Secondary Metabolites Of Three Plants And Their Biological Functions

This dissertation is composed of four chapters.Chapter 1 described the investigation on the secondary metabolites of the glandular trichomes,aerial part and root of Colquhounia elegans and their biological functions.Chapter 2 elaborated the research on the secondary metabolites of the latex and whole plant of Euphorbia helioscopia and their biological functions.Chapter 3 illustrated the metabolic profiling of the latex of Euphorbia peplus and exploration of its nematicidal activity and molecular mechanism.Chapter 4 reviewed the recent progress on the anti-inflammatory activity of natural diterpenoids.By multiple chromatographic techniques and modern spectroscopic methods,a total of 98 compounds of different structure types,including triterpenoid saponins,diterpenoids,sesquiterpenoids,flavonoids,were isolated and identified from C.elegans and E.helioscopia.Among them,47 terpenoids were new,including labdanes,triterpenoid saponins with a novel rearranged skeleton,jatrophanes and lathyranes.Chapter 1 Secondary metabolites of Colquhounia elegans and their biological functionsGlandular trichomes produce large amounts of various types of secondary metabolites with diverse structures and extensive biological activities.Though utilizing laser microdissection(LMD)coupled with UPLC-MS/MS techniques,three labdane diterpenoids(ce-1-3)including two new ones,elegansnoids A(ce-2)and B(ce-3),were identified from the peltate glandular trichomes of C.elegans.Compound ce-1 showed moderate immunosuppressive activity against IL-2 production with IC50 of 26.46±0.98?M,but no obvious nematicidal and antifeedant activities were observed.A detailed phytochemical investigation on the aerial part of C.elegans was also carried out,which led to the isolation and identification of 47 compounds including 27 new diterpenoids.The new compounds were mainly labdane diterpenoids possessing either a furan moiety,a spiro ring moiety,an a,p-unsaturated lactone or an?,?-unsaturated lactam in the C-9 side chain.Compound ce-22 showed significant inhibitory activity against angiotensin converting enzyme.Compound ce-10 showed obvious immunosuppressive activity on the release of IL-2,with IC50 of 16.12±0.85?M.Compounds ce-4 and ce-7 exhibited obvious inhibitory activity on the release of cytokine TNF-?,with IC50 of 15.23±0.01?M and 18.22±1.94?M,respectively Meanwhile,both compounds showed weak cytotoxic activity against four cancer cell lines NCI-H1975,XLA-07,PC9,and HepG2.Further phytochemical investigation on the roots of C.elegans uncovered 16 compounds,including 14 new triterpenoid saponins.Among them,ce-51 and ce-52 were triterpenoid saponins possessing an unusual rearranged 6/6/6/6 carbon skeleton,which were confirmed by single crystal X-ray diffraction.Interestingly,ce-51,ce-54,ce-60 and ce-62 showed significant immunostimulating activity by enhancing TNF-?and IL-6 production in macrophages in the presence of lipopolysaccharide(LPS).This is the first report of the secondary metabolites of Colquhounia elegans and their biological functions,which provided scientific basis for sustainable utilization of this plant resource.Chapter 2 Secondary metabolites of Euphorbia helioscopia and their biologicalfunctionsPlant latex,a milk-like endogenous fluid,is stored in laticifer cells and secreted immediately upon plant tissues being damaged and has been suggested to be a defense system for plants.We found that the latex of E.helioscopia displayed significant nematicidal activity against Caenorhabditis elegans,with LD50 value of 5.60 ?g/mL.Five compounds(eh-1-5)were isolated and identified from the latex,including a new jatrophane diterpenoid(eh-3)and four known macrocyclic diterpenoids.Compound eh-4 exhibited significant nematicidal activity with LD50 value of 18.25 ?g/mL,suggesting that eh-4 was the major active nematicidal compound in the latex.In addition,eh-4 exhibited moderate inhibitory activity on the release of cytokine TNF-?(IC50=23.69±1.71 ?M),and chemokine MCP-1(IC50=33.69±3.78 pM)in LPS induced RAW 264.7 macrophages.In addition,weak cytotoxicity was also observed for eh-4.Further chemical investigation on the whole plant of E.helioscopia uncovered 27 compounds,including two new jatrophanes and a new lathyrane.All three new compounds showed significant antifeedant activity against a generalist plant-feeding insect,Helicoverpa armigera,with EC50 values ranging from 2.05±0.11 to 4.34±0.16?/cm2.In addition,compound eh-7 showed moderate cytotoxicity,and compounds eh-13,eh-16 and eh-21 exhibited weak cytotoxicity.Our finding not only revealed the secondary metabolites of latex of E.helioscopia,but also identified the major nematicidal component in the latex,which provided reference value for sustainable utilization of this rich plant resource.Chapter 3 Biological function of secondary metabolites of E.peplusEuphorbia is the largest genus in the family Euphorbiaceae,and it is well known representative species containing latex.Euphorbia peplus is a small annual weed with milky latex.Through TLC,HPLC and UPLC-MS/MS analyses and comparison with the authentic compounds isolated from the whole plant,a total of 13 terpenoids including twelve diterpenoids and an acyclic triterpene alcohol were identified in the latex of E.peplus.20-Deoxyingenol-3-angelate(ep-12)exhibited significant nematicidal activity,with LD50 value of 15.70 ?g/mL against C.elegans.The preliminary nematicidal mechanism of ep-12 was explored in the animal model C.elegans.Exposure of C.elegans to ep-12 led to decrease pharyngeal pumping rate,brood size and delay larval development.Interestingly,N-(?-D-glucopyranosyl)indole,was isolated as a nematode growth retardation maker from the nematodes induced by ep-12,indicating that ep-12 could greatly affect the metabolism of C.elegans.RNA-seq analysis revealed ep-12 treatment caused significantly upregulated or downregulated expression of a series of genes related to cellular processes,metabolism and genetic information processing was in C.elegans.Further quantitative reverse transcription-PCR(qRT-PCR)results indicated that the expression of genes reltaed to apoptosis was upregulated,while the expression of genes related to metabolism was downregulated,in the nematodes after ep-12 treatment.The results not only enriched the chemical diversity of terpenoids in E.peplus latex,but also disclosed the major latex nematicidal compound and its preliminary mechanism of action,which heralded the potential application of latex-producting plants in natural pesticides.Chapter 4 Recent progress on the anti-inflammatory diterpenoidsDiterpenoids are a class of terpenoids composed of four isoprene units.A variety of structural types such as labdane,kaurane,abietane,ingenane,clerodane,and so on,have been reported from various organisms.Many of them possessed significant biological activities,including anti-tumor,antimicrobial,anti-inflammatory,and so forth.New structural diterpenoids and their biological activities have been a continuing hot topic in natural product chemistry.This chapter reviewed the advances in anti-inflammatory diterpenoids in the past ten years,which might help for the development and utilization of natural diterpenoids.