Abstracts of papers read at the Winter Meeting, 1961

Herdmania litoralis is a heterotrophic, sand-dwelling dinoflagellate with morphological characters that do not provide clear evidence for its systematic position in any existing family of dinoflagellates. Protoperidinium minutum is a heterotrophic, planktonic species that has a typical tabulation for the genus Protoperidinium. In order to infer the phylogenetic positions of these two species more confidently, we characterized the thecal plate patterns and determined small-subunit and large-subunit ribosomal DNA sequences (SSU rDNA and LSU rDNA, respectively) from both species. Intraindividual and intraspecific diversity of SSU and LSU rDNA data were characterized in H. litoralis using a combination of single-cell PCR approaches and analyses of PCR clones derived from multi-cell DNA extractions. The results of the molecular phylogenetic analyses demonstrated a novel, well-supported clade comprising both sand-dwelling species (H. litoralis and Thecadinium dragescoi) and planktonic species (P. minutum). Because the establishment of this clade also demonstrated that P. minutum is not a member of Protoperidinium, we reinstated and emended the genus Archaeperidinium Jörgensen 1912 Jörgensen, E. 1912. Bericht über die von der schwedischen hydrographisch-biologischen Kommission in den schwedischen Gewässern in den Jahren 1909–10 eingesammelten Planktonproben. Svenska Hydrograph.-Biol. Komm. Skr., 4: 1–20.  [Google Scholar].     

Abstracts of papers read at the Winter Meeting at the University of Surrey

A novel member of the Calyptrosphaeraceae surrounded by a hyaline sheath, with only one chloroplast and without an emergent haptonema, is tentatively described as a species of Calyptrosphaera Lohmann. It has an interrupted body to its single pyrenoid and a unique microtubular root complement with only one crystalline root (CR) nucleating on root 2 (R2). At preprophase it produces a CR1, lending support to the idea that these crystalline roots contribute to the mitotic spindle. It shares numerous ultrastructural traits with the heterococcolithophorid Cruciplacolithus neohelis but, although it is tempting to consider it a haploid phase of this organism, is distinct from it.     

Abstracts of Papers read at the Winter Meeting at Queen Elizabeth College,London

A new heterotrophic armoured dinoflagellate is described from sand habitats in eastern Australia. Cabra matta gen. nov., sp. nov., lacks plastids and an eyespot. The thecal plate formula is Po 4′ 4” ‘x’ 3c ?s 5′′’ 1′′”. Its plate pattern differs from all currently described dinoflagellate genera, but is most similar to the genus Roscoffia. Cabra matta shows some similarity to species currently placed in the family Podolampaceae, however its evolutionary affinities and hence its position within the dinoflagellate systematic hierarchy remain unresolved.     

雨生红球藻在不同培养基的生长比较

研究了雨生红球藻(Haematococcuspluvialis)3个不同品系CH-1、UTEX-16和CS-321分别在3种不同培养基BBM、BG-11、JM中的生长情况。结果表明:在3种培养基中,CH-1的最高细胞密度、生物量和虾青素含量都要高于另外2株雨生红球藻,其中以BBM培养的CH-1生长情况最好,其最终营养细胞密度可达到59.8×104cell?mL-1,干重为0.527g?L-1,细胞密度最高时虾青素含量为3.55mg?L-1。     

雨生红球藻八氢番茄红素合成酶基因的克隆及表征

雨生红球藻是一种单细胞绿藻,在多种逆境胁迫条件下能够大量合成并迅速积累虾青素,其积累量最高可达细胞干重的4%,从而成为目前最理想的天然虾青素合成工具.八氢番茄红素合成酶(PSY)是虾青素合成途径中第一个限速酶.分离了八氢番茄红素合成酶基因(psy)的全长cDNA及基因组DNA.其全长cDNA包括1200个碱基,编码400个氨基酸,基因组DNA包括5个外显子,4个内含子.系统发育分析结果显示,绿藻的八氢番茄红素合成酶基因形成一个进化枝,它们与高等植物的psy亲缘关系比较近.通过GenomeWalking的方法,分离了psy基因约1kb的5′侧翼序列.将含有TATA-box和CAAT-box的297bp的序列与LacZ报告基因构成嵌合的表达载体,用基因枪法转化雨生红球藻.lacZ的瞬间表达检测结果表明,这段上游序列能够驱动lacZ表达,具有启动子活性.     

活性氧对雨生红球藻生长及虾青素含量的影响

在雨生红球藻培养液中分别添加活性氧1O2、H2O2和·OH的诱生剂,通过测定细胞密度、叶绿素a含量、虾青素含量,研究了这三种活性氧诱生处理对雨生红球藻生长和虾青素含量的影响,初步探索了利用活性氧诱生剂提高雨生红球藻虾青素含量的可行性。实验结果表明,适当浓度的MB能够促进虾青素含量增加,当MB浓度为10-7mol·L-1时,虾青素含量达到5.27μg·mL-1,比对照显著提高。活性氧诱生剂对雨生红球藻生长有抑制作用,但MB的抑制作用小于H2O2和·OH诱生剂。     

乙酸钠诱导雨生红球藻合成虾青素的机理

添加乙酸钠时,雨生红球藻细胞内硝酸还原酶(NR)的活性提高了1.6倍,培养液中硝酸盐浓度的迅速下降。另一方面,乙酸钠的存在和氮缺乏,又抑制了叶绿素的合成和1,5-二磷酸核酮糖羧化酶(Rubisco)的活性。在虾青素开始合成的第4d,硝酸盐的浓度,叶绿素含量和Rubisco活性分别下降了96.7%,71.9%和80%。相比之下,在未添加乙酸钠的对照培养液中,实验结束时硝酸盐的浓度,叶绿素含量和Rubisco活性仅下降了47.2%,27.3%和4.4%,培养过程中没有虾青素积累。     

Extraction of astaxanthin from Haematococcus pluvialis by supercritical carbon dioxide fluid with ethanol modifier

Conventional solvent extraction methods cannot attain high‐quality antioxidant extracts from microalgae and also require solvent recovery and posttreatment. In this study, we utilized environmental friendly supercritical carbon dioxide fluid extraction (SFE‐CO₂) techniques to obtain pigment (i.e. astaxanthin) from Haematococcus pluvialis. The effects of key operating parameters on the extraction efficiency of astaxanthin were investigated, giving an optimal condition of H. pluvialis weight, 6.5 g; CO₂‐flow rate, 6.0 NL/min; extraction time, 20 min; extraction pressure, 4500 psi; volume of ethanol modifier added, 9.23 mL/g; extraction temperature, 50°C; modifier composition, 99.5%. Under these optimum conditions, the astaxanthin yield was 73.9% (10.92 mg/g dry H. pluvialis powder) after eight cycle of extraction cycles. The saponification index (C_S/C₀, representing the ratio of astaxanthin concentration after and before the saponification procedures) of the extract could be increased from 1 to 12.78 by saponification with 3.5 M NaOH.     

Enhancement of astaxanthin production from Haematococcus pluvialis under autotrophic growth conditions by a sequential stress strategy

Abstract

The study of microalgal culture has been growing in recent decades, because the cellular structure of microalgae has diverse highly valuable metabolites that have attract attention of numerous companies and research groups. The pigment astaxanthin is considered one of the most powerful antioxidants in nature. The microalga Haematococcus pluvialis was proposed as one of the best natural astaxanthin sources, because it can accumulate high amount of the pigment. In this work, we studied different stress treatments on H. pluvialis growth cultures as well as astaxanthin production under autotrophic growth conditions. The results showed that extending nitrogen starvation before increasing radiation intensity up to 110?μmol photons m^?2 s^?1 during late the palmella cell phase incremented the astaxanthin concentration up to 2.7% of dry biomass with an efficient light energy utilization during the stress stage.     

Cell wall proteomics of the green alga Haematococcus pluvialis (Chlorophyceae)

The green microalga Haematococcus pluvialis can synthesize and accumulate large amounts of the ketocarotenoid astaxanthin, and undergo profound changes in cell wall composition and architecture during the cell cycle and in response to environmental stresses. In this study, cell wall proteins (CWPs) of H. pluvialis were systematically analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) coupled with peptide mass fingerprinting (PMF) and sequence-database analysis. In total, 163 protein bands were analyzed, which resulted in positive identification of 81 protein orthologues. The highly complex and dynamic composition of CWPs is manifested by the fact that the majority of identified CWPs are differentially expressed at specific stages of the cell cycle along with a number of common wall-associated 'housekeeping' proteins. The detection of cellulose synthase orthologue in the vegetative cells suggested that the biosynthesis of cellulose occurred during primary wall formation, in contrast to earlier observations that cellulose was exclusively present in the secondary wall of the organism. A transient accumulation of a putative cytokinin oxidase at the early stage of encystment pointed to a possible role in cytokinin degradation while facilitating secondary wall formation and/or assisting in cell expansion. This work represents the first attempt to use a proteomic approach to investigate CWPs of microalgae. The reference protein map constructed and the specific protein markers obtained from this study provide a framework for future characterization of the expression and physiological functions of the proteins involved in the biogenesis and modifications in the cell wall of Haematococcus and related organisms.     

Abstracts of papers read at the winter meeting at the University of Bristol

Summaries of changes since 1965 in the phytoplankton fora of Lake Mälren, the third largest lake in Sweden, are presented. The phosphorus load was reduced by 90% in 1970–1973 and a fertilizer factory on the lake decreased its N-outlet by 80%. The phosphorus concentrations in the water decreased by up to 40% and the total nitrogen concentration by 50% between 1970 and 1983. But in one part of the lake the N-concentration increased by 50% due to drainage from surrounding land and atmospheric deposition. With changes in the ratios of N/P and Si/P as well as the turbidity of the water there have been drastic alterations in species composition and total phytoplankton biomasses.

There was a rise in the number of species over the period of study at all times of the year. Colonial species decreased while small and large flagellates of chrysophytes, cryptomonads and dinoflagellates increased. In the most polluted basins cyanobacterial blooms have halved their peak biomasses or have almost disappeared.

At times during the 19 years of the study the summer phytoplankton communities have been tested to fits of a log-normal distribution, assuming such a distribution to be valid in well-balanced ecosystems. The general trend indicates an increasing adjustment to a log-normal distribution. Phytoplankton changes in the lake are associated with effects known to cause reversed eutrophication, although climatically induced variations may also be identified.     

Encapsulation of Haematococcus pluvialis using chitosan for astaxanthin stability enhancement

Astaxanthin is receiving commercial interest due to its use as a preferred pigment in aquaculture feeds. Its antioxidant activity is approximately 100 times higher than that of β-tocopherol, and can be used as a potential agent against cancer. Astaxanthin can easily be degraded by thermal or oxidative processes during the manufacture and storage. In this study, astaxanthin and its biological activity were protected against oxidative environmental conditions by encapsulating the homogenized cells in chitosan. Haematococcus pluvialis were formed into beads, which were then coated with 5 layers of chitosan film, resulting in chitosan-algae capsules that have a mean diameter of 0.43 cm and the total film thickness of approximately 100 μm. No significant loss in the amount of astaxanthin content in H. pluvialis was found due to the process of encapsulation. However, approximately 3% loss of antioxidant activity of the H. pluvialis was observed after encapsulation. The results of stability under different storage conditions showed that although encapsulation caused 3% loss of antioxidant activity, the longer term stability of the dried algae biomass, beads, and capsules indicated that encapsulation of H. pluvailis in chitosan film was capable of protecting the algae cells from oxidative stresses.     

外源2,6-二叔丁基对甲酚对非生物胁迫条件下雨生红球藻虾青素积累的影响

【背景】雨生红球藻是天然虾青素的最佳来源,广泛应用于虾青素的工业化生产。【目的】探究外源添加不同浓度的2,6-二叔丁基对甲酚(Butylated hydroxytoluene,BHT)对雨生红球藻虾青素积累的影响,以期建立BHT提高雨生红球藻虾青素产量的技术体系。【方法】选用不含硝态氮的BBM培养基,辅以强光照,培养雨生红球藻(Haematococcus pluvialis)LUGU,测试不同浓度BHT对雨生红球藻生物量、虾青素含量、活性氧、抗氧化系统和虾青素合成相关酶基因的影响。【结果】在0-3 mg/L BHT范围内,2 mg/L BHT对雨生红球藻虾青素积累的促进效果最佳,达到31.66 mg/g。2 mg/L BHT有效降低了雨生红球藻内的活性氧水平,增加了细胞内NO水平,提高了藻细胞内过氧化氢酶(Catalase,CAT)、过氧化物酶(Peroxidase,POD)和超氧化物歧化酶(Superoxidedismutase,SOD)活性以及谷胱甘肽(Glutathione,GSH)的含量,诱导了虾青素合成关键酶基因chy和lcy的高效表达。【结论】非生物胁迫条件下,外源添加适量的BHT能促进雨生红球藻中虾青素的积累,且与藻细胞内的信号分子活性氧(Reactive oxygen species,ROS)、NO水平及虾青素合成相关基因的表达调控相关。     

温度对雨生红球藻(Haematococcus pluvialis)生物量和虾青素产量的影响

在用环形培养池模拟系统培养雨生红球藻的过程中,研究了温度对雨生红球藻生物量及虾青素产量的影响。结果表明,在15～25℃的范围内,不同温度下雨生红球藻生物量和虾青素含量及产量都经历了一个上升—最高—下降的过程。25℃与22℃时红球藻的虾青素产量、虾青素含量(干重)均显著高于其他温度的(P<0.01),但两者间差异不显著(P>0.05)。15℃时,红球藻生物量、虾青素含量和虾青素产量均最低,分别为1.4g、0.54%和2.49mg/L;25℃时,红球藻生物量和虾青素产量最高,分别为2.68g和13.53mg/L;22℃时,虾青素含量最高,为1.52%。