First draft for Applied and Environmental Microbiology



Methane biofiltration of methane from ruminants gas effluent using autoclaved aerated concrete as the carrier material

Giovanni Ganendraa,d, Daniel Mercado Garciaa, Emma Hernandez-Sanabriaa, Nico Peirenb, Sam De Campeneereb, Adrian Hoc, Nico Boon a,#

aLaboratory of Microbial Ecology and Technology, Ghent University, Coupure Links 653, B-9000, Gent, Belgium

bInstitute for Agriculture and Fisheries Research, Animal Science Unit, Scheldeweg 68, Melle 9090, Belgium

cDepartment of Microbial Ecology, Netherlands Institute of Ecology (NIOO-KNAW),

Droevendaalsesteeg 10, 6708 PB, Wageningen, The Netherlands

dSIM vzw, Technologiepark 935, BE-9052, Zwijnaarde, Belgium

* Correspondence to: Nico Boon, Ghent University; Faculty of Bioscience Engineering; Laboratory of Microbial Ecology and Technology (LabMET); Coupure Links 653; B-9000 Gent, Belgium; phone: +32 (0)9 264 59 76; fax: +32 (0)9 264 62 48; E-mail: Nico.Boon@UGent.be; Webpage: labmet.Ugent.be.

Abstract

The capacity of Methane-Oxidizing Bacteria (MOB) immobilized on Autoclaved Aerated Concrete (AAC) in a biofilter setup to remove methane from ruminants’ gas waste was investigated. Two dairy cows (C165 and C201) were employed in respiration chambers for two days where the exhaust gas from the chambers was used as the biofilter feed. MOB comsumed methane at a removal efficiency (RE) of 17.52 ± 11.7 % and elimination capacity (EC) of 67.3 ± 53.7 g m-3 d-1. Compared to our previous lab scale study, several factors caused the lower RE and EC, namely, the lower methane mixing ratio in the real livestock waste gas, the presence of ammonia in the gas, and higher gas flow rate into the biofilter. By using AAC as carrier material, carbon dioxide in the waste gas as well as bacterial-derived carbon dioxide were removed by carbonation reaction with AAC thus, complete carbon sequestration from methane was achieved. Taken together, our results showed a more sustainable process when using abiofilter with MOB immobilized in ACC as carrier material to remove methane gas.

Keywords: Methane biofiltration, Autoclaved aerated concrete, Methane-Oxidizing Bacteria, ruminant gas waste, respiration chamber

Introduction

Among different anthropogenic sources, methane emitted from livestock contributes up to 40 % of the total global anthropogenic methane emission [1]. Effective livestock methane emission mitigation strategies should focus on methane emission originating from the ruminants as it accounts for ~90% of the total livestock methane emitted [2]. Up until now, several strategies include improving feed quality to reduce methane generation in the ruminant (e.g., addition of feed supplement) and management practices to improve meat/milk productivity (e.g., usage of antibiotics) [2-4]. However, methane is constantly produced in the gut of ruminants and retained in the waste gas. Considering the low methane mixing ratio in the waste gas, biotechnological applications to treat the gas can be a viable option as it is economically beneficial and environmentally friendly [5, 6].

Biofiltration has been applied to mitigate ruminant methane emission. Methane-Oxidizing Bacteria (MOB) are the biocatalysts used to degrade methane in the biofilter. In a biofilter setup, methane is removed by MOB immobilized on a carrier material in a fixed bed system. By possessing the Methane Mono-Oxygenase (MMO) enzyme, MOB can oxidize methane and utilize it as a carbon and energy source [7]. Although full scale biofilter application has not been established yet, several lab scale tests have been conducted to remove methane emission from piggery and dairy cows [8-10].

Different carrier materials have been demonstrated to have an optimum methane removal by the MOB. Previously, we show that Autoclaved Aerated Concrete (AAC) could be used to remove methane at low methane mixing ratio (i.e., ~1000 ppmv) in a biofilter setup. AAC is a lightweight building material exhibiting porosity up to 80% pore volume where more than 40% of the pore diameter is in the range of 5 to 100 µm [11]. With bacteria diameter around 1 to 2 µm, this makes AAC a suitable carrier material to immobilize MOB in the biofilter setup as it can accommodate a high number of bacteria per gram of material [12].

Here, we used a MOB biofilter using AAC as the carrier material previously tested in lab scale environment to remove methane from a cattle livestock gas waste. The aim of this study is to investigate the capacity of MOB immobilized on AAC in a biofilter setup to mitigate methane emission from livestock. Several challenges foreseen compared to our previous study include: (1) lower atmospheric methane mixing ratio (~50 – 100 ppmv), (2) the presence of other components in the gas waste (e.g., ammonia) that might influence the methane removal capacity of MOB, and (3) the dynamic of gas load throughout the biofilter operation (e.g., higher methane mixing ratio during rumination period).

Materials and methods

2.1 Methane biofilter

Methane biofilter A (MBF-A) from our previous test was used in this research. The biofilter was filled with AAC specimens previously inoculated with MOB mixed culture enriched from moderately alkali soil originating from Ghent, Belgium [11]. In our previous study, methane gas at ~1000 ppmv mixing ratio was used to feed the filter for 127 days with an average methane removal efficiency of 24.6 ± 18.1 %. The present study was conducted one week after the lab scale test.

2.2 Biofiltration in ruminant respiration chamber

The biofilter test was done in a ruminant respiration facility situated in the Institute for Agriculture and Fisheries Research (ILVO) in Melle, Belgium. The facility was dedicated to investigate greenhouse gas emission from livestock. In short, in this facility, the dynamic of gas emitted from ruminants under different conditions such as the feeding and milking period is investigated.

The facility consists of six different chambers where the ruminants could be placed but only two chambers (i.e., one cow for each chamber) were used in this study (Fig. 1.). Each chamber dimension was 4 m (length) x 1.55 m (width) x 2.8 m (height) and they were made from polypropylene (50 mm thick; Paneltim, Belgium) mounted on an internal stainless steel frame (total effective volume: 12.3 m3). Detail description of the facility construction can be found in [13]. During the test, the chambers were operated at slightly below atmospheric pressure. Air flew from the front door (67 cm x 37 cm) of each chamber into the exhaust system located on top of the chambers via the rear part of the chamber’s roof which was equipped with a ventilator (diameter: 35 cm; Fancom, The Netherlands). The gas was subsequently released into the atmosphere via a roof panel where an axial exhaust fan was fixed (Fancom, The Netherlands). The exhaust fan generated the air flow throughout the chamber.

Inside the exhaust system, the biofilter was placed horizontally between the roof opening of one of the chambers and the roof panel (Fig 1.). The biofilter had four ports that were used as the waste gas inlet/outlet (2 ports) and sampling gas lines (2 ports). Gas entered the biofilter via the bottom part and left from the top part of the filter (Fig 1.). The gas outlet port was connected to a ME2C pump (Vacuubrand, Germany) to create a gas flow inside the biofilter. The gas outlet after the pump was placed relatively far from the biofilter to prevent mixing with the inlet gas. The outlet gas lines (i.e., sampling and process) were made using gas leak-proof PFA tubes (Cole-Palmer, USA).

Two dairy cows (code number C165 and C201) were the ruminants employed for the test and they were bred in ILVO. One day prior to the test, C165 had the entire rumen content removed and re-inoculated with a quarter volume of his own rumen content. Both cows were in the second lactation phase and had been fistulated. C165 and C201 were 42 and 39 months old, respectively.

All biofilter connections were opened an hour before both cows entered the chambers. This was done so that the MOB in the biofilter could acclimatize to the environmental conditions around the exhaust system. The summary of the biofilter process and technical parameters can be seen in Table 1. The test was performed from the 20th (5 pm) to the 22nd of August 2014 (4 pm). The cows entered the chambers on the 20th of August at ~6 pm. During the measurement campaign, cleaning the manure trays and feeding/milking of the cows were done at three separate periods. For C165, they were at 8.30 am (21st August), 17.40 pm (21st August), 8 am (22nd August) whereas for C201, they were at 8 am (21st August), 17.10 pm (21st August), 7.45 am (22nd August). The summary of the cows’ feed composition can be seen in Table 2. After the test, both cows were guided out from the chambers and the biofilter was removed from the exhaust.

2.3 Gas composition analysis

Gas sample measurements started as soon as the connections to the biofilter ports were opened (i.e., 20th august at 5 pm). Gas samples taken from the sampling ports were fed directly to the gas analyzer. Additionally, two sampling ports were placed at the exhaust fan of each chamber to analyze the gas composition of the waste gas from each cow. The gas analyzer was a photoacoustic multi-gas MonitorInnova 1312 (LumaSense Technologies, Denmark) and it was used to measure methane, carbon dioxide, nitrous oxide, ammonia, and water mixing ratio. The gas sampling lines from the biofilter were first connected to a Vacu-Guard filter (Whatman, UK) before going to the analyzer. In the analyzer, the tubes were connected to an eight-channel multi sampler (AP2E, France) via a PFA tubing. The parameters used for the biofilter evaluations were methane removal efficiency (RE) and elimination capacity (EC). Additionally, the methane emission rate from the chambers were evaluated. The RE was calculated using the following equation:

RE(%) = ((Cin – Cout)/Cin) X 100% (1)

where Cin and Cout were the methane inlet (ppmv) and outlet concentrations (ppmv), respectively, from the biofilter. The EC was calculated using the following equation:

EC (g m-3 h-1) = (Qb/Vf) X (Cin – Cout) (2)

where Vf is the filter bed volume (m3) and Qb is the inlet gas flow rate (m3 h-1). The methane emission rate was calculated from the following equation:

Methane emission (g d-1) = Qf X (Cchamber – Cbackground) X 10-6 X M/Vm (3)

where Cchamber was the methane mixing ratio analyzed from each chamber (ppmv), Qf was the exhaust fan flow rate (m3 h-1), Cbackground was the background methane mixing ratio (ppmv), M was the methane molecular weight (16 g mol-1), and Vm was the molar volume of methane at 19.1 °C and 0.89 atm (26.9 L mol-1).

3. Results and discussion

3.1 Ruminants gas emission from the respiration chambers

Similar gas mixing ratio profile was examined in the exhaust gas from both chambers during the measurement campaign (Fig 2.). Hence, similar average mixing ratio of the gasses was observed: 55.5 ± 23.6 ppmv (C165) and 54.3 ± 21.7 ppmv (C201) for methane, 1.4 ± 0.4 ppmv (C165 and C201) for ammonia, 0.3 ± 0.3 ppmv (C165 and C201) for nitrous oxide. Higher average carbon dioxide mixing ratio was analyzed in the gas emission from C165 chamber (1068.3 ± 581 ppmv) than the one of C201 (1032.6 ± 532 ppmv). C165 and C201 emitted 306.8 ± 134.5 g methane d-1 and 301.2 ± 121.4 g methane d-1, respectively.

Ruminants produce methane as a result of the microbial digestion of the food in the rumen and large intestines [14]. Protein, starch, and other polysaccharides are hydrolyzed and fermented partly to hydrogen and acetic acid. These components are subsequently converted to methane by methanogens. Methane represents energy loss from the food digestion process and it is emitted primarily by eructation [15]. Methane can also be emitted by flatulation or methanogenesis in the anoxic part of manure, however, they do not represent a significant methane emission from the ruminant. Carbon dioxide is produced from both respiration and eructation. Ammonia and nitrous oxide are emitted from the ammonification and nitrification/denitrification processes in the manure [16]. These components were the main emission constituents in the ruminant waste gas.

The dynamic of the methane emission in both chambers was mainly influenced by eructation.

Higher methane mixing ratio in the waste gas was observed in between meals or in the evening (Fig. 2) which coincided with the rumination period (i.e., secondary ruminal contraction). [17]. In agreement with previous studies, lower eructation methane emission was analyzed before the morning feeding period [18, 19].

Only the dynamic of the carbon dioxide emission was identical with methane in both chambers (Fig. 2). Similar to methane emission dynamics, increasing carbon dioxide emission is usually observed during eructation [20]. Higher carbon dioxide mixing ratio during eructation was due to the added emission from the bacterial metabolism (e.g., fermentation) occurred in the rumen [18]. In contrary with carbon dioxide emission profile, the dynamic of ammonia and nitrous oxide emissions differed from methane as both ammonia and nitrous oxide were mostly emitted from manure and not from the cow itself. Moreover, the ammonia and nitrous oxide mixing ratio was only slightly lower compared to previous studies [20, 21]. In conclusion, the effluent gas from the chambers were comparable to the one typically observed from a ruminant in a respiration chamber.

3.2 The biofilter performance

Methane was removed from the cows’ gas waste in the biofilter throughout the test period with varying removal efficiency (Fig. 3). The methane mixing ratio in the biofilter inlet was relatively stable with an average concentration of 61.9 ± 15.6 ppmv. Prior to the cows’ presence in the chamber, the methane mixing ratio in the biofilter inlet was similar to the background concentration indicating that the additional methane entering the filter afterwards was originating from the cows.

The average RE (17.52 ± 11.7 %) and EC (67.3 ± 53.7 g m-3 d-1) were lower than the one observed in our lab scale test [ref] which was caused by several factors. Firstly, methane at much lower mixing ratio was fed into the filter compared to the previous study (~1000 ppmv). The activity of the immobilized MOB on building material followed the hyperbolic Michaelis-Menten kinetic model and at lower methane mixing ratio, smaller activity was exhibited by the bacteria [11]. Secondly, the biofilter operated with much lower residence time in this study as the gas flow rate was six times higher (Table 2). This decreased the component’s contact time with the bacteria and therefore the substrate conversion. Thirdly, the presence of ammonia may competitively inhibit the MMO. ; MMO is homologous to the Ammonium Mono-Oxygenase enzyme possessed by Ammonia-Oxidizing Bacteria [22]. According to the Michaelis-Menten kinetic model, a substrate competitive inhibition would lower the enzyme affinity to the substrate (1/Km) and at low substrate concentration, the kinetic model followed the first order kinetic [23]. Therefore lower conversion rate at the same substrate concentration would be obtained.

Beside methane, carbon dioxide and ammonia were also removed in the biofilter (Fig. 4). Methane is converted by MOB to synthesize new biomass and carbon dioxide [7], however, at low mixing ratio (0-100 ppmv), a complete methane conversion to carbon dioxide occurs [24]. However, higher carbon dioxide emission was not observed in the biofilter outlet (Fig. 4), presumably, due to the carbonation reaction of carbon dioxide with the binder material (i.e., tobermorite -1.1 nm) of AAC. Thus, a complete carbon sequestration from methane was achieved [11]. Moreover, due to the ammonia binding and conversion in the MMO, lower ammonia mixing ratio in the biofilter outlet was shown. Nitrous oxide was not removed from the waste gas whereas higher water mixing ratio was observed in the biofilter outlet. Higher water content in the filter outlet was observed due to the water evaporation from the biofilter. Future, research should investigate water addition into the biofilter to prevent the evaporation for water is essential in the bacterial metabolism.

4. Conclusions

Here, the capacity of MOB in a biofilter setup using AAC to remove methane from ruminants’ gas waste in a respiration chamber was investigated. MOB removed methane at a RE of (17.52 ± 11.7 %) and EC of (67.3 ± 53.7 g m-3 d-1). Compared to the lab scale test, several factors lowered the methane removal capacity of the bacteria, namely, the lower methane mixing ratio,higher gas flow rate into the biofilter, and the presence of ammonia in the waste gas. The use of AAC as the filter bed gave an added advantage compared to other materials by removing carbon dioxide resulting from bacterial metabolism and the ruminants by carbonation reaction with its binder material.

Acknowledgements

The project is funded by SIM-SHE SECEMIN project (SIM 2009-1). Special thanks to Thijs Mulder for the technical help and Amanda Luther for critically reviewing this paper. This publication is Publication No. xxx of the Netherlands Institute of Ecology.

References

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Figures

[pic]

Figure 1. Biofilter configuration in the exhaust system of the ruminant respiration chamber

[pic]

Figure 2. Mixing ratio of (a) methane, (b) carbon dioxide, (c) ammonia, and (d) nitrous oxide in waste gas from each chamber where cow C165 and C201 resided. Black lines indicate the change of day. Solid (C201) and dashed (C165) grey lines indicate cow feeding times and milking period.

[pic]

Figure 3. (a) The methane mixing ratio at the inlet and outlet of the biofilter and (b) the methane removal efficiency in the biofilter. Black lines indicate the change of day.

[pic]

Figure 4. Mixing ratio of (a) carbon dioxide, (b) ammonia, (c) nitrous oxide, and (d) water at the inlet and outlet of the biofilter. Black lines indicate the change of day.

Tables

Table 1. Overview of the biofilter operating and technical parameters

|Parameter |Value |

|Empty biofilter volume (m3) |0.0051 |

|Biofilter bed volume (m3) |0.0026 |

|Biofilter bed mass (kg) |1.02 |

|Flow rate biofilter (m3 h-1) |1.2 |

|Flow rate chambers’ exhaust fan (m3 h-1) |400 |

|EBRT (s) |15.3 |

|Temperature (° C)a |19.1 |

a Average chamber temperature during the measurement campaign

Table 2. Overview of the cows’ diet composition

|Feed |Amount (kg) |

|Maize silage |21.3 |

|Haylage |11.7 |

|Corn cob mixture |1.5 |

|Sugar beet pulp |5 |

|Balanced concentrate (F10-14)a |0.8 |

|Balanced concentrate (F09-08)a |0.9 |

|Soya |0.4 (C165) |

| |0.5 (C201) |

a F-10-14 and F09-08 were commercial concentrate products (Vermeulen, Belgium)

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