Aspirin eugenol ester (AEE) is a novel medicinal compound synthesized by esterifying aspirin with eugenol using the pro-drug principle. Pharmacological and pharmacodynamic experiments showed that AEE had excellent thromboprophylaxis and inhibition of platelet aggregation. This study aimed to investigate the effect of AEE on the liver of thrombosed rats to reveal its mechanism of thromboprophylaxis. Therefore, a multi-omics approach was used to analyze the liver. Transcriptome results showed 132 differentially expressed genes (DEGs) in the AEE group compared to the model group. Proteome results showed that 159 differentially expressed proteins (DEPs) were identified in the AEE group compared to the model group. Six proteins including fibrinogen alpha chain (Fga), fibrinogen gamma chain (Fgg), fibrinogen beta chain (Fgb), orosomucoid 1 (Orm1), hemopexin (Hpx), and kininogen-2 (Kng2) were selected for parallel reaction monitoring (PRM) analysis. The results showed that the expression of all six proteins was upregulated in the model group compared with the control group. In turn, AEE reversed the upregulation trend of these proteins to some degree. Metabolome results showed that 17 metabolites were upregulated and 38 were downregulated in the model group compared to the control group. AEE could reverse the expression of these metabolites to some degree and make them back to normal levels. The metabolites were mainly involved in metabolic pathways, including linoleic acid metabolism, arachidonic acid metabolism, and the tricarboxylic acid (TCA) cycle. Comprehensive analyses showed that AEE could prevent thrombosis by inhibiting platelet activation, decreasing inflammation, and regulating amino acid and energy metabolism. In conclusion, AEE can have a positive effect on thrombosis-related diseases.

Platelet activation is closely related to thrombosis. Aspirin eugenol ester (AEE) is a novel medicinal compound synthesized by esterifying aspirin with eugenol using the pro-drug principle. Pharmacological and pharmacodynamic experiments showed that AEE has excellent anti-inflammatory, antioxidant, and inhibitory platelet activation effects, preventing thrombosis. However, the regulatory network and action target of AEE in inhibiting platelet activation remain unknown. This study aimed to investigate the effects of AEE on platelets of thrombosed rats to reveal its regulatory mechanism via a multi-omics approach. The platelet proteomic results showed that 348 DEPs were identified in the AEE group compared with the model group, of which 87 were up- and 261 down-regulated. The pathways in this result were different from previous results, including mTOR signaling and ADP signaling at P2Y purinoceptor 12. The metabolomics of heart and abdominal aortic tissue results showed that the differential metabolites were mainly involved in steroid biosynthesis, the citric acid cycle, phenylalanine metabolism, phenylalanine, tyrosine, and tryptophan biosynthesis, and glutathione metabolism. Molecular docking results showed that AEE had a better binding force to both the COX-1 and P2Y12 protein. AEE could effectively inhibit platelet activation by inhibiting COX-1 protein and P2Y12 protein activity, thereby inhibiting platelet aggregation. Therefore, AEE can have a positive effect on inhibiting platelet activation.

Rifaximin and rifampicin are good broad-spectrum antimicrobials. The irrational use of antimicrobial drugs in veterinary clinics could threaten public health and food safety. It is necessary to develop a reliable detection method of the residue for enhancing the rational supervision of the use of such drugs, reducing and slowing down the generation of bacterial resistance, and promoting animal food safety and human health. So, this study developed an LC-MS/MS method for the detection of rifaximin and rifampicin residues in animal-origin foods. The residual rifaximin and rifampicin of homogenized test materials were extracted with acetonitrile-dichloromethane solution or acetonitrile in the presence of anhydrous sodium sulfate and vitamin C, purified by dispersible solid phase extraction, determined by LC-MS/MS, and quantified by the internal standard method. The specificity, sensitivity, matrix effect, accuracy, and precision of the method were investigated in the edible tissues of cattle, swine, or chicken. In addition, the stability of the standard stock solution and the standard working solution was also investigated. The method was suitable for the muscle, liver, kidney, fat, milk, and eggs of cattle, swine, or chicken, as well as fish and shrimp. The specificity of the method was good, and the detection of the analytes was not affected by different matrices. Both the LOD and LOQ of the two analytes were 5 μg/kg and 10 μg/kg, respectively. The results of matrix effects in each tissue were in the range of 80–120%; there were no significant matrix effects. The average accuracy of rifaximin and rifampicin in different foodstuffs of animal origin was between 80% and 120%, and the method precision was below 20% (RSD). The proposed method showed good performance for determination, which could be employed for the extraction, purification, and detection of residual rifaximin and rifampicin in edible animal tissues. The pretreatment procedure of tissue samples was simple and feasible. The method was highly specific, stable, reliable, and with high sensitivity, accuracy, and precision, which met the requirements of quantitative detection of veterinary drug residues.

Gastric ulcer (GU) is a common upper gastrointestinal disorder characterized by oxidative stress and inflammatory responses, which significantly impact the patient’s quality of life and pose a serious challenge to public health. Excessive and chronic alcohol consumption are considered primary contributing factors to gastric ulcer. Pharmacodynamic and pharmacological experiments showed that aspirin eugenol ester (AEE) had good anti-inflammatory and anti-oxidant effects. Therefore, it was speculated that AEE could alleviate ethanol-induced gastric mucosal injury through anti-inflammatory and antioxidant pathways. This study aimed to systematically evaluate the effect of AEE on ethanol-induced gastric mucosal injury using in vivo and in vitro experiments. In a gastric injury model induced by ethanol, H&E staining, AB-PAS staining, RT-PCR, immunohistochemistry, and a series of other molecular biological assays and omics techniques were employed to investigate AEE potential mechanisms. The results revealed extensive necrosis in the gastric mucosa of the ethanol group with a marked reduction in mucus secretion on the mucosal surface and a significantly decreased expression of ZO-1, claudin-1, and occludin, while AEE exhibited a significant protective effect when compared to ethanol group. AEE inhibited NF-κB pathway activation and reduced the expression of inflammatory cytokines. AEE significantly enhanced superoxide Dismutase (SOD) levels and reduced the ethanol-induced increases in reactive oxygen species (ROS), malondialdehyde (MDA), and lipid peroxidation (LPO) levels by reversing the ethanol-induced decline in Nrf-2 expression. AEE also mitigated cellular damage by inhibiting ferroptosis in the cells. AEE significantly improved the metabolic profiles of gastric tissue and serum. AEE exerts gastric protective effects by synergistically modulating multiple pathways and biological processes. AEE can enhance antioxidant capacity and inhibit ferroptosis by activating the Nrf-2/GPX4 pathway, alleviate inflammatory responses by suppressing the NF-κB pathway, and simultaneously maintain gastric mucosal barrier integrity through regulation of metabolic reprogramming and enhancement of tight junction function.

Non-alcoholic fatty liver disease (NAFLD) is a highly prevalent condition worldwide and represents a major global health challenge. Pharmacological and pharmacodynamic results indicate that aspirin eugenol ester (AEE) performs various pharmacological activities. However, it is unclear whether AEE can ameliorate the NAFLD. This study investigated the ameliorative effects of AEE on glucose and lipid metabolism disorders by in vitro and in vivo experiments. In the cellular model, TC increased to 0.104 μmol/mg and TG increased to 0.152 μmol/mg in the model group, while TC decreased to 0.043 μmol/mg and TG decreased to 0.058 μmol/mg in the AEE group. In the model group, the area occupied by lipid droplets within the visual field was significantly elevated to 17.338%. However, the administration of AEE resulted in a substantial reduction in this area to 10.064%. AEE significantly reduced the lipid droplet area and TC and TG levels (p < 0.05), increased bile acids in the cells and in the medium supernatant (p < 0.05), and significantly up-regulated the expression of LRH-1, PPARα, CYP7A1, and BSEP mRNA levels (p < 0.05) compared to the model group. In the animal model, different doses of AEE administration significantly down-regulated the levels of TC, TG, LDL, GSP, and FBG (p < 0.05) compared to the high-fat-diet (HFD) group, and 216 mg/kg of AEE significantly improved hepatocellular steatosis, attenuated liver injury, and reduced the area of glycogen staining (p < 0.05). In the HFD group, the glycogen area within the visual field exhibited a significant increase to 18.250%. However, the administration of AEE resulted in a notable reduction in the glycogen area to 13.314%. Liver and serum metabolomics results show that AEE can reverse the metabolite changes caused by a HFD. The major metabolites were involved in seven pathways, including riboflavin metabolism, glycerophospholipid metabolism, tryptophan metabolism, primary bile acid biosynthesis, biosynthesis of unsaturated fatty acids, nicotinate and nicotinamide metabolism, and tryptophan metabolism. In conclusion, AEE had a positive regulatory effect on NAFLD.