Anaerobic digestion of cattle waste at mesophilic and thermophilic temperatures

Methanogenesis was studied using stirred, bench-top fermentors of 3-1 working volume fed on a semi-continuous basis with waste obtained from cattle fed a high grain, finishing diet. Digestion was carried out at 40 and 60°C. CH₄ production was 11.8, 18.3, 61.9 and 84.5% higher in the thermophilic than the mesophilic digestor at the 3, 6, 9 and 12 g volatile solids (VS) l^–1 reactor volume loading rates, respectively. When compared on an energetic basis CH₄ production was 7.4, 18.3, 72.9 and 107.3 kJ day higher in the thermophilic than the mesophilic digestor. CH₄ production decreased more rapidly with each increase in VS loading rate and decrease in retention time (RT) in the mesophilic than the thermophilic digestor. When expressed as l g^–1 VS fed or as kJ kJ^–1 fed, the amount of CH₄ was 49% less at the highest compared to the lowest loading rate in the mesophilic digestor. In the thermophilic digestor the decrease was only 16%. Propionate accumulated in the mesophilic digestor at the two highest loading rates, reaching concentrations of about 50 mM, but were only about 13 mM in the thermophilic digestor. Isobutyrate, isovalerate plus 2-methylbutyrate, and valerate also accumulated at the higher loading rates.     

The influence of sulphate on substrate utilization in a thermophilic sewage sludge digestor

Summary Bacterial sulphate reduction and the interaction between sulphate reduction and methane production was studied in an unadapted and sulphate-adapted thermophilic anaerobic sludge digestor. Addition of sulphate to a concentration of 5 mm (100 times the background level) did not influence gas production or volatile fatty acid concentration compared to the control digestor. When sulphate reduction was not limited by the sulphate concentration, the sulphate-adapted digestor had a sulphate reduction rate of 910 mol l^–1 day compared with 17 mol l^–1 day in the control digestor. The results indicate that the potential for sulphate reduction is low in a thermophilic sewage sludge digestor receiving a low sulphate concentration. Counts of sulphate-reducing bacteria and methanogens showed that sulphate-reducing bacteria were found only in significant numbers in the sulphate-adapted digestor and only with H₂/CO₂ as substrate. Only low numbers of acetate-utilizing sulphate-reducing bacteria were found in both digestors. When using radio-labelled acetate, the relative percentage of 2-labelled acetate converted to CO₂ was two to four times higher in the sulphate-adapted digestor compared to the control digestor. These results suggest that oxidation of acetate seems to play a larger role in the sulphate-adapted digestor.Offprint requests to: B. K. Ahring     

Influence of trace elements on biogas production from mango processing waste in 1.5 m3 KVIC digesters

Summary The influence of trace elements (Co²⁺, Ni²⁺ and Fe³⁺) in varying concentrations and combination, was studied in 1.5 m³ Khadi & Village Industries Commission (KVIC) digesters for biogas generation from mangopeel. Addition of these trace metals enhanced the biogas yield and methane content moderately, the maximum being with the iron fed digester. The digesters were always found to be stable without much variation in total volatile fatty acids (VFA), pH, total alkalinity and other parameters. A methane content of 62% and biogas yield of 0.49 m³/kg VS added was obtained with 4000 mg/L FeCl₃ supplemented mangopeel fed digester as compared to control having biogas yield of 0.22 m³/kg VS added with a methane content of about 48–50%.     

A kinetic study of the anaerobic digestion of piggery waste using down-flow stationary fixed film reactors

Summary In this paper the behaviour of the down-flow stationary fixed film digesters is studied at laboratory and bench scale. Several organic loading rates are applied to the reactors in order to examine the support surface behaviour. Specific support surfaces of about 50 m²/m³ void volume seems to be optimal. A set of experiments carried out in a continuous stirred reactor is used to fit the kinetic constants of the Chen and Hashimoto's model. The model is then used to assess its applicability to the DSFF digesters. The results show that its application, is possible as a first approximation.Nomenclature B₀ Ultimate methane yield (m³ CH₄/Kg VS) - B Specific methane production (m³ CH₄/Kg VS) - CSTR Continuous stirred tank reactor - DSFF Down-flow Stationary Fixed Film - HRT Hydraulic retention time (days) - K Kinetic constant of the Chen and Hashimoto model (dimensionless) - S Biodegradable substrate concentration (g/l) - SLR Superficial loading rate (Kg VS/m²·d) - SSS Specific support surface (m² support surface/m³ digester void volume) - S₀ Initial substrate concentration (g/l) - VS Volatile solids (g/l) - VFA Volatile Fatty Acids (mg/dm³) - Microorganisms specific growth (day^-1) - _m Kinetic constant of Chen and Hashimoto's model (day^-1) - Retention time (days) - _m Minimum retention time to avoid microorganisms washout (days)     

Single-Phase and Two-Phase Anaerobic Co-Digestion of Residues from the Treatment Process of Waste Vegetable Oil and Pig Manure

The co-digestion of residues from the pre-treatment process of waste vegetable oil (OW) and pig manure (PM) was performed under different OW/PM feed ratios (1:0, 1:1 and 1:3 v/v) and at organic loading rates ranging from 0.25 to 3.1 kg VS m^?3 day^?1 in lab-scale single-phase (SP) and two-phase (TP) systems. From the experiments, it was observed that digestion of OW alone was inhibitory for the anaerobic degradation. Mixing OW with PM neutralized the negative effects of lipids accumulation and high VS removal efficiencies were realized in both systems (63 and 71 % in SP system and 69 and 72 % in TP system, at 1:1 and 1:3 OW/PM mixtures, respectively). Under the same operational conditions, the methane yield was 0.30 and 0.22 m³ CH₄ kg^?1 VS removed for the SP anaerobic digester and 0.30 and 0.27 m³ CH₄ kg^?1 VS removed for the TP configuration. Additionally, TP digestion presented more stable operation and higher treatment capacity.     

A structured model for the kinetics of fungal amylase production

Summary The cell is considered to be divided into nucleic acids, proteinaceous material and storage compounds. The enzyme is believed to be constitutive but repressed by the rate of catabolism. A structured model is developed to describe the growth, amylase production and dissolved oxygen profile in the batch culture ofAspergillus oryzae.Nomenclature CA Conc. of enzyme SKB units/m³ - C_D,C_E,C_G,C_O,C_S Conc. of D.E.G. mass, oxygen, substrate kg/m³ - C_O* Mean oxygen conc. in gas-liquid interface kg/m³ - C_X Total cell conc. kg/m³ - D D-mass (proteins) kg - E E-mass (storage carbon) kg - G G-mass (necleic acids) kg - K_E Rate of usage of E-mass kg/m³/h - K_EO K_E with oxygen limitation kg/m³/h - Q Active fraction of promotor genes - - R_D,R_E,R_G Rate of production of D,E,G-mass kg/m³/h - R_O,R_S Rate of usage of oxygen, substrate kg/m³/h - R_DO,R_EO,R_GO R_D,R_E and R_G with oxygen limitation kg/m³/h - S Substrate (carbon) concentration kg/m³ - t Time h - t_o Time at which C_S = 0 h - K₁,K₂,K₃,K₂₄,K₂₅ Stoichiometric constants - - K₄,K₅,K₆,K₇ Rate constants h^–1 - K₈-K₁₂,K₁₈,K₂₀-K₂₃ Michaelis-Menten constants kg/m³ - K₂₆,K₂₆ Absorption coefficient, K₂₆ at C_X = 0 h^–1 - K₂₇ Empirical constant kg/m³ - K₁₅ Rate of enzyme formation SKB units/kg - K₁₆,K₁₇ Equilibrium constants m³/kg - K₁₉ Decay constant for mRNA h^–1     

A real-time feed rate control system for pilot plant baker's yeast production

Summary An on-line feed rate control system for baker's yeast production using the molasses uptake rate as a feeding index was developed. The optimal feed rate was obtained by maximising the feeding index. The experiments were performed to test this control system in fermenter of 30 m³ total capacity. In baker's yeast process 2760 kg M₅₀ was consumed and 2852 kg compressed yeast (D_c) was produced. Cell yield, final molasses dilution and final yeast concentration were 1.0 kg D_c/kg M₅₀, 1:6.5 and 52 g D₁₀₀/l, respectively. These results found that the developed feed rate control system is to be successful.     

Productivity and nutrient uptake of water hyacinth,Eichhornia crassipes I. Effect of nitrogen source

Water hyacinth,Eichhornia crassipes, growth and nutrient uptake rates, as influenced by different N sources and N transformations, were measured using microcosm aquaculture systems. Net productivity was highest in the system receiving equal amounts of NH₄ ⁺ and NO₃ ^- (at 10 mg N 1^-1 each) and decreased in the order of NO₃ ^-, NH₄ ⁺, urea (added at 20 mg N 1^-1 each), and methane digestor effluent (at 6 mg N 1^-1). During the first 7-wk study (average ambient air temperature was 26–28°C), biomass yields were in the range of 19–53 g dry wt m^-2 day^-1, while between the 8th and 12th wk (average ambient air temperature was 16–22°C), biomass yields were in the range of 10–33 g dry wt m^-2 day^-1. In the systems with either NH₄ ⁺ or NO₃ ^-, or both added in equal proportions, about 14–20% of the total yield was contributed by roots, whereas in the system with urea and digestor effluent, roots contributed about 23 and 44% of the total yield, respectively. Nitrogen and P uptake per unit area followed trends similar to biomass yields. Nitrogen uptake rates were in the range of 533–2, 161 mg N m^-2 day^-1 for the systems receiving NH₄ ⁺, NO₃ ^-, and urea, while uptake rates were in the range of 124–602 mg N m^-2 day^-1 for the system receiving methane digestor effluent. Phosphorus uptake rates were found to be in the range of 59–542 mg P m^-2 day^-1. Under the most favorable conditions, maximum recorded biomass yield was 53 g dry wt m^-2 day^-1, with N and P removal rate of 2,161 mg N m^-2 day^-1 and 542 mg P m^-2 day^-1, indicating the potential of water hyacinth to produce large amounts of biomass which can be potentially used as a feedstock to produce methane.     

Plant regeneration from hypocotyl-derived protoplasts of Brassica juncea (L.) Czern and Coss

Protoplasts isolated from etiolated hypocotyls of 6-day-old seedlings of Brassica juncea cv RLM 198 were cultured in a modified V₄₇ medium containing 7% mannitol, 2% sucrose, 1.0 mg/l 2,4-D, 0.1 mg/l NAA and 0.4 mg/l BAP, at a density of 5×10⁴ protoplasts per ml of medium. Cultures were incubated in the dark at 25+1°C. After 7 d of culture, cell colonies were diluted with 8_p medium containing 5% mannitol and a similar hormone combination as described earlier. After 14 d, cell colonies were embedded in 8_p medium containing agarose and 3.5% mannitol. Immediately upon gelling, liquid 8_p medium was added to each Petri dish as an overlayer, and cultures were incubated in the light. After a total of 3 to 4 weeks in culture, microcalli were obtained. A modified MS medium with 2% sucrose, 1.0 mg/l 2,4-D and 0.1 mg/l kinetin solidified with 0.5% agarose was used for growing microcalli into callus lines. On MS medium containing 2% sucrose, 0.1 mg/l IAA, 2.0 mg/l zeatin riboside and 2.0 mg/l BAP, solidified with 0.5% agarose, about 35% of the calli regenerated multiple shoots. The time required from culture of protoplasts to multiple shoot regeneration was about 10 weeks. Regenerated shoots were rooted and plants were re-established in a growth chamber at high frequency.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - IAA Indole-3-acetic acid - NAA -naphthaleneacetic acid - BAP 6-benzylaminopurine - IBA Indole-3-butyric acid     

Cotton yarn and fabric finishing wastewater treatment in upflow anaerobic filter

Summary Wastewater from cotton yarn and fabric finishing was successfully treated in an upflow anaerobic filter at 35°C up to a COD loading of 1 Kgr COD/m³ · day; the COD removal varied from 50 to 90% and production of biogas was 0,2–0.4 L/g influent COD, having 70–80% CH₄. At higher COD loading biogas production stopped although COD removal remainedca 50%.     

Metabolic Activity of Fatty Acid-Oxidizing Bacteria and the Contribution of Acetate, Propionate, Butyrate, and CO2 to Methanogenesis in Cattle Waste at 40 and 60°C

The quantitative contribution of fatty acids and CO₂ to methanogenesis was studied by using stirred, 3-liter bench-top digestors fed on a semicontinuous basis with cattle waste. The fermentations were carried out at 40 and 60°C under identical loading conditions (6 g of volatile solids per liter of reactor volume per day, 10-day retention time). In the thermophilic digestor, acetate turnover increased from a prefeeding level of 16 μM/min to a peak (49 μM/min) 1 h after feeding and then gradually decreased. Acetate turnover in the mesophilic digestor increased from 15 to 40 μM/min. Propionate turnover ranged from 2 to 5.2 and 1.5 to 4.5 μM/min in the thermophilic and mesophilic digestors, respectively. Butyrate turnover (0.7 to 1.2 μM/min) was similar in both digestors. The proportion of CH₄ produced via the methyl group of acetate varied with time after feeding and ranged from 72 to 75% in the mesophilic digestor and 75 to 86% in the thermophilic digestor. The contribution from CO₂ reduction was 24 to 29% and 19 to 27%, respectively. Propionate and butyrate turnover accounted for 20% of the total CH₄ produced. Acetate synthesis from CO₂ was greatest shortly after feeding and was higher in the thermophilic digestor (0.5 to 2.4 μM/min) than the mesophilic digestor (0.3 to 0.5 μM/min). Counts of fatty acid-degrading bacteria were related to their turnover activity.     

Filtration of a yeast suspension using an enclosed microporous metal filter

Summary A microporous (3 m) metal filter was very efficient for the recovery of Saccharomyces cerevisiae from a suspension. The filtration could be described by a cake filtration model, the cake resistance being dependent on the pressure drop applied and the concentration of bodyfeed added. The mean filtration capacity was 0.4 m³/m² h.     

Mathematical modeling of production of microbial lipids

In the microbial lipid production system using the yeast Rhodotorula gracilis, CFR-1, kinetics of lipid accumulation and substrate utilisation at initial substrate concentrations in the range of 20–100 kg/m³ were investigated using shake flask experiments. A mathematical representation based on logistic model for biomass and Luedeking-Piret model for lipid accumulation gave reasonably good agreement between the theoretical and experimental values for substrate concentration less than 60 kg/m³. The kinetic expressions and parameters obtained through shake flask studies were directly applied to experiments in the laboratory fermentors also and the models were found to hold good for the prediction of the change of biomass, product as well as substrate with time. The attainment of a saturation in the intracellular lipid accumulation with time, however, was not predicted by the model which was shown to be an inherent feature of the Luedeking-Piret model.List of Symbols S ₀, P ₀ kg/m³ Initial concentrations of sugar and lipid respectively - S, S(t) kg/m³ Concentrations of sugar and lipid respeclively at any timet - p,p(t) L kg/m³ Maximum concentration of lipid produced - E % Maximum sugar utilised - dP/dt kg/(m³ · h) Rate of lipid production - -dS/dt kg/(m³ · h) Rate of sugar utilisation - _max h^–1 Maximum specific growth rate - X _max kg/m³ Maximum biomass reached in a run - P _max kg/m³ Maximum product concentration - m, n Constants used in Luedeking-Piret model in eq. (7) - , Constants used to predict residual sugar - k _e maintainance coefficient - Y _x g/g Biomass yield based on sugar consumed - Y _p g/g Lipid yield based on sugar consumed - (dP/d t)_stat kg/(m³ · h) Rate of lipid production at stationary phase - (dS/dt)_stat kg/(m³ · h) Rate of sugar utilisation at stationary phase     

Ethanol production with Zymomonas mobilis with synthetic medium in batch operation

Ethanol was produced with Zymomonas mobilis Z6 (ATCC 29191), in batch culture with synthetic medium on glucose as substrate and in the presence of aspartate. The concentrations of glucose, phosphate, ammonium, ethanol and dissolved O₂ and CO₂ in the medium and O₂ and CO₂ in the outlet gas as well as the cell mass by culture fluorescence were measured on-line. Cell mass, glucose and aspartate concentrations were measured off-line. In the presence of a sufficient amount of aspartate, the ethanol inhibition effect can be reduced considerably. However, the improvement with yeast extract is more incisive. The relationship between the intensity of culture fluorescence and cell mass concentration is linear, if sufficient aspartate is present.List of Symbols ASP kg/m³ aspartate concentration - CTR kg/(m³ · h) CO₂ transfer rate - N, NH₄ kg/m³ nitrogen concentration from NH ₄ ⁺ - P kg/m³ product (ethanol) concentration - p% product (ethanol) yield - PO₄ kg/m³ phosphate concentration - Q _E kg/(kg · h) specific ethanol production rate - kg/(kg · h) specific nitrogen uptake rate from NH ₄ ⁺ - Q _P kg/(kg · h) specific phosphate uptake rate - Q _s kg/(kg · h) specific substrate (glucose) uptake rate - S kg/m³ glucose concentration - S _O kg/m³ initial glucose concentration - Y _x/s kg/kg yield coefficient - h^–1 specific growth rate     

Mathematical modeling of production of microbial lipids

As a part of the investigations on the microbial lipid production using the yeast Rhodotorula gracilis, CFR-1, kinetics of the biomass synthesis has been studied using shake flask experiments. Using a medium containing a carbon to nitrogen ratio of 701, the rates of biomass production were followed at different initial substrate concentrations in the range of 20–100 kg/m³. A logistic model was found to be reasonably adequate to describe the kinetics of the growth of biomass; the maximum specific growth rate of 0.105 h^–1 was applicable for substrate concentrations less than 60 kg/m³, which gave reasonable agreement between predicted and actual biomass concentration values.List of Symbols S ₀, X ₀ kg/m³ Initial concentrations of sugar, non lipid biomass respectively - X, X(t) kg/m³ Concentrations of non lipid biomass at any time t - dX/dt kg/(m³ · h) Rate of biomass growth - h^–1 Specific growth rate - _max h^–1 Maximum specific growth rate - K _s mol/dm³ Monods constant - X _max kg/m³ Maximum biomass reached in a run     

Production of virus resistant and insect tolerant transgenic tobacco plants

The cucumber mosaic virus coat protein (CMV-CP) gene and a modified Bacillus thuringiensis -endotoxin (Bt toxin) gene were cloned into plant expression vector pE3. Tobacco (Nicotiana tabacum cv. G28) leaf discs were transformed with Agrobacterium tumefaciens A12 carrying recombinant pE14. Transgenic r₀ and R₁ tobacco plants expressing CMV-CP and Bt toxin genes were protected from CMV infection as well as feeding damage of Manduca Sexta (tobacco hornworm) larvae. These results demonstrate that it is feasible to breed new cultivars with multiple resistances via genetic engineering.Abbreviations CMV cucumber mosaic virus - Bt toxin Bacillus thuringiensis -endotoxin - CaMV cauliflower mosaic virus - NOS nopaline synthase - Kan Kanamycin - Spe spectinomycin - Carb Carbenicillin     

Methane from coffee pulp juice: Experiments using polyurethane foam reactors

Summary Methanogenic bioreactors packed with polyurethane foams were used for the conversion of acidified coffee pulp juice into a methane-rich biogas. Gas productivities of the order of 3 v/v day were obtained with loads ranging from 9 to 22 kg VS/m³ day.     

Lactase production in continuous culture by Trichoderma reesei Rut-C30

Summary Trichoderma reesei Rut-C30 was found to produce extracellular lactase when grown on lactose medium. Maximum enzyme levels in continuous culture were observed at dilution rates (D) between 0.02 and 0.027 hr^-1. The enzyme productivity reached 27.3 U/L hr at D = 0.027 hr^-1. Lactase synthesis appears to be inducible and subject to catabolite repression. Optimal growth temperature and pH for enzyme production were 28°C and pH 5. Maximum enzyme activity was observed at 63°C and pH 4.6. The apparent K_m, based on the nitrophenyl-galactopyranoside assay was estimated as 0.4 mM. The enzyme is suitable for lactose hydrolysis in acid whey.     

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.