Effect of chlorinated aliphatic hydrocarbons on the acetoclastic methanogenic activity of granular sludge

The toxicity of chlorinated aliphatic hydrocarbons on acetoclastic methanogens in anaerobic granular sludge was determined using a standardized anaerobic bioassay method. Most of the chloroaliphatics tested were strong inhibitors of methanogenesis. Tri- and tetrachloride derivatives of methane and ethane were the most highly toxic compounds tested, with concentrations of less than 18 mg/l resulting in 50% inhibition (IC₅₀) of the methanogenic activity. Dichlorinated compounds were less toxic, with IC₅₀ values ranging from 40 mg/l to 100 mg/l. On the other hand, perchlorinated derivatives of ethane and ethene were scarcely inhibitory at concentrations near their maximum water solubility. The toxicity caused by chlorinated aliphatic hydrocarbons was reversible. The comparison of structurally related compounds indicated that unsaturated chloroaliphatics were less toxic than their saturated counterparts. A reverse correlation between the electric dipole moment of these compounds and their methanogenic toxicity is discussed. Received: 9 July 1996 / Received revision: 11 October 1996 / Accepted: 18 October 1996     

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     

Inhibition of mesophilic solid-substrate anaerobic digestion by ammonia nitrogen

This work focused on determining the effects of ammonia-nitrogen supplementation on the mesophilic solid-substrate anaerobic digestion of municipal wastes and waste activated sludge (biosolids). Bench-scale, semi-continuous, mesophilic reactors were operated with a 21-day mass-retention time and dosed with NH₄Cl, such that the corresponding chemical O₂ demand (COD)/N ratios in their feeds were 90, 80, 65 and 50 (reactors R1 or control, R2, R3 and R4 respectively). Reactor performance was evaluated in terms of the efficiency of volatile solid removal (efficiency for short), biogas productivity, methane content in the biogas, pH and volatile organic acid contents, among other monitoring and analytical parameters. The feedstock was a mixture of urban wastes with biosolids. It was found that the process performance deteriorated at increasing dosages of ammonia N, the process practically ceasing at COD/N = 50 (R4). Inhibition was characterized by efficiency and biogas productivity decreases and a more sudden drop of methane content in biogas and pH. A significant rise of propionic, butyric and valeric acid was found in reactors receiving the highest doses of ammonia N (R3 and R4). This suggested that inhibition of the syntrophic bacteria present in the anaerobic consortia also occurred. Luong and Pearson inhibition models were fitted to the data. Both models represented very well the acute effects of N supplementation on solid-substrate anaerobic digestion. However, the Luong model could also represent the process ceasing at a critical ammonia N concentration of 2800 mg/kg mixed solids. Received: 12 April 1996 / Received revision: 23 July 1996 / Accepted: 5 August 1996     

Oxygen consumption and sulfide detoxification in the lugworm Arenicola marina (L.) at different ambient oxygen partial pressures and sulfide concentrations

The lugworm Arenicola marina is a typical inhabitant of intertidal flats. In its L-shaped burrow the animal is exposed to varying concentrations of O₂ and toxic sulfide depending on the tides. The lugworm is able to detoxify sulfide through its oxidation to thiosulfate. When exposed to declining O₂ tensions Arenicola marina reacted as an oxyconformer. In the presence of 25 μmol · l⁻¹ sulfide the respiration was not affected. In contrast, the lugworm consumed significantly less O₂ at any Po₂ in the presence of 200 μmol · l⁻¹ sulfide. Without sulfide anaerobic metabolism started at a Po₂ of approximatedly 10 kPa. Even at high O₂ tensions animals exposed to sulfide produced significantly more anaerobic metabolites compared with the controls. Accordingly the critical value Pc_M, the ambient Po₂ below which anaerobic metabolism starts, was shifted towards normoxia. Since O₂ supply was sufficient for aerobic metabolism, anaerobiosis was induced by sulfide. An influx of sulfide was observed at 25 as well as at 200 μmol · l⁻¹ sulfide. The main product of sulfide detoxification in the lugworm was thiosulfate. Its synthesis increased with ambient Po₂ and depended on the sulfide concentration. Sulfide and thiosulfate were detected in the coelomic fluid, the blood, and the body wall of Arenicola marina. Only about 2% of the ambient O₂ was used for sulfide detoxification at 25 μmol · l⁻¹ sulfide and about 50% at 200 μmol · l⁻¹ sulfide, respectively. Even at the low sulfide concentration Arenicola marina's capacity to detoxify sulfide was too low to maintain a complete aerobic metabolism. Accepted: 19 February 1997     

Inhibition of methane production from whey by heavy metals – protective effect of sulfide

A whey solution was used as a substrate for methane production in an anaerobic fixed-bed reactor. At a hydraulic retention time of 10 days, equivalent to a space loading of 3.3 kg (m³day)⁻¹, 90% of the chemical oxygen demand was converted to biogas. Only a little propionate remained in the effluent. Toxicity tests with either copper chloride, zinc chloride or nickel chloride were performed on effluent from the reactor. Fifty per cent inhibition of methanogenesis was observed in the presence of ≥10 mg CuCl₂ l⁻¹≥40 mg ZnCl₂ l⁻¹ and ≥60 mg NiCl₂ l⁻¹, respectively. After exposure to Cu²⁺, Zn²⁺ or Ni²⁺ ions for 12 days, complete recovery of methanogenesis by equimolar sulfide addition was possible upon prolonged incubation. Recovery failed, however, for copper chloride concentrations ≥40 mg l⁻¹. If the sulfide was added simultaneously with the three heavy metal salts, methanogenesis was only slightly retarded and the same amount of methane as in non-inhibited controls was reached either 1 day (40 mg ZnCl₂ l⁻¹) or 2 days later (10 mg CuCl₂ l⁻¹). Up to 60 mg NiCl₂ l⁻¹ had no effect if sulfide was present. Sulfide presumably precipitated the heavy metals as metal sulfides and by this means prevented heavy metal toxicity. Received: 8 October 1999 / Received revision: 3 January 2000 / Accepted: 4 January 2000     

Anaerobic digestion of secondary residuals from an anaerobic bioreactor at a brewery to enhance bioenergy generation

Many beer breweries use high-rate anaerobic digestion (AD) systems to treat their soluble high-strength wastewater. Biogas from these AD systems is used to offset nonrenewable energy utilization in the brewery. With increasing nonrenewable energy costs, interest has mounted to also digest secondary residuals from the high-rate digester effluent, which consists of yeast cells, bacteria, methanogens, and small (hemi)cellulosic particles. Mesophilic (37 °C) and thermophilic (55 °C) lab-scale, low-rate continuously-stirred anaerobic digestion (CSAD) bioreactors were operated for 258 days by feeding secondary residuals at a volatile solids (VS) concentration of ∼40 g l⁻¹. At a hydraulic retention time (HRT) of 15 days and a VS loading rate of 2.7 g VS l⁻¹ day⁻¹, the mesophilic bioreactor showed an average specific volumetric biogas production rate of 0.88 l CH₄ l⁻¹ day⁻¹ and an effluent VS concentration of 22.2 g VS l⁻¹ (43.0% VS removal efficiency) while the thermophilic bioreactor displayed similar performances. The overall methane yield for both systems was 0.21 l CH₄ g⁻¹ VS fed and 0.47–0.48 l CH₄ g⁻¹ VS removed. A primary limitation of thermophilic digestion of this protein-rich waste is the inhibition of methanogens due to higher nondissociated (free) ammonia (NH₃) concentrations under similar total ammonium (NH₄ ⁺) concentrations at equilibrium. Since thermophilic AD did not result in advantageous methane production rates or yields, mesophilic AD was, therefore, superior in treating secondary residuals from high-rate AD effluent. An additional digester to convert secondary residuals to methane may increase the total biogas generation at the brewery by 8% compared to just conventional high-rate digestion of brewery wastewater alone. JIMB-2008: BioEnergy—Special issue.     

Isolation and characterization of Rhodococcus rhodochrous for the degradation of the wastewater component 2-hydroxybenzothiazole

2-Hydroxybenzothiazole (OBT) is present in wastewaters from the industrial production of the rubber vulcanization accelerator 2-mercaptobenzothiazole (MBT). We have achieved the first isolation of axenic bacterial cultures capable of the degradation of OBT and growth on this substrate as the sole source of carbon, nitrogen and energy. All isolates had similar characteristics corresponding to one particular isolate, which was studied in more detail and identified as Rhodococcus rhodochrous. The strains were also capable of degrading benzothiazole (BT) but not MBT or benzothiazole-2-sulphonate (BTSO₃). OBT was degraded at a concentration of up to 600 mg · l⁻¹. BT was toxic above 300 mg · l⁻¹. MBT inhibited OBT degradation. Growth on OBT was not significantly different at pH values of between 6.3 and 7.9 or salt concentrations between 1 % and 3 %. In shake flasks the cells clumped together, which resulted in a lower rate of oxygen transfer and slower degradation as compared to cells grown on OBT in a stirred reactor. Received: 22 August 1996 / Received revision: 29 November 1996 / Accepted: 29 November 1996     

Insulin and glucagon immunoreactivity during high-intensity exercise under opiate blockade

Eight fit men [maximum oxygen consumption (O_2max) 64.6 (1.9) ml · kg⁻¹ · min⁻¹, aged 28.3 (1.7) years (SE in parentheses) were studied during two treadmill exercise trials to determine the effect of endogenous opioids on insulin and glucagon immunoreactivity during intense exercise (80% O_2max). A double-blind experimental design was used with subjects undertaking the two exercise trials in counterbalanced order. Exercise trials were 20 min in duration and were conducted 7 days apart. One exercise trial was undertaken following administration of naloxone (N; 1.2 mg; 3 ml) and the other after receiving a placebo (P; 0.9% NaCl saline; 3 ml). Prior to each experimental trial a flexible catheter was placed into an antecubital vein and baseline blood samples were collected. Immediately after, each subject received either a N or P bolus injection. Blood samples were also collected after 20 min of continuous exercise (running). Glucagon was higher (P < 0.05), while insulin was lower (P < 0.05), during exercise compared with pre-exercise values in both trials. However, glucagon was higher (P < 0.05) in the P than in the N exercise trial [141.4 (8.3) ng · l⁻¹ vs 127.2 (7.6) ng · l⁻¹]. There were no differences in insulin during exercise between the P and N trials [50.2 (4.3) pmol · l⁻¹ vs 43.8 (5) pmol · l⁻¹]. These data suggest that endogenous opioids may augment the glucagon response during intense exercise. Accepted: 15 June 1996     

Mesophilic and thermophilic anaerobic digestion of source-sorted organic wastes: effect of ammonia on glucose degradation and methane production

The wet organic fraction of household wastes was digested anaerobically at 37 °C and 55 °C. At both temperatures the volatile solids loading was increased from 1 g l⁻¹ day⁻¹ to 9.65 g l⁻¹ day⁻¹, by reducing the nominal hydraulic retention time from 93 days to 19 days. The volatile solids removal in the reactors at both temperatures for the same loading rates was in a similar range and was still 65% at 19 days hydraulic retention time. Although more biogas was produced in the thermophilic reactor, the energy conservation in methane was slightly lower, because of a lower methane content, compared to the biogas of the mesophilic reactor. The slightly lower amount of energy conserved in the methane of the thermophilic digester was presumably balanced by the hydrogen that escaped into the gas phase and thus was no longer available for methanogenesis. In the thermophilic process, 1.4 g/l ammonia was released, whereas in the mesophilic process only 1 g/l ammonia was generated, presumably from protein degradation. Inhibition studies of methane production and glucose fermentation revealed a K _i (50%) of 3 g/l and 3.7 g/l ammonia (equivalent to 0.22 g/l and 0.28 g/l free NH₃) at 37 °C and a K _i (50%) of 3.5 g/l and 3.4 g/l ammonia (equivalent to 0.69 g/l and 0.68 g/l free NH₃) at 55 °C. This indicated that the thermophilic flora tolerated at least twice as much of free NH₃ than the mesophilic flora and, furthermore, that the thermophilic flora was able to degrade more protein. The apparent ammonia concentrations in the mesophilic and in the thermophilic biowaste reactor were low enough not to inhibit glucose fermentation and methane production of either process significantly, but may have been high enough to inhibit protein degradation. The data indicated either that the mesophilic and thermophilic protein degraders revealed a different sensitivity towards free ammonia or that the mesophilic population contained less versatile protein degraders, leaving more protein undegraded. Received: 26 March 1997 / Received revision: 13 May 1997 / Accepted: 19 May 1997     

The use of silica gel prepared by sol-gel method and polyurethane foam as microbial carriers in the continuous degradation of phenol

A mixed microbial culture was immobilized by entrapment into silica gel (SG) and entrapment/ adsorption on polyurethane foam (PU) and ceramic foam. The phenol degradation performance of the SG biocatalyst was studied in a packed-bed reactor (PBR), packed-bed reactor with ceramic foam (PBRC) and fluidized-bed reactor (FBR). In continuous experiments the maximum degradation rate of phenol (q _s ^max) decreased in the order: PBRC (598 mg l⁻¹h⁻¹) > PBR (PU, 471 mg l⁻¹h⁻¹) > PBR (SG, 394 mg l⁻¹h⁻¹) > FBR (PU, 161 mg l⁻¹h⁻¹) > FBR (SG, 91 mg l⁻¹ h⁻¹). The long-term use of the SG biocatalyst in continuous phenol degradation resulted in the formation of a 100–200 μm thick layer with a high cell density on the surface of the gel particles. The abrasion of the surface layer in the FBR contributed to the poor degradation performance of this reactor configuration. Coating the ceramic foam with a layer of cells immobilized in colloidal SiO₂ enhanced the phenol degradation efficiency during the first 3 days of the PBRC operation, in comparison with untreated ceramic packing. Received: 2 December 1999 / Revision received: 2 February 2000 / Accepted: 4 February 2000     

An Analysis of the Energy Potential of Anaerobic Digestion of Agricultural By-Products and Organic Waste

Anaerobic digestion is a promising option for recycling agricultural by-products and some organic wastes. While both agricultural by-products and wastes have no direct commercial value, their management is both complicated and costly. One option to simplify by-product management and reduce the costs associated with biogas plant feedstock is to substitute dedicated crops with vegetal by-products. Given that the chemical composition of some of these by-products can differ considerably from more typical biogas plant feedstock (such as maize silage), more complete knowledge of these alternatives to produce environmentally friendly energy is warranted. To this end, batch trials under mesophilic conditions were conducted to evaluate the potential biogas yield of many agricultural by-products: maize stalks, rice chaff, wheat straw, kiwi fruit, onions, and two expired organic waste products (dairy and dry bread) from the retail mass-market. Among the considered biomasses, the highest methane producer was the expired dairy product mixture, which yielded 554 l_NCH₄ kg⁻¹ volatile solids (VS). Maize stalks and wheat straw produced the lowest yields of 214 and 285 l_NCH₄ kg⁻¹VS, respectively. An assessment of the biogas and methane yields of each biomass was also undertaken to account for the specific chemical composition of each biomass as it can affect the anaerobic digestion operating system. Finally, the total Italian green energy production that might be derived from feeding all these biomasses to a biogas digester was estimated, in order to understand its potential impact.     

The use of amiloride to uncouple branchial sodium and proton fluxes in the brown trout, Salmo trutta

Resting proton, ammonium and sodium fluxes in Salmo trutta were 492.6 ± 19.5 (n = 29); 122.9 ± 34.2 (n = 28) and 277.1 ± 18.5 (n = 50) μmol · kg⁻¹ · h⁻¹, respectively. The resting transepithelial potential was found to be composed of three successive potentials, the outermost averaging −7.36 ± 0.19mV, the second, −14.3 ± 1.4 mV and the third −37 ± 1.7 mV. Amiloride inhibits the proton, ammonium and sodium fluxes in a dose-dependent manner at concentrations of 0.5 mmol · 1⁻¹ and 0.1 mmol · l⁻¹, but at 0.01 mmol · l⁻¹, proton and ammonium fluxes remained at control levels whilst the sodium was reduced to 70.59 ± 7.29 μmol · kg⁻¹ · h⁻¹. The trans-epithelial potential was effected in a bi-phasic manner by 0.5 mmol · l⁻¹ amiloride. An initial hyperpolarisation of ca. 6 mV was followed by a sustained depolarisation of ca. 14 mV (towards zero) which persisted until the amiloride was washed off the gill. The initial hyperpolarisation was thought to reflect a rapid inhibition of a positive inward sodium current and the subsequent depolarisation was due to the inhibition of a positive outward current (proton) which would abolish the transepithelial potential. However, at 0.01 mmol ·  l⁻¹ only the hyperpolarisation was seen, due to the inhibition of only the inward sodium current. Acetazolamide (0.1 mmol · l⁻¹) was found to have no significant effect on the proton, ammonium and sodium fluxes. These results indicate that the proton and sodium fluxes across the gill of the freshwater trout are not tightly linked. While this suggests that the trout gill resembles the model of Ehrenburg et al. (1985) of sodium uptake in frog skin, the apical potentials measured in the pavement epithelial cell(s) are too low to account for sodium uptake unless the activity of the sodium in the cells is very low. Accepted: 8 August 1996     

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     

Active calcium ion transport in Xenopuslaevis skin

The skin of intact, free-swimming Xenopus laevis transports Ca²⁺ inwardly in a manner that is proportional to the external [Ca²⁺] up to about 0.3 mmol · l⁻¹, saturates above 0.3 mmol · l⁻¹, and is opposed to the electrochemical gradient. Efflux is relatively constant at external concentrations between 0.016 and 0.6 mmol · l⁻¹; net flux which is negative below 0.125 mmol · l⁻¹ becomes positive above this external [Ca²⁺]. Allometric analysis suggests that both Ca²⁺ influx and efflux scale to the 2/3 power approximately like surface area. There were no significant differences in influx between summer and fall animals; however, efflux was greater in the fall and this resulted in a change from positive balance in the summer to negative balance in the fall. Isolated skins were shown to support a Ca²⁺ uptake rate of nearly 30 nmol · cm⁻² · h⁻¹. The phenylalkylamine verapamil in the apical bathing solution significantly inhibited this at 25 μmol · l⁻¹. The benzothiazepine diltiazem was also effective at 50 μmol · l⁻¹ while the dihydropyradine nifedipine was ineffective up to 100 μmol · l⁻¹. The inorganic ion La³⁺ was effective at blocking Ca²⁺ uptake at 300 μmol · l⁻¹; Ni²⁺ was also effective at 500 μmol · l⁻¹ but Co²⁺ was ineffective up to 500 μmol · l⁻¹. These results suggest that apical calcium channels in Xenopuslaevis skin have properties similar to mammalian L-channels and fish gill Ca²⁺ channels. Accepted: 23 January 1997     

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     

Haemolymph oxygen transport and acid-base status in Glyptonotus antarcticus Eights

Haemolymph samples were withdrawn from routinely active male intermoult Glyptonotus held at 0 ± 0.5°C, and analysed for blood-gas and acid-base variables. In both the arterialised (a) and venous (v) haemolymph, over 50% of the oxygen was transported as dissolved oxygen at PaO₂ and PvO₂ levels of 12.0 ± 1.15 and 7.70 ± 1.89 kPa, respectively. The maximum oxygen-carrying capacity of the haemocyanin (C^maxHcO₂) was relatively low at 0.19 ± 0.05 mmol l⁻¹, accompanied by relatively low protein and [Cu²⁺] levels indicating low circulating haemocyanin concentrations. Arterialised haemolymph had a mean pH of 7.88 ± 0.02(6) at a PCO₂ of 0.12 ± 0.01(6) kPa and a bicarbonate level of 12.95 ± 0.80(6) mequiv l⁻¹ with small differences in PCO₂ and pH between arterial and venous haemolymph. The non-bicarbonate buffering capacity of Glyptonotus haemolymph was low at −2.0 mequiv l⁻¹ HCO₃ ⁻ pH unit⁻¹. Haemolymph [l-lactate] and [d-glucose] levels were similar at < 1 mmol l⁻¹ in animals held in the laboratory and those sampled in Antarctica. The blood-gas and acid-base status of Glyptonotus haemolymph may be a reflection of the low and stable temperatures experienced by this Antarctic crustacean. Received: 14 August 1996 / Accepted: 3 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     

Characteristics of dipeptide transport in pig jejunum in vitro

 Characteristics of dipeptide transport in pig jejunum were investigated in vitro by applying the Ussing-chamber technique and mucosal uptake studies. Addition of both glycyl-l-glutamine and glycyl-l-sarcosine (20 mmol · l⁻¹) to the mucosal buffer solution significantly increased the short-circuit current by 2.60 ± 0.15 and 1.57 ± 0.20 μeq · cm⁻² · h⁻¹, respectively. Concentration-dependent changes in short-circuit current followed Michaelis-Menten kinetics with similar affinity constants for both dipeptides. From unidirectional flux rates for radiolabelled glycyl-l-sarcosine, a net flux rate for glycyl-l-sarcosine of 49.8 ± 6.7 nmol · cm⁻² · h⁻¹ was calculated. In mucosal uptake experiments, the apical influx of ¹⁴C-labelled glycyl-l-sarcosine into isolated porcine mucosa was pH dependent and significantly inhibited by glycyl-l-glutamine. Moreover, RT-PCR studies with primers derived from rabbit PepT1 identified two PCR fragments of identical size to rabbit PepT1 from pig intestinal mRNA preparations. In conclusion, our studies revealed key features of mammalian intestinal peptide transporters and give evidence for a PepT1-like transporter in the pig jejunum that could significantly contribute to the overall amino acid absorption from the gut. Accepted: 30 June 1999     

Nitrate removal from drinking water using a membrane-fixed biofilm reactor

Biological treatment of drinking water is a cost-effective alternative to conventional physico/chemical processes. A new concept was tested to overcome the main disadvantage of biological denitrification, the intensive post-treatment process to remove microorganisms and remnant carbon source. The biological reaction zone and carbon supply were separated from the raw water stream by a nitrate-permeable membrane. Denitrification takes place in a biofilm, which is immobilized at the membrane. In a series of bench-scale runs, different types of membranes and reactor configurations were investigated. The best denitrification rates achieved were 1230 mg NO₃ ⁻-N m⁻² day⁻¹. In one run, raw water containing 100 mg NO₃ ⁻ l⁻¹ was completely freed from nitrate. The membrane and the attached biofilm also represent a barrier against the passage of the C source and nutrients into the raw water. At concentrations of 20 mg l⁻¹ ethanol and 15 mg l⁻¹ phosphate in the bioreactor no diffusion through the membrane into the treated water was observed. Without any post-treatment, the effluent met nearly all the relevant criteria for drinking water; only the colony count was slightly increased. Received: 18 December 1996 / Received last revision: 14 April 1997 / Accepted: 19 April 1997     

Cellular volume regulation in freshwater bivalves

The effect of ambient osmolality on the height of lateral ciliated cells from the gills of two freshwater bivalve species (Dreissena polymorpha, Toxolasma texasensis) was directly observed microscopically. The addition of 1 mmol · l⁻¹ KCl to an artificial pondwater (APW) superfusion medium resulted in an increase in cell height. When the superfusion solution was made hyperosmotic (∼90 mmol · kg⁻¹ H₂O) by the addition of 45 mmol · l⁻¹ NaCl to APW, the cell height decreased by about 20–30% and there was no evidence of a regulatory volume increase over 20–30 min. In contrast, when 1 mmol · l⁻¹ KCl was added to the hyperosmotic medium the cell height always partially (40–50%) recovered. When the gill tissue was returned to APW following the hyperosmotic treatment the cells returned to the original cell height. Bivalve gills superfused with the hyperosmotic NaCl and KCl solution in the presence of 1 mmol · l⁻¹ ouabain experienced a similar 25% decrease in cell height. When the ouabain-treated tissues were returned to APW the cells swelled, overshooting the original cell height. These results indicate these freshwater bivalves have a limited ability for cellular volume regulation using inorganic ions, but depend on a suitable balance of Na⁺ and K⁺ in the environment to effect regulatory volume changes. Accepted: 17 October 1997