Kinetic studies of acetate fermentation by Methanosarcina sp. MSTA-1

Summary The effect of three parameters (initial acetate concentration, temperature and pH) on the acetoclastic reaction was studied with the thermophilic methanogenic bacterium Methanosarcina sp. MSTA-1. The optimum temperature for growth ranged around 55° C, and optimum pH was 6.5–7.5, giving a minimum generation time of 12.6–13.9 h (µ_max = 0.050–0.055 h^–1) and a maximum value of the specific acetate consumption rate (q _infs ^supps ) of 14–20 mmol/g cells per hour. Contrary to the methane yield, the growth yield was found to be dependent on culture conditions, especially on incubation temperature. Methanosarcina sp. MSTA-1 showed a low affinity for acetate substrate. Growth at 55° C and at constant pH 7 resulted in a K _m value and a threshold acetate concentration of 10.7 mM and 0.7 mM, respectively. Offprint requests to: R. Moletta     

Acetate treatment in 70°C upflow anaerobic sludge-blanket (UASB) reactors: start-up with thermophilic inocula and the kinetics of the UASB sludges

This study focused on the use of thermophilic anaerobic granulae in the start-up of 70°C acetate-fed upflow anaerobic sludge-blanket (UASB) reactors and the kinetics of granulae grown at 70°C. In the UASB reactors, chemical oxygen demand removal commenced within 48 h of the start-up. The maximum reduction in chemical oxygen demand was 84% with the feed containing yeast and 71% without a yeast supplement. In the bioassays, the yeast-grown sludge converted 98% of the acetate consumed to methane as compared to 92% for the sludge grown without yeast. The highest initial specific methane production rate (µ_{CH 4}) of the UASB sludges grown at 70°C was 0.088 h^–1 at an acetate concentration of 4.6 mM. The higher initial acetate concentration was found to prolong the lag-phase in methane production significantly and to decrease the µ_{CH 4}. The half-saturation constant (K _s), the inhibition constant (K _i), the inhibition response coefficient (n), and the µ_{CH 4} ^max, calculated according to a modified Haldane equation, were 1.5 mM, 2.8 mM, 0.8, and 0.28 h^–1, respectively. The prolonged starvation of the 70°C sludge (15 days) decreased the µ_{CH 4} from about 0.022 h^–1 to 0.011 h^–1 and increased the lag phase in methane production from 6 h to 24 h as compared to non-starved sludge.     

Growth and lactic acid production ofPediococcus pentosaceus at different gas environments,temperatures, pH values and nitrite concentrations

Summary In order to obtain a better understanding of the behaviour ofPediococcus pentosaceus in food products as well to facilitate the designing of industrial production processes for the organism, the growth and lactic acid production ofPediococcus pentosaceus in a complex glucose medium was followed in batch cultures at different gas environments (CO₂, air, N₂ and static cultures without gasflow), temperatures (10–50°C), pH (4.3–7.3) and nitrite concentrations (0–700 ppm). Optimal growth was obtained in CO₂ at 40°C and pH 6.3 and resulted in a maximum specific growth rate ( _max) of 1.27 h^–1. In static culture at 40°C and pH 6.3 the _max was 1.21 h^–1. The _max was, compared with static culture, reduced in air (12%) and nitrogen (26%). At 10°C the _max was reduced by 99% and at 50°C by 88%. The reduction at pH 4.3 and 7.3 was 65% and 57%, respectively. Nitrite did not affect the _max at any pH but increased the lag phase at pH 4.3 by a factor of 12. The lactic acid production was linked to the growth. The total amount of lactic acid produced was the same in all the tested gases and nitrite concentrations and also within the wide temperature range (15–45°C) and pH range (5.3–7.3). Mainly L(+)-lactic acid was produced during the exponential growth phase, but after this growth declined about 30% of the L(+)-lactic acid was converted to D(–)-lactic acid. The lactic acid product yield and the cellmass yied were both affected by the temperature but not by the pH.     

Respiration of individual honeybee larvae in relation to age and ambient temperature

The CO₂ production of individual larvae of Apis mellifera carnica, which were incubated within their cells at a natural air humidity of 60–80%, was determined by an open-flow gas analyzer in relation to larval age and ambient temperature. In larvae incubated at 34 °C the amount of CO₂ produced appeared to fall only moderately from 3.89±1.57 µl mg^–1 h^–1 in 0.5-day-old larvae to 2.98±0.57 µl mg^–1 h^–1 in 3.5-day-old larvae. The decline was steeper up to an age of 5.5 days (0.95±1.15 µl mg^–1 h^–1). Our measurements show that the respiration and energy turnover of larvae younger than about 80 h is considerably lower (up to 35%) than expected from extrapolations of data determined in older larvae. The temperature dependency of CO₂ production was determined in 3.5-day-old larvae, which were incubated at temperatures varying from 18 to 38 °C in steps of 4 °C. The larvae generated 0.48±0.03 µl mg^–1 h^–1 CO₂ at 18 °C, and 3.97±0.50 µl mg^–1 h^–1 CO₂ at 38 °C. The temperature-dependent respiration rate was fitted to a logistic curve. We found that the inflection point of this curve (32.5 °C) is below the normal brood nest temperature (33–36 °C). The average Q₁₀ was 3.13, which is higher than in freshly emerged resting honeybees but similar to adult bees. This strong temperature dependency enables the bees to speed up brood development by achieving high temperatures. On the other hand, the results suggest that the strong temperature dependency forces the bees to maintain thermal homeostasis of the brood nest to avoid delayed brood development during periods of low temperature.Abbreviations m body mass - R rate of development or respiration - T_I inflexion point of a logistic (sigmoid) curve - T_L lethal temperature - T_O temperature of optimum (maximum) developmentCommunicated by G. Heldmaier     

Nitrogen and phosphorus uptake by the Brazilian kelp Laminaria abyssalis (Phaeophyta) in culture

The uptake of ammonium, nitrate and phosphate by laboratory-grown young sporophytes of Laminaria abyssalis was measured in a perturbed system (batch mode) at 18 °C and 35 ± 5 µE m^–2 s^–1 photon flux density. Uptake of all appeared to follow saturation-type nutrient uptake kinetics. The NO _inf3 ^sup– (K _s = 14.0 µM, V _max = 5.0 µmol h^–1 g^–1 dry wt) and NH _inf4 ^sup+ (K _s = 4.6 µM, V _max= 2.0 µmol h^–1 g^–1 dry wt) were taken up simultaneously, although NH _inf4 ^sup+ was taken up more rapidly. Values of K ₃ and V _max for phosphate were, respectively, 2.21 µM and 0.83 µmol h^–1 g^–1 dry wt. Nitrate and phosphate were both consumed in similar rates (V _max /Ks 0.37) at low concentrations. NH _inf4 ^sup+ , thus, might be a more efficient form of N fertilizer if artificial enrichment of seawater is used.     

Growth kinetics of Thermus thermophilus

Summary The nonsporulating extreme thermophile Thermus thermophilus was grown in continuous culture at dilution rates up to 2.65 h^–1 at 75°C and pH 6.9 on complex medium. Concomitantly very low yield (Y=0.12 g cell dry weight g^–1 utilized organic carbon) and incomplete substrate utilization (always less than 45%) were found. In batch cultures T. thermophilus could be grown with _max =h^–1, in shake flasks only with _max =h^–1 with the same low yield and incomplete substrate utilization. Stable steady states at 84C and 45°C were realized at a dilution rate of 0.3 h^–1 whereas at 86°C and 40°C no growth could be detected. Artefacts arising from wall growth (in bioreactors) or improper materials must be ruled out. Inhibition of growth by organic substrates was demonstrated at low concentrations: a decrease in the yield obtained was found when more than 0.7 gl^–1 of meat extract were supplied in the medium. The maintenance requirement for oxygen is potentially very high and was determined to be 10 to 15 mmol g^–1 h^–1.     

Nitrite uptake by Chlamydomonas reinhardtii cells immobilized in calcium alginate

When an initial cell loading of about 30–40 µg chlorophyll (Chl)·g^–1 gel and alginate suspension of 3% (w/v) were used for immobilization of Chlamydomonas reinhardtii, the resulting cell beads showed optimum nitrite uptake rate, at 30° C and pH 7.5, of 14 µmol NO _inf2 ^sup– ·mg^–1 Chl·h^–1, the photosynthetic and respiratory activities being about 120 µmol O₂ produced·mg^–1 Chl·h^–1, and 40 µmol O₂ consumed ·mg^–1 Chl·h^–1, respectively. The nitrite uptake activity required CO₂ in the culture and persisted after 8 days of cells immobilization, or in the presence of 0.2 mm ammonium in the medium. Our data indicate that alginate-entrapped C, reinhardtii cells may provide a stable and functional system for removing nitrogenous contaminants from waste-waters.Correspondence to: C. Vílchez     

Optimization and maintenance of soluble methane monooxygenase activity inMethylosinus trichosporium OB3b

Soluble methane monooxygenase (sMMO) maximization studies were carried out as part of a larger effort directed towards the development and optimization of an aqueous phase, multistage, membrane bioreactor system for treatment of polluted groundwater. A modified version of the naphthalene oxidation assay was utilized to determine the effects of methane:oxygen ratio, nutrient supply, and supplementary carbon sources on maximizing and maintaining sMMO activity inMethylosinus trichosporium OB3b.Methylosinus trichosporium OB3b attained peak sMMO activity (275–300 nmol of naphthol formed h^–1 mg of protein^–1 at 25°C) in early stationary growth phase when grown in nitrate mineral salts (NMS) medium. With the onset of methane limitation however, sMMO activity rapidly declined. It was possible to define a simplified nitrate mineral salts (NMS) medium, containing nitrate, phosphate and a source of iron and magnesium, which allowed reasonably high growth rates (_max 0.08 h^–1) and growth yields (0.4–0.5 g cells/g CH₄) and near maximal activities of sMMO. In long term batch culture incubations sMMO activity reached a stable plateau at approximately 45–50% of the initial peak level and this was maintained over several weeks. The addition of d-biotin, pyridoxine, and vitamin B₁₂ (cyanocobalamin) increased the activity level of sMMO in actively growing methanotrophs by 25–75%. The addition of these growth factors to the simplified NMS medium was found to increase the plateau sMMO level in long term batch cultures up to 70% of the original peak activity.Abbreviations sMMO soluble methane monooxygenase - pMMO particulate methane monooxygenase - NMS nitrate mineral salts - TCE trichloroethene - NADH reduced nicotinamide adenine dinucleotide     

Growth yield determination in a chemostat culture of the marine microalgaIsochrysis galbana

The growth yield of the PUFA-producing marine microalgaIsochrysis galbana ALII-4 grown in a light limited chemostat, was measured under a wide variety of conditions of incident irradiance (I _O ) and dilution rates (D). The experiments were conducted under laboratory conditions at 20 °C under continuous light. D ranged from 0.0024 to 0.0410 h^–1 at three intensities of Io (820, 1620 and 3270 µmol photon m^–2 s^–1) close to those found in outdoor cultures. A maximum efficiency _max = 0.616 g mol photon^–1 was obtained at I _O = 820 µmol photon m^–2 s^–1 and D = 0.030 h^–1 and the maximum capacity of the biomass to metabolize the light harvested was found to be 13.1 µmol photon g^–1 s^–1. Above this value, a significant drop in the system efficiency was observed. A new approach based in the averaged irradiance is used to assess the photon flux absorbed by the biomass.     

The effect of carbon dioxide on the growth kinetics of fructose-limited chemostat cultures of Acetobacterium woodii DSM 1030

The growth of the anaerobic acetogenic bacterium Acetobacterium woodii DSM 1030 was investigated in fructose-limited chemostat cultures. A defined medium was developed which contained fructose, mineral salts, cysteine · HCl and Ca pantothenate (1 mg · 1^–1) supplied in a vitamin supplement. Growth at high dilution rates was dependent on the presence of CO₂ in the gas phase. The ^max was found to be 0.16 h^–1 and the fructose maintenance requirement was 0.1 to 0.13 mmol fructose · (g dry wt)^–1 · h^–1. A growth yield of 61 g dry wt · (mol fructose)^–1, corrected for the cell maintenance requirement and for incorporation of fructose carbon into cell biomass, was determined from the fructose consumption. A corresponding growth yield of 69 g dry wt · (mol fructose)^–1 was calculated from the acetate production assuming that fructose fermentation was homoacetogenic. A Y_ATP of 12.2 to 13.8 g dry wt · (mol ATP)^–1 was calculated from these growth yields using a value of 5 mol ATP · (mol fructose)^–1 as an estimate of the amount of ATP synthesised from fructose fermentation. The addition of yeast extract (0.5 g · 1^–1) to the medium did not influence the ^max or cell yield. After prolonged growth under fructose-limited conditions the requirement of the culture for CO₂ in the gas phase was reduced.Abbreviations YE yeast extract - IC inorganic carbon - D fermenter dilution rate : h^–1 - M_X maintenance requirement for X: mmol X · (g dry wt)^–1 · h^–1 - X may be fructose (Fruct), fructose consumed in energy metabolism (Fruct [E]), acetate (Ac) - ATP CO₂, NH _inf4 ^sup+ or Pi - qX specific rate of utilisation or consumption of X: mmol X · (g dry wt)^–1 · h^–1 - V fermenter volume: litre - rC · Cell, fermenter cell carbon production: mmol C · h^–1 - Y_X yield of cells on X: g dry wt · (mol X)^–1 - Y _infx ^supmax the yield corrected for cell maintenance: g dry wt · (mol X)^–1 - S_ATP stoichiometry of ATP synthesis from fructose: mol ATP · (mol frucose)^–1 - x cell concentration: g dry wt · 1^–1 - specific growth rate : h^–1 - ^max maximum specific growth rate: h^–1     

The growth behaviour of Thermus aquaticus in continuous cultivation

Summary The growth of the yellow pigmented non-sporulating caldoactive bacterium Thermus aquaticus was investigated in different culture vessels and using differnt culture techniques. Each combination of these two variables led to very specific characteristic behaviour of the culture. A synthetic medium for a white cell type of T. aquaticus was optimized by means of pulse and medium-shift techniques. The main kinetic parameters of the organism have been determined to be =1.62h^–1 and Y (glucose)=0.4 g g^–1 at T=68 °C and pH=7.3. In complex medium only a mixed population of white and yellow cells could be cultivated. The cell yield was shown to be very low (Y=0.02 g g^–1) due to incomplete substrate utilisation, but a very high maximal specific growth rate was determined ( _max=3.5h^–1) at 75 °C and pH=7.3. The maintenance coefficient for oxygen uptake was approximately Mo=16 mMol g^–1 h^–1. A discussion of the problems arising in the cultivation of thermophilic microorganisms with respect to their physiology and stability is given.     

Phenol degradation by a psychrotrophic strain of Pseudomonas putida

Summary Cell growth and phenol degradation kinetics were studied at 10°C for a psychrotrophic bacterium, Pseudomonas putida Q5. The batch studies were conducted for initial phenol concentrations, S_o, ranging from 14 to 1000 mg/1. The experimental data for 14<=S_o<=200 mg/1 were fitted by non-linear regression to the integrated Haldane substrate inhibition growth rate model. The values of the kinetic parameters were found to be: _m=0.119 h^–1, K _S=5.27 mg/1 and K _I=377 mg/1. The yield factor of dry biomass from substrate consumed was Y=0.55. Compared to mesophilic pseudomonads previously studied, the psychrotrophic strain grows on and degrades phenol at rates that are ca. 65–80% lower. However, use of the psychrotrophic microorganism may still be economically advantageous for waste-water treatment processes installed in cold climatic regions, and in cases where influent waste-water temperatures exhibit seasonal variation in the range 10–30°C.Nomenclature K _S saturation constant (mg/l) - K _I substrate inhibition constant (mg/l) - specific growth rate (h^–1) - _m maximum specific growth rate without substrate inhibition (h^–1) - _max maximum achievable specific growth rate with substrate inhibition (h^–1) - S substrate (phenol) concentration (mg/l) - S_o initial substrate concentration (mg/l) - S_max substrate concentration corresponding to _max (mg/l) - t time (h) - X cell concentration, dry basis (mg DW/l) - X_f final cell concentration, dry basis (mg DW/l) - X_o initial cell concentration, dry basis (mg DW/l) - Y yield factor (mg DW cell produced/mg substrate consumed)     

Mass production of lipase by fed-batch culture of Pseudomonas fluorescens

Summary High concentration production of an extracellular enzyme, lipase, was achieved by a fed-batch culture of Pseudomonas fluorescens. During the cultivation, temperature, pH and dissolved oxygen concentration wwre maintained at 23°C, 6.5 and 2–5 ppm, respectively. Olive oil was used as a carbon source for microbial growth. To produce lipase effectively the specific feed rate of olive oil had to be maintained in a range of 0.04–0.06 (g oil) · (g dry cell)^-1 · h^-1. The CO₂ evolution rate was monitored to estimate the requirement of olive oil. The ratio of feed rate of olive oil to the CO₂ evolution rate was varied in the range of 20–60 g oil/mol CO₂. The higher value of the ratio accelerated microbial growth, but did not favour lipase production. Once the high cell concentration of 60 g/l had been achieved, the ratio was changed from 50 to 30 g oil/mol CO₂ to accelerate the lipase production. By this CO₂-dependent method a very high activity of lipase, 1980 units/ml, was obtained. Both the productivity and yield of lipase were prominently increased compared with a conventional batch culture.     

Effects of growth temperature on maximal specific growth rate,yield, maintenance,and death rate in glucose-limited continuous culture of the thermophilic Bacillus caldotenax

Summary The influence of temperature on the growth of the theromophilic Bacillus caldotenax was investigated using chemostat techniques and a chemically defined minimal medium. All determined growth constants, that is maximal specific growth rate, yield and maintenance, were temperature dependent. It was striking that the very large maintenance requirement was about 10 times higher than for mesophilic cells under equivalent conditions. A death rate, which was very substantial at optimal and supraoptimal growth temperatures, was estimated by comparing the maintenance for substrate and oxygen. There was no indication for a thermoadaptation as postulated by Haberstich and Zuber (1974).Symbols D Dilution rate (h^–1) - D_c=_max Critical dilution rate (h^–1) - E Temperature characteristic (J mol^–1) - k Organism constant - k_d Death rate coefficient (h^–1) - k_m Maintenance substrate coefficient estimated from M_O (h^–1) - M_O Maintenance respiration, mmol O₂ per g dry biomass and h (mmol g^–1h^–1) - M_O Maintenance respiration, taking k_d into account - m_S Maintenance substrate coefficient, g glucose per g dry biomass and h (h^–1) - OD Optical density at 546 nm - QO₂ Specific O₂-uptake rate (mmol g^–1h^–1) - Q _O2 ^V Specific O₂-uptake rate for viable portion of biomass (mmol g^–1 h^–1) - Q_S Specific glucose uptake rate (h^–1) - Q _S ^V Specific glucose uptake rate for viable portion of biomass (h^–1) - R Gas constant 8.28 J mol^–1K^–1 - S Substrate concentration in reactor (g l^–1) - S_O Influent substrate concentration (g l^–1) - T_max Maximal growth temperature (°C) - T_min Minimal growth temperature (°C) - X Dry biomass (g l^–1) - X_tOt=X Dry biomass containing dead and viable cells - X_v Viable portion of biomass - Y _O ^m Potential yield for O₂ corrected for maintenance respiration (g mol^–1) - Y _S ^m Potential yield for substrate corrected for maintenance requirement, g biomass per g glucose (–) - Specific growth rate (h^–1) - _max Maximal specific growth rate (h^–1)     

Planktonic bacterial biomass and seasonal pattern of the heterotrophic uptake and respiration of glucose and amino acids in the shallow sandpit lake of Créteil (Paris Suburb,France)

Biomass and activity of planktonic bacteria were investigated during a one year study in a shallow sandpit lake. The shallowness of the lake helped keep the water column homogeneous regarding bacterioplankton. Small free-living bacteria (0.03 µm³ cell^–1) dominated the populations throughout the period studied. Bacterial abundances varied from 1 to 11 × 10⁶ cells ml^–1. Kinetic parameters (V _max, K + S and T) were determined with ¹⁴C labelled compounds (glucose and amino acids mixture). V _max values were high and averaged 0.056 and 0.050 µgCl^–1 h^–1 for glucose and amino acids respectively. Maximal V _max values were observed in summer at the highest temperatures, but also in early spring. T values were much greater in winter. K + S values were significantly higher for amino acids (3 µg Cl^–1) than for glucose (1 µg Cl^–1). A low percentage of mineralization (about 25% for both tracers) could be the expression of the high growth efficiency expected when bacteria are growing at the expense of low molecular weight compounds as phytoplankton exudates.     

Purification and characterization of a high molecular mass serine carboxypeptidase from <Emphasis Type="Italic">Monascus pilosus</Emphasis>

Two serine carboxypeptidases, MpiCP-1 and MpiCP-2, were purified to homogeneity from Monascus pilosus IFO 4480. MpiCP-1 is a homodimer with a native molecular mass of 125 kDa composed of two identical subunits of 61 kDa, while MpiCP-2 is a high mass homooligomer with a native molecular mass of 2,263 kDa composed of about 38 identical subunits of 59 kDa. This is unique among carboxypeptidases and distinguishes MpiCP-2 as the largest known carboxypeptidase. The two purified enzymes were both acidic glycoproteins. MpiCP-1 has an isoelectric point of 3.7 and a carbohydrate content of 11%, while for MpiCP-2 these values were 4.0 and 33%, respectively. The optimum pH and temperature were around 4.0 and 50°C for MpiCP-1, and 3.5 and 50°C for MpiCP-2. MpiCP-1 was stable over a broad range of pH between 2.0 and 8.0 at 37°C for 1 h, and up to 55°C for 15 min at pH 6.0, but MpiCP-2 was stable in a narrow range of pH between 5.5 and 6.5, and up to 50°C for 15 min at pH 6.0. Phenylmethylsulfonylfluoride strongly inhibited MpiCP-1 and completely inhibited MpiCP-2, suggesting that they are both serine carboxypeptidases. Of the substrates tested, benzyloxycarbonyl-l-tyrosyl-l-glutamic acid (Z-Tyr-Glu) was the best for both enzymes. The K_m, V_max, K_cat and K_cat/K_m values of MpiCP-1 for Z-Tyr-Glu at pH 4.0 and 37°C were 1.33 mM, 1.49 mM min^–1, 723 s^–1 and 545 mM^–1 s^–1, and those of MpiCP-2 at pH 3.5 and 37°C were 1.55 mM, 1.54 mM min^–1, 2,039 s^–1 and 1,318 mM^–1 s^–1, respectively.     

Modeling of the pyruvate production with<Emphasis Type="Italic"> Escherichia coli</Emphasis> in a fed-batch bioreactor

A family of 10 competing, unstructured models has been developed to model cell growth, substrate consumption, and product formation of the pyruvate producing strain Escherichia coli YYC202 ldhA::Kan strain used in fed-batch processes. The strain is completely blocked in its ability to convert pyruvate into acetyl-CoA or acetate (using glucose as the carbon source) resulting in an acetate auxotrophy during growth in glucose minimal medium. Parameter estimation was carried out using data from fed-batch fermentation performed at constant glucose feed rates of q_VG=10 mL h^–1. Acetate was fed according to the previously developed feeding strategy. While the model identification was realized by least-square fit, the model discrimination was based on the model selection criterion (MSC). The validation of model parameters was performed applying data from two different fed-batch experiments with glucose feed rate q_VG=20 and 30 mL h^–1, respectively. Consequently, the most suitable model was identified that reflected the pyruvate and biomass curves adequately by considering a pyruvate inhibited growth (Jerusalimsky approach) and pyruvate inhibited product formation (described by modified Luedeking–Piret/Levenspiel term).List of symbols c_A acetate concentration (g L^–1) - c_A,0 acetate concentration in the feed (g L^–1) - c_G glucose concentration (g L^–1) - c_G,0 glucose concentration in the feed (g L^–1) - c_P pyruvate concentration (g L^–1) - c_P,max critical pyruvate concentration above which reaction cannot proceed (g L^–1) - c_X biomass concentration (g L^–1) - K_I inhibition constant for pyruvate production (g L^–1) - K_I^A inhibition constant for biomass growth on acetate (g L^–1) - K_P saturation constant for pyruvate production (g L^–1) - K_P inhibition constant of Jerusalimsky (g L^–1) - K_S^A Monod growth constant for acetate (g L^–1) - K_S^G Monod growth constant for glucose (g L^–1) - m_A maintenance coefficient for growth on acetate (g g^–1 h^–1) - m_G maintenance coefficient for growth on glucose (g g^–1 h^–1) - n constant of extended Monod kinetics (Levenspiel) (–) - q_V volumetric flow rate (L h^–1) - q_VA volumetric flow rate of acetate (L h^–1) - q_VG volumetric flow rate of glucose (L h^–1) - r_A specific rate of acetate consumption (g g^–1 h^–1) - r_G specific rate of glucose consumption (g g^–1 h^–1) - r_P specific rate of pyruvate production (g g^–1 h^–1) - r_P,max maximum specific rate of pyruvate production (g g^–1 h^–1) - t time (h) - V reaction (broth) volume (L) - Y_P/G yield coefficient pyruvate from glucose (g g^–1) - Y_X/A yield coefficient biomass from acetate (g g^–1) - Y_X/A,max maximum yield coefficient biomass from acetate (g g^–1) - Y_X/G yield coefficient biomass from glucose (g g^–1) - Y_X/G,max maximum yield coefficient biomass from glucose (g g^–1) - growth associated product formation coefficient (g g^–1) - non-growth associated product formation coefficient (g g^–1 h^–1) - specific growth rate (h^–1) - _max maximum specific growth rate (h^–1)     

Enzyme formation by a yeast cell wall lytic Arthrobacter sp.: formation of \(1,3)-glucanase

The kinetics of \-1,3-glucanase (EC 3.2.1.39; 1,3-\-d-glucan-glucano-hydrolase) formation by a yeast cell wall lytic Arthrobacter species was studied. Yeast glucan as a substrate yielded 360 units (U)/l, but it appeared to be unsuitable for fermentation purposes because of its insolubility and its residual content of glycogen. Growth on water-soluble \(1,3)-glucan [maximum specific growth rate (µ_max)=0.19 h^–1] was governed by different saccharides liberated by enzyme action on glucan. Enzyme formation was repressed by glucose and derepressed by its restricted availability during late exponential and stationary growth. At least 380 U/l of \(1,3)-glucanase were formed. Lactose and lactulose were detected as precursors of potent inducers for \(1,3)-glucanase, the first being a cheap and easily available substrate for large-scale cultivations. Growth rates were reduced (µ_max=0.18 h^–1 and µ_max=0.13 h^–1, respectively), enzyme synthesis occurred only during post-logarithmic growth. The \(1,3)-glucanase levels (260 U/l) formed were comparable to that attained with glucan as a substrate. In continuous culture no enzyme was formed under steady-state conditions but it occurred during transient states after shifting the dilution rate to lower values. Correspondence to: W. Hampel     

Studies on the variables and kinetics of SCP production from nopal fruit juice

Summary Submerged batch cultivation under controlled environmental conditions of pH 3.8, temperature 30°C, and K_La200 h^–1 (above 180 mMO₂ l ^–1 h^–1 oxygen supply rate) produced a maximum (12.0 g·l ^–1) SCP (Candida utilis) yield on the deseeded nopal fruit juice medium containing C/N ratio of 7.0 (initial sugar concentration 25 g·l ^–1) with a yield coefficient of 0.52 g cells/g sugar. In continuous cultivation, 19.9 g·l ^–1 cell mass could be obtained at a dilution rate (D) of 0.36 h^–1 under identical environmental conditions, showing a productivity of 7.2 g·l ^–1·h^–1. This corresponded to a gain of 9.0 in productivity in continuous culture over batch culture. Starting with steady state values of state variables, cell mass (C_X–19.9 g·l ^–1), limiting nutrient concentration (C_ln–2.5 g·l ^–1) and sugar concentration (C_S–1.5 g·l ^–1) at control variable conditions of pH 3.8, 30°C, and K_La 200 h^–1 keeping D=0.36 h^–1 as reference, transient response studies by step changes of these control variables also showed that this pH, temperature and K_La conditions are most suitable for SCP cultivation on nopal fruit juice. Kinetic equations obtained from experimental data were analysed and kinetic parameters determined graphically. Results of SCP production from nopal fruit juice are described.Nomenclature C_ln concentration of ammonium sulfate (g·l ^–1) - C_S concentration of total sugar (g·l ^–1) - C_X cell concentration (g·l ^–1) - D dilution rate (h^–1) - K_ln Monod's constant (g·l ^–1) - m maintenance coefficient (g ammonium sulfate cell^–1 h^–1) - m_(S) maintenance coefficient (g sugar g cell^–1 h^–1) - t time, h - Y yield coefficient (g cells/g ammonium sulfate) - Y_m maximum of Y - Y_S yield coefficient based on sugar consumed (g cells · g sugar^–1) - Y_S(m) maximum value of Y_S - ^µm maximum specific growth rate constant (h^–1)     

Growth and physiology of potassium-limited chemostat cultures of Paracoccus denitrificans

In potassium-limited chemostat cultures of Paracoccus denitrificans the maximum specific growth rate (µ_max) was found to depend on the input potassium concentration: At 0.21mM µ_max was 0.10–0.11 h^-1; at 0.44 mM 0.15–0.16 h^-1 and at 0.66 mM 0.20–0.21 h^-1. The plots of the specific rates of oxygen-, succinate-and potassium consumption against gave straight lines. The intracellular potassium concentration was a linear function of and varied from 1% (0.13 M) at a value of 0.034 h^-1 to 2.2% (0.29 M) at =0.26 h^-1; the potassium concentration gradient and the potassium concentration in the culture fluid in the steady state were dependent on the input potassium concentration. The potassium concentration gradient varied from 8,900-1,200. At all values 20–25% of the total energy production was used for potassium transport. 350,100 and 30 ATP molecules were calculated to be required to maintain one potassium ion intracellular during 1 h at values of 0.034, 0.197 and 0.257 h^-1 respectively. It is concluded that the amount of circulation of potassium is dependent on the potassium concentration gradient or on the potassium concentration in the culture in the steady state. The dependency of µ_max on the input potassium concentration was explained by the assumption that at low input potassium concentrations the net uptake of potassium (influx-efflux) is not rapidly enough to maintain the high potassium gradient in the existing cells and to establish it in the newly formed cells. At high values and at high input potassium concentrations µ_max is limited by the specific rate of oxygen consumption, which was found to be 11–12 mmol O₂ g dry weight^-1 h^-1 at µ_max for potassium-, succinate-and sulphate-limited chemostat cultures.