Growth kinetics and fucoxanthin production of <Emphasis Type="Italic">Phaeodactylum tricornutum</Emphasis> and <Emphasis Type="Italic">Isochrysis galbana</Emphasis> cultures at different light and agitation conditions

Fucoxanthin is a carotenoid that exerts multiple beneficial effects on human health. However, reports comparing microalgae culture conditions and their effect on growth and fucoxanthin production are still limited. Isochrysis galbana and Phaeodactylum tricornutum cultures in different light (62.0, 25.9, 13.5, or 9.1 μmol photons m^-2 s^-1), mixing conditions (1 vvm aeration or 130 rpm agitation), and media compositions (F/2 and Conway medium) were studied for comparison of cellular growth and fucoxanthin production on F/2 medium. I. galbana showed a better adaptation to tested culture conditions in comparison with P. tricornutum, reaching 2.15?×?10⁷?±?4.07?×?10⁶ cells mL^-1 and a specific growth rate (μ) of 1.12?±?0.05 day^-1 under aerated conditions and 62.0 μmol photons m^-2 s^-1 light intensity. Fucoxanthin concentration was about 25 % higher in P. tricornutum cultures under 13.5 μmol photons m^-2 s^-1 light intensity and aerated conditions, but the highest fucoxanthin total production was higher in I. galbana, where 3.32 mg can be obtained from 1 L batch cultures at the 16th day under these conditions. Moreover, higher cell densities (~32.41 %), fucoxanthin concentration (~42.46 %), and total production (~50.68 %) were observed in I. galbana cultures grown in Conway medium, if compared with cultures grown in F/2 medium. The results show that the best growth conditions did not result in the best fucoxanthin production for either microalgae, implying that there is not a direct relationship between cellular growth and fucoxanthin production. Moreover, the results suggest that I. galbana cultures on Conway medium are strong candidates for fucoxanthin production, where 1.2 to 15 times higher fucoxanthin concentration are observed in comparison to macroalgal sources.     

Oil production from five marine microalgae for the production of biodiesel

Marine microalgae were studied as potential resources for the production of biodiesel. Five marine microalgae, Tetraselmis suecica, Phaeodactylum tricornutum, Chaetoceros calcitrans, Isochrysis galbana, and Nannochloropsis oculata were cultured in f/2 media, 12:12 L:D cycle at 20 ± 1°C with a light intensity of 36.3 μmol/m²/sec using a 15-L circular cylindrical photobioreactor. The dry cell weight, specific growth rate, biomass productivity, oil content and fatty acid composition of palmitic acid, stearic acid, oleic acid, linoleic acid, and linolenic acid of microalgae were determined. T. suecica, I. galbana, and N. oculata showed high dry cell weights of 0.58, 0.57, and 0.57 g/L, respectively. The culture period of T. suecica to reach the stationary phase was 9 days. On the other hand, N. oculata showed the longest culture period of 28 days to reach the stationary phase. T. suecica absorbed nitrate at the initial stages of cell growth, decreasing the nitrate concentration to 0.5 mg/L on day-7 of the culture. The highest oil contents were observed in P. tricornutum with a 25.31% dry cell weight and I. galbana with a 23.15% dry cell weight on day-9 after the stationary phase. I. galbana showed 417.33 mg of palmitic acid per g oil and T. suecica showed 235.61 mg of oleic acid per g oil. Stearic acid, linoleic acid, and linolenic acid did not exceed 30.02 mg/g oil in any of the microalgae. T. suecica showed the shortest culture period of 9 days to reach the stationary phase. Therefore, the highest biomass production of 0.58 g/L was obtained and I. galbana showed high biomass production of 0.57 g/ L and oil content of 23.15% of dry cell weight. Therefore, T. suecica and I. galbana can be selected as a potential candidate for the production of biodiesel.     

Under low irradiation,the light regime modifies growth and metabolite production in various species of microalgae

The effects of continuous light exposure (24L:0D) and a 12 h:12 h light/dark regime (12L:12D) were compared on the growth and carotenoid, protein, sugar, lipid, and fatty acid contents in Chlorella vulgaris, Nannochloropsis sp., Isochrysis galbana, and Dunaliella salina cultured in a batchwise facility. These microalgae were grown axenically under a low photon flux density (PFD) of 27 μmol photons m^?2 s^?1. C. vulgaris, Nannochloropsis sp., and I. galbana exhibited the highest cell densities when cultured under 24L:0D, whereas D. salina grew better under the alternating light/dark regime. I. galbana accumulated high levels of proteins, sugars, and lipids and exhibited the highest carotenoid content under 24L:0D. Protein production was enhanced in C. vulgaris under 24L:0D. The highest total lipid content was recorded for D. salina, reaching 74.6 % of total proteins, sugars, and lipids in cells at the stationary phase when grown under 12L:12D. The light/dark regime at low PFD was sufficient to stimulate the accumulation of monounsaturated and polyunsaturated fatty acids in all four algae. Their levels, like those of saturated fatty acids, did not differ significantly under the two light regimes. D. salina was an important source of tetradecenoic acid 14:1(n-5). Nannochloropsis sp. produced a large amount of the essential eicosapentaenoic acid, which reached 20 % of total fatty acids under 12L:12D, while I. galbana exhibited the highest level of docosahexaenoic acid, which reached 21 % under both light regimes. This study demonstrated the feasibility of culturing microalgae under low PFD in order to produce large quantities of valuable metabolites, especially various lipids with neutraceutical value.     

Growth inhibition of the eight species of microalgae by growth inhibitor from the culture of Isochrysis galbana and its isolation and identification

A type of growth inhibitor was successfully isolated and purified from cell-free filtrates of cultural medium at the death phase of Isochrysis galbana, and its chemical structure was confirmed by the methods of FABMS, UV, ¹H-NMR, ¹³C-NMR and 2D NMR, which was 1-[hydroxyl-diethyl malonate]-isopropyl dodecenoic acid, C₂₂H₃₈O₇. The results showed that the growth-inhibitor strongly inhibited the growth of Isochrysis galbana, and the growth of the eight species of microalgae (Dunaliella salina, Platymonas elliptica, Chlorella vugralis, Nitzschia closterium, Chaetoceros muelleri, Chaetoceros gracilis, Nitzschia closterium minutissima, Phaeodactylum tricornutum) also could be regulated by the growth-inhibitor in a concentration-dependent manner. The further investigation found that the synthesis process of chlorophyll and protein in the cells of all test microalgae could be inhibited by the growth inhibitor, and the content of chlorophyll and protein significantly decreased.     

Limitations on microalgal growth at very low photon fluence rates: the role of energy slippage

The lower limits of photosynthetically useable radiation at which growth and photosynthesis can occur establish the lower boundaries for the extent of photolithotrophy in the biosphere. Photolithotrophic growth denotes the capacity to grow with photons as the sole energy input. Slippage in terms of photosynthetic energy conversion implies a less than theoretical stoichiometry of energy-transduction process(es) such as the dissipation of intermediates of O₂ evolution and of ATP synthesis (H⁺/e⁻ and H⁺/ATP ratios). Slippage is particularly important in limiting the growth of photolithotrophic organisms at very low photon fluence rates. We found that Dunaliella tertiolecta and Phaeodactylum tricornutum avoid such reductions in photon use efficiency by increasing the size and number of their photosynthetic units, respectively, and by altering Q_A reduction kinetics on the reducing side of PS II. P. tricornutum is also less susceptible to slippage in terms of the breakdown of intermediates in its O₂ evolution pathway than D. tertiolecta. Minimizing H⁺ leakage through the CF₀–CF₁ ATP synthetase (and other H⁺ porters) is also discussed briefly. In combination, strategies employed by P.␣tricornutum effectively allow it to grow and photosynthesize at lower rates of energy input than D. tertiolecta, consistent with our observations. Differences in the responses of the photosynthetic apparatus of these two marine microalgae are mechanistic and probably representative of evolutionary divergences associated with strategies for dealing with environmental perturbations.     

Carbon dioxide utilisation of Dunaliella tertiolecta for carbon bio-mitigation in a semicontinuous photobioreactor

Bio-fixation of carbon dioxide (CO₂) by microalgae has been recognised as an attractive approach to offset anthropogenic emissions. Biological carbon mitigation is the process whereby autotrophic organisms, such as microalgae, convert CO₂ into organic carbon and O₂ through photosynthesis; this process through respiration produces biomass. In this study Dunaliella tertiolecta was cultivated in a semicontinuous culture to investigate the carbon mitigation rate of the system. The algae were produced in 1.2-L Roux bottles with a working volume of 1 L while semicontinuous production commenced on day 4 of cultivation when the carbon mitigation rate was found to be at a maximum for D. tertiolecta. The reduction in CO₂ between input and output gases was monitored to predict carbon fixation rates while biomass production and microalgal carbon content are used to calculate the actual carbon mitigation potential of D. tertiolecta. A renewal rate of 45 % of flask volume was utilised to maintain the culture in exponential growth with an average daily productivity of 0.07 g L^?1 day^?1. The results showed that 0.74 g L^?1 of biomass could be achieved after 7 days of semicontinuous production while a total carbon mitigation of 0.37 g L^?1 was achieved. This represented an increase of 0.18 g L^?1 in carbon mitigation rate compared to batch production of D. tertiolecta over the same cultivation period.     

Effect of oxygen supply on biomass and helvolic acid production in submerged fermentation of Cordyceps taii

The entomogenous fungus Cordyceps taii, a traditional Chinese medicinal mushroom, exhibits potent important pharmacological effects and it has great potential for health foods and medicine. In this work, the effects of oxygen supply on production of biomass and bioactive helvolic acid were studied in shake-flask fermentation of C. taii mycelia. The value of initial volumetric oxygen transfer coefficient (K_La) within 10.1–33.8 h⁻¹ affected the cell growth, helvolic acid production and expression levels of biosynthetic genes. The highest cell concentration of 17.2 g/L was obtained at 14.3 h⁻¹ of initial K_La. The highest helvolic acid production was 9.6 mg/L at 10.1 h⁻¹ of initial K_La. The expression levels of three genes encoding hydroxymethylglutaryl-CoA synthase, hydroxymethylglutaryl-CoA reductase and squalene synthase were down-regulated on day 2 and day 8 but up-regulated on day 14 at an initial K_La value of 10.1 h⁻¹ vs. 33.8 h⁻¹, which well corresponded to the helvolic acid biosynthesis in those conditions. The information obtained would be helpful for improving the biomass and helvolic acid production in large-scale fermentation of C. taii.     

Protein turnover in relation to maintenance metabolism at low photon flux in two marine microalgae

Acclimation to very low photon fluxes involves adjusting a suite of physiological characteristics that collectively elicit a physiological response. Facilitating such changes is pro‐tein turnover. Dunaliella tertiolecta (Butcher) and Phaeodactylum tricornutum (Bohlin) were grown in turbidostats at a range of photon fluxes between 2 and 300 µmol photons m^?2 s^?1. The kinetics of pulse‐chase labelling of the protein with ³H showed that (1) two protein pools were present, one of which turned‐over rapidly (hours), and a second which turned over more slowly (days); and (2) protein turnover rates were slower in P. tricornutum than in D. tertiolecta. Phaeodactylum tricornutum had a lower maintenance coefficient for protein turnover than D. tertiolecta, and correspondingly a smaller proportion of its respiratory demands (30%) were associated with protein turnover than in D. tertiolecta (36%). There appears to be a correlation between lower metabolic activity, requiring lower protein concentrations, and an associated decreased cost of maintenance processes in P. tricornutum compared to D. tertiolecta. Differences between protein turnover rates and maintenance metabolic costs may be one of the photo‐acclimation strategies that determine which photon niches microalgae can successfully exploit.     

Optimisation and scale-up of α-glucuronidase production by recombinant Saccharomyces cerevisiae in aerobic fed-batch culture with constant growth rate

α-Glucuronidase (EC 3.2.1.139) of family GH 115 from Scheffersomyces stipitis is a valuable enzyme for the modification of water-soluble xylan into insoluble biopolymers, due to its unique ability to act on polymeric xylans. The influence of growth rate on the production of α-glucuronidase by recombinant Saccharomyces cerevisiae MH1000pbk10D-glu in glucose-limited fed-batch culture was studied at 14 and 100 L scale. At and below the critical specific growth rate (μ_crit) of 0.12 h⁻¹ at 14 L scale, the biomass yield coefficient (Y_x/s) remained constant at 0.4 g g⁻¹ with no ethanol production, whereas ethanol yields relative to biomass (k_eth/x) of up to 0.54 g g⁻¹ and a steady decrease in Y_x/s were observed at μ > 0.12 h⁻¹. Production of α-glucuronidase was growth associated at a product yield (k_α-glu/x) of 0.45 mg g⁻¹, with the highest biomass (37.35 g/L) and α-glucuronidase (14.03 mg/L) concentrations, were recorded during fed-batch culture at or near to μ_crit. Scale-up with constant k_La from 14 to 100 L resulted in ethanol concentrations of up to 2.5 g/L at μ = 0.12 h⁻¹. At this scale, α-glucuronidase yield could be maximised at growth rates below μ_crit, to prevent localised high glucose concentration pockets at the feed entry zone that would induce oxido-reductive metabolism. This is the first report where recombinant production of α-glucuronidase (EC 3.2.1.139) by S. cerevisiae was optimised for application at pilot scale.     

Ingestion rates and dietary lipids affect growth and fatty acid composition of Dendraster excentricus larvae

This study investigated the effect of single and mixed algal diets on growth, fatty acid composition and ingestion rates for Dendraster excentricus larvae. Larvae were assigned to three single algal diet treatments Isochrysis galbana, Dunaliella tertiolecta or Rhodomonas sp. and four mixed algal diet treatments D. tertiolecta and Rhodomonas, I. galbana and D. tertiolecta, I. galbana and Rhodomonas, D. tertiolecta, Isochrysis galbana and Rhodomonas sp. Small amounts (0.36-0.6%) of stearic acid (18:0) were seen in the three algae used but a relatively large percentage (7-25%) of this SAFA was found in Dendraster larvae. The alga D. tertiolecta had the highest percentage (51.7%) of the short chain polyunsaturated fatty acid (PUFA) linolenic acid 18:3(n-3) and trace amounts (0.02-0.14%) of the long chain PUFAs eicosapentanoic (EPA, 20:5(n-3) and docosahexanoic acids (DHA, 22:6(n-3)). However, sand dollar larvae demonstrated the ability to elongate and desaturate shorter chain (18 carbon) polyunsaturated fatty acids (PUFA) to longer chain (20 carbon) n-3 PUFA. Thus high levels of 18:3(n-3) in D. tertiolecta led to high levels of EPA and low levels of 18:3(n-3) in Dendraster larvae fed this diet. Rhodomonas sp. had the highest percentage of stearidonic acid (18:4(n-3), 38.14%) and EPA (10.6%). Despite high levels of 18:4(n-3) in Rhodomonas sp. this acid was absent or found at very low levels in larvae fed this alga, or any combination of this alga. I. galbana had the highest percentage of DHA(14.3%) but was almost devoid of EPA (0.43%). Although Rhodomonas sp. and I. galbana had high levels of EPA and DHA sand dollar larvae did not incorporate higher levels of these long chain PUFAs into their lipids compared to those fed the alga D. tertiolecta. Dendraster larvae synthesized a number of 20 and 22 carbon non-methylene interrupted dienes (NMID), with levels increasing with larval stage. Higher ingestion rates were observed for Dendraster larvae fed single algal diets (Rhodomonas sp. or D. tertiolecta) and lower ingestion rates for those fed mixed algal diets. The highest ingestion rates were for 8-arm Dendraster larvae fed the large alga Rhodomonas sp. presented as a single algal diet. When fed a combination of three algae, selection of particles varied slightly depending on stage with 8-arm larvae ingesting slightly more of the larger algal cell in the mixture than 6-arm larvae. The present study suggests that regardless of the ratios given larvae might have an optimum ratio of different sized particles at which they can feed. The mixed algal diet of I. galbana and D. tertiolecta was the best algal diet leading to significantly larger larvae with high survival and development to metamorphosis. The single algal diets of Rhodomonas sp. or Dunaliella tertiolecta were the second and third best algal diets based on growth and survival to metamorphosis.     

The effect of cadmium and lead on the growth of two species of marine phytoplankton with particular reference to the development of tolerance

Decreases in salinity (<10%) increased the growth rates ofPhaeodactylum tricornutum and Dunaliella tertiolecta. Increasinglevels of cadmium (1–50 ppm (mg 1^–1)) reduced thegrowth rates of both species. 100 ppm cadmium was lethal toD. tertiolecta but not to P. tricornutum. Lead (1 –4 ppm)initially increased the growth rate of D. tertiolecta but thencaused all but the 1 ppm culture to die. Lead (1–4 ppm)caused a decrease in growth rate of P. tricornutum. After exposureto 1 ppm cadmium, cultures of D. tertiolecta showed an increasedtolerance to levels of cadmium, and a changed response to levelsof lead. Exposure of P. tricornutum to either cadmium or lead,or exposure of D. tertiolecta to lead caused no change in responseto either metal.     

Unmarked insertional inactivation in the gfo gene improves growth and ethanol production by Zymomonas mobilis ZM4 in sucrose without formation of sorbitol as a by-product,but yields opposite effects in high glucose

Sorbitol, one of the main by-products of growth on high sucrose concentrations, is catalyzed by glucose-fructose oxidoreductase (GFOR, EC 1.1.99.28) in Zymomonas mobilis, which decreases the ethanol yield. In this study, an unmarked gfo mutant from Z. mobilis ZM4 was constructed using a site-specific FLP recombinase, and growth and ethanol production were evaluated with or without the addition of sorbitol to the media. The inactivation of gfo had contrasting effects in different substrates, especially at high concentrations. The maximum specific growth rate (μ_m) and theoretical ethanol yield value (Y_m) increased from 0.065 h⁻¹ and 60.56% to 0.094 h⁻¹ and 83.87% in 342 g/L sucrose, respectively. Conversely, in 200 g/L glucose, gfo inactivation decreased μ_m and Y_m from 0.15 h⁻¹ and 89.85% to 0.10 h⁻¹ and 67.59%, respectively, and prolonged the lag period from 16 h to 40 h. The addition of sorbitol slightly accelerated growth and sucrose hydrolysis by the gfo mutant in 342 g/L sucrose; however, addition of sorbitol restored the μ_m and Y_m of the gfo mutant in 200 g/L glucose to 0.14 h⁻¹ and 82.50%, respectively. Inactivation of gfo had a small effect on fructose utilization, and a positive one on mixture of glucose and fructose similar to that on sucrose. These results provide further understanding of the osmoregulation mechanisms in Z. mobilis and may help to exploit the biotechnological applications of this industrially important bacterium.     

The optimal growth conditions for the biomass production of Isochrysis galbana and the effects that phosphorus, Zn2+, CO2, and light intensity have on the biochemical composition of Isochrysis galbana and the activity of extracellular CA

The effects of phosphorus, Zn²⁺, CO₂, and light intensity on growth, biochemical composition, and the activity of extracellular carbonic anhydrase (CA) in Isochrysis galbana were investigated. A significant change was observed when the concentration of phosphorus in the medium was increased from 5 μmol/L to 1000 μmol/L affecting I. galbana’s cell density, biochemical composition, and the activity of extracellular CA. Phosphorous concentration of 50 μmol/L to 500 μmol/L was optimal for this microalgae. The Zn²⁺ concentration at 10 μmol/L was essential to maintain optimal growth of the cells, but a higher concentration of Zn²⁺ (≥ 1000 μmol/L) inhibited the growth of I. galbana. High CO₂ concentrations (43.75 mL/L) significantly increased the cell densities compared to low CO₂ concentrations (0.35 mL/L). However, the activity of extracellular CA decreased significantly with an increasing concentration of CO₂. The activity of extracellular CA at a CO₂ concentration of 43.75 mL/L was approximately 1/6 of the activity when the CO₂ concentration was at 0.35 mL/L CO₂. Light intensity from 4.0 mW/cm² to 5.6 mW/cm² was beneficial for the growth, biochemical composition and the activity of extracellular CA. The lower and higher light intensity was restrictive for growth and changed its biochemical composition and the activity of extracellular CA. These results indicate that phosphorus, Zn²⁺, CO₂, and light intensity are important factors that impact growth, biochemical composition and the activity of extracellular CA in I. galbana.     

Continuous high-ethanol fermentation from cane molasses by a flocculating yeast

Repeated-batch fermentation by a flocculating fusant, Saccharomyces cerevisiae HA 2, was done in a molasses medium that contained 20% (w/v) total sugar, at 30°C in an automatically controlled fermentor, and the effects of ethanol concentration on the specific growth rate and the specific production rate of ethanol were studied. Both the specific growth rate and the specific production rate of ethanol fell with increase of ethanol concentration, and there was a linear correlation between each rate and the concentration of thanol. The maximum specific growth rate (μ_max) and the maximum specific production rate of ethanol (q_max) were 0.12 h⁻¹ and 0.1 g ethanol/10⁹ cells·h, respectively. The specific growth rate and the specific production rate of ethanol fell to zero at ethanol concentration of 89 g/l and 95 g/l, respectively. The number of viable cells, calculated from the linear inhibition equation, was 1.3 × 10⁹ cells/ml for production of 85 g/l ethanol at a dilution rate (D₁) of 0.2 h⁻¹. Based on this estimation, a laboratory-scale continuous fermentation, using two fermentors in series, was done. In the second fermentor, 85 g/l ethanol was produced at a dilution rate (D₁) of 0.2 h⁻¹ by the active feedig of the fermented mash from the first fermentor into the second fermentor by pumping (hereafter called active feeding). To maintain the number of viable cells above 10⁹ cells/ml in the second fermentor, a active feeding ratio of more than 23% was required. Under these conditions, 81 g/l ethanol was produced in the second fermentor at a dilution rate (D_t) of 0.25 h⁻¹, and the high ethanol productivity of 20.3 g/l·h could be achieved. A bench-scale continuous fermentation, using two fermentors in series, with a active feeding ratio of 25% was done. An ethanol concentration of 84 g/l in the second fermentor at a dilution rate (D_t) of 0.25 h⁻¹ was achieved, just as it was in the laboratory-scale fermentation test.     

Fatty acid composition and biological activities of Isochrysis galbana T-ISO,Tetraselmis sp. and Scenedesmus sp.: possible application in the pharmaceutical and functional food industries

Organic and water extracts of Isochrysis galbana T-ISO (=Tisochrysis lutea), Tetraselmis sp. and Scenedesmus sp. were evaluated for their antioxidant activity, acetylcholinesterase (AChE) inhibition, cytotoxicity against tumour cell lines, and fatty acids and total phenolic content (TPC). I. galbana T-ISO had the highest TPC (3.18 mg GAE g^?1) and radical scavenging activity, with an IC₅₀ value of 1.9 mg mL^?1 on the acetone extract. The extracts exhibited a higher ability to chelate Fe²⁺ than Cu²⁺, and the maximum Fe²⁺ chelating capacity was observed in the hexane extract of Scenedesmus sp. (IC₅₀=0.73 mg mL^?1) and Scenedesmus sp. (IC₅₀?=?0.73 mg mL^?1). The highest ability to inhibit AChE was observed in the water and ether extracts of Scenedesmus sp., with IC₅₀ values of 0.11 and 0.15 mg mL^?1, respectively, and in the water extract of I. galbana (IC₅₀?=?0.16 mg mL^?1). The acetone extract of I. galbana T-ISO significantly reduced the viability of human hepatic carcinoma HepG2 cells (IC₅₀?=?81.3 μg mL^?1) as compared to the non-tumour murine stromal S17 cell line, and displayed a selectivity index of 3.1 at the highest concentration tested (125 μg mL^?1). All species presented a highly unsaturated fatty acids profile. Results suggest that these microalgae, particularly I. galbana T-ISO, could be a source of biomolecules for the pharmaceutical industry and the production of functional food ingredients and can be considered as an advantageous alternative to several currently produced microalgae.     

Optimization of Citric Acid Production from Glucose by Yarrowia lipolytica

Citric acid (CA) is mainly produced in a biotechnological process using Aspergillus niger. In this process, large amounts of wastes have to be removed. Since the use of Yarrowia lipolytica for CA production is an environmental compatible alternative method, the CA production was optimized in regard to growth temperature and pH as well as substrate and product inhibition. The highest value of the maximum specific growth rate at pH 6.5 was found to be μ_max = 0.192 h^–1, whereas the largest amount of CA of 24.91 g/L as well as the highest selectivity of the bioprocess (89.9 % CA) and the maximum yield (0.22 g_CA/g_Glucose) were obtained at pH 6.0. During the growth phase, the temperature optimum was found to be in the range of 30–34 °C (μ_max = 0.132 h^–1). Nevertheless, the highest concentration of CA during the production phase was obtained at 30 °C (41 g/L CA, 93.1 % CA, 0.55 g_CA/g_glucose). In studying the substrate inhibition of the process, a clear tendency of decrease in the maximum specific growth rate was detected when the initial glucose concentration was increased from 50 g/L (μ_max = 0.17 h^–1) to 200 g/L (μ_max = 0.055 h^–1). The addition of 120 g/L CA to the culture broth at the start of the production phase reduced the production of CA from 32.1 g/L to 7.4 g/L.     

Changes in growth and composition of the marine microalgae <Emphasis Type="Italic">Phaeodactylum tricornutum</Emphasis> and <Emphasis Type="Italic">Nannochloropsis salina</Emphasis> in response to changing sodium bicarbonate concentrations

Oils, carbohydrates, and fats generated by microalgae are being refined in an effort to produce biofuels. The research presented here examines two marine microalgae, Nannochloropsis salina (green alga) and Phaeodactylum tricornutum (diatom), when grown with 0 (no addition), 0.5, 1.0, 2.0, and 5.0 g L^?1 NaHCO₃ added to an f/2 medium during the growth phase (GP) and a nutrient induced (nitrate limitation) lipid formation phase (LP). We hypothesize that the addition of NaHCO₃ is a sustainable and practical strategy to increase cellular density and concentrations of lipids in microalgae as well as the rate of lipid accumulation. In N. salina, final cell densities were significantly (p?<?0.05) higher in the NaHCO₃-treated cells than the control while in P. tricornutum the cell densities were higher with >[NaHCO₃] during the GP. During the LP, cell densities were generally higher in the NaHCO₃-treated cells compared with controls. F _V/F _M (efficiency of photosystem II) patterns paralleled those for cell density with generally higher values with higher concentrations of NaHCO₃ and significantly different values between controls and 5.0 g L^?1 NaHCO₃ at the end of the GP (p?<?0.05). F _V/F _M was variable between treatments in P. tricornutum (0.3–0.65) but less so in N. salina for (0.5–0.7) regardless of [NaHCO₃]. The lipid index (measured with Nile red), used as a proxy for triacylglycerides (TAGs), was 10.2?±?6.5 and 4.4?±?2.9 (fluorescence units/OD cells ×1000) for N. salina and P. tricornutum, respectively, at the end of the GP. At the end of the LP, the lipid index was eight and four times higher than during the GP in the corresponding 5.0 g L^?1 NaHCO₃ treatments, revealing that N. salina was accumulating more lipid than P. tricornutum. Dry weights essentially doubled during LP compared with GP for N. salina; this was not the case for P. tricornutum. In general, the percentage of ash in dry weights was significantly higher in the LP relative to the corresponding GP treatments for P. tricornutum; this was not the case for N. salina. During the LP, there was also less soluble protein in N. salina compared to GP; differences were not significant in cells growing with 2.0 or 5.0 g L^?1 NaHCO₃. In P. tricornutum, faster growing cells had more soluble protein during the GP and LP; differences between treatments were significant. P. tricornutum generally accumulated significantly more crude protein than N. salina at higher [NaHCO₃]; there was three times more crude protein in the highest NaHCO₃ (5.0 g L^?1) treatment compared with the controls. C:N ratios (mol:mol) were similar across treatments during GP: 7.03?±?0.12 and 10.16?±?0.41 for N. salina and P. tricornutum, respectively. Further, C:N ratios increased with increasing [NaHCO₃] during LP. Species-specific fatty acid methyl ester (FAMEs) profiles were observed. While C16:0 was lower in P. tricornutum compared to N. salina, the diatom produced more C16:1 and C14 but not C18:3. Monounsaturated fatty acids (MUFA) significantly increased in N. salina in the LP compared to GP and in response to increasing [NaHCO₃] (t tests; p?<?0.05). Saturated fatty acids (SFA) responded similarly but to a lesser degree. There were more polyunsaturated fatty acids (PUFA) in N. salina than MUFAs or SFAs. In P. tricornutum, there were generally more SFAs, MUFAs and PUFAs in P. tricornutum during LP than GP in the corresponding NaHCO₃ treatments. These findings reveal the importance of considering NaHCO₃ as a supplemental carbon source in the culturing marine phytoplankton in large-scale production for biofuels.     

A comparative study of the use of inorganic carbon resources by Chara aspera and Potamogeton pectinatus

We studied the potential role of dissolved inorganic carbon (DIC) in determining vegetation dominance of Potamogeton pectinatus L. and Chara aspera Deth. ex Willd. by monitoring the seasonal dynamics of DIC in a shallow lake and comparing the use of DIC of the two species. The HCO₃⁻-concentration in summer dropped from 2.5 to <0.5 mM with seasonally increasing Chara biomass, whereas outside the vegetation concentrations remained at 2.5 mM. Inside Potamogeton spp. vegetation DIC decreased from 2.5 to ca. 0.75 mM HCO₃⁻. A growth experiment showed ash-free biomass for P. pectinatus was nearly two times as high as for C. aspera at 3 mM HCO₃⁻, but almost two times lower at 0.5 mM than at 3.0. In a separate experiment, P. pectinatus precultured at a relatively low HCO₃⁻-level had a lower net photosynthetic rate (P_max, 0.1 mmol O₂ g⁻¹ DW h⁻¹) than C. aspera (P_max, 0.1 mmol O₂ g⁻¹ DW h⁻¹) over the range of HCO₃⁻-concentrations tested (P_max, 0.14 mmol O₂ g⁻¹ DW h⁻¹). In response to CO₂ no significant differences between the compensation points (P. pectinatus, 28 mM; C. aspera 66 mM), were observed, but the photosynthetic rate increased faster than for C. aspera than for P. pectinatus. Under field conditions, the use of CO₂ is not important since inside vegetation CO₂-concentrations were below 10 μM, and thus, not available for photosynthesis of either species during the main part of the growth season. It is suggested that C. aspera may be a better competitor for HCO₃⁻ than P. pectinatus in conditions with a low HCO₃⁻ supply. As HCO₃⁻ is a strong limiting factor for growth inside the vegetation and probably the only carbon source available, the superior ability of C. aspera to use HCO₃⁻ may be an important factor explaining its present dominance in Veluwemeer.     

The effect of glycerol mixed substrate on the heterologous production of a Rhizopus oryzae lipase in Pichia pastoris system

A recombinant Rhizopus oryzae lipase producing Mut^s Pichia pastoris strain was used as a model organism to study the effect of mixed substrates (glycerol and methanol) on the specific product productivity. Different fed-batch cultivations were performed under three constant specific growth rates (0.02, 0.05 and 0.1 h⁻¹), maintaining a constant methanol concentration of 2 g l⁻¹.At the lowest μ tested (0.02 h⁻¹), the specific productivity was 1.23 and 1.61 fold higher and the specific methanol consumption rate (q_sMeOH) was 3 and 3.5 fold higher than values obtained when μ was 0.05 and 0.1 h⁻¹, respectively. This implies a relation between the q_sMeOH and the specific productivity, yielding higher specific productivities whenever the consumption of methanol is higher. Although glycerol was maintained under limiting conditions in all μ tested, when the relation between the μ_Gly and μ_MeOH was larger than 4, an important decrease on the maximal activity value was observed.Finally, a comparison under the same conditions using glycerol or sorbitol as co-substrates was also performed, obtaining better specific productivity when sorbitol was used. In addition, protease activity was detected when glycerol was used as co-substrate.     

Kinetic model for growth of Phaeodactylum tricornutum in intensive culture photobioreactor

A kinetic model has been developed to estimate the specific growth rate of Phaeodactylum tricornutum in batch cultures. The cultures were carried out in a laboratory scale photobioreactor. Some factors like pH, temperature and irradiance were studied. In the first case, an optimum pH of 7.8 and a specific growth rate of 0.064 h⁻¹ were achieved for certain nitrate conditions and illumination. The temperature influence has been modelled by a modified Sinclair model. The optimum temperature was achieved at 20.4 °C in aerated cultures and at 22.3 °C in non-aerated cultures. Better adaptation to low temperatures than high ones has been obtained. The experiments carried out with different irradiances drive to a simple Monod's equation for the irradiance influence on growth, with semi-saturation irradiance of 10.2 μEinstein⁻² s⁻¹ in aerated cultures and of 6.8 μEinstein m⁻² s⁻¹ in non-aerated cultures.