Identifying targets for increased biogas production through chemical and organic matter characterization of digestate from full-scale biogas plants: what remains and why?

BACKGROUND: This study examines the destiny of macromolecules in different full-scale biogas processes. From previous studies it is clear that the residual organic matter in outgoing digestates can have significant biogas potential, but the factors dictating the size and composition of this residual fraction and how they correlate with the residual methane potential (RMP) are not fully understood. The aim of this study was to generate additional knowledge of the composition of residual digestate fractions and to understand how they correlate with various operational and chemical parameters. The organic composition of both the substrates and digestates from nine biogas plants operating on food waste, sewage sludge, or agricultural waste was characterized and the residual organic fractions were linked to substrate type, trace metal content, ammonia concentration, operational parameters, RMP, and enzyme activity. RESULTS: Carbohydrates represented the largest fraction of the total VS (32-68%) in most substrates. However, in the digestates protein was instead the most abundant residual macromolecule in almost all plants (3-21 g/kg). The degradation efficiency of proteins generally lower (28-79%) compared to carbohydrates (67-94%) and fats (86-91%). High residual protein content was coupled to recalcitrant protein fractions and microbial biomass, either from the substrate or formed in the degradation process. Co-digesting sewage sludge with fat increased the protein degradation efficiency with 18%, possibly through a priming mechanism where addition of easily degradable substrates also triggers the degradation of more complex fractions. In this study, high residual methane production (> 140 L CH4/kg VS) was firstly coupled to operation at unstable process conditions caused mainly by ammonia inhibition (0.74 mg NH3-N/kg) and/or trace element deficiency and, secondly, to short hydraulic retention time (HRT) (55 days) relative to the slow digestion of agricultural waste and manure. CONCLUSIONS: Operation at unstable conditions was one reason for the high residual macromolecule content and high RMP. The outgoing protein content was relatively high in all digesters and improving the degradation of proteins represents one important way to increase the VS reduction and methane production in biogas plants. Post-treatment or post-digestion of digestates, targeting microbial biomass or recalcitrant protein fractions, is a potential way to achieve increased protein degradation.

Microbiological Surveillance of Biogas Plants: Targeting Acetogenic Community.

Acetogens play a very important role in anaerobic digestion and are essential in ensuring process stability. Despite this, targeted studies of the acetogenic community in biogas processes remain limited. Some efforts have been made to identify and understand this community, but the lack of a reliable molecular analysis strategy makes the detection of acetogenic bacteria tedious. Recent studies suggest that screening of bacterial genetic material for formyltetrahydrofolate synthetase (FTHFS), a key marker enzyme in the Wood-Ljungdahl pathway, can give a strong indication of the presence of putative acetogens in biogas environments. In this study, we applied an acetogen-targeted analyses strategy developed previously by our research group for microbiological surveillance of commercial biogas plants. The surveillance comprised high-throughput sequencing of FTHFS gene amplicons and unsupervised data analysis with the AcetoScan pipeline. The results showed differences in the acetogenic community structure related to feed substrate and operating parameters. They also indicated that our surveillance method can be helpful in the detection of community changes before observed changes in physico-chemical profiles, and that frequent high-throughput surveillance can assist in management towards stable process operation, thus improving the economic viability of biogas plants. To our knowledge, this is the first study to apply a high-throughput microbiological surveillance approach to visualise the potential acetogenic population in commercial biogas digesters.

Inoculum Source Determines Acetate and Lactate Production during Anaerobic Digestion of Sewage Sludge and Food Waste.

Acetate production from food waste or sewage sludge was evaluated in four semi-continuous anaerobic digestion processes. To examine the importance of inoculum and substrate for acid production, two different inoculum sources (a wastewater treatment plant (WWTP) and a co-digestion plant treating food and industry waste) and two common substrates (sewage sludge and food waste) were used in process operations. The processes were evaluated with regard to the efficiency of hydrolysis, acidogenesis, acetogenesis, and methanogenesis and the microbial community structure was determined. Feeding sewage sludge led to mixed acid fermentation and low total acid yield, whereas feeding food waste resulted in the production of high acetate and lactate yields. Inoculum from WWTP with sewage sludge substrate resulted in maintained methane production, despite a low hydraulic retention time. For food waste, the process using inoculum from WWTP produced high levels of lactate (30 g/L) and acetate (10 g/L), while the process initiated with inoculum from the co-digestion plant had higher acetate (25 g/L) and lower lactate (15 g/L) levels. The microbial communities developed during acid production consisted of the major genera Lactobacillus (92-100%) with food waste substrate, and Roseburia (44-45%) and Fastidiosipila (16-36%) with sewage sludge substrate. Use of the outgoing material (hydrolysates) in a biogas production system resulted in a non-significant increase in bio-methane production (+5-20%) compared with direct biogas production from food waste and sewage sludge.

Ammonia threshold for inhibition of anaerobic digestion of thin stillage and the importance of organic loading rate.

Biogas production from nitrogen-rich feedstock results in release of ammonia (NH3), causing inhibition of the microbial process. The reported threshold ammonia value for stable biogas production varies greatly between studies, probably because of differences in operating conditions. Moreover, it is often difficult to separate the effect of ammonia inhibition from that of organic loading rate (OLR), as these two factors are often interrelated. This study attempted to distinguish the effects of ammonia and OLR by analysis of two laboratory-scale biogas reactors operating with thin stillage and subjected to an increase in free ammonia (from 0.30 to 1.1 g L(-1)) either by addition of an external nitrogen source (urea) or by increasing the OLR (3.2-6.0 g volatile solids L(-1) d(-1)). The results showed that ammonia concentration was detrimental for process performance, with the threshold for stability in both processes identified as being about 1 g NH3-N L(-1), irrespective of OLR. Analysis of the methanogenic community showed limited differences between the two reactors on order level and a clear increase in the abundance of Methanomicrobiales, particularly Methanoculleus sp., in response to increasing ammonia concentration. Further comprehensive molecular analysis revealed that diverse Methanoculleus species dominated in the reactors at a given ammonia level at different OLR. The acetogenic community was clearly affected by both ammonia concentration and OLR, suggesting that the volatile fatty acid load in relation to the higher OLR was important for the dynamics of this community.

Co-digestion of manure and industrial waste--The effects of trace element addition.

Manure is one of the most common substrates for biogas production. Manure from dairy- and swine animals are often considered to stabilize the biogas process by contributing nutrients and trace elements needed for the biogas process. In this study two lab-scale reactors were used to evaluate the effects of trace element addition during co-digestion of manure from swine- and dairy animals with industrial waste. The substrate used contained high background concentrations of both cobalt and nickel, which are considered to be the most important trace elements. In the reactor receiving additional trace elements, the volatile fatty acids (VFA) concentration was 89% lower than in the control reactor. The lower VFA concentration contributed to a more digested digestate, and thus lower methane emissions in the subsequent storage. Also, the biogas production rate increased with 24% and the biogas production yield with 10%, both as a result of the additional trace elements at high organic loading rates. All in all, even though 50% of the feedstock consisted of manure, trace element addition resulted in multiple positive effects and a more reliable process with stable and high yield.

Comparison of operating strategies for increased biogas production from thin stillage.

The effect of increasing organic loading rate (OLR) and simultaneously decreasing hydraulic retention time (HRT) during anaerobic digestion of sulphur- and nitrogen-rich thin stillage was investigated during operation of continuously stirred tank laboratory reactors at two different temperatures. The operating strategies and substrate were set in order to mimic an existing full-scale commercial biogas plant in Sweden. The reactors were operated for 554-570 days with a substrate mixture of thin stillage and milled grain, resulting in high ammonium concentrations (>4.5gL(-1)). Initially, one reactor was operated at 38°C, as in the full-scale plant, while in the experimental reactor the temperature was raised to 44°C. Both reactors were then subjected to increasing OLR (from 3.2 to 6.0gVSL(-1)d(-1)) and simultaneously decreasing HRT (from 45 to 24 days) to evaluate the effects of these operational strategies on process stability, hydrogen sulphide levels and microbial composition. The results showed that operation at 44°C was the most successful strategy, resulting in up to 22% higher methane yield compared with the mesophilic reactor, despite higher free ammonia concentration. Furthermore, kinetic studies revealed higher biogas production rate at 44°C compared with 38°C, while the level of hydrogen sulphide was not affected. Quantitative PCR analysis of the microbiological population showed that methanogenic archaea and syntrophic acetate-oxidising bacteria had responded to the new process temperature while sulphate-reducing bacteria were only marginally affected by the temperature-change.

The effect of substrate and operational parameters on the abundance of sulphate-reducing bacteria in industrial anaerobic biogas digesters.

This study evaluated the effects of operational parameters and type of substrate on the abundance of sulphate-reducing bacteria in 25 industrial biogas digesters using qPCR targeting the functional dissimilatory sulphite reductase gene. The aim was to find clues for operational strategies minimizing the production of H2S. The results showed that the operation, considering strategies evaluated, only had scarce effect on the abundance, varying between 10(5) and 10(7) gene copies per ml. However, high ammonia levels and increasing concentration of sulphate resulted in significantly lower and higher levels of sulphate-reducing bacteria, respectively.