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PubMed2006

High quality biodiesel production from a microalga Chlorella protothecoides by heterotrophic growth in fermenters.

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.

Biomass↗
PubMed2004

Recovery in aqueous two-phase systems of lutein produced by the green microalga Chlorella protothecoides.

In this study the potential use of aqueous two-phase system (ATPS) to establish a viable process for the recovery of lutein from the green microalga Chlorella protothecoides is evaluated. The partitioning behaviour of lutein, a representative model of natural compounds of commercial interest, was investigated in a polyethylene glycol (PEG)-phosphate system. An evaluation of system parameters including PEG molecular mass, the concentrations of PEG, phosphate and product concentration was conducted, to estimate conditions under which lutein partitions preferentially to the top phase whilst cell debris partition to the opposite phase. The necessary addition of ethanol to the ATPS for the dissolution of lutein affected the phase formation and such effect was evaluated using the change in the volume ratio produced. ATPS extraction comprising Vr = 1.0, PEG 8000 22.9% (w/w) and phosphate 10.3% (w/w), pH 7.0 provided the conditions for the concentration of lutein into the upper phase and the cell debris preferentially to the bottom phase. The use of ATPS resulted in a primary recovery process to obtain lutein with an overall product yield of 81.0 +/- 2.8%. The findings reported here demonstrate the potential of ATPS for the further development of a prototype process to recover lutein from C. protothecoides as a first step for the generic application of this technique.

Chlorella↗
PubMed2004

Removal and recovery of nickel(II) from aqueous solution by loofa sponge-immobilized biomass of Chlorella sorokiniana: characterization studies.

The biosorption process for the removal of nickel(II) by loofa sponge-immobilized biomass of Chlorella sorokiniana (LIBCS), a newly developed immobilized biosorbent, was characterized. Effects of environmental factors on metal uptake capacity of LIBCS were studied and compared with free biomass of C. sorokiniana (FBCS). Nickel(II) removal by LIBCS was found to be influenced by pH of the solution, initial metal concentration, and biomass concentration. The biosorption of nickel(II) ions by both LIBCS and FBCS increased as the initial concentration of nickel(II) ions increased in the medium. No loss to biosorption capacity of LIBCS for nickel(II) was found due to the presence of loofa sponge, indeed as compared to FBCS an increase of 25.3% was noted in the biosorption capacity of LIBCS. Maximum biosorption capacities for FBCS and LIBCS were found as 48.08 and 60.38 mg nickel(II)/g, respectively, whereas the amount of nickel(II) ions adsorbed on the plain loofa sponge was 6.1mg/g. During these biosorption studies, LIBCS exhibited excellent physical and chemical stability without any significant release/loss of microalgal biomass from loofa sponge matrix. The kinetics of nickel(II) removal was extremely fast reaching at equilibrium in about 15 min for LIBCS and 20 min for FBCS. The biosorption equilibrium was well described by the Langmuir and Freundlich adsorption isotherms. The biosorption capacities were found to be solution pH dependent and the maximum adsorption was found at a solution pH 4-5. The LIBCS could be regenerated using 75 mM HCl, with up to 98% recovery. The LIBCS were shown to be robust and stable with little decrease in the nickel(II) uptake capacity when used in consecutive seven biosorption-desorption cycles. Continuous removal of nickel(II) from electroplating effluent by LIBCS packed in fixed bed column bioreactor confirm the possibility of developing a biological treatment process for the removal of toxic metals from authentic wastewater.

Adsorption↗
PubMed2006

Photoreduction of chromium(VI) in the presence of algae, Chlorella vulgaris.

In this thesis, the photochemical reduction of hexavalent chromium Cr(VI) in the presence of algae, Chlorella vulgaris, was investigated under the irradiation of metal halide lamps (lambda = 365 nm, 250 W). The affecting factors of photochemical reduction were studied in detail, such as exposure time, initial Cr(VI) concentration, initial algae concentration and pH. The rate of Cr(VI) photochemical reduction increased with algae concentration increasing, exposure time increasing, initial Cr(VI) concentration decreasing and the decrease of pH. When pH increased to 6, the rate of Cr(VI) photochemical reduction nearly vanished. When initial Cr(VI) concentration ranged from 0.4 to 1.0 mg L(-1) and initial algae concentration ranged from ABS(algae) (the absorbency of algae) = 0.025 to ABS(algae) = 0.180, According to the results of kinetic analyses, the kinetic equation of Cr(VI) photochemical reduction in aqueous solution with algae under 250 W metal halide lamps was V0 = kC(0)(0.1718)A(algae)(0.5235) (C0 was initial concentration of Cr(VI); A(algae) was initial concentration of algae) under the condition of pH 4.

Biodegradation, Environmental↗
PubMed2006

Metal toxicity to Chlorella pyrenoidosa assessed by a short-term continuous test.

This study presents the metal toxicity as revealed by Chlorella pyrenoidosa using a short-term continuous test and the conventional batch test. Toxicity tests were conducted in a chemostat with a test duration of 1 day. Toxicity data of five different metals (Cd, Pb, Co, Ni, and Zn) derived from the continuous test were compared against results from the conventional batch tests. The batch test is characterized by its saturated nutrient status for algal growth and, on the other hand, the continuous test is conducted at a nutrient-limiting condition simulating more closely to the aquatic field environment. The continuous test consistently yielded smaller EC(50) values, which indicated that alga in the chemostat is approximately four to six times more sensitive than those from the batch culture. Similar phenomena between the two types of tests were observed based on the NOEC values. Compared to literature data (US EPA ECOTOX Database) from the same species of alga, differences in test sensitivity between the batch tests and chemostat technique varied from 9.25 to 160 times. Such distinct differences indicate that current toxicity database may not adequately reflect the impact of various metal toxicants on C. pyrenoidosa. The continuous test method presented herein is a sensitive and rapid assessment technique and, therefore, has a potential for more general applications such as routine checkup for regulatory purposes.

Chlorella↗
PubMed2006

Biosorption and bioreduction of Cr(VI) by a microalgal isolate, Chlorella miniata.

The ability and mechanism of a microalgal isolate, Chlorella miniata to remove Cr(VI) were investigated. Kinetic studies indicated that both biosorption and bioreduction were involved in the Cr(VI) removal. The adsorbed Cr(VI) was reduced to Cr(III), and desorption studies indicated that Cr(III) occupied most of the adsorption sites on the biomass. The equilibrium time for Cr(VI) removal was dependent on various factors including initial pH, biomass and Cr(VI) concentrations. Equilibrium study showed that the Cr(VI) removal capacity was negatively related to the initial pH, and the biosorption capacity of total Cr [Cr(III) and Cr(VI)] reached the maximum at initial pH of 3.0. The spectrum of Fourier Transform Infrared Spectrometer analysis (FTIR) further confirmed that amino group on the algal biomass was the main adsorption site for Cr(VI) biosorption in acidic pH while the reduced Cr(III) was mainly sequestered by carboxylate group. The comparison between biosorption-bioreduction and direct bioreduction kinetic models proved that biosorption of Cr(VI) was the first step, followed by Cr(VI) bioreduction and Cr(III) biosorption on the algal biomass.

Adsorption↗
PubMed2005

Immunostimulatory principles from Chlorella pyrenoidosa--part 1: isolation and biological assessment in vitro.

Our proprietary preparation obtained by extraction of Chlorella pyrenoidosa cells, ONC-107 (Respondin), was recently found to selectively boost antibody response to the influenza vaccine in a human clinical trial. Respondin is a potent stimulator of mouse B cell proliferation and an activator of macrophages. Bioactivity-guided resolution concluded that Respondin is composed of a mixture of immunostimulatory principles of different chemical nature. A combination of size exclusion, anion exchange and hydrophobic interaction chromatography revealed that the bulk of the immunostimulatory activity resides in polysaccharide/protein complexes with molecular masses larger than 100 kDa that are composed primarily of galactose, rhamnose and arabinose.

Animals↗
PubMed2004

Protective role of 20-hydroxyecdysone against lead stress in Chlorella vulgaris cultures.

Treatment of cultured C. vulgaris cells with 10(-6)-10(-4) M lead decreased their growth and chemical composition during the first 48 h of cultivation. However, at concentrations above 10(-4) M, lead is cytotoxic to Chlorella vulgaris cells, resulting in cellular fragmentation and lysis. In contrast, at concentrations below 10(-6) M lead had no influence on the growth and metabolism of C. vulgaris cells. 20-Hydroxyecdysone (20E) (10(-10)-10(-8) M) increased growth and chemical composition of C. vulgaris cells over a concentration range. Levels per cell of chlorophylls, protein, sugars are all increased by 20E treatment, when compared to non-treated control cells. However, the cultures treated with 20E and lead show a lower stimulation than the cultures treated with 20E alone. The effects of 20E mixed with lead on the growth and the level of cellular lead, chlorophyll, sugar and protein in C. vulgaris are also reported. The decreased growth and composition of C. vulgaris cells treated with lead was restored by the 20E. Application of 20E to C. vulgaris cultures reduced the impact of lead stress on growth, prevented chlorophyll, sugar and protein loss and increased phytochelatins synthesis. Furthermore, 20E did not restore toxic effect of lead on C. vulgaris cells. The combined treatment with lead and 20E appeared to have a stimulatory effect on the above parameters during the 48 h of cultivation, as compared to the control. 20E reduced the toxicity of lead and the growth recovered to the level of cells treated with 20E alone. Concentration-dependent stimulation was observed with increasing concentration of 20E and decreasing concentration of lead.

Algal Proteins↗
PubMed2004

Enhancement of photosynthetic O2 evolution in Chlorella vulgaris under high light and increased CO2 concentration as a sign of acclimation to phosphate deficiency.

The photosynthetic oxygen evolution of Chlorella vulgaris (Beijer.) cells taken from phosphate-deficient (-P) and control cultures was measured during 8 days of culture growth. Under inorganic carbon concentration (50 microM) in the measuring cell suspension and irradiance (150 micromol m(-2) s(-1)), the same as during culture growth, there were no marked differences in the photosynthetic O2 evolution rate between the -P cells and the controls. The much slower growth of -P cultures indicated that the utilization of absorbed photosynthetically active radiation (PAR) in the CO2 assimilation and biomass production were in -P cells less efficient than in the controls. Alga cells under the phosphorus stress utilized more of the absorbed PAR in the nitrate reduction than the control cells. However, under conditions of more efficient CO2 supply (inorganic carbon concentration 150 microM, introducing of exogenous carbonic anhydrase to the measuring cell suspension) and under increased irradiance (500 micromol m(-2) s(-1)), the photosynthetic O2 evolution in -P cells reached a higher rate than in the controls. The results suggest that in -P cells the restricted CO2 availability limits the total photosynthetic process. But under conditions more favorable for the CO2 uptake and under high irradiance, the -P cells may reveal a higher photosynthetic oxygen evolution rate than the controls. It is concluded that an increased potential activity of the photosynthetic light energy absorption and conversion in the C. vulgaris cells from -P cultures is a sign of acclimation to phosphorus stress by a sun-type like adaptation response of the photosynthetic apparatus.

Carbonic Anhydrases↗
PubMed2006

Toxicity of hexachlorobenzene and its transference from microalgae (Chlorella kessleri) to crabs (Chasmagnathus granulatus).

The aim of this work was to study the transference of hexachlorobenzene from a green alga (Chlorella kessleri) to an estuary crab (Chasmagnathus granulatus), and to analyze the toxic effects that the xenobiotic has on the latter. The effect of hexachlorobenzene uptake was evaluated measuring oxidative stress, Uroporphyrinogen decarboxylase activity and morphometric parameter alteration, and also performing a histological analysis of crab hepatopancreas. Results demonstrated that hexachlorobenzene enters the alga, is accumulated in it, and then transferred into the crab, causing a decrease in Uroporphyrinogen decarboxylase activity in both organisms. The high malondialdehyde levels detected in crab hepatopancreas after the toxic treatment suggested the existence of hexachlorobenzene-induced lipid peroxidation. Antioxidant defenses such as superoxide dismutase activity and reduced glutathione content fell below normal values on the fourth week of treatment. At the same time, the hepatosomatic index, used as a morphometric parameter, reduced 20% with respect to the control. The histological analysis revealed epithelium disorganization in hepatopancreas tubules, confirming the existence of structural damage caused by hexachlorobenzene.

Animals↗
PubMed2006

C-terminal repetitive motifs in Vp130 present at the unique vertex of the Chlorovirus capsid are essential for binding to the host Chlorella cell wall.

Previously, Vp130, a chloroviral structural protein, was found to have host-cell-wall-binding activity for NC64A-viruses (PBCV-1 and CVK2). In this study, we have isolated and characterized the corresponding protein from chlorovirus CVGW1, one of Pbi-viruses that have a different host range. In NC64A-viruses, Vp130 consists of a highly conserved N-terminal domain, internal repeats of 70-73 aa motifs and a C-terminal domain occupied by 23-26 tandem repeats of a PAPK motif. However, CVGW1 was found to have a slightly different Vp130 construction where the PAPK repeats were not in the C-terminus but internal. Immunofluorescence microscopy with a specific antibody revealed that the C-terminal region containing the Vp130 repetitive motifs from PBCV-1 and CVK2 was responsible for binding to Chlorella cell walls. Furthermore, by immunoelectron microscopy and immunofluorescence microscopy, Vp130 was localized at a unique vertex of the chlorovirus particle and was found to be masked through binding to the host cells. These results suggested that Vp130 is localized at a unique vertex on the virion, with the C-terminal repetitive units outside for cell wall binding.

Amino Acid Sequence↗
PubMed2006

Chlorella viruses contain genes encoding a complete polyamine biosynthetic pathway.

Two genes encoding the putative polyamine biosynthetic enzymes agmatine iminohydrolase (AIH) and N-carbamoylputrescine amidohydrolase (CPA) were cloned from the chloroviruses PBCV-1, NY-2A and MT325. They were expressed in Escherichia coli to form C-terminal (His)6-tagged proteins and the recombinant proteins were purified by Ni2+-binding affinity chromatography. The biochemical properties of the two enzymes are similar to AIH and CPA enzymes from Arabidopsis thaliana and Pseudomonas aeruginosa. Together with the previously known virus genes encoding ornithine/arginine decarboxlyase (ODC/ADC) and homospermidine synthase, the chloroviruses have genes that encode a complete set of functional enzymes that synthesize the rare polyamine homospermidine from arginine via agmatine, N-carbamoylputrescine and putrescine. The PBCV-1 aih and cpa genes are expressed early during virus infection together with the odc/adc gene, suggesting that biosynthesis of putrescine is important in early stages of viral replication. The aih and cpa genes are widespread in the chlorella viruses.

Amidohydrolases↗
PubMed2000

The C-terminal tetrapeptide HWFW of the Chlorella HUP1 hexose/H(+)-symporter is essential for full activity and an alpha-helical structure of the C-terminus.

C-terminal tails of plant hexose/H(+)-symporters of the major facilitator superfamily contain a highly conserved motif of four amino acids: HWFW. A deletion of these four amino acids in the Chlorella HUP1 protein leads to a decrease in transport activity by a factor of 3-4. The mutated tail is highly sensitive to trypsin; it does not show alpha-helical conformation in contrast to the wild type C-terminal peptide with an alpha-helical content of at least 15%. The production of monoclonal antibody 416B8 recognizing an epitope within the central loop of HUP1 protein has been a prerequisite for the experiments described.

Amino Acid Sequence↗
PubMed2003

Are chlorella viruses a rich source of ion channel genes?

Plaque-forming dsDNA (>330 kb) viruses that infect certain unicellular, eukaryotic chlorella-like green algae contain approximately 375 protein-encoding genes. These proteins include a 94 amino acid K+ channel protein, called Kcv, as well as two putative ligand-gated ion channels. The viruses also encode other proteins that could be involved in the assembly and/or function of ion channels, including protein kinases and a phosphatase, polyamine biosynthetic enzymes and histamine decarboxylase.

Amino Acid Sequence↗
PubMed2001

High molecular weight lipids from the trilaminar outer wall (TLS)-containing microalgae Chlorella emersonii, Scenedesmus conmmunis and Tetraedron minimum.

High molecular weight lipids were isolated from Chlorella emersonii, Scenedesmus communis and Tetraedron minimum, thin trilaminar outer wall (TLS)-containing freshwater microalgae producing an insoluble non-hydrolysable biopolymer (i.e. algaenan). Molecular weight determination by gel permeation chromatography indicated that their molecular weights range from ca. 400 to 2000 Da. Flash pyrolysis with in situ methylation using tetramethylammonium hydroxide (TMAH) and alkaline hydrolysis showed that the high molecular weight lipids isolated from C. emersonii and S. communis are mainly composed of saturated n-C26 and n-C28 fatty acids and alcohols and of saturated n-C30 and n-C32 alpha,omega-diols and omega-hydroxy acids. In contrast the high molecular weight lipids isolated from T. minimum are predominantly composed of long-chain fatty acids and omega-hydroxy acids. Aromatic moieties were also identified in small amounts in the thermochemolysate and in the hydrolysate. Chemical structural models containing long-chain mono- and polyesters were proposed for the high molecular weight lipids isolated from the three microalgae in agreement with analytical and spectroscopic data. Structural similarity between the outer cell wall of these microalgae and the cuticular membrane of higher plants is suggested.

Cell Wall↗
PubMed1995

Analysis of 45 kb of DNA located at the left end of the chlorella virus PBCV-1 genome.

Forty-five kilobases of DNA, including the previously sequenced 2.2-kb inverted repeat region, located at the left termini of the 330-kb Chlorella virus PBCV-1 genome were sequenced and analyzed. Eighty-five complete open reading frames (ORFs) larger than 195 nucleotides were identified. Thirty-seven of the 85 ORFs, which are densely packed on both strands of the DNA, were considered major ORFs. Fifteen of the major ORFs have similarity to genes in the databases, including bacterial glycerophosphoryl diester phosphodiesterase, bacteriophage T4 endonuclease V, D-isomer specific 2-hydroxyacid dehydrogenases, and beta-alanine synthetase and bacterial nitrilases. Two major ORFs resemble the virus major capsid protein. Three major ORFs contain three or more ankyrin-like repeat elements and four ORFs encode proline-rich proteins.

Amino Acid Sequence↗
PubMed2002

Removal of ammonium and phosphorus ions from synthetic wastewater by the microalgae Chlorella vulgaris coimmobilized in alginate beads with the microalgae growth-promoting bacterium Azospirillum brasilense.

Coimmobilization of the freshwater microalga Chlorella vulgaris in alginate beads with the microalgae growth-promoting bacterium Azospirillum brasilense under semi-continuous synthetic wastewater culture conditions significantly increased the removal of ammonium and soluble phosphorus ions compared to immobilization of the microalgae alone. In continuous or batch cultures removal of these ions followed a similar trend but was less efficient than in semi-continuous culture. It is proposed that coimmobilization of a microalgae with microalgae growth-promoting bacteria can serve as a tool in devising novel wastewater treatments.

Alginates↗