两株小球藻培养基的优化及其产EPA性能的研究

采用L18(37)正交法优化小球藻C95和小球藻C97的生长培养基,经15 L规模培养并收获小球藻;碱性乙醇法萃取游离脂肪酸,再经脲素包埋提取多不饱和脂肪酸,并用气相色谱检测二十碳五烯酸(eicosapentaenoic acid,EPA)的含量。研究结果表明:小球藻C95最优培养基(微量元素母液0.8 m L/L,维生素母液2.4 mL/L,NaH2PO4·2H2O6.0 g/L,Na2SiO3·9H2O 9.0 g/L,NaNO360.0 g/L,FeC6H5O7·5H2O 3.0 g/L);小球藻C97最优培养基(微量元素母液1.6 m L/L,维生素母液2.4 m L/L,NaH2PO4·2H2O 4.0 g/L,Na2SiO3·9H2O 11.0 g/L,NaNO380.0 g/L,FeC6H5O7·5H2O 3.0 g/L);小球藻C95和小球藻C97对数末期(分别是第4天和第5天)达到的细胞密度分别为9.53×10^7个/mL和8.91×10^7个/mL,EPA含量分别占总脂肪酸的30.2%和29.5%,小球藻C95的EPA含量要高于小球藻C97。     

污水处理中小球藻的自絮凝特性

以BG11(+N)培养基和模拟生活污水为底物,考察了污水处理小球藻(Chlorella vulgaris)的自絮凝特性,探究了自絮凝机理。结果表明:以短沉淀时间运行,小球藻在模拟生活污水中自然形成絮体,沉降性良好;藻细胞表面粗糙,含有丝状、胶状胞外聚合物;絮体内部含有大量短杆菌、球菌;絮体具有相对较高的Na、P、Fe、Ca、Mg元素含量,无机物中主要物质形态为Ca4O(PO4)2。污水处理小球藻的絮凝机理为:基质条件及反应器运行参数等促使藻细胞生理过程发生改变,藻细胞分泌更多絮凝性胞外聚合物增强黏附,生化过程富集P、Fe、Ca、Mg元素改善沉降性,絮凝性细菌大量繁殖促进菌藻共絮凝。     

菹草对普通小球藻和铜绿微囊藻的化感作用

选取沉水植物菹草与浮游藻类普通小球藻、铜绿微囊藻为研究对象,在菹草共培养胁迫下测定2种藻单独存在和按1∶1混合情况下的光密度、叶绿素a(Chl-a)、最大光化学量子产量、相对电子传递速率、可溶性糖、丙二醛以及超氧化物歧化酶活性的变化。结果表明,在菹草存在的条件下,普通小球藻、铜绿微囊藻和二者混合藻的生长被快速、强烈地抑制,抑制率最大值分别为74.38%、80.98%和89.63%。3个处理组的光密度值、叶绿素a、可溶性糖和超氧化物歧化酶均低于对照组,且随时间呈明显的下降趋势,说明其光合能力逐渐减弱;而丙二醛含量在0~6 d却高于对照组,表明可能发生了浮游藻类膜脂过氧化过程。     

小球藻对奶牛场沼液处理能力及生物质生产的探究

为资源化处理奶牛场沼液、探究小球藻Chlorella vulgaris NIES-227对奶牛场沼液的处理能力以及生物质利用潜力，在柱式光生物反应器中利用小球藻处理沼液占比分别为25%、50%、75%和 100% 4种不同浓度的未灭菌污水。研究结果显示，各浓度污水中总氮、总磷和COD的去除率分别为36.0%～92.5%、42.8%～100%和6.9%～32.2%。在沼液占比为25%的污水中氮磷的去除率最高，氨氮、总氮和总磷的去除效率分别可达99.9%、91.0%和100%。微藻在低浓度沼液（25%～50%）中生长状态良好，在沼液占比为50%的污水中可获得最高生物质产率393.6 mg/(L·d)。但是在高浓度沼液（75%～100%）中微藻生长受到一定抑制，导致氮磷的去除效果变差。培养期间细菌的数量增长显著，促进了COD的去除。各浓度沼液生物质中总脂、总糖和蛋白质含量分别为13.2%～32.2%、12.3%～27.6%和16.2%～30.9%。实验数据表明，低浓度沼液能产生更多高能量组分的生物质，适合用作生物燃料的开发；高浓度沼液能产生含较多蛋白质的生物质，更适合用作动物饲料。     

The Discrepancy of Fatty Acid Composition of Astaxanthin Esters and Total Fatty Acids in Photoautotrophic and Heterotrophic Chlorella zofingiensis

Chlorella zofingiensis has great potential for astaxanthin and fatty acid production. This study investigated the accumulation of astaxanthin and total fatty acids (TFA), the fatty acid compositions of astaxanthin esters and TFA in photoautotrophic and heterotrophic C. zofingiensis, respectively. The results displayed that in photoautotrophic cells, 18:3 was the most predominant fatty acid, followed by 16:0, 18:2, and 18:1, while astaxanthin mono-esters (mono-AE) contained only 16:0 and 18:0, and astaxanthin di-esters (di-AE) contained 16:0 mostly, followed by 18:1, 18:0, 18:2, and 18:3. C. zofingiensis accumulated the highest level of astaxanthin and TFA under heterotrophic conditions fed with 30 g L⁻¹ glucose. The percentage of 18:1 in TFA, mono-AE, and di-AE increased from 10.05%, 0%, and 17.76% to 35.80%, 14.84%, and 50.12%, respectively; in contrast, the percentages of 18:3 in TFA and di-AE were observed to decrease from 40.42% and 7.59% to 9.56% and 1.30%, respectively, in comparison with those in photoautotrophic ones. The RNA sequencing (RNA-seq) results showed that the expressions of the genes in astaxanthin and fatty acid biosynthetic pathways were increased under this heterotrophic condition. These results indicated differential accumulation of AE and TFA, especially fatty acid compositions of TFA and AE in photoautotrophic and heterotrophic cells, suggesting a positive correlation between astaxanthin and fatty acid biosynthesis.     

珠磨法破碎小球藻提取类胡萝卜素的动力学及能量消耗分析

采用珠磨法破碎小球藻冷冻干燥藻和湿藻细胞，以叶绿素释放量为指标评价细胞破碎程度，分析其细胞破碎过程的动力学情况，同时结合能量消耗情况分析细胞破碎程度对类胡萝卜素提取效果的影响。结果表明：一级动力学方程可较好地拟合小球藻细胞珠磨破碎过程。在类胡萝卜素提取过程中，将冷冻干燥藻和湿藻细胞的破碎率分别控制在70%和90%时，其最佳的类胡萝卜素提取量可分别达6.38 mg/g和6.26 mg/g。通过破碎过程的能量消耗分析，发现湿藻细胞破碎过程消耗的能量要高于冷冻干燥藻；冷冻干燥藻细胞质量浓度在40～160g/L时，只要输入相同的比能量消耗，均可达到较为一致的细胞破碎率及类胡萝卜素提取效果；而湿藻细胞质量浓度在80g/L时，可在较低的比能量消耗情况下达到较高的细胞破碎率及类胡萝卜素提取量。本实验结果可为小球藻的细胞破碎过程及其有效成分的开发利用提供一定的理论参考和实验依据。     

优化营养方式强化蛋白核小球藻生物量及蛋白质和叶绿素生产

本文通过优化营养方式和培养条件,提高了蛋白核小球藻生物量及细胞内蛋白质和叶绿素含量。结果表明,葡萄糖浓度能显著影响蛋白核小球藻混养和异养生长的生物量浓度(p0.05),但不同硝酸钠浓度影响不显著。在葡萄糖浓度50 g/L、硝酸钠浓度3.75 g/L条件下,小球藻异养培养4 d,即可达到最高生物量浓度为21.31 g/L,显著高于盐度15‰条件下混养所得最高生物量浓度(14.32 g/L)(p0.05)。当葡萄糖耗尽,将小球藻细胞经含氮且不含糖的培养基适当稀释后进行光诱导培养,并优化硝酸钠浓度。发现在高异养细胞密度(11.09 g/L)下,采用硝酸钠浓度(3.75 g/L)和低光强(6,411±532 Lux)诱导培养藻细胞48 h,即可获得高蛋白质含量(54.10%)和高叶绿素含量(3.14%)的小球藻,相对于光诱导培养前,分别提高了207.74%和342.25%,蛋白质和叶绿素产率也显著高于单独混养或异养培养的最高值,分别达到1.32 g/(L·d)和0.09 g/(L·d)。     

塑料薄膜材料在微藻培养环境中的稳定性和生物附着行为

考察了6种薄膜材料在次氯酸钠(NaClO)溶液、HCl溶液(pH=2)和NaOH溶液(pH=12)中的稳定性及其在普通小球藻培养体系中的生物附着行为。结果表明，聚氯乙烯(PVC)在3种溶液中浸泡后420 nm处的透光率下降约50%，聚氨酯(PU)在NaClO溶液中、乙烯?醋酸乙烯酯(EVA)在NaOH溶液中浸泡后420 nm处的透光率下降10%?15%，聚乙烯(PE)、聚丙烯(PP)和聚对苯二甲酸乙二醇酯(PET)在3种溶液中浸泡后的透光率没有明显变化；6种薄膜材料在普通小球藻培养体系中均有微藻明显附着，浸泡7 d后透光率明显下降，其中PVC表面附着最严重，浸泡7 d后透光率接近0，附着物量随浸泡时间延长而增加，浸泡45 d后达3069 ?g/cm2；其它5种薄膜表面附着物量先增加后降低，PU和EVA在第15 d时、PE, PP和PET在第30 d时附着物量最大，分别为292, 375, 292, 194和236 ?g/cm2。     

The enhancement mechanism of hydrogen photoproduction in Chlorella protothecoides under nitrogen limitation and sulfur deprivation

The aim of this study was to understand the enhancement mechanism of H₂ photoproduction in Chlorella protothecoides under simultaneous nitrogen limitation and sulfur deprivation (LNS). Nitrogen limitation (LN) rather than sulfur deprivation significantly inhibited relative variable fluorescence at K-step (W_K) and J-step (V_J), photochemical efficiency of PSII (photosystem II), F_v/F_m, during the process of incubation in the light. Under such conditions, photosynthetic O₂ evolution decreased and the anaerobiosis was established after 12 h of incubation. The algae generated large amounts of H₂ under nitrogen limitation but generated only trace amounts under sulfur deprivation. Obviously, nitrogen limitation rather than sulfur deprivation was the decisive factor that induced H₂ photoproduction in C. protothecoides under LNS. The LNS culture generated much more H₂ than the LN culture in the presence of DCMU during incubation, suggesting that a PSII-independent electron source contributed many more electrons for transfer to hydrogenase in the LNS culture. PSII electron transport includes linear electron flow (LEF) and cyclic electron flow (CEF) of PSII in C. protothecoides. In the PSII-dependent electron source for H₂ photoproduction, PSII supplies electrons to hydrogenase through the LEF. The LNS culture showed much higher LEF and lower CEF than the LN culture during the H₂ photoproduction phase, as indicated by the large lower quantum yield of PSII electron transport (Φ_PSII) in the LNS culture in the presence of DCMU. Therefore, compared with nitrogen limitation, simultaneous nitrogen limitation and sulfur deprivation enhanced H₂ photoproduction in C. protothecoides mainly due to enhanced PSII-dependent and -independent electron sources.