Cultivation of the brown alga Hizikia fusiformis (Harvey) Okamura: stress resistance of artificially raised young seedlings revealed by chlorophyll fluorescence measurement

Commercial farming of the intertidal brown alga Hizikia fusiformis (Harvey) Okamura in China and South Korea in the sea depends on three sources of seedlings: holdfast-derived regenerated seedlings, young plants from wild population and zygote-derived seedlings. Like many successfully farmed seaweed species, the sustainable development of Hizikia farming will rely on a stable supply of artificial seedlings via sexual reproduction under controlled conditions. However, the high rate of detachment of seedlings after transfer to open sea is one of the main obstacles, and has limited large-scale application of zygote-derived seedlings. To seek the optimal condition for growing seedlings on substratum in land-based tanks for avoidance of detachment in this investigation, young seedlings were grown in both outdoor tanks exposed directly to sunlight and in indoor raceway tanks in reduced, filtered sunlight. Results showed that young seedlings, immediately after fertilization, could withstand a daily fluctuation of direct solar irradiance up to a level of 1800 mu mol photons m(-1)s(-1), and maintained a faster growth rate than seedlings grown in indoor tanks. Detailed experiments by use of chlorophyll fluorescence measurements further demonstrated that the overnight (12 h) recovery of optimal fluorescence quantum yield (F-v/F-m) of seedlings after 1 h treatment at 40 degrees C was 98%, and the 48 h recovery of F-v/F-m of seedlings after 1 h exposure to 1800 mu mol m(-2)s(-1) was 92%. Forty-one-day-old seedlings showed no significant decrease of optimal fluorescence quantum yield at salinity ranging from 30 to 5 ppt for a treatment up to 17 h. Six-hour desiccation treatment did not have any influence on the optimal fluorescence quantum yield. Exposure to 18 mmol L-1 sodium hypochlorite for 10 min did not damage the PSII efficiency, and thus could be used to remove epiphytic algae. The strong tolerance of young seedlings to high temperature, high irradiance, low salinity and desiccation found in this investigation supports the view that mass production of Hizikia seedlings should be performed in ambient light and temperature instead of in shaded greenhouse tanks.

Temperature and light tolerance of representative brown, green and red algae in tumble culture revealed by chlorophyll fluorescence measurements

Laminaria japonica, Undaria pinnatifida, Ulva lactuca, Grateloupia turuturu and Palmaria palmata are Suitable species that fit the requirements of a seaweed-animal integrated aquaculture system in terms of their viable biomass, rapid growth and promising nutrient uptake rates. fit this investigation, the responses of the optimal chlorophyll fluorescence yield of the five algal species in tumble Culture were assessed at a temperature range of 10 similar to 30 degrees C. The results revealed that Ulva lactuca was the most resistant species to high temperature, withstanding 30 degrees C for 4 h without apparent decline in the optimal chlorophyll fluorescence yield. While the arctic alga Palmaria palmata was the most vulnerable one, showing significant decline in the optimal chlorophyll fluorescence yield at 25 degrees C for 2 h. The cold-water species Laminaria japonica, however, demonstrated strong ability to cope with higher temperature (24 similar to 26 degrees C) for shorter time (within 24 h) without significant decline in the optimal chlorophyll fluorescence yield. Grateloupia turuturu showed a general decrease in the optimal chlorophyll fluorescence yield with the rising temperature from 23 to 30 degrees C, similar to the temperate kelp Undaria pinnatifida. Changes of chlorophyll fluorescence yields of these algae were characterized differently indicating the existence of species-unique strategy to cope with high light. Measurements of the optimal chlorophyll fluorescence yield after short exposure to direct solar irradiance revealed how long these exposures could be without significant photoinhibition or with promising recovery in photosynthetic activities. Seasonal pattern of alternation of algal species in tank culture in the Northern Hemisphere at the latitude of 36 degrees N was proposed according to these basic measurements.

Temperature tolerance of young sporophytes from two populations of Laminaria japonica revealed by chlorophyll fluorescence measurements and short-term growth and survival performances in tank culture

The cold-water subtidal brown alga Laminaria japonica has been commercially fanned in the Far East and has been on top of all marine-fanned species in terms of farming area and annual output worldwide. The successful trials of transplantation of young sporophytes from the north to the south in winter along the Chinese coast in the 1950s led to the spreading of cultivation activities down to a latitude of 25-26 degrees N. Up to today, nearly 50% of the annual output of this farmed alga, as a cold-water species, comes from the sub-tropical south in China. The demand to have high-temperature-tolerant strains/ecotypes in farming area calls for a practical method to judge and select the desired parental plants for breeding programs and for seedling production. In this paper, we report our results on using chlorophyll fluorescence measurement and short-term growth performance in tank culture to estimate the temperature tolerance of offspring from two populations, Fujian Farmed Population (FFP) sampled from Fujian province (latitude: 25-26 degrees N) in subtropical area and Qingdao Wild Population (QWP) sampled from Qingdao (latitude: 36 degrees N). Contrary to what has been usually thought, the results revealed that offspring from Qingdao wild population in the north showed better performance both in short-term growth and survival rates and in optimal quantum efficiency (F-v/F-m) when exposed to higher temperature (20-25 degrees C). This result was further confirmed by fluorescence quenching analysis. QWP distributed along the southern distribution limit at a latitude of 36 degrees N in the Pacific west coast is thus taken as a more ideal one than the fanned population in subtropical region as a source of parental plants for breeding high-temperature-tolerant varieties. (c) 2006 Elsevier B.V. All rights reserved.

Circadian rhythms in the growth and reproduction of the brown alga Undaria pinnatifida and gametogenesis under different photoperiods

Circadian growth rhythm of the juvenile sporophyte of the brown alga Undaria pirznatifida was measured with the computer-aided image analysis system in constant florescent white light under constant temperature (10 degrees C). The growth rhythm persisted for 4 d in constant light with a free-running period of 25. 6 h. Egg release from filamentous gametophytes pre-cultured in the light - dark regime was evaluated for six consecutive days at fixed time intervals in constant white light and 12 h light per day. Egg release rhythm persisted for 3 d in both regimes, indicating the endogenous nature. Temporal scale of egg release and gametogenesis in 18, 16, 12 and 8 h light per day were evaluated respectively using vegetatively propagated filamentous gametophytes. Egg release occurred 2 h after the onset of dark phase and peaked at midnight. Evaluation of the rates of oogonium formation, egg release or fertilization revealed no significant differences in four light-dark regimes, indicating; the great plasticity of sexual reproduction. No photoperiodic effect in gametogenesis in terms of oogonium formation and egg release was found, but fertilization in short days was significantly higher than in long days. Results of this investigation further confirmed the general occurrence of circadian rhythms in inter-tidal seaweed species.

Evaluation of 28 marine algae from the Qingdao coast for antioxidative capacity and determination of antioxidant efficiency and total phenolic content of fractions and subfractions derived from Symphyocladia latiuscula (Rhodomelaceae)

The extracts obtained from 28 species of marine algae were evaluated for their antioxidant activity (AA) versus the positive controls butylated hydroxytoluene (BHT), gallic acid (GA), and ascorbic acid (AscA). Most of the tested samples displayed antioxidant activity to various degrees. Among them, the extract of Symphyocladia latiuscula exhibited the strongest AA, which was comparable to BHT, GA, and AscA in radical scavenging activity, as shown in the DPPH (alpha,alpha-diphenyl-beta-picrylhydrazyl) assay, and higher than those of the positive controls in beta-carotene-linoleate assay system. In addition, the ethyl acetate-soluble fraction isolated from the crude extract of S. latiuscula exhibited the highest antioxidant activity in both assay systems. This fraction was further fractionated into seven subfractions (F1-F7) by vacuum liquid chromatography (VLC). F1 and F4 were found to be the most effective subfractions in scavenging DPPH radical assay and in the beta-carotene-linoleate assay, respectively. The total phenolic content (TPC) and reducing power (RP) for all of the extracts, fractions, and subfractions (F1-F7) were also determined. The TPC of the 28 extracts ranged from 0.10 to 8.00 gallic acid equivalents (mg/g seaweed dry weight) while the RP ranged from 0.07 to 11.60 ascorbic acid equivalents (mg center dot g(-1) seaweed dry weight). Highly positive relationships between AA and TPC as well as between AA and RP were found for the extracts and fractions, while for the subfractions F1-F7 only weak or no such relations were found. The results obtained from this study indicate that further analysis is needed of those marine algal species that contain the most antioxidant activity in order to identify the active principles.

Composition, Abundance, and Diversity of the Peracarida on Different Vegetation Types in the Qi'ao-Dan'gan Island Mangrove Nature Reserve on Qi'ao Island in the Pearl River Estuary, China

Chang-Fu Wang, Xian-Qiu Ren, and Run-Lin Xu (2010) Composition, abundance, and diversity of the Peracarida on different vegetation types in the Qi'ao-Dan'gan Island Mangrove Nature Reserve on Qi'ao Island in the Pearl River estuary, China. Zoological Studies 49(5): 608-615. Almost nothing is known about the Peracarida in the Pearl River estuary. This is the 1st report to study the composition, abundance, and diversity of the Peracarida in the Qi'ao-Dan'gan I. Mangrove Nature Reserve on Qi'ao I., in the Pearl River estuary, southern China. Bimonthly samplings were carried out in 3 representative vegetation types (mangrove arbor, emergent plants, and seaweed) for 2 yr. Using a Peterson grab, 1940 individuals (id.) were collected in total, including 11 species of 6 genera, 5 families, and 3 orders (Amphipoda, Isopoda, and Tanaidacean). Discapseudes mackiei Bamber 1997 was the dominant species with the highest density of 1,432 incl./m(2). The effect of temperature on the abundance of Peracarida was significant (p < 0.01), and the optimum temperature was 22-23 degrees C in both the mangrove arbor and seaweed. The results showed that the abundance of the Peracarida was higher in the mangrove arbor, while the diversity, especially Amphipoda diversity, was higher in the seaweed. In contrast, emergent plants provided no suitable habitats for the Peracarida. http://zoolstud.sinica.edu.tw/Journals/49.5/608.pdf

Effects of temperature and irradiance on early development of Chondrus ocellatus Holm (Gigartinaceae, Rhodophyta)

Chondrus is a type of commercially produced red seaweed that widely used for food and carrageen extraction. Although the natural life history of the alga had been well understood, the factors influencing development of the tetraspore and carpospore remain poorly understood. In the perspective of seedling resources, the regulation of early development is crucial for the seedling nursing; therefore, it is necessary to understand the physiological influences during its early development. In this study, we studied the effects of temperature and irradiance on the early development of Chondrus ocellatus Holm under laboratory conditions. The released tetraspores and carpospores were cultivated at different temperatures (10-28 degrees C) and irradiances ( 10, 60 mu mol photons m(-2)s(-1)) with a photoperiod of 12L:12D. The results indicate that both tetraspores and carpospores are tolerant to temperatures of 10-25 degrees C, and have the highest relative growth rate at 20 degrees C. Irradiance variances influenced the growth of the discoid crusts, and the influence was more significant with increasing temperature; 60 mu mol photons m(-2)s(-1) was more suitable than 10 mu mol photons m(-2)s(-1). The optimum temperature and irradiance for the development of seedlings was 20 degrees C and 60 mu mol photons m(-2)s(-1), respectively.

Cultivation of the brown alga Hizikia fusiformis (Harvey) Okamura: controlled fertilization and early development of seedlings in raceway tanks in ambient light and temperature

Commercial cultivation of the dioecious brown macroalga Hizikia fusiformis (Harvey) Okamura in East Asia depends on the supply of young seedlings from regenerated holdfasts or from wild population. Recent development of synchronized release of male and female gametes in tumble culture provides a possibility of mass production of young seedlings via sexual reproduction. In this paper, we demonstrate that controlled fertilization can be efficiently realized in ambient light and temperature in a specially designed raceway tank in which the sperm-containing water has been recirculated. The effective fertilization time of eggs by sperm was found to be within six hours. Fast growth and development of the young seedlings relied on the presence of water currents. Velocity tests demonstrated that young seedlings of 2-3 mm in length could withstand a water current of 190 cm s(-1) stop without detachment. Culture experiments at 24 h postfertilization showed that elongation of both the seedlings and their rhizoids were not hampered by high irradiance up to 600 mu mol photons m(-2) stop s(-1) stop. However, growth was slightly retarded if cultured at a temperature of 16 degrees C compared to other culture temperatures of 22, 25 and 29 degrees C. No seedling detachment was observed after transfer of the young seedlings to raft cultivation in the sea after one and 1.5 months post-fertilization, indicating the feasibility of obtaining large quantity of seedlings in such a system.

Evidences of the intertidal red alga Grateloupia turuturu in turning Vibrio parahaemolyticus into non-culturable state in the presence of light

Grateloupia turuturu, previously known as Grateloupia doryphora, has been widely reported to be an invasive algal species. There are no studies to relate the impact of its existence on its surrounding environment. In this paper, we present our results to show that about 70% of individuals collected from the field could turn Vibrio parahaemolyticus into non-culturable state on both selective (TCBS) and non-selective (2216E) culture medium in 24 h in the presence of light in live algal culture. Total bacteria counts on TCBS and 2216E plates dropped from the initial 565 (174) and 1192 (60) cfu ml(-1) respectively to zero in 24 h. This effect disappeared when the alga was grown in darkness. The same effect was not found in two other intertidal macroalgae Laminaria japonica and Palmaria palmata. Further tests showed that the settlement ability of bacteria in seawater was impaired significantly in the presence of this alga in comparison with three other algal species. (c) 2006 Elsevier B.V. All rights reserved.

The dominant Ulva strain of the 2008 green algal bloom in the Yellow Sea was not detected in the coastal waters of Qingdao in the following winter

The region of Qingdao, China, experienced the world's largest green tide from May to July 2008. More than one million tons of fresh algal biomass of the green alga Ulva prolifera was harvested, while more was suspected to have sunk to the bottom. The original source of this seaweed was suspected to be from the south as revealed by satellite images. The floating biomass drifted with the water current northward and flourished in nearshore waters around Qingdao. However, direct biological evidence for "seed" source is lacking. It is still unclear whether this alga could survive the Qingdao local coastal environment and pose future danger of potential blooming. Systematic and seasonal sampling of waters in the intertidal zone at six collection sites along the Qingdao coast was conducted from December 2008 to April 2009. Forty-eight water samples were analyzed. From these, nine different morphotypes of Ulva were grown in the laboratory under standard temperature and light regimes. Growth of Ulva was observed in all water samples. However, molecular phylogenetic analyses revealed that the dominant U. prolifera strain of the 2008 bloom was absent in all the water-derived cultures during the sampling period. These results provide evidence that the dominant bloom-forming alga was unlikely able to survive the coastal waters (the minimal surface water temperature in February is 2A degrees C) in winter conditions in Qingdao, even though all the sampling locations were heavily covered by this alga in June 2008.

A comparative study of thermolysis characteristics and kinetics of seaweeds and fir wood

Thermal analysis and thermolysis kinetics of three kinds of seaweeds and fir wood (M. glyptostriboides Huet Cheng), a kind of typical land plant, had been conducted. The results showed that thermal stability follows the order of Grateloupia filicina < Ulva lactuca < Dictyopteris divaricata < fir wood. A notable difference on heat flow between seaweeds and fir wood during thermolysis was that the former were mainly connected with exothermic processes at relatively lower temperature regimes. while the latter was connected with an apparent endotherm at a relatively higher temperature regime followed by a maximum exothermic peak. This suggested that the heat coupling might be realized if co-thermolysis of seaweeds and fir wood were carried out. The main devolatilization phase of each seaweed could be described by Avrami-Erofeev equation, which indicated that thermolysis of seaweeds follows the mechanism of random nucleation and nuclei growth, whereas that of fir wood by Z-L-T equation and its thermolysis mechanism was three-dimensional diffusion. The activation energies calculated for both seaweeds and fir wood increase as conversion increases. However, those for the former have wider distribution. (c) 2006 Elsevier Ltd. All rights reserved.

The genetic analysis and germplasm identification of the gametophytes of Undaria pinnatifida (Phaeophyceae) with RAPD method

Eleven pairs of Undaria pinnatifida (Harv.) Suringar gametophytes were identified with random amplified polymorphic DNA (RAPD) technique. After screening 100 primers, 20 ten-base primers were determined for the RAPD analysis. A total of 312 polymorphic loci were obtained, of which 97.7% were polymorphic. The primer S198 was found to distinguish all the selected Undaria pinnatifida gametophytes. The genetic distances between each two of the twenty-two U. pinnatifida gametophytes ranged from 0.080 to 0.428, while the distances to the Laminaria was 0.497 on average. After reexamination, two sequences characterized amplification region (SCAR) markers were successfully converted, which could be applied to U. pinnatifida germplasm identification. All these results demonstrated the feasibility of applying RAPD markers to germplasm characterization and identification of U. pinnatifida gametophytes, and to provide a molecular basis for Undaria breeding.

Growth of Gracilaria lemaneiformis under different cultivation conditions and its effects on nutrient removal in Chinese coastal waters

Gracilaria lemaneiformis (Bory) Daws has been extensively cultivated as a source of commercial agar and the ecomaterials in Shenao Bay, Guangdong Province, Jiaozhou Bay, Shandong Province and other waters in China. This paper examines the in situ suspended farming of G. lemaneiformis using raft cultivation under different conditions and its effects on nutrient removal in the laboratory. The results showed that cultivated Gracilaria grew well in both Shenao Bay and Jiaozhou Bay. The biomass of Gracilaria increased from 50 to 775 g m(-1) (fresh weight) during 28 days, with special growth rate (SPG) 13.9% d(-1) under horizontal cultivation in Jiaozhou Bay. Light, temperature, nutrient supply, as well as cultivation treatments such as initial density, and depth of suspension seaweed were important to the growth of Gracilaria. The highest biomass production was observed in the horizontal culture condition (0.0 m) and 0.5-1.5 m deep layer in Jiaozhou Bay. However, the highest growth rate in Shenao Bay appeared under the lowest initial stocking density treatment. In the laboratory, the aquarium experiments (fish and seaweed culture systems) demonstrated that Gracilaria was able to remove inorganic nutrients effectively. The concentration of NH4+-N decreased by 85.53% and 69.45%, and the concentration of PO4-P decreased 65.97% and 26.74% in aquaria with Gracilaria after 23 days and 40 days, respectively. The results indicate that Gracilaria has the potential to remove excess nutrient from coastal areas, and the large-scale cultivation of G. lemaneiformis could be effective to control eutrophication in Chinese coastal waters. (c) 2005 Elsevier B.V. All rights reserved.

贝藻混养系统中贝类食物来源的定量分析

应用稳定碳、氮同位素比值法和脂肪酸标志分析法，较为系统地研究了贝藻混养系统中滤食性贝类的食物来源，评估了大型藻类对混养系统及滤食性贝类的物质贡献。主要研究结果如下： 1.综述了典型生态系统中大型藻类和滤食性贝类各自的生态学地位和作用，大型藻类与滤食性贝类不仅在水体营养盐方面存在互利关系，二者在物质循环与收支方面同样具有耦合性，大型藻类提供的颗粒态有机质可以为滤食性贝类提供饵料来源。 2.总结了稳定同位素比值法和脂肪酸标志法在海洋生态系统食物来源及食物网分析中的应用，并建立了两种方法的具体操作规程。 3.分析了栉孔扇贝Chlamys farreri和海带Laminaria japonica混养系统中海带碎屑形成及释放不同阶段的生态学特征，评估了碎屑对扇贝的饵料贡献。海带在6周内释放了自身约27％的碳；碎屑形成及释放过程中C:N比值显著下降，同时伴随着旺盛的细菌降解，碎屑中也发现有大量硅藻类和原生动物存在。稳定同位素分析证实海带碎屑是混养期间扇贝的主要食物来源。 4.查明了春季胶州湾潮间带自然分布的长牡蛎Crassostrea gigas、紫贻贝Mytilus galloprovincialis和湾内浅海筏式养殖栉孔扇贝的可能食物来源。湾内栉孔扇贝饵料组成中浮游硅藻类为最主要部分，同时混杂有陆源有机质和细菌类物质；潮间带自然生长的牡蛎和贻贝饵料组成中，浮游植物占86.2-89.0％，种类组成中除硅藻外还包括一定比例的金藻和甲藻类；潮间带繁盛的孔石莼Ulva pertusa藻床为两种贝类提供了8.7-11.0％的补充食物来源。 5.揭示了桑沟湾贝藻混养海区春、夏季栉孔扇贝饵料来源组成情况及其季节变化，评估了海带养殖区碎屑碳量季节变化及海带来源碳对扇贝组织碳的贡献。结果表明，湾内贝藻混养区碎屑碳量为75.52-265.19 μg l-1，其在水体总颗粒态有机碳中的比例为25.6-73.8％。海带来源碎屑碳对栉孔扇贝组织碳的贡献比例为14.1-42.8％，且与水体碎屑碳比例的季节变化存在极显著相关性(F=0.992, P=0.004)。5月份湾外海带养殖区水体碎屑碳量为110.12-144.71 μg l-1，显著高于湾内无海带区（75.52 μg l-1），湾外养殖的扇贝组织中海带来源碳比例为22.0-24.1％，显著高于湾内单养区扇贝（9.6％）。估算结果表明，桑沟湾每年收获的6967吨（总湿重）栉孔扇贝中，海带提供了约57.1吨碳，换算为海带干物质为219.6吨。脂肪酸标志分析结果表明，2月份至8月份硅藻类在扇贝饵料组成中比例逐渐下降，而细菌类比例逐渐升高。整个采样期间，EPA/DHA比例较低，说明扇贝饵料组成中可能包括高DHA含量的组分。

羟基自由基降解琼胶及其产物的活性研究

琼胶是一种从石花菜等红藻中提取的，目前生产工艺和结构等方面研究比较成熟的海藻多糖，广泛应用于医药、仪器等行业。但是，海藻多糖因为具有分子量大，粘度大，溶解度较小的等特点，而使其应用范围受到限制。利用降解的手段对其进行修饰，降低分子量和粘度，改善溶解性，可以拓展其应用范围。并且根据文献报道，琼 胶寡糖具有一些特殊的生物活性，如抗氧化性，抗炎症等。因此，对琼胶降解的研究具有生要意义。本研究中，为了选择一种合适的降解方法，进行了几种水解方法的尝试，其中包括在不同湿度和酸度下盐酸水解，过氧化氢和醋酸催化水解，Fenton体系羟基自由基降解。对于酸水解和Fenton体系氧化还原降解方法，通过粘度法对反应的速度进行了比较，表明氧化还原降解反应中琼胶的粘度降低比较快，并且具有代表性和新意，确定为本实验的降解琼胶的方法并对氧化还原降解所得的产物进行了活性实验。通过模仿自然界普遍存在的氧化还原降解反应，利用Vc诱导的Fenton体系产生的羟基自由基氧化还原降解琼胶得到低分子量的琼胶。降解产物经过高速离心、60％乙醇沉淀，除去分子量比较大的降解产物和磷酸盐，得到可溶于60％乙醇的分子量估计小于3000的降争产物，其产率为85％。利用经Sephadex-G25凝胶色谱分离所香的不同分子量的级分进行分子量和α－葡萄糖苷酶抑制活性关系的实验。降解产物对α－葡萄糖苷酶的抑制率和各级分的浓度呈线性正相关，并且各级分的IC_(50)则随着分子量的降低而降低。另外，对所得的降解产物混合物进行了红外吸收光谱、质子去偶核磁共震碳谱和负离子基质辅助激光诱导－飞行时间质谱结构分析。结果表明，氧化还原降解反应的专一性差，在得到寡糖的同时，在光谱图中出现一些比较复杂的副产物的结构信息。最后，根据MTT法的原理，以有体皮肤成纤维细胞为材料，通过紫外线辐射产生自由基造成氧化损伤，研究降解产物对成纤维细胞的保护作用。当无紫外线辐射时，降解产物对成纤维细胞具有显著的促进生长增殖作用：当经UVa、UBb辐射时则可以显著地表现出对损伤的保护作用，并且这种促进生长和保护作用呈显著的量效关系，表明降解产物具有清除基自由基的作用。但是，因为氧化还原降解以应的机理尚不十分明的以及琼羟胶的特殊结构，使得反应的副产物很难预测，也就使得分离工作难以进行，所以，根据目前所得的信息，尚不能确定是降解产物的什么级分产生的以上两种生物活性。