Effect of Light Intensity and pH on Cell Density Assessed by Spectrophotometry for the Unicellular Algae Chlorella vulgaris

In this study, an effective environment for Chlorella vulgaris growth is sought after. As a substitute source of food and feed, increasing the cell density of Chlorella culture is one of the keys to ensuring sustainability. It can be showed from different studies that optimum light intensity and pH could increase cell density. In this study, the effects of light and pH on the growth rate of C. vulgaris were observed in photobioreactor. A specific wavelength (682 nm) was determined by UV-Vis Spectrophotometry to carry out the further analysis. The light intensities were set at 7409, 9261 and 11,113 lux;pH values were set at 7, 8 and 9 respectively. The experimental results depicted the light intensity of 9261 lux as the best due to the higher number of cells (48.56 × 106 cells/mL) obtained using this intensity. In terms of pH, without pH control, cell numbers were found to be highest under the light intensity of 9261 lux. When pH was controlled, it was found that under the optimum light intensity, pH control between 7.0 and 7.5 was the optimum range for the growth of C. vulgaris. Moreover, this method of study may possibly be a promising source of low cost culture for Chlorella vulgaris.

Vitreoscilla hemoglobin gene(vgb)improves lutein production in Chlorella vulgaris

Vitreoscilla hemoglobin is an oxygen-binding protein that promotes oxygen delivery and reduces oxygen consumption under low oxygen conditions to increase the effi ciency of cell respiration and metabolism.In this study,we introduced a Vitreoscilla hemoglobin gene(vgb)into Chlorella vulgaris by Agrobacterium tumefaciens-mediated transformation(ATMT).PCR analysis confi rmed that the vgb gene was successfully integrated into the Chlorella vulgaris genome.Analysis of biomass obtained in shake fl asks revealed transformant biomass concentrations as high as 3.28 g/L,which was 38.81% higher than that of the wild-type strain.Lutein content of transformants also increased slightly.Further experiments recovered a maximum lutein yield of 2.91 mg/L from the transformants,which was 36.77% higher than that of the wild-type strain.The above results suggest that integrated expression of the vgb gene may improve cell growth and lutein yield in Chlorella vulgaris,with applications to lutein production from Chlorella during fermentation.

Effect of Cu and Ni on Growth, Mineral Uptake,Photosynthesis and Enzyme Activities of Chlorella vulgaris at Different pH Values

A pH dependent reduction in growth, Pigment, ATP content, O,' evolution, carbonfixation, Photosynthetic electron transport system, nutrient uptake (NO,' and NH4+),nitrate reductase, and ATPase activities and increase in K+ emux of Chlorella vulgaris wasnoticed following supplementation of Cu and Ni to the culture medium. PS II was foundto be more sensitive to both pH and metals than PS I. Though, nitrate reductase (NR) wasmore sensitive to both PH and metals, the ATPase was however, more sensitive t0 metalsbut less sensitive to acidic pH. Acid PH was found to dst the nutrient (NO,' and NH4+)uptake and nitrate reductase in a non-competitive manner. The inhibition Pnduced by thetest metals alone was of noncompetitive type for NO3' uptake, nitrate reductase andATPase and competitive for NH'+ uptake. Acidity not only inhibited the metabolicvariables directly but also through facilitated uptake of metals and increased membranepermeability. A very low sensitivity of ATPase to acidic pH seems to be resgnsible forthe survival of algae in acid environment.

Photodegradation of 17α-Ethynylestradiol in Aqueous Solution with Nitzschiahantzschiana or Chlorella vulgaris and Fe^(3+)

The photodegradation of 17α-ethynylestradiol (EE2) induced by highpressure mercury lamp (λ≥313 nm, 250 W) in aqueous solution with algae (e. g.Nitzschia hantzschiana andChlorella vulgaris) and Fe3+ was ineestigated initially. The affecting factors on the photodegradation were studied and described in details, such as algae concentration, Fe3+, exposure time, and so on. The concentration of EE2 in distilled water was determined using fluorescence spectrophotometer. The photodegradation of EE2 in aqueous solution exposed to 250 W high-pressure mercury lamp was evident in the presence of algae and Fe3+. With the algae concentration increasing, photodegradation rate increased. Fe3+ could accelerate the photocatalytic degradation of EE2 in aqueous solution with algae. The colloidal ferric hydroxide particles that might have absorbed on the algae cells could enhance the photocatalytic degradation of EE2 by algae. The catalysis in photocatalytic degradation reaction mainly resulted from the active oxygen (H2O2,1O2 and ·OH) that was caused by algae and Fe3+ under 250 W HPML. In this paper, the mechanism of photocatalytic degradation of EE2 by algae and Fe3+ is discussed theoretically in details. Key words 17α-ethynylestradiol - photodegradation - high-pressure mercury lamp - Nitzschia hantzschiana - Chlorella vulgaris - Fe3+ CLC number X 131. 2 Foundation item: Supported by the Scientific Research Foundation of Wuhan Environmental Protection Bureau and the National Natural Science Foundation of China (20177017)Biography: Liu Xian-li (1965-), male, Ph. D candidate, Associate professor, research direction: environmental chemistry

Interaction between Chlorella vulgaris and bacteria:interference and resource competition

Research of interaction mechanism between Chlorella vulgaris and two bacterial strains（Z-QD08 and Z-QS01） were conducted under laboratory conditions.Growth rates of bacteria and C.vulgaris were tested under co-culture conditions to evaluate the effects of concentrations of C.vulgaris and bacteria on their interactions.To test whether the availability of inorganic nutrients,vitamins and trace metals affects the interactions between C.vulgaris and bacteria,experiments were performed with or without the culture medium filtrate of C.vulgaris or bacteria.The results showed that the growth of C.vulgaris was promoted at low concentrations of bacteria（5×106cells/ml）,and expressed a positive correlation with the bacteria density,whereas opposite trend was observed for treatments with high bacteria density（10×106cells/ml and 20×106cells/ml）.The growth rate of bacteria decreased with the increasing concentrations of C.vulgaris.The growth of bacteria Z-QD08 was inhibited by C.vulgaris through interference competition,while the mechanism for interaction between bacteria Z-QS01 and C.vulgaris was resource competition.The influence of cell density on the interaction between microalgae and bacteria was also discussed.These experiments confirm some elements of published theory on interactions between heterotrophic bacteria and microalgae and suggest that heterotrophic bacteria play an important role in the development of blooms in natural waters.

Genetic diversity analysis with ISSR PCR on green algae Chlorella vulgaris and Chlorella pyrenoidosa

In the present study, genetic polymorphism and diversity in unicellular clones of Chlorella vulgaris Beijerinck and Chlorella pyrenoidosa Chick were studied with Inter Simple Sequence Repeats PCR （ISSR PCR）. Samples including four clones of C. vulgaris and three clones of C. pyrenoidosa were purified by single-clone-choice method. For four C. vulgaris unicellular clones, the total number of the bands scored for 18 primers was 298; and the number of the polymorphic bands was 118, of which 39.6% were polymorphic. The size of PCR products ranged from 200 to 2 500 bp. The total number of bands scored for 18 primers, the number of polymorphic bands and the percentage of three C. pyrenoidosa unicellular clones was 194.83 and 30.8%, respectively. POPGENE analysis show that the average Nei genetic diversity （h^＊） and Shannon index of diversity （I^＊） in the four C. vulgaris unicellular clones was 0.2181 and 0.3208, respectively, which is slightly higher than those of the three C. pyrenoidosa unicellular clones （0.190 3 and 0.274 8）, which agreed with the percentage of polymorphic bands in the mixed samples of the two species. The results suggest that ISSR is a useful method to Chlorella for intra-species genetic analysis.

Feasible design for electricity generation from Chlorella vulgaris using convenient photosynthetic conditions

Many recent studies are concerned with low cost,easy to handle and alternative renewable energy as a feasible solution for the upcoming crisis of energy shortage.Microalgae are unicellular entities the can only depend on CO_(2),water and solar power to cover their nutritional needs.The current study is concerned with using algal cells in a polymeric hydrogel,as a cheap source of energy for electricity generation.Chlorella vulgaris has been proved to be a promising algal species for electricity generation,as compared with Micractinium reisseri.PVA hydrogel has been used for the immobilization of both algal species in order to protect them from the adverse surrounding conditions in addition to its ability to slowly release the required water molecules according to needs.Under these conditions,C.vulgaris showed the ability to generate 60 mV compared with 15 mV generated by M.reisseri.Scanning electron micrographs showed nano-threads that bind the C.vulgaris cells to each other,indicating the ability of algae to create nanowires that facilitate the electron transfer among algal cells and from cells to the nearest electrode.However,we would expect an increase in the produced potential with simultaneous amendment of environmentally polluted water,such as sewage or waste water.Both of FTIR and raman spectroscopy proved the presence of the characteristic groups of PVA hydrogel and proved the proper integration of the algal cells inside the hydrogel cavities.

Metal Induced Inhibition of Photosynthesis,Photosynthetic Electron Transport Chain and ATP Content of Anabaena doliolum and Chlorella vulgaris:Interaction with Exogenous ATP

This study demonstrates a concentration dependent inhibition of carbon fixation, O2 evolution, photosynthetic electron transport chain and ATP content of A. doliolum and C. vulgaris by Cu, Ni and Fe. Although the mode of inhibition of photosynthetic electron transport chain of both the algae was similar, PS II depicted greater sensitivity to the test metals used. The toxicity in both organisms was Cu > Ni > Fe. A. doliolum was, however, more sensitive to Cu and Ni, and C. vulgaris to Fe. Toxicity was generally dependent on metal uptake, which in turn was dependent on their concentrations in the external medium. A partial restoration of nutrient uptake, carbon fixation, and enzyme activities following supplementation of exogenous ATP suggests that ATP regulates toxicity through chelation.

Anisotropic transport of microalgae Chlorella vulgaris in microfluidic channel

In this work, we study the regional dependence of transport behavior of microalgae Chlorella vulgaris inside microflu- idic channel on applied fluid flow rate. The microalgae are treated as spherical naturally buoyant particles. Deviation from the normal diffusion or Brownian transport is characterized based on the scaling behavior of the mean square displacement （MSD） of the particle trajectories by resolving the displacements in the streamwise （flow） and perpendicular directions. The channel is divided into three different flow regions, namely center region of the channel and two near-wall boundaries and the particle motions are analyzed at different flow rates. We use the scaled Brownian motion to model the transitional characteristics in the scaling behavior of the MSDs. We find that there exist anisotropic anomalous transports in all the three flow regions with mixed sub-diffusive, normal and super-diffusive behavior in both longitudinal and transverse directions.

Biosorption of chromium from electroplating and galvanizing industrial effluents under extreme conditions using Chlorella vulgaris

Hexavalent chromium [Cr(Ⅵ)] is a toxic oxidized form and an important metal pollutant in the water bodies. Biosorption of chromium(VI)offers a potential alternative to conventional metal removal methods. Dried biomass of Chlorella vulgaris was used as biosorbent for the removal of Cr(VI) from electroplating and galvanizing industry effluents as a function of biosorbent dosage, contact time, pH, salinity and initial metal ion concentration. Batch experiments were conducted for biosorption and the optimum conditions were 1 g/L biomass, 4 h contact time, pH 2 and 2.893 mS/cm of electrical conductivity. The chromium biosorption was strictly pH dependent with a maximum Cr removal of 63.2 mg/L at pH 2. Highest Cr removal at a concentration of 81.3 mg/L was observed at Electrical conductivity(EC) value of 2.893 mS/cm. A comparison of Langmuir and Freundlich isotherm models revealed that Freundlich isotherm model fitted the experimental data based on R^2, q_(max) and standard error values. The results suggest that C. vulgaris biomass could be considered a promising low-cost biosorbent for the removal of Cr(VI) from electroplating and galvanizing industry effluents.

Effects of 1-Naphthol on Physiological Parameters of Chlorella vulgaris

[ Objective] This study aimed to investigate the effects of 1-naphthol on primary productivity in aquatic ecosystems. [ Method] The effects of 1-naphthol on biomass, chlorophyll a content, malondialdehyde （MDA） content and soluble protein content of Chlorella vulgaris were investigated. [ Result] 1-Naphthol could significantly inhibit the growth of Chlorella vulgaris. On the 7^th d, the biomass of Chlorella vulgaris exposed to 1.0, 3.0, 6.0, 12.0 and 18.0 mg/L 1-naphthol was 60.26%, 48.15%, 41.32%, 28.59% and 27.10% of that in control respectively, exhibiting a significant dose-response relationship. Chlorella vulgaris was exposed to different concentrations of 1-naphthol （0, 1,5 and 10 mg/L） for 7 d ; with the increase of 1-naphthol concentration, chlorophyll a content of Ch/orella vulgaris was reduced, MDA content increased gradually, soluble protein content increased first and then declined. [ Conclusion] According to the experimental results, 1-naphthol can significantly inhibit the growth of Chlorella vulgaris, thus reducing primary productivity in aquatic ecosystems.

Increasing the Protein Amount of Chlorella vulgaris [Chlorphyta] Strains Isolated from Different Fresh Water Ponds

Chlorella vulgaris is a single-cell, spherical green algae and one of the microalgae on which many applied studies are conducted. In the present study, five strains displaying fast and efficient reproduction were chosen among 11 C. vulgaris strains isolated from different fresh water ponds and their cell numbers and the amount of chlorophyll a, protein, lipid, cellulose and carbohydrate were examined. The main goal of the study is to investigate increasing the biochemical contents especially the protein content of C. vulgaris strains in different mediums. In the present study, cell densities were determined through cell count for five days. In parallel with cell count, their chlorophyll a content was determined. The highest cell density was observed with C. vulgaris TOH （Tourism and Hotel Management Pond） strain as 5.5 × 104 h/mL, and the chlorophyll a content as 4.3 × 102 mg/m3. The highest intracellular protein amount was determined with C. vulgaris GUH （Gazi University Rectorship Pond） （0.061 g/100 mL） and the highest lipid amount was attained with C. vulgaris UIK （Ulus Construction Well） strain as 0.019 g/100 mL. The process of increasing the intracellular protein amount in C. vulgaris GUH strain was carried out in Prat, Yagojinski and Chlorella medium. The results indicated that Chlorella medium increased the intracellular protein amount.

The Effects of Cells Density Arrangement in Chlorella vulgaris Culture to C02 Fixation and Essential Substances Production

Global warming that triggered the climate change is largely due to increased CO2 concentrations. Utilization of Chlorella sp. to reduce CO2 gas is a promising potential. Chlorella can efficiently reduce CO2 and easily be adapted into the photobioreactor system engineering. In this research, the type of microalgae which is used is Chlorella vulgaris in Benneck medium. The system of used reactor is mid-scale bubble column photobioreactor flowed by air which contains 5% CO2. Chlorella vulgaris biomass production will be increased by adjusting the cell density in the photobioreactor. These arrangements will be implemented through a continuous treatment of cell entrapment. The arrangement of cell density in continuous reactor has been proven to increase production of Chlorella vulgaris biomass about 1.25 times more than cultivation without arrangement of cell density by using the same number of inoculums. The results also have shown that the average rate of CO2 fixation and Carbon Transfer Rate （CTR） are obtained at cell entrapment condition about 17 times larger. Continuous cellular entrapment method is very potential to be developed as a method for the production of biomass. Lipids and carotene that have been produced from Ch. vulgaris respectively are 18.24% and 9.42 ppm.

Acute Toxicity Effects of 2,2',4,4',5,5'-Hexabromodiphenyl Ether on Chlorella vulgaris

[Objectives] This study was conducted to investigate the toxic effects of 2,2',4,4',5,5'-hexabromobiphenyl ether on Chlorella vulgaris and provide basic data for protecting aquatic ecosystems. [Methods] The acute toxicity effects of 2,2',4,4',5,5'-hexabromodiphenyl ether on C. vulgaris was investigated by the semi-static water contacting acute toxicity method. [Results] The 48,72 and 96 h-EC(50) of 2,2',4,4',5,5'-hexabromodiphenyl ether to C. vulgaris were23. 58,18. 71 and 14. 75 μg/L,respectively,and the safe concentration of 2,2',4,4',5,5'-hexabromodiphenyl ether to C. vulgaris was 1. 475μg/L. For the water solubility of 2,2',4,4',5,5'-hexabromodiphenyl ether is extremely low( 1 μg/L),it could not cause the acute poisoning death of C. vulgaris. According to the grading standards for the assessment of the toxicity on algae,2,2',4,4',5,5'-hexabromodiphenyl ether was extremely highly toxic to C. vulgaris. [Conclusions]The extremely high toxicity of 2,2',4,4',5,5'-hexabromodiphenyl ether to C. vulgaris shows that it has heavy potential harm to aquatic ecosystems and the maximum residue limit standards of 2,2',4,4',5,5'-hexabromodiphenyl ether in water should be formulated to better protect aquatic ecosystems.

Assessment of Biomass Productivities of Chlorella vulgaris and Scenedesmus obliquus in Defined Media and Municipal Wastewater at Varying Concentration of Nitrogen

Microalgae are emerging as one of the most promising long-term sustainable sources of renewable energy. Studies were conducted on two freshwater Chlorophytes, Chlorella vulgaris and Scenedesmus obliquus to evaluate heterotrophic growth rate and biomass productivity in filter-sterilized defined medium (BG 11) and municipal wastewater, both with varying concentrations of nitrogen (N). For each isolate, cultures were separately incubated in triplicate at room temperature with constant agitation on a shaker at 150 rpm for 9 days. In 0.25 mg N/L BG11 medium, the growth rate and biomass productivity of C. vulgaris were 0.28 day-1 and 3.5 g·L-1, respectively. In wastewater, the same amount of N addition resulted in a higher growth rate 0.44 day-1 and associated biomass productivity of 4.96 g·L-1. Increasing N levels to 0.5 mg N/L in BG11 caused an increase in growth rate (0.37 day-1) and biomass productivity (4.28 g·L-1), while the increase in N in wastewater caused growth to decline to 0.32 day-1 with decreased biomass productivity of 2.19 g·L-1. A further increase in N to 1.0 mg N/L in BG11 caused an increase in the growth rate (0.43 day-1) and a decrease in biomass productivity (3.64 g·L-1), while in wastewater, growth rate and productivity of C. vulgaris were 0.32 day-1 and 2.31 g·L-1, respectively. Overall, C. vulgaris grew faster and produced greater biomass than S. obliquus under comparable conditions. Based on high growth rate and biomass productivity of C. vulgaris, it could be a potential candidate for further consideration for simultaneous wastewater treatment and biofuel production.

Determination of the Concentration of Chlorella vulgaris Beij by Visible Spectrophotometer

[Objective]This study aimed to provide technical parameters for large-scale production and industrial development of Chlorella vulgaris Beij.[Method]By using 721 visible spectrophotometer,the transmitted light intensity of different concentrations of C.vulgaris liquid under different wavelengths was measured to normalize the absorbance data,thus determining the optimal wavelength for the growth of C.vulgaris and the concentration of C.vulgaris liquid under the optimal wavelength.The regression equation of absorbance at the optimal wavelength and C.vulgaris concentration was obtained to derive the formula between equivalent concentration C and transmitted light intensity I,thus calculating the equivalent concentration of 0.25-fold C.vulgaris liquid under different transmittance.[Result]The transmitted light intensity of C.vulgaris reached the peak at 670 nm,indicating that 670 nm is the optimal wavelength for measuring C.vulgaris concentration.The regression equation between absorbance and C.vulgaris concentration was y=-1.479 7＋0.987x,with a correlation coefficient of 0.991,suggesting a high correlation between these two factors.The formula of equivalent concentration C and transmitted light intensity I was C=-ln（I/I0）/LK,where L indicates the optical path length（cuvette width）and K indicates the average cross-sectional area of C.vulgaris cells.[Conclusion]It is feasible to investigate indirectly the concentration of C.vulgaris with a spectrophotometer,which can be used to determine the optimal dehydration time of C.vulgaris.

Removal of Ammonia from Wastewater Effluent by Chlorella Vulgaris

The capability of Chlorella vulgaris to remove nitrogen in the form of ammonia and/or ammonium ions from wastewater effluent in a local wastewater treatment plant （i.e., the Mill Creek Plant in Cincinnati, Ohio, U.S.A.） was studied. The wastewater effluent leaving the plant was found to include high concentra- tions of nitrogen （7.7±0.19 mg/L） （ammonia （NH3） and/or ammonium ion （NH4＋）） and total inorganic carbon （58.6±0.28 mg/L） at pH 7, and to be suitable for growing Chlorella vulgaris. When Chlorella vulgaris was cul- tivated in a batch mode under a closed system, half of the nitrogen concentration was dramatically removed in 48 h after a 24-h lag-phase period. Total inorganic carbon concentration also concomitantly decreased during the rapid growth-phase. The total biomass weight gained during the entire cultivation period balanced out well with the total amount of inorganic carbon and nitrogen removed from the culture medium. These results indicate that wastewater can be synergistically used to polish residual nutrients in wastewater as well as to cultivate microalgae for biofuel production.

Inhibition effects of benzalkonium chloride on Chlorella vulgaris induced corrosion of carbon steel

In this work,a surfactant,benzalkonium chloride(BAC),was used to study its effects on both the growth of Chlorella vulgaris and the corrosion caused by its biofilm.Experimental results indicated that BAC at a low concentration of 3 mg/L suppressed C.vulgaris growth and achieved 81%corrosion inhibition based on weight loss reduction.The inhibition effects increased when the BAC dosage was increased.At 30 mg/L,the corrosion inhibition increased to 95%.Electrochemical results supported surface pitting analysis,weight loss results data and confirmed the corrosion inhibition.

Chlorella vulgaris triggers apoptosis in hepatocarcinogenesis-induced rats

Chlorella vulgaris （CV） has been reported to have antioxidant and anticancer properties. We evaluated the effect of CV on apoptotic regulator protein expression in liver cancer-induced rats. Male Wistar rats （200-250 g） were divided into eight groups： control group （normal diet）, CDE group （choline deficient diet supplemented with ethionine in drinking water to induce hepatocarcinogenesis）, CV groups with three different doses of CV （50, 150, and 300 mg/kg body weight）, and CDE groups treated with different doses of CV （50, 150, and 300 mg/kg body weight）. Rats were sacrificed at various weeks and liver tissues were embedded in paraffin blocks for immunohistochemistry studies. CV, at increasing doses, decreased the expression of anti-apoptotic protein, Bcl-2, but increased the expression of pro-apoptotic protein, caspase 8, in CDE rats, which was correlated with decreased hepatoctyes proliferation and increased apoptosis as determined by bromodeoxy-uridine （BrdU） labeling and terminal deoxynucleotidyl transferase mediated dUTP nick-end labeling （TUNEL） assay, respectively. Our study shows that CV has definite chemopreventive effect by inducing apoptosis via decreasing the expression of Bcl-2 and increasing the expression of caspase 8 in hepatocarcinogenesis-induced rats.

Influence of CaO and HZSM-5 on oxygen migration in Chlorella vulgaris polysaccharide pyrolysis

With polysaccharides of Chlorella pyrenoidosa as the raw material,the effects of CaO and HZSM-5 on the yield of bio-oil and the oxygen content in each phase in the pyrolysis of Chlorella vulgaris polysaccharides(CVP),which occurred in a tube furnace at 600℃,were comprehensively investigated.The reaction path of deoxidation was also analyzed by TG,GC and GC-MS.The GC-MS analysis of liquids showed that liquids from the pyrolysis of chlorella polysaccharides included a range of light oxygenated compounds(e.g.,furans,ketones and phenols),and the oxygen content of furan compounds decreased significantly with CaO and HZSM-5.Compared with the direct pyrolysis of polysaccharides(CVP),the catalytic pyrolysis contributed to the decrease in the oxygen content of organic components by 7.32%and 5.76%.The GC analysis showed that there was a remarkable downtrend in the release of oxygen-containing gas(CO and CO_(2)),and the emission of CO_(2) decreased from 53.11%to 32.92%.The results of the thermogravimetric analysis indicated that the reaction paths of deoxidation in the pyrolysis process of polysaccharides(CVP)with CaO and HZSM-5 varied from those of the direct pyrolysis process:the catalytic pyrolysis with HZSM-5 promoted the conversion of carbohydrate from furans to aromatics over strong acid sites,which was consistent with previous studies;CaO not only acted as a catalyst but also participated in the reaction to change the reaction paths.All results and findings can help to further understand the thermochemical utilization of CPP for bio-oil.