The Mechanism of CeCl<Subscript>3</Subscript> on the Activiation of Alanine Aminotransferase from Mice

The activity of alanine aminotransferase (ALT; E.C. 2.6.1.2) is often changed upon inflammatory responses in animals. Rare earths was shown to provoke various inflammatory responses both in rats and mice; however, the molecular mechanism by which rare earths exert its toxicity has not been completely understood, especially, we know little about the mechanism of the interaction between CeCl₃ and ALT. In this report, we investigated the mechanisms of CeCl₃ on ALT activity in vivo and in vitro. Our results showed that Ce³⁺ could significantly activate ALT in vivo and in vitro; the kinetics constant (Km) and Vmax were 0.018 μM and 1,380 unit mg⁻¹ protein min⁻¹, respectively, at a low concentration of Ce³⁺, and 0.027 μM and 624 unit mg⁻¹ protein min⁻¹, respectively, at a high concentration of Ce³⁺. By UV absorption and fluorescence spectroscopy assays, the Ce³⁺ was determined to be directly bound to ALT; the binding site of Ce³⁺ to ALT was 1.72, and the binding constants of the binding site were 4.82 × 10⁸ and 9.05 × 10⁷ L mol⁻¹. Based on the analysis of the circular dichroism spectra, it was concluded that the binding of Ce³⁺ altered the secondary structure of ALT, suggesting that the observed enhancement of ALT activity was caused by a subtle structural change in the active site through the formation of the complex with Ce³⁺.     

Effect of Mg2+ on the Structure and Function of Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase

Mg²⁺ in various concentrations was added to purified Rubisco in vitro to gain insight into the mechanism of molecular interactions between Mg²⁺ and Rubisco. The enzyme activity assays showed that the reaction between Rubisco and Mg²⁺ was two order, which means that the enhancement of Rubisco activity was accelerated by low concentration of Mg²⁺ and slowed by high concentration of Mg²⁺. The kinetics constant (K _m) and V _max was 1.91 μM and 1.13 μmol CO₂ mg⁻¹ protein∙min⁻¹, respectively, at a low concentration of Mg²⁺, and 3.45 μM and 0.32 μmol CO₂∙mg⁻¹ protein∙min⁻¹, respectively, at a high concentration of Mg²⁺. By UV absorption and fluorescence spectroscopy assays, the Mg²⁺ was determined to be directly bound to Rubisco; the binding site of Mg²⁺ to Rubisco was 0.275, the binding constants (K _A) of the binding site were 6.33 × 10⁴ and 5.5 × 10⁴ l·mol⁻¹. Based on the analysis of the circular dichroism (CD) spectra, it was concluded that the binding of Mg²⁺ did not alter the secondary structure of Rubisco, suggesting that the observed enhancement of Rubisco carboxylase activity was caused by a subtle structural change in the active site through the formation of the complex with Mg²⁺.     

Ab initio studies on the decomposition kinetics of CF<Subscript>3</Subscript>OCF<Subscript>2</Subscript>O radical

The present study deals with the decomposition of CF₃OCF₂O radical formed from a hydrofluoroether, CF₃OCHF₂ (HFE-125), in the atmosphere. The study is performed using ab initio quantum mechanical methods. Two plausible pathways of decomposition of the titled species have been considered, one involving C-O bond scission and the other occurring via F atom elimination. The geometries of the reactant, products and transition states involved in the decomposition pathways are optimized and characterized at DFT (B3LYP) level of theory using 6-311G(d,p) basis set. Single point energy calculations have been performed at G2M(CC,MP2) level of theory. Out of the two prominent decomposition channels considered, the C-O bond scission is found to be dominant involving a barrier height of 15.3 kcal mol⁻¹ whereas the F-elimination path proceeds with a barrier of 26.1 kcal mol⁻¹. The thermal rate constants for the above two decomposition pathways are evaluated using canonical transition state theory (CTST) and these are found to be 1.78 × 10⁶ s⁻¹ and 2.83 × 10⁻⁷ s⁻¹ for C-O bond scission and F-elimination respectively at 298 K and 1 atm pressure. Transition states are searched on the potential energy surfaces involved during the decomposition channels and each of the transition states is characterized. The existence of transition states on the corresponding potential energy surface is ascertained by performing intrinsic reaction coordinate (IRC) calculation.     

Sucrose monolaurate synthesis with Protex 6L immobilized on electrospun TiO<Subscript>2</Subscript> nanofiber

TiO₂ nanofibers with uniform diameter about 125 nm were prepared based on sol–gel process and electrospinning technology. Protex 6L, an industrial alkaline protease, was covalently immobilized on TiO₂ nanofiber through γ-aminopropyltriethoxysilane modification and glutaraldehyde crosslinking. With 2 (v/v)% glutaraldehyde as crosslinker, the enzyme loading is about 201 mg (g nanofiber membrane)⁻¹, and the specific activity of the immobilized Protex 6L is 2.45 μmol h⁻¹ ml⁻¹ mg⁻¹ protein for synthesis of sucrose monolaurate from sucrose and vinyl laurate. The optimal condition for sucrose monolaurate production is 5% (v/v) water content in DMSO/2-methyl-2-butanol solvent mixture and 50°C. Under this condition, 97% conversion was achieved within 36 h by nanofibrous Protex 6L, which is corresponding to a productivity 34 times higher than that of most widely used Novozym 435. After 10 cycles reuse, nanofibrous Protex 6L retained 52.4% of its original activity.     

The Interactions of Glutathione-Capped CdTe Quantum Dots with Trypsin

Due to their unique fluorescent properties, quantum dots present a great potential for biolabelling applications; however, the toxic interactions of quantum dots with biopolymers are little known. The toxic interactions of glutathione-capped CdTe quantum dots with trypsin were studied in this paper using synchronous fluorescence spectroscopy, fluorescence emission spectra, and UV–vis absorption spectra. The interaction between CdTe quantum dots and trypsin resulted in structure changes of trypsin and inhibited trypsin's activity. Fluorescence emission spectra revealed that the quenching mechanism of trypsin by CdTe quantum dots was a static quenching process. The binding constant and the number of binding sites at 288 and 298 K were calculated to be 1.98 × 10⁶ L mol⁻¹ and 1.37, and 6.43 × 10⁴ L mol⁻¹ and 1.09, respectively. Hydrogen bonds and van der Waals' forces played major roles in this process.     

Determination of shelf life of <Emphasis Type="Italic">Spirulina platensis</Emphasis> (MI<Subscript>2</Subscript>) grown in the Philippines

Gamma linolenic acid (GLA) degradation in Spirulina followed first-order reaction kinetics. At an accelerated temperature range of 45 to 55°C, the degradation rate constants (k _r) of GLA obtained were 4.0 × 10⁻² to 8.8 × 10⁻² day⁻¹. The energy of activation (E _a) was 16.53 kcal mol⁻¹, and the Q₁₀ was 2.22. Based on 20% GLA degradation, the shelf life of sun-dried Spirulina at 30°C is 263 days or 8.6 months using the Arrhenius plot, and 258 days or 8.5 months using the Q ₁₀ approach. Presented at the 6th Meeting of the Asia Pacific Society of Applied Phycology, Manila, Philippines.     

In vitro synthesis of poly(3-hydroxydecanoate): purification and enzymatic characterization of type II polyhydroxyalkanoate synthases PhaC1 and PhaC2 from Pseudomonas aeruginosa

For the first time, the purification has been achieved of the type II polyhydroxyalkanoate (PHA) synthases PhaC1 and PhaC2 from Pseudomonas aeruginosa applying N-terminal His₆-tag fusions and metal chelate affinity chromatography. In vivo His₆-tagged PHA synthase activity was confirmed by functional expression of the corresponding genes in Escherichia coli, and PHA synthase activity could also be measured in vitro with the enzymes. The specific enzyme activity of PHA synthases PhaC1 and PhaC2 was 0.039 U mg⁻¹ and 0.035 U mg⁻¹ protein, respectively. Kinetic studies showed a lag phase for both PHA synthases using (R,S)-3-hydroxydecanoyl-CoA as substrate. Specific enzyme activity was increased to 0.055 U mg⁻¹ when the phasin GA24 from Ralstonia eutropha was added to the assay. CoA inhibited PHA synthase activity, and a K _i of 85 μM was determined. A two-enzyme system was established, employing commercially available acyl-CoA synthetase and PHA synthase, which allowed the in vitro de novo PHA granule formation and the in vitro synthesis of poly(3-hydroxydecanoate) exhibiting a weight average molar mass of 9.8 × 10⁴ g mol⁻¹, and which occurred independently of pre-existing PHA granules. Received: 3 December 1999 / Revision received: 10 January 2000 / Accepted: 14 January 2000     

Gas exchange and photosynthetic performance of the tropical tree <Emphasis Type="Italic">Acacia nigrescens</Emphasis> when grown in different CO<Subscript>2</Subscript> concentrations

The photosynthetic responses of the tropical tree species Acacia nigrescens Oliv. grown at different atmospheric CO₂ concentrations—from sub-ambient to super-ambient—have been studied. Light-saturated rates of net photosynthesis (A _sat) in A. nigrescens, measured after 120 days exposure, increased significantly from sub-ambient (196 μL L⁻¹) to current ambient (386 μL L⁻¹) CO₂ growth conditions but did not increase any further as [CO₂] became super-ambient (597 μL L⁻¹). Examination of photosynthetic CO₂ response curves, leaf nitrogen content, and leaf thickness showed that this acclimation was most likely caused by reduction in Rubisco activity and a shift towards ribulose-1,5-bisphosphate regeneration-limited photosynthesis, but not a consequence of changes in mesophyll conductance. Also, measurements of the maximum efficiency of PSII and the carotenoid to chlorophyll ratio of leaves indicated that it was unlikely that the pattern of A _sat seen was a consequence of growth [CO₂] induced stress. Many of the photosynthetic responses examined were not linear with respect to the concentration of CO₂ but could be explained by current models of photosynthesis.     

Na<Superscript>+</Superscript>/H<Superscript>+</Superscript> exchange activity in the alkaliphile halotolerant cyanobacterium <Emphasis Type="Italic">Aphanothece halophytica</Emphasis>

The activity of Na⁺/H⁺ exchanger to remove toxic Na⁺ is important for growth of organisms under high salinity. In this study, the halotolerant cyanobacterium Aphanothece halophytica was shown to possess Na⁺/H⁺ exchange activity since exogenously added Na⁺ could dissipate a pre-formed pH gradient, and decrease extracellular pH. Kinetic analysis yielded apparent K _m (Na⁺) and V _max of 20.7 ± 3.1 mM and 3,333 ± 370 nmol H⁺ min⁻¹ mg⁻¹, respectively. For cells grown under salt-stress condition, the apparent K _m (Na⁺) and V _max was 18.3 ± 3.5 mM and 3,703 ± 350 nmol H⁺ min⁻¹ mg⁻¹, respectively. Three cations with decreasing efficiency namely Li⁺, Ca²⁺, and K⁺ were also able to dissipate pH gradient. Only marginal exchange activity was observed for Mg²⁺. The exchange activity was strongly inhibited by Na⁺-gradient dissipators, monensin, and sodium ionophore as well as by CCCP, a protonophore. A. halophytica showed high Na⁺/H⁺ exchange activity at neutral and alkaline pH up to pH 10. Cells grown at pH 7.6 under high salinity exhibited higher Na⁺/H⁺ exchange activity than those grown under low salinity during 15 days of growth suggesting a role of Na⁺/H⁺ exchanger for salt tolerance in A. halophytica. Cells grown at alkaline pH of 9.0 also exhibited a progressive increase of Na⁺/H⁺ exchange activity during 15 days of growth.     

Effects of elevated CO<Subscript>2</Subscript> and/or O<Subscript>3</Subscript> on hormone IAA in needles of Chinese pine

This study examined the effects of season-long exposure of Chinese pine (Pinus tabulaeformis) to elevated carbon dioxide (CO₂) and/or ozone (O₃) on indole-3-acetic acid (IAA) content, activities of IAA oxidase (IAAO) and peroxidase (POD) in needles. Trees grown in open-top chambers (OTC) were exposed to control (ambient O₃, 55 nmol mol⁻¹ + ambient CO₂, 350 μmol mol⁻¹, CK), elevated CO₂ (ambient O₃ + high CO₂, 700 μmol mol⁻¹, EC) and elevated O₃ (high O₃, 80 ± 8 nmol mol⁻¹ + ambient CO₂, EO) OTCs from 1 June to 30 September. Plants grown in elevated CO₂ OTC had a growth increase of axial shoot and needle length, compared to control, by 20% and 10% respectively, while the growth in elevated O₃ OTC was 43% and 7% less respectively, than control. An increase in IAA content and POD activity and decrease in IAAO activity were observed in trees exposed to elevated CO₂ concentration compared with control. Elevated O₃ decreased IAA content and had no significant effect on IAAO activity, but significantly increased POD activity. When trees pre-exposed to elevated CO₂ were transferred to elevated O₃ (EC–EO) or trees pre-exposed to elevated O₃ were transferred to elevated CO₂ (EO–EC), IAA content was lower while IAAO activity was higher than that transferred to CK (EC–CK or EO–CK), the change in IAA content was also related to IAAO activity. The results indicated that IAAO and POD activities in Chinese pine needles may be affected by the changes in the atmospheric environment, resulting in the change of IAA metabolism which in turn may cause changes in Chinese pine’s growth. An erratum to this article can be found at     

Kinetics of CO conversion into H<Subscript>2</Subscript> by <Emphasis Type="Italic">Carboxydothermus hydrogenoformans</Emphasis>

The objective of this study was to improve the biological water–gas shift reaction for producing hydrogen (H₂) by conversion of carbon monoxide (CO) using an anaerobic thermophilic pure strain, Carboxydothermus hydrogenoformans. Specific hydrogen production rates and yields were investigated at initial biomass densities varying from 5 to 20 mg volatile suspended solid (VSS) L⁻¹. Results showed that the gas–liquid mass transfer limits the CO conversion rate at high biomass concentrations. At 100-rpm agitation and at CO partial pressure of 1 atm, the optimal substrate/biomass ratio must exceed 5 mol CO g⁻¹ biomass VSS in order to avoid gas–liquid substrate transfer limitation. An average H₂ yield of 94 ± 3% and a specific hydrogen production rate of ca. 3 mol g⁻¹ VSS day⁻¹ were obtained at initial biomass densities between 5 and 8 mg VSS⁻¹. In addition, CO bioconversion kinetics was assessed at CO partial pressure from 0.16 to 2 atm, corresponding to a dissolved CO concentration at 70°C from 0.09 to 1.1 mM. Specific bioactivity was maximal at 3.5 mol CO g⁻¹ VSS day⁻¹ for a dissolved CO concentration of 0.55 mM in the culture. This optimal concentration is higher than with most other hydrogenogenic carboxydotrophic species.     

Production of β-galactosidase from thermophilic fungus Rhizomucor sp

A thermostable β-galactosidase was produced extracellularly by a thermophilic Rhizomucor sp, with maximum enzyme activity (0.21 U mg⁻¹) after 4 days under submerged fermentation condition (SmF). Solid state fermentation (SSF) resulted in a nine-fold increase in enzyme activity (2.04 U mg⁻¹). The temperature range for production of the enzyme was 38–55°C with maximum activity at 45°C. The optimum pH and temperature for the partially purified enzyme was 4.5 and 60°C, respectively. The enzyme retained its original activity on incubation at 60°C up to 1 h. Divalent cations like Co²⁺, Mn²⁺, Fe²⁺ and Zn²⁺ had strong inhibitory effects on the enzyme activity. The K _m and V _max for p-nitrophenyl-β- _D-galactopyranoside and o-nitrophenyl-β - _D-galactopyranoside were 0.39 mM, 0.785 mM and 232.1 mmol min⁻¹ mg⁻¹ respectively. The K _m and V _max for the natural substrate lactose were 66.66 μM and 0.20 μ mol min⁻¹ mg⁻¹. Received 10 March 1997/ Accepted in revised form 17 July 1997     

A new manganese superoxide dismutase identified from Beauveria bassiana enhances virulence and stress tolerance when overexpressed in the fungal pathogen

A superoxide dismutase (SOD) was characterized from Beauveria bassiana, a fungal entomopathogen widely applied to insect control. This 209-aa enzyme (BbSod2) showed no more than 71% sequence identity to other fungal Mn-SODs, sharing all conserved residues with the Mn-SOD family and lacking a mitochondrial signal. The SOD activity of purified BbSod2 was significantly elevated by Mn²⁺, suppressed by Cu²⁺ and Zn²⁺ but inhibited by Fe³⁺. Overexpressing the enzyme in a BbSod2-absent B. bassiana strain enhanced its SOD activity (107.2 ± 6.1 U mg⁻¹ protein) by 4–10-fold in different transformants analyzed. The best BbSod2-transformed strain with the SOD activity of 1,157.9 ± 74.7 U mg⁻¹ was 93% and 61% more tolerant to superoxide-generating menadione in both colony growth (EC₅₀ = 2.41 ± 0.03 versus 1.25 ± 0.01 mM) and conidial germination (EC₅₀ = 0.89 ± 0.06 versus 0.55 ± 0.07 mM), and 23% more tolerant to UV-B irradiation (LD₅₀ = 0.49 ± 0.02 versus 0.39 ± 0.01 J cm⁻²). Its virulence to Spodoptera litura larvae was enhanced by 26% [LT₅₀ = 4.5 (4.2–4.8) versus 5.7 (5.2–6.4) days]. Our study highlights for the first time that the Mn²⁺-cofactored, cytosolic BbSod2 contributes significantly to the virulence and stress tolerance of B. bassiana and reveals possible means to improving field persistence and efficacy of a fungal formulation by manipulating the antioxidant enzymes of a candidate strain.     

Characterization of a recombinant endo-type alginate lyase (Alg7D) from <Emphasis Type="Italic">Saccharophagus degradans</Emphasis>

A gene, alg7D, from Saccharophagus degradans, coding for a putative alginate lyase belonging to the family of polysaccharide lyase-7, was overexpressed in Escherichia coli. The properties of the recombinant Alg7D were characterized. The enzyme endolytically depolymerized alginate by β-elimination into oligo-alginates with degrees of polymerization of 2–5. Its activity was maximal at 50°C and pH 7 and was slightly increased in the presence of Na⁺. The K _M , V _max , k _cat , and k _cat /K _M values were: 3 mg ml⁻¹, 6.2 U mg⁻¹, 1.9 × 10⁻² s⁻¹, and 6.3 × 10⁻³ mg⁻¹ ml s⁻¹, respectively.     

Recent improvements in the development of A<Subscript>2B</Subscript> adenosine receptor agonists

Adenosine is known to exert most of its physiological functions by acting as local modulator at four receptor subtypes named A₁, A_2A, A_2B and A₃ (ARs). Principally as a result of the difficulty in identifying potent and selective agonists, the A_2B AR is the least extensively characterised of the adenosine receptors family. Despite these limitations, growing understanding of the physiological meaning of this target indicates promising therapeutic perspectives for specific ligands. As A_2B AR signalling seems to be associated with pre/postconditioning cardioprotective and anti-inflammatory mechanisms, selective agonists may represent a new therapeutic group for patients suffering from coronary artery disease. Herein we present an overview of the recent advancements in identifying potent and selective A_2B AR agonists reported in scientific and patent literature. These compounds can be classified into adenosine-like and nonadenosine ligands. Nucleoside-based agonists are the result of modifying adenosine by substitution at the N ⁶-, C²-positions of the purine heterocycle and/or at the 5′-position of the ribose moiety or combinations of these substitutions. Compounds 1-deoxy-1-{6-[N′-(furan-2-carbonyl)-hydrazino]-9H-purin-9-yl}-N-ethyl-β-D-ribofuranuronamide (19, hA₁ K _i = 1050 nM, hA_2A K _i = 1550 nM, hA_2B EC₅₀ = 82 nM, hA₃ K _i > 5 μM) and its 2-chloro analogue 23 (hA₁ K _i = 3500 nM, hA_2A K _i = 4950 nM, hA_2B EC₅₀ = 210 nM, hA₃ K _i > 5 μM) were confirmed to be potent and selective full agonists in a cyclic adenosine monophosphate (cAMP) functional assay in Chinese hamster ovary (CHO) cells expressing hA_2B AR. Nonribose ligands are represented by conveniently substituted dicarbonitrilepyridines, among which 2-[6-amino-3,5-dicyano-4-[4-(cyclopropylmethoxy)phenyl]pyridin-2-ylsulfanyl]acetamide (BAY-60–6583, hA₁, hA_2A, hA₃ EC₅₀ > 10 μM; hA_2B EC₅₀ = 3 nM) is currently under preclinical-phase investigation for treating coronary artery disorders and atherosclerosis.     

Down-regulation of Rubisco activity under combined increases of CO<Subscript>2</Subscript> and temperature minimized by changes in Rubisco k<Subscript>cat</Subscript> in wheat

Increases in growth temperature have been observed to affect photosynthesis differently under long-term exposure to ambient- and twice ambient-air CO₂ concentrations. This study investigates the causes of this interaction in wheat (Triticum aestivum L.) grown in the field over two consecutive years under temperature gradient chambers in ambient (370 μmol mol⁻¹) or elevated (700 μmol mol⁻¹) atmospheric CO₂ concentrations and at ambient or ambient +4°C temperatures, with either a low or a high nitrogen supply. The photosynthesis-internal CO₂ response curves and the activity, activation state, k_cat and amount of Ribulose-1, 5-bisphosphate carboxylase/oxygenase (Rubisco) were measured, as well as the soluble protein concentration in flag leaves at ear emergence and 8–15 days after anthesis. A high nitrogen supply increased V_cmax, the Rubisco amount and activity and soluble protein contents, but did not significantly change the Rubisco k_cat. Both elevated CO₂ and above ambient temperatures had negative effects on V_cmax and Rubisco activity, but at elevated CO₂, an increase in temperature did not decrease V_cmax or Rubisco activity in relation to ambient temperature. The amounts of Rubisco and soluble protein decreased with elevated CO₂ and temperature. The negative impact of elevated CO₂ on Rubisco properties was somewhat counteracted at elevated temperatures by an increase in k_cat. This effect can diminish the detrimental effects on photosynthesis of combined increases of CO₂ and temperature.     

High constitutive peroxidase activity and constitutive thermotolerance in <Emphasis Type="Italic">Neurospora crassa</Emphasis>

 During the isolation of mutations in the heat-inducible hsp70-1 gene of Neurospora crassa by RIP (repeat-induced point mutations), several transformants were generated by electroporation of conidia with a plasmid harboring an incomplete copy of this gene. One isolate, designated E-45, containing ectopically integrated hsp70-1 DNA, exhibited a slow growth rate, low-temperature sensitivity, constitutive thermotolerance (without prior heat shock), and high constitutive peroxidase activity. The constitutive form of peroxidase (CP) was distinguishable from the heat-inducible form (HIP) by immunoinactivation employing polyclonal antiserum against the latter enzyme and by electrophoretic resolution in nondenaturing polyacrylamide gels. This enzyme was purified to near homogeneity and some of its properties examined. The relative molecular mass of native CP was in the range of 118–136 kDa, as estimated by gel filtration analysis on size exclusion matrices, whereas SDS-PAGE analysis yielded a size of ∼37 kDa for the polypeptide. Substrate saturation kinetics studies were conducted using ABTS [2,2′-azino-bis (3-ethylbenzthiazole-6-sulfonic acid)] and H₂O₂ as substrates: K _m, V _max, and K _cat values for H₂O₂ were ∼22 μM, ∼447 nmol mg⁻¹, and 0.33 s⁻¹, respectively, and those for ABTS were ∼55 μM, ∼453 nmol mg⁻¹, and 0.3 s⁻¹, respectively. Guaiacol was not used as a substrate by this enzyme. CP peroxidase was shown to be a heme-containing enzyme, stable at temperatures up to 58°C. Received: August 5, 2002 / Accepted: January 22, 2003 Acknowledgments This work was supported by an operating grant from the Natural Sciences and Engineering Research Council (NSERC) of Canada (to M.K.). The financial support provided to A. M. in the form of a graduate studentship award by the AHFMR (Alberta Heritage Foundation for Medical Research) and of a graduate teaching assistantship to A. S. by the Department of Biological Sciences, University of Calgary, is gratefully acknowledged. Correspondence to:M. Kapoor     

Phenol biodegradation by the thermoacidophilic archaeon <Emphasis Type="Italic">Sulfolobus solfataricus</Emphasis> 98/2 in a fed-batch bioreactor

Toxic at low concentrations, phenol is one of the most common organic pollutants in air and water. In this work, phenol biodegradation was studied in extreme conditions (80°C, pH = 3.2) in a 2.7 l bioreactor with the thermoacidophilic archaeon Sulfolobus solfataricus 98/2. The strain was first acclimatized to phenol on a mixture of glucose (2000 mg l⁻¹) and phenol (94 mg l⁻¹) at a constant dissolved oxygen concentration of 1.5 mg l⁻¹. After a short lag-phase, only glucose was consumed. Phenol degradation then began while glucose was still present in the reactor. When glucose was exhausted, phenol was used for respiration and then for biomass build-up. After several batch runs (phenol < 365 mg l⁻¹), specific growth rate (μ_X) was 0.034 ± 0.001 h⁻¹, specific phenol degradation rate (q_P) was 57.5 ± 2 mg g⁻¹ h⁻¹, biomass yield (Y_X/P) was 52.2 ± 1.1 g mol⁻¹, and oxygen yield factor ( \textY_{\textX/\textO 2} ) \left( {{\text{Y}}_{{{\text{X}}/{\text{O}}_{ 2} }} } \right) was 9.2 ± 0.2 g mol⁻¹. A carbon recovery close to 100% suggested that phenol was exclusively transformed into biomass (35%) and CO₂ (65%). Molar phenol oxidation constant ( \textY_{\textO 2 /\textP} ) \left( {{\text{Y}}_{{{\text{O}}_{ 2} /{\text{P}}}} } \right) was calculated from stoichiometry of phenol oxidation and introducing experimental biomass and CO₂ conversion yields on phenol, leading to values varying between 4.78 and 5.22 mol mol⁻¹. Respiratory quotient was about 0.84 mol mol⁻¹, very close to theoretical value (0.87 mol mol⁻¹). Carbon dioxide production, oxygen demand and redox potential, monitored on-line, were good indicators of growth, substrate consumption and exhaustion, and can therefore be usefully employed for industrial phenol bioremediation in extreme environments.     

Continuous butanol production with reduced byproducts formation from glycerol by a hyper producing mutant of <Emphasis Type="Italic">Clostridium pasteurianum</Emphasis>

Butanol, a four-carbon primary alcohol (C₄H₁₀O), is an important industrial chemical and has a good potential to be used as a superior biofuel. Bio-based production of butanol from renewable feedstock is a promising and sustainable alternative to substitute petroleum-based fuels. Here, we report the development of a process for butanol production from glycerol, which is abundantly available as a byproduct of biodiesel production. First, a hyper butanol producing strain of Clostridium pasteurianum was isolated by chemical mutagenesis. The best mutant strain, C. pasteurianum MBEL_GLY2, was able to produce 10.8 g l⁻¹ butanol from 80 g l⁻¹ glycerol as compared to 7.6 g l⁻¹ butanol produced by the parent strain. Next, the process parameters were optimized to maximize butanol production from glycerol. Under the optimized batch condition, the butanol concentration, yield, and productivity of 17.8 g l⁻¹, 0.30 g g⁻¹, and 0.43 g l⁻¹ h⁻¹ could be achieved. Finally, continuous fermentation of C. pasteurianum MBEL_GLY2 with cell recycling was carried out using glycerol as a major carbon source at several different dilution rates. The continuous fermentation was run for 710 h without strain degeneration. The acetone–butanol–ethanol productivity and the butanol productivity of 8.3 and 7.8 g l⁻¹ h⁻¹, respectively, could be achieved at the dilution rate of 0.9 h⁻¹. This study reports continuous production of butanol with reduced byproducts formation from glycerol using C. pasteurianum, and thus could help design a bioprocess for the improved production of butanol.     

Utilization of <Emphasis Type="Italic">Anabaena</Emphasis> sp. in CO<Subscript>2</Subscript> removal processes

This paper focuses on modelling the growth rate and exopolysaccharides production of Anabaena sp. ATCC 33047, to be used in carbon dioxide removal and biofuels production. For this, the influence of dilution rate, irradiance and aeration rate on the biomass and exopolysaccharides productivity, as well as on the CO₂ fixation rate, have been studied. The productivity of the cultures was maximum at the highest irradiance and dilution rate assayed, resulting to 0.5 g_bio l⁻¹ day⁻¹ and 0.2 g_eps l⁻¹ day⁻¹, and the CO₂ fixation rate measured was 1.0 gCO₂ l⁻¹ day⁻¹. The results showed that although Anabaena sp. was partially photo-inhibited at irradiances higher than 1,300 μE m⁻² s⁻¹, its growth rate increases hyperbolically with the average irradiance inside the culture, and so does the specific exopolysaccharides production rate. The latter, on the other hand, decreases under high external irradiances, indicating that the exopolysaccharides metabolism hindered by photo-damage. Mathematical models that consider these phenomena have been proposed. Regarding aeration, the yield of the cultures decreased at rates over 0.5 v/v/min or when shear rates were higher than 60 s⁻¹, demonstrating the existence of thus existence of stress damage by aeration. The behaviour of the cultures has been verified outdoors in a pilot-scale airlift tubular photobioreactor. From this study it is concluded that Anabaena sp. is highly recommended to transform CO₂ into valuable products as has been proved capable of metabolizing carbon dioxide at rates of 1.2 gCO₂ l⁻¹ day⁻¹ outdoors. The adequacy of the proposed equations is demonstrated, resulting to a useful tool in the design and operation of photobioreactors using this strain.