Influence of oxygen concentration on the metabolism of Penicillium chrysogenum

In large-scale bioreactors, there is often insufficient mixing and as a consequence, cells experience uneven substrate and oxygen levels that influence product formation. In this study, the influence of dissolved oxygen (DO) gradients on the primary and secondary metabolism of a high producing industrial strain of Penicillium chrysogenum was investigated. Within a wide range of DO concentrations, obtained under chemostat conditions, we observed different responses from P. chrysogenum: (i) no influence on growth or penicillin production (>0.025 mmol L⁻¹); (ii) reduced penicillin production, but no growth limitation (0.013–0.025 mmol L⁻¹); and (iii) growth and penicillin production limitations (<0.013 mmol L⁻¹). In addition, scale down experiments were performed by oscillating the DO concentration in the bioreactor. We found that during DO oscillation, the penicillin production rate decreased below the value observed when a constant DO equal to the average oscillating DO value was used. To understand and predict the influence of oxygen levels on primary metabolism and penicillin production, we developed a black box model that was linked to a detailed kinetic model of the penicillin pathway. The model simulations represented the experimental data during the step experiments; however, during the oscillation experiments the predictions deviated, indicating the involvement of the central metabolism in penicillin production.     

Hypoxia tolerance of the mummichog: the role of access to the water surface

Low dissolved oxygen (DO) had a significant effect on specific growth rate (G_S), length increment (I_L) and haematocrit (Hct) of the mummichog Fundulus heteroclitus. Regardless of access to the water surface, F. heteroclitus maintained high growth rates (G_S and I_L) at DO concentrations as low as 3 mg O₂ l^?1. With access to the water surface, both G_S and I_L of F. heteroclitus decreased by c. 60% at 1·0 mg O₂ l^?1 compared to all higher DO treatments. When denied access to the water surface, a further decrease in G_S (c. 90%) and I_L (c. 75%) was observed at 1 mg O₂ l^?1. There was no effect of diel‐cycling DO (1–11 mg O₂ l^?1) with or without surface access on G_S, I_L or Hct of F. heteroclitus. Similar trends between G_S and faecal production across DO treatments suggest that decreased feeding contributed significantly to the observed decrease in growth rate. Haematocrit was significantly elevated at 1 mg O₂ l^?1 for fish with and without access to the water surface. Increased Hct, however, was not sufficient to maintain high G_S or I_L at severely low DO. When permitted to respire in the surface layer, however, F. heteroclitus was capable of maintaining moderate growth rates at DO concentrations of 1 mg O₂ l^?1 (c. 15% saturation). Although aquatic surface respiration (ASR) was not quantified in this study, F. heteroclitus routinely swam in contact with the water surface and performed ASR at DO concentrations ≤3 mg O₂ l^?1. No hypoxia‐related mortality was observed in any DO or surface access treatment for as long as 9 days. This study demonstrates that surface access, and thus potential for ASR, plays an important role in providing F. heteroclitus substantial independence of growth rate over a wide range of low DO conditions commonly encountered in shallow estuarine environments.     

Behavioral responses of red hake,Urophycis chuss,to decreasing concentrations of dissolved oxygen

Synopsis Laboratory experiments were conducted to examine changes in behavior of red hake,Urophycis chuss, under decreasing concentrations of dissolved oxygen (DO). Since the ecological requirements of this species change with age, responses were measured for three different groups: (1) age 0+, = 89 mm total length (TL); (2) age 1+, = 238 mm TL; and (3) age 2–3+, = 397 mm TL. As DO decreased from 8–10 mg l^-1 to < 0.5 mg l^-1, changes were evident in active time, water column activity, range of horizontal movement, food searching, and agonistic behavior. Age 0+ fish were most sensitive, moving up into the water column and swimming continuously as DO levels fell below 4.2 mg l^-1. Age 2–3+ fish were the least responsive, remaining on the substrate and increasing only their range of movement at concentrations below 3 mg l^-1. Responses of age 1 + fish were variable, possibly reflecting a transition stage between the younger and older fish. Common to all groups was the decrease and eventual cessation of food searching.     

Ventilatory and behavioural responses of the marbled sole Pseudopleuronectes yokohamae to progressive hypoxia

This study identified ventilatory and behavioural responses in the marbled sole Pseudopleuronectes yokohamae under experimentally induced progressive decreases in dissolved oxygen (DO) levels. Ventilation frequency showed an increase with decreasing DO levels from normoxia to 2·75 mg O₂ l^?1, followed by a decrease in ventilation frequency at decreased DO levels from 2·00 to 0·75 mg O₂ l^?1. At DO levels below 2·00 mg l^?1, behaviours at the bottom were suppressed, whereas avoidance behaviours increased. A decrease in avoidance behaviours was observed from 1·00 to 0·75 mg O₂ l^?1. Upside‐down reversal and incapacitation at DO levels of 1·00–0·75 mg O₂ l^?1 suggested that sublethal effects on P. yokohamae were induced. The responses observed before the sublethal DO level could be interpreted as an effort to maintain oxygen uptake, reduce routine activities and facilitate avoidance. The observed DO level thresholds that induce behavioural responses, in addition to sublethal effects, indicate hypoxia‐tolerance that is important for understanding the effects of hypoxia on coastal ecosystems.     

Alginate production by Azotobacter vinelandii DSM576 in batch fermentation

The kinetics of growth and alginate production from glucose in a nitrogen and phosphate-rich medium by Azotobacter vinelandii DSM576 were studied in a laboratory fermenter at pH 7 and 35°C. Batch fermentations were carried out both without control of dissolved oxygen concentration (DO) and at 1, 2, 5 and 10% DO. Although growth was faster at higher DO, maximum biomass concentration was lower. No alginate was produced at 10% DO. Alginate production was faster at 5 and 2% DO but higher alginate concentrations and yields were obtained without DO control. Alginate production was growth-associated at 5% DO, but significant amounts of alginate were produced after growth had stopped at lower DO values. In fermentations without DO control the molecular weight of the polymer reached a maximum (11–17.6 × 10⁴) when specific growth rate was between 0.02 and 0.04 h⁻¹ and residual concentration of ammoniacal nitrogen was between 0.01 and 0.02 g L⁻¹ and then sharply decreased. Received 15 August 1997/ Accepted in revised form 08 January 1998     

The effect of dissolved oxygen,sediment, and sewage treatment plant discharges upon growth,survival and density of Asiatic clams

The biology of Corbicula fluminea, the Asiatic clam, in the Vermilion River, Louisiana, as affected by sediment, dissolved oxygen (DO) levels, and sewage treatment plant (STP) effluents was investigated. A point source of high DO water to the Vermilion River established a gradient of DO that decreased as the river moved towards the Gulf of Mexico. Lowering DO levels were exacerbated by municipal sewage treatment plant discharges in the 20 km reach studied. Low dissolved oxygen was associated with reduced Corbicula density in the river and 30-day in-stream growth studies (weight and length) demonstrated that low DO inhibited growth. Generally, if DO was < 1.0 mg l^–1 in sediment pore water and/or < 3.0 mg l^–1 at the sediment-water interface, growth was significantly impaired (p < 0.05). Corbicula experienced substantial mortality near the STP discharges (up to 70% in 30 days) and laboratory toxicity tests with Ceriodaphnia dubia, a sensitive cladoceran, also strongly suggested discharges were chronically toxic at 6.25–25.0% effluent. Respiration experiments along with environmental measurements of DO, temperature, and STP discharge chemistry support a hypothesis that clam populations are adversely affected by the suite of environmental conditions present in the Vermilion River. Further, growth studies were consistent with observed population densities in situ.     

Characterizing the escape response of juvenile summer flounder Paralichthys dentatus to diel‐cycling hypoxia

Swimming speed, angular correlation and expected displacement were measured in juvenile summer flounder Paralichthys dentatus acclimated to either oxygen saturation (c. 7·8 mg O₂ l^?1; saturation‐acclimated fish) or diel‐cycling hypoxia (cycling between 11·0 and 2·0 mg O₂ l^?1) for 10 days and subsequently exposed to more severe diel‐cycling hypoxia (cycling between 7·0 and 0·4 mg O₂ l^?1). Saturation‐acclimated P. dentatus exhibited an active response to declining dissolved oxygen (DO) by increasing swimming speed, angular correlation and expected displacement to peak levels at 1·4 mg O₂ l^?1 that were 3·5, 5·5 and 4·2 fold, respectively, greater than those at DO saturation. Diel‐cycling hypoxia‐acclimated P. dentatus also exhibited an active response to declining DO, although it was relatively less pronounced. Diel‐cycling hypoxia‐acclimated P. dentatus swimming speed, however, still doubled as DO decreased from 7·0 to 2·8 mg O₂ l^?1. Diel‐cycling hypoxia‐acclimated P. dentatus did not recover as well from low DO exposure as did saturation‐acclimated fish. This was reflected in their relatively more random swimming (low angular correlation between successive moves) and poor maintenance of rank order between individuals during the recovery phase. Even saturation‐acclimated P. dentatus did not resume swimming at speeds observed at saturation until DO was 4·2 mg O₂ l^?1. Paralichthys dentatus were very sensitive to decreasing DO, even at DO levels that were not lethal or growth limiting. This sensitivity and their poor recovery may preclude juvenile P. dentatus from using highly productive nursery habitats affected by diel‐cycling hypoxia.     

Improvement of Photorhabdus temperata bioinsecticides production in low‐cost media through adequate fermentation technology

To develop a cost effective process for bioinsecticides production by Photorhabdus temperata, dissolved oxygen (DO) requirements were investigated in both the complex and the optimized media using diluted seawater as a source of micronutrients. By varying DO concentrations, tolerance to hydrogen peroxide was shown to be medium dependant. Indeed, P. temperata cells grown in the complex medium, exhibited higher tolerance than cells grown in the optimized medium (OM). Tolerance to H₂O₂ was shown to be related to intracellular reactive oxygen species (ROS) accumulation during soya bean meal or glucose assimilation, as shown by flow cytometry analysis. To avoid oxidative stress damages in P. temperata cells cultured in the OM, DO concentration should be constant 50% saturation throughout the fermentation. However, a DO‐shift control strategy was demonstrated to be beneficial for P. temperata bioinsecticide production in the complex medium. By using such a strategy biomass, culturability, and oral toxicity reached 16.5 × 10⁸, 1.15 × 10⁸ cells/mL and 64.2%, respectively, thus was 16.19, 26.37, and 12.2% more than in the cultures carried out at a constant 50% saturation. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012     

Partial nitrification to nitrite using low dissolved oxygen concentration as the main selection factor

Partial nitrification to nitrite (nitritation) can be achieved in a continuous process without sludge retention by wash out of nitrite oxidising bacteria (NOB) while retaining ammonia oxidising bacteria (AOB), at elevated temperatures (the SHARON process) and, as demonstrated in this paper, also at low dissolved oxygen (DO) concentrations. Enriched AOB was attained at a low DO concentration (0.4 mg l⁻¹) and a dilution rate of 0.42 day⁻¹ in a continuous process. A higher oxygen affinity of AOB compared to NOB seemed critical to achieving this. This was verified by determining the oxygen half saturation constant, K _o, with similar oxygen mass transfer resistances for enriched AOB and NOB as 0.033 ± 0.003 mg l⁻¹ and 0.43 ± 0.08 mg l⁻¹, respectively. However, the extent of nitritation attained was found to be highly sensitive to process upsets.     

Role of oxygen in the utilization of phenol by Pseudomonas CF600 in continuous culture

The dissolved oxygen (DO) level is the key factor which decides the rate of degradation of the organic load in aerobic growth conditions. In this study the role of DO levels on the utilization of phenols has been reported using the continuous culture system. A phenol-utilizing strain, Pseudomonas CF600 has been used as a model. Its phenol-degrading capacity was studied using continuous cultivation for a period of 60 days. The bioreactor was kept at a dilution rate of 0.006 h^–1 with DO levels maintained at 2, 3, and 4 ppm keeping all the other cultivation conditions constant. Physiological variations under the cultivation conditions were studied by monitoring off-line phenol utilization and respirometric analysis of harvested culture against different substrates. It was observed that the accumulation of 2-hydroxymuconate semialdehyde (HMS), an intermediate in the phenol degradation pathway, depends on the DO level. The maximum level of HMS in the medium observed was 3.92 M when DO was maintained at 2 ppm whereas with 3 ppm of DO, HMS level was below 0.4 M. Oxygen uptake data of the cells harvested from cultures grown at different DO levels showed that the uptake was highest at 3 ppm DO for all the substrates tried. When phenol was used as substrate, the oxygen uptake rate was 42.66, 66.36 and 35.55 nM/min/mg dry weight of cells at 4, 3 and 2 ppm DO respectively. Results show that DO levels influence the rate of phenol utilization in Pseudomonas CF600.     

Influence of agitation and aeration on growth and antibiotic production by <Emphasis Type="Italic">Xenorhabdus nematophila</Emphasis>

The effect of agitation and aeration on the growth and antibiotic production by Xenorhabdus nematophila YL001 grown in batch cultures were investigated. Efficiency of aeration and agitation was evaluated through the oxygen mass transfer coefficient (K _L a). With increase in K _L a, the biomass and antibiotic activity increased. Activity units of antibiotic and dry cell weight were increased to 232 U ml⁻¹ and 19.58 g l⁻¹, respectively, productivity in cell and antibiotic was up more than 30% when K _L a increased from 115.9 h⁻¹ to 185.7 h⁻¹. During the exponential growth phase, DO concentration was zero, the oxygen supply was not sufficient. So, based on process analysis, a three-stage oxygen supply control strategy was used to improved the DO concentration above 30% by controlling the agitation speed and aeration rate. The dry cell weight and activity units of antibiotic were further increased to 24.22 g l⁻¹ and 249 U ml⁻¹, and were improved by 24.0% and 7.0%, compared with fermentation at a constant agitation speed and a constant aeration rate (300 rev min⁻¹, 2.5 l min⁻¹).     

Oxygen consumption and ammonia accumulation in the hypolimnion of Walker Lake, Nevada

Walker Lake (area = 140 km², Z _mean = 19.3 m) is a large, terminal lake in western Nevada. As a result of anthropogenic desiccation, the lake has decreased in volume by 75% since the 1880s. The hypolimnion of the lake, now too small to meet the oxygen demand exerted by decaying matter, rapidly goes anoxic after thermal stratification. Field and laboratory studies were conducted to examine the feasibility of using oxygenation to avoid hypolimnetic anoxia and subsequent accumulation of ammonia in the hypolimnion, and to estimate the required DO capacity of an oxygenation system for the lake. The accumulation of inorganic nitrogen in water overlaying sediment was measured in laboratory chambers under various DO levels. Rates of ammonia accumulation ranged from 16.8 to 23.5 mg-N m^–2 d^–1 in chambers with 0, 2.5 and 4.8 mg L^–1 DO, and ammonia release was not significantly different between treatments. Beggiatoa sp. on the sediment surface of the moderately aerated chambers (2.5 and 4.8 mg L^–1 DO) indicated that oxygen penetration into sediment was minimal. In contrast, ammonia accumulation was reversed in chambers with 10 mg L^–1 DO, where oxygen penetration into sediment stimulated nitrification and denitrification. Ammonia accumulation in anoxic chambers (18.1 and 20.6 mg-N m^–2 d^–1) was similar to ammonia accumulation in the hypolimnion from July through September of 1998 (16.5 mg-N m^–2 d^–1). Areal hypolimnetic oxygen demand averaged 1.2 g O₂ m^–2 d^–1 for 1994–1996 and 1998. Sediment oxygen demand (SOD) determined in experimental chambers averaged approximately 0.14 g O₂ m^–2 d^–1. Continuous water currents at the sediment-water interface of 5–6 cm s^–1 resulted in a substantial increase in SOD (0.38 g O₂ m^–2 d^–1). The recommended oxygen delivery capacity of an oxygenation system, taking into account increased SOD due to mixing in the hypolimnion after system start-up, is 215 Mg d^–1. Experimental results suggest that the system should maintain high levels of DO at the sediment-water interface (10 mg L^–1) to insure adequate oxygen penetration into the sediments, and a subsequent inhibition of ammonia accumulation in the hypolimnion of the lake.     

Response surface optimization of dissolved oxygen and nitrogen sources for the biodegradation of MTBE and BTEX

Response surface methodology (RSM) using central composite design was applied to obtain the optimal dissolved oxygen (DO) and nitrogen (N) concentrations for biodegrading MTBE (Methyl tert-butyl ether) and BTEX (benzene, ethylbenzene, toluene, p-xylene). Moreover, the effects of DO, N, and their interaction on the degradation process were evaluated. It was found that N, N², DO and DO² have significant effects on the efficiency of MTBE and BTEX removal. The removal efficiency when using biostimulation with bioaugmentation (BwB) is higher than with other processes, being greater than 82% at concentrations of 12 and 48 mg l⁻¹ for DO and N, respectively. However, it was also found that the interaction term of DO × N has no significant effect on the degradation processes.     

The effects of progressive hypoxia and re-oxygenation on cardiac function, white muscle perfusion and haemoglobin saturation in anaesthetised snapper (Pagrus auratus)

The effects of progressive hypoxia and re-oxygenation on cardiac function, white muscle perfusion and haemoglobin saturation were investigated in anaesthetised snapper (Pagrus auratus). White muscle perfusion and haemoglobin saturation were recorded in real time using fibre optic methodology. A marked fall in heart rate (HR) was evoked when the water bath dissolved oxygen (DO) concentration decreased below 1.5 mg L⁻¹. This bradycardia deepened over the subsequent 20 min of progressive hypoxia and noticeable arrhythmias occurred, suggesting that hypoxia had direct and severe effects on the cardiac myocytes. Perfusion to the white muscle decreased below a DO concentration of 3 mg L⁻¹, and oxyhaemoglobin concentration decreased once the DO fell below ca. 2 mg L⁻¹. During re-oxygenation, heart rate and white muscle perfusion increased as the DO concentration exceeded 1.9 ± 0.1 mg L⁻¹, whereas haemoglobin saturation increased once the external DO concentration reached 2.9 mg L⁻¹. These changes occurred in anaesthetised fish, in which sensory function must be impaired, if not abolished. As white muscle perfusion both fell and increased prior to changes in white muscle oxyhaemoglobin saturation, a local hypoxia is more likely to be the consequence than the cause of the reduced blood delivery, and changes upstream from the tail vasculature must be responsible. HR and tissue haemoglobin concentrations did increase simultaneously on re-oxygenation suggesting an increased cardiac output as the cause.     

Continous ethyl acetate production by Kluyveromyces fragilis on whey permeate

The synthesis of ethyl acetate by Kluyveromyces fragilis on diluted whey permeate was studied. Ethanol, lactose and O₂ are the direct precursors for ethyl acetate synthesis by this yeast. Ethyl acetate production is affected by many parameters, particularly the carbon/nitrogen (C/N) ratio, Tween 80 and iron. Ethyl acetate synthesis is optimum for C/N = 45. Tween 80 lowered slightly the level of ethyl acetate whereas iron completely stopped ethyl acetate production. The level of ethanol in the feed, the dissolved O₂ (DO) and dilution rate (D) were also optimised. Thus at D = 0.24 h ^–1, for 4 g/l of ethanol in the feed and 40% DO, the productivity of ethyl acetate was optimal (0.7 g/l per hour). Correspondence to: A. Miclo     

Renal O2 consumption during progressive hemorrhage

Most mammalian tissues regulate O2 utilization such that O2 consumption (VO2) is relatively constant at O2 delivery (DO2) higher than a critical value (DO2c). We studied the relationship between VO2 and DO2 of kidney and whole body during graded progressive exsanguination. The relationship between whole body VO2 and DO2 was biphasic, and whole body VO2 decreased by 5.6 +/- 14.4% (P = NS) from the initial value to the value nearest whole body DO2c. Kidney DO2 decreased in direct proportion to whole body DO2 such that the average R2 value describing the linear regression of kidney DO2 vs. whole body DO2 was 0.94 +/- 0.02. The relationship between kidney, like whole body, VO2 and DO2 appeared biphasic; however, kidney VO2 decreased by 63.3 +/- 10.4% (P less than 0.0001) from the initial value to the value nearest kidney DO2c. Renal O2 extraction ratio was relatively constant over a wide range of kidney DO2, whereas whole body O2 extraction ratio increased progressively at all whole body DO2 values as whole body DO2 decreased. However, final values of O2 extraction ratio were indistinguishable for whole body (0.86 +/- 0.1) and kidney (0.86 +/- 0.06) (P = NS). We conclude that the pattern of kidney and whole body VO2 response to decreasing DO2 differs during hemorrhage, particularly in the range of DO2 normally associated with tissue wellness.     

Effect of pH,aeration and sucrose feeding on the invertase activity of intactS. cerevisiae cells grown in sugarcane blackstrap molasses

S. cerevisiae was grown in a blackstrap molasses containing medium in batch and fed-batch cultures. The following parameters were varied: pH (from 4.0 to 6.5), dissolved oxygen (DO) (from 0 to 5.0 mg O₂L^–1) and sucrose feeding rate. When glucose concentration (S) was higher than 0.5 g L^–1 a reduction in the specific invertase activity of intact cells (v) and an oscillatory behavior of v values during fermentation were observed. Both the invertase reduction and the oscillatory behavior of v values could be related to the glucose inhibitory effect on invertase biosynthesis. The best culture conditions for attainingS. cerevisiae cells suitable for invertase production were: temperature=30°C; pH=5.0; DO=3.3 mg O₂L^–1; (S)=0.5 g L^–1 and sucrose added into the fermenter according to the equations: (V–V_o)=t²/16 or (V–V_o)=(V_f–V_o)·(e^0.6t–1)/10.This work was supported by FAPESP     

Effect of NaCl on the association of thrombin with heparin

The association of heparin with thrombin was investigated by fluorometric titration. A maximum of 25% of the fluorescence of fluorescein-labeled heparin (FTC-heparin) was quenched at thrombin saturation in the absence of NaCl. FTC-heparin (H) associated tightly with thrombin (T) and the association constant of the ternary complex, H₂T, formed in the absence of NaCl, was calculated to be 1.7 × 10⁸M^?1. However, the association was strongly influenced by the NaCl concentration, and the association constant of the equimolar complex, HT, formed in 0.15M NaCl was found to be 1 × 10⁶M^?1. The first-order rate constant, k_app, for inactivation of thrombin by antithrombin III (AT III) and low-affinity heparin (LA-heparin) was comparable with that of high-affinity heparin (HA-heparin) in the absence of NaCl, but decreased with an increase in the concentration of NaCl. The decreased enhancement of the thrombin-AT III reaction by LA-heparin at high NaCl concentration appeared to result from a decreased association of thrombin with LA-heparin, thus reducing the formation of the ternary complex, thrombin-LA-heparin-AT III.     

Continuous fermentation to produce fungal protein. Effect of growth rate on the biomass yield and chemical composition of Fusarium moniliforme

Fusarium moniliforme was grown on a carob aqueous extract in a chemostat for fungal protein production. The substrate was adjusted to provide 0.5% carob sugars supplemented with inorganic salts. The dilution rate varied from 0.086 to 0.227 hr^?1 under constant conditions of temperature (30°C), pH (4.5), and oxygen saturation (60–80%). A yield of 0.709 g dry mycelium/g consumed carob sugar and a productivity value of 0.687 g dry mycelium/liter hr^?1 were obtained at μ = 0.205 hr^?1. The maintenance coefficient was 0.077 g carob sugar/g dry mycelium hr^?1. While the carbohydrate and purine content of dry mycelium increased at μ values from 0.114 to 0.205 hr^?1 both true (Lowry) and crude (N × 6.25) protein contents decreased at the same μ range. Maximum values of 36.3% true and 47.9% crude protein of dry mycelium were obtained at μ = 0.114 hr^?1, whereas a minimum purine content of 99.8 μmol/g corresponding to 6.42% nucleic acids was recorded at μ = 0.086 hr^?1. It was concluded that a continuous fermentation of carob aqueous extract using F. moniliforme should be operated at growth rates of approximately 0.205 hr^?1 in order to maximize protein production.