Integrated untargeted and targeted metabolomics to reveal the mechanisms of herbal medicine HLXLD on Alzheimer's disease

Huo-Luo-Xiao-Ling-Dan (HLXLD), a Chinese herbal medicine consisting of 11 herbs, has been shown to be effective in alleviating cognitive and memory impairment in Alzheimer's disease (AD). However, the underlying mechanisms require further investigation. This study aimed to clarify potential therapeutic mechanisms of HLXLD in the treatment of AD from a metabolic perspective. A rat model of AD induced by AlCl₃ and D-gal was established, the Morris water maze (MWM) test and hippocampal histopathology were used to evaluate the pharmacological effect of the HLXLD on AD. Subsequently, untargeted metabolomics of brain tissues samples was performed by UHPLC-Q-Exactive-Orbitrap-MS, followed by multivariate statistical analysis. Targeted metabolomics by UHPLC-QQQ-MS was further employed to validate and supplement the untargeted metabolomics finding, involving neurotransmitters, amino acids, arachidonic acid and lipids, to elucidate the relationship between disease, herbal medicine and metabolism pathway. The study found that HLXLD could relieve the progression of AD and regulate metabolic imbalances. The levels of 26 metabolites decreased and 6 increased in brain tissues including lysine, taurine, fumaric acid, prostaglandin E2, choline and so on, these altered metabolites were primarily associated with amino acid metabolism, TCA cycle, arachidonic acid metabolism and lipid metabolism. The targeted metabolomics results showed that compared with the model group, the levels of 8 neurotransmitters, 21 amino acids, 7 arachidonic acids and 16 lipids in brain tissue, 9 neurotransmitters, 20 amino acids, 6 arachidonic acids and 2 lipids in plasma were changed. In summary, HLXLD could improve the levels of endogenous metabolites by regulating multiple metabolic pathways and play a role in energy supply, anti-neuroinflammatory and neuroprotective effects in AD treatment.