Productivity analysis of outdoor chemostat culture in tubular air-lift photobioreactors

Net productivity and biomass night losses in outdoor chemostat cultures ofPhaeodactylum tricornutum were analyzed in two tubular airlift photobioreactors at different dilution rates, photobioreactor surface/volume ratios and incident solar irradiance. In addition, an approximate model for the estimation of light profile and average irradiance inside outdoor tubular photobioreactors was proposed. In both photobioreactors, biomass productivity increased with dilution rate and daily incident solar radiation except at the highest incident solar irradiances and dilution rates, when photoinhibition effect was observed in the middle of the day. Variation of estimated average irradiance vs mean incident irradiance showed two effects: first, the outdoor cultures are adapted to average irradiance, and second, simultaneous photolimitation and photoinhibition took place at all assayed culture conditions, the extent of this phenomena being a function of the (incident)¹ irradiance and light regime inside the culture. Productivity ranged between 0.50 and 2.04 g L^–1 d^–1 in the tubular photobioreactor with the lower surface/volume ratio (S/V = 77.5 m^–1) and between 1.08 and 2.76 g L^–1 d^–1 in the other (S/V = 122.0 m^–1). The optimum dilution rate was 0.040 h^–1 in both reactors. Night-time biomass losses were a function of the average irradiance inside the culture, being lower in TPB0.03 than TPB0.06, due to a better light regime in the first. In both photobioreactors, biomass night losses strongly decreased when the photoinhibition effect was pronounced. However, net biomass productivity also decreased due to lower biomass generation during the day. Thus, optimum culture conditions were obtained when photolimitation and photoinhibition were balanced.     

n-3 Polyunsaturated fatty acid productivity of the marine microalgaIsochrysis galbana. Growth conditions and phenotypic selection

Two set of isolates were obtained in an isolation/selection programme to select eicosapentaenoic acid (EPA) and/or docosahexaenoic acid (DHA) rich strains ofIsochrysis galbana. EPA content was improved up to an average of 5.3% (d.wt) for the second set of isolates. On the other hand, with the aeration rate, pH and irradiance kept at low levels, the growth rate was slow and EPA synthesis was enhanced, but productivity increased when growth rates were maximum. A model relating steady-state dilution rates in chemostat cultures ofI. galbana to internal average irradiance is proposed. The greatest productivities were obtained between 0.0295 h^–1 and 0.0355 h^–1 with 300 mg m^–3 h^–1 for EPA and 130 mg m^–3 h^–1 for DHA.     

Variation of fatty acid profile with solar cycle in outdoor chemostat culture ofIsochrysis galbana ALII-4

Outdoor chemostat cultures of the marine microalgaIsochrysis galbana at constant dilution rate (0.034 h^–1 ) have been carried out under different weather conditions. Steady-state biomass concentrations were 1.61±0.03 kg m^–3 in May and 0.95±0.04 kg m^–3 in July, resulting in biomass output rates of 0.54 kg m^–3 d^–1 and 0.32 kg m^–3 d^–1 in May and July, respectively. Two patterns of daily variation with the solar cycle were observed in the fatty acid content. Saturated and mono-unsaturated fatty acids (16:0 and 16:1n7) show significant variation with the solar cycle, associated with short-term changes in environmental factors. Palmitic and palmitoleic acids are generated during daylight and consumed during the dark period. However, polyunsaturated fatty acids do not show a significant response to the solar cycle and their changes are associated with long-term variation in environmental factors. The maximum EPA productivity was obtained in May, 14.1 g m^–3 d^–1, which is close to that found in the literature for indoor continuous cultures. Nonetheless, the outdoor EPA content (up to 2.61 % d.wt) was lower than the indoor EPA content from a previous study (5% d.wt).     

Optimization of eicosapentaenoic acid production byPhaeodactylum tricornutumin the flat panel airlift (FPA) reactor

Biomass and eicosapentaenoic acid (EPA) productivities were investigated in a flat panel airlift loop reactor ideally mixed by static mixers. Growth with ammonium, urea and nitrate as nitrogen source were performed at different aeration rates. Cultures grew on ammonium but the decay of pH strongly inhibited biomass increase. On urea biomass productivity reached 2.35 g L^–1d^–1at an aeration rate of 0.66 vvm (24 h light per day, 1000 mol photon m^–2s^–1). Aeration rates between 0.33 vvm and 0.66 vvm and maximal productivities on urea were linearly dependent. Productivity on nitrate never exceeded 1.37 g L^–1d^–1. In the range of maximum productivity photosynthesis efficiency of 10.6% was reached at low irradiance (250 mol photon m^–2s^–1). Photosynthesis efficiency decreased to 4.8% at 1000 mol photon m^–2s^–1. At these high irradiances the flat panel airlift reactor showed a 35% higher volume productivity than the bubble column. At continuous culture conditions the influence of CO₂concentration in the supply air was tested. Highest productivities were reached at 1.25% (v/v) CO₂where the continuous culture yielded 1.04 g L^–1d^–1(16 h light per day, 1000 mol photon m^–2s^–1). The average EPA content amounted to 5.0% of cell dry weight, that resulted in EPA productivities of 52 mg L^–1d^–1(continuous culture, 16 h light per day) or 118 mg L^–1d^–1(batch culture, 24 h light per day).     

Comparison of photobioreactors for cultivation of Undaria pinnatifida gametophytes

Undaria pinnatifida gametophytes were grown in 2.5 l bubble column and airlift reactor at 25 °C and light intensity of 40 mol m^–2 s^–1 for 6 days. With aeration at 1 l min^–1, the airlift reactor yielded higher growth rate (0.12 mg DW ml^–1 d^–1) than a bubble column (0.08 mg DW ml^–1 d^–1). The advantages were related to the more homogeneous fluid dynamic characteristics of the airlift reactor.     

Differentiation during asexual reproduction and regeneration in a microturbellarian

Affordable biological technology for the reclamation of wastes and water of the waste streams from intensive livestock units is important in a country short of water. This study tested the concept of reclamation of waste by Streptocephalus macrourus (Crustacea: Anostraca) from the effluent of a high rate algal pond processing livestock wastes. S. macrourus showed a growth efficiency of 39% to 74% when fed optimal rations and cultured at densities between 10 and 400 1^–1. The maximum daily growth rates (0.15–0.21) approximate the growth rates of cladoceran or rotifer cultures managed for maximal biomass production. S. macrourus' ability to withstand crowding enabled the production from S. macrourus cultures (up to 91.8 mg dry mass l^–1 d^–1, or 1241 mg wet mass l^–1 d^–1) to exceed production recorded from cladoceran or rotifer cultures. Temperature influenced growth rate, with the highest growth rate occurring at 24 °C. The dilution rate of continuously fed cultures influenced growth rate, with the optimum dilution rate tested being 10 ml organism ^–1 d^–1. Mass mortality occurred when organisms were held at a density of 4000 l^–1. S. macrourus is able to convert algae grown on livestock waste efficiently into anostracan biomass, and is able to give a very high daily production.     

Outdoor cultivation ofArthrospira platensis during autumn and winter in temperate climates

Arthrospira (Spirulina) platensis M2 was grown outdoors in 50-mm diameter tubular reactors under the climatic conditions of central Italy (Florence) from September to December 1995 and in March 1996. Except for September, the cultures temperature was regulated. Mean productivities of 0.83, 0.44 and 0.61 g dry wt L^–1 d^–1 were achieved in autumn (September–October), winter (November–December) and March, respectively. In autumn and winter, the photosynthetic efficiency of the cultures and the degree of correlation between productivity and solar irradiance were significantly greater than in summer. The effect of cell density and aeration rate on productivity was evaluated in September. The productivity of cultures operated at high supra-optimal population density was about 30% less at high aeration rate (1.0 LL^–1 min^–1), and 50% less at standard aeration rate (0.17 LL^–1 min^–1), than that of control cultures kept at optimal population density and standard aeration rate. The reduction of productivity in high-density cultures was due to lower daylight output rates and higher night biomass losses (the latter were particularly relevant under standard aeration conditions). The main factor limiting productivity in closed reactors during autumn was the night temperature. Heating the cultures during daylight hours on sunny days did not cause any significant increase of the yields, since under sunlight the unheated cultures also reached the optimal temperature for growth early in the morning. On cloudy days, the day-time temperature of the unheated cultures remained well below the optimum, however this had only a limited effect on productivity since algal growth was mainly light-limited.     

A closed solar photobioreactor for cultivation of microalgae under supra-high irradiance: basic design and performance

An account is given of the setting up and use of a novel type of closed tubular photobioreactor at the Academic and University Centre in Nove Hrady, Czech Republic. This "penthouse-roof" photobioreactor was based on solar concentrators (linear Fresnel lenses) mounted in a climate-controlled greenhouse on top of the laboratory complex combining features of indoor and outdoor cultivation units. The dual-purpose system was designed for algal biomass production in temperate climate zone under well-controlled cultivation conditions and with surplus solar energy being used for heating service water. The system was used to study the strategy of microalgal acclimation to supra-high solar irradiance, with values as much as 3.5 times the ambient value, making the approach unique. The cultivation system proved to be fully functional with sufficient mixing and cooling, efficient oxygen stripping and light tracking. Experimental results (measurement of the maximum photochemical yield of PSII and non-photochemical quenching) showed that the cyanobacterium Spirulina (= Arthrospira) platensis cultivated under sufficient turbulence and biomass density was able to acclimate to irradiance values as high as 7 mmol photon m^–2 s^–1. The optimal biomass concentration of Spirulina cultures in September ranged between 1.2 to 2.2 g L^–1, which resulted in a net productivity of about 0.5 g L^–1 d^–1 corresponding to a biomass yield of 32.5 g m^–2 d^–1 (based on the minimum illuminated surface area of the photobioreactor).     

Effect of dilution rate on eicosapentaenoic acid productivity ofPhaeodactylum tricornutum utex 640 in outdoor chemostat culture

Summary Eicosapentaenoic acid (EPA) volumetric productivity from an outdoor chemostat culture ofPhaeodactylum tricornutum UTEX 640 in a 50-l tubular photobioreactor varies with dilution rate, reaching a maximum of 47.8 mg l^–1 d^–1 at D=0.36 d^–1. Continuous culture at high dilution rates' is proposed as the most adequate operating mode to maximize polyunsaturated fatty acid production.     

Photolimitation and photoinhibition as factors determining optimal dilution rate to produce eicosapentaenoic acid from cultures of the microalga Isochrysis galbana

Eicosapentaenoic acid (EPA) productivity from continuous cultures of the marine microalga Isochrysis galbana was studied, taking into account the irradiance on the reactor surface, that is, the photolimitation/photoinhibition regime to which the cells are exposed. Experiments were conducted under a wide variety of operating conditions. The dilution rate ranged from 0.005 h⁻¹ to 0.040 h⁻¹ at five external irradiances (820, 1620, 2050, 2450 and 3270 μmol photons m⁻² s⁻¹) covering photolimited to photoinhibited growth. Under these conditions, the specific growth rate (μ) was found to be the main factor influencing EPA content (ranging from 2.35% to 5.23% dryweight) and productivity (up to 0.88 mg l⁻¹ h⁻¹). The fatty acid content was not significantly affected by the external irradiance, but was influenced by the state of growth of the microalga, depending on whether the light regime was photolimiting or photoinhibiting. It might be suggested that light should no longer be considered an isolated factor affecting EPA synthesis, but an indirect influence through the photolimitation/photoinhibition regime and growth rate. At a given dilution rate, EPA content and biomass concentration are lower under photoinhibiting external irradiances than those corresponding to photolimiting conditions, and consequently EPA productivity decays. Since the effect of photoinhibition is less marked at high biomass concentration, a strategy to optimize EPA productivity from microalgal cultures could consist of reducing the dilution rate when the external irradiance increases above the phoinhibition threshold. Received: 16 January 1998 / Revised revision: 27 March 1998 / Accepted: 27 March 1998     

Mixotrophic growth of Phaeodactylum tricornutum on glycerol: growth rate and fatty acid profile

Mixotrophic growth of the eicosapentaenoic acid (EPA)producing diatom Phaeodactylum tricornutum UTEX640 was carried out in 1-L batch cultures under anexternal irradiance of 165 mol photons m^-2s^-1 by supplementing the inorganic culture mediumwith glycerol. The effect on the growth and the fattyacid profile was studied for different initialglycerol concentrations (0–0.1 M). The optimalglycerol concentration was 0.1 M.A lag phase was observed at high glycerolconcentrations. The present study also shows thatsuccessive additions of glycerol at 0.1M concentrationand using ammonium chloride as a nitrogen sourceremarkably increased the maximum biomass concentration(16.2 g L^-1) and maximum biomass productivity(61.5 mg L^-1 h^-1). These values wererespectively 9 and 8-fold higher than in thephotoautotrophically grown control. The level ofsaponifiable lipids in mixotrophically cultured cellswas significantly higher than in photoautotrophicallycultured cells and increased with the glycerolconcentration in the medium. The concentration ofstorage lipids, saturated and monounsaturated fattyacids, were enhanced but the EPA content did notchange significantly. The EPA content was around 2.2%of biomass dry weight. The maximum EPA yield was33.5 mg L^-1 d^-1 and was obtained in aculture containing 0.1 M glycerol, supplementedperiodically by ammonium chloride. This productivitywas 10-fold higher than the EPA productivity obtainedunder mixotrophic conditions.     

Production potential of eicosapentaenoic acid by the diatom Nitzschia laevis

To investigate the production potential of eicosapentaenoic acid (EPA) by the diatom Nitzschia laevis, the growth characteristics and fatty acid composition of the cells were studied under photoautotrophic, mixotrophic and heterotrophic conditions of growth. The specific growth rate and maximum biomass concentration were respectively 0.466 d^–1 and 2.27 g l^–1 for mixotrophic culture, 0.344 d^–1 and 2.04 g l^–1 for heterotrophic culture, and 0.167 d^–1 and 0.5 g l^–1 for photoautotrophic culture, respectively. As for EPA production, the yield and productivity were respectively 52.32 mg l^–1 and 10.46 mg l^–1 d for mixotrophic culture, 35.08 mg l^–1 and 6.37 mg l^–1 d for heterotrophic culture, and 6.78 mg l^–1 and 3.39 mg l^–1 d for photoautotrophic culture, respectively. Results suggest that mixotrophic culture is the most suitable growth mode for the production of EPA by the diatom Nitzschia laevis. The results are useful for the development of a cost-effective fermentation process for EPA production by Nitzschia laevis.     

Annual productivity of Spirulina (Arthrospira) and nutrient removal in a pig wastewater recycling process under tropical conditions

An evaluation was made of the annual productivity of Spirulina (Arthrospira) and its ability to remove nutrients in outdoor raceways treating anaerobic effluents from pig wastewater under tropical conditions. The study was based at a pilot plant at La Mancha beach, State of Veracruz, Mexico. Batch or semi-continuous cultures were established at different seasons during four consecutive years. The protein content of the harvested biomass and the N and P removal from the ponds were also evaluated. Anaerobic effluents from digested pig waste were added in a proportion of 2% (v/v) to untreated sea-water diluted 1:4 with fresh water supplemented with 2 g L^–1 sodium bicarbonate, at days 0, 3 and 5. A straight filament strain of Spirulina adapted to grow in this complex medium was utilized. A pH value 9.5 ± 0.2 was maintained. The productivity of batch cultures during summer 1998 was significantly more with a pond depth of 0.10 m than with a depth 0.065 m. The average productivity of semi-continuous cultures during summer 1999 was 14.4 g m^–2 d^–1 with a pond depth of 0.15 m and 15.1 g m^–2 d^–1 with a depth of 0.20 m. The average annual productivity for semi-continuous cultures operating with depths of 0.10 m for winter and 0.15 and 0.25 m for the rest of the year, was 11.8 g m^–2 d^–1. This is the highest value reported for a Spirulina cultivation system utilising sea-water. The average protein content of the semi-continuous cultures was 48.9% ash-free dry weight. NH₄-N removal was in the range 84–96% and P removal in the range of 72–87%, depending on the depth of the culture and the season.     

The cell composition of Nannochloropsis sp. changes under different irradiances in semicontinuous culture

Nannochloropsis sp. was grown semicontinuously with a rate of daily renewal of the culture media of 40% of the volume of the culture under different irradiances (40, 60, 80, 220 and 480 mol quanta m^–2 s^–1). Under the conditions tested, light saturation was achieved at 220 mol quanta m^–2 s^–1 with no significant increase in steady-state cell density or of dry weight productivity with higher irradiance, reaching values of 115 × 10⁶ cells ml^–1 and 375 mg l^–1 day^–1 respectively. C/N ratios clearly indicated the point of light saturation, decreasing with increasing irradiance for light-limited conditions and increasing for light-saturated conditions. Under light-limited conditions, an increase in the irradiance produced an increase in the protein percentage of the organic fraction to the detriment of lipids and carbohydrates, while small changes were recorded under light-saturated conditions. The degree of unsaturation of fatty acids was lower with increasing irradiance, with a three-fold decrease of the percentage of total n–3 fatty acids, from 29 to 8% of total fatty acids, caused mainly by a decrease of eicosapentaenoic acid (EPA) (20:5n–3). The microalga reached its maximal value of dry weight productivity (375 mg l^–1 day^–1), EPA productivity (3.2 mg l^–1 day^–1) and maximal protein content (36% of the organic content) at the point at which light saturation was achieved. Results demonstrate the efficiency of the use of the irradiance for the modification of the biochemical composition of Nannochloropsis sp.     

Ulva rigida (Ulvales,Chlorophyta) tank culture as biofilters for dissolved inorganic nitrogen from fishpond effluents

Ulva rigida was cultivated in 7501 tanks at different densities with direct and continuous inflow (at 2, 4, 8 and 12 volumes d^–1) of the effluents from a commercial marine fishpond (40 metric tonnes, Tm, of Sparus aurata, water exchange rate of 16 m³ Tm^–1) in order to assess the maximum and optimum dissolved inorganic nitrogen (DIN) uptake rate and the annual stability of the Ulva tank biofiltering system. Maximum yields (40 g DW m^–2 d^–1) were obtained at a density of 2.5 g FW 1^–1 and at a DIN inflow rate of 1.7 g DIN m^–2 d^–1. Maximum DIN uptake rates were obtained during summer (2.2 g DIN M^–2 d^–1), and minimum in winter (1.1 g DIN m^–2 d^–1) with a yearly average DIN uptake rate of 1.77 g DIN m^–2 d^–1 At yearly average DIN removal efficiency (2.0 g DIN m^–2 d^–1, if winter period is excluded), 153 m² of Ulva tank surface would be needed to recover 100% of the DIN produced by 1 Tm of fish.Abbreviations DIN= dissolved inorganic nitrogen (NH _inf4 ^sup+ + NO _inf3 ^sup– + NO _inf2 ^sup– ); - FW= fresh weight; - DW= dry weight; - PFD= photon flux density; - V= DIN uptake rate     

Light/dark cyclic movement of algal culture (<Emphasis Type="Italic">Synechocystis aquatilis</Emphasis>) in outdoor inclined tubular photobioreactor equipped with static mixers for efficient production of biomass

Synechocystis aquatilis SI-2 was grown outdoors in a 12.5cm diam. tubular photobioreactor equipped with static mixers. The static mixers ensured that cells were efficiently circulated between the upper (illuminated) and lower (dark) sections of the tubes. The biomass productivity varied from 22 to 45g m^–2d^–1, with an average of 35g m^–2d^–1, etc which corresponded to average CO₂ fixation rate of about 57 g CO₂ m^–2 d^–1. The static mixers not only helped in improving the biomass productivities but also have a high potential to lower the photoinhibitory effect of light during the outdoor cultures of algae. Revisions requested 27 July 2004; Revisions received 12 November 2004     

n-3 PUFA productivity in chemostat cultures of microalgae

Eicosapentaenoic (EPA) and docosahexaenoic (DHA) acid productivities from chemostat cultures of an isolate of Isochrysis galbana have been studied. The productivities reached in the interval of dilution rates between 0.0295 h^–1 and 0.0355 h^–1 were 1.5mg·1^–1·h^–1 for lipids, 300 g·1^–1·h^–1 for EPA and 130g1·1^–1·h^–1 for DHA. Furthermore, light attenuation by mutual shading, and agitation speed influences on growth and fatty acid composition were analysed. A model relating steady-state dilution rates to internal average light intensity has been proposed, the parameter values of which obtained by non-linear regression were: maximum specific growth rate (_max)=0.0426 h^–1; the affinity of cells to light (I_k) = 10.92 W·m^–2; the exponent (n) = 5.13; regression coefficient (r ²)=0.9999. Correspondence to: E. Molina Grima     

Light response of D1 turnover and photosystem II efficiency in the seagrassZostera capricorni

Biochemical and biophysical parameters, including D1-protein turnover, chlorophyll fluorescence, oxygen evolution activity and zeaxanthin formation were measured in the marine seagrassZostera capricorni (Aschers) in response to limiting (100 mol·m^–2·^–1), saturating (350 mol·m^–2·s^–1) or photoinhibitory (1100 mol·m^–2·s^–1) irradiances. Synthesis of D1 was maximal at 350 mol·m^–2·s^–1 which was also the irradiance at which the rate of photosynthetic O₂ evolution was maximal. Degradation of D1 was saturated at 350 mol·m^–2·s^–1. The rate of D1 synthesis at 1100 mol·m^–2·s^–1 was very similar to that at 350 mol·m^–2·s^–1 for the first 90 min but then declined. At limiting or saturating irradiance little change was observed in the ratio of variable to maximal fluorescence (Fv/Fm) measured after dark adaptation of the leaves, while significant photoinhibition occurred at 1100 mol·m^–2·s^–1. The proportion of zeaxanthin in the total xanthophyll pool increased with increasing irradiance, indicative of the presence of a photoprotective xanthophyll cycle in this seagrass. These results are consistent with a high level of regulatory D1 turnover inZostera under non-photoinhibitory irradiance conditions, as has been found previously for terrestrial plants.We would like to thank Professor Peter Böger (Department of Plant Biochemistry, University of Konstanz, Germany) for the kind gift of D1 antibodies. This work was partly supported by a University of Queensland Enabling Grant to CC.     

Culture ofGelidium sesquipedale (Clem.) Born.et Thur. in a chemostat system. Biomass production and metabolic responses affected by N flow

Gelidium sesquipedale is the most important raw material used for extraction of agar in Spain. Based on chemostats, a system of culture for macroalgae with a continuous flow of culture medium has been developed. A stressed morphotype from the South of Spain was cultured, and the effects of different rates of NO ₃ ^– flow on growth and internal constituents were investigated in the laboratory. Cultivation was successful after optimizing factors affecting growth, such as irradiance level, renewal rate and water movement. Mass production was dependent on N supply. With a flow of 35 mol NO₃ ^– g^–1 DW d^–1, optimal values of growth (2.1% d^–1) and biomass yield were obtained. In these conditions, biomass yield resembled the values observed in natural populations (about 500 g DW m^–2 y^–1). When the flow of N was reduced to 15 mol NO ₃ ^– g^–1 DW d^–1, growth rate and biomass yield were reduced three-fold, and were null when N was supplied as 7 mol NO ₃ ^– g^–1 DW d^–1. C:N ratio was an index of the physiological status of the tissue, remaining low when N was sufficient and raised to critical values when N supply was limited. Phycobiliproteins, kept at a constant irradiance level, were affected by N supply, acting as an internal nitrogen reserve, unlike chlorophylla. An effective phycobiliprotein synthesis took place when the flow of N was sufficient. Agar yield, on dry weight basis, was similar as a function of N flow, whereas agar yield of the culture was higher when N was sufficient as a result of growth not being limited by N.This system of culture, commonly used in microalgal studies, may have an important use in macroalgae as a system to obtain biomass of high quality as well as a good tool for physiological studies in conditions of continuous and controlled flow of nutrients.     

Effect of cell density and irradiance on growth,proximate composition and eicosapentaenoic acid production ofPhaeodactylum tricornutum grown in a tubular photobioreactor

Growth and eicosapentaenoic acid (EPA) productivity of the diatomPhaeodactylum tricornutum grown semicontinuously in a helical tubular photobioreactor were examined under a range of irradiances (approximately 56 to 1712 µmol photons m^-2 s^-1) and cell densities (3 × 10⁶ to 18 × 10⁶ cells mL^-1). Self shading sets the upper limit of operational maximum cell density. Higher irradiance increases this upper limit and also increase the growth rate. Biomass productivity and EPA productivity were enhanced at those cell densities which support the fastest growth rate irrespective of irradiance. The cell protein content increased with increasing irradiance and the carbohydrate and lipid content increased with increasing cell density. EPA productivity was greatest at the highest irradiance. This study shows that biomass productivity and EPA productivity can be maximised by optimising cell density and irradiance, as well as by addition of CO₂.Author for correspondence