Detection Of Unique Peptides Alteration In Ischemic Stroke Patients And Determination Of Therapeutical Ingredients

Ischemic stroke(IS)has posed serious threat to the health of middle-aged and elderly people in our country.Recently,the Morbidity and mortality of the disease keep increasing.Some pharmacological researches discovered that rotundic acid has effects on the serum lipid reducing activity and the treatment of IS.Rotundic acid can be extracted and purified from Ilicis rotundae Cortex,it is worth to development because of good pharmacological effects.In the development of monomeric drugs,establishing a fast,accurate method to determinate of the content of extracted product and rotundic acid is the key to guarantee the safety and efficacy study of the drug.It has been proven that biomarkers play important roles in diagnosis of IS.At present,some protein biomarkers for IS have been discovered,however,there are still some limits to apply these biomarkers into clinical practice,because of low serum level,low sensitivity and specificity.Here,we collected the Mass spectrum information of APOA1-UP and TF-UP by MRM method,and studied the association between the serum level of two biomarkers and the acute onset of ischemic stroke.Moreover,good biomarkers could be used as the therapeutic indicators,providing important informention to drug mechanism study.The main contents and results are as follows:1)The optimized HPLC method could simultaneously determine syringin,peduncloside,rotundic acid in Ilicis rotundae Cortex and its purified products.Methodological verification shows excellent precision,reproducibility linearity,and average recovery rate.The proposed method can be used for the quality control of Ilicis rotundae Cortex and its extracts.It provides the experimental basis for rotundic acid development2)MRM-LRP method based on TQ-MS,was set up to make relative quantification.Statistical analysis showed that negative correlation of the APOA1-UP,TF-UP levels and IS incidence,As the plasma levels of APOA1-UP or TF-UP decrease,the possibility of the occurrence of IS increases.The optimal cutoff value of the serum APOA1-UP level was 1.8031 and the AUC was 0.9750,which yielded a sensitivity of 90.63% and a specificity of 97.14%.The optimal cutoff value of serum TF-UP level was 0.6317,and the AUC was 0.9565,which yielded a sensitivity of 91.49 % and a specificity of 88.57%.So,the level of APOA1-UP and TF-UP have good diagnostic ability of IS disease.Conclusions: The HPLC method is simple and accurate,and easily applied to quick quality evaluation of Ilicis rotundae Cortex and its purified products.The serum level of APOA1-UP and TF-UP could be diagnostic biomarker candidates for ischemic stroke with high sensitivity and specificity.But it is still necessary to further confirm the effectiveness of their clinical application by expanding the clinical sample size and verifying the mechanism in the disease development.This study provided an experimental basis for quality control and the mechanism study in the development of rotundic acid.