In this study， to illustrate the specific regulation targets and pathways of glycerol monolaurate (GML) on lipid metabolism， GML was used to treat hyperlipidemia model of liver tumor cells HepG2 induced by oleic acid， and explore the regulation effects of GML on lipid metabolism of liver cells and its potential targets and mechanisms based on bioinformatics prediction and transcriptome sequencing analysis. Bioinformatics analysis showed that the GO and KEGG enrichment analysis of potential GML targets obtained 519 and 144 signal pathways (P<0.05)， respectively. Molecular docking supported that GML might regulate lipid metabolism through regulating ALB， AKT1， EGFR and other core target proteins. The results of cell experiment showed that the level of lipid accumulation in the cell group treated with GML was improved compared with the high-fat control group. A total of 2 879 differentially expressed genes were screened by transcriptome analysis， with 1 363 up-regulated and 1 516 down-regulated genes， respectively. According to the enrichment analysis of KEGG signal pathway， HIF-1α pathway and fatty acid oxidative catabolism pathway were significantly enriched. The gene transcription expression levels of important proteins CPT1B， FABP1 in the pathway were up-regulated to 1.79 times and 2.81 times in the GML treatment group compared with those in the high-fat group， respectively， and the expression levels of main downstream regulation genes of HIF-1α were significantly increased (P＜0.05)， indicating that GML intervention improved the oxidative stress of high-fat cells through regulation of HIF-1α pathway， and improved the lipid metabolism disorder in high-fat cells mediated by fatty acid degradation pathway. This study provides a theoretical reference for the research on the regulation and mechanisms of functional lipid components in improving lipid metabolism disorder induced by high-fat diet.