Effects of bioaugmentation by an anaerobic lipolytic bacterium on anaerobic digestion of lipid-rich waste
Summary (3 min read)
INTRODUCTION
- Lipids (characterized as oil, grease, fat and free long chain fatty acids, LCFAs) can be a major organic component in wastewater.
- Studies have shown that the hydrolysis of lipids to glycerol and LCFAs can be rapid, and that the main problem during lipid digestion is the further degradation of LCFAs.
- 6, 7 However, the substrate interface area available for hydrolysis may be a limiting factor.
- The possibility of pretreatment with enzyme-producing aerobic microorganisms has been demonstrated with a lipolytic fungus 13 and with mixed bacterial cultures comprising lipase, protease and amylase producers.
Substrate
- This waste consisted of a one week basis sample from the waste produced in the restaurant.
- The amount of each component was based on the chemical oxygen demand (COD); 10% was the contribution from lipids, 45% from protein, and 45% from carbohydrate (30% from starch and 15% from cellulose) (Table 1 ).
- Nutrients with the following final composition were added to ensure that no nutrient deficiency would occur (mg dm −3 ): 18.
Bioaugmenting strain
- The bioaugmenting lipolytic strain (Clostridium lundense, DSM 17 049 T ) was isolated from bovine rumen fluid.
- The culture broth was washed once with oxygen-free distilled water to remove most of the remaining substrate.
- The amount of lipolytic strain biomass added corresponded to 1.3% of the VS of the methanogenic inoculum added.
Experimental set-up
- The effect of bioaugmentation on the overall biomethanation process was studied under two kinds of conditions: treatment (A), methanogenic conditions where the lipolytic strain biomass was added on day 0 together with the methanogenic inoculum; and treatment (B), initially acidogenic conditions followed by methanogenic conditions.
- Only the lipolytic strain biomass was added at day 0, and after 4 days the methanogenic inoculum was added.
- Control experiments were also performed with substrate and methanogenic inoculum in the presence (control 1) and absence of inactivated lipolytic strain biomass (control 2).
- Assays were run using 11 replicates and at liquid phase sampling, one vial was randomly taken for analysis.
- 1, 11 Three bottles were used for gas phase studies during the experiment and the liquid contents in these were analysed at the end of the experiment.
Analysis
- 20 Gas composition was analysed using a Varian 3350 GC-TCD (Walnut Creek, CA, USA) in accordance with the method of Mshandete et al.
- The neutral lipid fraction was eluted with 1.5 cm 3 chloroform and the eluate was evaporated under nitrogen.
- LCFA methyl esters were analysed using a Varian 3400 GC-FID as described by Lyberg et al.
- 25 The COD of the substrate components was determined using suspensions of each component, which were homogenized using a homogenizer Disp 25 (20 500 rpm; Inter Med, Roskilde Denmark).
- Total Kjeldahl nitrogen was determined after digestion of samples according to the manufacturer's instructions by colorimetric analysis using a FIAStar 5000 analyser coupled to a 5027 sampler (Foss Tecator AB H öganäs, Sweden).
Methane production
- For all conditions investigated, the percentage of substrate methanization was above 90%, i.e. the fraction of the theoretical methane yield that was obtained experimentally (Table 2 ).
- The methane yields obtained were similar for test samples and controls for both treatments (Table 2 ).
COD and pH
- The initial soluble COD was similar for test samples and controls (Fig. 2 ).
- In treatment A the soluble COD observed was slightly higher when the active lipolytic strain biomass was present.
- After day 15, the soluble COD decreased for the test sample and controls under treatment A conditions.
- In treatment B, no differences between the test sample and control with inactivated lipolytic strain biomass were observed.
- In treatment A the pH showed substantial differences between test sample and controls, with the pH always lower in the test sample than in controls.
Biomass-associated LCFAs and lipolytic activity
- The biomass-associated LCFAs were analysed only in treatment A since adding the lipolytic strain biomass produced significant effects.
- This observation was in agreement with the lipolytic activity data (data not shown).
- The highest activity was observed at the beginning of the experiment.
- After the initial high concentration, the oleate concentration decreased to low values, while stearate and palmitate concentrations increased to values above 7.5 and 40 mg g −1 d.w. (d.w., dry weight), respectively (Fig. 3 ).
- No significant differences were observed between the test sample and controls for myristate and the concentrations were the lowest observed, 2-4 mg g −1 d.w.
VFAs
- The concentration of acetate was higher in the test sample for treatment A. After day 15, the concentration of acetate decreased while a slight increase in propionate concentration was observed.
- Other VFAs showed similar levels in test sample and controls for both treatment conditions investigated (Fig. 4 ).
- The initial hydrolysis of the lipid fraction of the substrate had no significant effect on the VFAs produced (data not shown).
DISCUSSION
- When the effect of bioaugmenting the anaerobic digestion process with a specific microorganism is positive, it can result in two effects: an enhancement in methane yield and/or an increase in the methane production rate.
- In view of this, the effect of bioaugmentation of the process with an anaerobic lipolytic microorganism rather than just the enzyme, was seen as an interesting alternative.
- The results of a kinetic study of the influence of biomass-associated LCFA concentration on methane production rate conducted by Pereira and coauthors 27 indicated that for concentrations below approximately 1000 mg COD-LCFA g −1 VS, there was no inhibition of methane production rate.
- The slight increase in propionate concentration observed in treatment A cannot be explained by the simultaneous decrease in stearate and palmitate concentrations.
- Therefore, once these acids start to degrade, an increase in acetate concentration would be observed only if aceticlastic methanogenesis was inhibited.
CONCLUSIONS
- In this study the effect of bioaugmentation with an anaerobic lipolytic microorganism on the anaerobic digestion process was evaluated using a model substrate containing triolein as lipid.
- It was concluded that the addition of the bioaugmenting lipolytic strain led to an increase in the methane production rate and accordingly, a reduction in the digestion period required to obtain the same methane yield as the control.
- It was not possible to draw conclusions about the survival of the bioaugmenting lipolytic strain during the experiments.
- Overall the LCFAs appeared to limit the complete conversion of the substrate to methane and carbon dioxide.
- Ways of overcoming the problems related to β-oxidation must thus be further investigated.
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...…and some industrial wastewaters, such as those from slaughterhouses, edible oil processing industry, restaurant waste, dairy industries, fish industry or fat refineries, and sewage sludge (Gannoun et al., 2009; Luste et al., 2009; Fernandez et al., 2005; Perle et al., 1995; Cirne et al., 2006)....
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"Effects of bioaugmentation by an an..." refers methods in this paper
...…were measured using HPLC in accordance with the method of Mshandete et al.21 Lipase activity was measured in accordance with the method of Winkler and Stuckmann (1979).26 The assay, using p-nitrophenylpalmitate (0.30 g dm−3) (Sigma St Louis, MO, USA) as substrate in Sörensen phosphate…...
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Frequently Asked Questions (9)
Q2. What is the effect of bioaugmenting the anaerobic digestion process?
When the effect of bioaugmenting the anaerobic digestion process with a specific microorganism is positive, it can result in two effects: an enhancement in methane yield and/or an increase in the methane production rate.
Q3. What is the significance of the bioaugmentation strategy?
The presence of higher concentrations of stearate and palmitate throughout the experiment was a clear indication that the bioaugmentation strategy improved the hydrolysis.
Q4. What is the main drawback of bioaugmentation under anaerobic conditions?
The major drawback of using bioaugmentation under anaerobic conditions is the accumulation of LCFAs, which may inhibit the digestion process.
Q5. What is the effect of hydrolysis on the digestion of a poultry waste?
Salminen and co-workers11 reported that hydrolysis limited the digestion of a poultry slaughterhouse waste due to a high concentration of propionate, which was the consequence of the presence of LCFAs.
Q6. What is the effect of bioaugmentation on the digestion of wastes?
From a practical point of view, the effect of utilization of bioaugmentation as a strategy to improve the digestion of lipid-containing wastes is a decrease in the time required for digestion.
Q7. What was the problem with the approach adopted in this study?
The problem with the approach adopted in this study was that as hydrolysis occurred rapidly, the lipolytic activity could not be accurately related to the presence of an active lipolytic strain.
Q8. What was the abundant acid in the test sample?
Acetate (6–8 g dm−3) and propionate (2.5–4 g dm−3) were the most abundant acids for both treatment conditions investigated (Fig. 4).
Q9. What is the effect of bioaugmentation on the digestion of lipid-rich waste?
15,16In the present study, the effect of bioaugmentation by an anaerobic lipolytic strain as a means of improving hydrolysis and solubilization of lipids in the anaerobic digestion process of restaurant lipid-rich waste was studied using a model substrate.