Study On Chemical Constituents Of Mountain-cultivated Ginseng By NMR And UPLC-QTOF-MS/MS

Panax ginseng,belonging to panax of araliaceae,is a perennial.As a nourished plant and functional food,it is widely used in China,South Korea,Japan and other countries for many years.Ginseng cultivated in the garden field with 3-6 years growth period is usually called “cultivated ginseng”,and it gains weight quickly.Ginseng cultivated in the mountain field with 10-20 years growth period is usually called “mountain-cultivated ginseng”,and it gains weight slowly.Due to imitate the wild ginseng’s growing environment,mountain-cultivated ginseng is similar to wild ginseng.In 2008,the mountain-cultivated ginseng is classified as wild ginseng according to "wild ginseng identification and grading quality"(GB / T 18765-2008).As we all known,the different growth environment,the different quality and efficient.Although Panax ginseng is divided into cultivated ginseng and mountain-cultivated ginseng in the Newest Chinese Pharmacopoeia,there is no available criteria for distinguishing them.Combining the traditional technique of NMR and modern technique of UPLC-QTOFMS/MS,this paper focuses on the mountain-cultivated ginseng(10-20 years)systematic research,aiming at the identification of small molecule chemical composition,and developing an accurate and confidential method for the compound identification,and to provide a new thinking for quality control for the mountain-cultivated ginseng.The main achievements are as follows:1.Volatile oil and other volatile components of 20-year-old mountain-cultivated ginseng were determined for the first time by combining the HS-SPME method and GC-MS.As a result,a total of 58 compounds were identified mainly including unsaturated hydrocarbons(34.3%),alcohols(22.0%),esters(7.8%)and aromatic(11.7%).Among of the 58 compounds,oxacyclotetradeca-4,11-diyne(17.3%),falcarinol(11.9%)and dibutyl phthalate(11.5%)are of the higher content.2.A total of Sixteen compounds were isolated and purified from the 80% methanol extract of mountain-cultivated by silica gel column chromatography,TLC and semi-preparative HPLC chromatography.A total of ten compounds were identified byHR-MS and NMR,and they are butanol isobutanol diester,ginsenoside Rg6,-Rf、-Re、-Rg1、-Rh4、-F5、-Rg2、-Rc、-Rb1.It is worthy to mention that ginsenoside Rg6,-Rh4、-F5 and butanol isobutanol diester were isolated from mountain-cultivated ginseng for the first time.3.With the combining the UPLC-QTOF-MS/MS and UNIFI new information platform,the small molecules(molecular weight 100-1200 Da)in the 80% methanol extract of mountain-cultivated ginseng(10,12,20 years)and cultivated ginseng(4,5,6years)were rapidly analyzed and identified according to the RT and m/z of detected peaks.As a result,forty-six,forty-three and forty-one chemical constituents were identified from mountain-cultivated ginseng,and forty,forty-two and thirty-six chemical constituents were identified from the garden ginseng respectively.4.By using plant metabolomics techniques,UPLC-QTOF-MS/MS combining multivariate statistical analysis methods such as PCA and OPLS-DA,the metabolites of mountain-cultivated ginseng(30 batches)and cultivated ginseng(30 batches)were studied.The results are: PCA score figure shows that mountain-cultivated ginseng and garden ginseng are obviously divided into two categories,indicating that the chemical composition of two kinds of ginseng are significant different.Then,eleven compounds for mountain-cultivated ginseng and six compounds for cultivated ginseng were identified.Among the eleven compounds for mountain-cultivated ginseng,nine of them(structure unidentified)has been not detected in the cultivated ginseng,and there is a significant difference for another two compounds(quinquenoside R1 and malonyl-ginsenoside Rb2)content in the two kinds of ginseng.Meanwhile,the six compounds for cultivated ginseng has not been detected in the mountain-cultivated ginseng.