SULPHUR HEXAFLUORIDE TRACER TECHNIQUE FOR MEASURING METHANE DIRECTLY FROM RUMEN OF DAIRY COWS VALIDATED WITH RESPIRATION CHAMBERS

BAYAT, A.R.¹, STEFAŃSKI, T.¹, LUUKKONEN, T.¹, KAIRENIUS, P.¹, LESKINEN, H.¹, VILKKI, J.²

1 Milk Production Solutions, Green Technology, Natural Research Institute Finland (Luke), FI-31600 Jokioinen, Finland

2 Animal Genomics, Green Technology, Natural Research Institute Finland (Luke), FI-31600 Jokioinen, Finland

ABSTRACT: Traditionally respiration chambers are used to measure methane (CH4) emissions from individual animals but due to limitations, alternative techniques must be developed and validated. Sulphur hexafluoride (SF6) tracer technique was used to estimate CH4 emissions directly from the rumen of cannulated animals fed different diets. Ten cannulated dairy cows were used for 3-5 days to compare SF6 technique and respiration chambers simultaneously in an incomplete change-over design with three periods and four different diets resulting in 29 cow/period comparisons. The diet and period did not influence the ratio of CH4 measured by both techniques. The average CH4

emissions were 485 and 510 g/d for SF6 and chamber techniques, respectively.

Repeatability of daily CH4 measurement was 0.52 and 0.90 for SF6 and chamber techniques, respectively. Residual coefficient of variation was 11.2 and 3.24%, for daily CH4 emissions and 5.79 and 3.25 for cow/period averages for SF6 and chamber techniques, respectively. Lin’s concordance correlation coefficient between two techniques was 0.62 and 0.76 for daily or cow/period average CH₄ emissions, respectively. It is concluded that SF6 technique can be used as an alternative method to estimate CH4 emissions using rumen gas sampling from cannulated cows.

Keywords: Dairy cow, CH4, Measuring method, Sulphur hexafluoride, Respiration chambers

INTRODUCTION: Generally respiration chambers are considered as gold standard due to the accuracy for measuring CH4 emissions from individual animals. However, they cannot be exploited under grazing conditions, are expensive and laborious, and can measure a limited number of animals over time. Therefore alternative methods must be developed and validated. Sulphur hexafluoride (SF6) tracer technique was first used by Johnson et al. (1994) using breath samples in ruminants. The technique has been used mainly under grazing conditions but a variant of the technique is used in animals fitted with rumen cannulae allowing collection of gas samples directly and continuously from the rumen (Boadi et al., 2002; Bayat et al., 2015). The objective of this study was to compare CH4 emissions measured directly from rumen using SF6 tracer technique and those measured using respiration chambers simultaneously.

1. MATERIAL AND METHODS: ten dairy cows in mid-lactation fitted with rumen cannulae (#1c, i.d. 100 mm; bar diamond, inc., parma, id) were used. the study was conducted as incomplete change-over design with three periods resulting in 29

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cow/period comparisons. the cows received four diets including low, medium and high forage diets based on grass (forage to concentrate ratio 30:70, 50:50 and 70:30 on dm basis, respectively) or red clover (50:50 on a dm basis) silages for 14 days before sampling (overall 6, 8, 8 and 7 cows per diet, respectively). milk yield, dry matter (dm) intake and ch4 emissions were measured over 3-5 days from day 15 of each experimental period. the ch4 measurements were conducted for both techniques simultaneously inside the chambers. ruminal ch4 emissions were measured using the sf6

tracer technique (boadi et al., 2002). gases in the rumen headspace were drawn continuously (1.7 ml/min) over every 24-h period into evacuated 5.5 l air-tight canisters (-0.9 and -0.4 bar in the beginning and end of sampling, respectively) through sampling tubes (4-mm i.d.) and 100 cm of capillary tubing (peek 1.6 mm × 0.13 mm i.d., vici valco instruments co, houston, tx, usa). a filter with 0.2 µM was used to prevent rumen liquor entering the capillary tube. a t-shape connection was used for the tip of the sampling tube to ensure a better flow of the rumen gas to the sampling line and the open heads were covered with nylon cloth (17-µM pore size) to prevent the entrance of and blockage by rumen particulates. tubes used to collect the ruminal gas were anchored securely to the neck of the rumen cannula allowing gas to be collected at approximately 5 cm above the rumen mat. the sf6 releasing tubes (measured rate of release 1.11±0.31 mg/d) were suspended in the rumen via cannulae. sub-samples of ruminal gases were analysed in triplicate for ch₄ and sf₆ concentrations by gas chromatography (agilent 6890n, agilent technologies, santa clara, ca, usa). daily ruminal ch4 emissions were calculated based on the measured sf6 release rate in the rumen over the course of experiment and concentrations of CH4 and SF6 in analysed rumen gases.

Four open-circuit respiration chambers, located in the Minkiö dairy barn, Jokioinen, Finland, were used to measure CH4 within each period for 3-5 consecutive days.

Concentrations ofCH4 in the inlet and exhaust airflow were measured using dedicated analysers (Columbus Instruments., Columbus, OH, USA) with 3.5 min interval for each chamber and the reference air. Gas analysers were calibrated using the standard gases in the beginning of each measurement. Air flow was measured for every chamber using mass flow meter (HFM-200, Teledyne Hastings Instruments, Hampton, VA, USA) while corrected for temperature and pressure. An adjustable air conditioning system (Flow 500-2000 L/min; cooling capacity 2.9 kW; heating capacity 3.2 kW) allowed mixing the air inside the chambers and environmental control of temperature across a range of 12-22°C and a relative humidity of 50-70% monitored using electronic sensors. For cow welfare and safety, an emergency door sensitive to electricity failure or high CO2

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from further calculations. Lin’s concordance coefficient (LCC) for continuous variables was calculated to evaluate the equivalence between the techniques (http://services.niwa.co.nz/services/statistical/concordance). Linear regression analysis was used to find the best equation fitting to the data.

2. RESULTS AND DISCUSSION: feed intake, milk yield and ch4 emissions measured by SF6

or respiration chamber techniques across all periods, are presented in table 1. The overall average of CH4 emission (g d^-1) and intensity (g kg^-1 milk) were lower (p<0.01 482 vs. 510 and 12.5 vs. 13.2, respectively) and CH4 yield (g kg^-1 dm intake) tended to be lower (p=0.07; 20.8 vs. 22.0) for SF6 compared with chambers. lower CH4 emissions from SF6 compared with chambers (5.5%) can be expected as the SF6 technique measures only the emissions from rumen. it was estimated that about 3% of total enteric CH4 is excreted via rectum of cattle (grainger et al., 2007; muñoz et al., 2012). repeatability of daily CH4 measurements was 0.52 and 0.90 for SF6 tracer and chamber techniques, respectively.

The CV of daily CH4 emissions for SF6 and chamber techniques were 11.2 and 3.24%, and the respective values were 5.79 and 3.25% for cow/period average CH4 emissions, respectively. These results confirm the previous findings of CH4 emissions measured by SF6 tracer technique having greater CV compared with respiration chambers (Grainger et al., 2007; Muñoz et al., 2012). The LCC between two techniques was 0.62 and 0.76 for daily or cow/period average CH4 emissions, respectively. The LCC improved for CH4 yield and intensity (0.87 and 0.85, respectively for cow/period average CH4 emissions). These findings based on ruminal gas analysis are consistent with those from breath sampling of animals (Muñoz et al., 2012; Deighton et al., 2013).

Figure 1. The relationship between cow/period averages of a) methane emissions (g d^-1) and b) methane yield (g kg^-1DM intake) measured simultaneously by SF6 and respiration chamber techniques for individual

cows. The solid line depicts the line with 1:1 fit and the dotted line depicts the best linear line fitting to the data.

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Table 1. Summary statistics of feed intake, milk yield, and methane emissions during the sulphur hexafluoride (SF6) technique and chamber measurements (n=29).

Item Mean SD Minimum Maximum

DM intake (kg d^-1) 23.5 3.17 17.1 31.6

Milk yield (kg d^-1) 39.1 5.28 31.6 50.3

Methane (g d^-1)

SF6 technique 482 63.2 343 637

Chamber 510 54.8 384 605

Methane yield (g kg^-1 DM intake)

SF6 technique 20.8 3.34 14.3 29.1

Chamber 22.0 3.01 16.0 29.1

Methane intensity (g kg^-1 milk)

SF6 technique 12.5 2.13 8.84 17.2

Chamber 13.2 2.04 9.71 16.6

3. CONCLUSION: The SF6 tracer technique can be used as an alternative method to estimate CH4 emissions using rumen gas sampling from cannulated cows even though the technique has more variability compared with the respiration chambers.

Acknowledgements. The financial support from Academy of Finland to conduct this study as part of Global Network project is appreciated.

REFERENCES:

Bayat, A.R., Kairenius, P., Stefański, T., Leskinen, H., Comtet-Marre, S., Forano, E., Chaucheyras-Durand, F., Shingfield, K.J., 2015. Effect of camelina oil or live yeasts (Saccharomyces cerevisiae) on ruminal methane production, rumen fermentation, and milk fatty acid composition in lactating cows fed grass silage diets. J. Dairy Sci. 98, 3166-3181.

Boadi, D.A., Wittenberg, K.M., Kennedy, A.D., 2002. Validation of the sulphur hexafluoride (SF6) tracer gas technique for measurement of methane and carbon dioxide production by cattle. Can. J. Anim. Sci. 82, 125-131.

Deighton, M.H., Williams, S.R.O., Eckard, R.J., Boland, T.M., Moate, P.J., 2013. High concordance of CH4 emissions is possible between the SF6 tracer and respiration chamber techniques. Advances in Animal Biosciences 4, 411.

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