Influence of Growth and High Mould Concentration on the Pressure Drop in Solid State Fermentations

Aspergillus niger was grown on Amberlite IRA-900 imbibed with a solution containing high concentrations of sucrose (S_i = 100, 200, 300 and 400 g/litre) in static aerated fermentors. Growth was followed in dry biomass, biomass protein, CO₂ production and pressure drop (DP). The DP allowed the monitoring of germination, vegetative growth, limitation and the onset of sporulation for the four concentrations of sucrose studied. Concentrations up to 103 mg dry biomass/g dry support were obtained with S_i = 400 g/litre and these reduced the relative intrinsic permeability to 0·0125. Under this condition the mould occupies 34% of the free space. DP increase was related to CO₂ production.     

Mathematical modelling of the aerobic degradation of two-phase olive mill effluents in a batch reactor

A laboratory-scale study was conducted on the aerobic degradation of two-phase olive mill effluents (TPOME) made up of the mixture of the washwaters derived from the initial cleansing of the olives and those obtained in the washing and purification of virgin olive oil. The process was carried out in a 1-l working volume stirred tank reactor operating in batch mode at room temperature (25 °C). The reactor was operated at influent substrate concentrations of 2.80 g COD/l (TPOME 25%), 5.45 g COD/l (TPOME 50%), 8.18 g COD/l (TPOME 75%) and 10.90 g COD/l (TPOME 100%). After five days of operation time, total and soluble COD removal efficiencies of 64.3% and 66.6% were achieved respectively for the most concentrated influent used (TPOME 100%). A simplified kinetic model for studying the hydrolysis of insoluble organic matter, oxidation of soluble substrate and biomass production was proposed on the basis of the experimental results obtained. The following kinetic constants with their standard deviations were obtained for the above stages in the case of the most concentrated influent used (TPOME 100%): k₁ (kinetic constant for hydrolysis of suspended organic matter): 0.11 ± 0.01 l/(g VSS day); k₂ (kinetic constant for total consumption of soluble substrate): 0.30 ± 0.02 l/(g VSS day); k₃ (endogenous metabolism constant): 0.07 ± 0.01 per day). Finally, the biomass yield coefficient was found to be 0.30 g VSS/g COD_removed. The values of non-biodegradable total and soluble CODs obtained from the model were found to be 3 and 2 g/l, respectively. The kinetic constants obtained and the proposed equations were used to simulate the aerobic degradation process of TPOME and to obtain the theoretical values of non-soluble and soluble CODs and biomass concentration. The small deviations obtained (equal or lower than 10%) between the theoretical and experimental values suggest that the parameters obtained represent and predict the activity of the microorganisms involved in the overall aerobic degradation process of this wastewater.     

Medium improvement by orthogonal array designs for cholesterol oxidase production by Rhodococcus equi No. 23

Medium improvement for the production of cholesterol oxidase (CO, EC 1.1.3.6) by Rhodococcus equi No. 23 was investigated using an orthogonal array design in two steps. Results revealed that yeast extract, Tween 80 and zinc sulphate had positive effects on CO production, but magnesium sulphate had an inhibitory effect. In addition, interaction between cholesterol and sodium chloride also had a significant effect on enzyme production. The improved medium consisted of 2·0 g/litre cholesterol, 8·0 g/litre yeast extract, 1·0 g/litre NH₄Cl, 1·0 g/litre NaCl, 0·50 g/litre KH₂PO₄, 0·25 g/litre Na₂HPO₄, 0·10 g/litre -valine, 0·15 g/litre -tyrosine, 0·15 g/litre MgSO₄·7H₂O, 0·01 g/litre ZnSO₄·7H₂O, 0·10 g/litre FeSO₄·7H₂O and 4·0 ml/litre Tween 80. CO production at 60 h (about 0·24 units/ml) was about four-fold greater than with the control medium.     

Kinetic behaviour of waste tyre rubber as microorganism support in an anaerobic digester treating cane molasses distillery slops

The kinetics of anaerobic digestion of cane molasses distillery slops was investigated using a continuous-flow bioreactor which contained waste tyre rubber as support, to which the microorganisms became immobilized. Hydraulic retention times (HRT) ranging from 1 to 10 days were investigated at an average influent chemical oxygen demand (COD) concentration of 47.7?g/l. The maximum substrate utilization rate, k, and half saturation coefficient, K _L, were determined to be 1.82?kg COD_removed/kg VSS day and 0.33?kg COD/kg VSS day. The yield coefficient, Y, and sludge decay rate coefficient, K _d, were also determined to be 0.06?kg VSS/kg COD_removed and 0.05?day^-1, respectively. Methane production was maximum (6.75?l/l day) at a 2 day HRT corresponding to a biomass loading rate of 2.578?kg COD/kg VSS day. Biogas yield ranged between 0.51?l/g COD (HRT=2 days) and 0.25?l/g COD (HRT=1?day). In addition, the methane percentage in the biogas varied between 70.5% (HRT=10?days) and 47.5% (HRT=1?day). The close relationship between biomass loading rate and specific substrate utilization rate supported the use of Monod equations. Finally, the experimental values of effluent substrate concentration were reproduced with deviations equal to or less than 10% in every case.     

The Production of Polyunsaturated Fatty Acids by Microalgae: from Strain Selection to Product Purification

A selection programme to increase the cellular eicosapentaenoic acid (EPA) content has been carried out with the microalga Isochrysis galbana. The selection process involved two stages of single selection. EPA content continuously increased from 2·4% dry weight (d.w.) of the ‘parent’ culture to an average value of 5·3% d.w. in the final stage. The proportion of total EPA variation attributable to the genetic variation (heritability in a broad sense) was 0·99 showing the importance of the genome in the determination of this fatty acid. The growth and fatty acid profile of an EPA-rich isolate grown as a chemostat in a cylindrical photobioreactor have been studied. A decrease in EPA content was observed (5·21% w/w to 2·8% w/w) at the lowest dilution rate D = 0·024 h⁻¹, up close to the maximum growth rate, D = 0·038 h⁻¹. At the same time, the biomass concentration also decreased from 1015 mg/litre to 202 mg/litre over the abovementioned range of dilution rate (D). Nonetheless, the EPA productivity increases with D, with a maximum of 15·26 mg/litre/day at D = 0·0208 h⁻¹. Furthermore, steady-state dilution rates may be related to average internal light intensity. Reverse-phase, high-pressure liquid chromatography (HPLC) on octadecylsilyl semi-preparative columns was used to separate stearidonic acid (SA), EPA and docosohexaenoic acid (DHA) in polyunsaturated fatty acid concentrate obtained by the urea complexation method from a fatty acid solution previously obtained by direct saponification of biomass. Isolate SA, EPA and DHA fraction purity was 94·8, 96·0 and 94·9%, respectively, with yields of 100·0, 99·6 and 94·0%.     

Pretreatment of beet molasses to increase pullulan production

Pretreatment of beet molasses with cation exchange resin, sulphuric acid, tricalcium phosphate, potassium ferrocyanide, and ethylenediaminetetraacetic acid and disodium salt (EDTA) to increase the production of pullulan was investigated. Among the above techniques used for the removal of heavy metals, sulphuric acid treatment gave better results regarding polysaccharide concentration, polysaccharide yield, and sugar utilization. Aureobasidium pullulans grown on beet molasses produced a mixture of pullulan and other polysaccharides. The pullulan content of the crude polysaccharide was 30–35%. The addition of nutrients improved the production of polysaccharide. A maximum polysaccharide concentration (32·0±1·0 g litre⁻¹) was achieved in molasses solution (70 g litre ¹ initial sugar concentration, pH 6·5–7·5) treated with sulphuric acid and supplemented with K₂HPO₄ 0·5%, -glutamic acid 1%, olive oil 2·5% and Tween 80 0·5%. In this case, the highest values of biomass dry weight (33·8±1·0 g litre⁻¹), polysaccharide yield (63·5±2·5%), and sugar utilization (97·5±1·5%) were obtained at pH 6·5, 3·5, and 4·5–7·5, respectively.     

Determination of kinetic constants for a two-stage anaerobic upflow packed-bed reactor for dairy wastewater

A two-stage upflow packed-bed (reactors in series) system was used for the treatment of dairy wastewater. Nylon pads were used as supporting media for the biomass. This investigation aimed at the determination of various kinetic constants for substrate, biomass and biogas based on various models. The maximum loadings that could be applied to reactor I and reactor II were 14·29 and 5·0 kg of chemical oxygen demand (COD) per m³ per day, respectively. The maximum COD removal efficiencies at various loading rates were in the ranges of 93·8–98·5% and 72·5–84% for the two reactor systems, respectively. The combined biogas yield was between 0·196 and 0·386 m³ gas/kg COD_a.     

Characterization of gliadin-galactomannan incubation mixtures by analytical ultracentrifugation—Part I. Sedimentation velocity

The aim of this work is to examine the possible interaction and extent thereof of the polysaccharide galactomannan (GAL) with the cereal protein gliadin (GLI) and a peptic-tryptic degraded gliadin (PT-GLI) by analytical ultracentrifugation. The work is part of a series of investigations into the field of coeliac disease (gluten-induced enteropathy) as gliadins are known to be toxic for patients with this disease.

The molecular integrity of the GAL and GLI preparations was first checked by sedimentation velocity and sedimentation equilibrium. Sedimentation velocity showed single boundaries indicating homogeneity and low-speed sedimentation equilibrium gave plausible apparent weight average molar masses of 180,000 g/ mol for GAL and 20,000 g/mol for GLI. PT-GLI, GLI and GAL in phosphate buffer (pH 6.5) and the incubated mixtures (stirred for 3 h at 37 °C; PT-GLI: GAL = 3.53:1, wt.wt.; GLI:GAL = 0.23 and 0.55:1, wt.wt.) were then investigated by sedimentation velocity at a temperature of 20 °C. The plots of 1/s₂₀ vs. c of GAL, PT-GLI-GAL and GLI-GAL mixtures after incubation show a significantly different shape suggesting the presence of interactions. According to the equation 1/s₂₀ = 1/s^o₂₀(1 + k_sc), values for {s^o₂₀, k_s} of {(4.02 ± (490.9 ± 28.9) ml/g, {(5.92 ± 0.24) S, (1152 ± 44) ml/g} and {(5.38 ± 0.19) S, (1141 ± 38) ml/g} for GAL and PT-GLI-GAL and GLI-GAL mixtures, respectively, were obtained. The concentration of GAL ranged from 0.75–3.0 mg/ml for GAL alone and from 0.34–1.50 mg/ml in the incubated mixtures. This apparent indication for a weak non-covalent protein-polysaccharide interaction was further supported by UV absorption spectrometry and gel filtration.     

Influence of phosphorus concentration on the growth kinetics and stoichiometry of the microalga Scenedesmus obliquus

The growth of the freshwater microalga Scenedesmus obliquus was studied at 30°C in a mineral culture medium with phosphorus concentrations of between 0 and 372 μ . The values for the specific growth rates, between and , fitted a semistructured substrate-limitation model with μ_m1 = 0·0466 h⁻¹, μ_m2 = 0·0256 h⁻¹ and . The specific uptake rate of phosphorus reached a maximum value of q_Sm1 = 658·01 × 10⁻⁴ μmol P mg⁻¹ biomass h⁻¹.     

Operation strategies for biohydrogen production with a high-rate anaerobic granular sludge bed bioreactor

Long-term operation for biohydrogen production with an efficient carrier-induced granular sludge bed (CIGSB) bioreactor had encountered problems with poor biomass retention at a low hydraulic retention (HRT) as well as poor mass-transfer efficiency at a high HRT or under a prolonged operation period. This work was undertaken to develop strategies enabling better biomass retention and mass-transfer efficiency of the CIGSB reactors. Supplementation of calcium ion was found to enhance mechanical strength of the granular sludge. Addition of 5.4–27.2 mg/l of Ca²⁺ also led to an over three-fold increase in biomass concentration and a nearly five-fold increase in the H₂ production rate (up to 5.1 l H₂/h/l). Two reflux strategies were utilized to enhance the mass-transfer efficiency of the CIGSB system. The liquid reflux (LR) strategy enhanced the H₂ production rate by 2.2-fold at an optimal liquid upflow velocity of 1.09 m/h, which also gave a maximal biomass concentration of ca. 22 g VSS/l. Similar optimal H₂ production rate was also obtained with the gas reflux (GR) strategy at a rate of 1.0–1.49 m/h, whereas the biomass concentration decreased to 2–7 g VSS/l and thereby the specific H₂ production rate was higher than that with LR. The operation strategies applied in this work were effective to allow stable and efficient H₂ production for nearly 100 days.     

Power consumption of three impeller combinations in mixing Xanthan fermentation broths

Xanthan gum fermentation represents a good model for the study of the mixing of rheologically complex culture broths. Most of the previous work on power consumption dealt with ‘standard’, single impellers and used model fluids to simulate xanthan broths. This work describes the characterization of three dual-impeller combinations (D/T = 0·53) for the mixing of dehydrated—reconstituted fermentation broths of Xanthomonas campestris that had matched rheology to the actual broths. The bottom impeller was a Rushton turbine (RT) and the top impeller was another RT, a 45° pitched blade turbine (PT) or an A-310 Lightnin mixer (A310). The experiments were carried out in a tank of 0·0094 m³ working volume equipped with an air bearing dynamometer. The power was measured in a wide range of xanthan concentrations (5–40 kg m⁻³) in aerated (0·25, 0·5 and 1·0 vvm) and unaerated conditions. Unaerated power number (Po) vs. Reynolds number (Re) curves showed similar trends for the three combinations. Exponents close to −1 were obtained in the laminar region. A minimum in Po (Po_min) occurred at Re = 30–40, then increasing to a plateau value which was evident at Re> 200. In the transition region Po_min values were 4·3 (RT and RT), 3·6 (RT and PT) and 2·4 (RT and A310). The aerated power data for (RT and PT) and (RT and A-310) showed higher torque instabilities than the dual RT combinations at higher xanthan concentrations. The higher the xanthan concentrations, the higher the drop in power and the less important the effect of the aeration rate. Among the combinations tested, when using Rushton turbines, the well-mixed ‘cavern’ reached the tank wall (i.e., fluid motion was observed) at the lowest volumetric power input. High     

Solid substrate cultivation of Gibberella fujikuroi on an inert support

Growth of Gibberella fujikuroi on Amberlite, an inert support, and gibberellic acid (GA₃) production was studied in glass columns under different conditions of temperature and water activity (a_w). Maximum biomass concentration and GA₃ production were respectively 40 (mg/g inert support) and 0.73 (mg/g inert support). While high specific growth rates were obtained, low initial nitrogen resulted in low biomass concentrations. Maximum GA₃ (31°C, a_w=0.985) was not produced by the maximum concentration of biomass (25°C, a_w=0.992). Peaks in the rate curves of either outlet gas, CO₂ or O₂, occurred on exhaustion of urea indicating, for future works, just when to feed the culture additional nitrogen.     

Ethanol production from glucose and xylose by separated and co-culture processes using high cell density systems

Media containing xylose and/or glucose were tested utilizing Zymomonas mobilis or Saccharomyces diastaticus and Pichia stipitis. The best fermentation results were obtained in separated glucose (180 g/litre) and xylose (80 g/litre) fermentations utilizing Zymomonas mobilis and Pichia stipitis strains, respectively. In these conditions, the maximum ethanol concentrations achieved were 86·2 g/litre and 29 g/litre, respectively. The complete conversion of a glucose and xylose mixture (50 g/litre) was obtained using a respiratory deficient mutant of Saccharomyces diastaticus co-cultivated with Pichia stipitis in continuous culture. Using the co-culture process, the maximum ethanol concentration was 21·5 g/litre (Yp/s=0·45 g/g) and the maximum volumetric ethanol productivity was 4·3 g/(litre × h).     

Competitive product inhibition of aromatase by natural estrogens

In order to better understand the function of aromatase, we carried out kinetic analyses to asses the ability of natural estrogens, estrone (E₁), estradiol (E₂), 16-OHE₁, and estriol (E₃), to inhibit aromatization. Human placental microsomes (50 μg protein) were incubated for 5 min at 37°C with [1β-³H]testosterone (1.24 × 10³ dpm ³H/ng, 35–150 nM) or [1β-³H,4-¹⁴C]androstenedione (3.05 × 10³ dpm ³H/ng, ³H/¹⁴C = 19.3, 7–65 nM) as substrate in the presence of NADPH, with and without natural estrogens as putative inhibitors. Aromatase activity was assessed by tritium released to water from the 1β-position of the substrates. Natural estrogens showed competitive product inhibition against androgen aromatization. The K_i of E₁, E₂, 16-OHE₁, and E₃ for testosterone aromatization was 1.5, 2.2, 95, and 162 μM, respectively, where the K_m of aromatase was 61.8 ± 2.0 nM (n = 5) for testosterone. The K_i of E₁, E₂, 16-OHE₁, and E₃ for androstenedione aromatization was 10.6, 5.5, 252, and 1182 μM, respectively, where the K_m of aromatase was 35.4 ± 4.1 nM (n = 4) for androstenedione. These results show that estrogens inhibit the process of andrigen aromatization and indicate that natural estrogens regulate their own synthesis by the product inhibition mechanism in vivo. Since natural estrogens bind to the active site of human placental aromatase P-450 complex as competitive inhibitors, natural estrogens might be further metabolized by aromatase. This suggests that human placental estrogen 2-hydroxylase activity is catalyzed by the active site of aromatase cytochrome P-450 and also agrees with the fact that the level of catecholestrogens in maternal plasma increases during pregnancy. The relative affinities and concentration of androgens and estrogens would control estrogen and catecholestrogen biosynthesis by aromatase.     

On the ecological energetics of 0-group Carcinus maenas (L.) from a shallow sandy bottom in gullmar fjord, Sweden

0-group Carcinus maenas (L.) was investigated from June 1975 to September 1976 on a shallow sandy bottom at Kvarnbukten Bay, Gullmar Fjord (58° 15′N: 11°28′E), Sweden, at an average salinity of 25% and a range of monthly mean temperatures of −0.3 to 197. °C.

The new year-class settles from August to early September at a carapace breath of 2 to 3 mm and a calorific content of 32 cal. The distribution is restricted to clusters of the mussel Mytilus edulis L. Depth, type of substratum, and patches of the eel-grass Zostera marina L. are of no importance for their spatial distribution. There is no migration to deeper water in the autumn. The carapace breadth is ≈ 9.5 mm after one year of benthic life. Sexual maturity is reached after two years. Growth occurs at temperatures above 10 °C, i.e., from August to October and from May to July. During the first year of benthic life the animals moult 7 times. The 0-group seems to be micro-carnivores feeding on the sediment meiofauna.

The individual energy budget for the first year of benthic life is: consumption (C_c) 905 cal., production (P_1c) 236 cal., cast carapaces (P_2c) 153 cal., respiration (R_c) 404 cal., and rejectiction (F_c) 112 cal. The assimilation efficiency (U_c⁻¹) is 88%, the gross growth efficiency (K_1c) 43%, and the net growth efficiency (K_2c) 49%.

At Kvarnbukten Bay there are large variations in size between the separate year-classes. The energy content of the food consumed by the 1975/76 cohort was used as follows: 4% was stored in living biomass after one year, 36% was released to other trophic levels as dead animals and cast carapaces, 13% rejected as faeces, and 47% was lost through respiration.     

Influence of substrate concentration on the macroenergetic parameters of anaerobic digestion of black-olive wastewater

Summary The macroenergetic parameters of the anaerobic digestion of black-olive wastewater, i.e. the yield coefficient for the biomass (Y. g VSS/g COD) and the specific rate of substrate uptake for cell maintenance (m, g COD/g VSS-day) decreased 6 limes and increased 5 times. respectively, when the influent substrate concentration increased from 1.1 to 4.4 g COD/l. This was significant at 95% confidence level. The use of the Guiot kinetic model allows a more accurate prediction of growth yield to be made as it relates substrate utilization to product formation.     

Investigations on Nickel Biosorption and its Remobilization

Immobilized preparations of the bacteria (Pseudomonas aeruginosa and Rhodopseudomonas BHU strain 1) and the cyanobacterium ( Anacystis nidulans) exhibited significant Ni adsorption in the order 91%, 72%, 75%, respectively, within 2 h contact with aqueous NiCl₂ (7·05 μg Ni/0·1 mg biomass). The immobilizing agent (Ca-alginate, 1·5%, w/v) absorbed more Ni (43%) than the exopolysaccharide of cyanobacteria, Rivularia sp. (40%) or Aphanothece sp. (30%). Ni remobilization from different adsorbed systems was maximum (84%) for Ca(NO₃)₂ over NaCl (4·3%) at equimolar concentrations (12 m , each). Extracts from forest soil (organic C, 2-3%) were more effective in Ni remobilization (22·65%) than similar preparations from garden soil (18%) with organic C in the range of 0·98-1·1%.     

Determination of effective diffusion coefficient of substrate in gel particles with immobilized biocatalyst

A method of determining of the effective diffusion coefficient of substrate in a particle, where the diffusion and consumption of substrate by biocatalytic reaction are present simultaneously, was designed and experimentally verified. The method is based on measuring the overall rate of heterogeneous biocatalytic reaction in particles of varying diameter. The effective diffusion coefficient, D_e, was determined by fitting the measured reaction rates with the solution of the reaction-diffusion equation. The method is tailored for cases where the enzyme reaction is governed by the Michaelis-Menten kinetics. The value of K_m required for the solution of the mathematical model was adopted from the measurement of the kinetics of free cells, whereas the rate parameter, k₂, was optimized together with D_e. As an experimental model, the sucrose hydrolysis catalyzed by Ca-alginate-entrapped yeast cells was examined. The particle diameter varied in the range of 1.2–3.9 mm and the initial reaction rates were measured in a batch-stirred reactor at a sucrose concentration of 100 m . The D_e of sucrose at 30°C was found to be 2.9 · 10⁻¹⁰ m²s⁻¹.     

Production d'NH4 par la crevette Crangon crangon L. dans deux écosystémes côtiers. approche expérimentale et étude de l'influence du sédiment sur le taux d'excrétion

Estimation of the ammonia production of the shrimp C. crangon in two littoral ecosystems (oligotrophic sand and eutrophic mud) was determined in winter and summer conditions from laboratory observations in experimental microcosms. The ammonia excretion rate of C. crangon was not influenced by either the sediment type or the ammonia concentration of the overlying water; on the other hand, the mean excretion rate and the response to initial handling stress increased markedly as shrimp were deprived of soft substratum.

The daily ammonia production of C. crangon was 16 μmol NH₃ · g ⁻¹ wet wt · day ⁻¹ in winter and 40 μmol in summer. A gross production of 12 μmol NH₃ · m⁻² · day ⁻¹ and 300–700 μmol μ m⁻² · day⁻¹, respectively, could be expected in the two ecosystems studied. This would account for 5% (winter) and 2–4% (summer) of the total NH⁺₄ flux at the sediment-water interface. The contribution of the excretion of all macrofauna to the NH⁺₄ flux from the sediment is discussed.     

Crystal and molecular structure of polymeric [MnCl2(bpy)]n

A crystal and molecular structure determination of MnCl₂(bpy) showed that it exists as polymeric octahedral [MnCl₂(bpy)]_n. [MnCl₂(bpy)_n crystallizes in the monoclinic space group I2/a with A = 7.007(1), B = 9.200(1), C = 16.495(1) Å, β = 91.313(5)° and Z = 4. On the basis of 979 unique observed reflections with I 2.5σ(I) the structure was refined to R = 0.032.