Studies On The Chemical Constituents And Bioactivities Of Justicia Procumbens L

Justicia procumbens L.(Acanthaceae), known as “Juechuang” in China, is acommonly used traditional herbal medicine embodied in Chinese Pharmacopoeia1977version, with clinical effects of reducing fire, removing toxic heat. The wholeplant of J. procumbens mainly distributed in South China and Taiwan has long beenused to treat cold, fever, cough, pain, inflammation of the throat and indigestion inchildren, etc. Several arylnaphthalide lignans showing cytotoxic and antivirusactivities have been reported as main constituents of this plant.Preliminary tests on66Chinese herbs for anti-H5N1virus activity in vitroshowed that the ethanol extract, CHCl3extract and BuOH extract of J. procumbenspossessed obvious activity of virus-resistance. Based on the review on the chemicalconstituents and bioactivities research of the Justicia, chemical constituents of J.procumbens were studied systematically and deeply, their anti-H5N1virus andantitumor activities were tested to obtain compounds with novel structures orsignificant bioactivities.In our investigation, by means of many different chromatographic methods suchas silica gel CC, reversed phase C18, Sphadex LH-20, MPCC, preparative HPLC etc,58compounds isolated from the60%ethanol extract of the herbs of J. procumbenswere elucidated on the basis of chemical and spectral evidence (IR、NMR、MS). Theywere identifie as: Pronaphthalide A (1), Procumbenoside J (2), Procumbiene (3),Juspurpudin (4), Procumbenoside H (5), Procumbenoside I (6), Procumbenoside K (7),Procumbenoside L (8), Procumbenoside M (9),Justicianene A (10), Justicianene B(11), Justicianene C (12), Justicianene D (13), Justicidin C (14), Justicidin D (15),Justicidin E (16),5′-methoxy retrochinensin (17), Detetrahydroconidendrin (18),Justicidinoside A (19), Justicidin A (20), Justicidin B (21),6ˊ-hydroxy justicidin A(22),6ˊ-hydroxy justicidin B (23), Diphyllin (24), Taiwanin C (25), Tuberculatin (26),Cleistanthin B (27), Justicidinoside B (28), Justicidinoside C (29), procumbenoside A(30), procumbenoside B (31), Rostellulin A (32),(+)-Pinoresinol (33),(﹣)-syringaresinol (34),(﹣)-medioresinol (35), epipinoresinol (36), Isoscopletin (37),Vanillovanillone (38), Vanillosyringone (39), Chrysophanol (40),1,7-dihydroxy-3,8- dimethoxy xanthone (41), Ursolic acid (42), β-Sitosterol (43), stigmasterol (44),β-Daucosterin (45),4-hydroxy benzoic acid (46), vanillic acid (47), Octadeca-9,16-dioxo-10(E),12(Z),14(E)-trienoic acid (48), Pinellic acid (49), quercetin (50),Apigenin-7-O-β-D-(6"-p-coumaroyl)-glucopyranoside (51), Apigenin-7-O-β-D-glucopyranoside (52), acacetin-7-O-β-D-lutinoside (53), Lumichrome (54),3-indole-acid (55),5-Hydroxypyrrolidine-2-one (56), adenosine (57), urea (58), respectively.Compounds1-13were new compounds, including9lignans and4cyclopeptidealkaloids, and19of them were first isolated from Justicia，they were compounds17,18,27,34,36,38-41,46,48-49,51,53-58, compounds33and35were first isolatedfrom J. procumbens.The compounds obtained from J. procumbens referred to32lignans,4cyclopeptide alkaloids,4flavones,5nitrogen containing compounds and13otherstructural types. Four cyclopeptide alkaloids from the50%ethanol elution of theBuOH extract from J. procumbens were first discovered from Justicia. Previousstudies on the chemical constituents of J. procumbens home and abroad focusedmainly on the lignans extracted with high concentration methanol or ethanol, but onlya few alkaloids were isolated from the other species of Justicia, the cyclopeptidealkaloids and other nitrogen containing compounds of the BuOH extract from the60ethanol extract of J. procumbens would open up a new approach to the study onchemical constituents for species of Justicia.By CPE method, the in vitro anti-H5N1virus activity of some compounds,mainly including15lignans, isolated from the CHCl3extract of J. procumbens wasassayed and evaluated. The tests showed that compounds26and33exhibitedantiviral activity at some degree. At the effective dose of37.5and150μg/mL,respectively, compounds33and26abundant in J. procumbens, exerted potentanti-H5N1virus activity in MDCK cell cultures which did not show obviouscytopathic effect on cell viability, proliferation or metabolism, and the exact effect ormechanism should be proven further by hemagglutination test and in vivo studies.On the basis of the pharmacological tests which showed the lignans of J.procumbens possessed significant cytotoxic activity, two tumor cell lines of humanLoVo colon carcinoma cells and human BGC-823gastric carcinoma cells wereselected to test and evaluate cytotoxicity preliminarily of ligans from J. procumbensin vitro with MTT method, consequently, compounds1,2,20,21,22,23,24and26showed significant cytotoxicity, among them, the IC50value of compound1was0.13 and0.11μM against Human LoVo and BGC-823cell lines, respectively; that ofcompound2was0.53and0.20μM, respectively; the IC50values of compound20were0.04μM on both cell lines; the IC50value of compound21was6.08and0.18μM, respectively; that of compound22was0.15and0.42μM, respectively; the valueof compound23was2.36and4.68μM, respectively; that of compound24was8.12and8.08μM, respectively, and the IC50value of compound26was1.63μM againstHuman BGC-823cell line. Compounds5and27showed moderate cytotoxicityagainst Human LoVo cell line selectively with the IC50value of17.91and21.48μM,respectively, others showed no or weak cytotoxicity. Statistical analysis betweengroups showed the cytotoxicity against Human LoVo cell line of compounds1,2,20,22and23was significantly higher than that of Diphyllin (24) as the positive control,and the cytotoxicity against Human BGC-823cell line of compounds1and20wassignificantly higher than that of JR6(22) as the positive control, in addition, thecytotoxicity of compound20was significantly higher than all the other compounds (P<0.01). The structure-activity relationship (SAR) of the arylnaphthalide lignans of J.procumbens was discussed preliminarily on the basis of the MTT assay, the analysisrevealed that the parent structure of6,7-dimethoxy-1-arylnaphthalide lactone was theessential element for the cytotoxicity against cancer cell lines of this type ofcompounds, other different functional groups played role in the cytotoxicity to avariable extent.