David L. Parry

Effect of temperature on growth, chemical composition and fatty acid composition of tropical Australian microalgae grown in batch cultures

The growth and nutritional content of four tropical Australian microalgal species, diatom Chaetoceros sp. (CS256), two cryptomonads, Rhodomonas sp. (NT15) and Cryptomonas sp. (CRFI01) and unidentified prymnesiophyte (NT19), cultured at five different temperatures, were studied. Commercially available Isochrysis sp. (clone T.ISO) was included in the study for comparison. Microalgae were grown in laboratory 1.5-l batch cultures in F/2 medium at 25, 27, 30, 33 and 35 °C (salinity 25‰; pH 8.3; photon flux density 80 μmol photon m−2 s−1; 12:12 h light:dark cycle). Microalgal cells were harvested in late logarithmic growth phase and analysed for protein, carbohydrate, lipid, chlorophyll a, inorganic matter (ash) and fatty acid composition. The optimum temperature for growth was 25–27 °C for Rhodomonas sp. (specific growth rate, μ=0.27 day−1), and 27–30 °C for prymnesiophyte NT19, Cryptomonas sp., Chaetoceros sp. and Isochrysis sp. (μ=0.56, 0.33, 0.87 and 0.97 day−1, respectively). Only Chaetoceros sp. grew well at 33 and 35 °C (μ>0.78 day−1). All tropical Australian species had significantly lower percentages of protein when cells were grown at temperatures above 27 °C, but there was no consistent trend in the percentages of carbohydrate. Chaetoceros sp. had highest percentage of lipid (16.8% dry weight; P<0.01), when cells were cultured at 25 °C, while Rhodomonas sp., Cryptomonas sp., NT19 and Isochrysis sp., had significantly higher amounts of lipid at temperatures within the range 27–30 °C (15.5, 12.7, 21.4, and 21.7% dw, respectively; P<0.05 in each case). Considering all species together, there was no overall relationship between percentage of protein, carbohydrate or lipid and temperature, but there was an overall, linear relationship between percentage of ash (inorganic matter) and temperature (r2=0.42, P≤0.05). Highest calculated energy values were found in Chaetoceros sp. (21.9 kJ g−1) and Tahitian Isochrysis sp. (22.5 kJ g−1) cultured at 27–30 °C. There was no significant change in chlorophyll a (range 1.2–1.68 pg cell−1) for any species over the temperature range studied. The highly unsaturated fatty acid (HUFA), eicosapentaenoic acid, 20:5n−3, was present in all species, with highest amounts in prymnesiophyte NT19 (19.9% total fatty acids). Percentages of 20:5n−3 were slightly lower at highest growth temperatures for all species. The control, Isochrysis sp., had the highest amount of docosahexaenoic acid, 22:6n−3 (6.6% total fatty acids). All species had lower percentages of 22:6n−3 at higher growth temperatures. Chaetoceros sp. and NT19 had moderate amounts of arachidonic acid, 20:4n−6 in the fatty acid profile (2.7–5.4% total fatty acids). Highest percentages were associated with growth temperatures within the range 27–30 °C. Only Chaetoceros sp. grew well at 35 °C, maintaining moderate percentages of protein, carbohydrate, lipid, PUFA and HUFA (9.6% total fatty acids), at that temperature. All tropical Australian species performed better than Isochrysis sp., in terms of percentage of HUFA, over the range of growth temperatures.

The gross chemical composition and fatty acid composition of 18 species of tropical Australian microalgae for possible use in mariculture

Abstract The gross chemical composition and fatty acid composition of 18 species of tropical Australian microalgae (2 cryptomonads, 8 diatoms, 3 prasinophytes, 4 prymnesiophytes and a rhodophyte) grown under standard conditions (pH 8.3±0.2, temperature 25±1°C, photon flux density 80±2 μ mol m −2 s −1 and salinity 25±1‰ and harvested in late log phase of growth), were determined as part of a program to find species for use as feed in tropical aquaculture. Protein was the major component of all species (range 24.2–47.1% dry weight) except diatoms (range 19.4–36.7% dw). Highest lipid contents were found in Cryptomonas sp. CRFI01 and Isochrysis sp. NT14 (22.0 and 23.4% dw, respectively). The highest percentage of soluble carbohydrate was found in Rhodosorus sp. CS249 (20.2% dw), while diatoms had the highest amounts of ash (range 21.2–38.8% dw). Moderate levels of the polyunsaturated fatty acids (PUFA) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) were found in Cryptomonas sp. CRFI01 (12 and 6.6% of total fatty acids, respectively). Moderate levels of EPA were also found in Chaetoceros sp. CS256 and in Skeletonema species GOC27 and GOC36. DHA was most abundant in Isochrysis sp. NT14 (9.9% total fatty acids) and Cryptomonas sp. CRFI01 (6.6% total fatty acids). The highest amount of arachidonic acid (AA) was found in Fragilaria sp. GOC52.

Effect of light intensity on the proximate biochemical and fatty acid composition of Isochrysis sp. and Nannochloropsis oculata for use in tropical aquaculture

The total protein, carbohydrate, lipid and ash compositions, and fatty acid contents of two species of marine microalgae, the eustigmatophyte Nannochloropsis oculata (formerly ‘Chlorella sp., Japan’) and the chrysophyte Isochrysis sp. (Tahitian) used in tropical Australian mariculture, were studied. The microalgae were grown under a range of culture conditions (41 and 601 laboratory culture, 3001 bag culture, and 80001 outdoor culture) and four light regimes (100 to 107 µ E m−2 s−1, 240 to 390 µ E m−2 s−1, 340 to 620 µ E m−2 s−1, and 1100 to 1200 µE m−2 s−1 respectively) to determine the effect of light intensity on the chemical composition of large scale outdoor cultures. Laboratory and bag cultures were axenic and cultured in Walne medium while outdoor cultures were grown in a commercial medium designed for optimum nutrition in tropical outdoor aquaculture operations. Change in growth medium and photon flux density produced only small changes in the proximate biochemical composition of both algae. N. oculata and Isochrysis sp. both showed a trend towards slightly lower carbohydrate and higher chlorophyll a in shaded outdoor culture. Isochrysis sp. showed significant concentrations of the essential polyunsaturated fatty acid 22:6(n−3) (docosahexaenoic acid) from 5.3 to 10.3% of total fatty acid, and 20:5(n−3) (eicosapentaenoic acid) ranged from 0.6 to 4.1%. In contrast, N. oculata had high concentrations of 20:5(n−3) (17.8 to 39.9%) and only traces of 22:6(n−3). The fatty acid composition of Isochrysis sp. grown at high photon flux density (1100–1200 µE m−2 s−1) under outdoor culture showed a decrease in the percentage of several highly unsaturated fatty acids, including 20:5(n−3), and an increase in 22:6(n−3). N. oculata showed a similar decrease in the percentage of 20:5(n−3). High light intensity caused a decrease in the ratio of total C16 unsaturated fatty acids to saturated 16:0 in N. oculata, and a decrease in the ratio of total C18 unsaturated fatty acids to saturated 18:0 together with a decrease in the ratio of total unsaturated fatty acids to total saturated fatty acids in both microalgae.

Effect of temperature on the growth, total lipid content and fatty acid composition of recently isolated tropical microalgae Isochrysis sp., Nitzschia closterium, Nitzschia paleacea, and commercial species Isochrysis sp. (clone T.ISO)

The effect of temperature from 10 °C to 35 °C on the growth, total lipid content, and fatty acid composition of three species of tropical marine microalgae, Isochrysis sp., Nitzschia closterium, N. paleacea (formerly frustulum), and the Tahitian Isochrysis sp. (T.ISO), was investigated.Cultures of N. closterium, Isochrysis sp. and T.ISO grew very slowly at 35 °C, while N. closterium did not grow at temperatures higher than 30 °C or lower than 20 °C. N. paleacea was low-temperature tolerant, with cells growing slowly at 10 °C. N. paleacea produced the highest percentage of lipids at 10 °C, while the other species produced maximum amounts of lipid at 20 °C. None of the species maintained high levels of polyunsaturated fatty acids (PUFAs) at high growth temperature and there was a significant inverse relationship between the percentage of PUFAs and temperature for N. paleacea. A curved relationship was found between temperature and percentage of PUFA for N. closterium and tropical Isochrysis sp., with the maximum production of PUFA at 25 °C and 20 °C, respectively. The two Nitzschia species produced higher levels of the essential fatty acid eicosapentaenoic acid [20:5(n-3)] at lower growth temperatures, but the two Isochrysis species had little change in percentage of 20:5(n-3) with temperature. Only T.ISO had the highest percentage of 22:6(n-3) at lowest growth temperature (11.4% total fatty acids at 10 °C).

Microalgae for use in tropical aquaculture I: Gross chemical and fatty acid composition of twelve species of microalgae from the Northern Territory, Australia

Twelve species of microalgae, isolated from north Australian marine, freshwater and hypersaline environments, were grown under controlled conditions of temperature, pH, photon flux density and salinity, and analysed for ash, total protein, water soluble carbohydrates, chlorophylla, total lipids, total fatty acids and fatty acid composition. Highest levels of the polyunsaturated fatty acid eicosapentaenoic acid [20:5(n-3)] were found in the marine diatoms.Nitzschia (frustulum) andN. closterium (23.1% and 15.2% of total fatty acids, respectively). None of the species studied had levels of docosahexaenoic acid [22:6(n-3)] greater than 1.1 % of total fatty acids. None of the chlorophyte species contained significant levels of either 20:5(n-3) or 22:6(n-3). The highest total fatty acid concentration of all species in the study was found in the freshwater chlorophyte speciesScenedesmus dimorphus (105 mg g−1 dry wt). The hypersaline speciesDunaliella salina had the highest total lipid content (28.1% dry wt), followed byN. closterium, N. (frustulum) andNavicula sp. (24.2–27.8% dry wt).Chlamydomonas sp. had the highest protein content (66.9% dry wt).N. (frustulum) was highlighted as a possible useful source of lipids and polyunsaturated fatty acids in mixed microalgal diets for mariculture organisms used in tropical aquaculture.

Microalgae for use in tropical aquaculture II: Effect of salinity on growth, gross chemical composition and fatty acid composition of three species of marine microalgae

The influence of salinity on the growth, gross chemical composition and fatty acid composition of three species of marine microalgae,Isochrysis sp.,Nannochloropsis oculata andNitzschia (frustulum), was investigated. There was no significant change in growth rate ofIsochrysis sp. andN. (frustulum) over the experimental range of salinity (10–35 ppt), whileN. oculata had a significantly slower growth rate only at 35 ppt. The ash content of all three species increased with increasing salinity. Two species,Isochrysis sp. andN. oculata, showed significant linear increases in total lipid content with increasing salinity over the range 10 to 35 ppt.N. (frustulum) showed significant linear decrease in total lipids, with the highest percentage at low salinity within the range 10–15 ppt. Variation in salinity had only a slight effect on the total protein, the soluble carbohydrate and chlorophylla content of all species. All species responded to change in salinity by modifying their cellular fatty acid compositions. Significant positive correlations were observed between increase in salinity and increase in the percentage ofcis-9-hexadecenoic acid [16:1 (n-7)] over the entire experimental range inN. (frustulum) and between 25–35 ppt inN. oculata. There were curved relationships between salinity and percentage of hexadecanoic acid [16:0] inN. oculata andN. (frustulum), with maxima within the range 25–30 ppt for both species. A curved relationship was found between salinity and percentage of eicosapentaenoic acid [20–5(n-3)], forN. (frustulum), with lowest percentages of the fatty acid within the range 25–30 ppt. There was no consistent pattern in the percentages of other major fatty acids as functions of salinity. The Northern Territory isolateN. (frustulum) was unusual in having a substantial increase in total fatty acids with decreasing salinity (85 mg g−1 dry wt at 10 ppt compared with 33 mg g−1 at 35 ppt). The optimum salinities for the production of maximum amount of lipids and the essential fatty acids 20:5(n-3) and/or 22:6(n-3) were as follows:25 ppt forIsochrysis sp. [22:6(n-3)]; 20–30 ppt forN. oculata [20:5(n-3)]; 10–15 ppt forN. (frustulum) [20:5(n-3) and 22:6(n-3)].

Evaluation of recently isolated Australian tropical microalgae for the enrichment of the dietary value of brine shrimp, Artemia nauplii

Thirteen species of tropical Australian microalgae (both benthic and planktonic) were compared in terms of survival and growth of Artemia and enrichment of polyunsaturated fatty acids (PUFAs), including eicosapenaenoic acid (EPA; 20:5n−3) and docosahexaenoic acid (DHA; 22:6n−3) in Artemia. Artemia nauplii showed enhanced growth beyond the unfed control value after 24 h feeding with all species of microalgae except Isochrysis NT14. However, when the growth was compared against the commercial northern hemisphere species Isochrysis galbana TISO, only Artemia fed Cryptomonas CRFI01 showed better growth (significance at P<0.05). Artemia fed Chaetoceros sp. CS256, Nephroselmis GOC52, Tetraselmis sp. NT18 and Nitzschia palacea NT7 showed similar growth over 24 h to those fed TISO. Artemia consumed Isochrysis NT14 at a very low rate. In 7 day feeding trials, CRFI01, CS256 and GOC52 performed much better than TISO (P<0.05) as food sources for Artemia. The species Isochrysis sp. PS11, Tetraselmis spp. TEQL01 and NT18, and NT7 showed similar results to TISO. Survival of Artemia at the end of a 7 day feeding with seven species of microalgae (NT7, PS11, CRFI01, NT18, CS256, TEQL01, and GOC52) ranged from 65 to 97%, with NT18 showing a wide range of survival results, due to the crash of some of the cultures. The suitability of these tropical Australian microalgae as a food source for Artemia was related to the fatty acid composition of the microalgae, rather than the taxonomic group, size or proximate chemical composition. Artemia fed CRFI01 had significantly higher levels of EPA and DHA (P<0.05) than those fed TISO. Artemia fed CS256 and NT7 had highest levels of EPA, however, only Artemia fed CS260 had higher amounts of DHA than Artemia fed TISO. The results showed that one species of tropical Australian microalgae CRFI01 was superior to the commercial northern hemisphere TISO as a food source for Artemia. NT7, PS11, CS256 and GOC52 could be used as a replacement or supplementary feed to TISO.