Chlorella Minutissima Research Articles

Implication of Industrial Waste for Biomass and Lipid Production in Chlorella minutissima Under Autotrophic, Heterotrophic, and Mixotrophic Grown Conditions.

Following the diminishing hopes from the first and second generation biofuels, mainly due to the limitations of land availability, feed stock requirements, and complicated pre-treatments, third generation biofuels from microalgae are becoming a priority in the current scenario. The present study focuses on comparison and optimization of lipid accumulation efficiency in algal strain Chlorella minutissima grown under autotrophic, heterotrophic, and mixotrophic modes of nutrition, employing various carbon sources obtained from cheap industrial wastes such as glucose, acetate, and glycerol. Other pertinent factors such as the effect of various nitrogen sources, effect of salinity on the cell growth, and lipid accumulations in the algal cells were also studied. The results suggested that C. minutissima can grow efficiently under autotrophic, heterotrophic, and mixotrophic modes of nutrition. C. minutissima cells were capable of utilizing other non-popular carbon sources such as glycerol and acetate collected as waste products from different industries along with commonly used glucose. The maximum biomass concentration (8.9 g/L) and lipid content (36.19 %) were found in heterotrophic mode of nutrition. Our findings indicated that C. minutissima can efficiently utilize these cheaper carbon sources from industrial waste products for its growth and the production cost of various bioenergy sources can be reduced significantly.

Elevated CO2 concentration impacts cell wall polysaccharide composition of green microalgae of the genus Chlorella.

The effect of CO2 concentration on the relative content of starch, lipid and cell wall carbohydrates in microalgal biomass was investigated for the four following Chlorella strains: C.vulgaris (UTEX 259), C.sorokiniana (UTEX 2805), C.minutissima (UTEX 2341) and C.variabilis (NC64A). Each strain had a different response to CO2 concentration. The starch content was higher in UTEX259 and NC64A cultured with 2% CO2 in the air supply than in cells cultured with ca. 0·04% CO2 (ambient air), while starch content was not affected for UTEX 2805 and UTEX 2341. The lipid content was higher in Chlorella minutissima UTEX 2341 cultured in 2% CO2 than in cells cultured in ambient air, but was unchanged for the other three strains. All four Chlorella strains tended to have a higher percentage of uronic acids and lower percentage of neutral sugars in their cell wall polysaccharide complement when grown with 2% CO2 supply. Although the percentage of neutral sugars in the cell walls varied with CO2 concentration, the relative proportions of different neutral sugar constituents remained constant for both CO2 conditions. The results demonstrate the importance of considering the effects of CO2 on the cell wall carbohydrate composition of microalgae. Microalgae have the potential to produce products that will reduce society's reliance on fossil fuels and address challenges related to food and feed production. An overlooked yet industrially relevant component of microalgae are their cell walls. Cell wall composition affects cell flocculation and the recovery of intracellular products. In this study, we show that increasing CO2 level results in greater cell wall polysaccharide and uronic acid content in the cell walls of three strains of microalgae. The results have implications on the management of systems for the capture of CO2 and production of fuels, chemicals and food from microalgae.

Phycoremediation potential of Chlorella minutissima on primary and tertiary treated wastewater for nutrient removal and biodiesel production

The integrated approach of wastewater treatment and microalgal biodiesel production based on phycoremediation is a promising process. The experiment was set up to study the phycoremediation potential of Chlorella minutissima to remove the pollution load from primary treated IARI’s (Indian Agricultural Research Institute) wastewater and tertiary treated CETP (Common Effluent Treatment Plant) wastewater and evaluation of biomass for biodiesel after harvesting. The physical and chemical parameters of wastewater quality such as nitrate, phosphate, potassium, EC, TDS, BOD, COD, etc., were studied. C. minutissima removed about 90–98% TDS, 70–80% N, 60–70% P and 45–50% K from the wastewater within 12 days. The level of BOD and COD were reduced by 60 and 75%, respectively. The algal biomasses were harvested for biodiesel extraction. The highest specific lipid productivity for CETP and IARI wastewater was 0.171 and 0.132g-lipidsg-cell−1day−1, respectively. The oleic acid had increased 59.6% in CETP wastewater in comparison to IARI wastewater. The results of this study concluded that algal strain C. minutissima is not only a means for remediation of pollutant load, but it can also be used as potential driving force for biodiesel production.

An advanced hybrid medium optimization strategy for the enhanced productivity of lutein in Chlorella minutissima

The biosynthesis of lutein in microalgae is critically influenced by the media components, which either increase the key enzyme concentrations or act as cofactors for the enzymes involved in lutein metabolic pathway. In this study, an advanced medium optimization strategy, involving Plackett–Burman design for screening the Bold's Basal Medium (BBM) components, followed by artificial neural network modeling and particle swarm optimization (ANN–PSO), was implemented for improving the productivity of lutein in Chlorella minutissima. In this two-step hybrid optimization endeavor, Plackett–Burman analysis helped select NaNO3, KH2PO4, MnCl2·4H2O and CuSO4·5H2O as critical components that were subsequently optimized by ANN–PSO technique. On experimental validation, the optimized medium resulted in three-fold greater lutein productivity, as compared to BBM. Furthermore, the maximum lutein productivity of 3.45±0.07mgL−1d−1 was obtained upon cultivation of C. minutissima in a 2-L airlift photobioreactor using the optimized medium with appropriate process conditions. Thus, the hybrid optimization approach was instrumental in enhancing lutein productivity by about 187%, which was significantly higher than the relevant values reported in literature. To our knowledge, this is the first report on the application of an integrated optimization strategy for the enhanced lutein productivity in C. minutissima as a model photo-autotrophic organism.

Lipid production combined with biosorption and bioaccumulation of cadmium, copper, manganese and zinc by oleaginous microalgae Chlorella minutissima UTEX2341

Algae lipid production combined with heavy metal removal is a cost-effective and environment-friendly method for algae biofuel production and hazardous waste treatment. Chlorella minutissima UTEX 2341 had strong resistance to cadmium, copper, manganese and zinc ions under heterotrophic culture condition and could efficiently remove them through intracellular accumulation and extracellular immobilization. Meanwhile, lipid accumulation was not inhibited by heavy metals. Instead, the algae lipid content significantly increased by 21.07% and 93.90%, respectively with the addition of cadmium and copper. Furthermore, the heavy metal residue in lipid was within μg range and satisfied the commercial standard. This artificial wastewater-algae biofuel-heavy-metal integrated utilization technology offered a new alternative solution to the problems of energy shortage and environmental pollution.

A novel bioflocculant produced by a salt-tolerant, alkaliphilic and biofilm-forming strain Bacillus agaradhaerens C9 and its application in harvesting Chlorella minutissima UTEX2341

A novel bioflocculant MBF-C9 produced by a salt-tolerant, alkaliphilic Bacillus agaradhaerens C9 was investigated in this study. The effects of culture conditions such as initial pH, carbon source, nitrogen source, C/N ratio, and NaCl concentrations on MBF-C9 production were studied. The result showed that 4.65g/L purified MBF-C9 was extracted with the following optimized conditions: 10g/L glucose as carbon source, 10g/L yeast extract as nitrogen source and initial pH 10.2. The MBF-C9 contained 65.42% polysaccharides, 4.70% proteins, and 1.65% nucleic acids. The highest flocculating rate of 95.29% for kaolin suspension was achieved at a dosage of 1.5mg/L, pH 6.53 and 29°C, and the highest flocculating rate of 80.63% for Chlorella minutissima UTEX2341 cells was observed at a dosage of 8mg/L, which is much lower than that used in previous study. In addition, C9 could form biofilm under alkaline condition. Therefore, C9 has the potentials to be used in harvesting microalgae or as a bacterial agent to strengthen the treatment of alkaline wastewater.

Cellular and molecular damage caused by high UV-C irradiation of the cave-harvested green alga Chlorella minutissima: Implications for cave management

Algal biofilms in caves visited by tourists are a significant threat to karstic formations. They cause a dirty greenish appearance on cave formations and provoke biophysical and biochemical erosion. Seeking an alternative to the use of chemicals, we investigated the effects of high doses of UV-C irradiation on the green algae Chlorella minutissima-dominating biofilms in the Moidons Cave (Jura, France). The algae were harvested and subjected to laboratory subculturing under conditions similar to the conditions in the cave (low temperatures and a short photoperiod with weak light intensities). After one or two doses of UV-C of 150 or 300 kJ m−2, the samples were incubated for 21 days, and the physiological parameters (pigment concentrations and photosynthetic activity) were monitored every seven days. Hydrogen peroxide (H2O2) and malondialdehyde (MDA) concentrations were also measured to detect oxidative stress, and analyses of DNA integrity were performed. The results showed that all UV-C treatments caused chlorophyll bleaching with completely inhibited metabolic activity. In addition, high UV-C doses caused oxidative stress and programmed cell death. Irradiation was tested on a natural biofilm that proliferated in the Moidons Cave. Colorimetric measurements of the treated biofilm indicated that UV-C treatment caused significant chlorophyll bleaching.

Effects of nitrate and ph sphate on the growth and total lipid contents of Chlorella oocystoides and Chlorella minutissima ( Chlorophyceae)

The effects of different concentrations (0.1, 0.5, 1.5 and 4.5 ppm) of nitrate (NO3-) as NaNO3 and phosphate ( PO4---) asK2HPO4 on growth parameters (dry weight, optical density, total pigments), total lipid, total carbohydrate and total protein contents of Chlorella oocystoides and Chlorella minutissima cultured were followed for 7 days.The growth parameters (dry weight, optical density and total pigments), total carbohydrate and total protein contents ofChlorella oocystoides and Chlorella minutissima were significantly increased up to the concentration 1.5 ppm of NO3- andPO4--- Under relatively higher concentration 4.5 ppm of NO3- and PO4--- all these parameters were significantly decreased. On the contrary, the total lipid contents were significantly increased up to the 0.1 ppm of NO3- and PO4--- However, under relatively higher concentration 4.5 ppm of NO3- and PO4---, the total lipid contents were significantly decreased

Effects of Escherichia coli on mixotrophic growth of Chlorella minutissima and production of biofuel precursors.

Chlorella minutissima was co-cultured with Escherichia coli in airlift reactors under mixotrophic conditions (glucose, glycerol, and acetate substrates) to determine possible effects of bacterial contamination on algal biofuel production. It was hypothesized that E. coli would compete with C. minutissima for nutrients, displacing algal biomass. However, C. minutissima grew more rapidly and to higher densities in the presence of E. coli, suggesting a symbiotic relationship between the organisms. At an initial 1% substrate concentration, the co-culture produced 200-587% more algal biomass than the axenic C. minutissima cultures. Co-cultures grown on 1% substrate consumed 23–737% more of the available carbon substrate than the sum of substrate consumed by E. coli and C. minutissima alone. At 1% substrate, total lipid and starch productivity were elevated in co-cultures compared to axenic cultures indicating that bacterial contamination was not detrimental to the production of biofuel precursors in this specific case. Bio-fouling of the reactors observed in co-cultures and acid formation in all mixotrophic cultures, however, could present challenges for scale-up.

Effect of light on growth and endogenous hormones in Chlorella minutissima (Trebouxiophyceae)

Plant growth regulators (PGRs) play an important role in mediating growth and stress responses in plants. Light influences PGRs concentrations in vascular plants. The effect of light on growth and endogenous PGR concentrations in microalgae was investigated in the present study. Chlorella minutissima MACC 360 was grown in 14:10 h light:dark (L:D), continuous dark (CD) and continuous dark with the addition of 5 g L−1 glucose (CD + G) for 48 h. Cultures were synchronized in the L:D cultures, increasing in size during the light period and dividing during the dark period. C. minutissima cells did not increase in size or undergo cell division in CD cultures. In CD + G conditions, the cultures were no longer synchronized but did continue to increase in cell size and constantly underwent cell division although fewer cells divided than in the L:D cultures. Endogenous auxin and cytokinin concentrations increased and gibberellin concentrations decreased over time in the actively growing cultures (L:D and CD + G) but did not increase in the CD cultures. The largest increase in indole content was in the CD + G cultures while the L:D cultures had the largest cytokinin increase. Brassinosteroid concentrations decreased over time in all the cultures including those grown in CD conditions. Abscisic acid (ABA) concentrations were low and only increased in the CD cultures. These results show that endogenous PGRs were affected by the light regime and/or culture growth.

TOXICITY OF SOME MICROALGAE TO THE CITRUS BROWN MITE EUTETRANYCHUS ORIENTALIS (KLEIN)

The citrus brown mite Eutetranychus orientalis is a common spider mite, which is a harmful pest in citrus orchards in Egypt. The toxic activities of some microalgae isolated from Egyptian soils and water are studied against E. orientalis. The toxicity of whole cultures (culturing broth and cell) of seven algal strains was tested against E. orientalis. Three strains out of the tested seven strains showed higher toxicity with mortality percentage reached up to 99 % after 72 h recorded against adult individuals of E. orientalis with a strain of Chlorella minutissima. The toxic activity was also tested against immature individuals and eggs of E. orientalis. The highest mortality percent against immature individuals was observed with C. oocystoids. Eggs were not affected with any of the tested algal strains. When cells and culturing broth were tested separately the activity was higher in culturing broth which means that this activity is due to some extracellular metabolites secreted by algae in their culturing medium. When the organic solvent n-butanol was used to extract the active constituents from the whole cultures (culturing broth and cell), the activity was found to be higher in aqueous phase compared to the butanolic phase.

One step closer to eliminating the nomenclatural problems of minute coccoid green algae: Pseudochloris wilhelmii, gen. et sp. nov. (Trebouxiophyceae, Chlorophyta)

‘Chlorella’ and ‘Nannochloris’ were traditional genera of minute coccoid green algae with numerous species described in the past century, including isolates used as experimental test organisms. In the last few years, the introduction of DNA-based phylogenetic analyses resulted in a large number of taxonomic revisions. We investigated and reclassified a taxonomically problematic group within the Trebouxiophyceae (comprising ‘Nannochloris eucaryotum’ UTEX 2502, ‘N. eucaryotum’ SAG 55.87 and ‘Chlorella minutissima’ SAG 1.80), distantly related to the recently described Chloroparva isolates (97.5–97.9 % 18S rRNA gene pairwise similarity). Cryopreserved material of SAG 55.87 was selected as holotype for a novel species – Pseudochloris wilhelmii Somogyi, Felföldi & Vörös – whose phylogenetic position confirmed the proposal of a new genus. Pseudochloris wilhelmii had spherical to oval cells with an average diameter of 2.6 × 2.8 µm and a simple ultrastructure characteristic of small green algae. Vegetative cells sometimes contained several lipid droplets occupying a large portion of the cells. The cell wall consisted of an outer trilaminar layer and an inner microfibrillar sheet. Cells divided by autosporulation, forming two or four daughter cells per autosporangium. The pigment composition was typical of green algae, with chlorophylls a and b, and lutein as the dominant carotenoid.

Chromatographic characterization of triacylglycerides and fatty acid methyl esters in microalgae oils for biodiesel production

The fatty acid methyl ester and triacylglyceride content of the oils of seven microalgae species were analyzed by gas chromatography in order to predict the influence of fatty acid content on the physicochemical properties of biodiesel. Among the seven species examined, Chlorella minutissima and Arthrospira platensis showed biodiesel production potential, while fatty acids from Chlorella vulgaris, Cyclotella sp., Cylindrotheca closterium, Entomoneis alata, and A. platensis have great economic values, with application in the pharmaceutical and food industries. The oxidative stability of the biodiesel produced from C. minutissima will be high, but there will be problems with cold filter plugging point temperatures. These can be solved by mixing C. minutissima biodiesel with a biodiesel like that from Ankistrodesmus gracilis that is rich in di- or tri-unsaturated fatty acids. Triacylglyceride analysis made it possible to tell if a microalgae oil is composed mainly of triacylglycerides or of some other class of lipids.

Biochemical composition of green alga Chlorella minutissima in mixotrophic cultures under the effect of different carbon sources

Mixotrophic growth of Chlorella minutissima with carbon supplements such as glucose, glycerol, succinate, molasses and press mud resulted in maximum biomass accumulation in glucose supplemented culture. Lipid content was maximum with molasses followed by press mud, fatty acid compositions of which also were best suited for biodiesel production.

Effect of nitrate on lipid production by T. suecica, M. contortum, and C. minutissima

Abstract Microalgae are an alternative and sustainable source of lipids that can be used as a feedstock for biodiesel production. Nitrate is a good nitrogen source for many microalgae and affects biomass and lipid yields of microalgae. In this study, the effect of nitrate on cell growth and lipid production and composition in Monoraphidium contortum, Tetraselmis suecica, and Chlorella minutissima was investigated. Nitrate affected the production of biomass and the production and composition of lipids of the three microalgae tested. Increasing the nitrate concentration in the culture medium resulted in increased biomass production and higher biomass productivity. Furthermore, increasing the nitrate concentration resulted in a reduction in lipid content and productivity in M. contortum; however, the opposite effect was observed in T. suecica and C. minutissima cultures. C. minutissima and M. contortum lipids contain high levels of oleic acid, with values ranging from 26 to 45.7% and 36.4 to 40.1%, respectively. The data suggest that because of its high lipid productivity (13.79 mg L−1 d−1) and high oleic acid productivity (3.78 mg L−1 d−1), Chlorella minutissima is a potential candidate for the production of high quality biodiesel.

Improvement in microalgae lipid extraction using a sonication-assisted method

Oil extraction from microalgae stands as a huge challenge for bioproducts and biofuels production, mainly due to high costs and energy requirement. In this study, maximum cell density, growth curve, lipid content, daily biomass and oil productivities were determined for Chlorella minutissima, Thalassiosira fluviatilis and Thalassiosira pseudonana, strains maintained at the Instituto Oceanográfico of the Universidade de São Paulo. The microalgal dry biomass was used to evaluate the lipid extraction methodology regarding a sonication bath as pretreatment cell disruption technique followed by vortex mixing and n-hexane as solvent. The results showed a lipid content of 15.5% (% dry weight) for C. minutissima, 40.3% for T. fluviatilis and 39.5% for T. pseudonana. To determine the feasibility of using microalgae into the biofuel industry is essential to consider the daily biomass and oil productivities along with the lipid content, as could be evidenced in C. minutissima. The results indicate that the sonication-assisted method with n-hexane was efficient for lipid extraction in these strains, suggesting a favorable potential for biodiesel production.

Effect of carbon dioxide on the rheological behavior of submerged cultures of Chlorella minutissima in stirred tank reactors

The objective of this study was to quantify the effect of algal biomass concentration on the rheology of the algal culture broth. Batch cultivations of Chlorella minutissima were carried out with air and carbon dioxide in a stirred tank bioreactor with a working volume of 1.8 L. The apparent viscosity of the culture broth was significantly affected by the cell mass concentrations in the bioreactor. Culture broth containing 50 g/L cell mass from air fed was found to exhibit an apparent viscosity of 1.52 mPa.s. The apparent viscosity of the carbon‐dioxide‐fed cultivations was found to increase by 20% at a shear rate of 100 s−1. The flow behavior of the system was adequately described by the Herschel–Bulkley model with a small yield stress.

Quantitative and Qualitative Aspects of Bacterial Communities Associated with Cultures of Chlorella minutissima

Microbiological conditions in 200 L cultures of microalgae Chlorella minutissima in polyethylene sleeves were examined. The influence of addition of antibiotic (nitrofurantoin) was studied. Samples were taken 2, 5, 9, and 14 d after inoculation and were spread on solid Zobell medium and on thiosulfate citrate bile salts sucrose agar for the enumeration of total numbers of bacteria and numbers of presumptive Vibrio bacteria, respectively. Microalgae grew well in the cultures and high numbers of culturable bacteria (about 107 bacteria/mL) were present in all microalgae cultures. Addition of antibiotic in 200‐L cultures of C. minutissima did not result in a significant decrease of total counts of bacteria per unit volume (P > 0.05), while the numbers of presumptive Vibrio were higher in cultures added antibiotic compared with the control treatment (P < 0.05). The numbers of presumptive Vibrio per unit volume and the percentage of fast‐growing bacteria were higher in microalgae supernatant than in noncentrifuged samples. Representative colonies were taken from all samples and in total 649 bacterial strains were isolated during this study. A range of phylotypes was identified by amplification and sequencing of the 16S rDNA gene and phylogenetic trees were constructed. Several members of the Roseobacter clade were dominant among the culturable isolates.

Erratum to: Accumulation of Lipid Production in Chlorella minutissima by Triacylglycerol Biosynthesis-Related Genes Cloned from Saccharomyces cerevisiae and Yarrowia lipolytica

Erratum to: Accumulation of Lipid Production in Chlorella minutissima by Triacylglycerol Biosynthesis-Related Genes Cloned from Saccharomyces cerevisiae and Yarrowia lipolytica

Microalgae of different phyla display antioxidant, metal chelating and acetylcholinesterase inhibitory activities

Methanol and hexane extracts from Tetraselmis chuii, Nannochloropsis oculata, Chlorella minutissima and Rhodomonas salina were evaluated for total phenolic contents, radical scavenging activity (RSA), metal chelating potential against copper and iron ions and acetylcholinesterase (AChE) inhibition. Only the methanol extracts contained phenolic compounds. The hexane extracts had the highest RSA. The extracts had a higher capacity to chelate Fe2+ ions, more pronounced in the lowest concentration of the hexane extracts with values ranging from 73.3±3.3% (R. salina) to 97.5±1.1% (N. oculata). The highest AChE inhibitory activity was found in the hexane extracts at 10mg/ml of C. minutissima (79.3±1.9%), T. chuii (85.7±0.7%) and R. salina (81.5±7.5%). GC-MS analysis indicated polyunsaturated fatty acids and steroids as the most abundant compounds in the hexane extracts. The species under study provide a valuable source of antioxidants, metal chelators and AChE inhibitors.