Dan Yuan

Demethoxycurcumin, a natural derivative of curcumin attenuates LPS-induced pro-inflammatory responses through down-regulation of intracellular ROS-related MAPK/NF-κB signaling pathways in N9 microglia induced by lipopolysaccharide

Our previous report has showed that demethoxycurcumin (DMC), a natural derivative of curcumin (Cur), exhibited stronger inhibitory activity on nitric oxide (NO) and tumor necrosis factor-alpha (TNF-alpha) production compared with Cur in lipopolysaccharide (LPS) activated rat primary microglia. In the present study, the effect and possible mechanism of DMC on the production of pro-inflammatory mediators in LPS-activated N9 microglial cells were further investigated. The results showed that DMC significantly suppressed the NO production induced by LPS in N9 microglial cells through inhibiting the protein and mRNA expression of inducible NO synthase (iNOS). DMC also decreased LPS-induced TNF-alpha and IL-1beta expression at both transcriptional and protein level in a concentration-dependent manner. Further studies revealed that DMC blocked IkappaBalpha phosphorylation and degradation, inhibited the phosphorylation of mitogen-activated protein kinases (MAPKs). Moreover, the level of intracellular reactive oxygen species (iROS) was significantly increased by LPS, which is mainly mediated by the up-regulated expression of gp91phox, the catalytic subunit of nicotinamide adenine dinucleotide phosphate reduced (NADPH) oxidase. Both DMC and Cur could markedly decrease iROS production and the expression of NADPH oxidase induced by LPS, with more potent inhibitory activity of DMC. In summary, these data suggest that DMC exerts its in vitro anti-inflammatory effect in LPS-activated N9 microglial cells by blocking nuclear factor-kappaB (NF-kappaB) and MAPKs activation, which may be partly due to its potent down-regulation of the NADPH-derived iROS production.

Alkaloids from the leaves of Uncaria rhynchophylla And Their Inhibitory Activity on NO production in lipopolysaccharide-activated microglia

Two new isomeric alkaloids, 18,19-dehydrocorynoxinic acid B (1) and 18,19-dehydrocorynoxinic acid (2), were isolated from the CHCl3 extract of the leaves of Uncaria rhynchophylla, together with four known rhynchophylline-type alkaloids, corynoxeine (3), isocorynoxeine (4), rhynchophylline (5), and isorhynchophylline (6), and an indole alkaloid glucoside, vincoside lactam (7). The structures of compounds 1 and 2 were elucidated by spectroscopic methods including UV, IR, HREIMS, 1D and 2D NMR, and CD experiments. The activity assay showed that compounds 3-6, with a C-16 carboxylic ester group, and 7 exhibited inhibitory activity on lipopolysaccharide (LPS)-induced NO release in primary cultured rat cortical microglia (IC 50: 13.7-19.0 microM). However, only weak inhibitory activity was observed for compounds 1 and 2, with a C-16 carboxylic acid group (IC 50: >100 microM).

Oleanane-type triterpene oligoglycosides with pancreatic lipase inhibitory activity from the pericarps of Sapindus rarak

The methanolic extract from the pericarps of Sapindus rarak DC. was found to show pancreatic lipase inhibitory activity (IC50=ca. 614 microg/mL). From the extract, oleanane-type triterpene oligoglycosides, rarasaponins I-III (1-3), and raraoside A (4), were isolated together with 13 known saponins and four known sesquiterpene glycosides. Among them, several saponin constituents including rarasaponins I (1, IC50=131microM) and II (2, 172microM), and raraoside A (4, 151microM) inhibited pancreatic lipase activity, which were stronger than that of theasaponin E(1) (270microM).

Cytotoxic activity of flavonoids from the flowers of Chrysanthemum morifolium on human colon cancer Colon205 cells.

A new p-hydroxyphenylacetyl flavonoid, diosmetin 7-(6″-O-p-hydroxyphenylacetyl)-O-β-d-glucopyranoside (1), was isolated from the flowers of Chrysanthemum morifolium Ramat. ‘huaiju’ cv. nov. (Compositae), together with five known flavonoids, luteolin (2), diosmetin (3), diosmetin 7-O-β-d-glucopyranoside (4), diosmin (5), and scolimoside (6), and four known caffeoylquinic acid derivatives, macranthoin F (7), 3,5-dicaffeoylquinic acid (8), 1,3-dicaffeoyl-epi-quinic acid (9), and chlorogenic acid (10). The structure of 1 was elucidated by UV, IR, ESI-TOF-MS, 1D, and 2D NMR spectroscopic methods. Cytotoxic activity of compounds 1–5 against human colon cancer cell Colon205 was investigated using MTT assays. Compounds 2 and 3 showed significant cytotoxicities against Colon205, with their IC50 values being 96.9 and 82.9 μM, respectively. However, compounds 1, 4, and 5 showed little cytotoxic activity.

Comparison of inhibitory potency of three different curcuminoid pigments on nitric oxide and tumor necrosis factor production of rat primary microglia induced by lipopolysaccharide

Microglia are the resident innate immune cells in the central nervous system. Evidence supports that the unregulated activation of microglia results in the production of pro-inflammatory cytokines and chemokines that propagate neuronal injury and finally cause neurodegenerative diseases. Curcuminn (Cur), demethoxycurcumin (DMC), and bisdemethoxycurcumin (BDMC) are curcuminoid pigments extracted from turmeric (Curcuma longa L.). Cur has been reported to suppress the activation of microglia by reducing toxic factors production, but little is known about whether the two natural demethoxy derivatives of Cur, DMC and BDMC, have the similar effects as Cur. In the present study, we found that all of the three curcuminoid pigments significantly suppressed nitric oxide (NO) production by LPS-activated microglia and the relative potency was DMC>BDMC>Cur. This result was verified by RT-PCR analysis of iNOS mRNA. The NO-scavenging abilities of three curcuminoid pigments are very weak, which suggested that the indirect effect may not be critical in inhibiting NO production by LPS-activated microglia. Moreover, these three curcuminoid pigments attenuated the expression of mRNA and proteins of tumor necrosis factor-alpha (TNF-alpha) in a concentration-dependent manner and the relative potency was also DMC>BDMC>Cur. In conclusion, Cur, DMC and BDMC were found as potent microglia-activation inhibitors, and DMC exhibited the strongest inhibitory activity on NO and TNF-alpha production. These results provided an interesting clue for designing new compounds which could have better potential therapeutic implications for various neurodegenerative diseases.

Inhibitory activity of isoflavones of Pueraria flowers on nitric oxide production from lipopolysaccharide-activated primary rat microglia

Microglial activation plays an important role in alcohol-induced neuroinflammation. In search for natural medicines that may be of therapeutic potential for alcoholism, two new natural isoflavone glycosides, 6-hydroxybiochanin A-6,7-di-O-β-d-glucopyranoside (1) and 6-hydroxygenistein-7-O-β-d-glucopyranoside (2), were isolated from the ethanolic extract of the flowers of Pueraria thomsonii Benth., together with the seven known isoflavones, genistein (3), tectorigenin (4), irisolidone (5), genistin (7), tectoridin (8), tectorigenin-7-O-β-d-xylosyl-(1 → 6)-β-d-glucopyranoside (9), and 6-hydroxygenistein-6,7-di-O-β-d-glucopyranoside (11). Moreover, gehuain (6) and kakkalide (10) were obtained from the flowers of Pueraria lobata (Willd.) Ohwi. The structures of the new compounds were elucidated by UV, IR, HR-MS, and 1D and 2D NMR spectroscopic methods. Compounds 3–5 substantially inhibited the lipopolysaccharide-induced nitric oxide release from primary cultured rat cortical microglia (IC50: 1.3–9.3 μM). The inhibitory effects of compounds 6, 8, 9, and 11 (IC50: 38–62 μM) were significant but weaker than the above aglycones. However, compounds 1, 2, 7, and 10 showed little inhibitory activity. With regard to the structure–activity relationships of the isoflavonoids for the inhibition of microglial activation, the glycosylation at the C-7 hydroxyl group reduces the inhibitory activity. The methoxylation of 4′-hydroxyl group of 7-glycosylated isoflavonoids reduces the inhibitory activity, while the methoxyl group at the 6-position enhances the activity. The results suggest that isoflavonoids of Pueraria flowers may be of therapeutic potential in alcoholism related to microglial activation.

Urinary metabolites of isorhynchophylline in rats and their neuroprotective activities in the HT22 cell assay.

Isorhynchophylline is one of the major alkaloids from the Uncaria hook possessing the effects of lowered blood pressure, vasodilatation and protection against ischemia-induced neuronal damage. However, the metabolic pathway of isorhynchophylline has not been fully reported yet. In this paper, the metabolism of isorhynchophylline was investigated in rats. Five metabolites were isolated by using solvent extraction and repeated chromatographic methods, and identified by spectroscopic methods including UV, MS, NMR and CD experiments. Three new compounds were identified as 5-oxoisorhynchophyllic acid-22-O-β-D-glucuronide (M1), 17-O-demethyl-16,17-dihydro isorhynchophylline (M2) and 5-oxoisorhynchophyllic acid (M4) together with two known compounds isorhynchophylline (M0) and rhynchophylline (M3). Possible metabolic pathways of isorhynchophylline are proposed. Furthermore, the activity assay for all the metabolites showed that isorhynchophylline (M0) exhibited potent neuroprotective effects against glutamate-induced HT22 cell death. However, little or weak neuroprotective activities were observed for M1-M4. Our present study is important to further understand its metabolic fate and disposition in humans.

Alkaloids from the hook-bearing branch of Uncariarhynchophylla and their neuroprotective effects against glutamate-induced HT22 cell death.

One new alkaloid, 4-geissoschizine N-oxide methyl ether (1), was isolated from the EtOH extract of the hook-bearing branch of Uncariarhynchophylla, together with 10 known alkaloids, 3-epi-geissoschizine methyl ether (2) isolated from U.rhynchophylla for the first time, geissoschizine methyl ether (3), 4-hirsuteine N-oxide (4), hirsuteine (5), hirsutine (6), 3α-dihydro-cadambine (7), 3β-isodihydro-cadambine (8), cadambine (9), strictosamide (10), and akuammigine (11). The structures were elucidated by spectroscopic methods including UV, ESI-QTOF MS, NMR, and circular dichroism experiments. Neuroprotective effects of 1–9 were investigated against 3 mM glutamate-induced HT22 cell death. The activity assay showed that 2, 3, 5, and 6 exhibited potent neuroprotective effects against glutamate-induced HT22 cell death. However, only weak neuroprotective activities were observed for 1, 4, 7, 8, and 9.

Metabolic profile of irisolidone in rats obtained by ultra-high performance liquid chromatography/quadrupole time-of-flight mass spectrometry.

Irisolidone, a major isoflavone found in Pueraria lobata flowers, exhibits a wide spectrum of bioactivities, while its metabolic pathway in vivo has not been investigated. In this study, an ultra-high performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UHPLC/Q-TOF MS) method was employed to investigate the in vivo metabolism of irisolidone in rats. Plasma, bile, urine, and feces were collected from rats after a single 100mg/kg oral dose of irisolidone. Protein precipitation, solid phase extraction (SPE) and ultrasonic extraction were used to prepare samples of plasma, bile/urine, and feces, respectively. A total of 46 metabolites were detected and tentatively identified based on the mass spectral fragmentation patterns, elution order or confirmed using available reference standards. The metabolic pathways of irisolidone in rats included decarbonylation, reduction, demethylation, demethoxylation, dehydroxylation, hydroxylation, sulfation, and glucuronidation. The relative content of each metabolite was also determined to help understand the major metabolic pathways of irisolidone in rats.

Pharmacokinetics of kakkalide and its main metabolites in rat plasma determined by HPLC-DAD and LC–MSn

This study was undertaken to assess the plasma pharmacokinetic profile of kakkalide (KA), the major isoflavone found in extracts from the dried flower of Pueraria lobata. The main metabolites were identified using HPLC-DAD or LC/MS/MS method, and a HPLC-UV method for simultaneous quantification of the metabolites as well as the parent compound in plasma was developed. Rat plasma contained three glucuronide metabolites, irisolidone-7-O-glucuronide (Ir-7G), tectorigenin-7-O-glucuronide (Te-7G) and 6-OH biochanin A-glucuronide (6-OH BiA-G), as well as KA and trace amount of irisolidone (Ir) after oral administration of 200 mg/kg KA. The pharmacokinetics of KA and three glucuronide conjugates in rat plasma was determined for the first time using a simple, selective and accurate HPLC method. The AUC(0-t) values of the glucuronide metabolites are significantly greater than that of KA. They were detectable in rat plasma at different time points, indicating that glucuronidation during KA metabolism in vivo may occur in different sites, first in intestine and then in liver. Moreover, enterohepatic recirculation may result in the slow elimination of these glucuronide metabolites.