The Effects of Environmental Stressors on Aggregation in <i>Parachlorella kessleri</i>

The induction of biofilm formation has been explored as a means of harvesting microalgae for bioprocessing applications. Environmental stressors have been implicated in the induction of biofilm formation;however, it is unclear whether all stressors, or a select few, are responsible. This study aimed to investigate the effects of three stressors, nitrogen depletion, reduced or elongated day lengths, and increased culture turbulence on bio-film formation of Parachlorella kessleri. We also examined corresponding effects on growth and production of reactive oxygen species. Turbulence induced the greatest response in which a significant decrease in growth plus an increase in superoxide production and flocculation efficiency were seen for the 300-rpm treatment. For varying day lengths, stress response was not observed, however, a significant increase in EPS secretion was measured in both shorter and longer days. Nitrogen depletion induced a low-level stress response, in which superoxide production increased for the highest concentrations, while growth was not impacted. In contrast to previous studies on nitrogen depletion, a significant increase in EPS secretion was not observed. Results indicate that stress response varies according to type and magnitude. EPS production and thus biofilm formation are not linked to the stress indicators investigated. P. kessleri uses small quantities of EPS to contribute to cell stickiness, but not necessarily to the full formation of a biofilm;however, cell stickiness served as a mechanism for substrate adherence and cellular aggregation none the less.

凯氏拟小球藻Z6利用葛渣水解液合成油脂的工艺优化研究

【目的】筛选获得适合凯氏拟小球藻Z6(Parachlorella kessleri Z6)利用的木质纤维原料,通过单因素及响应面法优化该藻株培养条件,以期提高其油脂产量。【方法】通过探究葛渣、甘蔗渣、稻草秸秆及菌糠等4种水解液培养条件下凯氏拟小球藻Z6生物量与油脂产量变化,获得最优木质纤维原料;通过单因素、PB以及中心组合试验获得凯氏拟小球藻Z6最优培养工艺,并通过GC-MS技术分析优化前后凯氏拟小球藻Z6脂肪酸成分。【结果】(1)以葛渣水解液为发酵原料时,凯氏拟小球藻Z6的生物量与油脂产量最高;(2)利用PB试验获得葛渣水解液、硝酸钠、磷酸氢二钾和柠檬酸等4种成分为显著影响因素;(3)通过CCD中心组合试验确定凯氏拟小球藻Z6的最佳培养条件为70%葛渣水解液、0.70 g/L硝酸钠、0.02 g/L磷酸氢二钾、0.075 g/L七水硫酸镁、0.04 g/L二水氯化钙、0.008 g/L柠檬酸,其余成分与BG11培养基一致;(4)在最优组合条件下,凯氏拟小球藻Z6的油脂产量可达到2.22 g/L,相较于初始葛渣水解液培养基和异养培养基分别提升了43.23%和12.69%。【结论】葛渣水解液可作为凯氏拟小球藻Z6的发酵原料,经响应面法优化后的藻株Z6总脂量有显著提高,研究结果可为其他木质纤维素原料利用以及提高微藻产油性能提供有益的参考。

凯氏拟小球藻除锰过程及机理分析

与传统的废水中去除Mn^(2+)方法相比,生物修复具有更高的环境和经济效益。从安徽某铁硫矿山的酸性废水坑湖中分离纯化得到一株新型藻株——Parachlorella kessleri MP1,研究了其在不同pH值、Mn^(2+)浓度条件下的生长过程、培养基pH值变化和除锰效率,并分别通过测定细胞内外去除Mn^(2+)量、谷胱甘肽(GSH)含量、巯基含量以及傅里叶变化红外光谱(Fourier transform infrared spectroscopy,FTIR)分析初步探讨了除锰机制。结果表明,Parachlorella kessleri MP1可耐受pH值为3.5的酸性环境,且对Mn^(2+)具有良好去除效果。当Mn^(2+)初始浓度分别为5、15和30 mg/L时,pH=3.5和pH=7.0初始条件下可分别实现35.86%、30.82%、27.54%和93.73%、74.11%、52.22%的去除率。此外,该藻株具有产碱作用,pH=3.5试验组反应结束时pH值达到了9.04。所测胞外去除Mn^(2+)量远大于胞内去除,胞内GSH和巯基含量有所增加,FTIR分析得知细胞表面—NH_(2)、—OH、—COOH和C—O等官能团参与了除锰过程。因此,微藻除锰过程中胞外去除发挥主要作用。

凯氏拟小球藻己糖激酶基因的克隆及其在不同培养条件下的表达分析

为研究凯氏拟小球藻(Parachlorellakessleri)糖代谢分子机制,本研究采用RT-PCR和RACE技术从凯氏拟小球藻中克隆了己糖激酶基因CkeHK(GenBankID:AHF54566),并对其自养、异养、混养条件下的转录表达进行分析。结果表明,该序列的cDNA全长为1844bp,开放阅读框1389bp,编码462个氨基酸。该蛋白的相对分子质量为49.73,等电点为6.98。实时荧光定量PCR结果显示,以自养培养条件为对照,异养培养和混养培养条件下,CkeHK均能够发生明显上调,且混养条件下上调量比异养条件下上调量更多,说明CkeHK可能在凯氏拟小球藻利用外源糖的过程发挥重要作用,并且光信号对于凯氏拟小球藻利用外源糖可能存在调控作用。这些研究结果为进一步阐明CkeHK的功能及其作用机制奠定了分子基础。

一株新型自絮凝凯式拟小球藻的两步培养产油技术

以分离获得的一株新型自絮凝凯式拟小球藻(Parachlorella kessleri) F01为材料,自养单步培养法为对照,设计两步培养法,研究阶段Ⅰ添加葡萄糖兼养和阶段Ⅱ营养元素限制处理对藻细胞油脂积累及絮凝性能的影响。分别采用血球板计数法、干重法、脂染色法测定藻细胞浓度、生物量和总脂含量,三维荧光光谱分析藻细胞胞外聚合物(Extracellular polymeric substances, EPS)组分和含量。结果表明:(1)两步培养法阶段Ⅰ兼养培养最适葡萄糖浓度为10 g/L, 10d收获时藻细胞油脂产率204.25 mg/(L·d),是对照组的16.20倍;静置12h的藻细胞自絮凝率96.1%,与对照组差异不显著。(2)在阶段Ⅰ基础上,阶段Ⅱ进行不同元素限制处理培养1d,低糖组和低糖低氮协同处理组的藻细胞油脂产率分别为242.64和227.61 mg/(L·d),分别比阶段Ⅰ增加18.8%和11.4%;培养4d,低糖、无糖、低氮和低糖低氮协同4种处理组油脂产量显著高于对照组和阶段Ⅰ,其中低糖低氮协同处理组最高,达到3.08 g/L,是对照组的23.69倍,比阶段Ⅰ增加了51.0%,而且阶段Ⅱ中4种处理组藻细胞的自絮凝率基本在85.0%以上,能满足收获要求。(3) F01藻细胞EPS中蛋白类色氨酸物质含量高低与藻细胞自絮凝率大小密切相关,不同培养处理通过改变藻细胞EPS中蛋白类色氨酸物质的含量而影响其絮凝性能。自絮凝凯式拟小球藻F01是生物柴油生产的优良潜力藻种,两步培养法能大幅提升其产油效益。产油微藻的自絮凝优势与两步培养法结合,有望成为解决微藻生物柴油生产技术瓶颈的关键突破口。

Effect of storage conditions on long-term stability of Ag nanoparticles formed via green synthesis

Spherical Ag nanoparticles(AgNPs) with a diameter of 20 nm or smaller were biologically synthesized using algae Parachlorella kessleri. The effect of storage conditions on the long-term stability of AgNPs was investigated. UV/Vis spectrophotometry, transmission electron microscopy, and dynamic light scattering measurements revealed that the long-term stability of AgNPs was influenced by light and temperature conditions. The most significant loss of stability was observed for the AgNPs stored in daylight at room temperature. The AgNPs stored under these conditions began to lose their stability after approximately 30 d; after 100 d, a substantial amount of agglomerated particles settled to the bottom of the Erlenmeyer flask. The AgNPs stored in the dark at room temperature exhibited better long-term stability. Weak particle agglomeration began at approximately the 100 th day. The AgNPs stored in the dark at about 5℃ exhibited the best long-term stability; the AgNPs stored under such conditions remained spherical, with a narrow size distribution, and stable(no agglomeration) even after 6 months. Zeta-potential measurements confirmed better dispersity and stability of AgNPs stored under these conditions.

银纳米颗粒的绿色合成:藻类生命周期对银纳米颗粒合成和长期稳定性的影响（英文）

用分别培养了1、2、3和4周的凯氏拟小球藻的四种不同的提取物,生物合成球形银纳米颗粒(AgNPs)。研究藻类生命周期对AgNPs合成的影响和不同的储存条件对AgNPs长期稳定性的影响。结果发现,藻类的年龄会影响AgNPs的合成率和溶液中的数量,但不影响AgNPs的长期稳定性。UV–vis和TEM的结果显示,AgNPs的长期稳定性受储存温度的影响,低温更有利于其长期稳定性。储存在黑暗环境和5°C左右时,AgNPs表现出最佳的长期稳定性,而与藻类年龄无关,存储6个月以后,这些AgNPs仍然保持着球形,尺寸细小(5~20nm),且无团聚。