Betaine, a common methyl donor whose methylation is involved in the biosynthesis of carnitine and phospholipids in animals, serves as food and animal feed additive. The present study used liquid chromatography-mass spectrometry (LC-MS) to analyze the liver protein profile of mice on a high fat (HF) diet to investigate the mechanism by which betaine affects hepatic metabolism. Although betaine supplementation had no significant effect on body weight, a total of 103 differentially expressed proteins were identified between HF diet + 1% betaine group (HFB) and HF diet group by LC-MS (fold change > 2, p < 0.05). The addition of 1% betaine had a significant enhancement of the expression of enzymes related to fatty acid oxidation metabolism, such as hydroxyacyl-Coenzyme A dehydrogenase (HADHA), enoyl Coenzyme A hydratase 1 (ECHS1) (p < 0.05) etc., and the expression of apolipoprotein A-II (APOA2) protein was significantly reduced (p < 0.01). Meanwhile, the protein expression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and succinate-CoA ligase (SUCLG1) were highly significant (p < 0.01). Pathway enrichment using the Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed that the functions of differential proteins involved fatty acid catabolism, carbohydrate metabolism, tricarboxylic acid cycle (TCA) and peroxisome proliferator-activated receptor alpha (PPARα) signaling pathway. Protein–protein interaction (PPI) analysis discovered that acetyl-Coenzyme A acetyltransferase 1 (ACAT1), HADHA and ECHS1 were central hubs of hepatic proteomic changes in the HFB group of mice. Betaine alleviates hepatic lipid accumulation by enhancing fatty acid oxidation and accelerating the TCA cycle and glycolytic process in the liver of mice on an HF diet.

Betaine is more efficient than choline and methionine methyl donors, as it can increase nitrogen storage, promote fat mobilisation and fatty acid oxidation and change body fat content and distribution. Lipid is absorbed primarily in the small intestine after consumption, which is also the basis of lipid metabolism. This study was conducted to establish a mouse model of obesity in Kunming mice of the same age and similar body weight, and to assess the effect of betaine on the intestinal protein expression profile of mice using a proteomic approach. Analysis showed that betaine supplementation reversed the reduction in expression of proteins related to lipid metabolism and transport in the intestine of mice induced by a high-fat diet (HFD). For example, the addition of betaine resulted in a significant upregulation of microsomal triglyceride transfer protein (Mttp), apolipoprotein A-IV (Apoa4), fatty-acid-binding protein 1 (Fabp1) and fatty-acid-binding protein 2 (Fabp2) expression compared to the HFD group (p < 0.05), which exhibited accelerated lipid absorption and then translocation from the intestine into the body’s circulation, in addition to a significant increase in Acetyl-CoA acyltransferase (Acaa1a) protein expression, hastening lipid metabolism in the intestine (p < 0.05). Simultaneously, a significant reduction in protein expression of alpha-enolase 1 (Eno1) as the key enzyme for gluconeogenesis in mice in the betaine-supplemented group resulted in a reduction in lipid synthesis in the intestine (p < 0.05). These findings provide useful information for understanding the changes in the protein profile of the small intestine in response to betaine supplementation and the potential physiological regulation of diets’ nutrient absorption.