Reclamation of an activated-sludge microbial consortium by selective biostimulation

Our previous study showed that an activated-sludge process broke down at the phenol-loading rate of 1.5 g l⁻¹ day⁻¹, when non-flocculating bacteria (called R6T and R10) overgrew the sludge, resulting in a sludge washout. In this study, we attempted to circumvent this breakdown problem by reclaiming the consortium structure. Activated sludge was fed phenol, and the phenol-loading rate was increased stepwise from 0.5 g l⁻¹ day⁻¹ to 1.0 g l⁻¹ day⁻¹ and then to 1.5 g l⁻¹ day⁻¹. Either galactose or glucose (at 0.5 g l⁻¹ day⁻¹) was also supplied to the activated sludge from the phenol-loading rate of 1.0 g l⁻¹ day⁻¹. Pure culture experiments have suggested galactose to be a preferential substrate for a floc-forming bacterium (R6F) that predominantly degrades phenol under low phenol-loading conditions. Supplying galactose allowed sustainment of the R6F population and suppression of the overgrowth of R6T and R10 at the phenol-loading rate of 1.5 g l⁻¹ day⁻¹. This measure allowed the activated-sludge process to treat phenol at a phenol-loading rate up to 1.5 g l⁻¹ day⁻¹, although it broke down at 2.0 g l⁻¹ day⁻¹. In contrast, supplying glucose reduced the R6F population and allowed the activated-sludge process to break down at the phenol-loading rate of 1.0 g l⁻¹ day⁻¹. This study demonstrated that reclamation of the activated-sludge consortium by selective biostimulation of the floc-forming population improved the phenol-treating ability of the process. Received: 13 January 2000 / Received revision: 10 March 2000 / Accepted: 7 April 2000     

Fed-batch production of recombinant human calcitonin precursor fusion protein using Staphylococcus carnosus as an expression-secretion system

A pH-auxostatic fed-batch process was developed for the secretory production of a fusion protein consisting of the pro-part of Staphylococcus hyicus lipase and two synthetic human calcitonin (hCT) precursor repeats under the control of a xylose-inducible promotor from Staphylococcus xylosus. Using glycerol as the energy source and pH-controlled addition of yeast extract resulted in the production of 2000 mg l⁻¹ of the fusion protein (420 mg l⁻¹ of the recombinant hCT precursor) within 14 h, reaching 45 g l⁻¹ cell dry mass with Staphylococcus carnosus in a stirred-tank reactor. Product titer and space-time yield (30 mg calcitonin precursor l⁻¹ h⁻¹) were thus improved by a factor of 2, and 4.5, respectively, compared to Escherichia coli expression-secretion systems for the production of calcitonin precursors. Two hundred grams of the fusion protein was secreted by the recombinant S. carnosus on a 150-l scale (scale-up factor of 50) with a minimum use of technical-grade yeast extract (40 mg fusion protein g⁻¹ yeast extract). Received: 18 January 2000 / Received revision: 14 April 2000 / Accepted: 14 April 2000     

Simultaneous bioconversion of glucose and xylose to ethanol by Saccharomyces cerevisiae in the presence of xylose isomerase

Simultaneous isomerisation and fermentation (SIF) of xylose and simultaneous isomerisation and cofermentation (SICF) of a glucose/xylose mixture was carried out by Saccharomyces cerevisiae in the presence of xylose isomerase. The SIF of 50 g l⁻¹ xylose gave an ethanol concentration and metabolic yield of 7.5 g l⁻¹ and 0.36 g (g xylose consumed)⁻¹. These parameters improved to 13.4 g l⁻¹ and 0.40 respectively, when borate was added to the medium. The SICF of a mixture of 50 g l⁻¹ glucose and 50 g l⁻¹ xylose gave an ethanol concentration and metabolic yield of 29.8 g l⁻¹ and 0.42 respectively, in the presence of borate. Temperature modulation from 30 °C to 35 °C during fermentation further enhanced the above parameters to 39 g l⁻¹ and 0.45 respectively. The approach was extended to the bioconversion of sugars present in a real lignocellulose hydrolysate (peanut-shell hydrolysate) to ethanol, with a fairly good yield. Received: 14 May 1999 / Received revision: 27 September 1999 / Accepted: 2 October 1999     

Batch and continuous cultivation of Anaerobiospirillum succiniciproducens for the production of succinic acid from whey

Batch and continuous cultivation of Anaerobiospirillum succiniciproducens were systematically studied for the production of succinic acid from whey. Addition of 2.5 g l⁻¹ yeast extract and 2.5 g l⁻¹ polypeptone per 10 g l⁻¹ whey was most effective for succinic acid production from both treated and nontreated whey. When 20 g l⁻¹ nontreated whey and 7 g l⁻¹ glucose were used as cosubstrates, the yield and productivity of succinic acid reached at the end of fermentation were 95% and 0.46 g (l h)⁻¹, respectively. These values were higher than those obtained using nontreated whey alone [93% and 0.24 g (l h)⁻¹ for 20 g l⁻¹ whey]. Continuous fermentation of A. succiniciproducens at an optimal dilution rate resulted in the production of succinic acid with high productivity [1.35 g (l h)⁻¹], high conversion yield (93%), and higher ratio of succinic acid to acetic acid (5.1:1) from nontreated whey. Received: 23 July 1999 / Received revision: 17 November 1999 / Accepted: 24 December 1999     

Transition rate kinetics from ethanol oxidation to glucose utilisation within a structured model of baker’s yeast

 The transition rate kinetics from ethanol oxidation to glucose utilisation, within a structured model of baker’s yeast, described previously, were experimentally identified. The shift in metabolism has been assessed through glucose pulses during batch growth on ethanol. The influence of glucose concentration (between 0.25 g l^-1 and 0.90 g l^-1) and initial biomass concentration (between 0.61 g l^-1 and 1.44 g l^-1) on the transition rate was determined. The transition rate can not be described by a first-order saturation-type kinetics with respect to glucose only. A corrective term, which takes into account biomass concentration should be included. Received: 28 April 1995/Received revision: 6 July 1995/Accepted: 22 August 1995     

Inhibition of Clostridium butyricum by 1,3-propanediol and diols during glycerol fermentation

1,3-Propanediol inhibition during glycerol fermentation to 1,3-propanediol by Clostridium butyricum CNCM 1211 has been studied. The initial concentration of the 1,3-propanediol affected the growth of the bacterium more than the glycerol fermentation. μ _max was inversely proportional to the initial concentration of 1,3-propanediol (0–65 g l⁻¹). For glycerol at 20 g l⁻¹, the growth and fermentation were completely stopped at an initial 1,3-propanediol concentration of 65 g l⁻¹. However, for an initial 1,3-propanediol concentration of 50 g l⁻¹ and glycerol at 70 g l⁻¹, the final concentration (initial and produced) of 1,3-propanediol reached 83.7 g l⁻¹(1.1 M), with complete consumption of the glycerol. Therefore, during the fermentation, the strain tolerated a 1,3-propanediol concentration higher than the initial inhibitory concentration (65 g l⁻¹). The addition of 1,2-propanediol or 2,3-butanediol (50 g l⁻¹) in the presence of glycerol (50–100 g l⁻¹), showed that 2-diols reduced the μ _max in a similar way to 1,3-propanediol. The measurement of the osmotic pressure of glycerol solutions, diols and diol/glycerol mixtures did not indicate any differences between these compounds. The hypothesis of diol inhibition was discussed. Taking into account the strain tolerance of highly concentrated 1,3-propanediol during fermentation, the fermentation processes for optimising production were considered. Received: 15 November 1999 / Revision received: 1 February 2000 / Accepted: 4 February 2000     

Candida bombicola : production of novel alkyl glycosides based on glucose/2-dodecanol

Candida bombicola produces glycolipids containing sophorose and a glycosidically/esterically bound ω- or (ω−1)-hydroxy C₁₆₍₁₈₎ acid. Here we describe novel glycolipids from this source. Glucose and 2-dodecanol were used for the cultivation of the yeast, one part of the racemic secondary alcohol being connected directly with a glucose or a sophorose unit. A relatively high content of yeast extract, up to 4 g l⁻¹, and subsequently higher biomass concentrations favoured the production of novel products. The provision of 150 g l⁻¹ glucose and 15 g l⁻¹ 2-dodecanol resulted in maximum production of 22 g l⁻¹ novel alkyl glycosides (more than 90% novel products). The molecular structures were analysed by gas chromatography, fast atom bombardment/mass spectrometry, ¹H- and ¹³C-nuclear magnetic resonance and optical rotation studies. Sophorose and glucose were detected as carbohydrate moieties, (S)-(+)-2-dodecanol (88%) was found to be the major lipid moiety. The new glycolipids are suitable biosurfactants, reducing the surface tension of water from 72 mN m⁻¹ to 32–38 mN m⁻¹. Received: 8 December 1997 / Received revision: 19 March 1998 / Accepted: 20 March 1998     

Kinetic models for phycocyanin production by high cell density mixotrophic culture of the microalga Spirulina platensis

Phycocyanin production by high cell density cultivation of Spirulina platensis in batch and fed-batch modes in 3.7-L bioreactors with a programmed stepwise increase in light intensity program was investigated. The results showed that the cell density in fed-batch culture (10.2 g L⁻¹) was 4.29-fold that in batch culture (2.38 g L⁻¹), and the total phycocyanin production in the fed-batch culture (0.795 g L⁻¹) was 3.05-fold that in the batch culture (0.261 g L⁻¹). An unstructured kinetic model to describe the microalga culture system including cell growth, phycocyanin formation, as well as glucose consumption was proposed. The data fitted the models well (r ² > 0.99). Furthermore, based on the kinetic models, the potential effects of light limitation and photoinhibition on cell growth and phycocyanin formation can be examined in depth. The models demonstrated that the optimal light intensity for mixotrophic growth of Spirulina platensis in batch or fed-batch cultures using a 3.7-L bioreactor was 80160 μE m⁻² s⁻¹, and the stepwise increase in light intensity can be replaced by a constant light intensity mode. Received 28 July 1998/ Accepted in revised form 8 October 1998     

Pentachlorophenol biodegradation kinetics of an oligotrophic fluidized-bed enrichment culture

A fluidized-bed reactor (FBR) was used to enrich an aerobic chlorophenol-degrading microbial culture. Long-term continuous-flow operation with low effluent concentrations selected oligotrophic microorganisms producing good-quality effluent for pentachlorophenol(PCP)-contaminated water. PCP biodegradation kinetics was studied using this FBR enrichment culture. The results from FBR batch experiments were modeled using a modified Haldane equation, which resulted in the following kinetic constants: q _max = 0.41 mg PCP mg protein⁻¹ day⁻¹, K _S = 16 μg l⁻¹, K _i = 5.3 mg l⁻¹, and n = 3.5. These results show that the culture has a high affinity for PCP but is also inhibited by relatively low PCP concentrations (above 1.1 mg PCP l⁻¹). This enrichment culture was maintained over 1 year of continuous-flow operation with PCP as the sole source of carbon and energy. During continuous-flow operation, effluent concentrations below 2 μg l⁻¹ were achieved at 268 min hydraulic retention time (t _HR) and 2.5 mg PCP l⁻¹ feed concentration. An increase in loading rate by decreasing t _HR did not significantly deteriorate the effluent quality until a t _HR decrease from 30 min to 21 min resulted in process failure. Recovery from process failure was slow. Decreasing the feed PCP concentration and increasing t _HR resulted in an improved process recovery. Received: 10 October 1996 / Received revision: 21 January 1997 / Accepted: 24 January 1997     

Glucoamylase production in batch, chemostat and fed-batch cultivations by an industrial strain of Aspergillus niger

The Aspergillus niger strain BO-1 was grown in batch, continuous (chemostat) and fed-batch cultivations in order to study the production of the extracellular enzyme glucoamylase under different growth conditions. In the pH range 2.5–6.0, the specific glucoamylase productivity and the specific growth rate of the fungus were independent of pH when grown in batch cultivations. The specific glucoamylase producivity increased linearly with the specific growth rate in the range 0–0.1 h⁻¹ and was constant in the range 0.1–0.2 h⁻¹. Maltose and maltodextrin were non-inducing carbon sources compared to glucose, and the maximum specific growth rate was 0.19 ± 0.02 h⁻¹ irrespective of whether glucose or maltose was the carbon source. In fed-batch cultivations, glucoamylase titres of up to 6.5 g l⁻¹ were obtained even though the strain contained only one copy of the glaA gene. Received: 5 May 1999 / Received revision: 7 September 1999 / Accepted: 17 September 1999     

Kinetic models for astaxanthin production by high cell density mixotrophic culture of the microalga Haematococcus pluvialis

High cell density cultivation of Haematococcus pluvialis for astaxanthin production was carried out in batch and fed-batch modes in 3.7-L bioreactors with stepwise increased light intensity control mode. A high cell density of 2.65 g L⁻¹ (batch culture) or 2.74 g L⁻¹ (fed-batch culture) was obtained, and total astaxanthin production in the fed-batch culture (64.36 mg L⁻¹) was about 20.5% higher than in the batch culture (53.43 mg L⁻¹). An unstructured kinetic model to describe the microalga culture system including cell growth, astaxanthin formation, as well as sodium acetate consumption was proposed. Good agreement was found between the model predictions and experimental data. The models demonstrated that the optimal light intensity for mixotrophic growth of H. pluvialis in batch or fed-batch cultures in a 3.7-L bioreactor was 90–360 μmol m⁻² s⁻¹, and that the stepwise increased light intensity mode could be replaced by a constant light intensity mode. Received 24 December 1998/ Accepted in revised form 23 April 1999     

Enhanced l-lysine production in threonine-limited continuous culture of Corynebacterium glutamicum by using gluconate as a secondary carbon source with glucose

In order to improve the production rate of l-lysine, a mutant of Corynebacterium glutamicum ATCC 21513 was cultivated in complex medium with gluconate and glucose as mixed carbon sources. In a batch culture, this strain was found to consume gluconate and glucose simultaneously. In continuous culture at dilution rates ranging from 0.2 h⁻¹ to 0.25 h⁻¹, the specific l-lysine production rate increased to 0.12 g g⁻¹ h⁻¹ from 0.1 g g⁻¹ h⁻¹, the rate obtained with glucose as the sole carbon source [Lee et al. (1995) Appl Microbiol Biotechnol 43:1019–1027]. It is notable that l-lysine production was observed at higher dilution rates than 0.4 h⁻¹, which was not observed when glucose was the sole carbon source. The positive effect of gluconate was confirmed in the shift of the carbon source from glucose to gluconate. The metabolic transition, which has been characterized by decreased l-lysine production at the higher glucose uptake rates, was not observed when gluconate was added. These results demonstrate that the utilization of gluconate as a secondary carbon source improves the maximum l-lysine production rate in the threonine-limited continuous culture, probably by relieving the limiting factors in the lysine synthesis rate such as NADPH supply and/or phosphoenolpyruvate availability. Received: 16 May 1997 / Received revision: 28 August 1997 / Accepted: 29 August 1997     

Benzene/toluene/p-xylene degradation. Part II. Effect of substrate interactions and feeding strategies in toluene/benzene and toluene/p-xylene fermentations in a partitioning bioreactor

A two-phase aqueous/organic partitioning bioreactor scheme was used to degrade mixtures of toluene and benzene, and toluene and p-xylene, using simultaneous and sequential feeding strategies. The aqueous phase of the partitioning bioreactor contained Pseudomonas sp. ATCC 55595, an organism able to degrade benzene, toluene and p-xylene simultaneously. An industrial grade of oleyl alcohol served as the organic phase. In each experiment, the organic phase of the bioreactor was loaded with 10.15 g toluene, and either 2.0 g benzene or 2.1 g p-xylene. The resulting aqueous phase concentrations were 50 mg/l, 25 mg/l and 8 mg/l toluene, benzene and p-xylene respectively. The simultaneous fermentation of benzene and toluene consumed these compounds at volumetric rates of 0.024 g l⁻¹ h⁻¹ and 0.067 g l⁻¹ h⁻¹, respectively. The simultaneous fermentation of toluene and p-xylene consumed these xenobiotics at volumetric rates of 0.066 g l⁻¹ h⁻¹ and 0.018 g l⁻¹ h⁻¹, respectively. A sequential feeding strategy was employed in which toluene was added initially, but the benzene or p-xylene aliquot was added only after the cells had consumed half of the initial toluene concentration. This strategy was shown to improve overall degradation rates, and to reduce the stress on the microorganisms. In the sequential fermentation of benzene and toluene, the volumetric degradation rates were 0.056 g l⁻¹ h⁻¹ and 0.079 g l⁻¹ h⁻¹, respectively. In the toluene/p-xylene sequential fermentation, the initial toluene load was consumed before the p-xylene aliquot was consumed. After 12 h in which no p-xylene degradation was observed, a 4.0-g toluene aliquot was added, and p-xylene degradation resumed. Excluding that 12-h period, the microbes consumed toluene and p-xylene at volumetric rates of 0.074 g l⁻¹ h⁻¹ and 0.025 g l⁻¹ h⁻¹, respectively. Oxygen limitation occurred in all fermentations during the rapid growth phase. Received: 16 November 1998 / Received revision: 29 March 1999 / Accepted: 9 April 1999     

High alcohol production by repeated batch fermentation using an immobilized osmotolerant Saccharomyces cerevisiae

A repeated batch fermentation system was used to produce ethanol using an osmotolerant Saccharomyces cerevisiae (VS₃) immobilized in calcium alginate beads. For comparison free cells were also used to produce ethanol by repeated batch fermentation. Fermentation was carried for six cycles with 125, 250 or 500 beads using 150, 200 or 250 g glucose L⁻¹ at 30°C. The maximum amount of ethanol produced by immobilized VS₃ using 150 g L⁻¹ glucose was only 44 g L⁻¹ after 48 h, while the amount of ethanol produced by free cells in the first cycle was 72 g L⁻¹. However in subsequent fed batch cultures more ethanol was produced by immobilized cells compared to free cells. The amount of ethanol produced by free cells decreased from 72 g L⁻¹ to 25 g L⁻¹ after the fourth cycle, while that of immobilized cells increased from 44 to 72 g L⁻¹. The maximum amount of ethanol produced by immobilized VS₃ cells using 150, 200 and 250 g glucose L⁻¹ was 72.5, 93 and 87 g ethanol L⁻¹ at 30°C. Journal of Industrial Microbiology & Biotechnology (2000) 24, 222–226. Received 16 September 1999/ Accepted in revised form 22 December 1999     

Glycolipids of the smut fungus Ustilago maydis from cultivation on renewable resources

When grown on vegetable oils and their derivatives, the smut fungus Ustilago maydis (DSM 4500 and ATCC 14826) produces several glycolipids under nitrogen-limiting conditions. With 45 g l⁻¹ sunflower oil fatty acids (technical grade) a yield of 30 g l⁻¹ glycolipid was achieved. The resulting mixture contained predominantly mannosylerythritol lipids together with smaller amounts of cellobiose lipids. The production of the more polar cellobiose lipids was enhanced when glucose was used as carbon source. The molecular structure of the main components of the glycolipid mixture were elucidated by a combination of NMR spectroscopic and mass-spectrometric techniques. Received: 22 June 1998 / Received revision: 11 September 1998 / Accepted: 13 September 1998     

Isolation of <Emphasis Type="Italic">Aspergillus oryzae</Emphasis> mutants for heterologous protein production from a double proteinase gene disruptant

The recombinant Pichia pastoris harboring an improved methionine adenosyltransferase (MAT) shuffled gene was employed to biosynthesize S-adenosyl-l-methionine (SAM). Two l-methionine (l-Met) addition strategies were used to supply the precursor: the batch addition strategy (l-Met was added separately at three time points) and the continuous feeding strategies (l-Met was fed continuously at the rate of 0.1, 0.2, and 0.5 g l⁻¹ h⁻¹, respectively). SAM accumulation, l-Met conversion rate, and SAM productivity with the continuous feeding strategies were all improved over the batch addition strategy, which reached 8.46 ± 0.31 g l⁻¹, 41.7 ± 1.4%, and 0.18 ± 0.01 g l⁻¹ h⁻¹ with the best continuous feeding strategy (0.2 g l⁻¹ h⁻¹), respectively. The bottleneck for SAM production with the low l-Met feeding rate (0.1 g L⁻¹ h⁻¹) was the insufficient l-Met supply. The analysis of the key enzyme activities indicated that the tricarboxylic acid cycle and glycolytic pathway were reduced with the increasing l-Met feeding rate, which decreased the adenosine triphosphate (ATP) synthesis. The MAT activity also decreased as the l-Met feeding rate rose. The reduced ATP synthesis and MAT activity were probably the reason for the low SAM accumulation when the l-Met feeding rate reached 0.5 g l⁻¹ h⁻¹.     

Rhamnose lipids – biosynthesis, microbial production and application potential

Biosurfactants containing rhamnose and β-hydroxydecanoic acid and called rhamnolipids are reviewed with respect to microbial producers, their physiological role, biosynthesis and genetics, and especially their microbial overproduction, physicochemical properties and potential applications. With Pseudomonas species, more than 100 g l⁻¹ rhamnolipids were produced from 160 g l⁻¹ soybean oil at a volumetric productivity of 0.4 g l⁻¹h⁻¹. The individual rhamnolipids are able to lower the surface tension of water from 72 mN m⁻¹ to 25–30 mN m⁻¹ at concentrations of 10–200 mg l⁻¹. After initial testing, rhamnolipids seem to have potential applications in combating marine oil pollution, removing oil from sand and in combating zoosporic phytopathogens. Rhamnolipids are also a source of l-rhamnose, which is already used for the industrial production of high-quality flavor components. Received: 1 July 1998 / Received revision: 11 September 1998 / Accepted: 13 September 1998     

An attempt to channel the transformation of vanillic acid into vanillin by controlling methoxyhydroquinone formation in Pycnoporus cinnabarinus with cellobiose

The effects of adding cellobiose on the transformation of vanillic acid to vanillin by two strains of Pycnoporus cinnabarinus MUCL39532 and MUCL38467 were studied. When maltose was used as the carbon source in the culture medium, very high levels of methoxyhydroquinone were formed from vanillic acid. When cellobiose was used as the carbon source and/or added to the culture medium of P. cinnabarinus strains on day 3 just before vanillic acid was added, it channelled the vanillic acid metabolism via the reductive route leading to vanillin. Adding 3.5 g l⁻¹ cellobiose to 3-day-old maltose cultures of P. cinnabarinus MUCL39532 and 2.5 g l⁻¹ cellobiose to 3-day-old cellobiose cultures of P. cinnabarinus MUCL38467, yielded 510 mg l⁻¹ and 560 mg l⁻¹ vanillin with a molar yield of 50.2 % and 51.7 % respectively. Cellobiose may either have acted as an easily metabolizable carbon source, required for the reductive pathway to occur, or as an inducer of cellobiose:quinone oxidoreductase, which is known to inhibit vanillic acid decarboxylation. Received: 24 July 1996 / Received revision: 29 November 1996 / Accepted: 29 November 1996     

Poly-(3-hydroxybutyrate) production from whey by high-density cultivation of recombinant Escherichia coli

Recombinant Escherichia coli strain GCSC 6576, harboring a high-copy-number plasmid containing the Ralstonia eutropha genes for polyhydroxyalkanoate (PHA) synthesis and the E. coli ftsZ gene, was employed to produce poly-(3-hydroxybutyrate) (PHB) from whey. pH-stat fed-batch fermentation, using whey powder as the nutrient feed, produced cellular dry weight and PHB concentrations of 109 g l⁻¹ and 50 g l⁻¹ respectively in 47 h. When concentrated whey solution containing 210 g l⁻¹ lactose was used as the nutrient feed, cellular dry weight and PHB concentrations of 87 g l⁻¹ and 69 g l⁻¹ respectively could be obtained in 49 h by pH-stat fed-batch culture. The PHB content was as high as 80% of the cellular dry weight. These results suggest that cost-effective production of PHB is possible by fed-batch culture of recombinant E. coli using concentrated whey solution as a substrate. Received: 19 December 1997 / Received revision: 17 March 1998 / Accepted: 20 March 1998     

Effect of ammonia on the anaerobic degradation of protein by a mesophilic and thermophilic biowaste population

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