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Applied Microbiology and Biotechnology (v.49, #4)
Production of xanthan gum and ice-nucleating material from whey by Xanthomonas campestris pv. translucens by H. Kawahara; H. Obata (pp. 353-358).
Xanthomonas campestris pv. translucens IFO13599 could produce xanthan gum (18.5 mg/100 mg, lactose) with lactose as the growth substrate in spite of a low level of β-galactosidase. This productivity corresponded to one-fifth that with glucose. This strain could also produce ice-nucleating material having an ice-nucleating temperature, T 50, of −2.8 °C with xanthan gum in the culture broth. We found that this strain produced both materials in whey medium from which the insoluble components had been removed. The production of xanthan with ice-nucleating material reached a maximum after cultivation for 168 h under optimum conditions. Furthermore, the xanthan obtained had a low viscosity because of its variant structure revealed, by TLC and HPLC analyses, to be lacking pyruvic acid. Furthermore, we concluded that this mixture had considerable potential as a regeneratic agent, when compared to other regeneratic agents such as carboxymethylcellulose.
Production of 1,3-propanediol by Clostridium butyricum in continuous culture with cell recycling by A. Reimann; H. Biebl; W.-D. Deckwer (pp. 359-363).
The continuous fermentation of 1,3-propanediol from glycerol by Clostridium butyricum was subjected to cell recycling by filtration using hollow-fibre modules made from polysulphone. The performance of the culture system was checked at a retention ratio (dilution rate/bleed rate) of 5, dilution rates between 0.2 h−1 and 1.0 h−1 and glycerol input concentrations of 32 g l−1 and 56 g l−1. The near-to-optimum propanediol concentration of 26.5 g l−1 (for 56 g l−1 glycerol) was maintained up to a dilution rate of 0.5 h−1 and then decreased while the propanediol productivity was highest at 0.7 h−1. The productivity could be increased by a factor of four in comparison to the continuous culture without cell recycling. By application of the model of Zeng and Deckwer [(1995) Biotechnol Prog 11: 71–79] for cultures under substrate excess, it was shown that the limitations resulted exclusively from product inhibition and detrimental influences from the cell recycling system, such as shear stress, were not involved.
Growth-associated synthesis of recombinant human glucagon and human growth hormone in high-cell-density cultures of Escherichia coli by C. S. Shin; M. S. Hong; D. Y. Kim; H. C. Shin; J. Lee (pp. 364-370).
Synthesis of two recombinant proteins (human glucagon and human growth hormone) was investigated in fed-batch cultures at high cell concentrations of recombinant Escherichia coli. The glucose-limited growth was achieved without accumulation of metabolic by-products and hence the cellular environment is presumed invariable during growth and recombinant protein synthesis. Via exponential feeding in the two-phase fed-batch operation, the specific cell growth rate was successfully controlled at the desired rates and the fed-batch mode employed is considered appropriate for examining the correlation between the specific growth rate and the efficiency of recombinant product formation in the recombinant E. coli strains. The two recombinant proteins were expressed as fusion proteins and the concentration in the culture broth was increased to 15 g fusion growth hormone l−1 and 7 g fusion glucagon l−1. The fusion growth hormone was initially expressed as soluble protein but seemed to be gradually aggregated into inclusion bodies as the expression level increased, whereas the synthesized fusion glucagon existed as a cytoplasmic soluble protein during the whole induction period. The stressful conditions of cultivation employed (i.e. high-cell-density cultivation at low growth rate) may induce the increased production of various host-derived chaperones and thereby enhance the folding efficiency of synthesized heterologous proteins. The synthesis of the recombinant fusion proteins was strongly growth-dependent and more efficient at a higher specific growth rate. The mechanism linking specific growth rate with recombinant protein productivity is likely to be related to the change in cellular ribosomal content.
Microbial oxidative metabolism of diclofenac: production of 4′-hydroxydiclofenac using Epiccocum nigrum IMI354292 by R. Webster; M. Pacey; T. Winchester; P. Johnson; S. Jezequel (pp. 371-376).
The 4′-hydroxylated metabolite of diclofenac was produced by biocatalysis for probing specific human drug-metabolising enzymes (CYP2C9). An initial screen of 11 microorganisms was carried out (50 ml scale) to identify the organism best suited to the regioselective conversion of diclofenac to its 4′-hydroxylated metabolite. From this screen, the fungus Epicoccum nigrum IMI354292 was selected as the most suitable microorganism. Scale-up was carried out in a 30-l fermenter to which 2 g diclofenac was added. After 48 h, 50% of the diclofenac had been converted to it 4′-hydroxylated metabolite. The broth was then extracted with ethyl acetate and purified by chromatography and crystallisation. This yielded 0.3 g 4′-hydroxydiclofenac with a purity of at least 99%. The 4′-hydroxydiclofenac produced by E. nigrum was characterised by HPLC, mass spectrometry and NMR.
Doubly entrapped baker's yeast survives during the long-term stereoselective reduction of ethyl 3-oxobutanoate in an organic solvent by T. Kanda; N. Miyata; T. Fukui; T. Kawamoto; A. Tanaka (pp. 377-381).
To attain long-term bioreaction in organic solvents with living microorganisms, we tried to protect the microorganisms from the toxicity of the solvent by immobilization. In this study, baker's yeast, which is not tolerant to organic solvents such as isooctane, was selected as a model microorganism and the immobilized living yeast cells were examined for activity in the steroselective reduction of ethyl 3-oxobutanoate to ethyl (S)-3-hydroxybutanoate in isooctane; an activity that correlated well with the viability of the yeast cells. It was found that double entrapment, that is, further entrapment of calcium-alginate-gel-entrapped cells with a urethane prepolymer, made it possible for the yeast to remain viable in isooctane, although other conventional immobilization methods, such as single entrapment using polysaccharide or synthetic resin prepolymers, were insufficient for its protection. Furthermore, doubly entrapped living yeast cells could carry out the stereoselective reduction in isooctane repeatedly for a long period (more than 1200 h) with occasional cultivation. Thus, double entrapment enabled a microorganism sensitive to organic solvents to survive over long-term bioreaction in an organic solvent.
The bioemulsifier alasan: role of protein in maintaining structure and activity by S. Navon-Venezia; E. Banin; E. Z. Ron; E. Rosenberg (pp. 382-384).
Alasan, the bioemulsifier of Acinetobacter radioresistens KA53, is a high-molecular-mass complex of polysaccharide and protein. Enrichment culture was used to isolate a bacterial strain that grew on alasan as the sole source of carbon and energy, causing the loss of the protein portion of alasan, as well as the emulsifying activity. The degradation was mediated by extracellular proteinases/alasanases. One of these enzymes, referred to as alasanase II, was purified to homogeneity. Alasanase II, as well as pronase, inactivated alasan, whereas a polysaccharide-degrading enzyme mixture, snail juice, had no effect on emulsifying activity. Deproteinization of alasan with phenol yielded a viscous polysaccharide with no emulsifying activity. Heating alasan to 50 °C led to a 2.5-fold irreversible increase in viscosity with no change in emulsifying activity. Heating to 60°–90 °C caused a drop in viscosity and a 5.8-fold increase in emulsifying activity. The deproteinized alasan showed no increase in emulsifying activity and only small changes in viscosity when heated.
Secretion, purification, and characterisation of barley α-amylase produced by heterologous gene expression in Aspergillus niger by N. Juge; B. Svensson; G. Williamson (pp. 385-392).
Efficient production of recombinant barley α-amylase has been achieved in Aspergillus niger. The cDNA encoding α-amylase isozyme 1 (AMY1) and its signal peptide was placed under the control of the Aspergillus nidulans glyceraldehyde-3-phosphate dehydrogenase (gpd) promoter and the A. nidulans trpC gene terminator. Secretion yields up to 60 mg/l were obtained in media optimised for α-amylase activity and low protease activity. The recombinant AMY1 (reAMY1) was purified to homogeneity and found to be identical to native barley AMY1 with respect to size, pI, and immunoreactivity. N-terminal sequence analysis of the recombinant protein indicated that the endogenous plant signal peptide is correctly processed in A. niger. Electrospray ionisation/mass spectrometry gave a molecular mass for the dominant form of 44 960 Da, in accordance with the loss of the LQRS C-terminal residues; glycosylation apparently did not occur. The activities of recombinant and native barley α-amylases are very similar towards insoluble and soluble starch as well as 2-chloro-4-nitrophenol β-d-maltoheptaoside and amylose (degree of polymerisation = 17). Barley α-amylase is the first plant protein efficiently secreted and correctly processed by A. niger using its own signal sequence.
Secretion of the sweet-tasting protein thaumatin by recombinant strains of Aspergillus niger var. awamori by I. Faus; C. del Moral; N. Adroer; J. L. del Río; C. Patiño; H. Sisniega; C. Casas; J. Bladé; V. Rubio (pp. 393-398).
A recombinant form of the sweet-tasting protein thaumatin has been produced in the filamentous fungus Aspergillus niger var. awamori. Expression cassettes containing a synthetic gene encoding thaumatin II were prepared and used to transform Aspergillus niger var. awamori strain NRRL312. Several fungal strains capable of synthesizing and secreting thaumatin into the culture medium were generated, and their production capabilities were determined, first in shake flasks and later in a laboratory fermentor. We report the expression and secretion of thaumatin in concentrations of 5–7 mg/l. This recombinant thaumatin is sweet.
Cloning and expression of Candida guilliermondii xylose reductase gene (xyl1) in Pichia pastoris by C. Handumrongkul; D.-P. Ma; J. L. Silva (pp. 399-404).
A xylose reductase gene (xyl1) of Candida guilliermondii ATCC 20118 was cloned and characterized. The open reading frame of xyl1 contained 954 nucleotides encoding a protein of 317 amino acids with a predicted molecular mass of 36 kDa. The derived amino acid sequence of C. guilliermondii xylose reductase was 70.4% homologous to that of Pichia stipitis. The gene was placed under the control of an alcohol oxidase promoter (AOX1) and integrated into the genome of a methylotrophic yeast, Pichia pastoris. Methanol induced the expression of the 36-kDa xylose reductase in both intracellular and secreted expression systems. The expressed enzyme preferentially utilized NADPH as a cofactor and was functional both in vitro and in vivo. The different cofactor specificity between P. pastoris and C. guilliermondii xylose reductases might be due to the difference in the numbers of histidine residues and their locations between the two proteins. The recombinant was able to ferment xylose, and the maximum xylitol accumulation (7.8 g/l) was observed when the organism was grown under aerobic conditions.
High-level expression of the thermoalkalophilic lipase from Bacillus thermocatenulatus in Escherichia coli by M. L. Rúa; H. Atomi; C. Schmidt-Dannert; R. D. Schmid (pp. 405-410).
An efficient expression system for the previously only weakly expressed thermophilic lipase BTL2 (Bacillus thermocatenulatus lipase 2) was developed for the production of large amounts of lipase in Escherichia coli. Therefore, the gene was subcloned in the pCYTEXP1 (pT1) expression vector downstream of the temperature-inducible λ promoter PL. Three different expression vectors were constructed: (i) pT1-BTL2 containing the mature lipase gene, (ii) pT1-preBTL2 containing the prelipase gene and (iii) pT1-OmpABTL2 containing the mature lipase gene fused to the signal peptide of the OmpA protein, the major outer membrane protein of E. coli. With pT1-BTL2 and pT1-preBTL2, comparable expression levels of 7000–9000 U/g cells were obtained independently of the E. coli host. In contrast, with E. coli JM105 harbouring pT1-OmpABTL2, 660 000 soluble lipase U/g cells was produced, whereas, with E. coli DH5α and BL321, production levels of 30 000 U/g cells were achieved. However, most of the lipase remained insoluble but active after cell breakage because of the unprocessed OmpA signal peptide. A simple cholate extraction followed by proteinase K cleavage and ultrafiltration allowed the isolation of 1.15 × 106 units of 90% pure mature lipase/wet cells.
Partial-pKD1 plasmids provide enhanced structural stability for heterologous protein production in Kluyveromyces lactis by H.-P. Hsieh; N. A. Da Silva (pp. 411-416).
The stability of pKD1-based vectors in the yeast Kluyveromyces lactis was investigated during short- and long-term culture. The vectors carried an expression/secretion cassette consisting of the Saccharomyces cerevisiaeSUC2 gene under the control of the S. cerevisiaeα-factor promoter and leader. The first set of vectors contained the entire pKD1 sequence linearized at either the unique EcoRI or the unique SphI site of the pKD1 plasmid. During long-term sequential batch culture in selective medium with either vector, invertase activity rapidly dropped while the plasmid-bearing population increased from 60% to 100%. This apparently contradictory behavior was due to structural instability. The enzyme restriction patterns of recovered plasmid DNA retained the pKD1 band while the band containing the SUC2 cassette had decreased substantially in size. To overcome this structural instability, a vector carrying the pKD1 replication origin and the cis-acting stability locus (lacking the inverted repeats) was employed in a pKD1+ (but otherwise isogenic) strain. With this plasmid, invertase activity remained constant (for at least 70 generations). While the new vector was significantly more stable, initial invertase activity was substantially lower than that for the vectors containing the full pKD1 sequence. Southern hybridization confirmed that this decrease was primarily due to reduced copy number. The results indicate that full-pKD1 vectors may be preferred for batch culture, while partial-pKD1 vectors are more suitable for long-term (e.g. fed-batch or continuous) culture.
Construction of a food-grade multiple-copy integration system for Lactococcus lactis by K. Leenhouts; A. Bolhuis; G. Venema; J. Kok (pp. 417-423).
A food-grade vector system was developed that allows stable integration of multiple plasmid copies in the chromosome of Lactococcus lactis. The vector consists of the plus origin of replication (Ori+) of the lactococcal plasmid pWV01, the sucrose genes of the lactic acid bacterium Pediococcus pentosaceus PPE1.0 as selectable marker, a multiple-cloning site, and a lactococcal DNA fragment of a well-characterized chromosomal region. The system includes two L. lactis strains, LL108 and LL302, which produce the pWV01 RepA protein essential for replication of the Ori+ vectors. These helper strains allow the construction and isolation of the replicating form of the integration plasmids from a homologous background. Single-cross-over integration of the plasmids in L. lactis MG1363 resulted in amplifications to a level of approximately 20 copies/chromosome after selection of the transformants on medium containing sucrose as the only fermentable sugar. The amplifications were stable under selective growth conditions. In glucose-containing medium a limited loss of integrated plasmid copies was detected at a rate of (7.5–15) × 10−2 copies per generation. One strain, MG124, was isolated that had retained 11 integrated copies after a period of 120 generations of non-selective growth. These results show that the single-cross-over integration system described here represents a simple procedure for the engineering of stable food-grade strains carrying multiple copies of a gene of interest.
Influence of charge variation in the Streptomyces venezuelae a-amylase signal peptide on heterologous protein production by Streptomyces lividans by E. Lammertyn; S. Desmyter; S. Schacht; L. Van Mellaert; J. Anné (pp. 424-430).
With the aim of investigating determining factors for secretion of heterologous proteins by streptomycetes, we analysed the effect of charge variation in the Streptomyces venezuelae ATCC15068 α-amylase signal peptide on expression and secretion of mouse tumour necrosis factor α (mTNF) by Streptomyces lividans. To this end, the mTNF cDNA was fused to the wild-type α-amylase (aml) signal sequence and the fusion gene was expressed under the control of the S. venezuelae CBS762.70 subtilisin inhibitor gene (vsi) promoter, which has been shown to be very effective in initiating transcription. In addition, the number of positive charges in the N region of the α-amylase signal peptide was altered by in vitro mutagenesis. Secreted and intracellular mTNF levels were determined by sodium dodecyl sulphate/polyacrylamide gel electrophoresis and biologi-cal activity measurements. This revealed moderate amounts of secreted mTNF compared to the levels obtained in previous experiments using the vsi promoter in combination with the Vsi signal peptide. Levels of secreted mTNF could be increased sevenfold by introducing one extra positive charge in the N region of the signal peptide.
Poly(hydroxyalkanoate) biosynthesis from triglyceride substrates by R. D. Ashby; T. A. Foglia (pp. 431-437).
The biosynthesis of poly(hydroxyalkanoates) (PHA) by Pseudomonas resinovorans from triglyceride substrates was investigated. Each triglyceride, whether animal fat or vegetable oil, supported cellular growth to relatively high average cell yields (3.3 ± 0.2 g/l). PHA yields ranged from 1.1 g/l to 2.1 g/l, representing approximately 45% of the bacterial cell dry weight. The repeat-unit composition of the polymers was determined by gas chromatography (GC) and GC/mass spectrometry of the β-hydroxyalkanoate methyl esters from the hydrolyzed polymers. With the exception of PHA from soybean oil (PHA-soy), each polyester was composed of β-hydroxyacyl moieties with chain lengths ranging from C4 to C14, with C8 and C10 being the predominant species. PHA-soy contained an additional fraction (2%) of C16 monomers. The alkyl side-chains of the PHA contained varying degrees of unsaturation. PHA from coconut oil was composed entirely of saturated side-chains, whereas PHA-soy contained 4.2 mol% olefinic groups in its side-chains. The increase in the degree of side-chain unsaturation caused decreased melting temperatures, enthalpies of fusion, and glass transition temperatures. The molar masses of the polymers were relatively constant and ranged from 6.5 × 104 to 10.1 × 104 g/mol.
Influence of culture parameters on extracellular peroxidase activity and transformation of low-rank coal by Phanerochaete chrysosporium by J. P. Ralph; D. E. A. Catcheside (pp. 438-444).
The white-rot fungus Phanerochaete chrysosporium can degrade macromolecules in low-rank coal, offering the potential for converting coal to specific products. We investigated the influence of temperature, veratryl alcohol and oxygen on transformation of a solubilised fraction of Morwell brown coal (SWC6 coal) and on the activity of lignin peroxidase and manganese (Mn) peroxidase in N-limited cultures of P. chrysosporium. After 20 days, the mass and A 400 of SWC6 coal recovered from cultures containing 0.03% SWC6 coal, incubated at 28 °C under hyperbaric oxygen, were reduced by over 95%. The modal apparent molecular mass of the residuum was reduced by 50%. Addition of 2 mM veratryl alcohol had little effect on the transformation of SWC6 coal. The extent of transformation was reduced in cultures incubated at 37 °C or under air. In cultures under air, coal molecules were transiently polymerised. Decolourisation of SWC6 coal reflects conversion to products that cannot be recovered from the medium, not the destruction of chromophores within recoverable material. The activity of lignin peroxidase, measured in cultures free of SWC6 coal to avoid interference with the assay, correlates directly with the degradation of SWC6 coal as measured by the decline in A 400. The data suggest that lignin peroxidase is more important than Mn peroxidase in converting SWC6 coal to products that are assimilated by cells.
Biotransformation of isoquinoline, phenanthridine, phthalazine, quinazoline, and quinoxaline by Streptomyces viridosporus by J. B. Sutherland; J. P. Freeman; A. J. Williams (pp. 445-449).
Streptomyces viridosporus T7A (ATCC␣39115), during growth in tryptone/yeast extract broth, cometabolized five heterocyclic nitrogen-containing compounds. The metabolites produced from the azaarenes were identified by high-performance liquid chromatography, UV/visible absorption spectroscopy, and mass spectrometry. Isoquinoline was transformed to 1(2H)-isoquinolinone (14%), phenanthridine to 6(5H)-phenanthridinone (25%), phthalazine to 1(2H)-phthalazinone (46%), quinazoline to 2,4(1H,3H)-quinazolinedione (4%), and quinoxaline to 2(1H)-quinoxalinone (8%) and 1-methyl-2(1H)-quinoxalinone (12%).
Hydrogen production with high yield and high evolution rate by self-flocculated cells of Enterobacter aerogenes in a packed-bed reactor by M. A. Rachman; Y. Nakashimada; T. Kakizono; N. Nishio (pp. 450-454).
Continuous hydrogen gas evolution by self-flocculated cells of Enterobacter aerogenes, a natural isolate HU-101 and its mutant AY-2, was performed in a packed-bed reactor under glucose-limiting conditions in a minimal medium. The flocs that formed during the continuous culture were retained even when the dilution rate was increased to 0.9 h−1. The H2 production rate increased linearly with increases in the dilution rate up to 0.67 h−1, giving maximum H2 production rates of 31 and 58 mmol l−1 h−1 in HU-101 and AY-2 respectively, at a dilution rate of more than 0.67 h−1. The molar H2 yield from glucose in AY-2 was maintained at about 1.1 at dilution rates between 0.08 h−1 and 0.67 h−1, but it decreased rapidly at dilution rates more than 0.8 h−1.
Activity-dependent fluorescent labeling of bacteria that degrade toluene via toluene 2,3-dioxygenase by W. K. Keener; M. E. Watwood; W. A. Apel (pp. 455-462).
Alternative substrates for the toluene 2,3-dioxygenase pathway of several pseudomonads served as enzyme-activity-dependent fluorescent probes for the bacteria. Phenylacetylene and cinnamonitrile were transformed to fluorescent and brightly colored products by Pseudomonas putida F1, Pseudomonas fluorescens CFS215, and Burkholderia (Pseudomonas) strain JS150. Active bacteria transformed phenylacetylene, producing bright yellow solutions containing the putative product 2-hydroxy-6-oxo-7-octyn-2,4-dienoate. Transformation of cinnamonitrile resulted in bright orange solutions due to accumulation of the putative product 2-hydroxy-6-oxo-8-cyanoocta-2,4,7-trienoate. Chemical and physical properties of the products supported their identification, which indicated that the first three enzymes of the pathway catalyzed product formation. Phenylacetylene labeled bacteria with green fluorescence emission; bacteria were concentrated on black 0.2-μm-pore-size polycarbonate filters containing polyvinylpyrrolidone (PVP) as a wetting agent. Bacteria labeled with cinnamonitrile were fluorescent orange; labeling was effective with bacteria trapped on PVP-free polycarbonate filters. Production of the enzymes involved in labeling of P. putida F1 and P. fluorescens CFS215 was induced by growth (on arginine) in the presence of toluene; cells grown on arginine without toluene were not labeled. Labeling of P. putida F1 by phenylacetylene was inhibited by toluene, indicating that the same enzymatic pathway was required for transformations of both substrates. Bacteria expressing other toluene-degrading enzymatic pathways were not fluorescently labeled with phenylacetylene.
Nitrate and phosphate ion removal from water by Phormidium laminosum immobilized on hollow fibres in a photobioreactor by S. Sawayama; K. K. Rao; D. O. Hall (pp. 463-468).
Removal of nitrate and phosphate ions from water, by using the thermophilic cyanobacterium Phormidium laminosum, immobilized on cellulose hollow fibres in the tubular photobioreactor at 43 °C, was studied by continuously supplying dilute growth medium for 7 days and then secondarily treated sewage (STS) for 12 days. The concentrations of NO− 3 and PO3− 4 in the effluent from the dilute growth medium decreased from 5.0 mg N/l to 3.1 mg N/l, and from 0.75 mg P/l to 0.05 mg P/l respectively, after a residence time of 12 h. The concentrations of NO− 3 and PO3− 4 in the effluent from STS decreased from 11.7 mg N/l to 2.0 mg N/l, and from 6.62 mg P/l to 0.02 mg P/l respectively, after a residence time of 48 h. The removal rates of nitrogenous␣and phosphate ions from STS were 0.24 and 0.11 mmol day−1 l reactor−1 respectively, under the same conditions. Although, among nitrogenous ions, nitrate and ammonium ions were efficiently removed by P.␣laminosum, the nitrite ion was released into the effluent when STS was used as influent. Treatment of water with thermophilic P. laminosum immobilized on hollow fibres thus appears to be an appropriate means for the removal of inorganic nitrogen and phosphorus from treated wastewater.
Removal of organic pollutants and of nitrate from wastewater from the dairy industry by denitrification by G. Zayed; J. Winter (pp. 469-474).
The aim of this work was to remove nitrate-N and organic pollutants from wastewater of the dairy industry by denitrification. An artificially prepared wastewater, containing 250 mg/l nitrate-N and 1.5 g/l whey powder, was completely denitrified with removal of 90%–93% of the chemical oxygen demand (COD) of the whey powder by suspended or immobilized mixed cultures and by a suspended or immobilized pure culture that was isolated from the mixed culture inoculum. For the above COD/nitrate-N ratio of 6:1, the results indicated that the organic compounds of the wastewater served as electron donors for complete denitrification and that there was no need to add an external carbon source. In batch denitrification assays the suspended or immobilized mixed cultures proved to be more active and reacted faster than the isolated pure cultures. In continuous denitrification processes with immobilized pure or mixed cultures, the alginate beads, used for immobilization, were not stable for more than 12 days of incubation. The mixed free cultures removed the nitrate-N and COD continuously with no change of their activity for at least 15 days at an optimum hydraulic retention time of 0.27 days with a loading rate of 900 mg nitrate-N l−1 day−1.
Kinetics of biodegradation of gasoline and its hydrocarbon constituents by L. Yerushalmi; S. R. Guiot (pp. 475-481).
Aerobic biodegradation of gasoline and its constituents, benzene, toluene and ethylbenzene were studied by an enrichment from soil indigenous microbial population. The enrichment culture completely degraded 16.1–660 mg/l gasoline in 2.5–16 days respectively, without accumulation of any by-products. The kinetics of gasoline as well as benzene, toluene and ethylbenzene biodegradation was investigated with initial gasoline concentrations of 16.1–62.6 mg/l. The maximum specific rates of biodegradation of benzene, toluene and ethylbenzene were 0.12, 0.38 and 0.19 mg mg biomass−1 day−1 respectively. When benzene and toluene were used as sole substrate, the maximum specific rates of their biodegradation were 62.9 and 16.4 times greater than the corresponding values for a mixture (gasoline). The microbial culture was able to mineralize up to 200 mg/l pure toluene and benzene. Maximum mineralization efficiencies of benzene and toluene were 76.7 ± 5.1% and 76.8 ± 1.3% respectively. Self-inhibition and competitive inhibition patterns were observed during the biodegradation of benzene and toluene alone and in the mixture respectively. The observed kinetics was modeled according to Andrews' inhibition model.
Low-temperature bioremediation of a waste water contaminated with anionic surfactants and fuel oil by R. Margesin; F. Schinner (pp. 482-486).
We conducted a laboratory study at 10 °C on the biological decontamination of the waste water from a garage and car-wash that was contaminated with anionic surfactants (57 mg l−1) and fuel oil (184 mg hydrocarbons l−1). The indigenous microorganisms degraded both contaminants efficiently after biostimu- lation by an inorganic nutrient supply. After 7 days at 10 °C, the residual contaminations were 11 mg anionic surfactants l−1 and 26 mg hydrocarbons l−1. After 35 days, only the anionic surfactants had been further reduced to 3 mg l−1. Bioaugmentation of the unfertilized waste water with a cold-adapted inoculum, able to degrade both hydrocarbons (diesel oil) and anionic surfactants (sodium dodecyl sulphate), resulted in a significant increase of the hydrocarbon biodegradation during the first 3 days of decontamination, whereas biodegradation of anionic surfactants was inhibited during the first 21 days following inoculation. Bioaugmentation of the nutrient-amended waste water was without any effect.