Influence of pH shift and salting on the energetics of microalgae <Emphasis Type="Italic">Chlorella vulgaris</Emphasis> and <Emphasis Type="Italic">Dunaliella maritima</Emphasis>

The influence of pH shift and NaCl salting on the heat production, oxygen adsorption, and oxygen evolution rates of the unicellular green halotolerant microalga Dunaliella maritima and the freshwater microalga Chlorella vulgaris were investigated. In the growth process of both microalgae the alkalization of their culture medium was observed. And simultaneously it was shown that at increasing NaCl salting of cultural medium its acidification occurs. At alkalization and acidification of Chlorella medium the increase of heat production and respiration rates were observed. At alkalization and acidification of Dunaliella culture the adverse effect of decreasing heat production rate was observed. Acidification of culture medium of both algae led to short-term and sharp increase of photosynthesis measured by polarography and photomicrocalorimeter.     

The effect of zinc(II) on the growth of <Emphasis Type="Italic">E . coli</Emphasis> studied by microcalorimetry

By using an LKB2277 Bioactivity Monitor, stop-flow mode, the power-time curves of E . coli at 37°C effected by zinc(II) were determined. Some parameters, such as growth rate constants k, inhibitory ratio I, the maximum heat production rate P _max heat output Q and the time in the maximum heat production t _max were obtained. According these parameters, we found that a low concentration of zinc(II) had a promoting action on the growth of E. coli, but a high concentration of zinc(II) had an inhibitory action. The toxicity of zinc(II) can also be expressed as half inhibitory concentration IC ₅₀ of zinc(II), i.e., 50% effective in this inhibition. The value of IC ₅₀ of zinc(II) on E. coliis 28.09 µg mL^-1. The assay is quantitative, inexpensive and versatile.     

Microcalorimetric study on the growth and metabolism of <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis>

Microcalorimetry is an experimental technique which allows us to precisely measure the energy released as a consequence of any transformation process. All organisms produce heat as a consequence of metabolism. The rate of heat production is an adequate measurement of metabolic activity of organisms and their constituent parts, cells and sub-cellular levels. Microorganisms produce small amounts of heat, in the order of 1–3 pW per cell. Despite the low quantity of heat produced by bacteria, their exponential replication in culture medium allows their detection using microcalorimetry. This study is a microcalorimetric study of the growth and metabolism of the bacterium Pseudomonas aeruginosa, using the heat liberated as a consequence of bacterial metabolism. With this aim, we used a Calvet microcalorimeter, inside which two Teflon screw-capped stainless steel cells were located (sample and reference). Experiments were carried out at final concentrations of 10⁶, 10⁵, 10³ and 10 CFU/mL, and a constant temperature of 309.65 K was maintained within the microcalorimeter. Recording the difference in calorific potential over time we obtained P. aeruginosa’s growth curves. The shape of these curves is characteristic and has a single phase. Thus, the heat flow curves were mathematically studied to calculate the growth constant and generation time of this bacterium.     

Effect of Latent Heat in Boiling Water on the Synthesis of Gold Nanoparticles of Different Sizes by using the Turkevich Method

The Turkevich method, involving the reduction of HAuCl₄ with citrate in boiling water, allows the facile production of monodisperse, quasispherical gold nanoparticles (AuNPs). Although, it is well‐known that the size of the AuNPs obtained with the same recipe vary slightly (as little as approximately 4 nm), but noticeably, from one report to another, it has rarely been studied. The present work demonstrates that this size variation can be reconciled by the small, but noticeable, effect that the latent heat in boiling water has on the size of the AuNPs obtained by using the Turkevich method. The increase in latent heat during water boiling caused an approximately 3 nm reduction in the size of the as‐prepared AuNPs; this reduction in size is mainly a result of accelerated nucleation driven by the extra heat. It was further demonstrated that, the heating temperature can be utilized as an additional measure to adjust the growth rate of AuNPs during the reduction of HAuCl₄ with citrate in boiling water. Therefore, the latent heat of boiling solvents may provide one way to control nucleation and growth in the synthesis of monodisperse nanoparticles.     

Microcalorimetric study of the growth of <Emphasis Type="Italic">Enterococcus faecalis</Emphasis> in an enriched culture medium

Enterococcus faecalis is a Gram-positive bacteria, considered one of the most common causes of nosocomial infections. Bacterial cultures produce an exchange of energy as a result of the bacteria metabolisms. The rate of heat production is an adequate measure of the metabolic activity of the organisms and their constituent parts. Microorganisms produce small amounts of heat: 1–3 pW per cell. Although the heat produced by bacteria is very small, their exponential reproduction in a culture medium permits heat detection through microcalorimetry. In this study, we analyzed the microcalorimetric behavior of Enterococcus faecalis. A thermal Calvet microcalorimeter was used. The inside of the calorimeter contains two stainless steel cells (experimental and reference). Experiments were carried out at final concentrations of 10⁶,10⁵,10³, and 10 CFU/mL and a constant temperature of 309.65 K was maintained within the microcalorimeter. Recording the difference in calorific potential over time we obtained E. faecalis’s growth curves. Thermograms were analyzed mathematically allowing us to calculate the constant growth, generation time and the amount of heat exchanged over the culture time.     

Production of Cellulolytic Enzymes by <Emphasis Type="Italic">Aspergillus phoenicis</Emphasis> in Grape Waste using Response Surface Methodology

The production of cellulolytic enzymes by the fungus Aspergillus phoenicis was investigated. Grape waste from the winemaking industry was chosen as the growth substrate among several agro-industrial byproducts. A 2 × 2 central composite design was performed, utilizing the amount of grape waste and peptone as independent variables. The fungus was cultivated in submerged fermentation at 30 °C and 120 rpm for 120 h, and the activities of total cellulases, endoglucanases, and β-glucosidases were measured. Total cellulases were positively influenced by the linear increase of peptone concentration and decrease at axial concentrations of grape waste and peptone. Maximum activity of endoglucanase was observed by a linear increase of both grape waste and peptone concentrations. Concentrations of grape waste between 5 and 15 g/L had a positive effect on the production of β-glucosidase; peptone had no significant effects. The optimum production of the three cellulolytic activities was observed at values near the central point. A. phoenicis has the potential for the production of cellulases utilizing grape waste as the growth substrate.     

Kinetics of the Action of Na2SeO3 on Bacillus subtilis Growth as Studied by Microcalorimetry

Microcalorimetric bioassay for acute cellular toxicity is based on metabolic heat production from cultured cells.The biological response to toxicants is the inhibition of the heat production rate in cells,and toxicity is expressed as the concentration of toxicant that is 50% effective in this inhibition(IC50).In this paper,the effect of Na2SeO3 on Bacillus subtilis growth was investigated at 37℃ by microcalorimetry.The relationship between growth rate constants(k) and concentration of Na2SeO3(c) shows a logarithmic normal distribution,and IC50 is 20.3μg/mL.All these thermokinetic information is readily obtained by an LKB 2277-204 heat conduction microcalorimeter.Microcalorimetry is a quantitative,inexpensive,and versatile method for toxicology research.     

Overproduction of Glucoamylase by a Deregulated Mutant of a Thermophilic Mould <Emphasis Type="Italic">Thermomucor indicae-seudaticae</Emphasis>

Thermomucor indicae-seudaticae, a glucoamylase-producing thermophilic mould, was mutagenised using nitrous acid and gamma (⁶⁰Co) irradiation in a sequential manner to isolate deregulated mutants for enhanced production of glucoamylase. The mutants were isolated on Emerson YpSs agar containing a non-metabolisable glucose analogue 2-deoxy-d-glucose (2-DG) for selection. The preliminary screening for glucoamylase production using starch–iodine plate assay followed by quantitative confirmation in submerged fermentation permitted the isolation of several variants showing varying levels of derepression and glucoamylase secretion. The mutant strain T. indicae-seudaticae CR19 was able to grow in the presence of 0.5 g l⁻¹ 2-DG and produced 1.8-fold higher glucoamylase. As with the parent strain, glucoamylase production by T. indicae-seudaticae CR19 in 250-ml Erlenmeyer flasks attained a peak in 48 h of fermentation, showing higher glucoamylase productivity (0.67 U ml⁻¹ h⁻¹) than the former (0.375 U ml⁻¹ h⁻¹). A large-scale cultivation in 5-l laboratory bioreactor confirmed similar fermentation profiles, though the glucoamylase production peak was attained within 36 h attributable to the better control of process parameters. Although the mutant grew slightly slow in the presence of 2-DG and exhibited less sporulation, it showed faster growth on normal Emerson medium with a higher specific growth rate (0.138 h⁻¹) compared to the parent strain (0.123 h⁻¹). The glucoamylase produced by both strains was optimally active at 60 °C and pH 7.0 and displayed broad substrate specificity by cleaving α-1,4- and α-1,6-glycosidic linkages in starch, amylopectin, amylose and pullulan. Improved productivity and higher specific growth rate make T. indicae-seudaticae CR19 a useful strain for glucoamylase production.     

<Emphasis Type="Italic">Yarrowia lipolytica</Emphasis> Growth Under Increased Air Pressure: Influence on Enzyme Production

Improvement of microbial cell cultures oxygenation can be achieved by the increase of total air pressure, which increases oxygen solubility in the medium. In this work, a pressurized bioreactor was used for Yarrowia lipolytica batch cultivation under increased air pressure from 1 to 6 bar. Cell growth was strongly enhanced by the pressure rise. Fivefold and 3.4-fold increases in the biomass production and in specific growth rate, respectively, were observed under 6 bar. The increase of oxygen availability caused the induction of the antioxidant enzyme superoxide dismutase, which indicates that the defensive mechanisms of the cells against oxidative stress were effective and cells could cope with increased pressure. The pregrowth of Y. lipolytica under increased pressure conditions did not affect the lipase production ability of the cells. Moreover, the extracellular lipase activity increased 96% using a 5-bar air pressure instead of air at 1-bar pressure during the enzyme production phase. Thus, air pressure increase in bioreactors is an effective mean of cell mass and enzyme productivity enhancement in bioprocess based in Y. lipolytica cultures.     

Investigation of the Crystallization Kinetics of Zn(Leu)SO4·0.5H2O in Mixing Solvent by Microcalorimetry

The crystal growth process of Zn(Len)SO₄·0.5H₂O from water and acetone was investigated using a Calvet microcalorimeter. The heat and the rate of heat production during the crystal growth process at 293.15 K, 295.15 K, 298.15 K and 300.15 K were measured. On the basis of experimental and calculated results, the rate constant and the kinetic parameters (the activation energies, the pre‐exponential) during the crystal growth process were obtained. The results show that the crystal growth process accorded with the Burton‐Cabrera‐Frank dislocation theory.     

Biocalorimetry as a process analytical technology process analyser; robust in-line monitoring and control of aerobic fed-batch cultures of crabtree-negative yeast cells

Control of bioprocesses requires reliable and robust on- or in-line monitoring tools providing real-time information on process dynamics. Heat generation related to metabolic activity of living systems is currently gaining importance in bioprocess industry due to its non-invasive and essentially instantaneous characteristics. This study deals with monitoring and control of pure aerobic fed-batch cultures of three Crabtree-negative yeast strains, Kluyveromyces marxianus, Candida utilis and Pichia pastoris, based on in-line measured, metabolic heat flow signals. A high resolution biocalorimeter (BioRC1) was developed from a standard bench-scale heat flow calorimeter (RC1). The BioRC1 was equipped with in-line (dielectric spectroscopy, pH probe and dissolved oxygen probe) and at-line (exit gas analyser) sensors to characterise the growth behaviour of the yeast cells. Both metabolic heat flow and biomass profiles exhibited similar behaviour proving the significance of employing heat flow signal as a key-parameter for the system under investigation. A simple estimator for biomass concentration and specific growth rate was formulated based on heat flow values. In order to evaluate the potential of calorimetry as a reliable and powerful process monitoring tool, the robustness, reliability as well as the broad applicability of the developed estimators was assessed through comparison with off-line measurement techniques and showed promising results for general applicability with a wide range of bioprocesses.     

Investigation of the Crystallization Kinetics of Aqua Methioninezinc (II) Sulfate by Microcalorimetry

Summary.  The crystal growth process of aqua methioninezinc (II) sulfate (Zn(Met)SO₄·H₂O) from water and acetone was investigated using a Calvet microcalorimeter. The heat produced and the rate of heat production during the crystal growth process at 293.15, 295.15, 298.15, and 300.15 K were measured. On the basis of these results the rate constant, the Eyring parameters (the apparent activation enthalpy, the activation entropy, and the activation free energies), and the Arrhenius parameters (the activation energy, the pre-exponential factor) of the crystal growth process have been obtained. The results have shown that this crystal growth process accords with the Burton-Cabrera-Frank dislocation theory. Corresponding author. E-mail: weizhang@nwu.edu.cn Received August 16, 2002; accepted (revised) December 3, 2002 Published online May 15, 2003     

Using Multi-parameter Flow Cytometry to Monitor the Yeast <Emphasis Type="Italic">Rhodotorula glutinis</Emphasis> CCMI 145 Batch Growth and Oil Production Towards Biodiesel

Multi-parameter flow cytometry was used to monitor cell intrinsic light scatter, viability, and lipid content of Rhodotorula glutinis CCMI 145 cells grown in shake flasks. Changes in the side light scatter and forward light scatter were detected during the yeast batch growth, which were attributed to the different yeast growth phases. A progressive increase in the proportion of cells stained with PI (cells with permeabilized cytoplasmic membrane) was observed during the yeast growth, attaining 79% at the end of the fermentation. A high correlation between the Nile Red fluorescence intensity measured by flow cytometry and total lipid content assayed by the traditional gravimetric lipid analysis was found for this yeast, making this method a suitable and quick technique for the screening of yeast strains for lipid production and optimization of biofuel production bioprocesses. Medium growth optimization for enhancement of the yeast oil production is now in progress.     

Dynamic heat capacity and reaction enthalpy during the two-stage network growth in diepoxide–diamine compositions

The real and imaginary components of the dynamic heat capacity, C_p′ and C_p″, respectively, have been measured for a fixed frequency of 5 mHz during the polymerization of various compositions of a diepoxide–diamine, molecular liquid mixture to a network structure. The heat evolved during the polymerization was measured simultaneously. C_p′ decreased in two steps as the covalent bonds formed and the network structure grew. The steps became more separated when the amount of the already excess diepoxide was further increased. C_p″ showed a peak in its plot against the polymerization time, but only in the region where C_p′ showed a second step. This is attributed to the increase in the relaxation time leading to vitrification of the liquid. For the diepoxide-rich compositions, the enthalpy release also occurred in two steps and it was more for the second stage of the network's growth than for the first. Combined measurements of the exothermic effects and C_p′ and C_p″ thus delineated two stages of the network's growth by two chemical reactions. The nature of the second-stage network growth that ultimately vitrifies the stoichiometric liquid mixture is discussed. It is concluded that the second-stage growth is mass-controlled and occurs by an etherification reaction whose thermodynamic consequences have been elusive in past studies.     

Interspecies Interaction of Signal Peptide PapR Secreted by <Emphasis Type="Italic">Bacillus cereus</Emphasis> and Its Effect on Production of Antimicrobial Peptide

This study was carried out to investigate the interspecies interaction of PapR peptide secreted by Bacillus cereus on production of BSAP-254, an antimicrobial peptide produced by Bacillus subtilis SC-8 isolated from the Korean fermented soybean paste and exhibited narrow antagonistic activity against the B. cereus group, but not against other foodborne pathogens. PapR is a signal peptide that activates PlcR, which is a pleiotropic regulator controlling the expression of various virulence factors in B. cereus. When B. subtilis SC-8 was co-cultured with B. cereus, it completely inhibited the growth of B. cereus within 12 h, and the rate of BSAP-254 production was increased 34.2% at 12 h. Furthermore, 5 μM of synthetic PapR peptide added to the culture of B. subtilis SC-8 increased the rate of BSAP-254 production up to 59.7%. The growth of B. subtilis SC-8, however, was not significantly different with or without the addition of PapR. When B. cereus papR mutant was co-cultured with B. subtilis SC-8, the growth of the mutant was not inhibited and the rate of BSAP-254 production was decreased by 45%.     

Enthalpy relaxation of an epoxy–anhydride resin by temperature‐modulated differential scanning calorimetry

The enthalpy relaxation of an epoxy–anhydride resin was studied by physical aging and frequency‐dependence experiments with alternating differential scanning calorimetry (ADSC), which is a temperature‐modulated differential scanning calorimetry technique. The samples were aged at 80 °C, about 26 K below the glass‐transition temperature, for periods up to 3800 h and then scanned under the following modulation conditions: underlying heating rate of 1 K min⁻¹, amplitude of 0.5 K, and period of 1 min. The enthalpy loss was calculated by the total heat‐flow signal, and its variation with the log (aging time) gives a relaxation rate (per decade), this value being in good agreement with that calculated by conventional DSC. The enthalpy loss was also analyzed in terms of the nonreversing heat flow, revealing that this property is not suitable for calculating enthalpy loss. The effect of aging on the modulus of the complex heat capacity, |Cp*|, is shown by a sharper variation on the low side of the glass transition and an increase in the inflexional slope of |Cp*|. Likewise, the phase angle also becomes sharper in the low‐temperature side of the relaxation. The area under the corrected out‐phase heat capacity remains fairly constant with aging. The dependence of the dynamic glass transition, measured at the midpoint of the variation of |Cp*|, on ln(frequency) allows one to determine an apparent activation energy, Δh*, which gives information about the temperature dependence of the relaxation times in equilibrium over a range close to the glass transition. The values of Δh*, determined from ADSC experiments in a range of frequencies between 4.2 and 33 mHz and at an amplitude of 0.5 K, and an underlying heating rate of 1 K min⁻¹, were analyzed and compared with that obtained by conventional DSC from the dependence of the fictive temperature on the cooling rate. © 2000 John Wiley & Sons, Inc. J Polym Sci B: Polym Phys 38: 2272–2284, 2000     

Phase separation of off‐critical polymer blends of poly(styrene‐co‐maleic anhydride) and poly(methyl methacrylate). II. Morphology and mechanical properties

The effects of the phase‐separation temperature and time on the mechanical properties and morphology of poly(methyl methacrylate)/poly(styrene‐co‐maleic anhydride with 10 wt% ethyl acrylate) (SMA) blends were studied. Two compositions (20/80 and 40/60 w/w SMA/PMMAe) were prepared with a miniature twin‐screw extruder. Compared with those of the miscible blends, the Young's modulus values of the blends increased after the phase separation of the 40/60 SMA/PMMAe blend and within the early stage of spinodal decomposition of the 20/80 SMA/PMMAe blend. The mechanical properties, in terms of the tensile strength at break and the elongation, were better for the miscible blends than for the phase‐separation blends. This was believed to be the result of changes in the composition and molecular reorganization. The changes in the phase‐separating domains of both compositions, as observed by transmission electron microscopy, had no significant influence on the tensile moduli. Detailed studies of the morphology revealed a cocontinuous structure, indicating that the blends underwent spinodal decomposition. A morphological comparison of the two compositions illustrated the validity of the level rule. The growth rate of the droplet size was determined by approximation from the light scattering data and by direct measurements with transmission electron microscopy. The discrepancies observed in the droplet size growth rate were attributed to heat variations induced by the different sample thicknesses and heat transfer during the investigation. © 2004 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys 42: 886–897, 2004     

Calorimetric detection of the toxic effect of androgens on fission yeast

Calorimetry is evaluated for study of the toxic effect of environmental androgens on Schizosaccharomyces pombe cells. The results indicate that androstendione, androstandiendione and dehydrotestosterone inhibited S. pombe heat production rate. Although, the turbidimetric method showed that testosterone (TS) had no influence on growth of S. pombe, calorimetry revealed that there was a shift in growth period in samples with TS.     

Influence of Osmotic Stress on Fermentative Production of Succinic Acid by <Emphasis Type="Italic">Actinobacillus succinogenes</Emphasis>

This study investigated the influence of osmotic stress on succinic acid production by Actinobacillus succinogenes NJ113. Both cell growth and succinic acid production were inhibited with the increase in osmotic stress of the medium. The use of three different osmoprotectants in the production of succinic acid was studied in order to decrease the inhibitory effects of osmotic stress during fermentation. Results indicated that proline offers optimal osmoprotection in the production of succinic acid by A. succinogenes NJ113. In tests of batch fermentation, the maximum cell concentration was observed to be 5.36 g DCW/L after the addition of 25 mmol/L proline to the fermentation medium. The cell concentration was 24% higher than that noted for the control. A total quantity of 56.2 g/L of succinic acid was produced, with a production rate of 1 g/L per hour, after 56 h of fermentation. The concentration and productivity of succinic acid was observed to be increased by 22.2% and 22%, respectively, as compared with the control. The specific activity levels of key enzymes in the metabolic network was noted to be higher following the addition of proline, particularly in the later stages of fermentation. This method of enhancing succinic acid production by the addition of an osmoprotectant may potentially provide an alternative approach for enhanced production of other organic acids.     

Impact of sunflower and mustard leave extracts on the growth and dark respiration of mustard seedlings

The subject of the study was investigation of impact of extracts from sunflower and mustard leaves on growth of mustard seedlings. Seeds of mustard were germinated on water and then grew on aqueous extracts from sunflower or mustard leaves. The specific thermal power during seedlings growth was measured by isothermal calorimetry. Changes in the chemical composition stimulated by extracts were measured by FT-Raman spectroscopy and analyzed with the support of the cluster analysis. The heat production rate during growth of seedlings was related to the type of extracts. Crude sunflower and mustard extracts strongly inhibited the growth of seedlings when compared to non-treated control. FT-Raman spectroscopy confirms that allelopathic compounds have the greatest influence on the metabolism of fatty acids of mustard cotyledons. The obtained results indicate that sunflower and mustard extracts have varied impact on growth and heat production rate of mustard seedlings.