Las posturas, avances y desafíos en formación

In chapter 2, analysis of the performance parameters of AD showed that digesters pre- exposed to glycerol waste recovered 1.5 HRT faster than the control when exposed to glycerol waste again. Analysis of the P:A ratio indicated that changes in VFA production could be related to this improvement but that a deeper analysis of the VFA profile was required to understand this relationship.

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Digesters receiving glycerol waste in the feed for the first time (digesters 4-6, increase series 1 and digesters 1-3, increase series 2) showed very similar behaviour. Specifically there was an initial high production of lactic and iso butyric acid later substituted by a parallel production of propionic and acetic acid (Figure 4.4). After the second OLR increase series (increase series 2, digesters 4-6) the production of VFA was markedly different, showing that the processing of feedstock had changed in addition to (as shown in Chapter 3) the shifts in microbial community structure. Lactic acid was less than 6 % of the total organic acid fingerprint, in contrast to > 50 % after the first OLR (Figure 4.4). Accumulation of acetic and propionic acids occurred in parallel between day 91 and 105. After day 105 the production of these two acids occurred sequentially. This is in contrast with the results obtained for the digesters that were only exposed to one OLR change, where acetic and propionic acids accumulated in parallel throughout the trial. It is possible that after day 105, the propionic acid is converted into acetic acid resulting in faster recovery, as described in chapter 2. The concentration of iso-butyric acid, thought to be associated to high concentrations of lactic acid, was never above 1 g l-1, about 50 % lower than digesters only exposed to one OLR change. Therefore a lower accumulation of iso butyric acid was also linked to faster recovery. This demonstrates that after OLR change the metabolic function of the microbial community, in addition to the community resilience structure, is responsible for improved performances. A schematic for proposed pathway is presented in Figure 4.6.

After the second OLR change, when FOGs waste was used as co-digestant (digesters 13-15, OLR increase series 2) the digesters pre-exposed to glycerol waste processed VFA in the same way as digesters 10-12 not pre-exposed to glycerol waste, Figure 4.5. These results suggest that multiple OLR increase series using different feedstocks were

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not able to produce any change in the microbial metabolic pathways, which is consistent with the operational performances of the digesters observed in Chapter 2. However the microbial communities of these digesters were shown to be distinct in Chapter 3, demonstrating high functional redundancy in these microbial communities. These results highlights the value of understanding both microbial community structure and function in AD as a change in one may not always result in change of the other (Fernández et al. 1999; Fernández et al. 2000; Zumstein et al. 2000; Wang et al. 2010b; Wang et al. 2011). High functional redundancy could represent a barrier to microbial optimisation of AD. However research has shown that (including chapter 3) consistent links between performance and microbial community structure can be found in AD, particularly in the more specialist and less dominant groups of bacteria/archaea which show more correlation with physicochemical parameters (Briones and Raskin 2003; Rittmann 2006; McMahon et al. 2007; Werner et al. 2012). Therefore a deeper understanding of the microbial community using a combined approach of techniques such as lipid profiling, that can show changes in physiology of bacteria, and 454- pyroseasing, which can resolve the rare diversity, are key tools in microbial optimisation of AD.

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Figure 4.4. Concentrations of VFA in digesters 4-6 (exposed to both OLR increase

series). Red boxes indicates OLR increase series. acetic acid (black circles and solid lines) propionic acid (black circles and dashed lines) iso butyric acid (black squares, solid lines) n butyric acid (clear squares, dashed lines) and lactic acid (grey triangles, dashed lines). Triplicate digester average, error bars are not shown to improve legibility. average standard deviation for acetic acid = 0.3, propionic acid = 0.2, iso butyric = 0.08, n butyric = 0.2, lactic = 0.1, and % methane = 4.8.

Figure 4.5. Concentrations of VFA in digesters 13-15 (Exposed to both OLR increase

series). Red boxes indicates OLR increase series. acetic acid (black circles and solid lines) propionic acid (black circles and dashed lines) iso butyric acid (black squares, solid lines) n butyric acid (clear squares, dashed lines) and lactic acid (grey triangles, dashed lines). Triplicate digester average, error bars are not shown to improve legibility. average standard deviation for acetic acid = 0.4, propionic acid = 0.3, iso butyric = 0.2, n butyric = 0.1, lactic = 0, and % methane = 6.2.

10 30 50 70 90 0 1 2 3 4 5 6 7 8 0 10 20 30 40 50 60 70 80 90 100 110 120 % M e th an e g l- 1 Time (days) 10 30 50 70 90 0 1 2 3 4 5 6 7 0 10 20 30 40 50 60 70 80 90 100 110 120 % M e th an e g l- 1 Time (days)

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Figure 4.6. Schematic of digestion of glycerol waste during increase series 2 by

digesters exposed to repeated OLR increase series with glycerol waste.

Repeated and varied OLR increase using glycerol waste: The effect of an OLR

increase series using higher concentrations of glycerol waste was also investigated (digesters 7 to 9, increase series 1). The main difference between these digesters and those that received repeated OLR increase using glycerol waste at one concentration was that the production of acetic acid reached a higher concentration (5.8 ± 0.5, day 103) during the sequential phase of increase series 2, Figure 4.7. As reported in chapter 2, recovery times for biogas production and methane content were 3 days shorter in these reactors than in the one exposed to lower OLR. Although the difference was shown not to be significant (p > 0.05), it suggests than an even higher OLR increase in increase series 1 could result in further improved performance. A similar approach was used in previous research where much higher OLR were used to achieve higher resilience in AD microbial communities (18.8 kg VS m3 day-1) (McMahon et al. 2004). It is clear from these results that the effect of different OLRs on the microbial communities performances and composition needs further investigation

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Figure 4.7. Concentrations of VFA in digesters 7-9, red boxes indicate OLR increase

events, acetic acid (black circles and solid lines) propionic acid (black circles and dashed lines) iso butyric acid (black squares, solid lines) n butyric acid (clear squares, dashed lines) and lactic acid (gray triangles, dashed lines). Triplicate digester average, error bars are not shown to improve legibility. average standard deviation for acetic acid = 0.4, propionic acid = 0.2, iso butyric = 0.1, n butyric = 0.1, lactic = 0.1, and %

methane = 5.6.