Studies On The Chemical Constituents And Quality Control Of Perilla Frutescens (L.) Britt

The leaves, stems and fruits of perilla frutescens (L.) Britt.are used as traditional medicine in china, which are paid a lot of attention to for their remarkable bioactivities. In order to control their quality and utilize them better, this essay gives a comprehensive talk about research and analysis of Perilla frutescens in five aspects, including its growing, best harvest date, processing, chemical constituents research and preparation of mainly bioactive substance.To find out their best harvest time of the leaves, Perilla frutescens (L.) Britt. var. typica (Makino) Makino (abbr:PFT), Perilla frutescens (L.) Britt. var. acuta (Thunb.) Kudo (abbr: PFA) and Perilla frutescens (L.) Britt. var. acuta (Thunb.) Kudo form. discolor (Makino) Kudo (abbr:PFAD) were grown in Northwest University fruit garden in March, and their leaves were harvested from May to October on the second day of each month. Then the content of rosmarinic acid in them were determined by HPLC with Welch RP-C18 (250 mm×4.6 mm,5μm) column, using methanol-phosphonic acid solution (39:61) as mobile phase at flow rate of 1.0 mL/min. UV detection wavelength was set at 330 nm and column temperature was set at 28℃. As a results, the content of rosmarinic acid in leaves of PFAD and PFA reach the highest in August. The content of rosmarinic acid in leaf of PFT increases from May to August and reaches the highest in August too. So August is the best month for harvesting leaf of PFT, PFA and PFAD.The content of rosmarinic acid in the seeds harvested in October and its processed products of PFT, PFA and PFAD were determinated with HPLC. Consequently, the content of rosmarinic acid in roasted seeds of PFT is higher than in its seeds, but that of PFA and PFAD are lower; The content of rosmarinic acid in seeds of PFT, PFA and PFAD decline after the seeds are honey-pocessed and made into frostlike powder, and that of honey-processed one decline by a larger margin. Growing again,6.26kg dry leaves of PFAD were harvested in the middle of October. Silicon gel column chromatography was used to isolate the chemical constituents from the leaves of PFAD and five compounds were obtained. By means of spectroscopy (IR, MS, 1H-NMR, 13C-NMR) and TLC with reference substance, they were elucidated as phytol, stigmasterol, daucosterol,β-sitosterol and rosmarinic acid.High speed counter-current chromatography was applied for the preparation and purification of rosmarinic acid, the major bioactive constituent, from ethyl acetate part of the crude extract of PFAD. Petroleum ether-ethyl acetate-methanol-(0.5% acetic acid) water(2:5:2:5, V/V/V/V) was used as the two-phase solvent system. The upper phase was chosen as the stationary phase, and the lower phase was chosen as the mobile phase. The mobile phase was operated at a flow rate of 2.0 mL·min-1, while the apparatus rotated at 800 r·min-1 and the temperature was set at 25℃. As a result,25.7 mg rosmarinic acid with 98.6% purity is obtained from 200.4mg ethyl acetate part of the crude extract of PFAD by once separation with HSCCC, which is a perfect new method for the preparation and purification of rosmarinic acid.