Gerätesystem zur Massenspektrometrischen Prozeßüberwachung in der Fermentationstechnik

A quadrupol mass spectrometer QMS c 300 developed and manufactured from the workshops of the Institute of Nuclear Research of the Hungarian Academy of Sciences, Debrecen, has been cuppled with a 12 l laboratory steril fermenter. The QMS apparatus is provided with 3 different possibilities for sampling. Via a dynamic air input system the discharged air of the fermenter can be analyzed continuously. The static sampling input allows the off-line measurement of gaseous and volatile compounds. Special membran samplers are used for measurements of volatile and gaseous substrates and microbial metabolites in the fermentation broth. The samplers can be adopted in the reactor vessel directly analogous to the pH and pO₂-electrodes.     

Lycopene production from synthetic medium by <Emphasis Type="Italic">Blakeslea trispora</Emphasis> NRRL 2895 (+) and 2896 (−) in a stirred-tank fermenter

The dissolved oxygen tension of 20% of air saturation, pH-shift from 4.0 to 5.5 on day 3, and a moderate shear stress (calculated as an impeller tip speed, V_\texttip = 0. 9 2 6- 2. 1 6 1  \textm/\texts V_{\text{tip}} = 0. 9 2 6- 2. 1 6 1 \, {\text{m}}/{\text{s}} ) were identified to be the key factors in scaling-up the mated fermentation of Blakeslea trispora NRRL 2895 (+) and 2896 (−) for lycopene production from a shake flask to a stirred-tank fermenter. The maximal lycopene production of 183.3 mg/L was obtained in 7.5-L stirred-tank fermenter, and then the mated fermentation process was successfully step-wise scaled-up from 7.5- to 200-L stirred-tank fermenter. The comparability of the fermentation process was well controlled and the lycopene production was maintained during the process scale-up. Furthermore, with the integrated addition of 150 μmol/L abscisic acid on day 3, 0.5 g/L leucine and 0.1 g/L penicillin on day 4, the highest lycopene production of 270.3 mg/L was achieved in the mated fermentation of B. trispora in stirred-tank fermenter.     

Modelling of the process yields of a whey fermentation

Summary The biomass yields (y) and COD reduction efficiencies (η) of a whey fermentation by Kluyveromyces fragilis were studied in a 100-1 fermenter at various stirrer speeds and lactose concentrations, and compared to those obtained in 10-1 and 15-1 fermenters at constant values of the oxygen transfer coefficient (k_La) and air velocity. The empirical models previously constructed by using the 15-1 fermenter data could be used to predict the yields on the other scales by calculating for each run the 15-1 fermenter which would provide the same oxygen transfer coefficient measured by the sulphite method on each fermenter under study. To make this model independent of stirrer speeds used in each generic fermenter, the effect of aeration and mixing was incorporated into an overall parameter (k_La) and the values of y and η were correlated only with temperature, lactose level and k_L a, since these variables were approximately orthogonal. The validity of this model was finally checked against the yields reported by Wasserman et al. (1961) in a 6-m³ fermenter, thus confirming the capability of the model to provide a reliable basis for further scale-up on the production scale.     

Low-pressure airlift fermenter for single cell protein production: I. Design and oxygen transfer studies

The energy consumption of a fermenter constitutes a major part of the operating expense of a single cell protein process. A low-pressure airlift fermenter was designed to reduce this cost. In this new design, the fermenter broth is kept below 120 cm in depth, and air alone is employed to fulfill the need of supplying oxygen, and cooling and agitating the broth. The use of low-pressure air from air blowers instead of air compressors lowers the capital cost of air delivery and reduces the energy consumption in the fermenter section to below 1 kWh/kg protein, a saving of over 70% as compared to a conventional stirred tank fermenter. It also eliminates the investment of mechanical agitators, heat exchangers, and air compressors. Sulfite oxidation studies confirmed the design concepts.     

Evaporative temperature and moisture control in a rocking reactor for solid substrate fermentation

Summary In the solid substrate fermentation of cooked yellow corn grits with Rhizopus oligosporus in a rocking drum fermenter, temperature was controlled by blowing air through the substrate, forcing water evaporation. The rate of evaporation was controlled by the relative humidity of the air, according to the rate of heat generation during fermentation. Moisture content was maintained constant by spraying cold water on the substrate regulated by the water balance equation of the system. Both controls were operated by computer programs. The rocking motion in the reactor allowed even distribution of air and water in the substrate without disturbing the growing mycelia.     

Alginate production by <Emphasis Type="Italic">Pseudomonas mendocina</Emphasis> in a stirred draft fermenter

In this study alginate production by Pseudomonas mendocina in a laboratory-scale fermenter was investigated. In the experiments the effect of temperature (25–31°C) and agitation (500–620 rev min⁻¹) at a constant air flow of 10 v/v/h were evaluated in relation to the rate of glucose bioconversion to alginate using response surface methodology (RSM). The fermenter configuration was also adapted to a system with a screw mixer and draft tube, due to the change in rheological characteristics of the fermentation broth. The adjusted model indicates a temperature of 29.1°C and agitation of 553 rev min⁻¹ for optimum alginate synthesis. In this fermentation system a Y _p/s of 44.8% was achieved. The alginate synthesized by P. mendocina showed a partially acetylated pattern as previously reported for alginates obtained from other Pseudomonas spp and Azotobacter vinelandii.     

A kLa identification technique for a dynamic model of the coupled system: Air-lift fermenter — oxygen probes

The time delay of oxygen probe response to the signal from a fermenter makes identification of the volumetric oxygen transfer coefficient k_La by the dynamic method more complicated. A coupled model involving the transient-state oxygen balance of the fermenter together with the dynamic model of the oxygen probe must be then formulated, solved and identified. In this paper two simple models of air-lift loop fermenters have been proposed and a coupled mathematical model of the fermenter – oxygen probe system has been developed. The identification procedure was used to estimate k_La values in the fermenter with internal circulation flow on the basis of experimental measurements. A comparison of evaluated and experimental indications of the probes placed at various heights of the column proves that the model presented gives a possibility of the first-step approximation of k_La in loop fermenters.     

Growth and nicotinate biotransformation in batch cultured and airlift fermenter grown soybean cell suspension cultures

Summary Fermentation of a heterotrophic cell suspension culture of soybean (Glycine max) in a 15 l airlift fermenter for 10 to 12 days yielded appr. 4.67 kg fr. wt. (233.5 g dr. wt.) of biomass starting with an inoculum of 300 g fr. wt. Growth limiting factors were the conditions of preculturing the cells and the available amounts of phosphate and oxygen in the fermenter culture medium. Comparison of the metabolic activity of cells grown in 200 ml Erlenmeyer flasks and in the fermenter was performed by measuring the enzyme activities of nicotinamide-amidohydrolase and SAM: nicotinic acid-N-methyltransferase both in the presence and the absence of exogenous nicotinic acid during the biotransformation of nicotinate to trigonelline. The applied nicotinic acid was quantitatively taken up by the cells and an enhanced production of trigonelline could be observed. The activities of the aforementioned soybean enzymes were the same both in the batch cultured and in the airlift fermenter grown cells indicating good physiological conditions of the cells from the fermenter process.Abbreviations SAM S-adenosyl methionine - SCV sedimented cell volume - fr. wt. fresh weight - dr. wt. dry weight     

The negligible role of carbon dioxide and ethylene in ajmalicine production by Catharanthus roseus cell suspensions

Summary Removal of gaseous metabolites in an aerated fermenter affects ajmalicine production by Catharanthus roseus negatively. Therefore, the role of CO₂ and ethylene in ajmalicine production by C. roseus was investigated in 3 l fermenters (working volume 1.8 l) with recirculation of a large part of the exhaust air. Removal of CO₂, ethylene or both from the recirculation stream did not have an effect on ajmalicine production. Inhibition of ethylene biosynthesis in shake flasks with Co²⁺, Ni²⁺ or aminooxyacetic acid did not affect ajmalicine production. However, the removal of CO₂ did enhance the amount of extracellular ajmalicine.     

Physiological response ofCandida tropicalis grown on n-paraffin to mixing in a tubular closed loop fermenter

Summary A special tubular closed loop fermenter was used in order to simulate the particular mixing condition of a large scale recycle fermenter.Some mixing parameters of the system are characerized.During continuous cultivation ofCandida tropicalis on n-paraffin as a substrate the biomass yield with respect to carbon and oxygen increased, when a controlled oxygen limit was imposed on the culture.Mixing in the closed loop fermenter generates undamped short period oscillations in the respiration activity, in the dissolved oxygen tension and in the actual ATP content of the culture. These oscillations likely represent oscillations of allosteric feedback loops which manifest themselves by some synchronising action of the particular environmental transients in the closed loop fermenter.     

Cellulose degradation and carboxymethylcellulase production by Sporocytophaga myxococcoides grown in an air-lift fermenter

Summary Sporocytophaga myxcoccoides was grown in a 31 air-lift fermenter using a medium containing 2% w/v insoluble cellulose. The cellulose content of the medium reduced the k_La of the fermenter but during growth the dissolved oxygen concentration did not fall below 75% saturation. Rates of cellulose degradation and extracellular enzyme production were similar to those reported for a stirred-tank fermenter.     

Production of poly(3-hydroxybutyrate) from whey by fed-batch culture of recombinant Escherichia coli in a pilot-scale fermenter

Production of poly(3-hydroxybutyrate) [P(3HB)] from wheyby fed-batch culture of recombinant Escherichia coli CGSC 4401 harboring a plasmid containing the Alcaligenes latus polyhydroxyalkanoate (PHA) biosynthesis genes was examined in a 30 l fermenter supplying air only. With lactose below 2 g l^–1, cells grew to 12 g dry cell l^–1 with 9% (w/w) P(3HB) content. Accumulation of P(3HB) could be triggered by increasing lactose to 20 g l^–1. By employing this strategy, 51 g dry cell l^–1 was obtained with a 70% (w/w) P(3HB) content after 26 h. The productivity was 1.35 g P(3HB) l^–1 h^–1. The same fermentation strategy was used in a 300 l fermenter, and 30 g dry cell l^–1 with 67% (w/w) P(3HB) content was obtained in 20 h.     

Photoautotrophic growth of cell suspension cultures fromChenopodium rubrum in an airlift fermenter

Summary This communication describes the construction and operation of an airlift fermenter for the photoautotrophic growth of cell suspension cultures fromChenopodium rubrum. The basic batch culture unit provides a culture of 1.51 volume, sufficient to permit frequent aseptic sampling. It can be maintained at any desired temperature and aerated to different extents. Using an initial cell density of about 400,000 cells per ml suspension, the increase in cell number is 270% after a 14 days' growth period, although the stationary phase of growth is not yet reached. The transfer of photoautotrophic cell suspensions fromChenopodium rubrum from stationary growth into the large volume of fresh culture medium in the airlift fermenter results in an immediate protein formation, followed by an exponential phase of cell division, whereas rapid chlorophyll accumulation is delayed by 2 days.The growth capacities of photoautotrophic fermenter cultures including protein and chlorophyll formation as well asin vitro activities of the ribulosebisphosphate carboxylase and the phosphoenolpyruvate carboxylase are greatly lower as compared to photoautotrophic cells propagated in standard two-tier culture vessels using 30 ml culture medium. However the pattern of change in the activities of carboxylation enzymes is quite similar in both culture systems.Photoautotrophic cell suspensions fromChenopodium rubrum grown in an airlift fermenter assimilate about 90 mol CO₂/mg chlorophyll × hour. Dark CO₂ fixation is about 1.5% of the light values.Abbreviations PEF phosphoenolpyruvate - RuDP ribulosebisphosPhate - NS ground glass joints of standardized size made from Duran glass, Schott, Germany     

The optimization of L-lysine fermentation and a mass transfer model

The efficiency of L-lysine biosynthesis is essentially determined by the power input and aeration ratio in the stirred fermenter. A mass transfer model was developed by means of the results of lysine fermentations in four geometrically similar fermenters with working volumes of 10 1, 50 1, 100 1 and 2500 1 which allows the optimization of lysine fermentation from the energetical point of view. The usefullness of this k_La-model is demonstrated with an example where the power input for an unknown fermenter is calculated.     

Response of the thermophile Thermus sp. RQ-1 to hyperbaric air in batch and fed-batch cultivation

The effects of increased air pressure in a culture of the thermophilic microorganism Thermus sp. RQ-1 were investigated. Cell growth dependence on oxygen supply was investigated in a fermenter at atmospheric pressure. Total oxygen depletion from the medium for low values of k _La was observed during the exponential growth phase. It was possible with this strain to enhance the oxygen transfer rate by increasing the air pressure. Cell productivity was improved by pressurisation up to 0.56 MPa for batch cultivation; and an induction of the antioxidant enzymes, superoxide dismutase and catalase, was observed with the rise in pressure. Cell pre-cultivation under pressurised conditions conferred to the cells more resistance to an exposure to hydrogen peroxide and more sensitivity to paraquat (methyl viologen). The usefulness of bioreactor pressurisation on the cultivation of Thermus sp. RQ-1 was demonstrated for fed-batch operation, with the attainment of higher cell densities. A two-fold increase in cell mass productivity was obtained by the use of hyperbaric air (0.5 MPa). With the pressurisation of the head-space in the reactor, it was also possible to eliminate the loss of liquid by evaporation, which amounted to more than 10% at 70 °C and atmospheric pressure. Received: 16 July 1999 / Revision received: 26 November 1999 / Accepted: 28 November 1999     

Formation of gluconic acid at low pH-values by free and immobilized Aspergillus niger cells during citric acid fermentation

Summary A recently developed immobilization method, characterized by the adsorption of the mycelia onto a glass-carrier in a fixed-bed reactor, was applied for citric acid production by Aspergillus niger ATCC 9142, and compared with conventional culture techniques.In a fixed-bed reactor and in a stirred fermenter a rapid gluconic acid production started immediately after nitrate exhaustion, though the pH was below 2.5 During a second production phase a comparatively small amount of citric acid was formed.In surface and shaken-flask cultures nearly no gluconic acid could be found, whereas citric acid yields were significantly higher than in the fixed-bed reactor and in the stirred fermenter.Manganese (0.8×10^–7 Mol×dm^–3 after 6 days incubation) from the stainless steel parts of the vessel seemed to be responsible for both gluconic acid production and small citric acid yields in the stirred fermenter and in the fixed-bed reactor.     

Growth of Streptococcus faecalus in dense culture

A fermentation system was designed and constructed to study the growth characteristics of microorganisms at low and high cell concentrations. The technique used to develop high cell densities utilized a rotating microfiltration unit to permit the removal of cell-free product from the fermenter. The fermenter volume and the filter were contained in a single unit composed of a series of concentric cylinders. Annuli contained the fermenter volume while the second outermost cylinder supported a microfiltration membrane. Feed to the system was pumped at constant rates, and the internal pressure built up to a value, which would effect the required filtration rate. The system was operated batchwise and continuously with and without filtration. The anaerobie growth characteristics of Streptococcus faccalus were determined at 37°C and pH 7.0 for batch, continuous, and continuous with filtration modes of operation. The growth characteristics were unchanged when the cell density was increased. Changes in cell yield peer model of glucose consumed were clearly illustrated during thee course of single run by operating the fermenter in the unsteady state with filtration. No consumption of glucose for developed was 40% packed cell volume, a value 45 times larger than could be grown in simple batch culture.     

以麦秸为基质豌豆根瘤菌的压力脉动固态发酵

生物肥料的常规生产方法是经液态发酵后 ,再以固态基质吸附[1 ] 。其工艺繁琐 ,吸附时易引入大量杂菌 ,从而降低生物肥料的施用效果。而由于静止浅盘固态发酵存在着较大的氧气浓度和温度梯度[2 ,3 ] ,及易染菌等因素 ,所以利用固态发酵成功生产生物肥料的研究较少。近年来 ,固态发酵以其优于液态发酵的特点而成为人们研究的热点[4] ,但至今仍未能普遍应用于工业生产。其主要原因是固态发酵反应器在放大过程中传质、传热困难[3 ] 。Ｂａｊｒａｃｈａｒｙａ＆Ｍｕｄｇｅｔｔ[5 ] 指出 ,解决上述弊端的唯一可行的方法是对反应器中的气相进行控…     

Low-pressure airlift fermenter for single cell protein production: II. Continuous culture of pichia yeast

Experiments using Pichia yeast grown on n-paraffins have been conducted in laboratory 10-L airlift fermenters and in a 640-L module of commercial scale. Results confirmed the design concept of combining oxygen transfer and fermenter cooling with low-pressure air. However, in the absence of mass transport constraints, the build up of toxic factors in the fermenter appeared to be a major variable limiting cell productivity. Foaming in the large fermenter also presented a serious problem, which must be solved before low-pressure airlift fermenters become practical.     

Degradation of hydrocarbons and alcohols by Rhodococcus erythropolis DCL14: A comparison in scale performance

The degradation of alkanes and alcohols by Rhodococcus erythropolis DCL14 growing cells was tested at different reactor scales, ranging from a 2-L fermenter to 300 µL 96-well plates. The highest growth rates were attained in the fermenter, where pH, temperature and aeration rate, in particular, are monitored and controlled. However, the results acquired in reactors smaller than 100 mL indicate that these reactors may be successfully used in testing carbon sources, since the normalized results obtained with the different carbons sources were maintained in the 100–0.3 mL range. This study also shows the great potential of R. erythropolis DCL14 cells to degrade a wide range of alcohols and alkanes, which may be used in bioremediation of contaminated sites.