Evaluation Of Chemical Constituents In Perilla Frutescens(L.) Britt.

Perilla frutescens(L.)Britt.is an annual herb of Labiatae.The Pharmacopoeia of the People's Republic of China(2015)recorded three parts of Perilla(leaves,stems and seeds)as medicinal herbs,each part having its own efficacy.Modern studies have shown that volatile oils,flavonoids and phenolic acids are the main chemical constituents and active ingredients in Perilla.According to the relative content of the main components in the volatile oil of Perilla,it can be divided into different chemotypes.The main chemotype of Perilla resources in China is PK(perillaketone)type.However,in 1977,it was reported that perillaketone may have potentially toxic to hunman lungs.The correct chemotype of medicial Perilla should be PA type(the qualified perilla leaves are PA type according to Chinese Pharmacopoeia).Therefore,it is of great significance to find,research and cultivate the Perilla germplasms of PA type.The phenolic acids in Perilla are mainly rosmarinic acid and caffeic acid.The Chinese Pharmacopoeia stipulates that the content of Perilla stems and seeds is rosmarinic acid(no less than 0.1% and 0.25%,respectively).Flavonoids and their glycosides in Perilla are mainly monosaccharide and disaccharide glucuronide glycosides with scutellarein,luteolin and apigenin as the parent nucleus,which have antioxidant and antitumor activities.In this paper,the volatile oils in perilla leaves were extracted by steam distillation,headspace solid-phase microextraction,supercritical carbon dioxide extraction and organic solvent extraction.The composition and relative content of the volatile oils were determined by GC-MS.The effects of different extraction methods on the extraction of volatile oil from Perilla frutescens leaves of different germplasms were compared.The results showed that water distillation was the best method for extracting volatile oil.For the leaves of Perilla frutescens with different chemotypes of volatile oil,there were differences in the extraction rate of volatile components and the number of compounds obtained by different extraction methods.The extraction methods of volatile components from Perilla frutescens should be selected scientifically according to different needs.If the experimental materials are sufficient and the aim is to know the composition of volatile compounds in a comprehensive way,steam distillation should be chosen.If the experimental materials are limitted,supercritical extraction or solid-phase microextraction can be chosen to help extract volatile components quickly and analyze their main components.Non-PK type's germplasms were selected from 800 perilla germplasms by odor exclusion method,and the volatile oil from perilla leaves were extracted by steam distillation,and its chemical constituents were isolated and identified by GC-MS.62 germplasms of non-PK perilla were obtained and 38 traditional PA germplases and 2 PL germplases were identified from them.Three chemotypes were identified as new single-component chemotypes or subtypes,such as 1 PA-II germplasm with high content of D-limonene;3 PT-II germplasms with high content of(-)-piperitone,but no piperitenone;2 PS germplasms mainly contained sesquiterpenes,PS-c type dominated by caryophyllene oxide and PS-f type dominated by(Z,E)?-farnesene.Among the mixed chemotypes,there were 3 PAPK types and 6 new mixed types,including 1 PAPT type,1 PAPP type,1 PKPP type,2 PAPS types and 1 PPPS type.A method for simultaneous analysis of the contents of eight flavonoids and phenolic acids by UPLC was established.The content of three components of perilla,scutellarein 7-O-diglucuronide,luteolin 7-O-diglucuronide and apigenin 7-Odiglucuronide was first reported.72 samples of the perilla(leaves,stems,and seeds)samples were analyzed.The samples can be divide into three parts: the first part contained 8 germplasms,42 samples(PA,PK,PP-a,PP-m)and they were palnted in IMPLAD and collected from different harvest time;the second part is 5 PA type germplasms,17 samples and they were all planted in 4 different places(IMPLAD,Anhui Taihe,Henan Pingyi,Gansu Qingyang)respectively;the third part contains 18 new PA samples planted in IMPLAD.There were 10 samples of Perilla stems and perilla seeds respectively,corresponding to 10 germplasms in the third part of perilla leaf samples.The content of flavonoids and phenolic acids in Perilla leaves of different germplasms was significantly different;The content of phenolic acids in different germplasms of Perilla frutescens is different,the total content of flavonoids is not different,and the difference between LDG and LG in different germplasms is not significant.The phenolic acid content of Perilla frutescens from different germplasms was less than that of Perilla leaves,but the flavonoid content had little difference.The main effects of growth and development on the content of the two components were as follows: the content of phenolic acids in most perilla germplasms was higher in early panicle stage,lowest in early flowering stage,and then highest in fruit ripening stage;the content of flavonoids in some germplasms was highest in early flowering stage,and was negatively correlated with the metabolism of phenolic acids,however in some germplasms the content of flavonoids was not affected by development.It was found that environmental changes had a greater impact on the content of phenolic acid components,and different germplasms had different degrees of impact.The impact of environment on flavonoids in germplasms could be divided into two situations: one was that environmental factors had the same effect on all germplasms,the other was that the content of flavonoids did not change with the change of environment.In addition,it is suggested that rosmarinic acid,scutellarin-7-O-diglucuronide,scutellarin-7-Oglucuronide,luteolin-7-O-diglucuronide,luteolin-7-O-glucuronide,and apigenin-7-Odiglucuronide can be used as the quality standard components of Perilla leaves,and flavonoids should be used instead of rosmarinic acid as the quality standard component of Perilla stem.This study has certain guiding significance for selecting more scientific methods of volatile oil extraction.The new chemotypes selected and discovered provide valuable materials for the subsequent study of the anabolism of the main components in volatile oil.Quantitative analysis of flavonoids and phenolic acids in Perilla frutescens(leaves,stems and seeds)are of guiding significance to the selection of cultivation sites,selection of the best harvesting period of perilla leaves and selection of Perilla germplasms with high flavonoids and phenolic acids content.