Effects of growth morphology and time of harvesting on the chitosan yield of Absidia repens

Summary Absidia repens (CBS 102.32) was grown in a fermentor and the effects of growth morphology (due to different agitation) and harvest timing on chitosan yield were evaluated. The use of the titration rate as an on-line measure of growth rate was studied. Small pellets, 0.5 mm o.d. allowed the most efficient growth (highest growth rate and highest biomass yields from carbon and nitrogen sources) whereas growth as large pellets, 2–3 mm o.d., or as a viscous pulp exhibited limited growth. The differences were most pronounced during the later part of the cultivations. The chitosan content of the biomass remained essentially constant during active growth, irrespective of morphology, but during the stationary phase, this content continued to increase from 18% to 23% of the biomass, reaching 2.8 g/l. The titration rate of NaOH, in order to maintain constant pH, exceeded the growth rate in all cultures, and this was pronounced when growth was limited. Correspondence to: A. Persson     

Optimization of culture conditions and continuous production of chitosan by the fungi,Absidia coerulea

The production of chitosan from the mycelia ofAbsidia coerulea was studied to improve cell growth and chitosan productivity. Culture conditions were optimized in batch cultivation (pH 4.5 agitator speed of 250 rpm, and aeration rate of, 2 vvm) and the maximum chitosan concentration achieved was 2.3 g/L under optimized conditions. Continuous culture was carried out successfully by the formation of new growth spots under optimized conditions, with a chitosan productivity of 0.052 gL⁻¹ h⁻¹, which is the highest value to date, and was obtained at a dilution rate of 0.05 h⁻¹. Cell chitosan concentrations reached about 14% in the steady state, which is similar to that achieved in batch culture. This study shows that for the continuous culture ofAbsidia coerulea it is vital to control the medium composition.     

Fermentative production of 1,3-propanediol from glycerol by Clostridium butyricum up to a scale of 2m3

Summary The conversion of glycerol to 1,3-propanediol (PD) by Clostridium butyricum DSM 5431 was studied in anaerobic culture. Growth and product formation were optimal at pH = 7.0 and T = 35° C, while aeration rate and stirrer speed were found to have no significant influence. As increasing amounts of initial glycerol led to inhibition of growth, cultivations were done in fed-batch operation. Comparative cultivations were carried out in an air-lift (ALR) and a stirred-tank reactor (STR) having equal working volumes (V _L = 30 l) and no difference in product formation was found. The process was scaled up to reactor sizes of 1.2 m³ (ALR) and 2.0 m³ (STR). The same results were obtained irrespective of reactor volume as well as reactor type (STR/ALR). PD concentrations of approximately 50–58 g·l^–1 and overall productivities of 2.3–2.9 g·l^–1 ·h^–1 could be reached. Offprint requests to: W.-D. Deckwer     

Decolourization of anaerobically digested and polyaluminium chloride treated distillery spentwash in a fungal stirred tank aerobic reactor

Decolourization of anaerobically digested and polyaluminium chloride treated distillery spentwash was studied in a fungal stirred tank aerobic reactor without dilution of wastewater. Aspergillus niger isolate IITB-V8 was used as the fungal inoculum. The main objectives of the study were to optimize the stirrer speed for achieving maximum decolourization and to determine the kinetic parameters. A mathematical model was developed to describe the batch culture kinetics. Volumetric oxygen transfer coefficient (k _L a) was obtained using dynamic method. The maximum specific growth rate and growth yield of fungus were determined using Logistic equation and using Luedeking–Piret equation. 150 rpm was found to be optimum stirrer speed for overall decolourization of 87%. At the optimum stirrer speed, volumetric oxygen transfer coefficient (k _L a) was 0.4957 min⁻¹ and the maximum specific growth rate of fungus was 0.224 h⁻¹. The values of yield coefficient (Y _x/s) and maintenance coefficient (m _s) were found to be 0.48 g cells (g substrate)⁻¹ and 0.015 g substrate (g cells)⁻¹ h⁻¹.     

Microcalorimetric investigations of the metabolism of yeasts. Growth in chemostat cultures on glucose

Summary As part of a project on the production of penicillin, the penicillin production of two strains of Penicillium chrysogenum which have a different penicillin productivity was investigated in bubble column bioreactors and for comparison in stirred fermenters. The main interest of this study were the complicated interrelations between the stirrer speed, the stirrer type, the shear stress, the morphology of the mycelium and broth viscosity as well as the effect of the oxygen transfer behavior on antibiotic productivity.Stirred tank reactors with different turbine stirrers as well as with a draught tube and propeller were employed.The main variable investigated was the stirrer speed. At low stirrer speeds, gas dispersion is inadequate and the insufficient oxygen transfer rate is a limiting factor. At higher stirrer speeds, the oxygen supply of pulpy mycelia is improved and more cell mass is formed. This result is the same for both strains in all three reactors.If the oxygen partial pressure is near the lower cirtical value, a high percentage of the carbon source is converted into penicillin but the penicillin productivity is low due to a low percentage of penicillin producing cells. At oxygen partial pressures just above 8% saturation, the absolute penicillin productivity is maximal. At higher stirrer speeds and dissolved oxygen concentrations the penicillin production phase is shorter, cell growth is higher and a higher percentage of the carbon source is converted into CO₂.In reactors with a draught tube and propeller, a lower productivity is attained than in those with turbine stirrers.The behavior of the two strains is fairly similar. The higher producing strain, however, has a more distinct separation between its periods of growth and production than does the low producing one. At high stirrer speeds the increase in the cell growth rate is less significant and the substrate yield coefficients are higher for the high producing strain than for the low producing one.Symbols C Dissolved oxygen concentration (mg l^–1) - C^* C at saturation (mg l^–1) - k_La Volumetric mass transfer coefficient (h^–1) - OTR Oxygen transfer rate (mg l^–1 h^–1) - OUR Oxygen uptake rate (mg l^–1 h^–1) - rpm Impeller speed (min^–1) - X (Dry) biomass concentration (g kg^–1) - V_g Volumetric gas flow rate (Nl min^–1) - CMC Carboxymethyl cellulose     

Physiological diversity within the <Emphasis Type="Italic">kluyveromyces marxianus</Emphasis> species

The Kluyveromyces marxianus strains CBS 6556, CBS 397 and CBS 712^T were cultivated on a defined medium with either glucose, lactose or sucrose as the sole carbon source, at 30 and 37°C. The aim of this work was to evaluate the diversity within this species, in terms of the macroscopic physiology. The main properties evaluated were: intensity of the Crabtree effect, specific growth rate, biomass yield on substrate, metabolite excretion and protein secretion capacity, inferred by measuring extracellular inulinase activity. The strain Kluyveromyces lactis CBS 2359 was evaluated in parallel, since it is the best described Kluyveromyces yeast and thus can be used as a control for the experimental setup. K. marxianus CBS 6556 presented the highest specific growth rate (0.70 h⁻¹) and the highest specific inulinase activity (1.65 U mg⁻¹ dry cell weight) among all strains investigated, when grown at 37°C with sucrose as the sole carbon source. The lowest metabolite formation and highest biomass yield on substrate (0.59 g dry cell weight g sucrose⁻¹) was achieved by K. marxianus CBS 712^T at 37°C. Taken together, the results show a systematic comparison of carbon and energy metabolism among three of the best known K. marxianus strains, in parallel to K. lactis CBS 2359.     

The occurrence of oligochaetes in lake chains receiving pulp mill waste and their relation to eutrophication on the trophic scale

Earlier conceptions of oligochaetes as indicators of trophy were somewhat enlarged to include more littoral, meiofaunal and naidid species. The amplitudes of different species on the oligotrophy-eutrophy axis seem to be generally large. Certain species which probably originally occur mainly in very humic environments seem also to be able to endure very strong organic loading caused by pulp mills. Species of this kind seem to be Dero digitata, Specaria josinae, Slavina appendiculata and Aulodrilus pluriseta. Chaetogaster langi and Arcteonais lomondi are also species which live in areas receiving pulp mill wastes. A. lomondi and D. digitata seem to be species living in greater depths than the naidids generally. Tubifex tubifex densities as high as 600000 ind · m^-2 were found near a pulp mill when the meiofauna method was used. A third of the T. tubifex were found deeper than 5 cm from the sediment surface. S. josinae and S. appendiculata may be found down to depths between 5 and 10 cm.     

Influence of ultrasound amplitude and duty cycle on fungal morphology and broth rheology of <Emphasis Type="Italic">Aspergillus terreus</Emphasis>

Effects of ultrasound amplitude and duty cycle on cultures of Aspergillus terreus are reported in a 25 l slurry bubble column sonobioreactor. Fermentations were carried out batchwise. A 2^k-factorial design with added central points was used. Sonication at any cycle and amplitude level did not affect biomass growth rate and yield relative to nonsonicated control, but did affect growth morphology. Ultrasound disrupted fungal pellets and caused the biomass to grow mainly as dispersed hyphae. Production of lovastatin was reduced by medium- and high-cycle sonication. Sonication affected broth rheology. In view of these results, sonication can be used to modify growth morphology and broth rheology without affecting growth rate and yield of filamentous fungi.     

Oxidation of amines by yeasts grown on 1-aminoalkanes or putrescine as the sole source of carbon,nitrogen and energy

The maximum growth rate of Trichosporon cutaneum CBS 8111 in chemostat cultures was 0.185 h^-1 on ethylamine and 0.21 h^-1 on butylamine, that of Candida famata CBS 8109 was 0.32 h^-1 on putrescine.The amine oxidation pattern of the ascomycetous strains studied, viz. Candida famata CBS 8109, Stephanoascus ciferrii CBS 4856 and Trichosporon adeninovorans CBS 8244 was independent of the amine that had been used as the growth substrate. It resembled that of benzylamine/putrescine oxidase found in other ascomycetous yeasts. However, differences in pH optimum and substrate specificity were observed between the amine-oxidizing systems of these three species.The amine oxidation pattern of cell-free extracts of Trichosporon cutaneum CBS 8111 varied with the amine that was used as growth substrate. The enzyme system produced by Cryptococcus laurentii CBS 7140 failed to oxidize isobutylamine and benzylamine, and showed a high pH optimum.The synthesis of amine oxidase in the four yeast strains studied was not repressed by ammonium chloride and was weakly repressed by glucose but was strongly repressed if both compounds were present in the growth medium.     

Respiration‐induced weathering patterns of two endolithically growing lichens

The two endolithic lichen species Hymenelia prevostii and Hymenelia coerulea were investigated with regard to their thallus morphology and their effects on the surrounding substrate. The physiological processes responsible for the observed alterations of the rock were identified. Whereas the thallus surface of H. coerulea was level, H. prevostii formed small depressions that were deepest in the thallus center. In a cross‐section, both species revealed an algal zone consisting of algal cavities parallel to the substrate surface and a fungal zone below. However, H. prevostii revealed significantly larger cavities with more than twice the cell number and a denser pattern of cavities than H. coerulea, resulting in a biomass per surface area being more than twice as large. Below H. prevostii the layer of macroscopically visibly altered rock material was about twice as deep and within this layer, the depletion of calcium and manganese was considerably higher. In simultaneous measurements of the oxygen uptake/oxygen release and pH shift, the isolated algal strains of both lichens revealed respiration‐induced acidification of the medium in the dark. At higher light intensities, H. coerulea and to a lesser extent also H. prevostii alkalized the medium which may lessen the acidification effect somewhat under natural conditions. In a long‐term growth experiment, the isolated algal strains of both lichens revealed acidification of the medium to a similar extent. Neither acidic lichen substances nor oxalic acid was identified. The significant differences between the weathering patterns of both species are based on the same respiration‐induced acidification mechanism, with H. prevostii having a greater effect due to its higher biomass per area.     

Production of ψ-linolenic acid by the fungus Mucor rouxii on cheap nitrogen and carbon sources

Summary The production of -linolenic acid (GLA) by the fungus Mucor rouxii CBS 416.77 was studied on low budget nitrogen and carbon sources, i.e. rape meal, cocos expeller and two types of yeast extract (nitrogen sources), and starch, starch hydrolysate, beet molasses and cocos expeller (carbon sources). As references, Difco yeast extract and glucose were used. In flask cultivations the three yeast extracts were fully interchangeable, while the Difco yeast extract (the most expensive of those tested) gave a higher productivity of GLA in fermentor cultures (14 mg·l^–1·h^–1). The yield of lipids and GLA were increased in the order yeast extract < rape meal < cocos expeller. Thus the amount of lipid increased from 0.56 to 2.8 g·l^–1, and that of GLA from 0.15 to 0.33 g·l^–1. Use of beet molasses or cocos expeller as carbon sources gave poor growth. Starch and starch hydrolysate resulted in better productivity of GLA than glucose (4.7 and 4.9 compared to 3.4 mg·l^–1·h^–1). Offsprint requests to: A.-M. Lindberg     

Formal kinetics of poly-β-hydroxybutyric acid (PHB) production in Alcaligenes eutrophus H 16 and Mycoplana rubra R 14 with respect to the dissolved oxygen tension in ammonium-limited batch cultures

Summary Under chemolithoautotrophic growth conditions with the organism Alcaligenes eutrophus H16 the exponential growth phase is characterized by two different growth rates, each associated with different specific rates of ammonium consumption. On the basis of the analytical determination of Poly--hydroxybutyric acid (PHB), it can be conclusively shown that PHB is synthesized even during the exponential growth phase at a specific rate proportional to the specific growth rates of total biomass. After complete consumption of ammonium, the increase of biomass is exclusively due to PHB synthesis, whereas protein and rest biomass (cell dry weight minus PHB) remain constant. After an extended period of fermentation, the PHB content reaches a saturation value. The transient phase between the growth and the storage phase is very short in comparison to the duration of the whole fermentation. In the case of Alcaligenes eutrophus, strain H 16, high concentrations of dissolved oxygen strongly influence growth as well as PHB synthesis.Abbrevations ^cO₂,L concentration of oxygen in the liquid phase (dissolved oxygen tension: d.o.t) - ^cH₂,L concentration of hydrogen in the liquid phase - ^cCO₂,L concentration of carbon dioxide in the liquid phase - S limiting substrate, concentration of - X total biomass, concentration of; total cell dry weight - P product; PHB, concentration of - R rest biomass: X-P, concentration of - ^rX dX/dt growth rate - ^rP dP/dt rate of PHB synthesis - ^rR dR/dt rate of rest biomass production - ^r0 ^dcO₂,L/dt rate of oxygen consumption - X dX/dt·1/X=r_X·1/X specific growth rate - P dP/dt·1/P=r_P·1/P specific rate of product formation - R dR/dt·1/R=r_R·1/R specific rate of rest biomass formation - r₀/R specific respiration rate     

Effect of stirrer speed,aeration rate and cell mass concentration on volumetric oxygen transfer coefficients (KLa) in the cultivation of Curvularia lunata in a batch reactor

Summary Curvularia lunata was grown in a stirred and aerated reactor for the production of extracellular rifamycin oxidase. Volumetric oxygen transfer coefficients (K_La) were measured for various stirrer speeds, rates of aeration and cell mass concentrations in the reactor. Stirrer speed and aeration rate were optimized and it was found that stirrer speeds of 400–500 rpm and aeration rates of 0.75–1 vvm were optimum for the maximum amount of enzyme production. It was noticed that the increase in cell mass decreased the oxygen transfer coefficient. It was also noticed that the organism formed pellets rather than mycelia when grown on glucose and with an increase in the concentration of glucose in the reactor, there was heavy pellet formation.     

Four novel yeasts from decaying organic matter: Blastobotrys robertii sp. nov., Candida cretensis sp. nov., Candida scorzettiae sp. nov. and Candida vadensis sp. nov

Four novel yeast species are described, two from decaying mushrooms, viz. Candida cretensis and Candida vadensis, and two from rotten wood, viz. Blastobotrys robertii and Candida scorzettiae. Accession numbers for the CBS and ARS Culture Collections, and GenBank accession numbers for the D1/D2 domains of the large subunit of ribosomal DNA are: B. robertii CBS 10106^T, NRRL Y-27775, DQ839395; C. cretensis CBS 9453^T, NRRL Y-27777, AY4998861 and DQ839393; C. scorzettiae CBS 10107^T, NRRL Y-27665, DQ839394; C. vadensis CBS 9454^T, NRRL Y-27778, AY498863 and DQ839396. The GenBank accession number for the ITS region of C. cretensis is AY498862 and that for C. vadensis is AY498864. C. cretensis was the only species of the four that displayed fermentative activity. All four type strains grew on n-hexadecane. C. scorzettiae is the only one of the new species that assimilates some phenolic compounds, viz. 3-hydroxy derivatives of benzoic, phenylacetic and cinnamic acids, but not the corresponding 4-hydroxy acids. This is indicative of an operative gentisate pathway.     

Production of β-1.4-xylanase in continuous culture byAureobasidium Pullulans CBS 58475

Summary The fungusAureobasidium pullulans CBS 58475 can be fermented easily and economically in its yeast-like form. The enzyme production is strictly connected to biomass production. In continuous culture, at a dilution rate of D=0.085 h^–1, the fungus shows a volumetric -xylanase productivity of about 300 U/h·1.     

Solar heat gain in a desert rodent: unexpected increases with wind speed and implications for estimating the heat balance of free-living animals

We quantified metabolic power consumption as a function of wind speed in the presence and absence of simulated solar radiation in rock squirrels, Spermophilus variegatus, a diurnal rodent inhabiting arid regions of Mexico and the western United States. In the absence of solar radiation, metabolic rate increased 2.2-fold as wind speed increased from 0.25 to 4.0 m·s^-1. Whole-body thermal resistance declined 56% as wind speed increased over this range, indicating that body insulation in this species is much more sensitive to wind disruption than in other mammals. In the presence of 950 W·m^-2 simulated solar radiation, metabolic rate increased 2.3-fold as wind speed was elevated from 0.25 to 4.0 m·s^-1. Solar heat gain, calculated as the reduction in metabolic heat production associated with the addition of solar radiation, increased with wind speed from 1.26 mW·g^-1 at 0.25 m·s^-1 to 2.92 mW·g^-1 at 4.0 m·s^-1. This increase is opposite to theoretical expectations. Both the unexpected increase in solar heat gain at elevated wind speeds and the large-scale reduction of coat insulation suggests that assumptions often used in heat-transfer analyses of animals can produce important errors.Abbreviations absorptivity of coat to solar radiation - kinematic viscosity of air (mm²·s^-1) - reflectivity of coat to solar radiation - a r _B expected at zero wind speed (s·m^-1) - A _P projected surface area of animal on plane perpendicular to solar beam (cm²) - A _SKIN skin surface area (cm²) - b Coefficient describing change in r _B with change in square-root of wind speed (s^1.5·m^1.5) - d hair diameter (m) - d characteristic dimension of animal (m) - D _H thermal diffusivity of air (m²·s^-1) - E evaporative heat loss (W·m^-2) - I probability per unit coat depth that photon will strike hair - k constant equalling 1200 J·m^-3·°C^-1 - l _C coat depth m) - l _H hair length (m) - M metabolic rate (W·m^-2) - n density of hairs of skin (m^-2) - Q _A solar heat gain to animal (W·m^-2) - Q _I solar irradiance intercepted by animal (W·m^-2) - RQ respiratory quotient - r _A thermal resistance of boundary layer (s·m^-1) - r _B whole-body thermal resistance (s·m^-1) - r _E thermal resistance between animal surface and environment s·m^-1) - r _R radiative resistance (s·m^-1) - r _S sum of r _B and r _E at 0.25 m·s^-1 (s·m^-1) - r _T tissue thermal resistance s·m^-1) - T _AIR air temperature (°C) - T _B body temperature (°C) - T _E operative temperature of environment (°C) - T _ES standard operative temperature of environment (°C) - u wind speed (m·s^-1)     

Kinetics of α-amylase production in a batch and fed-batch culture of Bacillus subtilis with caseinate as nitrogen source and starch as carbon source

Summary Analysis of a large number of experimental data from the cultivation of Bacillus subtilis formed the basis for a kinetic model of the process explaining the effect of composition of the culture medium and of the growth rate on the rate of enzyme production. The resulting rate of formation of -amylase (EC 3.2.1.1) reflects the sum of the rate of enzyme production and the rate of its degradation as affected by the environment. The kinetic dependence confirms the previously described mechanism of regulation of enzyme biosynthesis. The mathematical model of the process served here to determine the optimal conditions for enzyme biosynthesis which were then verified in a fed-batch cultivation. The production of the enzyme in fed-batch culture was found to be twice that found in a batch cultivation.Symbols X biomass concentration, g·l^-1 - t time, h - S ₁ caseinate concentration, g·l^-1 - S ₂ starch concentration, g·l^-1 - P product concentration, U·ml^-1 - r _P specific rate of product formation, U·g^-1·h^-1 - R _P total rate of product formation, U·l^-1·h^-1 - Y yield coefficient - specific growth rate, h^-1     

Organic reduction of tapioca wastewaters using modified RBC

A modified Rotating Biological Contactor (RBC) was used for the treatability studies of synthetic tapioca wastewaters. The RBC used was a four stage laboratory model and the discs were modified by attaching porous nechlon sheets to enhance biofilm area. Synthetic tapioca wastewaters were prepared with influent concentrations from 927 to 3600 mg/l of COD. Three hydraulic loads were used in the range of 0.03 to 0.09 m³·m^–2·d^–1 and the organic loads used were in the range of 28 to 306 g COD· m^–2·d^–1. The percentage COD removal were in the range from 97.4 to 68. RBC was operated at a rotating speed of 18 rpm which was found to be the optimal rotating speed. Biokinetic coefficients based on Kornegay and Hudson models were obtained using linear analysis. Also, a mathematical model was proposed using regression analysis.List of Symbols A m² total surface area of discs - d m active depth of microbial film onany rotating disc - K _s mg ·l^–1 saturation constant - P mg·m^–2·^–1 area capacity - Q l·d^–1 hydraulic flow rate - q m³·m^–2·d^–1 hydraulic loading rate - S ₀ mg·l^–1 influent substrate concentration - S _e mg·l^–1 effluent substrate concentration - w rpm rotational speed - V m³ volume of the reactor - X _f mg·l^–1 active biomass per unit volume ofattached growth - X _s mg·l^–1 active biomass per unit volume ofsuspended growth - X mg·l^–1 active biomass per unit volume - Y _s yield coefficient for attachedgrowth - Y _A yield coefficient for suspendedgrowth - Y yield coefficient, mass of biomass/mass of substrate removed Greek Symbols hr mean hydraulic detention time - (_max)_A d^–1 maximum specific growth rate forattached growth - (_max)_s d^–1 maximum specific growth rate forsuspended growth - _max d^–1 maximum specific growth rate - d^–1 specific growth rate - _v mg·l^–1·hr^–1 maximum volumetric substrateutilization rate coefficient     

The influence of preculture on the process performance of penicillin V production in a 100-l air-lift tower loop reactor

Summary The influence of stirrer speed in the third preculture on the performance of penicillin V production by Penicillium chrysogenum in complex medium in a 100-l air-lift tower loop reactor was investigated. The process performance in the main culture was improved by increasing the stirrer speed from 500 to 750 rpm: the pellet size was reduced to half, the cell growth was influenced only slightly, but the production phase was extended considerably, and the final penicillin concentration was increased from 5.1 g 1^-1 to 10.4 g 1^-1.