Ilex Rotunda Thunb Research Articles

Resveratrol-Mg hydroxide complex display enhanced bioavailability: A possible application in DM-1 disease

Phytochemical studies on the barks of Ilex rotunda Thunb. had resulted in the isolation of seven previously undescribed triterpenoids, rotundinosides E-K, along with sixteen known ones. The structures of previously undescribed compounds were elucidated on the basis of extensive spectroscopic analysis and the sugar moieties were further identified by HPLC and GC after acid hydrolysis. Among the isolates, rotundinoside F featured a rare triterpene-phenylpropanoid hybrid structure and rotundinoside H was an uncommon triterpene saponin with α-linked glucopyranosyl moiety at C-3. The antiplatelet aggregation of all compounds were evaluated against ADP induced rat platelet aggregation in vitro, and five compounds exhibited moderate inhibitory effects with IC50 values ranging from 22.4 to 32.8 μM.

Rotundarpene inhibits TNF-α-induced activation of the Akt, mTOR, and NF-κB pathways, and the JNK and p38 associated with production of reactive oxygen species.

Ilex Rotunda Thunb has been shown to have anti-inflammatory and antioxidant effects. In human keratinocytes, we investigated the effect of rotundarpene (4-caffeoyl-3-methyl-but-2-ene-1,4-diol) on the TNF-α-stimulated production of inflammatory mediators in relation to the Akt, mTOR, and NF-κB pathways, and the JNK and p38-MAPK. Rotundarpene, Akt inhibitor, Bay 11-7085, rapamycin, and N-acetylcysteine inhibited the TNF-α-stimulated production of cytokines and chemokines, increase in the levels of p-Akt and mTOR, activation of NF-κB, and production of reactive oxygen species in keratinocytes. TNF-α treatment induced phosphorylation of the JNK and p38-MAPK. Inhibitors of the c-JNK (SP600125) and p38-MAPK (SB203580) reduced the TNF-α-induced production of inflammatory mediators, binding of NF-κB to DNA, and activation of the JNK and p38-MAPK in keratinocytes. The results show that rotundarpene may reduce the TNF-α-stimulated inflammatory mediator production by suppressing the reactive oxygen species-dependent activation of the Akt, mTOR, and NF-κB pathways, and activation of the JNK and p38-MAPK in human keratinocytes. Additionally, rotundarpene appears to attenuate the Akt, mTOR, and NF-κB pathways and the JNK and p38-MAPK-mediated inflammatory skin diseases.

Triterpenoid Saponins with Potential Cytotoxic Activities from the Root Bark of Ilex rotunda Thunb.

A new 19-oxo-18,19-seco-ursane-type triterpeonoid saponin, laevigin E (8), together with 17 known compounds (1 - 7 and 9 - 18) were isolated from the root bark of Ilex rotunda Thunb. Their structures were determined by various spectroscopic analysis. Among them, compounds 6, 9, 11, and 18 were isolated from this species for the first time, while compounds 10 and 12 were firstly isolated from the family Aquifoliaceae. Biological activity assay showed that all triterpenoids exhibit moderate cytotoxic activities against MCF7, A549, HeLa and LN229 cell lines. The four triterpenoid saponins (3, 4, 6, and 8) exhibit slightly better activities compared to the four triterpenoid sapogenins (1, 2, 5, and 7). Compound 8 showed the best cytotoxicity against A549, HeLa and LN229 cell lines with IC50 of 17.83, 22.58 and 30.98 μm, respectively.

Rotundarpene attenuates cholesterol oxidation product-induced apoptosis by suppressing the mitochondrial pathway and the caspase-8- and bid-dependent pathways

The extract of from the barks of Ilex Rotunda Thunb has demonstrated anti-inflammatory and anti-oxidant effects. Nevertheless, the effect of rotundarpene (4-caffeoyl-3-methyl-but-2-ene-1,4-diol) on the neuronal cell death induced by cholesterol oxidation products is unclear. We assessed the preventive effect of rotundarpene on the cholesterol oxidation product-induced apoptosis in neuronal cells using differentiated PC12 cells. 7-Ketocholesterol and 25-hydroxycholesterol induced a decrease in Bid, Bcl-2, Bcl-xL and survivin protein levels, increase in Bax levels, loss of the mitochondrial transmembrane potential, cytochrome c release, activation of caspases (-8, -9 and -3), cleavage of PARP-1 and an increase in the tumor suppressor p53 levels. Rotundarpene attenuated the cholesterol oxidation product-induced changes in the apoptosis-related protein levels, formation of reactive oxygen species, depletion of GSH, nuclear damage and cell death. The results show that rotundarpene may attenuate the cholesterol oxidation product-induced apoptosis in PC12 cells by suppressing the activation of the mitochondrial pathway and the caspase-8- and Bid-dependent pathways. The preventive effect appears to be attributed to its inhibitory effect on the formation of reactive oxygen species and depletion of GSH. Rotundarpene appears to attenuate cholesterol-oxidation product-mediated neuronal degeneration.

Antiplatelet aggregation triterpene saponins from the barks of Ilex rotunda

Four new triterpene saponins, rotundinosides A–D (1–4) and seven known triterpene saponins (5–11) were isolated from a methanol extract of the barks of Ilex rotunda Thunb. The new saponins were characterized as 3-O-β-d-glucopyranosy1-(1→2)-β-d-xylopyranosyl siaresinolic acid 28-O-β-d-glucopyranoside (1), 3-O-[β-d-glucopyranosy1-(1→2)-β-d-xylopyranosyl]-3β,19α-dihydroxyurs-12-en-28-oic-O-β-d-glucopranosy1ester (2), 3-O-[α-l-rhamnopyranosyl-(1→2)-β-d-glucopyranosy1-(1→2)-α-l-arabinopyranosyl]-3β,19α-dihydroxyurs-12-en-28-oic-O-β-d-glucopyranosy1 ester (3), and 3-O-α-l-rhamanopyranosyl-(1→2)-β-d-glucopyranosy1-(1→2)-α-l-arabinopyranosyl ilexgenin B 28-O-β-d-glucopyranosy1 ester (4), respectively. Their structures were established by extensive spectroscopic analysis, including HSQC, HMBC, 1H–1H COSY, NOESY and acid hydrolysis, and also by the comparison of their spectroscopic data with those of related compounds. The known compounds 5–11 were all obtained from this species for the first time. The biological activity of compounds 1–11 against ADP induced platelet aggregation in rabbit plasma was determined. Among the tested compounds 1, 3, 5 and 10 exhibited strong inhibition of platelet aggregation in vitro, with IC50 values of 11.4±2.2, 10.4±1.3, 13.2±2.4, and 15.1±3.4μM, respectively.

Rotundarpene prevents TRAIL-induced apoptosis in human keratinocytes by suppressing the caspase-8- and Bid-pathways and the mitochondrial pathway.

The extract and hemiterpene glycosides of Ilex rotunda Thunb have demonstrated antioxidant and anti-inflammatory effects. Nevertheless, the effect of rotundarpene on the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis in keratinocytes that may be involved in skin diseases has not been studied. In this respect, we investigated the effect of rotundarpene on TRAIL-induced apoptosis in human keratinocytes. TRAIL triggers apoptosis by inducing a decrease in the cytosolic levels of Bid, Bcl-2, Bcl-xL, and survivin proteins, increase in the cytosolic levels of Bax, and increase in the mitochondrial levels of VDAC1, loss of the mitochondrial transmembrane potential, release of cytochrome c, activation of caspases (-8, -9, and -3), cleavage of PARP-1, and an increase in the tumor suppressor p53 levels. Treatment with rotundarpene prevented TRAIL-induced changes in the levels of apoptosis-related proteins, formations of reactive oxygen species and nitric oxide, nuclear damage, and cell death. These results suggest that rotundarpene may reduce TRAIL-induced apoptosis in human keratinocytes by suppressing the activation of the caspase-8- and Bid-pathways and the mitochondria-mediated cell death pathway, which is associated with the formation of reactive oxygen species and reactive nitrogen species. These data suggest that rotundarpene appears to be effective in the prevention of TRAIL-induced apoptosis-mediated skin diseases.

ISOLATION OF FIVE GLYCOSIDES FROM THE BARKS OF ILEX ROTUNDA BY HIGH-SPEED COUNTER-CURRENT CHROMATOGRAPHY

Semi-preparative and preparative high-speed counter-current chromatography (HSCCC) were successfully used for isolation of five glycosides from 50% ethanol extract of the dried barks of Ilex rotunda Thunb. (Aquifoliaceae) by using a two-phase solvent system composed of ethyl acetate-n-butanol-water (1:6:7, v/v/v). From 1.0 g of the extract, syringaresinol 4′,4″-di-O-β-D-glucopyranoside ( I , 20.2 mg, 55.4% purity), syringin ( II , 56.8 mg, 98.1% purity), sinapaldehyde glucoside ( III , 26.2 mg, 68.6% purity), syringaresinol 4′-O-β-D-glucopyranoside ( IV , 20.4 mg, 95.3% purity), and pedunculoside ( V , 45.1 mg, 97.3% purity) were obtained by one run of TBE-1000A HSCCC instrument with 1000 mL of column volume. Glycosides I (9.8 mg, 95.5% purity) and III (14.2 mg, 97.8% purity) were further purified by preparative high performance liquid chromatography (HPLC). Their structures were identified by IR, MS, and 1H and 13C NMR studies. Glycoside I was isolated from this plant for the first time.

Rotundarpene inhibits Toll-like receptor 2 activation-induced production of inflammatory mediators in keratinocytes by suppressing the Akt and NF-κB pathways

Microbial components have been shown to be involved in the pathogenesis of inflammatory skin diseases. The extract of from the barks of Ilex rotunda Thunb has demonstrated anti-inflammatory and anti-oxidant effects. However, the effect of hemiterpene rotundarpene (4-caffeoyl-3-methyl-but-2-ene-1,4-diol) on the Toll-like receptor (TLR)-2 activation-induced production of inflammatory mediators in keratinocytes has not been studied. Using human keratinocytes, we investigated the effect of rotundarpene on the inflammatory mediator production in relation to the TLR-2-mediated-Akt and NF-κB pathways, which regulates the transcription genes involved in immune and inflammatory responses. Rotundarpene, Akt inhibitor, Bay 11-7085 and N-acetylcysteine each attenuated the lipoteichoic acid- or peptidoglycan-induced production of cytokines and chemokines, expression of TLR-2, activation of NF-κB and Akt, and formation of reactive oxygen species in keratinocytes. Cyclosporine A attenuated the bacterial component-induced production of inflammatory mediators but did not reduce the formation of reactive oxygen species. The results show that rotundarpene may attenuate the bacterial component-stimulated production of inflammatory mediators in keratinocytes by suppressing the TLR-2-mediated activation of the Akt and NF-κB pathways. The effect of rotundarpene may be attributed to its inhibitory effect on the formation of reactive oxygen species. Rotundarpene may exert a preventive effect against the bacterial component-mediated inflammatory skin diseases.

ENRICHMENT AND PURIFICATION OF PEDUNCULOSIDE AND SYRINGIN FROM THE BARKS OF ILEX ROTUNDA WITH MACROPOROUS RESINS

Pedunculoside and syringin are two bioactive compounds in herbal tea and Chinese drug Jiubiying, the dried barks of Ilex rotunda Thunb. In the present study, the enrichment and purification of pedunculoside and syringin were investigated on the macroporous resins. The evaluation results of performance and adsorption characteristics of eight macroporous resins indicated that SP825 was most suitable for the separation of pedunculoside and syringin. Based on the series of experiments of separation, dynamic adsorption, and desorption on the column of SP825 resin, the optimal conditions were determined as follows: processing volume: 2 BV; temperature: 25°C; for desorption of syringin: ethanol-water (30:70, v/v), 4 BV as an eluent, flow rate: 1.0 mL/min/cm2; for desorption of pedunculoside:ethanol–water (60:40, v/v), 5 BV as an eluent, flow rate: 1.0 mL/min/cm2. One-step treatment of 600 mL Jiubiying extract containing 4.346 mg/mL pedunculoside and 1.478 mg/mL syringin, yielded 7.04 g (29.6% purity) crude pedunculoside powder and 2.32 g (31.0% purity) crude syringin powder. Followed by repeated recrystallization in methanol, 1600 mg pedunculoside with 99% purity, and 533 mg syringin with 96% purity were obtained. The method provides a new approach for scale-up enrichment and purification of pedunculoside and syringin from Jiubiying extract.

Anti-oxidative and anti-inflammatory activities of caffeoyl derivatives from the barks of Ilex rotunda

The barks of Ilex rotunda Thunb. (IR) have been used for the treatment of common cold, tonsillitis and intestinal ulcer. As part of our continuous search for new anti-oxidative and anti-inflammatory agents from natural sources, we tried to isolate caffeoyl derivatives from the bark of IR. Activity guided isolations of the 80% MeOH extract yielded two new caffeoyl hemiterpenes (1 and 2), tentatively named rotundarpene and rotundarpenoside B, together with 6 caffeoyl derivatives which were methyl caffeic acid (3), 5-caffeoylquinic acid (4), 3,5-dicaffeoylquinic acid (5), 1,4-dicaffeoylquinic acid (6), 3,4.5-tricaffeoylquinic acid (7) and 3,5-dicaffeoyl shikimic acid (8), respectively.

Triterpenoid Saponins of Aquifoliaceous Plants. XII. Ilexosides XLVI-LI from the Leaves of Ilex rotunda Thunb.

Six new saponins, ilexosides XLVI-LI, were isolated from fresh leaves of Ilex rotunda, and their structures were elucidated on the basis of chemical and physicochemical evidence. Ilexosides XLVI, XLVII, L and LI are 3, 28-bisglycosides of ilexosapogenin A, spathodic acid, bredemolic acid, and siaresinokic acid, respectively. Ilexosides XLVIII and XLIX are 3, 28-bisglycosides of hederagenin.

Triterpenoid Saponins of Aquifoliaceous Plants. XI. Ilexosides XLI-XLV from the Leaves of Ilex rotunda Thunb.

Triterpenoid Saponins of Aquifoliaceous Plants. XI. Ilexosides XLI-XLV from the Leaves of Ilex rotunda Thunb.

Triterpenoid Saponins of Aquifoliaceous Plants. VIII. Ilexosides XXIX-XXXII from the Leaves of Ilex rotunda Thunb.

Triterpenoid Saponins of Aquifoliaceous Plants. VIII. Ilexosides XXIX-XXXII from the Leaves of Ilex rotunda Thunb.