Abstract
The purpose of this experiment was to study the effect of the physical form of rapeseed fat on methane (CH4) mitigation properties, feed digestion, and rumen fermentation. Four lactating ruminal-, duodenal-, and ileal-cannulated Danish Holstein dairy cows (143 d in milk, milk yield of 34.3 kg) were submitted to a 4 × 4 Latin square design with 4 rations: 1 control with rapeseed meal (low-fat, CON) and 3 fat-supplemented rations with either rapeseed cake (RSC), whole cracked rapeseed (WCR), or rapeseed oil (RSO). Dietary fat concentrations were 3.5 in CON, 5.5 in RSC, 6.2 in WCR, and 6.5% in RSO. The amount of fat-free rapeseed was kept constant for all rations. The forage consisted of corn silage and grass silage and the forage to concentrate ratio was 50:50 on a dry matter basis. Diurnal samples of duodenal and ileal digesta and feces were compiled. The methane production was measured for 4 d in open-circuit respiration chambers. Additional fat reduced the CH4 production per kilogram of dry matter intake and as a proportion of the gross energy intake by 11 and 14%, respectively. Neither the total tract nor the rumen digestibility of organic matter (OM) or neutral detergent fiber were significantly affected by the treatment. Relating the CH4 production to the total-tract digested OM showed a tendency to decrease CH4 per kilogram of digested OM for fat-supplemented rations versus CON. The acetate to propionate ratio was not affected for RSC and WCR but was increased for RSO compared with CON. The rumen ammonia concentration was not affected by the ration. The milk and energy-corrected milk yields were unaffected by the fat supplementation. In conclusion, rapeseed is an appropriate fat source to reduce the enteric CH4 production without affecting neutral detergent fiber digestion or milk production. The physical form of fat did not influence the CH4-reducing effect of rapeseed fat. However, differences in the volatile fatty acid pattern indicate that different mechanisms may be involved.
Key words
Introduction
Globally, agriculture accounts for 47% of total anthropogenic methane (CH4) emissions, with enteric fermentation contributing 32% of the total non-CO2 emissions from agriculture in 2005 (
Smith et al., 2007
). The CH4 production per animal varies depending on the feed composition, feed quality, and production level from 2 to 12% of the gross energy (GE) intake (- Smith P.
- Martino D.
- Cai Z.
- Gwary D.
- Janzen H.
- Kumar P.
- McCarl S.
- Ogle S.
- O’Mara F.
- Rice C.
- Scholes B.
- Sirotenko O.
Agriculture.
in: Metz B. Davidson O.R. Bosch P.R. Dave R. Meyer L.A. Climate Change 2007: Mitigation. Contribution of Working Group III to the Fourth Assessment Report of the Intergovernmental Panel on Climate Change. Cambridge University Press,
Cambridge, UK2007: 497-540
Johnson and Johnson, 1995
) under extreme circumstances, but values between 3 and 7% (Martin et al., 2008
) are more realistic in intensive dairy production.Numerous studies have discussed nutritional possibilities to reduce the enteric CH4 production (
Boadi et al., 2004
; Beauchemin et al., 2008
), and fat supplementation is among the most promising tools to depress CH4 production from ruminants (Martin et al., 2008
). Furthermore, fat is fed to dairy cows to increase the energy density of the ration or to alter the product quality (Beauchemin et al., 2007
). Several oils and oil seeds have been tested for their potential to reduce the CH4 production, and effects of chain length and saturation have been reported. The degree of saturation is important, as the negative effect on bacterial growth increases with the degree of unsaturation, inhibiting both fibrolytic bacteria and methanogens (Giger-Reverdin et al., 2003
). With reduced fiber digestibility and a shift in fermentation pattern, less hydrogen arises and, thus, less CH4 (Boadi et al., 2004
). Additionally, fat often replaces carbohydrates in the ration, thereby directly reducing rumen fermentation. Reduced fiber digestibility is associated with reduced DMI and milk production; therefore it has to be considered whether the overall reduction of CH4 production from the animal due to the addition of fat is accompanied by a reduction per kilogram of product or per kilogram of feed digested.Feeding whole seeds or cake, a by-product from the plant oil production, as well as pure oil, are tools to increase the dietary fat concentration, but the difference in physical form might influence the effect of fat in the rumen.
Czerkawski et al., 1966
showed that the effect of FA on CH4 production was stronger when the same amount was infused to the rumen once daily, compared with continuous infusion. Similarly, Machmüller et al., 2000
concluded that this could be of importance to achieve momentarily high fat concentrations in the rumen rather than a constant presence at a lower level. Oil in seeds is stored intracellularly, and the fat release depends on the digestion and breakdown of the cell wall, which leads to a slower release compared with feeding oil directly (Steele et al., 1971
). This indicates that pure oil may increase the rumen FA concentration faster and reduce the CH4 production more effectively compared with seeds or cake (Martin et al., 2008
).Oilseed rape (Brassica napus) is widely grown in many countries. The by-products, rapeseed meal and cake, remaining after oil extraction are common feed components in dairy cow rations. Rapeseed meal has a low crude fat concentration (about 4%) compared with rapeseed cake (10–20%) and whole seeds (approximately 50%). The aim of the current experiment was to study the effect of rapeseed fat and the physical form in which it was fed on enteric CH4 production, rumen fermentation, and digestion.
Materials and methods
Animals and Rations
The experiment complied with the guidelines of the Danish Ministry of Justice with respect to animal experimentation and care of animals under study. Four lactating Danish Holstein dairy cows (1 primiparous and 3 multiparous) were assigned to 1 of 4 rations over 4 periods according to a balanced Latin square design; each period consisted of 4 wk. One cow was omitted from the last period due to disease.
The cows were 143 DIM (SD = 74 d), had a milk yield of 34.3 kg (SD = 8.6 kg), and a BW of 592 kg (SD = 81 kg) at the beginning of the experiment. All animals were fitted with a ruminal cannula (#1C, Bar Diamond Inc., Parma, ID), a duodenal cannula (open T-piece placed 60 cm caudal to pylorus), and an ileal cannula (open T-piece placed 20 cm cranial to the cecum). The cows were housed in a tie stall with rubber mats and sawdust as bedding and had free access to water. They were milked and fed twice daily at 0500 and 1700 h. Total mixed rations were prepared once a day and fed to the cows on an ad libitum basis after milking. The feed intake was recorded on a daily basis. The animals were weighed at the start of the experiment as well as just before and after the respiration chamber measurements (the last week of each period).
The rations were a control ration (CON) and 3 high-fat rations with fat supplemented as either rapeseed cake (RSC), whole cracked rapeseed (WCR), or rapeseed oil (RSO), respectively. The amount of fat-free rapeseed was equal for all rations, as the basic rapeseed meal content in the CON was reduced according to the fat-free rapeseed which was supplemented with either cake or seed in the treatment rations. Rapeseed cake, whole rapeseed, and rapeseed oil were obtained from Danraps (DLG Food Oil, Dronninglund, Denmark). The rapeseed used in this study was double-00 rape, equivalent to what is known as canola in North America.
The chemical composition of ingredients is shown in Table 1. All rations were fed as TMR with a forage to concentrate ratio of 50:50 (Table 2). The forage consisted of 54% corn silage and 46% prewilted perennial ryegrass silage (on DM basis). The corn silage was stored in a bunker silo and the grass silage in bales.
Table 1Chemical composition (g/kg of DM unless otherwise noted) of feedstuffs.
| Item | Barley | Beet pulp | RS meal | RS cake | Whole RS | RS oil | Grass silage | Corn silage |
|---|---|---|---|---|---|---|---|---|
| DM, g/kg of fresh matter | 878 | 903 | 904 | 968 | 940 | 424 | 295 | |
| OM | 980 | 958 | 922 | 937 | 960 | 896 | 966 | |
| CP | 115 | 102 | 381 | 300 | 192 | 163 | 99 | |
| Crude fat | 30.0 | 4.8 | 54.7 | 173 | 479 | 1,000 | 30.8 | 30.0 |
| NDF | 161 | 408 | 260 | 201 | 150 | 420 | 392 | |
| INDF | 36.4 | 28.2 | 96.7 | 90.0 | 53.6 | 56.8 | 92.1 | |
| Gross energy, MJ/kg of DM | 18.2 | 17.5 | 19.6 | 22.3 | 28.7 | 39.6 | 17.8 | 18.7 |
| NEL, MJ/kg of DM | 9.03 | 7.90 | 8.68 | 9.99 | 14.35 | 22.3 | 6.55 | 7.00 |
| OMD,% | 75.2 | 72.7 | ||||||
| Fatty acids, g/kg of DM | ||||||||
| C16:0 | 6.30 | 1.99 | 3.53 | 8.44 | 19.0 | 43.7 | 2.93 | 3.73 |
| C18:0 | 4.20 | 0.06 | 0.74 | 2.46 | 6.35 | 15.5 | 0.30 | 3.60 |
| C18:1 | 2.98 | 0.82 | 20.2 | 80.3 | 218 | 537 | 0.85 | 4.28 |
| C18:2 | 10.8 | 1.19 | 11.7 | 33.4 | 80.4 | 191 | 2.72 | 9.10 |
| C18:3 | 0.96 | 0.23 | 3.46 | 13.7 | 38.2 | 91.4 | 10.4 | 2.39 |
| Total FA | 22.4 | 4.60 | 46.2 | 151 | 385 | 930 | 18.6 | 21.0 |
1 RS = rapeseed.
2 INDF = indigestible NDF.
3 OMD = in vitro OM digestibility.
4 Sum of fatty acids includes, beside those shown, C16:1, C20:0, C20:1, C20:2, and C22:0.
Table 2Ration ingredients and chemical composition (g/kg of DM unless otherwise noted).
| Item | Ration | |||
|---|---|---|---|---|
| CON | RSC | WCR | RSO | |
| Barley | 143 | 137 | 137 | 138 |
| Beet pulp dried | 143 | 137 | 137 | 138 |
| Rapeseed meal, 4% fat | 190 | 62 | 149 | 184 |
| Rapeseed cake, 17% fat | 0 | 156 | 0 | 0 |
| Rapeseed, cracked | 0 | 0 | 69 | 0 |
| Rapeseed oil | 0 | 0 | 0 | 33 |
| Corn silage | 238 | 232 | 232 | 232 |
| Grass silage | 286 | 275 | 275 | 275 |
| DM, g/kg of fresh matter | 479 | 500 | 492 | 494 |
| OM | 934 | 937 | 939 | 939 |
| CP | 169 | 171 | 168 | 171 |
| Crude fat | 35 | 55 | 62 | 65 |
| Fatty acids | 26 | 43 | 50 | 53 |
| NDF | 332 | 328 | 326 | 322 |
| Gross energy, MJ/kg of DM | 18.4 | 18.9 | 19.1 | 19.1 |
| NEL, MJ/kg of DM | 7.6 | 7.8 | 8.0 | 8.1 |
1 CON = control, RSC = rapeseed cake, WCR = whole cracked rapeseed, RSO = rapeseed oil.
2 Calculated values based on analysis of ingredients.
Measurements
The milk production and composition were measured once a week during morning and evening milkings. Weekly samples of the feed ingredients were stored (−20°C) and pooled during the whole experiment. Samples of TMR and refusals were taken daily in connection with the afternoon feeding, stored (−20°C), and pooled for each period from d 15 to 20.
Chromic oxide was used as a flow marker, and 10 g was administrated to the rumen via the ruminal cannula during each of the 2 daily feedings, except when the cows were in the respiration chambers.
Samples of duodenal chyme (600 mL), ileal chyme (300 mL), and feces (350 mL) were taken from d 15 to 19 at 1000, 1800 (d 15), 0200, 1200, 2000 (d 16), 0400, 1400, 2200 (d 17), 0600, 1600, 2400 (d 18), and 0800 h (d 19; 12 samples, representing every second hour of the day). Samples from the duodenum and ileum were taken in tube-formed plastic bags which were mounted to the cannulas with plastic knees. Duodenal, ileal, and fecal samples were added to the frozen pooled sample from previous samples at each sampling time. At the end of the period, representative subsamples from thawed material were taken and freeze-dried for chemical analysis. At the 12 sampling times, rumen liquid was sampled from the ventral ruminal sac with a collection tube (#RT, Bar Diamond Inc.). The rumen liquid pH was measured immediately, and two 8-mL samples were taken and frozen (−20°C) immediately for VFA and ammonia (NH3) analysis at each sampling time.
Chemical Analysis
Ash was determined by combustion at 525°C for 6 h. Nitrogen was determined by the Dumas principle (
Hansen, 1989
), and CP was calculated as N × 6.25. Crude fat was analyzed as Soxhlet extraction with petroleum ether (Soxtec 2050, Foss Analytical, Hillerød, Denmark) after hydrolyzing with HCl (Stoldt, 1952
). The NDF content was analyzed by neutral detergent extraction according to Mertens, 2002
with a Fibertec M6 System (Foss Analytical) using heat-stable amylase and corrected for ash. The indigestible NDF in freeze-dried ground (1.5mm) feed samples was determined as residual NDF after 288 h (12 d) of Dacron bag incubation in the rumen of 3 heifers fed a standard ration (Åkerlind et al., 2011
). The GE was determined by adiabatic bomb calorimeter (Parr 6300 Oxygen Bomb Calorimeter, Parr Instrument Company, Moline, IL).The concentrations of VFA were analyzed according to the method described by
Canibe et al., 2007
using a Hewlett Packard gas chromatograph (model 6890; Agilent Technologies Inc., Wilmington, DE) equipped with a flame ionization detector and a 30-m SGE BP1 column (Scientific Instrument Services, NJ). Fatty acids in feed were analyzed by GC after an acidic Bligh and Dyer extraction with hydrochloric acid-water-chloroform and methanol, and subsequent methylation as described by Jensen, 2008
. For determination of NH3, the rumen fluid was made alkaline with KOH, and NH3 was determined by titration after distillation.Chromic oxide was determined by colorimetry after oxidation to chromate (
Schürch et al., 1950
). The OM digestibility was determined in vitro for grass and corn silage as described by Tilley and Terry, 1963
. Milk concentrations of fat, protein, and lactose were analyzed by a Milkoscan Msc4000 infrared analyzer (Foss Analytical).Methane Measurements
During the fourth week of each period, the CH4 production was measured for 2 × 48 h in four 17-m3 open-circuit respiration chambers (
Hellwing et al., 2012
). The animals were housed individually. The chambers were covered with transparent polycarbonate and placed in a square so that the cows faced each other. The chambers were located in the barn where the cows were usually housed to minimize changes in the environment and the daily routines during the CH4 measurements were identical to the period outside the chambers. The mean ambient temperature in the chambers was 21.1°C, ranging from 15.6 to 29.5°C.The cows changed chambers diagonally after the first 48 h to balance out any differences in background levels of CH4 and CO2. Cow and chamber were confounded over periods and, therefore, every ration was tested in every chamber.
The chambers were opened twice daily, at 0500 and 1700 h, for about 20 min during milking and subsequent feeding. Methane was measured as the accumulated amount (L) over 24 h and is reported under standard conditions (0°C, 101.325 kPa). The measurements during the openings of the chambers for milking and feeding were deleted (about 60 min/d). The CH4 production during this period was assumed to correspond to the mean of the rest of the day.
The air flow was measured by a HFM-200 flow meter with a laminar flow element from Teledyne Hastings Instruments (Hampton, VA). The background (inlet air), as well as the chamber outlet air concentration of CH4, was measured every 12 1/2 min with an infrared analyzer. All instruments were from Columbus Instruments (Columbus, OH). The air flow was adjusted individually for every animal depending on the BW and milk yield to obtain a CO2 concentration in the chamber below, but close to 9,000 ppm. The instruments were calibrated every second day with zero gas (nitrogen) and a span gas with nitrogen and 20.55% O2, 5,000 ppm CO2, and 800 ppm CH4 (Yara Praxair AS, Oslo, Norway). The temperature, humidity, and CO2 concentration of the chamber air were monitored with independent sensors (Veng System A/S, Roslev, Denmark) for alarm purposes.
Calculations and Statistical Analysis
The net energy content in feeds was calculated in Scandinavian Feed Units (SFU;
Weisbjerg and Hvelplund, 1993
) and presented in megajoules of NEL by using a fixed conversion factor of 7.89 MJ of NEL/SFU, as described by Hvelplund et al., 2007
. Content of SFU is calculated based on content of ash, CP, crude fiber, crude fat, and in vitro OM digestibility.The apparent rumen digestibility was calculated as the feed intake minus the duodenal flow divided by feed intake. Apparent small intestine digestibility was calculated as the duodenal flow minus ileal flow divided by duodenal flow and, accordingly, apparent large intestine digestibility as ileal flow minus fecal flow divided by ileal flow.. Apparent total-tract digestibility was calculated as the feed intake minus fecal flow divided by feed intake.
Average ECM yield for each cow per period was calculated according to
Sjaunja et al., 1991
as follows: ECM = milk yield × (383 × fat % + 242 × protein % + 783.2)/3,140. For samples with repeated measures (VFA, NH3, and pH), an average for each cow-period was calculated before the statistical analysis.The data was evaluated with the MIXED procedure (SAS 9.2 version, SAS Institute Inc., Cary, NC) with treatment and period as fixed effects and cow as random effect.
The results are reported as LSM and SEM for each treatment. The SEM was different for RSO because one cow receiving RSO was omitted. Therefore, the RSO SEM is reported and a factor for the SEM of the other treatments is noted under each table. Apart from the mean treatment effect, the significance of orthogonal contrasts were calculated for CON versus fat supplement, RSC and WCR versus RSO, and RSC versus WCR. P-values < 0.05 were regarded as significant and P < 0.1 as a tendency.
Results
Milk Production
Average milk production and ECM were not significantly affected by the treatment (Table 3). The milk fat content was numerically lower in fat supplemented rations compared with CON. Milk protein content was numerically higher on RSC and WCR than on RSO (P = 0.71). Daily milk fat production in g/d was 1,187, 1,170, 1,164, and 968 for CON, RSC, WCR, and RSO, respectively. Daily milk protein production was 915, 1,030, 928, and 849 g/d for CON, RSC, WCR, and RSO. The yield of milk solids per day was not affected by the ration.
Table 3Milk production and composition.
| Item | Rations | SEM | P-value | Contrast (P-value) | |||||
|---|---|---|---|---|---|---|---|---|---|
| CON | RSC | WCR | RSO | CON vs. Fat | RSC and WCR vs. RSO | RSC vs. WCR | |||
| Milk, kg/d | 27.4 | 31.2 | 28.1 | 26.3 | 4.61 | 0.23 | 0.52 | 0.16 | 0.18 |
| Milk fat, g/kg | 40.4 | 39.1 | 41.2 | 37.9 | 4.39 | 0.90 | 0.78 | 0.62 | 0.64 |
| Milk protein, g/kg | 33.5 | 33.1 | 32.9 | 32.4 | 1.13 | 0.71 | 0.35 | 0.51 | 0.83 |
| Milk lactose, g/kg | 46.4 | 46.8 | 46.1 | 46.4 | 1.28 | 0.26 | 0.80 | 0.99 | 0.07 |
| ECM, kg/d | 27.2 | 30.2 | 25.1 | 28.4 | 4.37 | 0.12 | 0.61 | 0.04 | 0.29 |
1 CON = control, RSC = rapeseed cake, WCR = whole cracked rapeseed, RSO = rapeseed oil.
2 SEM for RSO; the SEM for the other treatments is the presented SEM × 0.93.
3 Fixed effect of treatment.
4 Linear orthogonal contrasts of treatment.
Feed Intake
The DMI was not affected by the treatments. The intake was numerically greater for cows consuming CON and RSC than WCR and RSO. The higher GE content in the 3 fat-supplemented rations compensated for the numerically lower DMI, and the GE intake was similar between CON and fat-supplemented rations. As planned, fat intake increased in the fat-supplemented rations, and NDF and CP intakes were not affected, as shown in Table 4. Adding rapeseed to the ration increased the intake of total FA from 471 g/d in CON to 804, 835, and 892 g/d for RSC, WCR, and RSO, respectively. Compared with CON, adding fat almost doubled the absolute intake of C18:0 (P = 0.001) and tripled the intake of C18:1 (P < 0.003).
Table 4Intake and apparent digestibility of nutrients.
| Ration | Contrast (P-value) | ||||||||
|---|---|---|---|---|---|---|---|---|---|
| Item | CON | RSC | WCR | RSO | SEM | P-value | CON vs. Fat | RSC and WCR vs. RSO | RSC vs. WCR |
| DM | |||||||||
| Intake, kg/d | 18.3 | 18.9 | 17.9 | 15.8 | 2.07 | 0.54 | 0.62 | 0.22 | 0.58 |
| OM | |||||||||
| Intake, kg/d | 17.1 | 17.7 | 16.8 | 14.9 | 1.96 | 0.56 | 0.64 | 0.23 | 0.61 |
| Duodenal flow, kg/d | 9.84 | 9.81 | 10.3 | 9.19 | 1.30 | 0.85 | 0.94 | 0.48 | 0.67 |
| Rumen digestibility, % | 42.5 | 44.6 | 38.1 | 40.6 | 2.34 | 0.23 | 0.57 | 0.79 | 0.06 |
| Total-tract digestibility, % | 73.2 | 74.3 | 72.0 | 71.9 | 1.36 | 0.41 | 0.71 | 0.41 | 0.16 |
| NDF | |||||||||
| Intake, kg/d | 6.10 | 6.20 | 5.83 | 5.18 | 0.74 | 0.59 | 0.54 | 0.27 | 0.60 |
| Duodenal flow, kg/d | 2.35 | 2.16 | 2.17 | 1.93 | 0.32 | 0.43 | 0.18 | 0.31 | 0.96 |
| Rumen digestibility, % | 61.7 | 65.5 | 62.7 | 63.9 | 2.57 | 0.22 | 0.14 | 0.93 | 0.13 |
| Hindgut digestibility, % | 1.55 | 9.64 | 1.28 | 1.83 | 6.39 | 0.61 | 0.66 | 0.63 | 0.29 |
| Total-tract digestibility, % | 61.7 | 63.3 | 60.4 | 61.1 | 2.37 | 0.60 | 0.98 | 0.72 | 0.23 |
| Crude fat | |||||||||
| Intake, kg/d | 0.65 | 1.03 | 1.11 | 1.00 | 0.10 | 0.01 | 0.003 | 0.48 | 0.38 |
| Duodenal flow, kg/d | 0.83 | 1.18 | 1.35 | 1.27 | 0.12 | 0.01 | 0.004 | 0.98 | 0.15 |
| Rumen digestibility, % | −27.5 | −15.7 | −22.5 | −26.4 | 6.21 | 0.46 | 0.37 | 0.36 | 0.39 |
| Small intestine digestibility, % | 75.0 | 75.0 | 75.0 | 69.9 | 3.18 | 0.33 | 0.44 | 0.10 | 0.99 |
| Total-tract digestibility, % | 65.4 | 67.8 | 65.1 | 58.0 | 4.29 | 0.25 | 0.59 | 0.07 | 0.50 |
| FA intake, g/d | 471 | 804 | 835 | 892 | 89.1 | 0.009 | 0.002 | 0.88 | 0.30 |
| CP | |||||||||
| Intake, kg/d | 3.09 | 3.23 | 3.00 | 2.70 | 0.35 | 0.53 | 0.68 | 0.23 | 0.48 |
| Duodenal flow, kg/d | 3.66 | 3.46 | 3.61 | 3.26 | 0.45 | 0.78 | 0.51 | 0.50 | 0.69 |
| Ruminal digestibility, % | −19.0 | −6.88 | −21.3 | −16.3 | 5.16 | 0.21 | 0.44 | 0.73 | 0.06 |
| Total-tract digestibility, % | 65.1 | 66.9 | 64.5 | 64.5 | 1.63 | 0.52 | 0.85 | 0.52 | 0.22 |
1 CON = control, RSC = rapeseed cake, WCR = whole cracked rapeseed, RSO = rapeseed oil.
2 SEM for RSO; the SEM for the other treatments is the presented SEM × 0.86.
3 Fixed effect of treatment.
4 Linear orthogonal contrasts of treatment.
Digestibility
Duodenal, ileal, and fecal flows were unaffected by the treatment, except for an increased fat flow in the fat-supplemented rations (data not shown). Neither total-tract nor ruminal apparent digestibility was affected by the treatment for any measured nutrient (Table 4). The flow of crude fat to the duodenum was higher than intake by an average of 210 g/d, or 22.7%; no difference was observed between rations in ruminal fat digestibility (P = 0.46). The NDF digestibility in the hind gut was close to zero for all rations except RSC.
VFA, Ammonia, and Rumen pH
Fat supplementation did not affect VFA concentrations in the rumen (P = 0.24). Bound-fat supplements (rapeseed cake, whole rapeseed) resulted in higher propionic acid proportion (P = 0.005), and tended to reduce the acetic acid proportion (P = 0.07) compared with free supplement (oil). Consequently, the acetate to propionate ratio was 2.95 for RSO and 2.76, 2.73, and 2.74 for CON, RSC, and WCR, respectively, resulting in a significant difference (P = 0.01) between the 2 bound-fat supplements and RSO (Table 5).
Table 5Rumen VFA, ammonia, and pH (mean of 12 diurnal samples).
| Item | Ration | SEM | P-value | Contrast (P-value) | |||||
|---|---|---|---|---|---|---|---|---|---|
| CON | RSC | WCR | RSO | CON vs. Fat | RSC and WCR vs. RSO | RSC vs. WCR | |||
| Total VFA, mmol/L | 109 | 105 | 103 | 98.6 | 3.52 | 0.24 | 0.10 | 0.20 | 0.63 |
| VFA, % of total VFA | |||||||||
| Acetic acid (A) | 60.5 | 59.8 | 60.1 | 61.0 | 0.67 | 0.22 | 0.68 | 0.07 | 0.50 |
| Butyric acid | 15.1 | 15.7 | 15.4 | 15.8 | 0.52 | 0.21 | 0.08 | 0.40 | 0.34 |
| Isobutyric acid | 0.74 | 0.78 | 0.72 | 0.84 | 0.06 | 0.32 | 0.39 | 0.17 | 0.29 |
| Propionic acid (P) | 22.1 | 22.1 | 22.1 | 20.8 | 0.93 | 0.02 | 0.10 | 0.005 | 0.91 |
| Valeric acid | 1.64 | 1.67 | 1.70 | 1.62 | 0.06 | 0.44 | 0.51 | 0.22 | 0.58 |
| A:P ratio | 2.76 | 2.73 | 2.74 | 2.95 | 0.15 | 0.05 | 0.34 | 0.01 | 0.89 |
| NH3-N, mg/100 g | 14.8 | 18.8 | 14.5 | 16.1 | 1.92 | 0.19 | 0.33 | 0.78 | 0.06 |
| Average pH | 6.32 | 6.28 | 6.26 | 6.28 | 0.05 | 0.73 | 0.31 | 0.75 | 0.73 |
1 CON = control, RSC = rapeseed cake, WCR = whole cracked rapeseed, RSO = rapeseed oil.
2 SEM for RSO; the SEM for the other treatments is the presented SEM × 0.91.
3 Fixed effect of treatment.
4 Linear orthogonal contrasts of treatment.
The ammonia concentration in the rumen fluid was not influenced by the ration (P = 0.19), but a tendency for a higher concentration in RSC than in WCR (P = 0.06) was noted. The rumen pH was within the normal physiological range: an average of 6.29 for the 16 observations ranging from 5.60 to 6.91.
Methane Production
Adding rapeseed fat to the ration reduced the enteric CH4 production significantly in liters per day and also related to the feed intake and ECM yield (Table 6). The average reduction (L of CH4 per kg of DMI per percent fat added) was 4.6, 4.8, and 3.8% for RSC, WCR, and RSO, respectively. A tendency was observed for a reduction in CH4 per kilogram of OM digested in the whole digestive tract (P = 0.08) when fat was added, but the ration had no effect on CH4 per kilogram of totally digested carbohydrate or NDF and per kilogram of ruminally digested OM and NDF.
Table 6Methane production.
| Ration | Contrasts (P-value) | ||||||||
|---|---|---|---|---|---|---|---|---|---|
| CH4production | CON | RSC | WCR | RSO | SEM | P-value | CON vs. Fat | RSC and WCR vs. Oil | RSC vs. WCR |
| L/d | 569 | 531 | 478 | 462 | 51.0 | 0.04 | 0.02 | 0.18 | 0.10 |
| L/kg of ECM | 20.4 | 19.0 | 16.9 | 16.7 | 2.03 | 0.008 | 0.003 | 0.11 | 0.02 |
| L/kg of DMI | 29.6 | 26.9 | 25.8 | 26.4 | 1.78 | 0.07 | 0.02 | 0.99 | 0.37 |
| % of GE intake | 6.32 | 5.60 | 5.31 | 5.40 | 0.03 | 0.002 | <0.001 | 0.04 | 0.02 |
| L/kg of total digested OM | 46.1 | 40.3 | 39.8 | 43.0 | 3.10 | 0.22 | 0.08 | 0.36 | 0.87 |
| L/kg of total digested NDF | 154 | 135 | 137 | 146 | 10.3 | 0.20 | 0.18 | 0.39 | 0.85 |
| L/kg of total digested CHO | 57.2 | 51.4 | 51.1 | 54.9 | 3.68 | 0.33 | 0.16 | 0.34 | 0.92 |
| L/kg of rumen digested OM | 79.8 | 69.9 | 79.1 | 79.3 | 7.49 | 0.54 | 0.57 | 0.55 | 0.26 |
| L/kg of rumen digested NDF | 154 | 132 | 132 | 140 | 12.3 | 0.45 | 0.16 | 0.60 | 0.99 |
1 CON = control, RSC = rapeseed cake, WCR = whole cracked rapeseed, RSO = rapeseed oil.
2 SEM for RSO; the SEM for the other treatments is the presented SEM × 0.88.
3 Fixed effect of treatment.
4 Linear orthogonal contrasts of treatment.
5 DMI during CH4 measurements.
6 GE = gross energy.
7 CHO = carbohydrate calculated as OM minus fat minus CP.
Discussion
Intake and Milk Production
The rations were calculated to have identical crude fat content. However, the fat content in the rapeseed cake was not as high as planned, which resulted in a lower fat content in RSC.
We hypothesized that fat supplementation at the present levels would only reduce CH4 production with out affecting feed energy intake and milk yield. Negative effects of fat supplementation on DMI have been reported in some (
Harvatine and Allen, 2006
; Martin et al., 2008
) but not all previous studies (Johnson et al., 2002
; Moate et al., 2011
). The effect of fat on DMI and GE intake depends on the level of supplementation. When cows are fed a low level of supplementation, DMI remains unaffected and GE intake increases, due to a higher energy density. When a moderate amount of fat is supplemented, DMI is reduced but GE intake is unchanged, whereas a further increase of the fat concentration in the ration can reduce the DMI to an extent where the GE intake is also reduced (Grainger and Beauchemin, 2011
). Neither DMI nor GE intake was affected significantly in the present study, indicating that the fat concentration in the rations (5.5–6.5% of DM) was within the nutritionally acceptable range. The rapeseed oil supplementation numerically reduced the DMI by 2.2 kg compared with CON, but the cows responded differently to the addition of oil, which is illustrated by the higher standard deviation between cows for DMI on RSO (5.3 kg) compared with the other rations (3.1 kg). This was also supported by heterogeneous variance for DMI, as a likelihood ratio test showed. Dry matter intake depression can be expected when the dietary fat concentration exceeds 6 to 7% (Beauchemin et al., 2007
). A fat concentration of 6.5% in RSO in the present study may therefore be at a borderline where it affects some, but not necessarily all, animals. This was illustrated by the pronounced depressing effect of oil supplementation on DMI for the 2 cows with the highest feed intake level (per kg of BW) compared with the 2 cows with the lowest feed intake level (values not shown).NDF Digestibility
Ruminal NDF digestibility was 63% on average, which is in accordance with previous studies with similar ration compositions (
Lund et al., 2007
). Rapeseed fat includes mainly monounsaturated FA and was presented at moderate concentrations in the present study; therefore, it did not affect NDF digestibility. The same effect has been observed with comparable additions of rapeseed (Chelikani et al., 2004
; Beauchemin et al., 2009
).Few studies have compared how different physical forms of the same fat affect digestion. Even though effects of the physical form and processing of whole seeds were reported for linseed (
Martin et al., 2008
), this might not be valid for rapeseed. Pallister and Smithard, 1987
found no effect of whole or extruded rapeseed or rapeseed oil on fiber digestion, using a comparable fat content in the control ration and slightly higher levels in the fat-supplemented rations compared with the present experiment. Ferlay et al., 1992
hypothesized that extrusion ruptures cell membranes and increases the availability of fat. However, in an experiment with feeding raw and extruded rapeseed to dairy cows, no effect on digestion was noted (Ferlay et al., 1992
). This might be due to the high fat content in rapeseed compared with other oilseeds (Ferlay et al., 1992
).Total-tract NDF digestibility was lower than the ruminal NDF digestibility for all fat-supplemented rations. That problem has been observed earlier in connection with flow markers (
Faichney, 1972
; Lund et al., 2007
) and different types of duodenal cannulas (Stensig and Robinson, 1997
). The hindgut digestibility of NDF was higher for RSC compared with the other rations, but still within the physiological range (Huhtanen et al., 2006
).Fat Digestibility
The fat digestibility in the rumen was negative for all rations.
Schmidely et al., 2008
reported, in a meta-analysis, a slightly positive FA balance from feed to duodenum (duodenal flow – intake). However, they found a net disappearance of FA with higher intake (50–120 g of FA/kg of DMI). In the present study, RSO only slightly exceeded the 50 g of FA/kg of DM, and the other rations had a lower FA content. Scollan et al., 2001
reported a positive net FA flow to the duodenum with an increase in rumen FA balance by 21.8% when steers receive rations with 60 g/kg of DM fat, which fits well with the present experiment where crude fat increased by 22.7% from feed to duodenum. As observed earlier (Doreau and Chilliard, 1997
), a net FA synthesis takes place in the rumen at low to moderate fat contents.The digestibility of fat in the small intestine is highly variable;
Doreau and Chilliard, 1997
reported variations from 55 to 92% in a literature survey. The digestibility of fat in the small intestine depends on the chain length and degree of unsaturation (Weisbjerg et al., 1992
; Doreau and Chilliard, 1997
). The present digestibility for fat with mainly C18 FA was between 70 and 75%, which is in agreement with earlier reported values (Schmidely et al., 2008
). With increasing fat consumption, the intestinal digestibility decreases because the absorptive capacity is limited, but absorption of FA can be higher than 1 kg/d (Doreau and Chilliard, 1997
). As the highest FA intake in the present experiment was 892 g/d, fat absorption was not affected by treatment.VFA and NH3
Molar concentration of VFA was in agreement with earlier studies with dairy cows receiving rapeseed fat supplementation (
Chelikani et al., 2004
; Beauchemin et al., 2009
). The higher acetate to propionate ratio for RSO compared with CON is surprising, because fat addition is believed to inhibit NDF but not starch fermentation, and therefore the propionic acid proportion was expected to increase as observed by Harvatine and Allen, 2006
. According to fermentation stoichiometry, a shift in fermentation pattern toward acetate, as observed for RSO, should result in enhanced CH4 production (Boadi et al., 2004
). However, cows fed RSO produced less CH4 than on CON. The higher propionate proportion could be due to reduction in total VFA production because less substrate was fermented in the rumen, or because differences were buffered, as the values shown are averages of day and night time measurements.The ammonia concentration in the rumen is considered to be the balance between entry sources (degradable feed N, N recycling) and outputs (incorporation into microbes, N absorption, ammonia N outflow;
Doreau and Ferlay, 1995
). In agreement with previous studies (Oldick and Firkins, 2000
), fat addition itself did not affect the protein digestion in the rumen.Methane Production
The CH4 production in absolute numbers (L/d), as well as in relation to intake, was comparable to previous studies (
Beauchemin et al., 2008
; Moate et al., 2011
). The reduction in CH4 loss expressed as the proportion of GE intake when rapeseed fat was supplemented was less than found by Beauchemin et al., 2009
with dairy cows (18 vs. 14% in the present study) or Machmüller et al., 2000
with lambs (22%). This was most likely due to a higher fat concentration (Beauchemin et al., 2009
) and a higher forage proportion (Machmüller et al., 2000
) compared with rations in the present study. The reduction in CH4 per kg of DMI compared with CON per percentage fat added was highest for WCR (4.8%) followed by 4.6 for RSC and 3.8% for RSO. The present reductions per kg of DMI were not as high as the 5.6% reduction found in a review by Beauchemin et al., 2008
, but close to the values presented in the review by Grainger and Beauchemin, 2011
; between 4.7 and 5.1% depending on fat level in the ration).Earlier studies with rapeseed found contradictory results;
Martin et al., 2011
added 3% fat as extruded rapeseed to a dairy cow ration without finding any effect on CH4. Beauchemin and McGinn, 2006
found a significant reduction in CH4 loss as proportion of GE when feeding a ration with 4.6% rapeseed oil added to heifers, but no effect per kg of DMI. In studies using dairy cows, Moate et al., 2011
found a significant reduction in CH4 per kg of DMI when adding 2.6% fat to the ration as a rapeseed-hominy meal mix, and Beauchemin et al., 2009
found a reduction for 1.7% fat added as crushed rapeseed compared with the control ration.Beauchemin and McGinn, 2006
observed a feed intake depression and therefore, no effect on CH4 per kg of DMI was observed when heifers received rapeseed oil. In a later study, Beauchemin et al., 2009
compared the CH4-reducing properties of different oil seeds and emphasized that rapeseed did not depress DMI in contrast to other oil seeds. The present results and those of Moate et al., 2011
confirm that rapeseed fat does not always depress feed intake. The depressed feed intake in heifers found by Beauchemin and McGinn, 2006
was probably due to the high fat supplementation level.We hypothesized that fat from seeds is released more slowly in the rumen, and therefore might affect both CH4 production and digestion differently than oil.
Martin et al., 2008
found a stronger effect on CH4 production the higher the fat content in the supplement (i.e., extruded linseed being less effective than whole linseed and whole linseed less effective than linseed oil). Conversely, Beauchemin et al., 2007
compared sunflower seeds and sunflower oil and found, in agreement with the present study, no difference in the CH4-depressing properties for different physical forms of the same fat source. The linseed fat in the study by Martin et al., 2008
was supplemented at a higher level, and linseed FA have a higher degree of unsaturation than rapeseed or sunflower. Furthermore, the different physical forms in the study by Martin et al., 2008
were confounded with fat contents, as the oil ration included 1.4 and 1.6% per kg of DM more fat than extruded and crude seed rations, respectively.The fat concentration in rapeseed cake was lower than planned, which resulted in a lower fat content in RSC than in WCR and RSO. A meta-analysis by
Beauchemin et al., 2008
showed a linear relationship between the percentage of fat added and the reduction in CH4. A rapeseed cake probably would have been more effective in reducing CH4 if it had had a higher fat concentration.Fat supplementation depresses CH4 production by lowering the quantity of OM degraded in the rumen, by influencing the microbial activity and ecosystem, and, to a very minor extent, by biohydrogenation of unsaturated FA (
Johnson and Johnson, 1995
). Although rumen digestibilities and VFA composition were not altered, the numeric reduction in CH4 per kg of digested carbohydrate indicates that fermentation pathways were affected by fat supplementation in addition to the effect of fat being nonfermentable.An optimal rumen microbial ecosystem is a prerequisite for efficient milk production, and it is well known that methanogens and fibrolytic bacteria may be hampered by addition of fat. The methanogen population of the rumen was studied in parallel to the present study by
Poulsen, 2012
; despite the fact that fat supplementation reduced the CH4 production, the total abundance of the methanogens in the rumen was unaffected by the ration.Conclusions
Supplementation with rapeseed fat up to 6 to 6.5% fat of ration of DM reduced GE loss of enteric CH4 by 14% without compromising the NDF digestibility or milk production. Adding fat by supplementing rapeseed in different forms decreased CH4 production and numerically increased milk and ECM yields in the present study, resulting in less CH4 per kg of product.
Acknowledgements
The work was funded by the Danish Ministry of Food, Agriculture and Fisheries, Mælkeafgiftsfonden (Aarhus, Denmark), and Aarhus University. The authors thank Torkild Jakobsen (Aarhus University, Foulum, Denmark) for skillful assistance during the experiment and Ingolf Nielsen (DLG Food Oil, Dronninglund, Denmark) for providing the rapeseed feeds.
Article info
Publication history
Published online: February 18, 2013
Accepted:
December 28,
2012
Received:
December 7,
2011
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© 2013 American Dairy Science Association. Published by Elsevier Inc.
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