Study On The Chemical Constituents And Anti-inflammatory And Immunomodulatory Activity In Vitro Of Litsea Coreana L.

Laoying tea is leaf of Litsea Coreana level which has been used as a tea in Guizhou, Sichuan and south of Anhui province in China. It has been reported that the extract from the leaves of Litsea Coreana L. exerts various pharmacological effects including antioxygen, antihyperlipidemic, antihypertensive, anti-inflammatory and immunomod- ulation. It contains many chemical compositions, such as flavonoid glycoside, saponin, organic acid, cumoric, anhydride and tannin et al. Total flavonoids is the effective component in the extract from leaves of Litsea Coreana L. We had studied on the chemical constituents of n-butanol extract obtained from leaves of Litsea coreana L. and obtioned four compounds which were accredited as quercetin-3-o-β-D-galactopyr- anoside, kaempferol-3-o-β-D-glucopyranoside,quercetin-3-o-β-D-glucopyranoside and kaempferol-3-o-β-D-galactopyranoside. The research was aimed to accumulate and purify the total compounds, separate and elucidate individual compounds from the acetoacetate extract, and then screen the anti-inflammatory and immunomodulatory activity of isolated flavonoids and their glycosides monomers in vitro.First Section:To accumulate and purify the chemical constituents and identify the components from the extracts of Litsea Coreana L.The leaves of Litsea coreana L.(10 kg)were extracted three times with 10-fold and 70% ethanol EtOH for 4h. The solution were combined concentrated under reduced pressure. The concentrated extract was distributed between petroleum and water, the water layer was further shakened with acetoacetate, and then the acetoacetate layer was concentrated under reduced pressure and subjected to chromatography through a silica gel column eluted with DAB-6-EtoAc and CHCl3- MeOH and then chromatographed through Sephadex LH-20 repeatedly and yielded compound 1, Quercetin, compound 2, Kaempferol, compound 3, (+)-Catechin, compound 4,β-Sitosterol, compound 5, Daucosterin. The compounds were identified by means of physical and chemical properties, TLC and spectroscopic analysis such as UV, 1H-NMR, 13C-NMR and MS.Second section: The activity screening of the monomers of flavonoids on anti-inflammatory and immunomodulatory in vitroThe experiment meaned to study the structure-function relationship of the monomers of flavonoids on immune function of rats with adjuvant arthritis. Adjuvant arthritis(AA) was induced on d0 by intradermal injection of Freund's Complete Adjuvant (FCA). On d 28 after immunization. AA rats were killed by cervical decollation. Immune cells(splenocytes and peritoneal macrophages) were obtained,then cultured in vitro with the five kinds of flavonoids we had choosed. Some immune indexes were detected respectively such as the productions of IL-1 and prostaglandin E2 (PGE2) were measured by the method of enzyme linked immunosorbent assay, the productions of IL-2 and tumor necrosis factor-α(TNF-α) was determined by radioimmunoassay, making a primary screening to the monomers of flavonoids on anti-inflammatory and immunomodulatory activity. We found that all the five kinds of flavonoids have activity on immune regulation , meanwhile, because of the differences of structure-function relationship between compounds, such as numbers of phenol–OH, types of glycosidic bond et al, there were differences on immunoregulatory activity. According to the date of 50% concentration of inhibition(IC50), the order was: kaempferol-3-o-β-D-glucose> kaempferol> quercetin> kaempferol-3-o-β-D-galactose> quercetin-3-o-β-D- galactose.