Source Identification,Quality Evaluation And MiRNA Analysis Of Milk Thistle(Silybum Mariauum(L.) Gaertn.)

Milk thistle was a natural and medicinal plant which belongs to the compositae plants.It has a high medicinal value and its seeds were used to extract silymarin.Silymarin was the main ingredient of many clinical drugs.It was commonly used to treat acute and chronic hepatitis,cirrhosis and other diseases.In recent years,silymarin has been found to has anti-tumor activity,and the study on its pharmacodynamics has entered the clinical trial stages.In this paper,the seeds of milk thistle from different batches were used as the research objective,and the source identification of milk thistle was studied and analyzed by different methods such as character identification,microscopic identification,DNA barcoding and high performance liquid chromatography.The contents of mositure,ash,extract and foreign organisms were determined by different methods.The contents of heavy metal in these samples was detected by atomic absorption spectroscopy.The contents of silymarin,silybin,silychristin,silydianin and isosilybin were determined by HPLC.Additionally,the miRNA database of stem,leaf and flower was established by high-throughput sequencing of milk thistle.The conserved miRNA and novel miRNAs were identified and analyzed in this paper.The detailed results were listed as following:1.A total of 10 batches of milk thistle samples were collected from Jilin,Inner Mongolia,Liaoning and Heilongjiang province,respectively.Besides,source identification was carried out by four methods,including character identification,microscopic identification,DNA barcoding and high performance liquid chromatography(HPLC).The results showed that milk thistle could be identified by the above four methods,however,the character identification was more simple,and the methods of DNA barcoding technology and high performance liquid chromatography were more accuracy.2.The contents of mositure,ash,extract and foreign organisms were detected in this paper.The average mositure of the 10 batches arrived 6.67% and the average ash content was 4.97%.The 10 batches average content of the extract reached 29.3% and the average content of foreign organisms was 1.39%.All these contents conformed the demand of Chinese Pharmacopoeia(CP)and United States Pharmacopoeia(USP).The average content of Pb,As,Cd,Cr,and Cu was 0.099 mg/kg,0.258 mg/kg,0.093 mg/kg,0.135 mg/kg,and 18.89 mg/kg,respectively.The atomic absorption spectrometry was used to detect the contents of heavy metals Pb,Cu,Cd,Cr and As.the result showed that all elements were in accordance with the United States Pharmacopoeia(USP).We found that the average content of silymarin was 3.179%,which was in accordance with the standard of the United States Pharmacopoeia,and the average concentration of silybin was 1.034%,in accordance with the provisions of the Chinese Pharmacopoeia(CP)standards.Additionally,the average content of isosilybin,silychristin and silydianin is 0.066%,0.063% and 0.035%,respectively.Then we analyzed the principal component by principal component analysis(PCA),and found that silymarin,silybin,and isosilybin were the first principal component of milk thistle.3.The small RNA libraries were built by high-throughput sequencing for the stem,leaf and flower of the milk thistle.A total number of 2776 conserved mature miRNAs generated from 2610 miRNA precursors and 303 novel mature mi RNAs generated from 196 miRNA precursors were found in this paper.Finally,12 conserved miRNAs and 12 novel miRNAs were examined by qRTPCR and the results were consistent with the sequencing report.