophic growth capacity, high turbidity, COD, nutrients and trace elements. In contrast, C. vulgaris showed poor heterotrophic and photoautotrophic growth under the highly turbid conditions of dairy effluent. Both Chlorella sp. and C. vulgaris showed similar total FAME (mg FAME/g algal cells). The fatty acid composition analysis revealed that both Chlorella sp. and C. vulgaris possessed major C18 and C20 fatty acids which will be used for biodiesel production. Overall, the novel microalga, Chlorella sp. isolated from the dairy effluent showed high potential for cost-effective algal cultivation and lipid production on dairy effluent without any modification of process.     

Optimization of environmental parameters for <Emphasis Type="Italic">Nannochloropsis salina</Emphasis> growth and lipid content using the response surface method and invading organisms

Algae biofuel has the potential to replace fossil fuels. However, cultivation and productivity of target algae need improvement, while controlling undesired organisms that can lower the efficiency of production systems. A central composite design and response surface model were utilized to predict cultivation optima of marine microalga, Nannochloropsis salina, under a suite of environmental parameters. The effects of salinity, pH, and temperature and their interactions were studied on maximum sustainable yield (MSY, a measure for biomass productivity), lipid content of N. salina, and invading organisms. Five different levels of each environmental predictor variable were tested. The environmental factors were kept within ranges that had previously been determined to allow positive N. salina growth (14.5–45.5 PSU; pH 6.3–9.7; 11–29 °C). The models created for this experiment showed that N. salina’s MSY and lipid content are not strongly affected over the broad range of salinity and temperature values. Calculated optima levels were 28 PSU/20 °C for MSY and 14.5 PSU/20 °C for lipid accumulation, but neither value significantly influenced the model. However, pH was the most important factor to influence algae productivity, and pH optimum was estimated around 8. Both MSY and lipid content were strongly reduced when pH deviated from the optimum. Occurrence of invading organisms seemed stochastic, and none of the environmental factors studied significantly influenced abundance. In conclusion, pH should be kept around 8 for maximum productivity of N. salina. Temperature and salinity should be kept around 20 °C and 28 PSU; however, moderate variations are not too much of a concern and might enhance lipid content of N. salina.     

Biomass productivity of two <Emphasis Type="Italic">Scenedesmus</Emphasis> strains cultivated semi-continuously in outdoor raceway ponds and flat-panel photobioreactors

Semi-continuous algal cultivation was completed in outdoor flat-panel photobioreactors (panels) and open raceway ponds (raceways) from February 17 to May 7, 2015 for side-by-side comparison of areal productivities at the Arizona Center for Algae Technology and Innovation in Mesa, AZ, USA. Experiments used two strains of Scenedesmus acutus (strains LB 0414 and LB 0424) to assess productivity, areal density, nutrient removal, and harvest volume across cultivation systems and algal strains. Panels showed an average biomass productivity of 19.0?±?0.6 g m^?2 day^?1 compared to 6.62?±?2.3 g m^?2 day^?1 for raceways. Photosynthetic efficiency ranged between 1.32 and 2.24 % for panels and between 0.30 and 0.68 % for raceways. Panels showed an average nitrogen consumption rate of 38.4?±?8.6 mg N L^?1 day^?1. Cultivation in raceways showed a consumption rate of 3.8?±?2.5 and 7.1?±?4.2 mg N L^?1 day^?1 for February/March and April/May, respectively, due to increase in biomass productivity. Excess nutrients were required to prevent a decrease in productivity. Daily biomass harvest volumes between 18 and 36 % from panels did not affect culture productivity, but density decreased with increased harvest volume. High cultivation temperatures above 30 °C caused strain LB 0414 to lyse and crash. Strain LB 0424 did not show any difference in biomass productivity when peak temperatures reached 34, 38, or 42 °C, but showed decreased productivity when the peak temperature during cultivation was 30 °C. Using algal strains with different temperature tolerances can generate increased annual biomass productivity.     

Optimization of aeration for biodiesel production by <Emphasis Type="Italic">Scenedesmus obliquus</Emphasis> grown in municipal wastewater

Despite the significant breakthroughs in research on microalgae as a feedstock for biodiesel, its production cost is still much higher than that of fossil diesel. One possible solution to overcome this problem is to optimize algal growth and lipid production in wastewater. The present study examines the optimization of pretreatment of municipal wastewater and aeration conditions in order to enhance the lipid productivity of Scenedesmus obliquus. Results showed that no significant differences were recorded in lipid productivity of S. obliquus grown in primary settled or sterilized municipal wastewater; however, ultrasound pretreatment of wastewater significantly decreased the lipid production. Whereas, aeration rates of 0.2 vvm significantly increased lipid content by 51 %, with respect to the non-aerated culture, which resulted in maximum lipid productivity (32.5 mg L^?1 day^?1). Furthermore, aeration enrichment by 2 % CO₂ resulted in increase of lipid productivity by 46 % over the CO₂ non-enriched aerated culture. Fatty acid profile showed that optimized aeration significantly enhanced monounsaturated fatty acid production, composed mainly of C18:1, by 1.8 times over the non-aerated S. obliquus culture with insignificant changes in polyunsaturated fatty acid proportion; suggesting better biodiesel characteristics for the optimized culture.     

The boosted lipid accumulation in microalga <Emphasis Type="Italic">Chlorella vulgaris</Emphasis> by a heterotrophy and nutrition-limitation transition cultivation regime

A model of heterotrophy and nutrition-limitation transition cultivation for efficient algal biomass and lipid production was proposed in this study, wherein sufficient robust heterotrophic-seed cells submitted into nitrogen-starvation induction for boosted lipid accumulation. The results demonstrated that heterotrophic-seed (HS) achieved specific growth rate of 1.35 day^?1 and biomass productivity of 1.93 mg/L/d, representing 6.42- and 32.16-fold, 2.01- and 2.75-fold more than that of photoautotrophic-seed (PS) and mixtrophic-seed (MS). Even though subsequent nutrition-limitation cultivation repressed the growth of HS, the overall lipid productivity caused by nitrogen-starvation was not offset by biomass loss. The most favorable lipid productivity (465.61 mg/L/d) of HS was 3.25 and 52.31 times higher than that of MS and PS. The high content of monounsaturated fatty acids (50.13%) over saturated and polyunsaturated fatty acids (totally 47.39%) in HS cells could provide superior oxidation stability and lower viscosity for biofuels generated from algal biomass feedstock. These findings suggested the feasibility of using heterotrophy and nutrition-limitation transition cultivation for enhancing the overall lipid productivity. Further, several critical enzymes (i.e. G3PDH, ME, and ACAD) were highly related to lipid accumulation and showed especially pronounced up-regulation or down-regulation expression in HS, which provide indications for shedding light on the molecular mechanisms of lipid accumulation and a prospective metabolic engineering for lipid production.     

Sucrose-enhanced biosynthesis of medicinally important antioxidant secondary metabolites in cell suspension cultures of <Emphasis Type="Italic">Artemisia absinthium</Emphasis> L.

Natural products are gaining tremendous importance in pharmaceutical industry and attention has been focused on the applications of in vitro technologies to enhance yield and productivity of such products. In this study, we investigated the accumulation of biomass and antioxidant secondary metabolites in response to different carbohydrate sources (sucrose, maltose, fructose and glucose) and sucrose concentrations (1, 3, 5, 7 and 9 %). Moreover, the effects of 3 % repeated sucrose feeding (day-12, -18 and -24) were also investigated. The results showed the superiority of disaccharides over monosaccharides for maximum biomass and secondary metabolites accumulation. Comparable profiles for maximum biomass were observed in response to sucrose and maltose and initial sucrose concentrations of 3 and 5 %. Maximum total phenolic and total flavonoid contents were displayed by cultures treated with sucrose and maltose; however, initial sucrose concentrations of 5 and 7 % were optimum for both classes of metabolites, respectively. Following 3 % extra sucrose feeding, cultures fed on day-24 (late-log phase) showed higher biomass, total phenolic and total flavonoid contents as compared to control cultures. Highest antioxidant activity was exhibited by maltose-treated cultures. Moreover, sucrose-treated cultures displayed positive correlation of antioxidant activity with total phenolics and total flavonoids production. This work describes the stimulatory role of disaccharides and sucrose feeding strategy for higher accumulation of phenolics and flavonoids, which could be potentially scaled up to bioreactor level for the bulk production of these metabolites in suspension cultures of A. absinthium.     

Early effect of sodium pentachlorophenate on photosynthetic activity of the alga <Emphasis Type="Italic">Chlorella pyrenoidosa</Emphasis> Chick. S-39

Early effects of 0.0001–10 mM sodium pentachlorophenate (PCP-Na) on the green alga Chlorella pyrenoidosa Chick. S-39 involves a rapid (within 1–2 min) decrease in the light-induced oxygen evolution by algal cells. The suppressed relative yield of variable chlorophyll fluorescence in C. pyrenoidosa in the presence of high PCP-Na concentrations and its dynamics provide evidence for rapid inactivation of photosystem 2, which is not observed at low concentrations of the toxicant. An analysis of the induction curve of delayed chlorophyll fluorescence in algal cells suggests that PCP-Na at low concentrations disturbs the coupling of electron transport and phosphorylation, whereas at high concentrations it inhibits electron transport and decreases the energy potential of photosynthetic membranes. The early toxic effect of PCP-Na is responsible for subsequent impairment of C. pyrenoidosa productivity.     

Process Engineering for High-Cell-Density Cultivation of Lipid Rich Microalgal Biomass of <Emphasis Type="Italic">Chlorella</Emphasis> sp. FC2 IITG

In the present study, process engineering strategy was applied to achieve lipid-rich biomass with high density of Chlorella sp. FC2 IITG under photoautotrophic condition. The strategy involved medium optimization, intermittent feeding of limiting nutrients, dynamic change in light intensity, and decoupling growth and lipid induction phases. Medium optimization was performed using combinations of artificial neural network or response surface methodology with genetic algorithm (ANN-GA and RSM-GA). Further, a fed-batch operation was employed to achieve high cell density with intermittent feeding of nitrate and phosphate along with stepwise increase in light intensity. Finally, mutually exclusive biomass and lipid production phases were decoupled into two-stage cultivation process: biomass generation in first stage under nutrient sufficient condition followed by lipid enrichment through nitrogen starvation. The key findings were as follows: (i) ANN-GA resulted in an increase in biomass titer of 157 % (0.95 g L^?1) in shake flask and 42.8 % (1.0 g L^?1) in bioreactor against unoptimized medium at light intensity of 20 μE m^?2 s^?1; (ii) further optimization of light intensity in bioreactor gave significantly improved biomass titer of 5.6 g L^?1 at light intensity of 250 μE m^?2 s^?1; (iii) high cell density of 13.5 g L^?1 with biomass productivity of 675 mg L^?1 day^?1 was achieved with dynamic increase in light intensity and intermittent feeding of limiting nutrients; (iv) finally, two-phase cultivation resulted in biomass titer of 17.7 g L^?1 and total lipid productivity of 313 mg L^?1 day^?1 which was highest among Chlorella sp. under photoautotrophic condition.     

pH and CO<Subscript>2</Subscript> effects on <Emphasis Type="Italic">Coelastrella</Emphasis> (<Emphasis Type="Italic">Scotiellopsis</Emphasis>) <Emphasis Type="Italic">rubescens</Emphasis> growth and metabolism

We studied effects of рН and СО₂ enrichment on the physiological condition and biochemical composition of a carotenogenic microalga Coelastrella (Scotiellopsis Vinatzer) rubescens Kaufnerová et Eliás (Scenedesmaceae, Sphaeropleales, Chlorophyceae), a promising source of natural astaxanthin. The microalga was grown at a constant pH (5, 6, 7 or 8) maintained by direct СО₂ injection. The air-sparged culture served as the control. Cell division rate and size, dry biomass productivity, the rates of nitrogen and phosphorus uptake as well as photosynthetic pigment and total lipid content and fatty acid composition were followed. С. rubescens possessed a narrow-range рН tolerance (the optimum рН 6–7). Under these conditions, the highest values of the maximum (1.0–1.1 1/day) and average (0.3–0.35 1/day) specific growth rate, chlorophyll а (4.8–4.9%) and total carotenoid dry weight percentages (1.7–1.8%) were recorded. Cell lipid fatty acid unsaturation index (1.851) and polyunsaturated fatty acid percentage (36–39%) and С18:3 ω3/С18:1 ω9 ratio (3.8–4.5) were also the highest under these conditions. A decline of рН to 5 brought about severe stress manifesting itself as a cell division cessation, photosynthetic apparatus reduction, two-fold increase in cell volume, accumulation of dry weight and lipids and a considerable decline in fatty acid unsaturation. Cultivation of С. rubescens without СО₂ enrichment resulted in a rapid alkalization of the medium to рН 9.5–10.5 impairing the physiological condition of the cells. Reasons of the deteriorative effects of suboptimal pH values on the physiological condition of C. rubescens are discussed.     

Comparison of the culture and harvesting of <Emphasis Type="Italic">Chlorella vulgaris</Emphasis> and <Emphasis Type="Italic">Tetradesmus obliquus</Emphasis> for the removal of pharmaceuticals from water

The objective of this work was to assess and compare the removal efficiency of paracetamol and salicylic acid from aqueous medium by a microalgae-based treatment, using either Chlorella vulgaris or Tetradesmus obliquus. Moreover, considering microalgae application in wastewater treatment, the influence of these pharmaceuticals in the algal nutrient removal capacity was evaluated. The removal of paracetamol by T. obliquus (>40 %) was larger than by C. vulgaris (>21 %) in batch culture, and this was also observed for salicylic acid (>93 % by T. obliquus and >25 % by C. vulgaris). Both strains removed nutrients (phosphate and nitrate) almost completely by the end of the batch culture, but T. obliquus showed the highest efficiency at the steady state conditions of the semicontinuous culture. In spite of this, under the flocculants here tested, the efficiency in the recovery of biomass was much higher for C. vulgaris. These results highlight the importance of strain selection in the application of microalgae for wastewater treatment and, particularly, for the removal of pharmaceuticals.