The past, present and future of phycology in China

Algae have been part of Chinese life for thousands of years. They are widely used as food and have been cited in Chinese literature as early as 2500 years ago. However, formal taxonomic studies on Chinese algae were initiated by foreign scientists only about 200 years ago, and by Chinese phycologists only about 90 years ago. This paper summarizes the history of modern phycological studies on Chinese algae and provides an overview of the achievements of phycological studies by Chinese scientists, especially on algal taxonomy, morphology, genetics, ecology and environmental research, physiology, biotechnology, algal culture, applied phycology and space phycology, in the last century. Recent development in phycological research focuses on algal floristic and molecular systematics, algal molecular biotechnology, applied phycology including micro and macroalgal cultivation and algal product development, and the roles of algae in environmental pollution control. These areas will also be the main focuses of Chinese phycological research in the foreseeable future.

Transient expression of lacZ in bombarded unicellular green alga Haematococcus pluvialis

This paper reports for the first time the transient expression of a reporter gene, LacZ, in the unicellular green alga Haematococcus pluvialis. By employing the micro-particle bombardment method, motile cells in the exponential phase showed transient expression of lacZ. This was detected in bombarded motile cells under the rupture-disc pressures of 3103 KPa and 4137 KPa. Transient expression of LacZ gene could not be observed in non-motile cells of this alga under the same transformation condition. No LacZ background was found in either the motile cells or the non-motile cells. The study suggests a promising potential of the SV40 promoter and the lacZ reporter gene in genetic engineering of unicellular green algae.

Isolation and purification of phycoerythrin from red alga Garcilaria verrucosa by expanded-bed-adsorption and ion-exchange chromatogaphy

R-phycoerythrin was isolated and purified from Gracilaria verrucosa on an expanded-bed adsorption column combined with ion-exchange chromatography, which can effectively solve the problem of blockage of chromatographic columns due to polysaccharides during isolation and purification of phycobiliproteins. 0.1 M (NH4)(2)SO4 proved best to elute R-phycoerythrin from the expanded-bed column, and desalted 0.1 M (NH4)(2)SO4 eluate was used on an ion-exchange column to purify the R-phycoerythrin. Using this two-stage chromatography, the purity (OD565/OD280) of the R-phycoerythrin from G. verrucosa is increased to 4.4, and the yield of purified R-phycoerythrin can reach 0.141 mg . g(-1) of the frozen alga.

Large-scale isolation and purification of R-phycoerythrin from red alga Palmaria palmata using the expanded bed adsorption method

R-phycoerythrin, a light-harvesting protein in some marine algae, and can be widely used in medicine, was isolated and purified from a red alga, Palmaria palmata (Lannaeus) Kuntze, using the streamline column (expanded bed adsorption) combined with ion-exchange chromatography. Because the crude extract was applied to the column upwardly, the column would not be blocked by polysaccharides usually very abundant in the extract of marine alga, this kind of blockage could hardly lie overcome in ordinary chromatographic column. After applying the crude extract containing 0.5 mol/L (NH4)(2)SO4, (NH4)(2)SO4 solution of different concentrations (0.2 mol/L, 0.1 mol/L and 0.05 mol/L) was used to elute the column downwardly and the eluates were collected and desalted. The desalted eluates were then applied onto all ion-exchange chromatographic column loaded with Q-sepharose for further purification of the R-phycoerythrin. Through these two steps, the purity (OD565/OD280) of the R-phycoerythrin from P. palmata was up to 3.5, more than 3.2, the commonly accepted criterion for purity, and the yield of the purified R-phycoerythrin could reach 0.122 mg/g of frozen P. palmata, much higher than that of phycobiliproteins purified with the previous methods. The result indicated that the cost of R-phycoerythrin will drop down with the method reported in this article.

Algal biotechnology industries and research activities in China

In old China there were very few people engaged in the study of the algae, but in new China, freshwater and marine algae are studied by over one hundred old and new phycologists. There is now an algal biotechnology industry consisting of an aquaculture industry, producing large amounts of the seaweeds Laminaria, Porphyra, Undaria, Gracilaria, eucheumoids, and the microalgae Dunaliella and Spirulina. There is also a phycocolloid industry, producing algin, agar and carrageenan; an industry producing chemicals and drugs, such as iodine, mannitol, phycocyanin, beta -carotene, PSS (propylene glycol alginate sulfate) and FPS (fucose-containing sulfated polysaccharides) and an industry producing food, feed and fertilizer. The Laminaria cultivation industry produces about 900,000 t dry Laminaria, probably the largest producer in the world and 13,000 t algin, undoubtedly one of the largest algin producer in the world.

Three-stage transformation of chlorophyll transient fluorescence pattern under sustained dehydration and the discovery of critical water content in seaweeds

The chlorophyll fluorescence kinetics of marine red alga Grateloupia turutunt Yamada, green alga Ulva pertusa Kjellm and brown alga Laminaria japonica Aresch during natural sustained dehydration were monitored and investigated. The pulse amplified modulation (PAM) system was used to analyze the distinct fluorescence parameters during thallus dehydration. Results proved that the fluorescence kinetics of different seaweed all showed three patterns of transformation with sustained water loss. These were: 1) peak kinetic pattern (at the early stage of dehydration fluorescence enhanced and quenched subsequently, representing a normal physiological state). 2) plateau kinetic pattern (with sustained water loss fluorescence enhanced continuously but quenching became slower, finally reaching its maximum). 3) Platform kinetic pattern (fluorescence fell and the shape of kinetic curve was similar to plateau kinetic pattern). A critical water content (CWC) could be found and defined as the percentage of water content just prior to the fluorescence drop and to be a significant physiological index for evaluation of plant drought tolerance. Once thallus water content became lower than this value the normal peak pattern can not be recovered even through rehydration, indicating an irreversible damage to the thylakoid membrane. The CWC value corresponding to different marine species were varied and negatively correlated with their desiccation tolerance, for example. Laminaria japonica had the highest CWC value (around 90%) and the lowest dehydration tolerance of the three. In addition, a fluorescence "burst" was found only in red algae during rehydration. The different fluorescence parameters F-o, F-v and F-v, F-m were measured and compared during water loss. Both F-o and F-v increased in the first stage of dehydration but F-v/F-m. kept almost constant. So the immediate response of in vivo chlorophyll fluorescence to dehydration was an enhancement. Later with sustained dehydration F-o increased continuously while F-v decreased and tended to become smaller and smaller. The major changes in fluorescence (including fluorescence drop during dehydration and the burst during rehydration) were all attributed to the change in F-o instead of F-v This significance of F-o indicates that it is necessary to do more research on F-o as well as on its relationship with the state of thylakoid membrane.

An acidic polysaccharide with xylose branches from Porphyra yezoensis

An acidic polysaccharide (PY3) was isolated from the hot water extract of the red algae Porphyra yezoensis by successive column chromatographies over DEAE-cellulose and Sephadex G-200. PY3 with an average molecular weight of 1.8x10(5) was demonstrated to be composed of galactose (Gal), 3,6-anhydrogalactose (3,6-AnGal), 6-OSO3-galactose (6-OSO3-Gal) and xylose (Xyl) in an approximate molar ratio of 25 : 15, 10, 1. In view of Smith degradation and methylation and on the basis of spectral evidence including those of IR, GC, GC-MS, and H-1 and C-13 NMR, the most probable repeating unit of PY3 could be proposed as [(1-->3)beta -D-Gal(1 --> 4)alpha -L-3,6-AnGal](3)-[(1 --> 3)beta -D-Gal(1 --> 4)alpha -L-6-OSO3-Gal](2) with a xylose moiety at the C-6 of one of every twenty-five beta -D-Gal residues. To our knowledge, PY3 was shown to be the first porphyran possessing occasional xylose branches.

Chemical species of iodine in some seaweeds II. Iodine-bound biological macromolecules

The distribution of iodine in various biological macromolecules in Sargassum kjellmanianum was studied using neutron activation analysis combined with chemical and biochemical separation techniques. The results indicate that iodine is mainly bound with protein, part of iodine with pigment and polyphenol, and little with polysaccharides, such as algin, fucoidan and cellulose. This result is significant for the mechanism of enriching iodine of algae and utilization of alga iodine.

Biological effect of R-phycoerythrin-mediated photosensitization on DNA.

R-phycoerythrin (R-PE) is one of important proteins involved in capturing light during photosynthesis in red algae, and it is highly fluorescent, and water-soluble chromophores. In vivo, it can transfer the light energy into photosynthetic center, however, it can deliver the captured light energy captured to the surrounding oxygen in vitro and produce reactive oxygen species such as singlet oxygen, which is toxic to tumor cells. R-PE was added to the culture medium of tumor cells, subsequently with irradiation of 488 nm, Argon laser of 25.6 J/cm(2). The result by MTT assay showed that the survival rate decreased with the increase of R-PE concentration from 1 to 100 mg/L. The result from H-3-TdR incorporation demonstrated that the synthesis of DNA reduced when the concentration of R-PE increased from 0.01 to 0.32 mg/L. Besides, pUC18 DNA showed a conversion from supercoiled into linear conformation. The conclusion comes that R-PE mediated PDT can influence the conformation of DNA, and it may be one of the mechanisms of R-PE mediated photodynamic therapy.

Diversity of pigment-protein complexes of photosynthetic organisms.

PS I, PS II and light-harvesting complexes (LHC) in oxygen evolving photosynthetic organisms were reviewed. These organisms include cyanobacteria, red algae, brown algae, diatoms, chrysophytes, dinophytes, xanthophytes, crypophytes, green algae and green plants. The diversity of pigment-protein complexes that fuel the conversion of radiant energy to chemical bond energy was highlighted, and the evolutionary relationships among the LHC structural polypeptides and the characteristics of the fluorescence emission of PS I at 77 K was discussed.

Assessment of calcareous alga Corallina pilulifera as elemental provider

Calcareous algae such as Corallina pilulifera a Postels et Ruprecht can be one of the most potential candidates to be used in biological carbon dioxide assimilation to reduce greenhouse effect because of its calcification capacity as well as photosynthesis if utilized extensively and properly. The major elemental composition in C. pilulifera is as follows: sodium 0.13%, chloride 1.75%, magnesium 4.37%, calcium 18.4%, iron 0.31%, and carbonate 28.5%. Calcareous algae can be used as elemental provider for livestock or agriculture. (C) 1999 Elsevier Science Ltd. All rights reserved.

A new model of phycobilisome in Spirulina platensis

Phycobilisomes (PBS) were isolated from blue-green alga Spirulina platensis. Scanning tunneling microscope was used to investigate the three-dimensional structure of PBS deposited on freshly cleaved highly oriented pyrolytic graphite (HOPG) in ambient condition at room temperature. The results showed that the rods of PBS radiated from the core to different directions in the space other than arrayed in one plane, which was different from the typical hemi-discoidal model structure. The diameter of PBS was up to 70 nm, and the rod was approximately 50 nm in length. Similar results were observed in Langmuir-Blodgett (LB) film of PBS. The dissociated PBS could reaggregate into rod-like structures and easily form two-dimensional membrane while being absorbed on HOPG, however, no intact PBS was observed. The filling-space model structure of PBS in Spirulina platensis with STM from three-dimensional real space at nanometer scale was found, which showed that this new structural model of PBS surely exists in blue-green algae and red algae. The function of this structural model of PBS was also discussed.

Integrated cultivation of the red alga Kappaphycus alvarezii and the pearl oyster Pinctada martensi

The effect of simultaneously cultivating the pearl oyster Pinctada martensi and the red alga Kappaphycus alvarezii on growth rates of both species was investigated in laboratory and field studies conducted from December 1993 to June 1995. The two study sites were in subtidal areas 100 km apart off the east coast of Hainan Island, China. Pearl oysters were cultivated in the center of an algal farm and red alga was cultivated in the center of the pearl oyster farm. These field experiments showed higher growth rates of both P. martensi and K. alvarezii in a co-culture system than in a monospecies culture system. Laboratory studies showed that the algae removed nitrogenous wastes released by pearl oysters. Algae treated with pearl oyster wastes grew much faster than those without oyster wastes. Algae treated with the seawater to which NH4Cl, NaNO3 and NaNO2 were added grew at the same rate as those treated with natural seawater containing oyster nitrogenous wastes, suggesting that enhanced growth of algae in the co-culture system was largely due to nitrogenous metabolites of the pearl oysters. In the co-culture, growth of pearl oysters was positively influenced by the presence of rapidly growing algae but when seawater temperature decreased below 20 degrees C, the algae grew slowly and there was no measurable benefit of mixed culture to either algae or pearl oyster.

Purification of High-quality Plastid DNA from the Brown Alga Laminaria japonica Sporophytes

Extracting DNA from a variety of algae is rather difficult because of high levels of polysaccharides, tannins, and phenolics as these interfere with DNA isolation and downstream applications. High-quality plastid DNA (ptDNA) purification is particularly difficult because of its small proportion in total genomic DNA. This report describes an improved protocol for ptDNA purification that efficiently produces high-quality ptDNA from sporophytes of Laminaria japonica and several other algae. This improved protocol simplifies procedures for ptDNA purification and improves yield to 150-200 mu g of ptDNA per 100 g of frozen algal tissue. Polymerase chain reaction (PCR) amplification of conserved sequences has been used to verify purity of the ptDNA product.

Phylogeography of the Northern Atlantic species Chondrus crispus (Gigartinales, Rhodophyta) inferred from nuclear rDNA internal transcribed spacer sequences

Eighteen isolates of the red algae Chondrus crispus were collected from Northern Atlantic sites, together with C. ocellatus, C. yendoi and C. pinnulatus from the North Pacific. The nuclear rDNA internal transcribed spacer (ITS) was sequenced and compared, spanning both the ITS regions and the 5.8S rRNA gene. Percentage of nucleotide variation for C. crispus ranged from 0.3% to 4.0%. Phylogenetic analyses were performed using maximum parsimony (MP), neighbor-joining (NJ) and minimum evolution methods. They showed that two main clades existed within the C. crispus samples examined and that suggested C. crispus had a single Atlantic origin. The clustering however did not follow the geographic origin. We hypothesized that the current distribution of C. crispus populations might be a result of three main factors: temperature boundaries, paleoclimate and paleoceanography. ITS data exhibited abundant molecular information not only for phylogeographical investigation but also for systematics studies.