Laboratory research

The aim of this study was to determine elementary processes related to the organic and nitrogen compounds removal in hybrid reactors with intermittent aeration, to assess removal efficiency under various organic and hydraulic loadings and organic and nitrogen compounds’ utilization rates. The utilitarian aim was to determine technological conditions which could make the process shorter and more economically efficient. The attempts to modeling using various technical parameters (together or separately) were conducted.

3.1. Laboratory model and methods

Carriers used in the research were corrugated cylindrical rings made of PP diameter and length of 13 mm, 0.98 g/m3 density and 0.86 porosity (figure 5).

The research studies were conducted in four stages of 10 months duration. Each stage was consisted of three or four series. In each stage three reactors worked simultaneously as continuous flow system in stage I and III and as a sequencing batch reactor in stage II. The volume of reactors was equal to 75 dm3 and volume of settler for the continuous flow was equal to 20 dm3.

The studies were focused on an intermittent aeration. The most attention was put on the last stage — with increased wastewater pH value using lime (Ca(OH)2). The aim of higher pH maintaining was to inhibit the second phase of nitrification by ammonia. The wastewater originated from one family household. The retention time before the sewage discharging into the reactors was relatively high — about 6 days (septic tank and retention tank). The activated sludge originated from Poznan Central WWTP and was inoculated to each reactor at the same amount in each stage beginning. Mixing and aeration of the reactors was made using large-bubble diffusers. The air was supplied by compressor of 0.1-2.0 m3/h capacity. The sludge recirculation was made using an air-lift cooperating with a membrane pump. Characteristic research parameters are shown in table 2.


Figure 5. Scheme of laboratory model

Continuous flow reactor (CFR)

Batch reactor (SBR)

Continuous flow reactor (CFR) with increased pH

Sewage volume per day, dm3/d

70 — 290

45 — 270


Number of series




Variable factor for series

time of aeration

length of reactor cycle

number of carriers

Variable factor for reactors

hydraulic load pollution load

active volume of reactor

pH value


1.31 + 0.09

1.67 + 0.10

1.39 + 0.07

Total solids, g/m3

34.95 + 6.65

62.30 + 3.30

37.00 + 5.20

Organic compounds as COD, g O2/m3

188.25 + 1.62

172.80 + 3.7

184.00 + 8.00

Nitrogen compounds as Ntot, g N/m3

46.56 + 1.35

41.65 + 1.09

52.92 + 2.43

Table 2. Technological characteristics of model investigation and average concentrations of pollutants in sewage

The characteristic feature of the used sewage was a low C/N ratio caused by pretreatment in a septic tank. The pollutants in sewage and suspended biomass concentrations were measured according to the standard methods. The attached biomass concentration was identified via the Kjeldahl nitrogen measurement: 1 g Ntkn corresponds to 0.11 gdm [50]; pH and oxygen were measured using calibrated electrodes.

The detailed description of research results of all experimental stages is included in the Makowska’s monography [11]. In this chapter only the most important processes and parameters related to the carbon and nitrogen removal efficiency are presented. Results related to the parameters like: biomass loading, pollutants’ removal efficiency and substrates utilization rates were analyzed statistically.

The "natural probiotic" effect of biofloc

Biofloc can be a novel strategy for disease management in contrast to conventional approaches such as antibiotic, antifungal, probiotic and prebiotic application. The "natural probiotic" effect in BFT could act internally and/or externally against, i. e., to Vibrio sp. and ectoparasites, respectively. This effect is promoted by large groups of microorganisms, but mainly bacteria that is considered the first trophic level in the system.

Internally, bacteria and its synthesized compounds could act similar to organic acids and might be effective bio-control agents, also given beneficial host’s microbial balance in the gut [68]. The regular addition of carbon in the water is known to select for polyhydroxyalkanoates (PHA) accumulating bacteria and other groups of bacteria that synthesize PHA granules. The microbial storage product poly-fi-hydroxybutyrate (PHB), a biodegradable polymer belonging to the polyesters class, is only one compound of a whole family of polyhydroxyalkanoates. PHB is produced by a widely variety of microorganisms such as Bacillus sp., Alcaligenes sp., Pseudomonas sp. from soluble organic carbon and is also involved in bacterial carbon metabolism and energy storage [68]. This polymer could comprise ~80% of the bacteria’s cell dry matter and up to 16% on biofloc dry weight [69]. Different carbon sources or structures of carbon substrate will result in varying types of PHA [69].

Such granules are synthesized under conditions of physiological and nutrient stress, i. e., when an essential nutrient like nitrogen is limited in the presence of an excess carbon source [68]. When these polymers are degraded in the gut, they could have antibacterial activity similar to short chain fatty acids (SCFAs) or organic acids. The breakdown of PHA inside the gastrointestinal tract can be carried out via chemical and enzymatic hydrolysis [70].

Chemical hydrolysis can be carried out by treating the polymers with, i. e., NaOH, in which could significantly accelerate its digestibility [70]. On the other hand, enzyme hydrolysis is generally carried out by extracellular depolymerases activities which are widely distributed among bacteria and fungi, acting as a preventive or curative protector against Vibrio sp. infections and stimulate growth and survival of shrimp and fish larvae [69].

The working mechanism of PHAs with respect to their antibacterial activity is not well understood [68]. As they could act similarly to SCFA, some studies speculated the working mechanism by (i) reduction of pH, in which antibacterial activity increases with decreasing pH value [71]; (ii) inhibiting the growth of pathogenic bacteria by interference on cell membrane structure and membrane permeability, as well as instability of internal protons balance, lowering ATP and depletion of cellular energy [72]; and (iii) down-regulate virulence factor expression and positively influence the gut health of animals [73]. Further research is need to maximizing PHA content in bioflocs applied, i. e., for fish/shrimp feed, characterizing and analyzing their bio-control efficacy in different host-microbe systems [68].

Externally, the working mechanism of biofloc microorganisms against pathogens seems to be by competition of space, substrate and nutrients. Some essentials nutrients such as nitrogen are required by both groups (i. e. heterotrophic bacteria vs Vibrio sp.) limiting their growth. Inhibiting compounds excreted by BFT microorganisms, light intensity and type of carbon source also could reduce pathogens growth. Unfortunately, limited information is available on this field. In a study with fish fingerlings [74] was reported that tilapia (initial weight 0.98 ± 0.1g) reared under BFT limited water-exchange condition (FLOC) presented less ectoparasites in gills and ectoderm’s mucous as compared to conventional water — exchange system (CW) after 60 days (Fig 5).


Figure 5. Number of total ectoparasites in gills and ectoderm’s mucous of fry tilapia reared under BFT limited water-exchange condition (FLOC) and conventional water-exchange system (CW) after 60 days (more details in [74])

Multienzyme complex cellulosome

The enzyme systems for the lignocellulose degradation by microorganisms can be generally regarded as non-complexed or complexed enzymes (Lynd et al., 2002). In the case of aerobic fungi and bacteria, the cellulase enzymes are free and mostly secreted. In such organisms, by the very nature of the growth of the organisms, they are able to reach and penetrate the cellulosic substrate and, hence, the secreted cellulases are capable of hydrolyzing the substrate. The enzymes in these cases are not organized into high molecular weight complexes and are called non-complexed (Fig. 3A). The polysaccharide hydrolases of the aerobic fungi are largely described based on the examples from Trichoderma, Penicillum, Fusarium, Humicola, Phanerochaete, etc., where a large number of the cellulases are encountered (Dashtban et al., 2009; Sanchez, 2009). In contrast, various cellulases and hemicellulases from several anaerobic cellulolytic microorganisms, are tightly bound to a scaffolding protein, as core protein and organized to form structures on the cell surfaces; these systems are called complexed enzymes or cellulosomes (Fig. 3B). The cellulosome is thought to allow concerted enzyme activities in close proximity to the bacterial cell, enabling optimum synergism between the enzymes presented on the cellulosome. Concomitantly, the cellulosome also minimizes the distance over which hydrolysis products must diffuse, allowing efficient uptake of these oligosaccharides by the host cells (Bayer et al., 1994; Schwarz, 2001; Lynd et al., 2002).

Biotechnological applications in terms of hydrolysis efficiency for complexed enzyme systems might have an advantage over non-complexed enzyme systems. The high efficiency of the cellulosome has been attributed to (i) the correct ratio between catalytic domains that optimize synergism between them, (ii) appropriate spacing between the individual


components to further favor synergism, (iii) the presence of different enzymatic activities (cellulolytic or hemicellulolytic enzymes) in the cellulosome that can remove "physical hindrances" of other polysaccharides in heterogeneous plant cell materials (Lynd et al., 2002), and (iv) the presence of carbohydrate-binding modules (CBMs) that can increase the rate of hydrolysis by bringing the cellulosome into intimate and prolonged association with its recalcitrant substrate (Shoseyov et al., 2006). Thus, the complexed enzyme system, cellulosome, may provide great potential for the degradation of plant biomass.

Figure 3. Simplified schematic of the hydrolysis of amorphous and microcrystalline celluloses by non — complexed (A) and complexed (B) cellulase systems. (This figure is adapted from Lynd et al., 2002).

The cellulosome was first identified in 1983 from the anaerobic, thermophilic, spore-forming Clostridium thermocellum (Lamed et al., 1983). The cellulosome of C. thermocellum is commonly studied along with cellulosomes from the anaerobic mesophiles, C. cellulovorans (Doi et al., 2003), C. josui (Kakiuchi et al., 1998) and C. cellulolyticum (Gal et al., 1997). All cellulosomes share similar characteristics, they all contain a large distinct protein, referred to as the scaffoldin which allows binding of the whole complex to microcrystalline cellulose via CBM. Also, the cellulosome scaffoldin expresses type I cohesins which allow binding of a wide variety of cellulolytic and hemicellulolytic enzymes within the complex via the expression of complementary type I dockerins on enzymes. Similarly, at the C-terminal the scaffoldin expresses type II cohesins, which allow the binding of the cellulosome to the cell through type II dockerins on surface layer homology proteins (SLH) (Fig. 4).

Cellulosomes are produced mainly by anaerobic bacteria, mostly from the class clostridia, and some anaerobic fungi such as genus Neocallimastix (Dalrymple et al., 1997), Piromyces (Teunissen et al., 1991) and Orpinomyces (Li et al., 1997). However, evidence suggests the presence of cellulosomes or cellulosome-like multienzyme complexes in a few aerobic microorganisms (Table 1). It is speculated that several other cellulolytic bacteria may also produce cellulosomes not yet described.



DockerinI U Dockerinll Cohesin I u Cohesin II

Catalytic domain


SLH domain




Figure 4. Simplified schematic of general cellulosome components and connection with cell surface based on knowledge of Clostridium cellulosome. (This figure is adapted from Bayer et al., 1994).















Ding et al., 1999

Bacillus circulans F-2

Potato starch granules

Kim and Kim, 1993






van Dyk et


lort et al., 2012

licheniformis SVD1

al., 2009




Ding et al., 2000

Paenibacillus curdlanolyticus B-6



Pason et al., 2006b

Bacteroides sp.


Ponpium et al.,



Hou, et al.,

strain P-1








Berger et al., 1990



Sabathe et






Lamed et al.,





Pages et al.,



Jiang et al.,



olivaceoviridis E-86





Sleat et al., 1984

Clostridium josui


Kakiuchi et al., 1998




Paper mill

Pohlschroder et al., 1994

Chaetomium sp. Nov. MS-017

Rotted wood

Ohtsuki et al., 2005



Lamed et al.,







Blair and Anderson, 1999b

Ruminococcus albus


Ohara et al., 2000




Ding et al., 2001



Phitsuwan et al.,

xylanilyticum BT14









Microorganism Source






Chimtong et al., 2011





Dalrymple et al., 1997

Orpinomyces joyonii


Qiu et al., 2000

Orpinomyces PC-2


Borneman et al., 1989

Piromyces equi


Teunissen et al., 1991

Piromyces E2


Teunissen et al., 1991

Table 1. Cellulosome and cellulosome-like multienzyme complexes from anaerobic and aerobic microorganisms. (This table is adapted from Doi & Kosugi, 2004).

Generation and Utilization of Microbial Biomass Hydrolysates in Recovery and Production of Poly(3-hydroxybutyrate)

Jian Yu, Michael Porter and Matt Jaremko

Additional information is available at the end of the chapter http://dx. doi. org/10.5772/52940

1. Introduction

In moving towards sustainable manufacturing with reduced carbon footprint, bio-based fuels, chemicals and materials produced from renewable resources have attracted great interest. Microbial cells, in working with other chemical and enzymatic catalysts, are often used in the conversion of feedstocks to desired products, involving different species of bacteria, yeast, filamentous fungi, and microalgae. A substantial amount of microbial biomass is generated in the industrial fermentations and often discarded as a waste. Because of a high cost associated with growth and disposal of the cell mass, reusing the microbial biomass may be an attractive alternative to waste disposal. In contrast to the biomass as energy storage (e. g. starch and oil) or plant structure (e. g. cellulose and hemi-cellulose), microbial biomass is biologically active, consisting primarily of proteins (10-60 wt%), nucleic acid (1-30 wt%), and lipids (1-15%) [1]. Few cases of reusing microbial biomass exist in industrial processes.

Poly(3-hydroxybutyrate) (PHB) is a representative polyhydroxyalkanoate (PHA) that is formed by many bacterial species as carbon and energy reserve [2,3]. Although the biopolyesters made from renewable feedstocks have the potential of replacing petroleum — based thermoplastics in many environmentally friendly applications, they are not widely accepted in the markets because of the high production cost [4]. Extensive research has been conducted to use cheap feedstocks [5,6], develop high cell density fermentation technology for high PHA productivity [7,8], and improve microbial strains that exhibit good performance under high osmotic pressure and environmental stress [9-11]. One major cost factor of PHA production is the recovery and purification of biopolyester for desired purity and material properties [4, 12]. Depending on strains and culture conditions, biopolyester may account for 50-80 wt% of cell mass [13]. They are stored in microbial cells as tiny

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amorphous granules [0.2-0.5 pm in diameter] and need a sophisticated treatment to separate them from the residual cell mass [14-16]. Two technologies, based-on solvent extraction or biomass dissolution, are usually adopted in PHA recovery. With solvent extraction, the PHA granules are dissolved in appropriate organic solvents, leaving cells or residual biomass intact [17,18]. With cell mass dissolution, the PHA granules are left intact while the non-PHA cell mass is decomposed and dissolved in aqueous solutions with help of biological and/or chemical agents [19,20]. Following either treatment, PHA and non-PHA cell mass can be separated with conventional solid/liquid separations. Separating biopolyester from the cells would generate a substantial amount of residual microbial biomass, 0.25 to 1 kg dry mass per kg of PHA resin, depending on the initial PHA content. As a mixture of proteins, nucleic acids, lipids, and wall fragments, the residual biomass has no market value and is discarded at an extra disposal cost.

According to the microbial structure and cellular composition [1], the residual microbial biomass is actually consisting of true biological compounds formed during cell growth while PHA is just a carbon storage material. In a conventional PHA fermentation, sufficient substrates and nutrients (C, N, P, minerals and some organic growth factors) are supplied to grow enough cell mass that in turn or simultaneously synthesize biopolyester from carbon substrates. A large portion of organic carbons and nutrients are therefore consumed to generate new cell mass that is going to be discarded as a solid waste after polymer recovery. Ideally, the residual biomass should be reused by microbial cells to generate new cell mass and/or PHA polymers. This would not only reduce the cost of waste treatment and disposal, but also save the cost of nutrients in PHA fermentation. The bacterial cells, however, cannot assimilate their cell mass because they lack appropriate enzymes to break down various biological macromolecules and their complex structure such as cellular walls and membranes [19]. If the cells or mutants from genetic engineering could easily assimilate their structural components, they might not be suitable to industrial PHA production because the cells would undergo autolysis under high environmental stress. It is highly possible, however, to make the residual biomass reusable during PHA recovery in which the non-PHA cell mass is decomposed and hydrolyzed in aqueous solutions [20]. Integration of PHA recovery with reusing of residual biomass in microbial PHA fermentation is a novel and challenging technology. This work shows the generation of biomass hydrolysates in a downstream PHB recovery and the beneficial utilization of the hydrolysates in cell growth and PHB formation.

Water management and Water Use Efficiency (WUE)

In NGP, soil water deficit occurs very frequently during crop growing season because of the highly variable and uneven distribution of seasonal precipitation. In general, biomass yield of switchgrass increased as the amount of seasonal precipitation increased. However, at a given seasonal precipitation level (e. g., 500-600 mm), switchgrass yield ranged from 2 to 25 Mg/ha (Figure 2) [71], indicating the importance of crop WUE and precipitation use efficiency. Ideally, the figure 2 should be converted to the biomass yield as a function of seasonal evapotranspiration (ET) or transpiration (T), not precipitation because crop yield is more closely related to ET or T. Although most field studies have included precipitation information in NGP, there is no detailed information of crop ET, transpiration and water — use efficiency (WUE). The quantification of ET and WUE in biofuel crops under various environmental conditions and management practices will lead to identify the best management strategies. Because both water and N are critical for crop growth, the interaction of water and N becomes important, particularly under dryland conditions. However, there are very few studies on the interactive effects of N and soil moisture on biomass yield and quality in biofuel crops.

Development of concept for multi-bale handling unit

3.4. Modulization of bales

Individual handling of bales (round or square) is not cost effective because of long loading and unloading times. Several concepts for a multi-bale handling unit are being developed. A verified multi-bale system is not available at this time to use as an example.

Permission was received [67] to present a concept that is far enough along in development that some field tests have been done. The concept was developed by a consortium led by FDC Enterprises and is shown in Figure 8. The self-loading trailer loads six stacks of six large rectangular bales, referred to "6-packs," for a total load of 36 bales. The bale size is 0.91 m x 1.22 m x 2.44 m. The length of the load is 6 x 2.44 = 14.6 m (48 ft), the height is 3 x 0.91 = 2.74 m (9 ft), and the width is 2 x 1.22 m = 2.44 m (8 ft).


Figure 8. Multi-bale handling unit concept developed for 3x4x8 large rectangular bales by FDC Enterprises-led Consortium (Reprinted with permission [67]).

The trailer built to implement the concept [68] is shown in Figure 9. Estimated load time is 5

min, which is about the same load time for the cotton module (Figure 7). The 36-bale unit

can be off-loaded by the truck directly onto the conveyor into a bioenergy plant, or it can be off-loaded into at-plant storage to be used later, just as is done with cotton modules at a cotton gin.

A similar concept known as the "Rack System" envisions that round bales will be loaded into a rack in-field or at an SSL. This rack is lifted off a trailer at the plant, emptied as the bales are needed, and then returned to be refilled. The racks are cycled multiple times each week within the closed logistics system. Cundiff et al. [59] developed the rack system concept, and it will now be used as an example to illustrate how the principles required for a logistics system are implemented.

All the various concepts and options cannot be discussed. However, we will apply a specific example to help the reader think through a process of developing a logistics system. The selection of the "Rack System" for this example implies no criticism of other ways of implementing the multi-bale handling unit concept currently being developed.


Figure 9. Self-loading trailer built by Kelderman Mfg. to implement multi-bale handling unit concept for 36-bale stack of large rectangular bales.

Quantitative calculation method of relationship between adsorption and biodegradation in BAC filter bed

3.3.2.I. Quantitative calculation method

This part compares O3-GAC with O3-BCF in which the biodegradation is dominant. By determining the iodide adsorption by BCF, only to find out the iodide adsorption is nearly zero, which may conclude that for BCF filter bed the removal of organics in water is mainly realized by biodegradation, and the BCF adsorption is extremely faint. Assuming the quantity of biodegradation is in proportion to biological activity, the more active the microbe is, the more organics degraded will be. Therefore, determining the quantity of organics being degraded by BAC can be conducted by calculating the ratio of BAC biological activity with BCF biological activity, which can also determine the roles played by BAC filter bed adsorption and biodegradation in the process of organic removal. The calculation in details is shown in Fig. 24. Shown from the simplified calculation diagram in Fig. 24, two kinds of indexes need to be assessed to quantitatively determine the relationship between BAC adsorption and biodegradation within the process of organics removal:

1. to evaluate the comprehensive effect of BAC and BCF on organics removal, the system is characterized by DOC and BDOC;

2. to compare and analyze the biological activity of carriers from each filter bed.


Figure 24. Illustration of quantification of the removal of organic matters by adsorption and biodegradation

Effects of earthworms on microbial communities during vermicomposting: a case study

Animal manures are microbe-rich environments in which bacteria constitute the largest fraction (around 70% of the total microbial biomass as assessed by PLFA analysis), with fungi mainly present as spores [24]. Thus, earthworm activity is expected to have a greater effect on bacteria than on fungi in these organic substrates in the short-term [79]. In line with this, a significant increase in the fungal biomass of pig manure, measured as ergosterol content, was detected in a short-term experiment (72 h) with the earthworm species E. fetida, and the effect depended on the density of earthworms [82]. A higher fungal biomass was found at intermediate and high densities of earthworms (50 and 100 earthworms per mesocosm, respectively), which suggests that there may be a threshold density of earthworms at which fungal growth is triggered. This priming effect on fungal populations was also observed in previous short-term experiments in the presence of the epigeic earthworms Eudrilus eugeniae and Lumbricus rubellus fed with pig and horse manure, respectively [16,86]. These contrasting short-term effects on bacterial and fungal populations are thus expected to have important implications on decomposition pathways during vermicomposting because important differences exist between both microbial decomposers related to resources requirements and exploitation [92]. This is based on the fact that fungi can immobilise great quantities of nutrients in their hyphal networks, whereas bacteria are more competitive in the use of readily decomposable compounds and have a more exploitative nutrient use strategy by rapidly using newly produced labile substrates [92].

The above-mentioned studies dealing with the effects of epigeic earthworms on microorganisms have focused on the changes before and after the active phase rather than those that occur throughout the whole vermicomposting process. Hence, in a current research study, and using a continuous-feeding vermicomposting system, we evaluated the different phases of interaction between earthworms and microorganisms and additionally, we monitored the stabilisation of the fresh manure during a period of 250 days. At the end of the experiment we obtained a profile of layers of increasing age, resembling a time profile, with a gradient of fresh-to-processed manure from the top to the bottom. This type of system allowed us to evaluate whether and when the samples reached an optimum value to be classified as vermicompost, as regards to the stabilisation of organic matter and the levels of microbial biomass and activity. Briefly, we used polyethylene reactors (n=5) with a volume of 1 m3, which were initially comprised of a 10 cm layer of mature vermicompost (a stabilised non- toxic substrate that serves as a bed for earthworms), on which earthworms (Eisenia fetida) were placed and a layer containing 5 kg of fresh rabbit manure, which was placed over a plastic mesh (5 mm pore size) to avoid sampling the earthworm bedding. New layers with the same amount of fresh manure were added to the vermireactor every fifty days according to the feeding activity of the earthworm population. This procedure allowed the addition of each layer to be dated within the reactors. The reactors were divided into 4 quadrants and two samples were taken at random from each quadrant with a cylindrical corer (8 cm diameter). Each corer sample was divided into five layers of increasing age and the samples from the same layer and each reactor were gently mixed to analyse the changes in microbial communities. The structure of the microbial communities was assessed by PLFA analysis; some specific PLFAs were used as biomarkers to determine the presence and abundance of specific microbial groups [93]. The sum of PLFAs characteristic of Gram-positive (iso/anteiso branched-chain PLFAs), and Gram-negative bacteria (monounsaturated and cyclopropyl PLFAs) were chosen to represent bacterial PLFAs, and the PLFA 18:2w6c was used as a fungal biomarker. Total microbial activity was also assessed by measuring the rate of evolution of CO2, as modified for [17] for samples with a high organic matter content. Dissolved organic carbon was determined colorimetrically in microplates after moist digestion (K2&2O7 and H2SO4) of aliquots of 0.5M K2SO4 extracts.

The earthworm species E. fetida had a strong effect in the decomposition of organic matter during vermicomposting, greatly modifying the structure of the microbial decomposer communities, as revealed by the phospholipid fatty acid analysis. The principal component analysis of the 27 identified PLFAs (10:0, 12:0, 13:0, 14:0, i14:0, 15:0, i15:0, a15:0, 16:0, i16:0, 17:0, a17:0, 18:0, 14:1w5c, 15:1w5c, 16:1w7c, 17:1w7c, 18:1w7c, 18:1w9c, 18:1w9t, 18:2w6c, 18:2w6t, 18:3w6c, 18:3w3c, cy17:0, cy19:0, 20:0) clearly differentiated between the samples in function of the age of layers, explaining 51% of the variance in the data (Figure 2). Thus, the upper layers (50 and 100 days old) along with the fresh manure were clearly distinguished from the intermediate (150 days old) and lower layers (200 and 250 days old) (Figure 2).


Figure 2. Changes in the microbial community structure throughout the process of vermicomposting assessed by the principal component analysis of the twenty-seven PLFAs identified in the layers of reactors fed with rabbit manure. The different layers represent different stages of the process. Values are means ± SE.

Such changes in the structure of microbial communities were accompanied by a decrease in the abundance of both Gram-positive and — negative bacterial populations with the depth of layers (Figure 3A, B), i. e. from upper to medium and lower layers; and the abundance of these groups were in the fresh rabbit manure 346 ± 49.0 and 336 ± 63 pg g-1 dw for Gram­positive and Gram-negative bacteria, respectively (Figure 3A, B). A similar trend was observed for fungal populations (Figure 3C), reaching an average value of 1.3 ± 0.1 at the end of the process (Figure 3C). These results are in accordance with previous studies based on PLFA profiles, with marked changes in the structure of microbial communities due to decreases in both bacterial and fungal populations throughout the process of vermicomposting [18, 89]. Recently, Fernandez-Gomez et al. [94] observed that the structure of fungal communities, assessed by DGGE profiles differed at the stage of maximum earthworm biomass the most, suggesting the existence of a strong gut passage effect on the microbial communities through a continuous-feeding vermicomposting system in the presence of E. fetida.

Decreases in microbial activity were also detected with depth of layer (Figure 4A) and, after a maturation period for 250 d, basal respiration values dropped below 100 mg CO2 kg-1 OM h-1 (Figure 4A), as previously shown by [18]. Accordingly, a reduction in the dissolved organic carbon content was detected from upper to lower layers (Figure 4B), reaching a value close to 7000 pg g-1 dw after 250 d of vermicomposting. In contrast, other authors [17] reported levels of DOC much more lower in a long-term experiment (252 days) with the epigeic earthworm E. fetida, with values below 1500 pg g-1 dw in the presence of earthworms. Such differences could be due to the composition of the parent material (pig slurry versus rabbit manure) and/or to the experimental setup conditions. Unlike compost — a limit value of 4000 mg kg-1 is suggested for a stable compost according to [34]- there is still no threshold level of DOC for which vermicompost is to be considered stable.


Figure 3. Changes in (a) Gram-positive bacterial, (b) Gram-negative bacterial and (c) fungal PLFAs in the layers of reactors fed with rabbit manure throughout the process of vermicomposting. The different layers represent different stages of the process. Different letters indicate significant differences between the layers based on post hoc test (Tukey HSD). Values are means ± SE.

Overall, in the present study a higher degree of stabilisation was reached in the rabbit manure after a period of between 200 and 250 days, as indicated by the lower values of microbial biomass and activity that are indicative of stabilized materials. These results underscore the potential of epigeic earthworms in the stabilisation of this type of organic substrates, which is of great importance for the application of animal manures as organic amendments into agricultural soils because, as already mentioned, it is widely recognised that the overproduction of this type of substrate has led to inappropriate disposal practices, which may result in severe risks to the environment [6]. Furthermore, these findings constitute a powerful tool for the development of strategies leading to a more efficient process for the disposal and/or management of animal manures, thereby highlighting the continuous-feeding vermicomposting system as an environmentally sound management option for recycling such organic wastes, as previously reported by [94] for treating tomato — fruit waste from greenhouses. Ultimately, it should be borne in mind that the functioning of this type of reactors can lead to the gradual accumulation of layers and compaction of the substrate, thus minimizing earthworm — induced aeration, which can promote pathogen survival [89].


Figure 4. Changes in (a) microbial activity assessed by basal respiration, and (b) dissolved organic carbon content in the layers of reactors fed with rabbit manure throughout the process of vermicomposting. The different layers represent different stages of the process. Different letters indicate significant differences between the layers based on post hoc test (Tukey HSD). Values are means ± SE.

Lignocellulosic residues

Lignocellulosic residues are composed by three main components: cellulose, hemicellulose and lignin. Cellulose and hemicellulose are polyssacharydes composed by units of sugar molecules [4]. Sugarcane bagasse is composed of around 50% cellulose, 25% hemicellulose and 25% lignin. It has been proposed that because of its low ash content (around 2-3%), this product offers numerous advantages in comparison to other crop residues (such as rice straw and wheat straw) when used for bio-processessing purposes. Additionally, in comparison to other agricultural residues, bagasse is considered a richer solar energy reservoir due to its higher yields on mass/area of cultivation (about 80 t/ha in comparison to about 1, 2, and 20 t/ha for wheat, other grasses and trees, respectively) [5].

1.1. Cellulose

Cellulose is composed of microfibrils formed by glucose molecules linked by p-1,4, being each glucose molecule reversed in relation to each other. The union of microfibrils form a linear and semicrystalline structure. The linearity of the structure enables a strong bond between the microfibrils. The crystallinity confers resistance to hydrolysis due to absence of water in the structure and the strong bond between the glucose chains prevents hydrolases act on the links p -1,4 [1].

1.2. Hemicellulose

Hemicellulose is a polysaccharide made of polymers formed by units of xylose, arabinose, galactose, manose and other sugars, that present crosslinking with glycans. Hemicellulose can bind to cellulose microfibrils by hydrogen bonds, forming a protection that prevents the contact between microfibrils to each other and yielding a cohesive network. Xyloglucan is the major hemicellulose in many primary cell walls. Nevertheless, in secondary cell wall, which predominate in the plant biomass, the hemicelluloses are typically more xylans and arabino-xylans. Typically, hemicellulose comprises between 20 to 50% of the lignocellulose polysaccharides, and therefore contributes significantly to the production of liquid biofuels

[1] . Sugarcane bagasse contain approximately 25-30% hemicelluloses [13,14].

Considerations for Sustainable Biomass Production in Quercus-Dominated Forest Ecosystems

Viktor J. Bruckman, Shuai Yan, Eduard Hochbichler and Gerhard Glatzel

Additional information is available at the end of the chapter http://dx. doi. org/10.5772/53518

1. Introduction

Our current energy system is mainly based on carbon (C) intensive metabolisms, resulting in great effects on the earth’s biosphere. The majority of the energy sources are fossil (crude oil, coal, natural gas) and release CO2 in the combustion (oxidation) process which takes place during utilization of the energy. C released to the atmosphere was once sequestered by biomass over a time span of millions of years and is now being released back into the atmosphere within a period of just decades. Fossil energy is relatively cheap and has been fuelling the world economy since the industrial revolution. To date, fossil fuel emissions are still increasing despite a slight decrease in 2009 as a consequence of the world’s economic crisis. Recently, the increase is driven by emerging economies, from the production and international trade of goods and services [1]."If we don’t change direction soon, we’ll end up where we’re heading" is the headline of the first paragraph in the executive summary of the World Energy Outlook 2011 [2]. It unfortunately represents systematic failure in combating climate change and the emphatic introduction of a "green society", leaving the fossil age behind. Certainly such far reaching transformations would take time, but recovery of the world economy since 2009, although uneven, again resulted in rising global primary energy demands [2]. It seems that more or less ambitious goals for climate change prevention are only resolved in phases of a relatively stable economy. Atmospheric carbon dioxide (CO2) is the second most important greenhouse warming agent after water vapour, corresponding to 26% and 60% of radiative forcing, respectively [3]. Together with other greenhouse gases (GHG’s) (e. g. methane (CH4), nitrous oxide (N2O) or ozone (Оз)) they contribute to anthropogenic global warming. The industrialization has been driven by fossil sources of energy, emerging in the 17th and 18th century in England as a historical singularity, but soon spreading globally.

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Today, our economies still rely on relatively cheap sources of fossil energy, mainly crude oil and natural gas, and consequently emitting as much as 10 PG C per year in 2010 [4]. The Mauna Loa Observatory in Hawaii carries out the most comprehensive and longest continuous monitoring of atmospheric CO2 concentration. It publishes the well-known Keeling Curve, representing the dynamic change since 1958. Observing the Keeling Curve, one can easily recognize the seasonal variability which is directly triggered by CO2 uptake of vegetation (biomass) in the northern hemisphere during the vegetation period and secondly, which is even more important in terms of global change, a steady increase of CO2 concentration from 315 ppmv in 1958 to 394 ppmv in March 2012 [5]. Earlier concentrations could still be derived from air occluded in ice cores. Neftel et al. [6] presents accurate gas concentration measurements for the past two centuries. However, the theoretical knowledge of the warming potential of CO2 in the atmosphere evolved in the late 19th century when a theory of climate change was proposed by Plass [7], pointing out the "influence of man’s activities on climate" as well as the CO2 exchange between oceans and atmosphere and subsequent acidification. He highlighted the radiative flux controlled by CO2 in the 12 to 18 micron frequency interval, agreeing with a number of studies published in the forthcoming decades, e. g. Kiehl and Trenberth [3]. In order to understand the fate of anthropogenic CO2 emissions, research soon focussed on estimating sources and sinks as well as their stability, since it was obvious that the atmospheric concentrations did not rise at the same magnitude as emissions. Available numbers on current fluxes are principally based on the work of Canadell et al., [8] and Le Quere et al., [1]. In their studies, it is emphasized that the efficiency of the sinks of anthropogenic C is expected to decrease. Sink regions (of ocean and land) could have weakened, source regions could have intensified or sink regions could have transitioned to sources [8]. Another explanation might be the fact that the atmospheric CO2 concentration is increasing at a higher rate than the sequestration rate of sinks [1]. Moreover, CO2 fertilization on land is limited as the positive effect levels off and the carbonate concentration which buffers CO2 in the ocean steadily decreases according to Denman, K. L. et al. [1]. Fossil fuel combustion and land use change (LUC) are the major sources for anthropogenic C emissions (Figure 1). Land use change is usually associated with agricultural practices and intensified agriculture triggers deforestation in developing countries [9] and consequently causes additional emissions.

Another consideration is the availability of fossil fuel, which is limited by the fact that it is a non-renewable and therefore finite resource. Since the fossil energy system is based on globally traded sources with centralized structures, the vulnerability to disturbance is high. Recent examples of price fluctuations caused by political crises or other conflicts in producing countries or along transportation lines demonstrate potential risks. Moreover, a shift towards alternative energy sources and a decentralization of the energy system may contribute to system resilience and create domestic jobs. It prevents capital outflow to unstable political regimes and it helps to protect the environment not only by reducing GHG emissions, but also by reducing impacts of questionable methods of extracting fossil sources of energy (e. g. tar sands exploitation, fracking etc.).






Fossil fuel combustion




Land use change








Figure 1. The fate of anthropogenic CO2 emissions in 2010, showing sources (left) and sinks (right). Presented numbers are Pg C yr-1. The values for 2010 were presented at the Planet under Pressure 2012 conference in London [4].

*includes cement production and flaring.

Biomass could play a significant role in the renewable energy mix. It is a feedstock for bioenergy production and currently thermal utilization (combustion) is by far the most important conversion process, but research activities are focussed on a range of different processes. This includes, for instance, the Fischer-Tropsch synthesis where any kind of biomass may be used as feedstock to produce liquid biofuels. This process is known as biomass — to — liquid (BtL). Research is pushed by national and international regulations (e. g. the EU’s 2020 bioenergy target) and commitments, as a climate change mitigation strategy.

This chapter focuses on aspects of sustainable woody biomass production in Quercus dominated forest ecosystems with emphasis on different silvicultural management systems. Short rotation woody crops (SRWC), coppice with standards (CS), high forest (HF) and Satoyama are characterized according to their biomass potential and sustainability considerations. CS and HF are directly compared based on our own research and links to similar systems (SRWC and Satoyama) are drawn in order to provide a holistic view of the current topic. The chapter aims at providing an interdisciplinary view on biomass production in forest ecosystems, considering impacts on C and nutrient metabolism as well as other effects (e. g. biodiversity, technical, silvicultural and cultural issues). Considerations for sustainable biomass production in Quercus dominated forest ecosystems are presented for each management system.