Tomasz Pokój

Theoretical and observed biogas production from plant biomass of different fibre contents

The methane productivity of silage of four crop species -Zeamays L., Sorghumsaccharatum, Miscanthusxgiganteus and Miscanthussacchariflorus - was investigated. The experiments revealed that at a hydraulic retention time of 60days the volumetric methane yields from the Z.mays L. or S.saccharatum silages were higher than those from the Miscanthusxgiganteus or M.sacchariflorus silages because of the higher crude fibre content in Miscanthus spp. However, at comparable lignin concentrations in the feedstock, methane productivity for M.sacchariflorus (0.19+/-0.08L/g volatile solids) was twice that of Miscanthusxgiganteus (0.10+/-0.03L/g volatile solids). The efficiency of cellulose conversion varied from 83.6% (S. saccharatum) to 52.1% (Miscanthusxgiganteus), and hemicellulose from 88.9% (Z. mays L.) to 59.7% (Miscanthusxgiganteus). Conversion of cellulose and hemicellulose depended on the ratio of these polysaccharides to the lignin concentration of the feedstock.

Molecular detection and diversity of medium-chain-length polyhydroxyalkanoates-producing bacteria enriched from activated sludge

Aims:  Knowledge of the species composition of complex bacterial communities is still very limited. The main objectives of this study were to identify medium‐chain‐length polyhydroxyalkanoates (mcl‐PHAs)‐producing bacteria from activated sludge fed with methanol as well as to characterize their PHA operon.

Semi-continuous anaerobic digestion of different silage crops: VFAs formation, methane yield from fiber and non-fiber components and digestate composition

This study presents the results of long-term semi-continuous experiments on anaerobic digestion at an HRT of 45d with ten silages: 2 annual and 4 perennial crops, and 4 mixtures of annual with perennial crops. The composition of substrates and digestates was determined with Van Soests fractionation method. Removal of non-fiber materials ranged from 49.4% (Miscanthus sacchariflorus) to 89.3% (Zea mays alone and mixed with M. sacchariflorus), that of fiber materials like lignin ranged from 0.005% (Z. mays alone and mixed with grasses at VS ratio of 90:10%) to 46.5% (Sida hermaphrodita). The lowest stability of anaerobic digestion, as confirmed by normalized data concentrations of volatile fatty acids, was reported for both miscanthuses and sugar sorghum. The methane yield coefficients for non-fiber and fiber materials were 0.3666 and 0.2556L/g, respectively. All digestate residues had high fertilizing value, especially those from mixtures of crops.

Molecular insight into activated sludge producing polyhydroxyalkanoates under aerobic–anaerobic conditions

One of the options enabling more economic production of polyhydroxyalkanoates compared to pure cultures is the application of mixed cultures. The use of a microbial community in a sequencing batch reactor has a few advantages: a simple process control, no necessity for sterile processing, and possibilities of using cheap substrates as a source of carbon. Nevertheless, while cultivation methods to achieve high PHAs biomass concentration and high productivity in wild and recombinant strains are defined, knowledge about the cultivation strategy for PHAs production by mixed culture and species composition of bacterial communities is still very limited. The main object of this study was to characterize on the molecular level the composition and activity of PHAs producing microorganism in activated sludge cultivated under oxygen limitation conditions. PHAs producers were detected using a PCR technique and the created PHA synthase gene library was analyzed by DNA sequencing. The obtained results indicate that PHAs-producers belonged to Pseudomonas sp., and possessed genes coding for mcl-PHA synthase. The kinetics of mcl-PHA synthase expression was relatively estimated using real-time PCR technology at several timepoints. Performed quantitative and qualitative analysis of total bacterial activity showed that there were differences in total activity during the process but differential expression of various groups of microorganisms examined by using DGGE was not observed.

ADM1-based modeling of anaerobic codigestion of maize silage and cattle manure – a feedstock characterisation for model implementation (part I) / Modelowanie kofermentacji kiszonki kukurydzy i obornika bydlęcego za pomocą ADM1 – charakterystyka wsadu surowcowego (część I)

Abstract This paper presents the results of fractionation of particulate and soluble organic matter in a mixture of maize silage and cattle manure (49:51% volatile solids) that was used as a feedstock for anaerobic digestion. The extended Weender’s analysis was adapted to measure raw protein, raw lipids, fraction of carbohydrates (including starch, cellulose, hemicelluloses) and lignin. The content of individual fractions in composite, Xc (as kg COD kg-1 COD) was: 0.111 proteins, 0.048 lipids, 0.500 carbohydrates and 0.341 inerts. The biodegradability of Xc was 68%. Based on material balance, the carbon concentration in Xc was 0.0326 kmol C kg-1 COD, whereas nitrogen concentration 0.0018 kmol N kg-1 COD. The estimated pH of the feedstock based on acid-base equilibrium corresponded to the actual value (pH 7.14). Streszczenie W pracy przedstawiono wyniki stężenia substancji chemicznych rozpuszczonych i nierozpuszczonych w mieszaninie kiszonki kukurydzy zwyczajnej i obornika bydlęcego (49:51% suchej masy organicznej), który wykorzystano jako substrat do wytwarzania biogazu. Do frakcjonowania nierozpuszczalnych związków organicznych, stanowiących kompozyt (Xc), wykorzystano metodę Weender’a. Udział poszczególnych frakcji (jako ChZT) w kompozycie wyniósł: białka - 0,111, tłuszcze - 0,048, węglowodany - 0,500 oraz związki inertne - 0,341. Stężenie związków biodegradowalnych w kompozycie wyniosło 68%. Na podstawie bilansu materiałowego węgla i azotu obliczono, że stężenie węgla w kompozycie wynosi 0,0326 kmol C kg-1 ChZT, zaś azotu 0,0018 kmol N kg-1 ChZT. Odczyn (pH) wsadu surowcowego wyznaczony z równowagi kwasowo-zasadowej odpowiadał rzeczywistemu, tj. 7,14.

ADM1-based modeling of anaerobic codigestion of maize silage and cattle manure – calibration of parameters and model verification (part II) / Modelowanie kofermentacji kiszonki kukurydzy i obornika bydlęcego za pomocą ADM1 – kalibracja i weryfikacja modelu (część II)

Abstract The aim of this study was to implement ADM1xp model to simulate behavior of anaerobic co-digestion of maize silage and cattle manure. The accuracy of ADM1xp has been assessed against experimental data of anaerobic digestion, performed at OLR = 2.1 gVS dm-3·d-1 and HRT = 45d. Due to the high number of parameters in ADM1xp, it was necessary to develop a customized procedure limiting the range of parameters to be estimated. The best fitting of experimental to simulated data was obtained after verification of 9 among 105 stoichiometric and kinetic parameters. The values of objective function (Jc) ranged between 0.003 (for valerate) and 211 (for biogas production). Streszczenie Celem pracy było wykorzystanie modelu ADM1xp do symulacji procesu kofermentacji kiszonki kukurydzy i obornika bydlęcego. Przydatność modelu oceniano wykorzystując dane z eksperymentu w skali laboratoryjnej. Badania prowadzono przy obciążeniu komory ładunkiem organicznym OLR = 2,1 gVS dm-3·d-1 oraz hydraulicznym czasie zatrzymania wsadu w fermentorze, HRT = 45d. Z powodu dużej liczby parametrów w modelu ADM1xp, zastosowano procedurę, która umożliwia zmniejszenie liczby weryfi kowanych parametrów podczas kalibracji. Najlepsze dopasowanie danych eksperymentalnych do modelowych uzyskano po weryfikacji 9 spośród 105 stechiometrycznych i kinetycznych parametrów. Wartości współczynnika dopasowania (Jc) zmieniały się w zakresie od 0,003 (kwas walerianowy) do 211 (produkcja biogazu).

Optimization of anaerobic digestion of a mixture of Zea mays and Miscanthus sacchariflorus silages with various pig manure dosages.

Digestion of crop silage (Zea mays L. and Miscanthus sacchariflorus) with 0%, 7.5%, 12.5% and 25% pig manure as co-substrate was performed in continuous stirred-tank reactors, for a constant hydraulic retention time of 45 d and organic load rate of 2.1 g L(-1)d(-1). A matrix of correlations between biogas/methane production and parameters of anaerobic digestion was created in order to estimate process stability. The values of the correlation coefficients indicated that the most stable anaerobic digestion was achieved using 7.5% and 12.5% pig manure. In contrast, the positive correlation between ammonium and volatile fatty acids (r=0.8698, p<0.001) at 25% pig manure showed process instability. Compared to crop silage alone, pig manure favored the production of biogas and methane; the highest production rates were obtained with 12.5% pig manure.

Exploring nutrient limitation for polyhydroxyalkanoates synthesis by newly isolated strains of Aeromonas sp. using biodiesel-derived glycerol as a substrate

Aeromonas spp. strains isolated from activated sludge in a municipal wastewater treatment plant were found to be able to synthesize polyhydroxyalkanoates (PHA) utilizing pure and crude glycerol. The 16S rRNA gene sequence of the isolates exhibited similarity to Aeromonas hydrophila, A. aquatica, and A. salmonicida. Our results confirmed that the adequate supply of nitrogen and phosphorus during culture in 250-ml shake flasks did not stimulate the synthesis of PHAs. The results indicate that the PHA content of cells was higher under a phosphorus-limiting environment compared to nitrogen starvation. In the two-stage cultivation using glucose (in the first step) and crude glycerol from biodiesel industry (in the second step) as a component of the growth medium, the analyzed strains grew to 3.06 g/l of cell dry weight containing up to 22% of PHAs. Furthermore, during the same culture strategy up to 42% of PHAs were extracted, when in the second step of the process, Aeromonas sp. AC_03 was grown on pure glycerol under phosphorus limitation. The purified biopolymer was confirmed to be polyhydroxybutyrate. Aeromonas sp. AC_02 was also capable to accumulate the poly(3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer when pure glycerol was added as a substrate under nitrogen-deficiency one-step bioprocess. Our results confirm that due to the biopolymer productivity, newly isolated strains could be exploited for obtaining valuable biopolymers using wastes generated from biodiesel industry.

Transcriptome remodeling of Pseudomonas putida KT2440 during mcl-PHAs synthesis: effect of different carbon sources and response to nitrogen stress

Bacterial response to environmental stimuli is essential for survival. In response to fluctuating environmental conditions, the physiological status of bacteria can change due to the actions of transcriptional regulatory machinery. The synthesis and accumulation of polyhydroxyalkanoates (PHAs) are one of the survival strategies in harsh environments. In this study, we used transcriptome analysis of Pseudomonas putida KT2440 to gain a genome-wide view of the mechanisms of environmental-friendly biopolymers accumulation under nitrogen-limiting conditions during conversion of metabolically different carbon sources (sodium gluconate and oleic acid). Transcriptomic data revealed that phaG expression is associated with medium-chain-length-PHAs’ synthesis not only on sodium gluconate but also on oleic acid, suggesting that PhaG may play a role in this process, as well. Moreover, genes involved in the β-oxidation pathway were induced in the PHAs production phase when sodium gluconate was supplied as the only carbon and energy source. The transition from exponential growth to stationary phase caused a significant expression of genes involved in nitrogen metabolism, energy supply, and transport system. In this study, several molecular mechanisms, which drive mcl-PHAs synthesis, have been investigated. The identified genes may provide valuable information to improve the efficiency of this bioprocess and make it more economically feasible.

PRODUCTION OF BIOGAS USING MAIZE SILAGE SUPPLEMENTED WITH RESIDUAL GLYCERINE FROM BIODIESEL MANUFACTURING

Abstract The aim of this study was to investigate the influence of residual glycerine (5 and 10% w/w) from the biodiesel industry, used as a co-substrate, on biogas production from maize silage. The experiments were conducted in a laboratory-scale, single-stage anaerobic digester at 39ºC and hydraulic retention time (HRT) of 60 d. Addition of 5% residual glycerine caused organic load rate (OLR) to increase to 1.82 compared with 1.31 g organic dry matter (ODM) L-1d-1 for maize silage alone. The specific biogas production rate and biogas yield were 1.34 L L-1d-1 and 0.71 L g ODM-1 respectively, i.e. 86% and 30% higher than for maize alone. Increasing the residual glycerine content to 10% increased OLR (2.01 g ODM L-1d-1), but clearly decreased the specific biogas production rate and biogas yield to 0.50 L L-1d-1 and 0.13 L g ODM-1 respectively. This suggested that 10% glycerine content inhibited methanogenic bacteria and organics conversion into biogas. As a result, there was accumulation of propionic and valeric acids throughout the experiment.