| Fever is a common clinical symptoms of many diseases, especially is accompanied by a variety of infectious diseases in modern medicine. Based on the point that heat is from the poison of body, clinical commonly use of detoxification methods to treat a variety of infectious diseases. Qing-kai-ling injection with detoxification, sedative effect, has a significant effect in the treatment of fever. It is the first batch of necessary emergency Chinese medicine in clinic. Currently, for the antipyretic effects of Qing-kai-ling injection mostly based on the clinical studies, while the mechanism research is still weak. In this paper, a urinary metabonomics study on biochemical changes in yeast-induced pyrexia rats to elucidating the biochemical basis of the febrile response using UPLC-Q-TOF/MS and HPLC-LTQ/Orbitrap is used. Then, a UPLC Q-TOF/MS-based metabolic profiling of urine is used to explore the perturbed compounds. Meanwhile, a metabolomic strategy for comprehensively elucidating the prototype components and metabolites of Qing-kai-ling injection in rats urine conducted by HPLC-LTQ-Orbitrap/MS was developed. The identification and structural elucidation of the prototype components and metabolites in rats urine could provide essential data for further pharmacological studies of Qing-kai-ling injection. Finally, copy the yeast-induced fever model and select the metabolites which were intervened by Qing-kai-ling injection using metabolomics method, reveal the potential pyretic mechanisms of QKL.injection. The main research contents and results are as follows:1. A urinary metabonomics study in yeast-induced pyrexia ratsEstablishment of yeast-induced pyrexia model, using ultra performance liquid chromatography coupled with a quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) and multivariate data analysis method, to reveal the subtle metabolic changes under the condition of fever and to explore its mechanism with metabonomics approach. And provide a basis for further research the antipyretic mechanism of Qing-kai-ling injection. The results showed that after subcutaneous injection yeast, there were significant differences between the control group and pyrexia group at the time points of5h,9h and13h. UPLC-Q-TOF/MS was used to detect. The acquired data were subjected to principal component analysis (PCA), and found a high relationship between the clustering characteristics and temperature. Based on this, the metabolites of the three points were selected by VIP>3and P<0.05. Sixteen, fifteen and fifteen metabolites in the time-points of5h,9h and13h with higher contributions to clustering analysis were picked out as the potential biomarkers. the rational biomarkers were verified by Pearson correlation analysis, and the predictive power was evaluated using receiver operator characteristic (ROC) curve. Metabolic network constructed by traditional Chinese medicine (TCM) grammar systems was used to validate the rationality of the verified biomarkers. Finally, five biomarkers, including indoleacrylic acid,3-methyluridine, tryptophan, nicotinuric acid and PI (37:3), were confirmed as rational biomarkers. This research provided informative data that the impaired tryptophan metabolism might be one of the important reasons in elucidating the biochemical basis of the febrile response.2. A pharmaco-metabonomics study in Qing-kai-ling injectionUsing pharmaco-metabonomics technology, completely unbiased search the metabolites of Qingkailing injection in rats urine after administration, so as to understand the metabolic characteristics of Qing-kai-ling injection in vivo, which lays the foundation for the antipyretic mechanisms research of Qing-kai-ling injection. A metabolomic strategy for comprehensively elucidating the prototype components and metabolites of Qing-kai-ling injection in rats urine conducted by HPLC-LTQ-Orbitrap/MS was developed.49chemical components in Qing-kai-ling injection,18prototype compounds and19metabolites of Qing-kai-ling injection in urine were found and verified.3. Study of the antipyretic mechanisms of Qing-kai-ling injection in urine with metabolomics strategyCopy the model of yeast-induced fever, selecting the metabolites which were intervened by Qing-kai-ling injection using metabolomics method. To explore the antipyretic mechanism of Qing-kai-ling injection by the disturbances of endogenous in rats after Qing-kai-ling injection administrated. The potential biomarkers were chosen based on VIP>1.5and P<0.05by PLS-DA. MetPA was used to find the possible metabolic pathways. The results indicated that the antipyretic effect of QKL injection on yeast-induced pyrexia rats was performed by repairing the perturbed metabolism of amino acids. |
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