Kalopanax septemlobus: its phytochemistry, pharmacology and toxicity (1966-2022).

Kalopanax septemlobus is a traditional herbal medicine for multiple medicinal sites (root, stem bark, bark, leaves) in East Asia, and its bark has a significant curative effect on rheumatoid arthritis. In the past 13 years (2009-2022), the research literature accounted for 50% of the total, and it is becoming a research highlight of the relevant international scholars (ACS, ScienceDirect, PubMed, Springer, and Web of Science). This paper is the first comprehensive review of its chemistry, pharmacology, and toxicity for more than half a century (1966-2022), in which the chemical studies include triterpenoids & saponins (86 compounds), and phenylpropanoids (26 compounds), involving 46 new structures and one biomarker-triterpenoid saponin (Kalopanaxsaponin A); According to the number of literature, the pharmacological effects and mechanisms are systematically divided into five aspects, such as: anti-inflammatory, anti-tumor, antioxidant, antifungal and anti-diabetic, etc., covering its toxicological progress. To provide literature support for the exploration of new drugs against related diseases, such as rheumatoid arthritis, which are becoming younger nowadays.

Chondroprotective Effects of a Standardized Extract (KBH-JP-040) from Kalopanax pictus, Hericium erinaceus, and Astragalus membranaceus in Experimentally Induced In Vitro and In Vivo Osteoarthritis Models.

The aim of this study was to investigate the chondroprotective effect of a standardized extract (KBH-JP-040) of the Korean traditional herbs Kalopanax pictus Castor-Aralia, Hericium erinaceus (Bull.) Persoon, and Astragalus membranaceus Schischkin on in vivo and in vitro osteoarthritis (OA) models. Cultured rat chondrocytes were pre-treated with KBH-JP-040 (50, 100 and 200 µg/mL) for 1 h, then recombinant human IL-1α (rhIL-1α) for 24 h. For the in vivo model, rabbits (n = 60) were equally divided into experimental groups: normal control (NC), a collagenase-induced OA group, and OA groups treated with KBH-JP-040 (75, 100, and 150 mg/kg body weight) and celecoxib (Cx, 100 mg/kg) orally for 28 days. Treatment with KBH-JP-040 significantly attenuated inflammatory cytokines and matrix metalloproteinases (MMPs), suppressed the expression of IκBα, NF-κB, and JNK/p38 mitogen-activated protein (MAP) kinase, and upregulated aggrecan and collagen type-II expression in rhIL-1α-stimulated chondrocytes. Furthermore, the serum and synovial levels of inflammatory cytokines of rabbits also decreased in the treatment groups when compared with the OA group. Improved magnetic resonance imaging and histopathological findings further confirmed the therapeutic efficacy of KBH-JP-040 against OA. In conclusion, these results indicate that KBH-JP-040 possesses chondroprotective effects, suppressing inflammation and MMPs, and downregulating IκBα, NF-κB, and JNK/p38 MAP kinase-signaling pathways. This might be a potential therapeutic candidate for OA treatment.

Transcriptomic Analysis of Kalopanax septemlobus and Characterization of KsBAS, CYP716A94 and CYP72A397 Genes Involved in Hederagenin Saponin Biosynthesis.

Kalopanax septemlobus, commonly named the castor aralia tree, is a highly valued woody medicinal tree belonging to the family Araliaceae. Kalopanax septemlobus contains approximately 15 triterpenoid saponins primarily constituted of hederagenin aglycones. Hederagenin is a representative precursor for hemolytic saponin in plants. In the present study, transcriptome analysis was performed to discover genes involved in hederagenin saponin biosynthesis in K. septemlobus. De novo assembly generated 82,698 unique sequences, including 17,747 contigs and 64,951 singletons, following 454 pyrosequencing. Oxidosqualene cyclases (OSCs) are enzymes that catalyze the formation of diverse triterpene skeletons from 2,3-oxidosqualene. Heterologous expression of an OSC sequence in yeast revealed that KsBAS is a ß-amyrin synthase gene. Cytochrome P450 genes (CYPs) make up a supergene family in the plant genome and play a key role in the biosynthesis of sapogenin aglycones. In total, 95 contigs and 110 singletons annotated as CYPs were obtained by sequencing the K. septemlobus transcriptome. By heterologous expression in yeast, we found that CYP716A94 was ß-amyrin 28-oxidase involved in oleanolic acid production from ß-amyrin, and CYP72A397 was oleanolic acid 23-hydroxylase involved in hederagenin production from oleanolic acid. Engineered yeast co-expressing KsBAS, CYP716A94 and CYP72A397 produced hederagenin. Kalopanax septemlobus CYP72A397 is a novel CYP enzyme that synthesizes hederagenin aglycone from oleanolic acid as a single product. In conclusion, we characterized three genes participating in sequential steps for hederagenin biosynthesis from ß-amyrin, which are likely to play a major role in hederagenin saponin biosynthesis in K. septemlobus.

Kalopanacis Cortex extract-capped gold nanoparticles activate NRF2 signaling and ameliorate damage in human neuronal SH-SY5Y cells exposed to oxygen-glucose deprivation and reoxygenation.

Recently, environment-friendly synthesis of gold nanoparticles (GNPs) has been extensively explored by biologists and chemists. However, significant research is still required to determine whether "eco-friendly" GNPs are beneficial to human health and to elucidate the molecular mechanisms of their effects on human cells. We used human neuronal SH-SY5Y cells to show that treatment with Kalopanacis Cortex extract-capped GNPs (KC-GNs), prepared via an eco-friendly, fast, one-pot synthetic route, protected neuronal cells against oxygen-glucose deprivation/reoxygenation (OGD/R)-induced damage. To prepare GNPs, Kalopanacis Cortex was used without any chemical reducing and stabilizing agents. Ultraviolet-visible spectroscopy showed maximum absorbance at 526 nm owing to KC-GN surface plasmon resonance. Hydrodynamic size (54.02±2.19 nm) and zeta potential (-20.3±0.04 mV) were determined by dynamic light scattering. The average diameter (41.07±3.05 nm) was determined by high-resolution transmission electron microscopy. Energy-dispersive X-ray diffraction spectroscopy and X-ray diffraction confirmed the presence of assembled GNPs. Fourier transform infrared analysis suggested that functional groups such as O-H, C-C, and C-N participated in KC-GN formation. Cell viability assays indicated that KC-GNs restored the viability of OGD/R-treated SH-SY5Y cells. Flow cytometry demonstrated that KC-GNs inhibited the OGD/R-induced reactive oxygen species production and mitochondrial membrane potential disruption. KC-GNs also inhibited the apoptosis of OGD/R-exposed cells. Western blot analysis indicated that the OGD/R-induced cellular apoptosis and simultaneous increases in the expression of cleaved caspase-3, p53, p21, and B-cell lymphoma 2-associated X protein were reversed by KC-GNs. The KC-GN-mediated protection against OGD/R-induced neurotoxicity was diminished by NRF2 and heme oxygenase-1 gene knockdowns. Collectively, these results suggested that KC-GNs exerted strong neuroprotective effects on human neuronal cells, which might be attributed to the attenuation of OGD/R-induced neuronal cell injury through the NRF2 signaling pathway.

Differences in drought- and freeze-induced embolisms in deciduous ring-porous plant species in Japan.

MAIN CONCLUSION: Deciduous ring-porous species in Japan shed all of their leaves under severe water stress before large vessels in earlywood are embolized, and embolization take place during winter. Water in deciduous ring-porous species is mainly conducted upward via large earlywood vessels of the current year. Water columns in large vessels are vulnerable to drought-induced and freeze stress-induced embolisms. Although a vulnerability curve can be created to estimate the hydraulic capacity of plants, it remains unclear why the loss of conductivity in potted plants of ring-porous species does not reach 100 % under severe drought stress. In this study, two deciduous ring-porous species in Japan (Kalopanax septemlobus and Toxicodendron trichocarpum) were used to explain the species-specific pattern in the water-conducting pathway of the stem. We monitored and visualized the spatial distribution of xylem embolisms in the stem of K. septemlobus saplings under drought stress and freeze stress using compact magnetic resonance imaging and cryo-scanning microscopy. In addition, we evaluated the water ascent in the stems of both species using a dye uptake method. Although embolisms of large vessels were observed under drought stress and in winter, all leaves were dropped to avoid fatal water loss after embolization of some large vessels. In contrast, all large vessels were embolized in winter. Larger-diameter vessels of latewood in T. trichocarpum tended to function in trees growing in the warm temperate zone. Thus, our results suggest that the unclear curve may be derived from a discrepancy between leaf water potential and actual water potential in the xylem under severe drought stress. The frequency of xylem embolisms in deciduous ring-porous species in Japan mainly depends on the number of freeze-thaw cycles.

Ethanol extract of Kalopanax septemlobus leaf induces caspase-dependent apoptosis associated with activation of AMPK in human hepatocellular carcinoma cells.

The Kalopanax septemlobus leaf (Thunb.) Koidz. has been used as a traditional medicine herb for the treatment of various human diseases for hundreds of years. In this study, we investigated the mechanism underlying the inhibitory effects of an ethanol extract of K. septemlobus leaf (EEKS) on proliferation of HepG2 hepatocellular carcinoma cells. For this study, cell viability and apoptosis were evaluated using the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay, DAPI (4,6-diamidino-2-phenylindole) staining, agarose gel electrophoresis, and flow cytometry. Measurements of the mitochondrial membrane potential (MMP), caspase activity assays and western blots were conducted to determine whether HepG2 cell death occurred by apoptosis. Treatment of HepG2 cells with EEKS concentration-dependently reduced cell survival while significantly increasing the ratio of apoptotic cells. EEKS treatment increased the levels of the death receptors (DRs), DR4 and DR5, and activated caspases, as well as promoting proteolytic degradation of poly(ADP-ribose)-polymerase associated with the downregulation of protein expression of members of the inhibitor of apoptosis protein family. Treatment with EEKS also caused truncation of Bid, translocation of pro-apoptotic Bax to the mitochondria, and loss of mitochondrial membrane permeabilization, thereby inducing the release of cytochrome c into the cytosol. However, treatment of HepG2 cells with a pan-caspase inhibitor reversed EEKS-induced apoptosis and growth suppression, indicating that EEKS appears to induce apoptosis though a caspase-dependent mechanism involving both intrinsic and extrinsic apoptotic pathways. In addition, the phosphorylation level of AMP-activated protein kinase (AMPK) was elevated when cells were exposed to EEKS. A specific inhibitor for AMPK attenuated the EEKS-induced activation of caspases, and consequently prevented the EEKS-induced apoptosis and reduction in cell viability. Overall, our findings suggest that EEKS inhibits the growth of HepG2 cells by inducing AMPK-mediated caspase-dependent apoptosis, suggesting the potential therapeutic application of EEKS in the treatment or prevention of cancers.

Biological synthesis of manganese dioxide nanoparticles by Kalopanax pictus plant extract.

Manganese dioxide (MnO2) nanoparticles were synthesised by the reduction of potassium permanganate (KMnO4) using Kalopanax pictus leaf extract at room temperature. A transparent dark-brown colour appeared after the addition of K. pictus leaf extract to the solution of permanganate. The time course of the reduction of KMnO4and synthesis of MnO2 nanoparticles was monitored by means of UV-Vis spectra. The reduction of KMnO4occurred after addition of plant extract with disappearance of KMnO4specific peaks and emergence of peak specific for MnO2nanoparticles. MnO2nanoparticles showed absorption maxima at 404 nm. The electron dispersive X-ray spectroscopy analyses confirmed the presence of Mn and O in the sample. X-ray photoelectron spectroscopy revealed characteristic binding energies for MnO2nanoparticles. Transmission electron microscopy micrographs revealed presence of uniformly dispersed spherical shaped particles with average size of 19.2 nm. The selected area electron diffraction patterns revealed the crystalline nature of MnO2nanoparticles. Fourier transform-infrared spectroscopy spectra of pure MnO2show the occurrence of O-Mn-O vibrational mode at around 518 cm⁻¹. The phyto-synthesised MnO2nanoparticles showed degradation ability of dyes (congo red and safranin O) similar to chemically synthesised MnO2nanoparticles. This study shows simple and eco-friendly synthesis of MnO2nanoparticles by plant extract and their utilisation for dye degradation for the first time.

Protective Effect of Liriodendrin Isolated from Kalopanax pictus against Gastric Injury

In this study, we investigated the inhibitory activities on gastritis and gastric ulcer using liriodendrin which is a constituent isolated from Kalopanax pictus. To elucidate its abilities to prevent gastric injury, we measured the quantity of prostaglandin E2 (PGE2) as the protective factor, and we assessed inhibition of activities related to excessive gastric acid be notorious for aggressive factor and inhibition of Helicobacter pylori (H. pylori) colonization known as a cause of chronic gastritis, gastric ulcer, and gastric cancer. Liriodendrin exhibited higher PGE2 level than rebamipide used as a positive control group at the dose of 500 microM. It was also exhibited acid-neutralizing capacity (10.3%) and H+/K+-ATPase inhibition of 42.6% (500 microM). In pylorus-ligated rats, liriodendrin showed lower volume of gastric juice (4.38 +/- 2.14 ml), slightly higher pH (1.53 +/- 0.41), and smaller total acid output (0.47 +/- 0.3 mEq/4 hrs) than the control group. Furthermore liriodendrin inhibited colonization of H. pylori effectively. In vivo test, liriodendrin significantly inhibited both of HCl/EtOH-induced gastritis (46.9 %) and indomethacin-induced gastric ulcer (46.1%). From these results, we suggest that liriodendrin could be utilized for the treatment and/or protection of gastritis and gastric ulcer.

Potential of Kalopanax septemlobus leaf extract in synthesis of silver nanoparticles for selective inhibition of specific bacterial strain in mixed culture.

Silver nanoparticles (AgNPs) were synthesised using Kalopanax septemlobus plant leaf extracts. UV-visible spectrophotometric, Fourier-transform infrared, electron dispersive X-ray spectroscopy, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) analyses confirmed synthesis of AgNPs. TEM micrographs revealed presence of well-dispersed AgNPs predominantly of small size and different shapes with an average particle size of 30.8 nm. Antimicrobial susceptibility tests of AgNP treatments revealed variability in sensitivity of bacteria Bacillus cereus and Saccharophagus degradans under study. Minimum inhibitory concentration (MIC) values of the AgNPs for B. cereus and S. degradans were found to be 30 and 10 µg/mL, respectively. The mixed culture of B. cereus and S. degradans treated with AgNPs at 10 µg/mL showed increase in growth with time, suggesting survival of bacteria in liquid media. The plating of mixed culture before AgNP treatment showed presence of both bacteria, but 24-h-old mixed culture treated with AgNPs at the concentration of 10 µg/mL showed presence of B. cereus colonies. SEM micrographs revealed damage to S. degradans cells but no effect on B. cereus cells after AgNP treatment. Confocal microscopic observations of AgNP-treated mixed cultures by Nile blue A staining indicated intact polyhydroxyalkanoates producing flourescent cells of B. cereus but damage and deformities in S. degradans cells. This study suggests that AgNPs can selectively inhibit growth of S. degradans and retain B. cereus at MIC of S. degradans. This report is a case study for selective inhibition of one bacteria and growth of the other in a culture using plant-synthesized silver nanoparticles.

Quantification and identification of bioactive metabolites from Kalopanacis Cortex by HPLC with evaporative light scattering detection and ESI quadrupole TOF MS.

A method based on HPLC coupled with an evaporative light scattering detection and ESI quadrupole TOF MS was established for the quantification and identification of phenolics and triterpene saponins in Kalopanacis Cortex using a gradient elution of acetonitrile with 0.1% formic acid and water with 0.1% formic acid on an RP C18 column (4.6 × 250 mm, 5 µm). Diverse validation parameters, such as the linearity, LOD and LOQ, accuracy, precision, repeatability, and stability, were successfully obtained. Additionally, the efficiencies of different extraction methods were compared. The developed method was applied for the quantitative analysis of twelve representative metabolites in 61 Kalopanacis Cortex samples. The quantitation results showed that coniferin, kalopanaxsaponin C, septemlosides II, III, C, and D exhibited distinct regional patterns in Kalopanacis Cortex samples. These six compounds including one new triterpene saponin show potential as marker compounds for evaluating the quality of Kalopanacis Cortex and the geographical variation in its chemical composition.

Inhibitory effects of oleanane-type triterpenes and saponins from the stem bark of Kalopanax pictus on LPS-stimulated pro-inflammatory cytokine production in bone marrow-derived dendritic cells.

Kalopanax pictus (Araliaceae) is a deciduous tree distributed in Korea, Japan, and China. The stem bark of K. pictus has been functionally used as a traditional crude drug for the treatment of various inflammatory diseases. In the present study, we describe the inhibitory effects of oleanane-type triterpenes and saponins isolated from the stem bark of K. pictus on production of pro-inflammatory cytokines in LPS-stimulated bone marrow-derived dendritic cells. Of the compounds tested, 16,23,29-trihydroxy-3-oxo-olean-12-en-28-oic acid (1), 4,23,29-trihydroxy-3,4-seco-olean-12-en-3-oate-28-oic acid (2), 3ß,6ß,23-trihydroxyolean-12-en-28-oic acid 28-O-ß-D-glucopyranoside (3), nipponogenin E (6), 3ß,6ß,23-trihydroxyolean-12-en-28-oic acid (7), and caulophyllogenin (19) significantly inhibited the production of IL-12 p40 and IL-6 with IC50 values ranging from 3.3 to 9.1 µM. Compounds 2, 3, 7, and 19 significantly suppressed the secretion of TNF-α with IC50 ranging from 8.8 to 20.0 µM. These data provide scientific support for the use of K. pictus stem bark and its triterpene and saponin components in the inhibition of pro-inflammatory cytokine secretion, including IL-12 p40, IL-6, and TNF-α, and for prevention and treatment of inflammatory diseases.

Kalopanaxsaponin A Exerts Anti-Inflammatory Effects in Lipopolysaccharide-Stimulated Microglia via Inhibition of JNK and NF-kappaB/AP-1 Pathways

Microglial activation plays an important role in the development and progression of various neurological disorders such as cerebral ischemia, multiple sclerosis, and Alzheimer's disease. Thus, controlling microglial activation can serve as a promising therapeutic strategy for such brain diseases. In the present study, we showed that kalopanaxsaponin A, a triterpenoid saponin isolated from Kalopanax pictus, inhibited inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and tumor necrosis factor (TNF)-alpha expression in lipopolysaccharide (LPS)-stimulated microglia, while kalopanaxsaponin A increased anti-inflammatory cytokine interleukin (IL)-10 expression. Subsequent mechanistic studies revealed that kalopanaxsaponin A inhibited LPS-induced DNA binding activities of NF-kappaB and AP-1, and the phosphorylation of JNK without affecting other MAP kinases. Furthermore, kalopanaxsaponin A inhibited the intracellular ROS production with upregulation of anti-inflammatory hemeoxygenase-1 (HO-1) expression. Based on the previous reports that JNK pathway is largely involved in iNOS and proinflammatory cytokine gene expression via modulating NF-kappaB/AP-1 and ROS, our data collectively suggest that inhibition of JNK pathway plays a key role in anti-inflammatory effects of kalopanaxsaponin A in LPS-stimulated microglia.

Cryopreservation of Kalopanax septemlobus embryogenic callus using vitrification and droplet-vitrification.

A cryopreservation protocol has been developed for embryogenic callus cultures of castor aralia (Kalopanax septemlobus), a deciduous tree which is widely used in oriental medicine and in landscape design. Three preculture treatments, four loading and six vitrification solutions were tested in a vitrification procedure. Preculture of embryogenic callus (EC) with high sucrose concentrations (up to 0.7 M) showed no effect on regrowth after cryopreservation. Loading for 20 min at ambient temperature improved regrowth of cryopreserved EC by 70-75 percent compared with non-loaded samples, regardless of the composition of the loading solution. Among vitrification solutions, the highest regrowth of 95-100 percent after cryopreservation was obtained after incubation of EC in a vitrification solution A3-80 percent comprising (w/v) 33.3 percent glycerol + 13.3 percent DMSO + 13.3 percent EG + 20.1 percent sucrose for 40 min at 0°C. Profiling of crystallization and recrystallization events using differential scanning calorimetry (DSC) confirmed that freezing injury was minimized in samples after loading and cryoprotection with this vitrification solution. Unlike many other papers, the droplet-vitrification protocol did not produce higher post-cryopreservation regrowth of Kalopanax EC, compared with the vitrification procedure. When samples are sufficiently cryoprotected during VS treatment, vitrification using cryovials may be preferred, since droplet-vitrification is more complex and requires skilled personnel. Cryopreserved callus grew rapidly and produced numerous somatic embryos, which developed similarly to embryos obtained from non-cryopreserved samples.

'Personalisation' of droplet-vitrification protocols for plant cells: a systematic approach to optimising chemical and osmotic effects.

Although an appropriate cryopreservation protocol is a prerequisite for basic studies and large-scale implementation as well as further cryopreservation studies, the process relies on trial and error. Among the vitrification-based cryopreservation techniques, droplet-vitrification produces high post-cryopreservation recovery. However, the protocol itself cannot solve the problems engaged in plant cryopreservation, prominently due to dehydration with cytotoxic vitrification solutions. This paper proposes a set of treatments to develop droplet-vitrification using a standard procedure associated with additional treatments and alternative vitrification solutions. The proposed standard protocol consists of a progressive preculture with 0.3 M sucrose for 31 h and with 0.7 M for 17 h, loading with vitrification solution C4-35% (17.5 percent glycerol + 17.5 percent sucrose, w/v) for 20 to 40 min, dehydration with vitrification solutions A3-90 percent (37.5 percent glycerol + 15% DMSO + 15 percent EG + 22.5 percent sucrose) for 10 to 30 min or B1-100 percent (PVS3) for 40 to 120 min at room temperature, cooling the samples using aluminum foil strips, rewarming by plunging into pre-heated (40 degree C) unloading solution (0.8 M sucrose) and further unloading for 20 to 60 min, depending on size and permeability of the materials. Using this systematic approach we can identify whether the material is tolerant or sensitive to chemical toxicity and to the osmotic stress of dehydration with vitrification solutions, thus revealing which is the main barrier in solution-based vitrification methods. Based on the sensitivity of samples we can design a droplet-vitrification procedure, i.e. preculture, loading, dehydration with vitrification solutions, cooling and rewarming. Using this approach, the development of appropriate droplet-vitrification protocol is facilitated.

Climate oscillation during the Quaternary associated with landscape heterogeneity promoted allopatric lineage divergence of a temperate tree Kalopanax septemlobus (Araliaceae) in East Asia.

We investigated the biogeographic history of Kalopanax septemlobus, one of the most widespread temperate tree species in East Asia, using a combined phylogeographic and palaeodistribution modelling approach. Range-wide genetic differentiation at nuclear microsatellites (G'(ST) = 0.709; 2205 samples genotyped at five loci) and chloroplast DNA (G(ST) = 0.697; 576 samples sequenced for 2055 bp at three fragments) was high. A major phylogeographic break in Central China corresponded with those of other temperate species and the spatial delineation of the two temperate forest subkingdoms of East Asia, consistent with the forests having been isolated within both East and West China for multiple glacial-interglacial cycles. Evidence for multiple glacial refugia was found in most of its current range in China, South Japan and the southernmost part of the Korean Peninsula. In contrast, lineage admixture and absence of private alleles and haplotypes in Hokkaido and the northern Korean Peninsula support a postglacial origin of northernmost populations. Although palaeodistribution modelling predicted suitable climate across a land-bridge extending from South Japan to East China during the Last Glacial Maximum, the genetic differentiation of regional populations indicated a limited role of the exposed sea floor as a dispersal corridor at that time. Overall, this study provides evidence that differential impacts of Quaternary climate oscillation associated with landscape heterogeneity have shaped the genetic structure of a wide-ranging temperate tree in East Asia.

Involvement of the p38 MAPK and ERK signaling pathway in the anti-melanogenic effect of methyl 3,5-dicaffeoyl quinate in B16F10 mouse melanoma cells.

Methyl 3,5-dicaffeoyl quinate (MDQ), an active compound present in Kalopanax pictus, Salicornia herbacea L., Aster oharai and Solidago virga-aurea var. gigantean, is a dicaffeoylquinic acid derivative esterified by methanol. Recent studies have revealed that MDQ possesses multiple pharmacological activities, such as antitumor, antioxidative and cytoprotective activities. To date, there has been no attempt to test the action of MDQ in melanocytes. In this study, we investigated the effect of MDQ on melanogenesis in B16F10 mouse melanoma cells. MDQ inhibited melanin production and tyrosinase activity in B16F10 mouse melanoma cells without a direct inhibitory effect on mushroom tyrosinase activity. Furthermore, we also found that MDQ decreased protein expression levels of microphthalmia-associated transcription factor (MITF) and tyrosinase in B16F10 melanin cells. Meanwhile, phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK) was significantly reduced after 6h MDQ treatment, and this expression recovered at 48 h. The phosphorylation of extracellular signal-regulated kinase (ERK) was significantly enhanced at 12-48 h, whereas no effect was observed in the phosphorylation of Akt. In addition, MDQ treatment did not significantly alter the expression levels of total p38 MAPK, ERK, and Akt. Thus, it seems that inhibition of phospho-p38 MAPK and activation of phospho-ERK may lead to the suppression of melanogenesis in MDQ-treated B16F10 mouse melanoma cells.

Kalopanaxsaponins A and B isolated from Kalopanax pictus ameliorate memory deficits in mice.

The stem-bark of Kalopanax pictus (KP, family Araliaceae), which contains triterpenoid saponins, has been shown to exhibit anticarcinogenic, antiinflammatory, antirheumatoid and antidiabetic activities. In a preliminary study, a KP methanol extract demonstrated acetylcholinesterase activity in vitro and memory enhancement in scopolamine-treated mice. Therefore, we isolated acetylcholinesterase inhibitors, kalopanaxsaponins A and B, from a KP butanol (BuOH) fraction, measured acetylcholinesterase activity in vitro, and investigated their memory-enhancing effects in a passive avoidance test, Y-maze test and Morris water maze test. These constituents inhibited acetylcholinesterase activity and significantly reversed scopolamine-induced deficits. They also increased brain-derived neurotrophic factor (BDNF) and phosphorylated cAMP response element binding (p-CREB) protein expression but reduced TNF-α increased by scopolamine. Based on these findings, kalopanaxsaponins A and B may ameliorate memory deficits by inhibiting acetylcholinesterase activity and inducing BDNF and p-CREB expression.

Effect of triterpenes and triterpene saponins from the stem bark of Kalopanax pictus on the transactivational activities of three PPAR subtypes.

Kalopanax pictus (Araliaceae) is a deciduous tree that grows in East Asian countries. Its stem bark and leaves have been used in traditional medicine to treat rheumatic arthritis, neurotic pain, and diabetes mellitus. A phytochemical study on a methanol extract of the stem bark of K. pictus resulted in the isolation of three new compounds, 6ß,16α-dihydroxy-hederagenin 3-O-ß-D-glucuronopyranoside (1), 3-O-ß-D-glucuronopyranosyl-28-O-ß-D-glucopyranosyl-6ß,16α-dihydroxy-oleanolic acid (2), and 3-O-ß-D-galactopyranosyl(1→3)-α-L-arabinopyranosyl hederagenin 28-O-ß-D-glucopyranosyl-(1→6)-ß-D-glucopyranosyl ester (3), along with eight known compounds (4-11). Their structures were established on the basis of chemical and spectroscopic methods (IR, 1D and 2D NMR, and HRESITOFMS). Compounds 1-6 and 8-10 upregulated PPARs transcriptional activity in a dose-dependent manner in HepG2 cells, with EC(50) values in the range 0.20-15.5 µM. Moreover, the specific PPAR transactivational effects of compounds 1-6 and 8-10 on separate PPAR subtypes, PPARα, -γ, and -ß(δ) were further investigated. Compounds 4, 5, 8, and 10 showed significant PPARα transactivational activity, with EC(50) values of 7.8, 8.0, 10.3, and 17.3 µM, respectively. Compounds 2, 4, 6, and 8-10 exhibited PPARγ dose-dependent transactivational activity, with EC(50) values of 14.7, 15.5, 14.8, 10.9, 17.1, and 16.3 µM, whereas compounds 8 and 10 significantly upregulated PPARß(δ) transcriptional activity, with EC(50) values of 15.7 and 17.7 µM, respectively.

[Studies on the chemical constituents from the roots of Kalopanax septemlobus].

OBJECTIVE: To investigate the chemical constituents of Kalopanax septemlobus. METHODS: Chromatographic techniques including silica gel, gel, semi-preparative HPLC and PTLC as well as recrystallization were employed in the isolation and purification, and the structures were elucidated by spectral analysis and physical and chemical properties. RESULTS: 6 compounds were identified as liriodendrin (1), (-) -syringarenol (2), trans-coniferyl aldehyde (3), trans-caffeic acid (4), beta-daucosterol (5), beta-sitosterol (6). CONCLUSION: Compounds 2 -5 are obtained from this genus for the first time.

Direct screening of G-quadruplex ligands from Kalopanax septemlobus (Thunb.) Koidz extract by high performance liquid chromatography.

G-quadruplex DNA structure is considered to be a very attractive target for antitumor drug design due to its unique role in maintaining telomerase activities. Therefore, discovering ligands with high stability of G-quadruplex structure is of great interest. In this paper, high-performance liquid chromatography (HPLC) was used for fast screening of G-quadruplex ligands from the crude extract of Kalopanax septemlobus (Thunb.) Koidz, a traditional Chinese medicine. Four potent G-quadruplex ligands were firstly selected through HPLC by comparing the peak profiles and absorption intensity of the crude sample before and after interaction with G-quadruplex DNA. Then the target compounds were isolated and purified by high-speed countercurrent chromatography (HSCCC) for further confirmation of their stabilities of G-quadruplex by temperature-dependent circular dichroism (CD). Four compounds were isolated and identified as 2,4-dihydroxybenzoic acid (I), chlorogenic acid (II), caffeic acid (III) and 5-feruloylquinic acid (IV) each by MS and NMR. Finally, compound I, II, III were each proved to be potent G-quadruplex ligands by decreasing the peak intensity in HPLC chromatogram after complexation with G-quadruplex, which stabilize G-quadruplex by 7±2 °C, 10±2 °C, and 3±2 °C respectively, based on CD analyses. However, compound IV showed no G-quadruplex stability. The decrease of peak absorption intensity in HPLC chromatogram is the most important signal to find G-quadruplex ligands. This provides a very promising strategy for fast screening G-quadruplex ligands from natural plant extracts.