Buenaventura Cabezas

Growth of the marine microalga Tetraselmis suecica in batch cultures with different salinities and nutrient concentrations

Abstract The marine microalga Tetraselmis suecica is known for its ability to tolerate a wide range of salt concentrations. Cultures were grown under 48 different nutrient concentration-salinity conditions, ranging from 2 to 64 mM NaNO3 and from 0 to 35‰ S. Salinity was more important for the growth rate of the microalgae when it was related to the nutrient concentration in the culture medium. Optimal growth conditions were between 25 and 35‰ salinity and nutrient concentrations of 2, 4 and 8 mM of NaNO3, resulting in 0.55 doublings/day and a maximum cellular density of 1.3 × 106 cells/ml. Variations in salinity and in nutrient concentration had a greater effect on the final biomass than on the growth velocity. The total protein of the culture and protein per cell increased when the salinity increased for a given nutrient concentration. The total protein of the cultures decreased when the nutrient concentration increased for a given salinity. Protein per cell decreased with increasing salinity up to 20‰ but from this point of the process was reversed. The nitrate-protein transformation rate increased with the salinity and decreased with increasing nutrient concentrations. The maximum rate was 64%.

Mass culture and biochemical variability of the marine microalga Tetraselmis suecica Kylin (Butch) with high nutrient concentrations

Abstract Mass cultures of Tetraselmis suecica were carried out with four nutrient concentrations, ranging from 2 to 16 mM of NaNO3 and salinity 35‰. An air flow of 15 l/min maintained a CO2 transference rate sufficient to keep the pH below 8.4. Using these cultural conditions equations were calculated, by a multiple non-linear least squares regression of order four, enabling predictions to be made of growth kinetics and chemical composition. Maximum cellular densities of 7.83 × 106 and 7.15 × 106 cells/ml were obtained with 8 and 16 mM of NaNO3, respectively. Growth velocity ranged between 0.53 and 0.63 doublings (dbl)/day, although 0.98 dbl/day were reached with 16 mM of NaNO3. Volume increased with nutrient concentration from 252 to 905 μm3. Protein content reached maximum values of 306 μg/ml or 59.8 pg/cell. In the logarithmic phase, protein was regulated by nutrient concentration and decreased according to this concentration. Maximum efficiency of transformation from nitrate to protein was 108%, obtained at 2 mM of NaNO3. Efficiency decreased, to 14%, when nutrient concentration increased. This fact indicates that the lowest cost of harvesting is obtained with a nutrient concentration of 2 mM NaNO3. Chlorophyll a cell reached values between 3.1 and 3.8 pg/cell in the stationary phase. There was a relationship between nutrient concentration and chlorophyll a cell in the logarithmic phase, with an increase from 2.15 pg/cell to 3.74 pg/cell. Changes in chlorophyll a level are related to nitrogen depletion. Carbohydrate/cell was constant at values of 19.84–28.68 pg/cell in the logarithmic and stationary phases and was not related to nitrogen depletion. RNA/cell ranged from 4.17 to 5.48 pg/cell, except at 2 mM of NaNO3 when it was 2.77 pg/cell, probably due to nitrogen depletion. The level of DNA/cell was constant in all the nutrient concentrations assayed and ranged from 0.1 to 1.09 pg/cell. Great variability in the chemical composition of T. suecica has been shown. Growth in mass cultures is closely coupled to changes in nutrient concentrations and variations occur in protein, chlorophyll a and RNA content, showing differences of 215%, 190% and 203%, respectively, in the stationary phase. This biochemical variability, mainly in protein content, must have a marked effect on the nutritive value of this microalga as feed in mariculture.

Biomass production and biochemical composition in mass cultures of the marine microalga Isochrysis galbana Parke at varying nutrient concentrations

Abstract Mass cultures of Isochrysis galbana were carried out with four nutrient concentrations ranging from 2 to 16 m M of NaNO 3 and salinity 35‰. An air flow of 15 l/min maintained a CO 2 transference rate sufficient to keep the pH below 8.4. Using these conditions, equations were calculated by a multiple non-linear least squares regression of order four, enabling predictions to be made of growth kinetics and chemical composition. Maximum cellular density of 65.5 × 10 6 cells/ml was obtained with 4 m M NaNO 3 . Cellular volume was constant in the different nutrient concentrations. Protein content reached a maximum value of 374 μg/ml at 4 m M of NaNO 3 , and this concentration also presented the maximum efficiency of transformation from nitrate to protein, i.e. 114%. As a result, lowest costs for harvesting are obtained at a nutrient concentration of 4 m M NaNO 3 . Efficiencies decreased to 15% as nutrient concentration increased. Maximum values of chlorophyll a (21.9 μg/ml) and carbohydrates (213 μg/ml) were also obtained with 4 m M NaNO 3 . In the logarithmic phase, the contents of protein, chlorophyll a , carbohydrates, RNA and DNA per cell were constant. Chlorophyll a reached values between 0.15 and 0.33 pg/cell in the stationary phase. Carbohydrate levels reached the maximum value of 3.16 pg/cell with 4 m M NaNO 3 in the stationary phase. The levels of RNA/cell and DNA/cell were constant in all the nutrient concentrations tested and in both growth phases, and ranged from 1.15 to 1.71 pg/cell for RNA and from 0.006 to 0.014 pg/cell for DNA. Growth in mass cultures is closely coupled to changes in nutrient concentrations and variations occur in protein, chlorophyll a and carbohydrate contents, showing differences of 177%, 220% and 136%, respectively, in the stationary phase. This biochemical variability, mainly in protein content, must have a marked effect on the nutritive value of this microalga as a feed in mariculture.

Growth, chlorophyll a and protein of the marine microalga Isochrysis galbana in batch cultures with different salinities and high nutrient concentrations

Abstract Cultures of the marine microalga Isochrysis galbana were grown under 56 different nutrient concentration-salinity conditions, ranging from 1 to 64 mM NaNO3 and from 0 to 35‰ salinity. Salinity and nutrient concentration were found to be closely related to I. galbana growth and to the biochemical composition. Optimal growth conditions were between 15 and 35‰ salinity and nutrient concentrations of 2, 4 and 8 mM NaNO3, resulting in one doubling/day and a maximum cellular density of 20 × 106 cells/ml. Variations in salinity and in nutrient concentration had a greater effect on the final biomass than on the growth velocity. Maximum values of chlorophyll a ml were found with 2, 4 and 8 mM NaNO3 and between 15 and 35‰ salinity. Chlorophyll a cell values were more homogeneously distributed between 15 and 35‰ salinity and 1 to 8 mM NaNO3, although maximum concentrations (37 pg chlorophyll a cell ) were reached at 10–15‰ with all the nutrient concentrations. Protein per ml of culture and protein per cell were closely related to salinity and nutrient concentration. Maximum values of 387 μg/ml and 18.6 pg/cell were obtained at 15–35‰ salinity and 4–8 mM NaNO3. The nitrate-protein transformation rate was related to nutrient concentration. Maximum rate was 84% at 15‰ salinity and 1 mM NaNO3. Nutrient concentrations higher than 16 mM NaNO3 produced a strong decrease in the efficiency at all salinities.

Biomass production and biochemical variability of the marine microalga Dunaliella tertiolecta (Butcher) with high nutrient concentrations

Abstract Mass cultures of Dunaliella tertiolecta were carried out in 10-l flasks with four nutrient concentrations in order to obtain a maximum biomass production and to find out its biochemical variability. Using these cultural conditions equations were calculated by a multiple non-linear least squares regression of order four, enabling predictions to be made of growth kinetics and chemical composition. Maximum cellular densities between 12.45 × 10 6 and 14.23 × 10 6 cells/ml were obtained with 4, 8 and 16 m M of NaNO 3 . Growth velocity ranged between 0.50 and 0.61 doublings/day. Protein content reached maximum values in the stationary phase of 442 μg/ml and 31 pg/cell at 16 m M of NaNO 3 . In the logarithmic phase protein concentration per cell was not related to nutrient concentration. Maximum efficiencies of transformation from nitrate to protein were 100%, obtained at 2 and 4 m M of NaNO 3 . Chlorophyll a/cell reached values between 1.03 and 1.95 pg/cell in the stationary phase. There was no relationship between nutrient concentration and chlorophyll a/cell in the logarithmic phase, the values for the different nutrient concentrations being very similar. The maximum value of carbohydrates, 185 μg/ml, was obtained with 4 m M of NaNO 3 . Carbohydrates/cell reached the maximum values of 8.94 and 10.05 pg/cell with 8 and 16 m M of NaNO 3 , respectively, in the stationary phase. RNA/cell ranged from 4.28 to 5.40 pg/cell in the logarithmic phase and from 4.72 to 5.80 pg/cell in the stationary. The level of DNA/cell was constant in all the nutrient concentrations tested and in both growth phases, and ranged from 0.05 to 0.12 pg/cell. Great variability in the chemical composition of D. tertiolecta has been shown. Growth in mass cultures is closely coupled to changes in nutrient concentration, and variations occur in protein, chlorophyll a , carbohydrates and RNA content, showing differences of 197%, 255%, 142% and 150%, respectively. This biochemical variability must have a marked effect on the value of this microalga as source of single cell protein, chemicals or as feed in mariculture.

Tris not only controls the pH in microalgal cultures, but also feeds bacteria

Tris (Tris(hydroxymethyl)amino methane), a compound often used as a buffer in microalgal culture media, sustains active bacterial growth in non-axenic microalgal cultures when sodium phosphate is present. The low pH levels caused by bacterial growth and probably the depletion of phosphorus in the medium caused the collapse ofPhaeodactylum tricornutum cultures resulting in a reduction of microalgal growth from 32 x 106 to 1.1 x 106 cells ml−1. This emphasizes the need for care when interpreting the results of non-axenic microalgae cultures in which Tris or other organic buffer is added.

Changes in protein, carbohydrates and gross energy in the marine microalga Dunaliella tertiolecta (Butcher) by nitrogen concentrations as nitrate, nitrite and urea

Cultures of the marine microalga Dunaliella tertiolecta were grown in nitrate, nitrite and urea at concentrations ranging from 0·25 to 16 mg atom. N/litre. Great biochemical variability has been shown in this microalga as a function of high nitrogen concentrations for all the sources used. Cellular protein and carbohydrates and gross energy per ml of culture increased proportionally to the increase in the N concentration, under conditions that maintain constant the NP ratio. Two kinds of cultures are defined: low nitrogen cultures 2 mg atom. N/litre. Variability mainly appears in the second type of cultures. Protein/cell values of up to 4·94, 5·47 and 1·41 times higher have been observed in nitrate, nitrite and urea cultures, respectively, when comparing protein/cell values obtained in high N cultures with those obtained in low N cultures. Similar variations have been observed in the carbohydrates/cell content, with values up to 3·16, 3·30 and 1·77 times higher in the high than in the low N cultures. Biochemical variability is greater in nitrate and nitrite cultures (inorganic sources of nitrogen) than in urea cultures (organic source of N). Lipid/carbohydrates ratio seems to be a convenient parameter for characterizing the physiological state of a microalgal population. This biochemical variability must have a marked effect on the value of this microalga as a source of single cell protein, chemicals or as feed in mariculture.

Approach to biomass production of the marine microalga Tetraselmis suecica (kylin) butch using common garden fertilizer and soil extract as cheap nutrient supply in batch cultures

Abstract We report the possibility of an economic cultivation of the marine microalga Tetraselmis suecica, using different mixtures of a common garden fertilizer, soil extract, micronutrients and vitamins. Maximum cellular densities were obtained with a nitrogen concentration of 14 μg/ml and maximum protein concentrations were obtained with 28 μg N/ml, in all cases. The mixture of fertilizer + soil extract + micronutrients gave the best results for obtaining maximum cellular densities and protein concentrations per ml of culture. An economic evaluation showed a maximum of 2770 g of microalgal protein/dollar and 210 × 1012 microalgal cells/dollar with the mixture of the fertilizer and soil extract.

Changes in the gross chemical composition of mass cultures of the marine microalga Dunaliella tertiolecta with different aeration rates

The effect of different aeration rates, in the range 0–6·51 l of air min−1 l of culture−1, and CO2 supply on the biochemical composition of mass cultures of the marine microalga Dunaliella tertiolecta was studied. The biochemical composition of D. tertiolecta was strongly affected by the aeration rate. There was a negative correlation between stationary-phase protein cellular content and air flow. Carbohydrate cellular content also decreased with aeration rate, a minimum being reached with 0·93 l of air min−1 l of culture−1. Maximum carbohydrate per volume unit was achieved with maximum aeration as the increase of carbohydrates per ml was directly proportional to air flow and therefore to CO2 availability. Maximum protein per ml was achieved with 1·86 l of air min−1 l of culture−1, keeping stable with higher air flows. The cultures supplied with CO2 showed carbohydrate and protein concentrations similar to the cultures with 1·86 l of air min−1 l of culture−1, indicating a correlation between available CO2 and not only carbon, but also nitrogen metabolism. Different factors seemed to limit cell division and nitrogen metabolism as maximum nitrogen transformation rate was achieved with an air flow of 1·86 l of air min−1 l of culture−1, lower than the 3·72 l of air min−1 l of culture−1 needed for maximum cell density.

Response of the Marine Microalga Dunaliella tertiolecta to Nutrient Concentration and Salinity Variations in Batch Cultures

Summary The marine microalga Dunaliella tertiolecta is known for its ability to tolerate a wide range of salt concentrations. Cultures were grown under 56 different nutrient concentration-salinity conditions. Optimal growth conditions were between 25 and 35 ‰ salinity and with nutrient concentrations between 8 and 32 times higher than the standard concentrations, resulting in maximum cellular densities between 8.41 x 10 6 and 16.74x 10 6 cells/ml. Growth is more affected by nutrient concentration than by salinity. No growth was obtained with the lowest salinities tested (0 and 5 ‰) at any of the nutrient concentrations used. Variations in salinity and in nutrient concentration had a greater effect on the final biomass than on the velocity of growth. Chlorophyll-a/ml was affected by salinity and nutrient concentrations and maximum values were found with 30 ‰ salinity and nutrient concentrations between 8 and 64 mM of NaNO 3 . Chlorophyll-a/cell reached maximum values between 2.02 and 3.51 pglcell and is only significantly affected by the nutrient concentration. These maximum values were reached with low nutrient concentrations (1-2 mM of NaNO 3 ). Protein per ml of culture and protein per cell were closely related to salinity and nutrient concentrations. Maximum protein per ml occurred at 20-25 ‰ salinity and 64mM of NaNO 3 , with values between 926 and 957 μml. Maximum protein/ cell concentrations were obtained also at 64 mM of NaNO 3 for all the salinities. The nitrate-protein transformation rate was related to nutrient concentration and was independent of salinity. Maximum rate was 100% at 20 ‰ salinity and 1 mM of NaNO 3 . This rate decreased as nutrient concentrations increased.