The climate-relevant enteric methane (CH₄) formation represents a loss of feed energy that is potentially meaningful for energetically undersupplied peripartal dairy cows. Higher concentrate feed proportions (CFP) are known to reduce CH₄ emissions in cows. The same applies to the feed additive 3-nitrooxypropanol (3-NOP), albeit through different mechanisms. It was hypothesised that the hydrogen not utilised for CH₄ formation through the inhibition by 3-NOP would be sequestered by propionate formation triggered by higher CFP so that it could thereby give rise to a synergistically reduced CH₄ emission. In a 2 × 2-factorial design, low (LC) or high (HC) CFP were either tested without supplements (CON_LC, CON_HC) or combined with 3-NOP (NOP_LC, 48.4 mg/kg dry matter (DM); NOP_HC, 51.2 mg 3-NOP/kg DM). These four rations were fed to a total of 55 Holstein cows from d 28 ante partum until d 120 post partum. DM intake (DMI) was not affected by 3-NOP but increased with CFP (CFP; p < 0.001). CH₄/DMI and CH₄/energy-corrected milk (ECM) were mitigated by 3-NOP (23% NOP_LC, 33% NOP_HC) (p < 0.001) and high CFP (12% CON, 22% 3-NOP groups) (CFP × TIME p < 0.001). Under the conditions of the present experiment, the CH₄ emissions of NOP_LC increased to the level of the CON groups from week 8 until the end of trial (3-NOP × CFP × TIME; p < 0.01). CO₂ yield decreased by 3-NOP and high CFP (3-NOP × CFP; p < 0.001). The reduced body weight loss and feed efficiency in HC groups paralleled a more positive energy balance being most obvious in NOP_HC (3-NOP × CFP; p < 0.001). ECM was lower for NOP_HC compared to CON_HC (3-NOP × CFP; p < 0.05), whereas LC groups did not differ. A decreased fat to protein ratio was observed in HC groups and, until week 6 post partum, in NOP_LC. Milk lactose and urea increased by 3-NOP (3-NOP; p < 0.05). 3-NOP and high CFP changed rumen fermentation to a more propionic-metabolic profile (3-NOP; CFP; p < 0.01) but did not affect rumen pH. In conclusion, CH₄ emission was synergistically reduced when high CFP was combined with 3-NOP while the CH₄ mitigating 3-NOP effect decreased with progressing time when the supplement was added to the high-forage ration. The nature of these interactions needs to be clarified.

Methane (CH₄) from ruminal feed degradation is a major pollutant from ruminant livestock, which calls for mitigation strategies. The purpose of the present 4 × 2 factorial arrangement was to investigate the dose–response relationships between four doses of the CH₄ inhibitor 3-nitrooxypropanol (3-NOP) and potential synergistic effects with low (LC) or high (HC) concentrate feed proportions (CFP) on CH₄ reduction as both mitigation approaches differ in their mode of action (direct 3-NOP vs. indirect CFP effects). Diet substrates and 3-NOP were incubated in a rumen simulation technique to measure the concentration and production of volatile fatty acids (VFA), fermentation gases as well as substrate disappearance. Negative side effects on fermentation regarding total VFA and gas production as well as nutrient degradability were observed for neither CFP nor 3-NOP. CH₄ production decreased from 10% up to 97% in a dose-dependent manner with increasing 3-NOP inclusion rate (dose: p < 0.001) but irrespective of CFP (CFP × dose: p = 0.094). Hydrogen gas accumulated correspondingly with increased 3-NOP dose (dose: p < 0.001). In vitro pH (p = 0.019) and redox potential (p = 0.066) varied by CFP, whereas the latter fluctuated with 3-NOP dose (p = 0.01). Acetate and iso-butyrate (mol %) decreased with 3-NOP dose, whereas iso-valerate increased (dose: p < 0.001). Propionate and valerate varied inconsistently due to 3-NOP supplementation. The feed additive 3-NOP was proven to be a dose-dependent yet effective CH₄ inhibitor under conditions in vitro. The observed lack of additivity of increased CFP on the CH₄ inhibition potential of 3-NOP needs to be verified in future research testing further diet types both in vitro and in vivo.

Methanogenesis in ruminants contributes to both greenhouse gas emissions and feed energy losses whereby the latter becomes specifically important in energy-deficient periparturient cows. It was hypothesized that increased concentrate feed proportions (CFP) and feeding with the methane inhibitor 3-nitrooxypropanol (3-NOP), as well as their potential synergism, improve the energy status of peripartal cows. Periparturient dairy cows were fed low or high dietary CFP either tested without or combined with 3-NOP. The GreenFeed system was used to calculate the metabolic respiration quotient (RQ_metabolic) and tissue energy retention (ER_tissue) by methods of indirect calorimetry. The calorimetrically estimated ER_tissue coincided with a conventionally calculated energy balance except for the antepartal period. Neither CFP nor 3-NOP affected the ultrasonographically assessed lipomobilization in adipose depots. In the group fed 3-NOP and a high concentrate feed proportion, the RQ_metabolic significantly rose over the course of the experiment and the ER_tissue was also increased. Serum non-esterified fatty acid concentrations were lower in the 3-NOP groups albeit ß-hydroxybutyrate (BHB) remained unaffected. Higher CFP reduced BHB and increased blood glucose levels. In conclusion, 3-NOP and high CFP improved the energy budget of the cows in an interactive manner, which was, however, not apparent in all of the examined parameters. The application of the GreenFeed system for indirect calorimetry is a promising approach, which needs further validation in the future.