Improved biomass production of cyanobacteria by reutilization of the culture medium

The biomass production of a cyanobacterium (Nostoc sp.) in a photoreactor with a low illuminated surface area to volume ratio was improved by the reutilization of the culture medium. After six succesive utilizations the growth ofNostoc sp. amounted to 2.15 g/l with an average content in phycobiliproteins of 14.4% on dry weight basis. The procedure reported allowed an 80% increase in biomass. The cellular self-sedimentation proved to be effective for biomass separation between reutilization steps.     

Microbial cleaning of waste gas containing volatile organic compounds in a bioreactor system with a closed gas circuit

The cleaning of the exhaust gases of a bioreactor containing volatile hydrocarbons in a bioreactor system with a closed gas circuit is described. The bioreactor system consisted of three different reactor types: a stirred tank which was filled with hydrocarbon-containing waste water to simulate the exhaust gases of a remediation process; a trickle-bed reactor for aerobic treatment of the exhaust gas from the stirred tank; and a photoreactor containing an algae culture which assimilated CO₂ from the trickle-bed reactor and also produced O₂. With this bioreactor system, it was possible to efficiently remove volatile organic compounds (VOC) from the waste gases. Depending on the type of waste water investigated, elimination rates of 41% to 93% of BTEX (benzene, ethylbenzene, toluene, xylene) and 29% to 53% of VCH (volatile chlorinated hydrocarbons) were obtained. Due to the photosynthesis of the algae in the system's photoreactor, oxygen concentrations between 12% and 18% [v/v], equivalent to about 57% to 83% DOT, were obtained. This concentration permitted the aerobic degradation to be carried out without having to add fresh air. The trickle-bed reactor and the photoreactor worked continuously, whereas the waste water in the stirred bioreactor was replaced in different batches. The accumulation of toxic compounds in the nutrient solutions of the trickle-bed (EC-50 > 30 g/l) and of the photoreactor (EC-50 > 35 g/l) was low. Carbon dioxide concentrations in the gas flow were higher than in fresh air (1% to 3% [vol/vol]), but no long-term accumulation of CO₂ occurred. This means that the algae in the photoreactor were active enough to assimilate the CO₂ which had been produced. They were also able to produce sufficient oxygen for aerobic hydrocarbon degradation. The system described is a first step towards treating waste gases which results from the bioremediation of hydrocarbon-contaminated media in a closed gas circuit without any emission (e.g. VOC, CO₂, germs).     

Isolation of a hydroxyproline-secreting pigment-deficient mutant ofNostoc sp. by metronidazole selection

Summary A pigment-deficient mutant ofNostoc sp. was induced by N-methyl-N-nitro-N-nitrosoguanidine and selected in a medium containing metronidazole. It formed chains of heterocysts, grew more slowly than the control, and excreted hydroxyproline into the medium, imparting a pinkish-brown colour to the cultures.

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Aislamiento de un mutante do Nostoc sp. deficiente en pigmentos y secretor de hidroxiprolina en un medio selectivo con metronidazol

Resumen Un mutante pigmento-deficiente deNostoc sp. fue inducido mediante N-metil-N-nitro-N-nitrosoguanidina y seleccionado en un medio conteniendo metronidazol. La cepa mutante creció más lentamente que la control formando cadenas de heterocistos y excretando hidroxiprolina al medio lo cual confirió una coloración rosa-parduzca a los cultivos.

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Isolement par sélection avec le métronidazole d'un mutant de Nostoc sp. excrétant de l'hydroxyproline et déficient en pigment

Résumé Un mutant deNostoc sp. déficient en pigment a été induit par la N-méthyl-N-nitro-N-nitrosoguanidine et sélectionné dans un milieu contenant du métronidazole. Ce mutant forme des chaînes d'hétérocystes, pousse plus lentement que le contrôle, et excréte de l'hydroxyproline dans le milieu, conférant aux cultures une coloration brun rosâtre.

     

Effect of nutritional factors on the culture of Nostoc sp. as a source of phycobiliproteins

Summary The effect of nutritional factors in a new culture medium (BW₃) is described for the cyanobacterium Nostoc sp. The growth of Nostoc sp. was higher in BW₃ than in control media currently used for cyanobacteria. With medium BW₃ the content of the pigment c-phycocyanin depended on the culture conditions employed, particularly the nature of the nitrogen and carbon sources. Higher amounts of c-phycocyanin amounting to 20.1% on a dry-weight basis were accumulated when both sources were supplied in the gas phase of the culture. The phycobiliproteins of Nostoc sp. were resolved into two components: c-phycocyanin (_max = 614 nm) and allophycocyanin (_max = 652 nm). The phycobiliprotein composition was 30% allophycocyanin and 70% c-phycocyanin. The culture of Nostoc sp. in BW₃ medium seems promising as a source of biomass for the production of natural dyes.     

Nitrogenase activity and dark CO2 fixation in the lichen Peltigera aphthosa Willd

The lichen Peltigera aphthosa consists of a fungus and green alga (Coccomyxa) in the main thallus and of a Nostoc located in superficial packets, intermixed with fungus, called cephalodia. Dark nitrogenase activity (acetylene reduction) of lichen discs (of alga, fungus and Nostoc) and of excised cephalodia was sustained at higher rates and for longer than was the dark nitrogenase activity of the isolated Nostoc growing exponentially. Dark nitrogenase activity of the symbiotic Nostoc was supported by the catabolism of polyglucose accumulated in the ligh and which in darkness served to supply ATP and reductant. The decrease in glucose content of the cephalodia paralleled the decline in dark nitrogenase activity in the presence of CO₂; in the absence of CO₂ dark nitrogenase activity declined faster although the rate of glucose loss was similar in the presence and absence of CO₂. Dark CO₂ fixation, which after 30 min in darkness represented 17 and 20% of the light rates of discs and cephalodia, respectively, also facilitated dark nitrogenase activity. The isolated Nostoc, the Coccomyxa and the excised fungus all fixed CO₂ in the dark; in the lichen most dark CO₂ fixation was probably due to the fungus. Kinetic studies using discs or cephalodia showed highest initial incorporation of ¹⁴CO₂ in the dark in to oxaloacetate, aspartate, malate and fumarate; incorporation in to alanine and citrulline was low; incorporation in to sugar phosphates, phosphoglyceric acid and sugar alcohols was not significant. Substantial activities of the enzymes phosphoenolpyruvate (PEP) carboxylase (EC 4.1.1.31) and carbamoyl-phosphate synthase (EC 2.7.2.5 and 2.7.2.9) were detected but the activities of PEP carboxykinase (EC 4.1.1.49) and PEP carboxyphosphotransferase (EC 4.1.1.38) were negligible. In the dark nitrogenase activity by the cephalodia, but not by the free-living Nostoc, declined more rapidly in the absence than in the presence of CO₂ in the gas phase. Exogenous NH ₄ ⁺ inhibited nitrogenase activity by cephalodia in the dark especially in the absence of CO₂ but had no effect in the light. The overall data suggest that in the lichen dark CO₂ fixation by the fungus may provide carbon skeletons which accept NH ₄ ⁺ released by the cyanobacterium and that in the absence of CO₂, NH ₄ ⁺ directly, or indirectly via a mechanism which involves glutamine synthetase, inhibits nitrogenase activity.Abbreviations CP carbamoyl phosphate - EDTA ethylenedi-amine tetraacetic acid - PEP phosphoenolpyruvate - RuBP ribulose 1,5 bisphosphate     

Methanogenic degradation of hydroquinone in an anaerobic fixed-bed reactor

Summary A 500-ml fixed-bed reactor filled with glass sinter spools was used to study the dynamics and potential of methanogenic hydroquinone degradation. The concentration of hydroquinone as sole energy and carbon source in the inflowing medium varied from 0.55 to 2.2 g/l. Degradation of hydroquinone to methane and CO₂ was complete at low flow rates (170 ml/h) and low hydroquinone concentrations (0.55 g/l). At higher hydroquinone concentrations and/or higher flow rates, acetate accumulated in the reactor, and traces of hydroquinone were detected in the outflowing medium. The maximum degradation rate was 0.3–0.4 hydroquinone/h per 500 ml reactor volume. The bacterial community that established in the reactor after several weeks of operation was fairly stable, and consisted primarily of three bacterial species: a rod-shaped bacterium responsible for the degradation of hydroquinone to acetate and hydrogen, and two species of methanogenic bacteria, Methanospirillum hungatei and Methanothix sp.     

Characteristics of Hormogonia Formation by Symbiotic Nostoc spp. in Response to the Presence of Anthoceros punctatus or Its Extracellular Products

Nostocacean cyanobacteria typically produce gliding filaments termed hormogonia at a low frequency as part of their life cycle. We report here that all Nostoc spp. competent in establishing a symbiotic association with the hornwort Anthoceros punctatus formed hormogonial filaments at a high frequency in the presence of A. punctatus. The hormogonia-inducing activity was produced by A. punctatus under nitrogen-limited culture conditions. The hormogonia of the symbiotically competent Nostoc spp. were characterized as motile (gliding) filaments lacking heterocysts and with distinctly smaller cells than those of vegetative filaments; the small cells resulted from a continuation of cell division uncoupled from biomass increase. An essentially complete conversion of vegetative filaments to hormogonia occurred within 12 h of exposure of Nostoc sp. strain 7801 to A. punctatus growth-conditioned medium. Hormogonia formation was accompanied by loss of nitrogen fixation (acetylene reduction) and by decreases in photosynthetic CO₂ fixation and in vivo NH₄⁺ assimilation of 30% and approximately 40%, respectively. The rates of acetylene reduction and CO₂ fixation returned to approximately the control rates within 72 to 96 h after hormogonia induction, as the cultures of Nostoc sp. strain 7801 differentiated heterocysts and reverted to the vegetative growth state. The relationship between hormogonia formation and symbiotic competence is discussed.     

Hydrogen uptake in Nostoc sp. strain PCC 73102. Cloning and characterization of a hupSL homologue

Structural genes encoding an uptake hydrogenase of Nostoc sp. strain PCC 73102 were isolated. From partial libraries of genomic DNA, two clones (pNfo01 and pNfo02) were selected and sequenced, revealing the complete sequence of both a hupS (960 bases) and a hupL (1,593 bases) homologue in Nostoc sp. strain PCC 73102. A comparison between the deduced amino acid sequences of HupS and HupL of Nostoc sp. strain PCC 73102 and Anabaena sp. strain PCC 7120 showed that the HupS proteins are 89% identical and the HupL proteins are 91% identical. However, the noncoding region between the genes in Nostoc sp. strain PCC 73102 (192 bases) is longer than that of Anabaena sp. strain PCC 7120 and of many other microorganisms. Southern hybridizations using DNA from both N₂-fixing and non-N₂-fixing cells of Nostoc sp. strain PCC 73102 and different probes from within hupL clearly demonstrated that, in contrast to Anabaena sp. strain PCC 7120, there is no rearrangement within hupL of Nostoc sp. strain PCC 73102. Indeed, 6 nucleotides out of 16 within the potential recombination site are different from those of Anabaena sp. strain PCC 7120. Furthermore, we have recently published evidence demonstrating the absence of the bidirectional/reversible hydrogenase in Nostoc sp. strain PCC 73102. The present knowledge, in combination with the unique characteristics, makes Nostoc sp. strain PCC 73102 an interesting candidate for the study of deletion mutants lacking the uptake-type enzyme. Received: 20 August 1997 / Accepted: 24 November 1997     

A culture method for microalgal forms using two-tier vessel providing carbon-dioxide environment: studies on growth and carotenoid production

A culture method was developed for photoautotrophic culture of Haematococcus pluvialis, Chlorella vulgaris, Scenedesmus obliquus, Spirulina platensis, Nostoc and Stigonema in a two-tier flask consisting of nutrient media in the upper chamber and CO₂ generating buffer mixture (KHCO₃/K₂CO₃) in the lower chamber. The concentration of buffer mixture was varied to obtain desired levels of CO₂. CO₂ at 2.0% (v/v) level enhanced growth and chlorophyll content over control cultures (without CO₂ supplementation) in all microalgal species. Haematococcus pluvialis culture in BBM and KM1 media showed 6.71- and 2.07-fold increase in biomass yields with astaxanthin productivity at 7.26 and 7.48 mg l^–1 level respectively. CO₂ supplementation to C. vulgaris and S. obliquus cultures resulted in 5.97- and 7.30-folds increase in biomass with 2–3 fold increase in chlorophyll and carotenoid contents over their respective controls. Similarly 2–3 fold increase in chlorophyll and carotenoid contents were observed in Sp. platensis, Nostoc and Stigonema spp. This culture methodology will provide information on CO₂ requirement for growth of algae and metabolite production and also facilitates studies on the influence of light and temperature conditions.     

Bubble free gaseous transfer in bioreactors using perfluorocarbons

In this work the concept of bubble-free oxygenation that was able to ensure oxygen supply and carbon dioxide extraction for a chemostat culture of Escherichia coli was experimentally demonstrated. It was operated at the dilution rate of 0.275 h^–1 under atmospheric pressure and at 35.5 °C. Foralkyl, a commercial perfluorocarbon, added in the influent medium under emulsified form and at 50% volumic fraction, was able to provide 0.17 g O₂/l/h and extract 0.23 g CO₂/l/h for a culture at 0.74 g/l of biomass. This oxygen supply was close to the maximum oxygenation Foralkyl was theoretically able to provide at this pressure when imposing a minimum oxygen concentration of 1 mg/l in the water phase. The quantification of transfer was not done from a direct measurement of oxygen transfer rates because conventional oxygen concentration measurement by membrane polarographic probe in an emulsion was not judged reliable. This evaluation was done by referring to conventional aerated culture whose measurable parameters (biomass and product concentrations) were found unaffected when shifting to the novel oxygenation device.List of Symbols C _LV ^* , C _LV g/l dissolved oxygen concentration in the vector phase at equilibrium and in the reactor - C _LW ^* , C _LW g/l dissolved oxygen concentration in the water phase at equilibrium and in the reactor - C _LWinput , C _LWoutput g/l dissolved oxygen concentrations in aqueous medium in the reactor input and output flow - D h^–1 dilution rate - ^DMM_{o 2}, MM_{co 2} g/mol oxygen and carbon dioxide molar masses - %O_2input, %O_2output oxygen percentages in the reactor input and output flow - %CO_2input, %CO_2output carbon dioxide percentages in the reactor input and output flow - %N _2input %N _2output nitrogen percentages in the reactor input and output flow - Q _G.i ⁿ Q _G.o ⁿ l/h gas flow rates at the reactor input and output at normal conditions (273 K, 1 atm) - Q _L l/h liquid flow rate - Q _LW , Q _LV l/h water and vector flow rates - r_{O 2} g/l/h oxygen consumption rate - r _x g/l/h biomass production rate - r _{CO 2} g/l/h carbon dioxide production rate - V _L l fermentor aqueous volume - V _LW , V _LV l water phase and vector phase volume - V _{O 2}, V _{CO 2}, l/mol oxygen and carbon dioxide molar volume under gaseous form at normal conditions (273 K, 1 atm) - Y _{O 2 x} gO₂/g cell oxygen consumption yield for biomass growth - Y _sx g glucose/g cell glucose consumption yield for biomass growth - vector volumic fraction - h^–1 growth rate This work was totally financed by the European Space Agency.     

Optimization of growth and fatty acid composition of a unicellular marine picoplankton,Nannochloropsis sp., with enriched carbon sources

A unicellular marine picoplankton, Nannochloropsis sp., was grown under CO₂-enriched photoautotrophic or/and acetate-added mixotrophic conditions. Photoautotrophic conditions with enriched CO₂ of 2800 l CO₂ l^–1 and aeration gave the highest biomass yield (634 mg dry wt l^–1), the highest total lipid content (9% of dry wt), total fatty acids (64 mg g^–1 dry wt), polyunsaturated fatty acids (35% total fatty acids) and eicosapentaenoic acid (EPA, 20:53) (16 mg g^–1 dry wt or 25% of total fatty acids). Mixotrophic cultures gave a greater protein content but less carbohydrates. Adding sodium acetate (2 mM) decreased the amounts of the total fatty acids and EPA. Elevation of CO₂ in photoautotrophic culture thus enhances growth and raises the production of EPA in Nannochloropsis sp.     

NOSTOC (CYANOPHYCEAE) GOES NUDE: EXTRACELLULAR POLYSACCHARIDES SERVE AS A SINK FOR REDUCING POWER UNDER UNBALANCED C/N METABOLISM1

Many species of the filamentous N₂‐fixing heterocyst‐forming Cyanobacteria of the genus Nostoc produce large amounts of extracellular polymeric substances (EPS), but hitherto no general model has been proposed of the factors that control their synthesis. Previously, we demonstrated a strong correlation between the presence of a glycocalyx (or EPS capsule) and diazotrophic growth in the genus Nostoc. When grown with nitrate, nude morphotypes lacking a glycocalyx were obtained for all the capsulated strains tested. CO₂ availability was pro‐posed as a key factor that controls the synthesis of the capsule. To test this hypothesis, Nostoc PCC 7936 was cultured diazotrophically (N₂) or with nitrate with different CO₂ supplies. By tuning the pH and the supply of CO₂, capsulated or nude mor‐photypes were obtained irrespective of the source of nitrogen. Exocellular polysaccharides were synthesized only when the fixed carbon exceeded the amount of nitrogen available. The glycocalyx is not needed for the optimal functioning of nitrogenase because diazotrophic cultures grew equally well, irrespective of whether they were capsulated or nude. Capsulated cultures possessed protein to carbohydrate ratios that ranged between 1 and 1.5, whereas in nude cultures the ratio ranged between 2 and 2.5. Low protein to carbohydrate ratios were indicative for either nitrogen‐limited or carbon‐oversaturated cultures. The results demonstrate that in Nostoc EPS serve as a sink for the excess fixed carbon under unbalanced C/N metabolism.     

On-line monitoring and modelling based process control of high rate nitrification — lab scale experimental results

This paper presents a new concept for the control of nitrification in highly polluted waste waters. The approach is based on mathematical modelling. To determine the substrate degradation rates of the microorganisms involved, a mathematical model using gas measurement is used. A fuzzy-controller maximises the capacity utilisation efficiencies. The experiments carried out in a lab-scale reactor demonstrate that even with highly varying ammonia concentrations in the influent, the nitrogen concentrations in the effluent can be kept within legal limits.List of Symbols c mg/l concentration - c mg/l gas concentration - H ₂ Henry-coefficient - k _L a 1/h mass transfer coefficient - mol/l dissociation constant - K _iS mg/l substrate inhibitor constant - k _iH mg/l inhibitor constant - k _S mg/l saturation constant - K _O2 mg/l oxygen saturation constant - r(B) mg/lh growth rate - r(S) mg/lh degradation reaction rate - t _v h retention time - T °C temperature - V 1 volume - V 1/h flow rate - Y g/g yield coefficient - k _b capacity utilisation efficiency - 1/h specific growth rate     

Predicting CO2 gain and photosynthetic light acclimation from fluorescence yield and quenching in cyano-lichens

Modulated chlorophylla fluorescence is useful for eco-physiological studies of lichens as it is sensitive, non-invasive and specific to the photobiont. We assessed the validity of using fluorescence yield to predict CO₂ gain in cyano-lichens, by simultaneous measurements of CO₂ gas exchange and chlorophylla fluorescence in five species withNostoc-photobionts. For comparison, O₂ evolution and fluorescence were measured in isolated cells ofNostoc, derived fromPeltigera canina (Nostoc PC). At irradiances up to the growth light level, predictions from fluorescence yield underestimated true photosynthesis, to various extents depending on species. This reflected the combined effect of a state transition in darkness, which was not fully relaxed until the growth light level was reached, and a phycobilin contribution to the minimum fluorescence yield (F_o). Above the growth light level, the model progressively overestimated assimilation, reflecting increased electron flow to oxygen under excess irradiance. In cyanobacteria, this flow maintains photosystem II centres open even up to photoinhibitory light levels without contributing to CO₂ fixation. Despite this we show that gross CO₂ gain may be predicted from fluorescence yield also in cyanolichens when the analysis is made near the acclimated growth light level. This level can be obtained even when measurements are performed in the field, since it coincides with a minimum in non-photochemical fluorescence quenching (NPQ). However, the absolute relation between fluorescence yield and gross CO₂ gain varies between species. It may therefore be necessary to standardise the fluorescence prediction for each species with CO₂ gas exchange.Abbreviations CCM CO₂-Concentrating mechanism - Chl chlorophyll - C_i inorganic carbon - 0 convexity (curvature of the light response curve) - ETR electron transport rate - F_o minimum fluorescence yield - F_m maximal fluorescence yield - F_s fluorescence yield at steady-state photosynthesis - F_v variable fluorescence yield - F_v/F_m dark ratio of variable to maximal fluorescence yield after dark adaptation - F_vF_mmax ratio of variable to maximal fluorescence yield in the absence of quenching - _CO2 maximum quantum yield of CO₂ assimilation - _PS quantum yield of photosystem II photochemistry - GP gross photosynthesis - I irradiance (mol quanta·m^–2·s^–1) - NPQ non photochemical fluorescence quenching - qp photochemical fluorescence quenching     

High Photobiological Hydrogen Production Activity of a <Emphasis Type="Italic">Nostoc</Emphasis> sp. PCC 7422 Uptake Hydrogenase-Deficient Mutant with High Nitrogenase Activity

We describe a strategy to establish cyanobacterial strains with high levels of H₂ production that involves the identification of promising wild-type strains followed by optimization of the selected strains using genetic engineering. Nostoc sp. PCC 7422 was chosen from 12 other heterocystous strains, because it has the highest nitrogenase activity. We sequenced the uptake hydrogenase (Hup) gene cluster as well as the bidirectional hydrogenase gene cluster from the strain, and constructed a mutant (ΔhupL) by insertional disruption of the hupL gene. The ΔhupL mutant produced H₂ at 100 μmoles mg chlorophyll a ^-1 h^-1, a rate three times that of the wild-type. The ΔhupL cells could accumulate H₂ to about 29% (v/v) accompanied by O₂ evolution in 6 days, under a starting gas phase of Ar + 5% CO₂. The presence of 20% O₂ in the initial gas phase inhibited H₂ accumulation of the ΔhupL cells by less than 20% until day 7.     

Chlorobium limicola forma thiosulfatophilum: Biocatalyst in the Production of Sulfur and Organic Carbon from a Gas Stream Containing H2S and CO2

Chlorobium limicola forma thiosulfatophilum (ATCC 17092) was grown in a 1-liter continuously stirred tank reactor (800-ml liquid volume) at pH 6.8, 30°C, saturated light intensity, and a gas flow rate of 23.6 ml/min from a gas cylinder blend consisting of 3.9 mol% H₂S, 9.2 mol% CO₂, 86.4 mol% N₂, and 0.5 mol% H₂. This is the first demonstration of photoautotrophic growth of a Chlorobium sp. on a continuous inorganic gas feed. A significant potential exists for applying this photoautotrophic process to desulfurization and CO₂ fixation of gases containing acidic components (H₂S and CO₂).     

Heterotrophic metabolism and diazotrophic growth ofNostoc sp. fromCycas circinalis

The cyanobiont ofCycas circinalis (identified asNostoc sp.) was isolated and its heterotrophic metabolism was studied in free culture under nitrogen-fixing conditions. Morphology, growth rate, nitrogenase activity, biochemical composition, efficiency of assimilation of organic carbon and molecular nitrogen were determined under different conditions of energy and carbon supply. The study has revealed the high potential of the heterotrophic metabolism in this symbiotic cyanobacterium. Although low rates of metabolic activities were attained under heterotrophic conditions, the efficiencies of organic carbon utilization (0.48 g cell-carbon per g glucose-carbon in chemoheterotrophy, from 0.65 to 0.74 under photoheterotrophy) and of N₂ assimilation (35.0 mg N₂ fixed per g glucose used in chemoheterotrophy, from 58.3 to 61.9 under photoheterotrophy) displayed by this organism were among the highest ever found in diazotrophically grown microorganisms. The isolate fromC. circinalis was able to grow indefinitely in the dark under nitrogen-fixing conditions, maintaining a well balanced biosynthetic activity and the capacity to resume photosynthetic metabolism quickly. The significance of the heterotrophic potential of this symbioticNostoc is discussed.     

The air‐lift photobioreactors with flow patterning for high‐density cultures of microalgae and carbon dioxide removal

A photobioreactor containing microalgae is a highly efficient system for converting carbon dioxide (CO₂) into biomass. Using a microalgal photobioreactor as a CO₂ mitigation system is a practical approach to the problem of CO₂ emission from waste gas. In this study, a marine microalga, Chlorella sp. NCTU‐2, was applied to assess biomass production and CO₂ removal. Three types of photobioreactors were designed and used: (i) without inner column (i.e. a bubble column), (ii) with a centric‐tube column and (iii) with a porous centric‐tube column. The specific growth rates (μ) of the batch cultures in the bubble column, the centric‐tube and the porous centric‐tube photobioreactor were 0.180, 0.226 and 0.252 day^?1, respectively. The porous centric‐tube photobioreactor, operated in semicontinuous culture mode with 10% CO₂ aeration, was evaluated. The results show that the maximum biomass productivity was 0.61 g/L when one fourth of the culture broth was recovered every 2 days. The CO₂ removal efficiency was also determined by measuring the influent and effluent loads at different aeration rates and cell densities of Chlorella sp. NCTU‐2. The results show that the CO₂ removal efficiency was related to biomass concentration and aeration rate. The maximum CO₂ removal efficiency of the Chlorella sp. NCTU‐2 culture was 63% when the biomass was maintained at 5.15 g/L concentration and 0.125 vvm aeration (volume gas per volume broth per min; 10% CO₂ in the aeration gas) in the porous centric‐tube photobioreactor.     

Algal nitrogen fixation in Californian streams: seasonal cycles

Using the acetylene reduction technique, nitrogen fixation was measured in Rocky Creek, a small seasonally dry Californian stream. In the 3 years since 1970 nitrogen fixation varied seasonally and spatially, being highest in the early stages of colony growth in shallow, clear regions where there was little shade. The annual rate of N₂-fixation was similar to that found recently for Arctic tundra, but was greater than rates for Antarctic rivulets and less than rates for temperate rocky shores. A hetero-cystous Nostoc appeared to be the sole organism responsible for this fixation. Nostoc occurred in variously sized gelatinous clumps on the stable boulders on which the co-dominant alga, Ulothrix zonata, also grew. Maximum nitrogenase activity yielded 70 nmoles C₂H₄ mg^-1 drywt day^-1 and 123 nmoles C₂H₄cm^-2 day^-1. Estimated annual amounts of nitrogen fixed in 1971 ranged from 42 in shade to 360 mg N fixed m^-2y^-1 in the most favourable areas. Nostoc biomass reached 33 g dry wt m^-2 and peeled off each year in June-July. Because Nostoc was confined to stable rocks the biomass per unit area of the stream as a whole was much less, ranging from 0.054 to 1.26 g dry wt m^-2 in the most favourable site. Nostoc was common throughout the length of Rocky Creek and also plentiful in eight out of ten adjacent streams but not in the main river (Eel River South Fork), probably due to high turbulence and turbidity. N₂-fixation in these streams makes a significant, but probably small contribution to the nitrogen income of the nitrogen-deficient Eel River system. Nostoc colony establishment appears to be controlled by a combination of reduced turbulence and firmer attachment of the colonies to the substrate. Nostoc colony disappearance in June is probably due to nutrient depletion.