Related Experiment Video
Updated: Jun 6, 2026

Analysis of Interactions between Endobiotics and Human Gut Microbiota Using In Vitro Bath Fermentation Systems
Published on: August 23, 2019
Isotopic H2 and NH4+ supplementation enhances volatile fatty acid and microbial protein synthesis during in vitro
Yurong Cao1,2, Xiumin Zhang3,4, Siyu Yi1,5
1State Key Laboratory of Forage Breeding-by-Design and Utilization, National Engineering Laboratory for Pollution Control and Waste Utilization in Livestock and Poultry Production, Institute of Subtropical Agriculture, Chinese Academy of Sciences, Changsha, Hunan, 410125, China.
Background:
The volatile fatty acid (VFA) and microbial protein production are tightly linked to hydrogen (H2) metabolism in rumen. The H2 can be split by hydrogenases into protons and electrons, and is regarded as the most primordial source of reducing equivalents within the rumen microecosystem. However, the quantitative role of H2 in VFA and microbial protein formation has not been thoroughly studied. Additionally, while rumen fermentation is influenced by ammonia-N availability, the combined effects of H2 and NH4+ supplementation on VFA and microbial protein synthesis have not been fully explored. The objective of this study was to explore the role of proton and electron transfer from H2 in VFA and microbial protein synthesis based on the synergistic effect of H2 and NH4+ supplementation. In vitro ruminal batch culture was employed for the complete block design with factorial arrangement of four treatments including H2 (0 or 40 mL 2H2) and NH4+ (0 or 8 mmol/L 15NH4+) supplementations.
Results:
2H2 supplementation increased dry matter (DM) degradation (P = 0.005), methane (CH4) production (P < 0.001), 2H abundance (P < 0.001) in microbial protein and individual VFAs. However, 2H abundance in microbial protein, acetate, propionate and butyrate are low with the value less than 2.0‰. The 15NH4+ supplementation increased microbial protein production (P = 0.001), microbial 15N abundance (P < 0.001), and microbial 15N incorporation (mg/g degraded DM, P < 0.001), and concurrently reduced CH4 production (P < 0.001). Moreover, 2H2 supplementation further enhanced the stimulatory effect of 15NH4+ on microbial protein production.
Conclusion:
These results indicate limited incorporation of protons for the synthesis of VFA and microbial protein, and the enhancement of VFA and microbial protein production may be associated with increased electron availability caused by the 2H2 supplementation. The isotopic dilution and hydrogen exchange with other pools cannot be ruled out as contributing factors, which should be explored in the future studies.
More Related Videos
Related Concept Videos
Microbial Interactions: Mutualism
¹H NMR of Labile Protons: Deuterium (²H) Substitution
Inorganic Nitrogen Assimilation
Microbes and Methanogenesis

