Mixotrophic and photoautotrophic cultivation of 14 microalgae isolates from Saskatchewan, Canada: potential applications for wastewater remediation for biofuel production

Northern regions are generally viewed as unsuitable for microalgal biofuel production due to unfavorable climate and solar insolation levels. However, these conditions can potentially be mitigated by coupling microalgal cultivation to industrial processes such as wastewater treatment. In this study, we have examined the biomass and lipid productivity characteristics of 14 microalgae isolates (Chlorophyta) from the Canadian province of Saskatchewan. Under both photoautotrophic and mixotrophic cultivation, a distinct linear trend was observed between biomass and lipid productivities in the 14 SK isolates. The most productive strain under cultivation in TAP media was Scenedesmus sp.-AMDD which displayed rates of biomass and fatty acid productivities of 80 and 30.7?mg?L^?1?day^?1, respectively. The most productive strain in B3NV media was Chlamydomonas debaryana-AMLs1b which displayed rates of biomass and fatty acid productivities of 51.7 and 5.9?mg?L^?1?day^?1, respectively. In 11 of the isolates tested, secondary municipal wastewater (MCWW) supported rates of biomass productivity between 21 and 33?mg?L^?1?day^?1 with Scenedesmus sp.-AMDD being the most productive. Three strains, Chlamydomonas debaryana-AMB1, Chlorella sorokiniana-RBD8 and Micractinium sp.-RB1b, showed large increases in biomass productivity when cultivated mixotrophically in MCWW supplemented with glycerol. High relative oleic acid content was detected in 10 of the 14 isolates when grown mixotrophically in media supplemented with acetate. There was no detectable effect on the fatty acid profiles in cells cultivated mixotrophically in glycerol-supplemented MCWW. These data indicate that biomass and lipid productivities are boosted by mixotrophic cultivation. Exploiting this response in municipal wastewater is a promising strategy for the production of environmentally sustainable biofuels.     

Inorganic carbon and pH effect on growth and lipid productivity of Tetraselmis suecica and Chlorella sp (Chlorophyta) grown outdoors in bag photobioreactors

There has been considerable interest in cultivation of green microalgae (Chlorophyta) as a source of lipid that can alternatively be converted to biodiesel. However, almost all mass cultures of algae are carbon-limited. Therefore, to reach a high biomass and oil productivities, the ideal selected microalgae will most likely need a source of inorganic carbon. Here, growth and lipid productivities of Tetraselmis suecica CS-187 and Chlorella sp were tested under various ranges of pH and different sources of inorganic carbon (untreated flue gas from coal-fired power plant, pure industrial CO₂, pH-adjusted using HCl and sodium bicarbonate). Biomass and lipid productivities were highest at pH 7.5 (320?±?29.9 mg biomass L^?1 day^?1and 92?±?13.1 mg lipid L^?1 day^?1) and pH 7 (407?±?5.5 mg biomass L^?1 day^?1 and 99?±?17.2 mg lipid L^?1 day^?1) for T. suecica CS-187 and Chlorella sp, respectively. In general, biomass and lipid productivities were pH 7.5?>?pH 7?>?pH 8?>?pH 6.5 and pH 7?>?pH 7.5?=?pH 8?>?pH 6.5?>?pH 6?>?pH 5.5 for T. suecica CS-187 and Chlorella sp, respectively. The effect of various inorganic carbon on growth and productivities of T. suecica (regulated at pH?=?7.5) and Chlorella sp (regulated at pH?=?7) grown in bag photobioreactors was also examined outdoor at the International Power Hazelwood, Gippsland, Victoria, Australia. The highest biomass and lipid productivities of T. suecica (51.45?±?2.67 mg biomass L^?1 day^?1 and 14.8?±?2.46 mg lipid L^?1 day^?1) and Chlorella sp (60.00?±?2.4 mg biomass L^?1 day^?1 and 13.70?±?1.35 mg lipid L^?1 day^?1) were achieved when grown using CO₂ as inorganic carbon source. No significant differences were found between CO₂ and flue gas biomass and lipid productivities. While grown using CO₂ and flue gas, biomass productivities were 10, 13 and 18 %, and 7, 14 and 19 % higher than NaHCO₃, HCl and unregulated pH for T. suecica and Chlorella sp, respectively. Addition of inorganic carbon increased specific growth rate and lipid content but reduced biomass yield and cell weight of T. suecica. Addition of inorganic carbon increased yield but did not change specific growth rate, cell weight or content of the cell weight of Chlorella sp. Both strains showed significantly higher maximum quantum yield (F_v/F_m) when grown under optimum pH.     

Extracellular β-d-xylosidase of Sclerotium rolfsii

Culture conditions are described for the production of extracellular β-d-xylosidase (xylobiase, exo-1,4-β-d-xylosidase, 1,4-β-d-xylan xylohydrolase, EC 3.2.1.37) in shake flasks by Sclerotium rolfsii. At the 1% cellulose level, a maximum activity of 0.82 U ml^?1is obtained in media containing either 1% corn steep liquor or 1% defatted coconut cake. The β-d-xylosidase has a molecular weight of 170 000 and catalyses the hydrolysis of 4-nitrophenyl-β-d-xylopyranoside optimally at pH 4.5 and 50°C. The energy of activation is 44 kJ mol^?1and the pI and K_mare 6.8 and 0.038 mm, respectively.     

Effect of photoperiod,light intensity and carbon sources on biomass and lipid productivities of Isochrysis galbana

Biomass and lipid productivities of Isochrysis galbana were optimized using nutrients of molasses (4, 8, 12 g l^?1), glucose (4, 8, 12 g l^?1), glycerol (4, 8, 12 g l^?1) and yeast extract (2 g l^?1). Combinations of carbon sources at different ratios were evaluated in which the alga was grown at three different light intensities (50, 100 and 150 μmol m^?2 s^?1) under the influence of three different photoperiod cycles (12/12, 18/6 and 24/0 h light/dark). A maximum cell density of 8.35 g l^?1 with 32 % (w/w) lipid was achieved for mixotrophic growth at 100 μmol m^?2 s^?1 and 18/6 h light/dark with molasses/glucose (20:80 w/w). Mixotrophic cultivation using molasses, glucose and glycerol was thus effective for the cultivation of I. galbana.     

Isolation of mutants of Talaromyces emersonii CBS 814.70 with enhanced cellulase activity

By a combination of genetic mutation and modification of growth medium the cellulase [see 1,4-(1,3;1,4)-β-d-glucan 4-glucanohydrolase, EC 3.2.1.4 etc.] activity of culture filtrates of Talaromyces emersonii CBS 814.70 has been increased four-fold to approximately 2 U ml^?1 and a productivity of 20–25 Ul^?1h^?1. At 50°C this system was completely stable for at least 24 h. At 60°C in static reaction mixtures 19% of the original activity was lost compared with 21% when mixtures were shaken. At 70°C the loss of activity after 4 h was 64% without shaking and 70% when shaken. The cellulase system from Trichoderma reesei was decidedly less stable than that of Talaromyces emersonii under each of the above conditions. The ability of each enzyme system, separately and together, to digest beet pulp was investigated.     

Photoautotrophic cultivating options of freshwater green microalgal Chlorococcum humicola for biomass and carotenoid production

Photoautotrophic cultivation of Chlorococcum humicola was performed in batch and continuous modes in different cultivating system arrangements to compare biomass and carotenoids’ concentration and their productivities. Batch result from stirred tank and airlift photobioreactors indicated the positive effect of increasing light intensity on growth and carotenoid production, whereas the finding from continuous cultivation indicated that carotenoid enhancement preferred high light intensity and nitrogen-deficient environment. The highest biomass (1.31?±?0.04?g?L^?1) and carotenoid (4.59?±?0.06?mg?L^?1) concentration as well as the highest productivities, 0.46?g?L^?1 d^?1 for biomass and 1.61?mg?L^?1 d^?1 for carotenoids, were obtained when maintaining high light intensity of 10 klx, BG-11 medium and 2% (v/v) CO₂ simultaneously, while the highest carotenoid content (4.84?mg?g^?1) was associated with high light intensity and nitrogen-deficient environment, which was induced by feed-modified BG-11 growth medium containing nitrate 20 folds lower than the original medium. Finally, the cultivating system arranged into smaller stirred tank photobioreactors in series yielded approximately 2.5 folds increase in both biomass and carotenoid productivities relative to using single airlift photobioreactor with equivalent working volume and similar operating condition.     

Immobilized Growth of the Peridinin-Producing Marine Dinoflagellate Symbiodinium in a Simple Biofilm Photobioreactor

Products from phototrophic dinoflagellates such as toxins or pigments are potentially important for applications in the biomedical sciences, especially in drug development. However, the technical cultivation of these organisms is often problematic due to their sensitivity to hydrodynamic (shear) stress that is a characteristic of suspension-based closed photobioreactors (PBRs). It is thus often thought that most species of dinoflagellates are non-cultivable at a technical scale. Recent advances in the development of biofilm PBRs that rely on immobilization of microalgae may hold potential to circumvent this major technical problem in dinoflagellate cultivation. In the present study, the dinoflagellate Symbiodinium voratum was grown immobilized on a Twin-Layer PBR for isolation of the carotenoid peridinin, an anti-cancerogenic compound. Biomass productivities ranged from 1.0 to 11.0 g m^?2 day^?1 dry matter per vertical growth surface and a maximal biomass yield of 114.5 g m^?2, depending on light intensity, supplementary CO₂, and type of substrate (paper or polycarbonate membrane) used. Compared to a suspension culture, the performance of the Twin-Layer PBRs exhibited significantly higher growth rates and maximal biomass yield. In the Twin-Layer PBR a maximal peridinin productivity of 24 mg m^?2 day^?1 was determined at a light intensity of 74 μmol m^?2 s^?1, although the highest peridinin content per dry weight (1.7 % w/w) was attained at lower light intensities. The results demonstrate that a biofilm-based PBR that minimizes hydrodynamic shear forces is applicable to technical-scale cultivation of dinoflagellates and may foster biotechnological applications of these abundant marine protists.     

Functional characteristics of two d-xylanases purified from Trichoderma harzianum

Two endo-1,4-β-d-xylanases (1,4-β-d-xylan xylanohydrolase, EC 3.2.1.8) were purified from Trichoderma harzianum culture filtrates. From kinetic analyses, apparent V_max and K_m values of 580 U mg^?1 protein and 0.16% d-xylan were obtained for the 20 000 dalton endo-1,4-β-d-xylanase, while values of 100 U mg^?1 protein and 0.066% d-xylan were obtained for the 29 000 dalton endo-1,4-β-d-xylanase. Substrate levels >1% (w/v) d-xylan were found to be inhibitory to both enzymes. Both d-xylanases were highly active against d-xylans obtained from various sources. Of the polymeric sugars tested, carboxymethyl cellulose was the only substrate which was hydrolysed to any extent. Little or no activity was observed against cellulose. Analyses by h.p.l.c. demonstrated the absence of hydrolytic activity by both d-xylanases on d-xylobiose. d-Xylotriose was cleaved to a limited extent by the 29 000 dalton d-xylanase only, while d-xylotetraose was hydrolysed by both. In the presence of d-xylotetraose, the 20 000 dalton d-xylanase had an associated transxylosidase activity which was not observed with the 29 000 dalton enzyme. When the solubilization assay was used, neither of the d-xylanases was inhibited by high concentrations of d-xylose and xylobiose.     

Preparation,characterization and laboratory-scale application of an immobilized glucoamylase

Glucoamylase (1,4-α-d-glucan glucohydrolase, EC 3.2.1.3) has been covalently immobilized on a polyacrylamide-type support containing carboxylic groups activated by water-soluble carbodiimide. The activity was 5.5– 6.0 units g^?1solid. The optimum pH for catalytic activity was pH 3.8. The apparent optimum temperature was found at 60°C. With soluble starch as substrate the K_m value was 14 mg ml^?1. The pH for maximum stability was pH 4.0–4.5. In the presence of 8 m urea the immobilized glucoamylase retained most of its catalytic activity but it was more susceptible to guanidinium hydrochloride than the soluble enzyme. The practical applicability of immobilized glucoamylase was tested in batch process and continuous operation.     

Improvement of biomass and fatty acid productivity in ocean cultivation of <Emphasis Type="Italic">Tetraselmis</Emphasis> sp. using hypersaline medium

In this study, hypersaline media were used for ocean cultivation of the marine microalga Tetraselmis sp. KCTC12432BP for enhanced biomass and fatty acid (FA) productivity. Hypersaline media (55, 80, and 105 PSU) were prepared without sterilization by addition of NaCl to seawater obtained from Incheon, Korea. The highest biomass productivity was obtained at 55 PSU (0.16 g L^?1 day^?1) followed by 80 PSU (0.15 g L^?1 day^?1). Although the specific growth rate of Tetraselmis decreased at salinities higher than 55 PSU, prevention of contamination led to higher biomass productivity at 80 PSU than at 30 PSU (0.03 g L^?1 day^?1). FA content of algal biomass increased as salinity increased to 80 PSU, above which it declined, and FA productivity was highest at 80 PSU. Ocean cultivation of Tetraselmis was performed using 50-L tubular module photobioreactors and 2.5-kL square basic ponds, closed- and open-type ocean culture systems, respectively. Culturing microalgae in hypersaline medium (80 PSU) improved biomass productivities by 89 and 152% in closed and open cultures, respectively, compared with cultures with regular salinity. FA productivity was greatly improved by 369% in the closed cultures. The efficacy of salinity shift and N-deficiency to enhance FA productivity was also investigated. Lowering salinity to 30 PSU with N-starvation following cultivation at 80 PSU improved FA productivity by 19% in comparison with single-stage culture without N-deficiency at 30 PSU. The results show that salinity manipulation could be an effective strategy to improve biomass and FA productivity in ocean cultivation of Tetraselmis sp.     

Optimization of the composition of the nutrient medium for cellulase and protein biosynthesis by thermophilic Aspergillus fumigatus NRC 272

An active strain of Aspergillus spp. has been selected for the production of cellulolytic enzymes and proteins when grown on peracetic acid-treated wheat straw. This strain produced a considerable amount of cellulase [see 1,4-(1,3;1,4)-β-d-glucan 4-glucanohydrolase, EC 3.2.1.4] in the extracellular supernatant and exhibited good overall cellulolytic activity, as measured using filter paper and Avicel as substrates. Also, under the same conditions the strain showed high activities of β-d-glucosidase (β-d-glucoside glucohydrolase, EC 3.2.1.21) and β-d-xylosidase (1,4-β-d-xylan xylohydrolase, EC 3.2.1.37). The maximum enzyme yields (carboxymethylcellulose activity 26.4 units ml^?1, filter paper activity 2.26 units ml^?1 and Avicel activity 16.82 units ml^?1; β-d-glucosidase 9.09 units ml^?1 and β-d-xylosidase 1.92 units ml^?1) were obtained after 96 h incubation at 45°C.     

Ulva lactuca and U. flexuosa (Chlorophyta,Ulvophyceae) cultivation in Brazilian tropical waters: recruitment,growth, and ulvan yield

Ulva spp. are used in a wide range of commercial applications, including bioremediation, food, bioenergy, pharmaceuticals, and agriculture. The sulfated polysaccharide ulvan obtained from Ulva spp. is of interest for triggering plant defenses against disease. However, the cultivation of Ulva spp. is still in its infancy. This study verified the feasibility of cultivating Ulva lactuca and Ulva flexuosa at two sites on the tropical Brazilian coast. We investigated the following: (a) methods to induce sporulation, (b) comparison of seeding ropes inoculated in vitro versus seeding at sea over 40 days, (c) production and harvest cycles at 15 and 30 days, (d) growth productivity of U. flexuosa at sea and in outdoor tanks, and (e) comparison of ulvan yields from biomass cultivated in tanks and the sea. High nutrient treatment was the most efficient method to induce sporulation (7,540?±?3,133 spores m^?1). Sea-based cultivation of U. flexuosa was only successful at one site. Seeding of ropes in vitro was more efficient than seeding at sea (0.31?±?0.20 g m^?2 day^?1), and 15-day harvest cycles were most efficient (20.1?±?1.8 % day^?1; 0.46?±?0.11 g m^?2 day^?1). Despite differences in plant growth in tanks (27.9?±?4.4 % day^?1) and at sea (20.1?±?1.8 % day^?1), the dry biomass and ulvan yields (17.7?±?5.0 %) did not differ between these systems. Cultivation of U. flexuosa was feasible at sea using in vitro seeding with a production cycle of 15 days in Brazilian tropical waters and tanks with high irradiance and enriched seawater.     

Patterns of growth and β-galactosidase production by Bifidobacteria

We investigated the patterns of growth and β-galactosidase production in the strains Bifidobacterium adolescentis GO-13, MS-42, 91-BIM, and 94-BIM and b. bifidum No.1, LVA-3, 791 on media with various carbon sources. The synthesis of β-galactosidase was shown to be associated with exponential growth of the cultures involved. The maximum specific rate of β-galactosidase synthesis of 0.20 U mg^?1 h^?1 was observed in B. bifidum LVA-3 after 3–6 h of cultivation. This value for B. adolescentis 91-BIM and 94-BIM was lower and amounted to 0.03–0.08 U mg^?1h^?1. On the medium with lactose, the highest specific growth rates for B. bifidum LVA-3 and B. bifidum No.1 were 0.38 and 0.60 h^?1, respectively, after 3–6 h of cultivation. For B. adolescentis 91-BIM and 94-BIM, this parameter peaked at 12–15 h of cultivation at 0.13 and 0.22 h^?1, respectively. The hydrolytic activity of β-galactosidase in the growth medium decreased during the stationary growth phase of the tested cultures.     

Utilization of cane molasses towards cost-saving astaxanthin production by a Chlorella zofingiensis mutant

The aim of the present study was to survey the growth and astaxanthin production of E17, an astaxanthin-rich mutant of Chlorella zofingiensis, through feeding the low-cost carbon source cane molasses. In heterotrophic batch cultivation, E17 fed with pretreated molasses achieved biomass (1.79 g L^?1 day^?1) and astaxanthin (1.99 mg L^?1 day^?1) productivities comparable to those with glucose, which were about 2- and 2.8-fold of those fed with untreated molasses, respectively. Molasses-induced astaxanthin accumulation may be attributed to the elicited expression of carotenogenic genes, in particular the genes specifically responsible for the ketolation and hydroxylation of β-carotene to form astaxanthin. A two-stage fed-batch strategy was employed to grow E17 and induce astaxathin accumulation, resulting in 45.6 g L^?1 biomass and 56.1 mg L^?1 astaxanthin, the highest volumetric astaxanthin yield ever reported for this alga. In addition, the astaxanthin production by E17 was tested with a semi-continuous culture method, where the directly diluted raw molasses (giving 5 g L^?1 sugar) was used as the carbon source. Little growth inhibition of E17 was observed in the semi-continuous culture with a biomass productivity of 1.33 g L^?1 day^?1 and an astaxanthin productivity of 0.83 mg L^?1 day^?1. The mixotrophic semi-continuous cultures enhanced the biomass and astaxanthin productivities by 29.3 % and 42.2 %, respectively. This study highlights the potential of using the industrially cheap cane molasses towards large-scale cost-saving production of the high-value ketocarotenoid astaxanthin.     

Endo-β-1,4-d-mannanase is efficiently produced by Sclerotium (Athelia) rolfsii under derepressed conditions

A number of wild-type isolates of Sclerotium (Athelia) rolfsii and S. coffeicola were studied for their ability to produce endo-β-1,4-mannanase, endo-β-1,4-xylanase, and endo-β-1,4-glucanase activity when grown on cellulose- or glucose-based media. Whereas the presence of the inducer cellulose was strictly necessary for increased xylanase and endoglucanase production by both S. rolfsii (208 and 599 U ml⁻¹, respectively) and S. coffeicola (102 and 330 U ml⁻¹, respectively), elevated activities of mannanase (up to 96.6 U ml⁻¹) were formed even when employing glucose as the only carbohydrate substrate. Significant production of mannanases as well as of auxiliary mannan-degrading enzymes (β-mannosidase, β-glucosidase, α-galactosidase, acetyl esterase) was only observed, however, under derepressed conditions, i.e. after glucose had been consumed from the medium. By applying a fed-batch strategy, in which a glucose solution was continuously fed to a cultivation of S. rolfsii CBS 191.62 so that the glucose concentration in the medium never exceeded a certain low, critical value, production of mannanase could be almost doubled as compared to a batch cultivation on glucose (462 versus 240 U ml⁻¹). Mannanase preparations produced by several S. rolfsii and S. coffeicola strains under inductive and noninductive conditions (i.e. using cellulose or glucose as the substrates, respectively) were further analyzed with respect to the patterns of isoformic mannanases formed under these different growth conditions. Multiple mannanases were secreted by all isolates investigated. Certain mannanase isoenzymes were only formed by S. rolfsii in the presence of the inducer cellulose, indicating a complex and separated regulation of the synthesis of mannanase isoenzymes in this strain.     

Functional expression of trypsin from Streptomyces griseus by Pichia pastoris

In the present study, the genes encoding trypsinogen and active trypsin from Streptomyces griseus were both cloned and expressed in the methylotrophic yeast Pichia pastoris with the α-factor secretion signal under the control of the alcohol oxidase promoter. The mature trypsin was successfully accumulated extracellularly in soluble form with a maximum amidase activity of 6.6?U?ml^?1 (batch cultivation with flask cultivation) or 14.4?U?ml^?1 (fed-batch cultivation with a 3-l fermentor). In contrast, the recombinant trypsinogen formed inclusion bodies and no activity was detected. Replacement of the trypsin propeptide Ala-Pro-Asn-Pro confirmed that its physiological function was as a repressor of activity. More importantly, our results proved that the propeptide inhibited the activity of trypsinogen after its successful folding.     

Kinetic characterization of a crude β-d-glucosidase from Aspergillus wentii Pt 2804

The kinetic characteristics of β-d-glucosidase (cellobiase, β-d-glucosidase glucohydrolase, EC 3.2.1.21) from the filtered broth of a well grown culture of Aspergillus wentii have been studied. Both cellobiose and 4-nitrophenyl-β-d-glucoside (4NPG) were used as substrates and values of K_m, V_max for both the substrates were determined. Activity was maximum over a pH range of 4.5–5.5 but declined sharply beyond 5.5 for both substrates. The optimum temperature was between 60 and 65°C. Half-life of the cellobiase was ～38.0 h at 60°C and ～6.3 h at 65°C. However, the enzyme was found to be quite stable at 50°C. The activation and deactivation energies for 4NPG hydrolysis were 33.2 and 111.3 kJ mol^?1 K^?1, and 43.6 and 63.7 kJ mol K^?1 for cellobiose hydrolysis. Product inhibition was found to be of the competitive type. Preliminary experiments showed that marked synergistic activity exists between Trichoderma reesei and A. wentii cellulases [see 1,4-(1,3;1,4)-β-d-glucan 4-glucanohydrolase, EC 3.2.1.4] for cellulose hydrolysis.     

Acetone–butanol–ethanol production with high productivity using Clostridium acetobutylicum BKM19

Conventional acetone–butanol–ethanol (ABE) fermentation is severely limited by low solvent titer and productivities. Thus, this study aims at developing an improved Clostridium acetobutylicum strain possessing enhanced ABE production capability followed by process optimization for high ABE productivity. Random mutagenesis of C. acetobutylicum PJC4BK was performed by screening cells on fluoroacetate plates to isolate a mutant strain, BKM19, which exhibited the total solvent production capability 30.5% higher than the parent strain. The BKM19 produced 32.5 g L^?1 of ABE (17.6 g L^?1 butanol, 10.5 g L^?1 ethanol, and 4.4 g L^?1 acetone) from 85.2 g L^?1 glucose in batch fermentation. A high cell density continuous ABE fermentation of the BKM19 in membrane cell‐recycle bioreactor was studied and optimized for improved solvent volumetric productivity. Different dilution rates were examined to find the optimal condition giving highest butanol and ABE productivities. The maximum butanol and ABE productivities of 9.6 and 20.0 g L^?1 h^?1, respectively, could be achieved at the dilution rate of 0.85 h^?1. Further cell recycling experiments were carried out with controlled cell‐bleeding at two different bleeding rates. The maximum solvent productivities were obtained when the fermenter was operated at a dilution rate of 0.86 h^?1 with the bleeding rate of 0.04 h^?1. Under the optimal operational condition, butanol and ABE could be produced with the volumetric productivities of 10.7 and 21.1 g L^?1 h^?1, and the yields of 0.17 and 0.34 g g^?1, respectively. The obtained butanol and ABE volumetric productivities are the highest reported productivities obtained from all known‐processes. Biotechnol. Bioeng. 2013; 110: 1646–1653. © 2013 Wiley Periodicals, Inc.     

Comparison of growth of Tetraselmis in a tubular photobioreactor (Biocoil) and a raceway pond

Microalgae cultivation systems can be divided broadly into open ponds and closed photobioreactors. This study investigated the growth and biomass productivity of the halophilic green alga Tetraselmis sp. MUR-233, grown outdoors in paddle wheel-driven open raceway ponds and in a tubular closed photobioreactor (Biocoil) at a salinity of 7 % NaCl (w/v) between mid-March and June 2010 (austral autumn/winter). Volumetric productivity in the Biocoil averaged 67 mg ash-free dry weight (AFDW) L^?1 day^?1 when the culture was grown without CO₂ addition. This productivity was 86 % greater, although less stable, than that achieved in the open raceway pond (36 mg L^?1 day^?1) grown at the same time in the autumn period. The Tetraselmis culture in the open raceway pond could be maintained in semi-continuous culture for the whole experimental period of 3 months without an additional CO₂ supply, whereas in the Biocoil, under the same conditions, reliable semi-continuous culture was only achievable for a period of 38 days. However, stable semi-continuous culture was achieved in the Biocoil by the addition of CO₂ at a controlled pH of ~7.5. With CO₂ addition, the volumetric biomass productivity in the Biocoil was 85 mg AFDW L^?1 day^?1 which was 5.5 times higher than the productivity achieved in the open raceway pond (15 mg AFDW L^?1 day^?1) with CO₂ addition and 8 times higher compared to the productivity in the open raceway pond without CO₂ addition (11 mg AFDW L^?1 day^?1), when cultures were grown in winter. The illuminated area productivities highlight an alternative story and showed that the open raceway pond had a three times higher productivity (3,000 mg AFDW m^?2 day^?1) compared to the Biocoil (850 mg AFDW m^?2 day^?1). Although significant differences were found between treatments and cultivation systems, the overall average lipid content for Tetraselmis sp. MUR-233 was 50 % in exponential phase during semi-continuous cultivation.     

Immobilization of glucoamylase on DEAE-cellulose activated with chloride compounds

Partially purified glucoamylase (1,4-α-d-glucan glucohydrolase, EC 3.2.1.3) from Aspergillus niger NRRL 330 has been immobilized on DEAE-cellulose activated with cyanuric chloride in 0.2 m acetate buffer, pH 4.2. In the matrix-bound glucoamylase, enzyme yield was 20 mg g^?1 of support, corresponding to 40 200 units g^?1 of DEAE support. Binding of the enzyme narrows the pH optimum from 3.8–5.2 to 3.6. Thermal stability of the bound glucoamylase enzyme was decreased although it showed a higher temperature optimum (70°C) than the free form (55°C). The rate of reaction of glucoamylase was also changed after immobilization. V_max values for free and bound enzyme were 36.6 and 22.6 μmol d-glucose ml^?1 min^?1 and corresponding K_m values were 3.73 and 4.8 g l^?1 respectively. Free and immobilized enzyme when used in the saccharification process gave 84 and 56% conversion of starch to d-glucose, respectively. The bound enzyme was quite stable and in the batch process it was able to operate for about five cycles without any loss of activity.