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1.
High-cell-density cultivation of microorganisms 总被引:29,自引:0,他引:29
High-cell-density cultivation (HCDC) is required to improve microbial biomass and product formation substantially. An overview
of HCDC is given for microorganisms including bacteria, archae and eukarya (yeasts). Problems encountered by HCDC and their
possible solutions are discussed. Improvements of strains, different types of bioreactors and cultivation strategies for successful
HCDC are described. Stirred-tank reactors with and without cell retention, a dialysis-membrane reactor, a gas-lift reactor
and a membrane cyclone reactor used for HCDC are outlined. Recently modified traditional feeding strategies and new ones are
included, in particular those for unlimited growth to very dense cultures. Emphasis is placed on robust fermentation control
because of the growing industrial interest in this field. Therefore, developments in the application of multivariate statistical
control, artificial neural networks, fuzzy control and knowledge-based supervision (expert systems) are summarized. Recent
advances using Escherichia coli– the pioneer organism for HCDC – are outlined.
Received: 20 October 1998 / Received revision: 18 December 1998 / Accepted: 21 December 1998 相似文献
2.
3.
During cassava starch production, large amounts of cyanoglycosides were released and hydrolysed by plant-borne enzymes, leading
to cyanide concentrations in the wastewater as high as 200 mg/l. For anaerobic degradation of the cyanide during pre-acidification
or single-step methane fermentation, anaerobic cultures were enriched from soil residues of cassava roots and sewage sludge.
In a pre-acidification reactor this culture was able to remove up to 4 g potassium cyanide/l of wastewater at a hydraulic
retention time (t
HR) of 4 days, equivalent to a maximal cyanide space loading of 400 mg CN− l−1 day−1. The residual cyanide concentration was 0.2–0.5 mg/l. Concentrated cell suspensions of the mixed culture formed ammonia and
formate in almost equimolar amounts from cyanide. Little formamide was generated by chemical decay. A concentration of up
to 100 mmol ammonia/l had no inhibitory effect on cyanide degradation. The optimal pH for cyanide degradation was 6–7.5, the
optimal temperature 25–37 °C. At a pH of 5 or lower, cyanide accumulated in the reactor and pre-acidification failed. The
minimal t
HR for continuous cyanide removal was 1.5 days. The enriched mixed culture was also able to degrade cyanide in purely mineralic
wastewater from metal deburring, either in a pre-acidification reactor with a two-step process or in a one-step methanogenic
reactor. It was necessary to supplement the wastewater with a carbon source (e.g. starch) to keep the population active enough
to cope with any possible inhibiting effect of cyanide.
Received: 29 April 1998 / Received revision: 8 June 1998 / Accepted: 14 June 1998 相似文献
4.
Minekus M Smeets-Peeters M Bernalier A Marol-Bonnin S Havenaar R Marteau P Alric M Fonty G Huis in't Veld JH 《Applied microbiology and biotechnology》1999,53(1):108-114
This paper introduces a new type of system to simulate conditions in the large intestine. This system combines removal of
metabolites and water with peristaltic mixing to obtain and handle physiological concentrations of microorganisms, dry matter
and microbial metabolites. The system has been designed to be complementary to the dynamic multi-compartmental system that
simulates conditions in the stomach and small intestine described by Minekus et al. [Minekus M, Marteau P, Havenaar R, Huis
in't Veld JHJ (1995) ATLA 23:197–209]. High densities of microorganisms, comparable to those found in the colon in vivo, were
achieved by absorption of water and dialysis of metabolites through hollow-fibre membranes inside the reactor compartments.
The dense chyme was mixed and transported by peristaltic movements. The potential of the system as a tool to study fermentation
was demonstrated in experiments with pectin, fructo-oligosaccharide, lactulose and lactitol as substrates. Parameters such
as total acid production and short-chain fatty acid (SCFA) patterns were determined with time to characterize the fermentation.
The stability of the microflora in the system was tested after inoculation with fresh fecal samples and after inoculation
with a microflora that was main-tained in a fermenter. Both approaches resulted in total anaerobic bacterial counts higher
than 1010 colony-forming units/ml with physiological levels of Bifidobacterium, Lactobacillus, Enterobacteriaceae and Clostridium. The dry matter content was approximately 10%, while the total SCFA concentration was maintained at physiological concentrations
with similar molar ratios for acetic acid, propionic acid and butyric acid as measured in vivo.
Received: 4 February 1999 / Received revision: 4 June 1999 / Accepted: 4 June 1999 相似文献
5.
Fibrobacter succinogenes S85 cultures that were cellobiose-limited converted cellobiose to succinate and acetate, produced little glucose or cellotriose,
maintained an intracellular ATP concentration of 4.1 mM and a membrane potential of 140 mV for 24 h, did not lyse at a rapid
rate once they had reached stationary phase, and had a most probable number of viable cells that was greater than 106/ml. When the cellobiose concentration was increased 6-fold (5 mM to 30 mM), ammonia was depleted and the cultures left 10 mM
cellobiose. Cultures provided with excess cellobiose produced succinate and acetate while they were growing, but there was
little increase in fermentation acids after the ammonia was depleted and growth ceased. The stationary-phase, cellobiose-excess
cultures had a lysis rate that was 7-fold faster than that of the cellobiose-limited cultures, and the most probable number
was only 3.3 × 103 cells/ml. The stationary-phase, cellobiose-excess cultures had 2.5 times as much cellular polysaccharide as the cellobiose-limited
cultures, but the intracellular ATP and membrane potential were very low (0.1 mM and 40 mV respectively). Methylglyoxal, a
potentially toxic end-product of carbohydrate fermentation, could not be detected, and fresh inocula grew rapidly in spent
medium that was supplemented with additional ammonia. Stationary-phase, cellobiose-excess cultures converted cellobiose to
glucose and cellotriose, but the apparent K
m of cellotriose formation was 15-fold lower than the K
m of glucose production (0.7 mM compared to 10 mM).
Received: 26 June 1997 / Received revision: 12 August 1997 / Accepted: 29 August 1997 相似文献
6.
It is critical that an inexpensive electron- donor/carbon-source be found for selenium bioremedia-tion using the selenate-respiring
bacterium, Thauera selenatis. Since acetate is a preferred substrate for growth of this organism, a method was developed for fermenting the lactose in
whey to large amounts of acetate. Indigenous whey microorganisms fermented the whey lactose in this manner when grown in continuous
culture at a very slow dilution rate (D = 0.05 h−1). The successful use of the fermented whey lactose as the carbon-source/electron-donor feed for a laboratory-scale selenium-bioremediation
reactor system, inoculated with T. selenatis, treating selenium-contaminated drainage water was also demonstrated. Selenium oxyanions and nitrate were reduced by 98%.
Received: 30 October 1998 / Received revision: 26 January 1999 / Accepted: 5 February 1999 相似文献
7.
Very good solvent formation rates were observed when Clostridium beijerinckii NRRL B592 was cultivated on different whole potato media. The increase in whole potato concentration contributed to the increased
final solvent concentrations, while the addition of yeast extract or mineral salts gave negative effects. To obtain good solvent
productivities and high final solvent concentrations during batch fermentation, no enzymatic hydrolysis of the potato starch
was necessary, indicating high activity of the clostridial amylases produced by the strain applied.
Received: 17 April 1998 / Received revision: 22 June 1998 / Accepted: 27 June 1998 相似文献
8.
The role of gravity in the autolysis of Bacillus subtilis and Escherichia coli was studied by growing cells on Earth and in microgravity on Space Station Mir. Autolysis analysis was completed by examining
the death phase or exponential decay of cells for approximately 4 months following the stationary phase. Consistent with published
findings, the stationary-phase cell population was 170% and 90% higher in flight B. subtilis and E. coli cultures, respectively, than in ground cultures. Although both flight autolysis curves began at higher cell densities than
control curves, the rate of autolysis in flight cultures was identical to that of their respective ground control rates.
Received: 3 December 1998 / Received revision: 23 February 1999 / Accepted: 14 March 1999 相似文献
9.
Mechanistic action of pediocin and nisin: recent progress and unresolved questions 总被引:14,自引:0,他引:14
Nisin and pediocin PA-1 are examples of bacteriocins from lactic acid bacteria (LAB) that have found practical applications
as food preservatives. Like other natural antimicrobial peptides, LAB bacteriocins act primarily at the cytoplasmic membranes
of susceptible microorganisms. Studies with in vivo as well as in␣vitro membrane systems are directed toward understanding
how bacteriocins interact with membranes so as to provide a mechanistic basis for their rational applications. The dissipation
of proton motive force was identified early on as the common mechanism for the lethal activity of LAB bacteriocin. Models
for nisin/membrane interactions propose that the peptide forms poration complexes in the membrane through a multi-step process
of binding, insertion, and pore formation. This review focuses on the current knowledge of: (1) the mechanistic action of
nisin and pediocin-like bacteriocins, (2) the requirement for a cell factor such as a membrane protein, (3) the influence
of membrane potential, pH, and lipid composition on the of specificity and efficacy of bacteriocins, and (4) the roles of
specific amino acids and structural domains of the bacteriocins in their action.
Received: 3 April 1998 / Received last revision: 27 July 1998 / Accepted: 29 July 1998 相似文献
10.
The maximum growth rate of Saccharomyces cerevisiae ATCC 96581, adapted to fermentation of spent sulphite liquor (SSL), was 7 times higher in SSL of hardwood than the maximum
growth rate of bakers' yeast. ATCC 96581 was studied in the continuous fermentation of spruce hydrolysate without and with
cell recycling. Ethanol productivity by ATCC 96581 in continuous fermentation of an enzymatic hydrolysate of spruce was increased
4.6 times by employing cell recycling. On-line analysis of CO2, glucose and ethanol (using a microdialysis probe) was used to investigate the effect of fermentation pH on cell growth and
ethanol production, and to set the dilution rate. Cell growth in the spruce hydrolysates was strongly influenced by fermentation
pH. The fermentation was operated in continuous mode for 210 h and a theoretical ethanol yield on fermentable sugars was obtained.
Received: 25 May 1998 / Received revision: 11 August 1998 / Accepted: 12 August 1998 相似文献
11.
Physiological aspects of continuous lactic acid fermentations at high dilution rates 总被引:1,自引:0,他引:1
U. Kulozik 《Applied microbiology and biotechnology》1998,49(5):506-510
The effects of the substrate conditions on the volumetric productivity of Lactobacillus helveticus at different cell densities up to 60 g l−1 in a continuous stirred-tank reactor with microfiltration to retain the biomass were investigated. At low dilution rates,
D, the steady-state volumetric productivity, r
p, gradually increased to a maximum at D = 1.2–1.5 h−1, because of reduced product inhibition. At higher D values, r
p unexpectedly decreased, although the substrate conditions further improved. The maxima of r
p at different cell densities coincided with a critical specific substrate utilization rate beyond which the cell metabolism
seems to be controlled through a catabolic modulator factor, and r
p decreases.
Received: 8 September 1997 / Received last revision: 31 December 1997 / Accepted: 2 January 1998 相似文献
12.
Response of fluoranthene-degrading bacteria to surfactants 总被引:1,自引:0,他引:1
P. A. Willumsen U. Karlson P. H. Pritchard 《Applied microbiology and biotechnology》1998,50(4):475-483
A prerequisite for surfactant-enhanced biodegradation is that the microorganisms survive, take up substrate and degrade it
in the presence of the surfactant. Two Mycobacterium and two Sphingomonas strains, degrading fluoranthene, were investigated for their sensitivity towards non-ionic chemical surfactants. The effect
of Triton X-100 and Tween 80 above their critical micelle concentration on mineralization of [14C]-glucose and [14C]-fluoranthene was measured in shaker cultures. Tween 80 had no toxic effect on any of the tested strains. The surfactant
inhibited fluoranthene mineralization by the hydrophobic Mycobacterium spp. slightly, but more than doubled that by the two less hydrophobic Sphingomonas strains. Triton X-100 inhibited fluoranthene mineralization by all strains, yet this was more pronounced for the Sphingomonas spp. Both surfactants caused cell wall permeabilization, as shown by transient colouring of surfactant-containing media.
Inhibition of glucose mineralization, indicating non-specific toxic effects of Triton X-100, was observed only for the Sphingomonas strains and the toxicity was caused by micelle-to-cell interactions. These strains, however, appeared to recover from initial
Triton X-100 toxicity within 50–500 h of exposure. The ratio of surfactant concentration to initial cell density was found
to determine critically the bacterial response to surfactants. For both Sphingomonas and Mycobacterium strains, this work indicates that fluoranthene solubilized in surfactant micelles is only partially available for mineralization
by the bacteria tested. However, our results suggest that optimal conditions for polycyclic aromatic hydrocarbon mineralization
can be developed by selection of the proper surfactant, bacterial strains, cell density and incubation conditions.
Received: 6 February 1998 / Received revision: 19 June 1998 / Accepted: 19 June 1998 相似文献
13.
Repeated application of carvone-induced bacteria to enhance biodegradation of polychlorinated biphenyls in soil 总被引:4,自引:0,他引:4
Carvone, the principal component of spearmint oil, induces biodegradation of polychlorinated biphenyls (PCB) by Arthrobacter sp. strain B1B. This study investigated the effectiveness of the repeated application of carvone-induced bacteria for bioremediation
of Aroclor-1242-contaminated soil. Control treatments compared a single inoculation of carvone-induced cells, repeated applications
of noninduced cells, and repeated applications of cell-free carvone/fructose medium. The results showed that repeated application
of carvone-induced bacteria was the most effective treatment for mineralizing PCB, resulting in 27 ± 6% degradation of Aroclor
1242 after 9 weeks; whereas a single application of cells resulted in no significant degradation. Addition of cell-free, carvone/fructose
medium resulted in 10% degradation of PCB, which suggests that this treatment stimulated biodegradation of PCB by the indigenous
microflora. The di- and trichlorobiphenyls were the most readily degraded congeners. More highly chlorinated congeners, which
had been previously shown to be degraded in liquid culture, were not substantially degraded in soil, indicating that low bioavailability
may have limited their degradation. With the development of new technology, which permits automated in situ fermentation and
delivery of degrader microorganisms, the repeated application of carvone-induced bacteria may facilitate bioremediation of
PCB-contaminated soils.
Received: 7 January 1998 / Received revision: 18 June 1998 / Accepted: 27 June 1998 相似文献
14.
F. Laborda I. F. Monistrol N. Luna M. Fernández 《Applied microbiology and biotechnology》1999,52(1):49-56
Several fundamental aspects of microbial coal liquefaction/solubilization were studied. The liquefied/solubilized products
from coal by microorganisms were analysed. The liquid products analysed by IR titration and UV/visible spectrometry showed
some alterations with regard to the original coal. Humic acids extracted from the liquefied lignite showed a reduction in
the average molecular weight and a increase in the condensation index, probably due to depolymerization caused by microorganisms.
The mechanisms implicated in coal biosolubilization by two fungal strains, M2 (Trichoderma sp.) and M4 (Penicillium sp.) were also studied. Extracellular peroxidase, esterase and phenoloxidase enzymes appear to be involved in coal solubilization.
Received: 15 June 1998 / Received revision: 23 November 1998 / Accepted: 29 November 1998 相似文献
15.
The sequencing batch reactor as a powerful tool for the study of slowly growing anaerobic ammonium-oxidizing microorganisms 总被引:145,自引:0,他引:145
M. Strous J. J. Heijnen J. G. Kuenen M. S. M. Jetten 《Applied microbiology and biotechnology》1998,50(5):589-596
Currently available microbiological techniques are not designed to deal with very slowly growing microorganisms. The enrichment
and study of such organisms demands a novel experimental approach. In the present investigation, the sequencing batch reactor
(SBR) was applied and optimized for the enrichment and quantitative study of a very slowly growing microbial community which
oxidizes ammonium anaerobically. The SBR was shown to be a powerful experimental set-up with the following strong points:
(1) efficient biomass retention, (2) a homogeneous distribution of substrates, products and biomass aggregates over the reactor,
(3) reliable operation for more than 1 year, and (4) stable conditions under substrate-limiting conditions. Together, these
points made possible for the first time the determination of several important physiological parameters such as the biomass
yield (0.066 ± 0.01 C-mol/mol ammonium), the maximum specific ammonium consumption rate (45 ± 5 nmol/mg protein/min) and the
maximum specific growth rate (0.0027 · h−1, doubling time 11 days). In addition, the persisting stable and strongly selective conditions of the SBR led to a high degree
of enrichment (74% of the desired microorganism). This study has demonstrated that the SBR is a powerful tool compared to
other techniques used in the past. We suggest that the SBR could be used for the enrichment and quantitative study of a large
number of slowly growing microorganisms that are currently out of reach for microbiological research.
Received: 14 May 1998 / Received last revision: 30 July 1998 / Accepted: 31 July 1998 相似文献
16.
The influence of temperature and pH on growth of Leuconostoc mesenteroides subsp. mesenteroides FR52 and production of its two bacteriocins, mesenterocin 52A and mesenterocin 52B, was studied during batch fermentation.
Temperature and pH had a strong influence on the production of the two bacteriocins which was stimulated by slow growth rates.
The optimal temperature was 20 °C for production of mesenterocin 52A and 25 °C for mesenterocin 52B. Optimal pH values were
5.5 and 5.0 for production of mesenterocin 52A and mesenterocin 52B respectively. Thus, by changing the culture conditions,
production of one bacteriocin can be favoured in relation to the other. The relationship between growth and specific production
rates of the two bacteriocins, as a function of the culture conditions, showed different kinetics of production and the presence
of several peaks in the specific production rates during growth.
Received: 13 February 1998 / Received revision: 27 May 1998 / Accepted: 1 June 1998 相似文献
17.
A two-phase organic/aqueous reactor configuration was developed for use in the biodegradation of benzene, toluene and p-xylene, and tested with toluene. An immiscible organic phase was systematically selected on the basis of predicted and experimentally
determined properties, such as high boiling points, low solubilities in the aqueous phase, good phase stability, biocompatibility,
and good predicted partition coefficients for benzene, toluene and p-xylene. An industrial grade of oleyl alcohol was ultimately selected for use in the two-phase partitioning bioreactor. In
order to examine the behavior of the system, a single-component fermentation of toluene was conducted with Pseudomonas sp. ATCC 55595. A 0.5-l sample of Adol 85 NF was loaded with 10.4 g toluene, which partitioned into the cell containing 1 l
aqueous medium at a concentration of approximately 50 mg/l. In consuming the toluene to completion, the organisms were able
to achieve a volumetric degradation rate of 0.115 g l−1 h−1. This system is self-regulating with respect to toluene delivery to the aqueous phase, and requires only feedback control
of temperature and pH.
Received: 16 November 1998 / Received revision: 28 March 1999 / Accepted: 9 April 1999 相似文献
18.
The influence of ammonia on the anaerobic degradation of peptone by mesophilic and thermophilic populations of biowaste was
investigated. For peptone concentrations from 5 g l−1 to 20 g l−1 the mesophilic population revealed a higher rate of deamination than the thermophilic population, e.g. 552 mg l−1 day−1 compared to 320 mg l−1 day−1 at 10 g l−1 peptone. The final degree of deamination of the thermophilic population was, however, higher: 102 compared to 87 mg NH3/g peptone in the mesophilic cultures. If 0.5–6.5 g l−1 ammonia was added to the mesophilic biowaste cultures, deamination of peptone, degradation of its chemical oxygen demand
(COD) and formation of biogas were increasingly inhibited, but no hydrogen was formed. The thermophilic biowaste cultures
were most active if around 1 g ammonia l−1 was present. Deamination, COD degradation and biogas production decreased at lower and higher ammonia concentrations and
hydrogen was formed in addition to methane. Studies of the inhibition by ammonia of peptone deamination, COD degradation and
methane formation revealed a K
i (50%) for NH3 of 92, 95 and 88 mg l−1 at 37 °C and 251, 274 and 297 mg l−1 at 55 °C respectively. This indicated that the thermophilic flora tolerated significantly more NH3 than the mesophilic flora. In the mesophilic reactor effluent 4.6 × 108 peptone-degrading colony-forming units (cfu)/ml were culturable, whereas in the thermophilic reactor effluent growth of only
5.6 × 107 cfu/ml was observed.
Received: 24 April 1998 / Received revision: 26 June 1998 / Accepted: 27 June 1998 相似文献
19.
K. Hayakawa Y. Ueno S. Kawamura T. Kato R. Hayashi 《Applied microbiology and biotechnology》1998,50(4):415-418
In order to test the possibility of utilizing high pressure in bioscience and biotechnology, a simple method for high-pressure
generation and its use for microbial inactivation have been studied. When a pressure vessel was filled with water, sealed
tightly and cooled to sub-zero temperatures, high pressure was generated in the vessel. The pressure generation was 60 MPa
at −5 °C, 103 MPa at −10 °C, and 140 MPa at −15 °C, −20 °C, and −22 °C. The high pressure generated inactivated microorganisms
effectively: yeasts (Saccharomyces cerevisiae and Zygosaccharomyces rouxii), bacteria (Lactobacillus brevis and Eschericia coli), and fungi (Aspergillus niger and Aspergillus oryzae) were completely inactivated when stored in sealed vessels −20 °C for 24 h. However, Staphylococcus aureus was only partly inactivated under the same conditions. This method opens up a new application of high pressure for storing,
transporting, and sterilizing of foods and biological materials.
Received: 28 July 1997 / Received last revision: 12 June 1998 / Accepted: 19 June 1998 相似文献
20.
A mixed culture of microorganisms able to utilize 4,6-dinitro-ortho-cresol (DNOC) as the sole source of carbon, nitrogen and energy was isolated from soil contaminated with pesticides and from
activated sludge. DNOC was decomposed aerobically in batch cultures as well as in fixed-bed column reactors. Between 65% and
84% of the substrate nitrogen was released as nitrate into the medium, and 61% of the carbon from uniformly 14C-labelled DNOC was recovered as 14CO2. The mixed microbial culture also decomposed 4-nitrophenol and 2,4-dinitrophenol but not 2,3-dinitrophenol, 2,6-dinitrophenol,
2,4-dinitrotoluene, 2,4-dinitrobenzoic acid or 2-sec-butyl-4,6-dinitrophenol (Dinoseb). Maximal degradation rates for DNOC by the bacterial biofilm immobilized on glass beads
in fixed-bed column reactors were 30 mmol day−1 (l reactor volume)−1, leaving an effluent concentration of less than 5 μg l−1 DNOC in the outflowing medium. The apparent K
s value of the immobilized mixed culture for DNOC was 17 μM. Degradation was inhibited at DNOC concentrations above 30 μM and
it ceased at 340 μM, possibly because of the uncoupling action of the nitroaromatic compound on the cellular energy-transducing
mechanism.
Received: 27 March 1997 / Received revision: 5 June 1997 / Accepted: 7 June 1997 相似文献