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In Vitro Evaluation of Enzymatically Transformed Alfalfa Saponins on Methane Reduction, Rumen Microbes and Metabolomics in Goats
by
, ,
Ran Zhang
,
Xinran Bao
,
Xingqi Shi
,
Shixuan Jin
,
Ying Meng
,
Zhiwei Li
,
Zhumei Du
and
Xuebing Yan
* College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China
*
Author to whom correspondence should be addressed.
Animals 2025, 15(11), 1516; https://doi.org/10.3390/ani15111516
Submission received: 10 April 2025
/
Revised: 17 May 2025
/
Accepted: 20 May 2025
/
Published: 22 May 2025
(This article belongs to the Section Small Ruminants)
Simple Summary
Alfalfa, a common livestock feed, contains natural compounds known as saponins, which may help reduce methane emissions from goats, a major contributor to climate change. This study investigated the use of a food-safe enzyme derived from Aspergillus niger to enhance the activity of alfalfa saponins. Saponins from 21 alfalfa cultivars were tested, showing stronger antioxidant activity after silage fermentation. Adding a small amount (0.10 mg/mL) of saponins to goat rumen fluid during in vitro fermentation reduced methane production and altered the rumen microbes linked to methane release. When the enzyme-treated saponins were used, methane levels dropped further, while beneficial microbes and nutrients linked to animal growth increased. The enzymatically enhanced saponins also influenced key metabolic pathways in the goats’ digestive processes, suggesting a dual benefit: lowering greenhouse gas emissions and improving feed efficiency. This eco-friendly approach demonstrates how enzymatically transformed alfalfa saponins could make livestock farming more sustainable by mitigating methane emissions.
Abstract
Alfalfa (Medicago sativa L.) saponins (AS), primarily pentacyclic triterpenoids, may reduce methane emissions from goats (Capra hircus L.). This study evaluated the methane-suppressing potential of Aspergillus niger β-glucosidase-modified AS using in vitro rumen fermentation (0.10 mg/mL inoculum, 24 h incubation, gas chromatography detection). Among the 21 alfalfa cultivars, Pegasis (fall dormancy 9) exhibited the highest antioxidant efficacy (half maximum effective concentration 2.13 mg/mL) and the lowest ferric-reducing activity (0.32 μM Fe2+/g) (p < 0.05). Fresh/silage AS reduced methane proportions to 4.50–5.21% of total gas, while enzymatic biotransformation further decreased it to 3.34–3.48% (p < 0.05). Methanogen abundance declined by 20.10–44.93%, and general anaerobic fungi declined by 34.22–44.66% compared to untreated AS (p < 0.05). Metabolomics linked methane suppression to six pathways, including zeatin biosynthesis (via nucleotide metabolites accumulation) and prolactin signaling pathway (via bioactive molecules downregulation), suggesting impaired methanogen energy metabolism and hydrogen flux redirection as mechanisms. Enzymatic AS also enhanced volatile fatty acid production, indicating improved fiber digestion. These in vitro findings demonstrate that enzyme-treated AS modulates rumen fermentation through dual methane mitigation and nutrient utilization enhancement, offering a sustainable feed additive strategy for livestock.
