| 1. Breeding High-lipid Excellent strains of green algaeThe six green algae (Halochlorococcum marinum C11, Nannochloris oculata C14, Oocystisminuta C16, Chlamydomonas sp. C30, Dunaliella sp. C46and Chlorella sp. C103) were culturedusing f/2medium with continuous aeration under (25±1)℃， the light intensity was100μmol/(m~2·s)，the salinity was31. Through comparisons of the chlorophyll fluorescence parameters(maximum photochemical efficiency of PS Ⅱ Fv/Fm, photochemical quenching qP, non-photochemical quenching NPQ, maximum efficiency of photosynthesis Pm, the initial slope of therapid curve α and half-saturation Ik), cell density, dry weight, relative growth ratio, chlorophyllcontent, total lipid content and total lipid productivity, the two excellent strains (H. marinum C11and N. oculata C14) that had rapid growth, high photosynthesis activity, high total lipid content andtotal lipid productivity were selected from six green algae. The effects of different media (f/4, f/2, f,2f,4f and8f) on the above parameters and fatty acid composition of the two green algae (C11andC14) were investigated. The results showed that different media had significant effects on thechlorophyll fluorescence parameters (P<0.05). Cell density, dry weight and relative growth ratio ofthe two algae in f treatment were the higher than the other treatments. The total lipid contents in ftreatment of C11and C14were29.27%and28.59%. The two algae cultured in f treatment had thehighest total lipid productivity, were0.012g/(L·d). The monounsaturated fatty acid (MUFA) of thetwo green algae were found to be decreased with the increase of nutrient concentration in media,while the polyunsaturated fatty acid (PUFA) content of these algae were increased. In summary, fmedium were the optimum medium of the two algae.2. Effects of different induction condition on the growth, chlorophyll fluorescenceparameters, total lipid content and fatty acid composition of the two green algaeC11and C14were cultured using f medium with continuous aeration under (25±1)℃，the lightintensity was100μmol/(m~2·s)，the salinity was31. Centrifugated and harvested400ml algae in theend of the exponential growth phase, then transferred to the same volume f medium, which did notcontain N, and cultured with continuous aeration for0,1,2,4and6d, respectively. The effect ofdifferent times of nitrogen deficiency on the growth, chlorophyll fluorescence parameters, total lipidcontent and fatty acid composition of the two green algae were studied. Nitrogen deficiency hadsignificant effects on the chlorophyll fluorescence parameters of the two green algae (P<0.05). Thecell density, dry weight, total lipid content and total lipid yield of the two algae were increased withthe increase of nitrogen deficiency time. The maximal total lipid content of C11and C14occurring in the6d treatment were47.34%and47.46%, respectively. The total lipid productivity of C11andC14were first increased then decreased with the increase of nitrogen deficiency time, the maximalvalues occurring in the2d treatment were0.02g/(L·d) and0.017g/(L·d), respectively. The MUFAof the two algae were increased with the increase of nitrogen deficiency time, while PUFA reduced.Therefore, the optimum nitrogen deficiency time of C11and C14were2d.Cultured and harvested C11and C14as the above way and transferred to the same volume fmedium with different sodium tangstate concentrations (0,0.5,1,2and4mmol/L) and culturedwith continuous aeration. The effect of sodium tangstate concentrations on the growth, chlorophyllfluorescence parameters, total lipid content and fatty acid composition of the two green algae wereinvestigated. The4thd, sodium tangstate had significant effects on the chlorophyll fluorescenceparameters in C11(P<0.05), but NPQ and α. While sodium tangstate had no significant effect onthe chlorophyll fluorescence parameters in C14(P>0.05), except for qP. Cell density and dry weightof the two algae were first increased and then decreased with the increase of sodium tangstateconcentration, the maximal values of C11and C14occurring in0.5mmol/L treatment and1mmol/L treatment, respectively. C11and C14had the highest total lipid content in1mmol/Ltreatment (31.78%and31.27%). The maximal total lipid productivity occurred in0.5mmol/Ltreatment [0.021g/(L·d)] of C11, and in1mmol/L treatment [0.017g/(L·d)] of C14. The MUFA ofC11and C14in1mmol/L treatment was higher than the other treatment. Sodium tangstateconcentration had no significant effect on the other fatty acids of the two algae (P>0.05). Insummary, the optimum sodium tangstate concentrations of C11and C14were0.5mmol/L and1mmol/L, respectively.Cultured and harvested C11and C14as the above way and transferred to the same volume fmedium with different citric acid concentrations (0,0.05,0.1,0.2and0.4g/L) and cultured withcontinuous aeration. The effects of citric acid concentrations on the growth, chlorophyllfluorescence parameters, total lipid content and fatty acid composition of the two green algae wereinvestigated. Citric acid had significant effects on the chlorophyll fluorescence parameters of thetwo algae (P<0.05). Cell density and dry weight of the algae were first increased then decreasedwith the increase in citric acid concentration. While the total lipid content of the two algaeincreased with the increase of citric acid concentration. Both of C11and C14had the highest totallipid content of35.2%and40.42%in0.4g/L treatment, and the highest total lipid productivity of0.018g/(L·d) and0.025g/(L·d) in0.1g/L treatment. MUFA of C11and C14was increased with theincrease of citric acid concentration, while16:3n-3of the two algae was decreased. In summary,the optimum citric acid concentrations of C11and C14were0.1g/L.C11and C14were cultured using f medium, which contained different IAA concentration(IAA concentration of C11were0,0.025,0.1and0.2mg/L; IAA concentration of C14were0, 0.05,0.1,0.2and0.4mg/L) with continuous aeration under (25±1)℃，the light intensity was100μmol/(m~2·s)，the salinity was31. The effects of different IAA concentration on the growth,chlorophyll fluorescence parameters, total lipid content and fatty acid composition of the two algaewere studied. The10thd, IAA had significant effects on the chlorophyll fluorescence parameters ofC11and C14(except for I_k)(P<0.05). Cell density, dry weight and chlorophyll content of C11andC14were first increased then decreased with the increase of IAA concentration. The total lipidcontent of C11in0.05mg/L treatment was lower than the other treatments, and C11had thehighest total lipid productivity in0.025mg/L [0.037g/(L·d)]. C14had the highest total lipidcontent and total productivity of36.37%and0.023g/(L·d) in0.1mg/L treatment. The18:2n-6ofC11in0.025mg/L treatment was higher than the other treatments. IAA had no significant effectson the other fatty acids of C11, and all fatty acids of C14(P>0.05). In summary, the optimum IAAconcentrations of C11and C14were0.025mg/L and0.1mg/L, respectively. |
