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The prepared mesoporous silica SBA-15 was modified by doping with La by an incipient wet impregnation method. XRD, TEM, NMR and FT–IR were used to evaluate the influence of La doping amounts on the structure and hydrothermal stability of SBA-15. The results showed that the introduction of La could significantly improve the hydrothermal stability of the mesoporous silica SBA-15. The silanol groups on the defect sites of pore surface are the main place for lanthanum to enter. After being treated with steam at 750 °C for 40 h, La-SBA-15 with appropriated amount of La loading maintained a highly ordered structure. In addition, a mesoporous Ni/5La-SBA-15 catalyst was prepared by an incipient wet impregnation method, and steam reforming of methane was carried out on this catalyst for 100 h at 750 °C with CH4:H2O:N2 = 4:12:1. During the reaction, the CH4 conversion rate remained unchanged, and the catalyst activity did not decrease, confirming that La-SBA-15 can be used as a hydrothermally stable support for catalytic reactions contained steam.

Microalgae are known to respond to salinity stress via mechanisms that include accumulation of compatible solutes and synthesis of antioxidants. Here, we describe a salinity-tolerance mechanism mediated by lipid droplets (LDs). In the alga Parachlorella kessleri grown under salt-stress conditions, we observed significant increases in cell size and LD content. LDs that were closely grouped along the plasma membrane shrank as the plasma membrane expanded, and some LDs were engulfed by vacuoles. Transcriptome analysis showed that genes encoding lysophospholipid acyltransferases (LPLATs) and phospholipase A2 were significantly up-regulated following salt stress. Diacylglycerol kinase and LPLAT were identified in the proteome of salt-induced LDs, alongside vesicle trafficking and plastidial proteins and histone H2B. Analysis of fatty acid composition revealed an enrichment of C18:1 and C18:2 at the expense of C18:3 in response to salt stress. Pulse-chase experiments further suggested that variations of fatty acid composition were associated with LDs. Acetate stimulation research further confirmed a positive role of LDs in cell growth under salt stress. These results suggest that LDs play important roles in salt-stress tolerance, through harboring proteins, participating in cytoplasmic component recycling, and providing materials and enzymes for membrane modification and expansion.

Saline-alkali soil has become an important environmental problem for crop productivity. One of the most effective approaches is to cultivate new stress-tolerant plants through genetic engineering. Through RNA-seq analysis and RT-PCR validation, a novel bZIP transcription factor ChbZIP1, which is significantly upregulated at alkali conditions, was obtained from alkaliphilic microalgae Chlorella sp. BLD. Overexpression of ChbZIP1 in Saccharomyces cerevisiae and Arabidopsis increased their alkali resistance, indicating ChbZIP1 may play important roles in alkali stress response. Through subcellular localization and transcriptional activation activity analyses, we found that ChbZIP1 is a nuclear-localized bZIP TF with transactivation activity to bind with the motif of G-box 2 (TGACGT). Functional analysis found that genes such as GPX1, DOX1, CAT2, and EMB, which contained G-box 2 and were associated with oxidative stress, were significantly upregulated in Arabidopsis with ChbZIP1 overexpression. The antioxidant ability was also enhanced in transgenic Arabidopsis. These results indicate that ChbZIP1 might mediate plant adaptation to alkali stress through the active oxygen detoxification pathway. Thus, ChbZIP1 may contribute to genetically improving plants’ tolerance to alkali stress.

Intrauterine adhesion (IUA) is a serious complication caused by excessive fibrosis resulting from endometrial repair after trauma. The traditional Chinese medicine Tiaoshen Tongluo recipe (TTR) contains ingredients associated with the alleviation of fibrosis. The transforming growth factor-β1 (TGF-β1)/Smad pathway is thought to mediate fibrosis in IUA. In this study, we evaluated the influence of TTR on endometrial fibrosis in a rat model of IUA and in TGF-β1-stimulated endometrial stromal cells (ESCs). TTR was found to alleviate the level of endometrial fibrosis in a rat model of IUA. A higher number of collagen fibers and greater damage were observed in the endometrial tissue of untreated rats compared to those treated with TTR. The expression of TGF-β1, Smad2, Smad3, and Smad4 was upregulated following IUA, whereas Smad7 expression was downregulated. TTR lowers the expression of TGF-β1, Smad2, Smad3, and Smad4 but increases the expression of Smad7 in vivo, indicating that TTR can modulate the expression of the TGF-β1/Smad pathway to mediate fibrosis. In ESCs, the phosphorylation of Smad2 and Smad3 and upregulation of Smad4 were induced by TGF-β1 whereas the expression of Smad7 was inhibited. Administration of TTR reduces the phosphorylation of Smad2 and Smad3, increases Smad4 expression induced by TGF-β1, and promotes the expression of Smad7. TTR modulates the TGF-β1/Smad pathway to alleviate the generation of fibrotic tissue in response to IUA.

Switching cultivation (mixotrophic-heterotrophic, 12h:12h) of Heynigia riparia SX01 was studied, the maximum biomass concentration of 3.55gL-1 and lipid yield of 1.45gL-1 were achieved after 8days cultivation. The extracellular polymeric substance (EPS) was developed as co-product. Addition of MgSO4 could enhance the production of EPS. The highest amount of 0.60gL-1 EPS was obtained with the addition of 2gL-1 MgSO4, the self-flocculation efficiency was as high as 83% at this condition. The total lipid and lipid fractions did not show differences with extra MgSO4. Based on the above results, a new biodiesel production model was proposed: culturing Heynigia riparia SX01 with extra 2gL-1 MgSO4 by switching cultivation and using self flocculation to collect microalgae for biodiesel production, while EPS was collected as valuable co-products.

Alkaline environments are abundant globally and cause damage to most organisms, while some microalgae can grow well and accumulate lipids under those conditions. Here the mechanisms of alkali resistance and lipid accumulation in the alkaliphilic microalgae Chlorella sp. BLD were explored using physiological-biochemical and transcriptome analysis. When cultivated at alkaline pH, Chlorella sp. BLD exhibited good alkali-resistance ability and increased biomass (0.97 g L−1). The biochemical composition of Chlorella sp. BLD changed significantly (lipid content increased 39% and protein content decreased 19.5%) compared with pH 7.5. Through transcriptome analysis, we found that pathways related to carbon metabolism such as photosynthesis, glycolysis, and the TCA cycle were significantly regulated under alkaline conditions. Genes that encoding the key enzyme in carbon-related metabolism such as Rubisco, AMY, PK, ME, CS, ACAT, KAS, and DGAT were identified. Transcriptional regulation of these genes results in carbon flow switching from starch and protein to cell wall metabolism, organic acid synthetic and lipid accumulation in response to alkaline conditions. These results reveal the alkali resistance mechanism of Chlorella sp. BLD and provide a theoretical basis for microalgae oil production under alkaline conditions.

Glyceroglycolipids, abundant in cyanobacteria&rsquo, s photosynthetic membranes, present bioactivities and pharmacological activities, and can be widely used in the pharmaceutical industry. Environmental factors could alter the contents and compositions of cyanobacteria glyceroglycolipids, but the regulation mechanism remains unclear. Therefore, the glyceroglycolipids contents and the transcriptome in Synechococcus elongatus PCC 7942 were analyzed under phosphate starvation. Under phosphate starvation, the decrease of monogalactosyl diacylglycerol (MGDG) and increases of digalactosyl diacylglycerol (DGDG) and sulfoquinovosyl diacylglycerol (SQDG) led to a decrease in the MGDG/DGDG ratio, from 4:1 to 5:3, after 12 days of cultivation. However, UDP&ndash, sulfoquinovose synthase gene sqdB, and the SQDG synthase gene sqdX, were down-regulated, and the decreased MGDG/DGDG ratio was later increased back to 2:1 after 15 days of cultivation, suggesting the regulation of glyceroglycolipids on day 12 was based on the MGDG/DGDG ratio maintaining glyceroglycolipid homeostasis. There are 12 differentially expressed transcriptional regulators that could be potential candidates related to glyceroglycolipid regulation, according to the transcriptome analysis. The transcriptome analysis also suggested post-transcriptional or post-translational regulations in glyceroglycolipid synthesis. This study provides further insights into glyceroglycolipid metabolism, as well as the scientific basis for glyceroglycolipid synthesis optimization and cyanobacteria glyceroglycolipids utilization via metabolic engineering.

Fatty acid profiles analyzed by gas chromatography (GC). Escherichia coli cells (a) and Saccharomyces cerevisiae cells (b) with Δ9FAD overexpressed. CFAs: cyclopropane fatty acids, including C17 and C19 cyclopropane fatty acid. Bars represent means and standard deviation (n=3).

Growth (a) and fatty acid composition (b) of Auxenochlorella pyrenoidosa under nitrate absence and nitrate replenishment cultures. Initially, Auxenochlorella pyrenoidosa was cultured at 0 g L-1 nitrate concentration. Nitrate replenishment to 1.5 g L-1 was performed after 8 days cultivation under nitrate absence. The arrow indicates when nitrate was replenished. Bars represent means and standard deviation (n=3).

Relative gene expression in response to different nitrate concentrations. The expressions of three genes Δ9FAD (a), Δ12FAD (b) and Δ15FAD (c) in three different growth phases lag (Lag), logarithmic (Log) and stationary phase (S) were detected by real time PCR. Algal cells were collected in lag (day 2), logarithmic (day 7) and stationary (day 12) phases under the nitrate concentrations of 0 and 1.5 g L-1. Bars represent means and standard deviation (n=3).

Alignment of the deduced amino acid sequences of the Δ9, Δ12 and Δ15 fatty acid desaturase enzyme from Auxenochlorella pyrenoidosa (KQ481983.1) and other public database.

Nitrogen consumption and biomass production of Auxenochlorella pyrenoidosa cultivated with initial sodium nitrate concentrations of 1.5 g L-1 (a) and 0 g L-1 (b). The residual nitrogen was detected within the whole cultivation by an automatic chemistry analyzer. Bars represent means and standard deviation (n=3).

Biodiesel quality associated with biochemical components of Chlorella pyrenoidosa and Scenedesmus obliquus under different nitrate levels were investigated. The highest lipid contents of 54.5% for C. pyrenoidosa and 47.7% for S. obliquus were obtained in nitrate absence. Carbohydrate peaked at 0.3gL(-1) with values of 40.7% for C. pyrenoidosa and 42.5% for S. obliquus. Protein content seemed species dependent, which decreased substantially to 11.2% in C. pyrenoidosa and 8.8% in S. obliquus under nitrate absence in present research. Better biodiesel quality (e.g. cetane number58, iodine value69) could be obtained from C. pyrenoidosa in nitrate absence and S. obliquus in 0.3gL(-1), where the highest saturated fatty acids (39.5 for C. pyrenoidosa, 31.2 for S. obliquus) and the lowest unsaturated fatty acids (60.5 for C. pyrenoidosa, 68.8 for S. obliquus) were obtained. These results suggest that microalgae grown in the presence of nitrogen may limit biodiesel quality.

Organisms use various adaptive strategies against phosphate stress, including lipid remodeling. Here, the response of major membrane lipids to phosphate stress was analyzed in Synechococcus sp. PCC 7942. Unlike plants and eukaryotic microalgae, no significant increases in neutral lipids were found, whereas glycolipids content increased to as high as 6.13% (of dry cell weight, DCW) and phospholipids decreased to 0.34% (of DCW) after 16 days of cultivation without phosphate. Glycolipids accumulation were mainly attributed to the significant increase of digalactosyldiacylglycerol (DGDG) by 50% and sulfoquinovosyldiaclglycerol (SQDG) by 90%, both of which acted as complementary lipids for phosphatidylglycerol (PG) in the cyanobacterial membrane. Also, a notable increase in content (by 48%) of C18 fatty acids (especially C18:1) was observed in all glycolipids at the expense of C12 and C14 (72%). These changes may contribute to membrane fluidity and photosynthetic activity for basic cell metabolism and phosphate stress adaptation. Lipidomic analyses showed the reduction of PG 18:1/16: 0 (by 52%) with the increase of DGDG 18:1/16:0 (133%) and SQDG 18:1/16:0 (245%), strongly suggesting a direct conversion of PG to DGDG and SQDG. Moreover, the decreasing amount of monogalactosyldiacylglycerol (MGDG) 16:1/16:0 (22%) was consistent with the increase of free fatty acids (125%) on day 2 of phosphate absence, which suggested that MGDG is more likely to provide a pool of fatty acids for de novo synthesis of glycolipids. This study provides valuable insight into cyanobacteria adaptation strategies to phosphate stress by membrane lipid remodeling and unveils the underlying acyl chain fluxes into glycolipids.

Haematococcus pluvialis is one of the most abundant sources of natural astaxanthin as compared to others microorganism. Therefore, it is important to understand the biorefinery of astaxanthin from H. pluvialis, starting from the cultivation stage to the downstream processing of astaxanthin. The present review begins with an introduction of cellular morphologies and life cycle of H. pluvialis from green vegetative motile stage to red non-motile haematocyst stage. Subsequently, the conventional biorefinery methods (e.g., mechanical disruption, solvent extraction, direct extraction using vegetable oils, and enhanced solvent extraction) and recent advanced biorefinery techniques (e.g., supercritical CO2 extraction, magnetic-assisted extraction, ionic liquids extraction, and supramolecular solvent extraction) were presented and evaluated. Moreover, future prospect and challenges were highlighted to provide a useful guide for future development of biorefinery of astaxanthin from H. pluvialis. The review aims to serve as a present knowledge for researchers dealing with the bioproduction of astaxanthin from H. pluvialis.

The aim of this work was to study salt stress effects on DNA content and oil production processes integrating harvesting, lipid accumulation and oil extraction. Salt-induced enlargement of Parachlorella kessleri cells, with increasing content of DNA and neutral lipid were found. The 34.77% neutral lipid content and biomass concentration of 0.83 g L−1 were obtained after 7 days of salt treatment, compared with that of 13.57% and 0.89 g L−1 cultivated under normal condition. Sedimentation efficiency increased markedly from 15% to 90% due to the cell enlargement. Disruption fraction and the recovery rate of total lipids of wet cells under salt stress were significantly higher than that of normal conditions (100% and 82.4% for salt stress vs.76.8% and 51.1% for normal conditions). This work demonstrated that salt-induced increase in cell size and DNA content was an effective strategy for the enhancement of oil production, microalgae harvesting and oil extraction.

Glycolipids had received increasing attention because of their uses in various industries like cosmetics, pharmaceuticals, food and machinery manufacture. Microalgae were competitive organisms to accumulate metabolic substance. However, using microalgae to produce glycolipid was rare at present. In this study, glycolipid content of Chlorella pyrenoidosa and Synechococcus sp. under different nitrate and hydrophosphate levels were investigated. The highest glycolipid contents of 24.61% for C. pyrenoidosa and 15.37% for Synechococcus sp. were obtained at nitrate absence, which were 17.19% for C. pyrenoidosa and 10.99% for Synechococcus sp. at 0.01 and 0 g L−1 hydrophosphate, respectively. Glycolipid productivities of two microalgae could reach at more than 10.59 mg L−1 d−1. Nitrate absence induced at least 8.5% increase in MGDG, DGDG and SQDG, while hydrophosphate absence resulted in over 21.2% increase in DGDG and over 48.4% increase in SQDG and more than 22.2% decrease in MGDG in two microalgae. Simultaneous nitrate and hydrophosphate limitation could make further improvement of glycolipid accumulation, which was more than 25% for C. pyrenoidosa and 21% for Synechococcus sp. These results suggest that nitrogen and phosphorus limitation or starvation should be an efficient way to improve microalgal glycolipid accumulation.

Rice straw hydrolysate was used as lignocellulose-based carbon source forChlorella pyrenoidosacultivation and the feasibility ofin situbiodiesel production was investigated. 13.7 g/L sugar was obtained by enzymatic hydrolyzation of rice straw.Chlorella pyrenoidosashowed a rapid growth in the rice straw hydrolysate medium, the maximum biomass concentration of 2.83 g/L was obtained in only 48 hours. The lipid content of the cells reached as high as 56.3%.In situtransesterification was performed for biodiesel production. The optimized condition was 1 g algal powder, 6 mLn-hexane, and 4 mL methanol with 0.5 M sulfuric acid at the temperature of 90°C in 2-hour reaction time, under which over 99% methyl ester content and about 95% biodiesel yield were obtained. The results suggested that the method has great potential in the production of biofuels with lignocellulose as an alternative carbon source for microalgae cultivation.

Microalgae are promising feedstocks for biodiesel, where the high proportion of monounsaturated fatty acid such as oleic acid (C18:1) is preferred. To regulate fatty acid desaturation in microalgae, the relationship among nitrate concentration, fatty acid composition and the expression levels of desaturase genes was explored. Dynamic variations of fatty acid profiles suggested nitrate could induce desaturation of C18 fatty acids. The content of C18:1 inAuxenochlorella pyrenoidosawas 30.88% at 0 g l−1nitrate concentration compared with 0.48% at 1.5 g l−1. The expressions of relative delta-9, 12 and 15 fatty acid desaturase genes (Δ9, Δ12 and Δ15FADs) were further investigated. The 330% upregulated expression of Δ9FAD in logarithmic phase at 0 g l−1resulted in C18:1 accumulation. Moreover, nitrate replenishment caused a sharp reduction of C18:1 from 34.79% to 0.22% and downregulation of Δ9FAD expression to 1% of the nitrate absence level, indicating the pivotal role of Δ9FAD in C18:1 accumulation. Finally, overexpression of Δ9FAD inEscherichia coliandSaccharomyces cerevisiaeresulted in an increase of C18:1, confirming its ability of desaturating C18:0. The results could provide a new approach and scientific guidance for the improvement of biodiesel quality and industrialization of high-valued chemicals by means of metabolic engineering.

High effective acidic transesterification catalyzed by trifluoroacetic acid for biodiesel production was studied in the present research. The results showed that the oil could be converted to biodiesel directly by one-step trifluoroacetic acid catalyze process without extreme temperature and pressure conditions. The optimum process combination was 2.0 M catalyst concentration with 20:1 M ratio of methanol to oil at temperature of 120 °C. It reduced product specific gravity from an initial value of 0.965 to a value of 0.878 in about 5 h of reaction time, and the methyl ester content reached as high as 98.4%. The present procedure represents a simple and mild method for biodiesel production in short reaction time and with high conversion rate, which would offer potential for an industrial process.

Aims: To obtain bacteria with PKS (polyketide synthase) genes and antimicrobial activity from sponges. Methods and Results: Eighteen bacteria with KS (ketosynthase) genes were identified by polymerase chain reaction (PCR) screening of 98 isolates from South China Sea sponges, Stelletta tenuis, Halichondria rugosa, Dysidea avara and Craniella australiensis. 16S rRNA gene-based Blast analysis indicated that 15 isolates belonged to the phylum Firmicutes, among which 14 isolates were closely related to genus Bacillus, and 1 to Staphylococcus lentus. Two isolates were identified as actinomycetes, and one as Alcaligenes sp. in the phylum Proteobacteria. The 18 KS domains belong to trans-AT type I PKS and match PKS of marine bacterial symbionts. The 18 bacteria exhibited broad-spectrum antimicrobial activities against fungi, gram-positive and gram-negative bacteria. A2 1AE8-kb PKS gene cluster fragment containing five modules was isolated from the Staphylococcus lentus isolate A75 by screening of a fosmid library. Conclusions: The PKS gene diversity and different antimicrobial spectra indicate the potential of bacteria associated with South China Sea sponges for diverse polyketide production. Significance and Impact of the Study: Combined with bioactivity assay the PKS gene-based approach can be applied to efficient screening of strains of pharmaceutical value and the prediction of related compounds.

In this paper, statistical Plackett–Burman design and Box–Behnken Response Surface Methodology were applied to optimize the medium components to improve the chitinase activity of Streptomyces sp. DA11 associated with South China Sea sponge Craniella australiensis. Firstly, galactose and peptone were found to be the suitable carbon and nitrogen sources for the growth and chitinase activity by single factor Seriatim–Factorial test. Secondly, galactose, colloidal chitin and MgSO4� 7H2O were proved to have remarkable effects on chitinase activity. Finally, an optimal medium was obtained by Box–Behnken

The cultivable bacterial communities associated with four South China Sea sponges-Stelletta tenuis, Halichondria rugosa, Dysidea avara, and Craniella australiensis in mixed cultures-were investigated by microbial community DNA-based DGGE fingerprinting and 16S rDNA phylogenetic analysis. Diverse bacteria such as alpha-, gamma-, delta-Proteobacteria, Bacteroidetes, and Firmicutes were cultured, some of which were previously uncultivable bacteria, potential novel strains with less than 95% similarity to their closest relatives and sponge symbionts growing only in the medium with the addition of sponge extract. According to 16S rDNA BLAST analysis, most of the bacteria were cultured from sponge for the first time, although similar phyla of bacteria have been previously recognized. The selective pressure of sponge extract on the cultured bacterial species was suggested, although the effect of sponge extract on bacterial community in high nutrient medium is not significant. Although alpha- and gamma-Proteobacteria appeared to form the majority of the dominant cultivable bacterial communities of the four sponges, the composition of the cultivable bacterial community in the mixed culture was different, depending on the medium and sponge species. Greater bacterial diversity was observed in media C and CS for Stelletta tenuis, in media F and FS for Halichondria rugosa and Craniella australiensis. S. tenuis was found to have the highest cultivable bacterial diversity including alpha-, gamma-, delta-Proteobacteria, Bacteroidetes, and Firmicutes, followed by sponge Dysidea avara without delta-Proteobacteria, sponge Halichondria rugosa with only alpha-, gamma-Proteobacteria and Bacteroidetes, and sponge C. australiensis with only alpha-, gamma-Proteobacteria and Firmicutes. Based on this study, by the strategy of mixed cultivation integrated with microbial community DNA-based DGGE fingerprinting and phylogenetic analysis, the cultivable bacterial community of sponge could be revealed effectively.

Culture-independent molecular techniques, 16S rDNA clone library alongside RFLP and phylogenetic analysis, were applied to investigate the bacterial diversity associated with three South China Sea sponges, Stelletta tenui, Halichondria rugosa and Dysidea avara. A wide bacterial diversity was detected according to total genomic DNA-based 16S rDNA clone library, abundant clones with low identify with sequences retrieved from database were found as well as uncultured sponge symbionts. The phylogenetic analysis shows that the bacterial community structure of Stelletta tenui is similar to that of Halichondria rugosa comprising gamma-Proteobacteria and Firmicutes. Whereas, alpha-Proteobacteria, gamma-Protebacteria, Bacteroidetes and uncultured sponge symbionts were found in sponge Dysidea avara, suggesting that Dysidea avara has the highest bacteria diversity among these sponges. A specific sponge–microbe association is suggested based on the difference of bacterial diversity among these three sponges from the same geography location and the observed sponge species-specific bacteria.

Effects of white light-emitting diodes (LEDs) with different light intensities at photoperiod of 18:6h on Chlorella pyrenoidosa growth were investigated. The microalgae exhibited the highest growth rate 89.0mgL(-1)d(-1) and growth efficiency 97.8mgL(-1)KWh(-1) at 110 and 90μmolm(-2)s(-1), respectively. Based on the discovery of this asynchronous phenomenon between growth rate and growth efficiency, influences of LEDs (red, blue and white) under capped daily energy consumption (0.80KWh d(-1)) with combinations of electric power (33.3, 44.4 and 66.6w) and photoperiod (24:0, 18:6 and 12:12h) were further investigated. The highest growth efficiency 106.4mgL(-1)KWh(-1) and growth rate 85.1mgL(-1)d(-1) were both obtained under white-33.3w-24h. Growth efficiency and growth rate were simultaneously improved 1.1 times through this method above. The order of growth efficiency under different LEDs were white>blue>red.

The aim of the study was to obtain high quality biodiesel production from a microalga Chlorella protothecoids through the technology of transesterification. The technique of metabolic controlling through heterotrophic growth of C. protothecoides was applied, and the heterotrophic C. protothecoides contained the crude lipid content of 55.2%. To increase the biomass and reduce the cost of alga, corn powder hydrolysate instead of glucose was used as organic carbon source in heterotrophic culture medium in fermenters. The result showed that cell density significantly increased under the heterotrophic condition, and the highest cell concentration reached 15.5 g L(-1). Large amount of microalgal oil was efficiently extracted from the heterotrophic cells by using n-hexane, and then transmuted into biodiesel by acidic transesterification. The biodiesel was characterized by a high heating value of 41 MJ kg(-1), a density of 0.864 kg L(-1), and a viscosity of 5.2 x 10(-4) Pa s (at 40 degrees C). The method has great potential in the industrial production of liquid fuel from microalga.

In the present study, fast pyrolysis tests of microalgae were performed in the fluid bed reactor. The experiments were completed at temperature of 500 degreesC with a heating rate of 600 degreesC s(-1) and a sweep gas (N-2) flow rate of 0.4 m(3) h(-1) and a vapour residence time of 2-3 s. In comparison with the previous studies on slow pyrolysis from microalgae in an autoclave, a greater amount of high quality bio-oil can be directly produced from continuously processing microalgae feeds at a rate of 4 g min(-1) in the present work, which has a potential for commercial application of large-scale production of liquid fuels. The liquid product yields of 18 and 24% from fast pyrolysis of Chllorella protothecoides and Microcystis aeruginosa were obtained. The saturated and polar fractions account for 1.14 and 31.17% of the bio-oils of microalgae on average, which are higher than those of bio-oil from wood. The H/C and O/C molar ratios of microalgae bio-oil are 1.7 and 0.3, respectively. The gas chromatograph analyses showed that the distribution of straight-chain alkanes of the saturated fractions from microalgae bio-oils were similar to diesel fuel. Bio-oil product from fast pyrolysis microalgae is characterized by low oxygen content with a higher heating value of 29 MJ/kg, a density of 1.16 kg l(-1) and a viscosity of 0.10 Pa s. These properties of bio-oil of microalgae make it more suitable for fuel oil use than fast pyrolysis oils from lignocellulosic materials. (C) 2003 Elsevier B.V. All rights reserved.

The use of renewable energy sources is becoming increasingly necessary to mitigate global warming. Recently much research has been focused on identifying suitable biomass species, which can provide high-energy outputs, to replace conventional fossil fuels. This paper reports an approach for increasing the yield of bio-oil production from fast pyrolysis after manipulating the metabolic pathway in microalgae through heterotrophic growth. The yield of bio-oil (57.9%) produced from heterotrophic Chlorella protothecoides cells was 3.4 times higher than from autotrophic cells by fast pyrolysis. The bio-oil was characterized by a much lower oxygen content, with a higher heating value (41 MJ kg(-1)), a lower density (0.92 kg l(-1)), and lower viscosity (0.02 Pas) compared to those of bio-oil from autotrophic cells and wood. These properties are comparable to fossil oil. The research could contribute to the creation of a system to produce energy from microalgae, and also could have great commercial potential for liquid fuel production.

The agp gene encoding ADP-glucose pyrophosphorylase is involved in cyanobacterial glycogen synthesis. By in vitro DNA recombination technology, agp deletion mutant (agp-) of cyanobacterium Synechocystis sp. PCC 6803 was constructed. This mutation led to a complete absence of glycogen biosynthesis. As compared with WT (wild type), a 60% decrease in ratio of the c-phycocyanine/chlorophyll a and no significant change in the carotenoid/chlorophyll a were observed in agp- cells. The agp- mutant had 38% less photosynthetic capacity when grown in light over 600 micromol m(-2) s(-1). Under lower light intensity, the final biomass of the mutant strain was only 1.1 times of that of the WT strain under mixotrophic condition after 6 d culture. Under higher light intensity, however, the final biomass of the WT strain under mixotrophic conditions was 3 times that of the mutant strain after 6 d culture and 1.5 times under photoautotrophic conditions. The results indicate that there is a minimum requirement for glycogen synthesis for normal growth and development in cyanobacteria.

Adaptive laboratory evolution (ALE) is an effective method to improve microalgal strains. The growth phenotypes of three strains (cc4324, cc4326 and cc4334) of green microalgae Chlamydomonas reinhardtii were enhanced by ALE. As a result, endpoint strains exhibited higher growth rates. Upon the utilisation of ALE strategy, the biomass concentrations of the endpoint strains of cc4324, cc4326 and cc4334 became 1.17, 1.33 and 1.48 times of those of the starting strains. The total lipid content of the original strains was increased gradually from 32% to 36.67% in the endpoint strain cc4326 and abruptly increased from 24.27% to 44.67% in the endpoint strain cc4334 by nitrogen starvation. Slight growth impairment was also observed in low-starch mutants exposed to nitrogen starvation stress. However, this impairment was quickly resolved after nitrogen was replenished. These findings demonstrated that the biomass concentration and lipid productivity of low-starch mutants can be enhanced by ALE.

A one-step process was applied to directly converting wet oil-bearing microalgae biomass ofChlorella pyrenoidosacontaining about 90% of water into biodiesel. In order to investigate the effects of water content on biodiesel production, distilled water was added to dried microalgae biomass to form wet biomass used to produce biodiesel. The results showed that at lower temperature of 90°C, water had a negative effect on biodiesel production. The biodiesel yield decreased from 91.4% to 10.3% as water content increased from 0% to 90%. Higher temperature could compensate the negative effect. When temperature reached 150°C, there was no negative effect, and biodiesel yield was over 100%. Based on the above research, wet microalgae biomass was directly applied to biodiesel production, and the optimal conditions were investigated. Under the optimal conditions of 100 mg dry weight equivalent wet microalgae biomass, 4 mL methanol, 8 mL n-hexane, 0.5 M H2SO4, 120°C, and 180 min reaction time, the biodiesel yield reached as high as 92.5% and the FAME content was 93.2%. The results suggested that biodiesel could be effectively produced directly from wet microalgae biomass and this effort may offer the benefits of energy requirements for biodiesel production.

In this study, a novel 96-well microplate swivel system (M96SS) was built for high-throughput screening of microalgal strains for CO2 fixation. Cell growth under different CO2 supply conditions (0.2, 0.4, 0.8, and 1.2 g L−1 d−1), residual nitrate, and pH value of Chlorella sp. SJTU-3, Chlorella pyrenoidosa SJTU-2, and Scenedesmus obliquus SJTU-3 were examined in the M96SS and traditional flask cultures. The dynamic data showed there was a good agreement between the systems. Two critical problems in miniature culture systems (intra-well mixing and evaporation loss) were improved by sealed vertical mixing of the M96SS. A sample screen of six microalgal species (Chlorella sp. SJTU-3, Chlorella pyrenoidosa SJTU-2, Selenastrum capricornutum, Scenedesmus obliquus SJTU-3, Chlamydomonas sajao, Dunaliella primolecta) was carried out in flasks and the M96SS. Chlamydomonas sajao appeared to be a robust performer (highest cell density: 1.437 g L−1) in anaerobic pond water with 0.8, and 1.2 g L−1 d−1 CO2. The reliability and efficiency of the M96SS were verified through a comparison of traditional flask culture, M96SS, Lukavský’s system, and a microplate shaker.

In this study, Scenedesmus obliquus SJTU-3 and Chlorella pyrenoidosa SJTU-2 were cultivated with 0.03%, 5%, 10%, 20%, 30%, 50% CO(2). The two microalgae could grow at 50% CO(2) (>0.69 g L(-1)) and grew well (>1.22 g L(-1)) under CO(2) concentrations ranging from 5% to 20%. Both of the two examined microalgae showed best growth potential at 10% CO(2). The maximum biomass concentration and CO(2) biofixation rate were 1.84 g L(-1) and 0.288 g L(-1) d(-1) for S. obliquus SJTU-3 and 1.55 g L(-1) and 0.260 g L(-1) d(-1) for C. pyrenoidosa SJTU-2, respectively. The main fatty acid compositions of the two examined microalgae were fatty acids with C(16)-C(18) (>94%) under different CO(2) levels. High CO(2) levels (30-50%) were favorable for the accumulation of total lipids and polyunsaturated fatty acids. The present results suggested that the two microalgae be appropriate for mitigating CO(2) in the flue gases and biodiesel production.

Static liquid culture of Ganoderma lucidum, a traditional Chinese medicinal mushroom, is a proven technology for producing ganoderic acids, which are secondary metabolites that possess antitumor properties. In this work, the addition of phenobarbital, a P450 inducer, was used to enhance the production of total and individual ganoderic acids in a two-stage cultivation involving a period of initial shake flask culture followed by static liquid culture of G. lucidum. The dosage and time of phenobarbital induction were critical for the enhanced production of ganoderic acids. The addition of 100 muM (final concentration) phenobarbital on day 5 after the shake flask culture was converted to the static liquid culture was found to be optimal, resulting in a maximal amount of total ganoderic acids of 41.4 +/- 0.6 mg/g cell dry weight and increases in the levels of ganoderic acid-Mk, -T, -S, and -Me in the treated cells by 47%, 28%, 36%, and 64%, respectively. Meanwhile, the accumulation of lanosterol, a key intermediate, was found to decrease and transcriptions of three key genes encoding 3-hydroxy-3-methylglutaryl coenzyme A reductase, squalene synthase, and lanosterol synthase in the triterpene biosynthetic pathway were up-regulated under phenobarbital induction. This work demonstrated a useful strategy for the enhanced production of ganoderic acids by G. lucidum.

Xylitol production from xylose by a self-isolated furfural and 5-hydroxymethyl furfural assimilating Pichia guilliermondii was studied under oxygen limitation. An extremely low initial volumetric oxygen transfer coefficient (0.075 h(-1)) was found most favorable to the xylitol production with yield of 0.61 g g(-1). Related enzymes activities were also investigated and discussed.

The gene cloning, purification, properties, kinetics, and antifungal activity of chitinase from marine Streptomyces sp. DA11 associated with South China sponge Craniella australiensis were investigated. Alignment analysis of the amino acid sequence deduced from the cloned conserved 451 bp DNA sequence shows the chitinase belongs to ChiC type with 80% similarity to chitinase C precursor from Streptomyces peucetius. Through purification by 80% ammonium sulfate, affinity binding to chitin and diethylaminoethyl-cellulose anion-exchange chromatography, 6.15-fold total purification with a specific activity of 2.95 Umg(-1) was achieved. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) showed a molecular weight of approximately 34 kDa and antifungal activities were observed against Aspergillus niger and Candida albicans. The optimal pH, temperature, and salinity for chitinase activity were 8.0, 50 degrees C, and 45 g per thousand psu, respectively, which may contribute to special application of this marine microbe-derived chitinase compared with terrestrial chitinases. The chitinase activity was increased by Mn(2+), Cu(2+), and Mg(2+), while strongly inhibited by Fe(2+) and Ba(2+). Meanwhile, SDS, ethyleneglycoltetraacetic acid, urea, and ethylenediaminetetraacetic acid were found to have significantly inhibitory effect on chitinase activity. With colloidal chitin as substrates instead of powder chitin, higher V (max) (0.82 mg product/min.mg protein) and lower K (m) (0.019 mg/ml) values were achieved. The sponge's microbial symbiont with chitinase activity may contribute to chitin degradation and antifungal defense. To our knowledge, it was the first time to study sponge-associated microbial chitinase.

Biomass, nutrient removal capacity, lipid productivity and morphological changes of Chlorella sorokiniana and Desmodesmus communis were investigated in mixed wastewaters with different CO2 concentrations. Under optimal condition, which was 1:3 ratio of swine wastewater to second treated municipal wastewater with 5% CO2, the maximum biomass concentrations were 1.22 g L-1 and 0.84 g L-1 for C. sorokiniana and D. communis, respectively. Almost all of the ammonia and phosphorus were removed, the removal rates of total nitrogen were 88.05% for C. sorokiniana and 83.18% for D. communis. Lipid content reached 17.04% for C. sorokiniana and 20.37% for D. communis after 10 days culture. CO2 aeration increased intracellular particle numbers of both microalgae and made D. communis tend to be solitary. The research suggested the aeration of CO2 improve the tolerance of microalgae to high concentration of NH4-N, and nutrient excess stress could induce lipid accumulation of microalgae.

The present study introduced an integrated method for the production of biodiesel from microalgal oil. Heterotrophic growth of Chlorella protothecoides resulted in the accumulation of high lipid content (55%) in cells. Large amount of microalgal oil was efficiently extracted from these heterotrophic cells by using n-hexane. Biodiesel comparable to conventional diesel was obtained from heterotrophic microalgal oil by acidic transesterification. The best process combination was 100% catalyst quantity (based on oil weight) with 56:1 molar ratio of methanol to oil at temperature of 30 degrees C, which reduced product specific gravity from an initial value of 0.912 to a final value of 0.8637 in about 4h of reaction time. The results suggested that the new process, which combined bioengineering and transesterification, was a feasible and effective method for the production of high quality biodiesel from microalgal oil.

The agp gene encoding ADP-glucose pyrophosphorylase is involved in cyanobacterial glycogen synthesis. By in vitro DNA recombination technology, agp deletion mutant (agp-) of cyanobacterium Synechocystis sp. PCC 6803 was constructed. This mutation led to a complete absence of glycogen biosynthesis. As compared with WT (wild type), a 60% decrease in ratio of the c-phycocyanine/chlorophyll a and no significant change in the carotenoid/chlorophyll a were observed in agp- cells. The agp- mutant had 38% less photosynthetic capacity when grown in light over 600 micromol m(-2) s(-1). Under lower light intensity, the final biomass of the mutant strain was only 1.1 times of that of the WT strain under mixotrophic condition after 6 d culture. Under higher light intensity, however, the final biomass of the WT strain under mixotrophic conditions was 3 times that of the mutant strain after 6 d culture and 1.5 times under photoautotrophic conditions. The results indicate that there is a minimum requirement for glycogen synthesis for normal growth and development in cyanobacteria.

The agp gene encoding the ADP-glucose pyrophosphorylase is involved in cyanobacterial glycogen synthesis and glucosylglycerol formation. By in vitro DNA recombination technology, a mutant with partial deletion of agp gene in the cyanobacterium Synechocystis sp. PCC 6803 was constructed. This mutant could not synthesize glycogen or the osmoprotective substance glucosylglycerol. In the mutant cells grown in the medium containing 0.9 M NaCl for 96 h, no glucosylglycerol was detected and the total amount of sucrose was 29 times of that of in wild-type cells. Furthermore, the agp deletion mutant could tolerate up to 0.9 M salt concentration. Our results suggest that sucrose might act as a similar potent osmoprotectant as glucosylglycerol in cyanobacterium Synechocystis sp. PCC 6803.