Potential Antiaging and Hepatoprotective Effects of Acanthopanax senticosus Extracts in Adult Rat Models.

Data regarding plant extracts with antiaging properties, particularly through the biological process involving telomeres and telomerase, are limited. Thus, this study aimed to investigate the effects of Acanthopanax senticosus extract (ASE) supplementation on leukocyte telomere length (LTL), telomerase, and inflammatory and metabolic markers in adult animal models. A freeze-dried product of ethanol extracts was prepared using a mixture product of stem and root ASE. In a 24-week experiment that included 24-week-old Sprague Dawley male rats, experimental rats (n = 10) were administrated with 7 mg/day of ASE dissolved in saline and control rats (n = 10) with saline. All rats had access to chow and tap water ad libitum. Their LTL and plasma levels of telomerase and inflammatory and metabolic markers were assayed and compared between the two groups. The experimental rats showed significantly longer LTL (p < 0.05) and lower plasma levels of alanine aminotransferase (p < 0.05) and aspartate aminotransferase (p = 0.08) compared with the control. In addition, LTL was correlated with the aforementioned biochemical parameters of liver function test among experimental rats only. No significant differences in plasma levels of telomerase and inflammatory and metabolic markers were observed. These findings indicate that ASE supplementation may attenuate LTL shortening and reduce liver biochemical parameters, indicating its potential antiaging and hepatoprotective effects without any adverse metabolic response.

Adsorption and Desorption Characteristics of Total Flavonoids from Acanthopanax senticosus on Macroporous Adsorption Resins.

We examined the application of six different resins with the aim of selecting a macroporous resin suitable for purifying Acanthopanax senticosus total flavonoids (ASTFs) from Acanthopanax senticosus crude extract (EAS) by comparing their adsorption/desorption capacities, which led to the selection of HPD-600. Research on the adsorption mechanism showed that the adsorption process had pseudo-second-order kinetics and fit the Freundlich adsorption model. Moreover, the analysis of thermodynamic parameters indicated that the adsorption process is spontaneous and endothermic. The optimal conditions for purification of ASTFs were determined as sample pH of 3, 60% ethanol concentration, and 3 BV·h-1 flow rate, for both adsorption and desorption, using volumes of 2.5 and 4 BV, respectively. The application of macroporous resin HPD-600 to enrich ASTFs resulted in an increase in the purity of total flavonoids, from 28.79% to 50.57%. Additionally, the antioxidant capacity of ASTFs was higher than that of EAS, but both were lower than that of L-ascorbic acid. The changes in ASTFs compositions were determined using ultra-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS), with the results illustrating that the levels of seven major flavonoids of ASTFs were increased compared to that in the crude extract.

Pharmacognostic Evaluation and HPLC-PDA and HS-SPME/GC-MS Metabolomic Profiling of Eleutherococcus senticosus Fruits.

Eleutherococcus senticosus (Rupr. et Maxim.) Maxim. is a medicinal plant used in Traditional Chinese Medicine (TCM) for thousands of years. However, due to the overexploitation, this species is considered to be endangered and is included in the Red List, e.g., in the Republic of Korea. Therefore, a new source of this important plant in Europe is needed. The aim of this study was to develop pharmacognostic and phytochemical parameters of the fruits. The content of polyphenols (eleutherosides B, E, E1) and phenolic acids in the different parts of the fruits, as well as tocopherols, fatty acids in the oil, and volatile constituents were studied by the means of chromatographic techniques [HPLC with Photodiode-Array Detection (PDA), headspace solid-phase microextraction coupled to gas chromatography-mass spectrometry (HS-SPME/GC-MS)]. To the best of our knowledge, no information is available on the content of eleutherosides and phenolic acids in the pericarp and seeds. The highest sum of eleutheroside B and E was detected in the whole fruits (1.4 mg/g), next in the pericarp (1.23 mg/g) and the seeds (0.85 mg/g). Amongst chlorogenic acid derivatives (3-CQA, 4-CQA, 5-CQA), 3-CQA was predominant in the whole fruits (1.08 mg/g), next in the pericarp (0.66 mg/g), and the seeds (0.076 mg/g). The oil was rich in linoleic acid (C18:3 (n-3), 18.24%), ursolic acid (35.72 mg/g), and α-tocopherol (8.36 mg/g). The presence of druses and yellow oil droplets in the inner zone of the mesocarp and chromoplasts in the outer zone can be used as anatomical markers. These studies provide a phytochemical proof for accumulation of polyphenols mainly in the pericarp, and these structures may be taken into consideration as their source subjected to extraction to obtain polyphenol-rich extracts.

Eleutheroside K isolated from Acanthopanax henryi (Oliv.) Harms suppresses methicillin resistance of Staphylococcus aureus.

Acanthopanax (A.) henryi (Oliv.) Harms contain many bioactive compounds commonly used in traditional Chinese medicine. The objective of the present study was to investigate the antibacterial activity of the single constituent, Eleutheroside K (ETSK) isolated from the leaves of A. henryi (Oliv.) Harms, against methicillin-resistant Staphylococcus (S.) aureus (MRSA). Broth microdilution assay was used to measure the minimal inhibitory concentration (MIC) and the MIC values of ETSK against eight clinical S. aureus strains were all 50 µg ml-1 . At sub-inhibitory concentrations, a synergistic effect between oxacillin (OXA) and ETSK was confirmed using checkerboard dilution assay and time-kill curve analysis. The bacteriostatic effect became more pronounced when ETSK was used in combination with detergent (Triton X-100) or ATPase inhibitor (N, N'-dicyclohexylcarbodiimide). According to western blot analysis, the down-regulated expression of Penicillin-binding protein 2a (PBP2a) further validated that the bacterial activity was inhibited when treated with ETSK in a dose-dependent manner. Results based on our study verified that ETSK significantly suppressed MRSA infections and emphasized the potential application of ETSK as a novel anti-MRSA natural drug.

Determination of Kaurenoic Acid in Acanthopanax trifoliatus by Ultra-High Performance Liquid Chromatography Coupled with Tandem Mass Spectrometry (UHPLC-MS/MS).

Acanthopanax trifoliatus (L.) Merr. (A. trifoliatus) belongs to the family Araliaceae, which is called "Le Cai", and is an indigenous plant to Guangdong Province that has been prevalently planted for years. A. trifoliatus is used in folk medicine and has ginseng-like activity. Kaurenoic acid ((-)-kaur-16-en-19-oic acid, KA) is a kaurane-type diterpenoid that is regarded as a major compound in A. trifoliatus. Early studies have reported the determination of KA by HPLC capillary electrophoresis. However, KA could not be completely separated from other components in the plant extract by HPLC because of their similar molecular structures and physical and chemical properties. UHPLC-MS/MS could be a useful tool to identify and quantify KA. In the present work, a UHPLC-ESI-MS/MS method for determining KA in A. trifoliatus was developed and validated. KA was extracted from lyophilized A. trifoliatus leaves by ultrasound-assisted extraction and further purified by solid phase extraction (SPE). KA was quantified and separated on an Accucore C18 LC column. Mass spectrometry with multi-reaction monitoring (MRM) and quantitative fragment ion/product ion (m/z: 301.3/301.3) in ESI negative mode was used for quantification. The intra-assay and inter-assay relative standard deviation (R.S.D.) were 2.8% and 3.2%, respectively. The inter-person R.S.D. on the same day was 3.6%. The inter-instrument R.S.D. with the same model on the same day was 2.9%. The recoveries evaluated upon spiking three different concentrations of KA were above 97%. A minor matrix effect of 94% was observed. This method has been applied successfully for the determination of KA in A. trifoliatus leaves.

[Effects of fungi fraction on growth and anti-oxidative activity of Eleutherococcus senticosus].

The present study was conducted to explore the effect of endophytic fungi fraction on growth and anti-oxidative activity of Eleutherococcus senticosus. The growth,yield,contents of MDA,and antioxidant activities were assessed in E. senticosus under five fungi fractions,namely BZ,MH,DT,JS,and XFZ. The results showed that fungi fractions and component significantly affected the growth,low concentration of DT fungi fraction significantly increased the biomass of E. senticosus,reduced the MDA content in cells,and the antioxidant activities of the aqueous extracts were superior to the others. The results indicated that low concentration of DT fungi fraction was the optimum fraction to achieve high yield and quality of E. senticosus.

LC-ESI-MS/MS profiling of phenolics in the leaves of Eleutherococcus senticosus cultivated in the West Europe and anti-hyaluronidase and anti-acetylcholinestarase activities.

Neither secondary metabolites of the spring leaves nor the autumn leaves of Eleutherococcus senticosus species cultivated in Poland, or the bioactivity are known. The richest in polyphenols was the autumn leaves (171.1 mg/g DE), while in flavonoids the spring leaves (107.9 mg/g DE). Using LC-ESI-MS/MS, protocatechuic acid has been identified as the most abundant compound in the spring and autumn leaves (200 and 70 µg/g DE, respectively). Amongst flavonoids, naringenin 7-O-glucoside occurred in the largest amount (20 and 10 mg/g DE in the spring and autumn leaves, respectively). The autumn leaves inhibited Hyal the strongest (74.3%), comparing to the spring leaves (33%). A weak inhibition was found towards AChE (0.64 and 5.8% for the autumn and spring leaves, respectively). To our best knowledge, no information was available on the phytochemical composition and activity of the leaves of E. senticosus cultivated in Poland.

Structural Modification of Stilbenoids from Acanthopanax leucorrhizus and Their Cytotoxic Activity.

A new cis-stilbenoid, 1,9-dihydroxy-10-methoxy-6H-dibenzo[b,f]oxocin-6-one (2) was isolated from the AcOEt extract of the stem barks of Acanthopanax leucorrhizus, along with three known stilbenoids, 9-hydroxy-10-methoxy-6H-dibenzo[b,f]oxocin-6-one (1), 5-O-methyl-(E)-resveratrol 3-O-ß-d-glucopyranoside (3), and (E)-resveratrol 3-O-ß-d-xylopyranoside (4). Two derivatives (2a and 2b) were synthesized by the structural modification of compound 2, which exhibited certain cytotoxic activities against HT-29 and HeLa cell lines in vitro. All compounds were structurally characterized by comprehensive analysis of their spectroscopic data and comparison with literature information, and evaluated for their cytotoxic activities against three human tumor cell lines (HL-60, HT-29, and HeLa) by the standard MTT assay in vitro. The results showed that derivatives 2a and 2b exhibited strong activities than compounds 2 against HT-29 and HeLa cell lines.

Eleutherococcus senticosus (Araliaceae) Leaf Morpho-Anatomy, Essential Oil Composition, and Its Biological Activity Against Aedes aegypti (Diptera: Culicidae).

The roots of Eleutherococcus senticosus (Rupr. & Maxim.) Maxim., a well-known medicinal plant from Eastern Asia, are used worldwide for their known beneficial medicinal properties. Recently, the leaves have been used as an alternative to the roots. The present study was aimed at exploring the leaf essential oil as a potential source of compounds for mosquito management. Gas chromatography/mass spectrometry analysis of the leaf essential oil revealed 87 compounds, constituting 95.2% of the oil. α-Bisabolol (26.46%), ß-caryophyllene (7.45%), germacrene D (6.87%), ß-bisabolene (4.95%), and α-humulene (3.50%) were five of the major constituents. The essential oil was subjected to biting deterrence and repellent activity against mosquito Aedes aegypti. The biting deterrence of the oil produced a proportion not biting (PNB) value of 0.62 at 10 µg/cm2 as compared with 0.86 of control DEET (N,N-diethyl-3-methylbenzamide) at a standard dose of 25 nmol/cm2. Among individually selected compounds present in the oil (α-bisabolol, ß-caryophyllene, α-humulene, and caryophyllene oxide), only α-bisabolol produced a PNB value of 0.80, equivalent to DEET at 25 nmol/cm2, whereas the others were not repellent. The artificial mixture (AMES-1) of these four selected compounds produced a relatively high PNB value of 0.80. The repellent activity measured by minimum effective dosage (MED) for α-bisabolol and α-humulene produced MED values of 0.094 and 0.104 mg/cm2, respectively, as compared with 0.023 mg/cm2 of DEET. The leaf essential oil, the artificial mixture (AMES-1), and other binary and tertiary combinations of major compounds showed no repellent activity. In addition, morpho-anatomical features of the leaf are provided for correct identification of the species.

Phytochemical Content and Pharma-Nutrition Study on Eleutherococcus senticosus Fruits Intractum.

In the past two decades public interest in herbal products has increased significantly in Europe, especially in the plant-based products from non-European traditions. Eleutherococcus senticosus has been used for the treatment of inflammatory diseases, anemia, and rheumatoid arthritis. The Eleutherococcus senticosus fruits intractum was examined for the content of phenolic acids (LC-ESI-MS/MS), minerals (AAS), TPC, and TFC (spectrophotometric assay). The antioxidant activity was determined using free radical scavenging assay and TLC-DB-DPPH∗ dot-blot test. An anti-Hyal activity was evaluated by the spectrophotometric assay method. Cytotoxicity towards HL-60, HL-60/MX1, HL-60/MX2, CEM/C1, and CCRF/CEM leukemic cell lines was done using trypan blue test. Among eight phenolic acids, trans-caffeic acid was found in the largest amount (41.2 mg/g DE). The intractum presented a high amount of macroelements (Ca, Mg, K; 1750, 1300, and 21000 mg/kg) and microelements (Fe, Mn; 32.7, 54.3 mg/kg), respectively. The content of TPC and TFC was 130 and 92 mg/g DE, respectively. The intractum showed anti-Hyal activity (2.16-60%) and an antioxidant capacity (EC50; 52 µg/mL). The intractum most strongly inhibited the growth of HL-60, HL-60/MX1, and CCRF/CEM. A better understanding of the intractum health benefits is important in order to increase its utility and enrich dietary sources of health promoting compounds.

Toxicogenomic Effects in Rat Blood Leukocytes and Chemoprophylaxis of Radiation-Induced Carcinogenesis.

Toxicogenomic parameters were studied in the blood of female rats after exposure to ionizing γ-radiation in a dose of 4 Gy and chemoprophylaxis with α-difluoromethylornithine, eleutherococcus or leuzea extracts, which were used in animals with morphological manifestations of tumor growth under conditions of radiation-induced carcinogenesis. Life-time evaluation of toxicogenomic effects was carried out by express method for measurements of blood nucleotid DNA - fluorescent indication. The level of hyperaneu/polyploidy increased in the blood leukocytes of control rats 30 days after radiation exposure. A significant decrease of genotoxicity as a result of drug treatment in comparison with the number and multiplicity of tumors in irradiated animals was found only in the endocrine and reproductive organs of rats treated by eleutherococcus extract.

Identification of six new lupane-type triterpenoids from Acanthopanax koreanum leaves and their tyrosinase inhibitory activities.

Chemical investigation of Acanthopanax koreanum leaves resulted in the isolation of 13 compounds (1-13), including six new (20,29)-dehydrolupane-type triterpenoids: 3α,11α,30-trihydroxylup-20(29)-en-23,28-dioic acid (1), 3α,11α,30-trihydroxylup-20(29)-en-28-oic acid (2), 3α,11α,30-trihydroxylup-23-al-20(29)-en-28-oic acid (3), 3α, 11α-dihydroxy-20-oxo-30-norlupane-23,28-dioic acid (5), (20S)-3α-hydroxy-30 oxolupane-23,28-dioic acid (8), (20S)-3ß,7ß,29-trihydroxy-lupane-23-al-28-oic acid (10), and one novel compound isolated for the first time, named 3α,20α,29-trihydroxylupane-23,28-dioic acid (9), together with six known compounds (4, 6, 7, and 11-13). Chemical structures of the isolated compounds were evaluated by analyzing and comparing spectroscopic data with those reported in the literature. These compounds were also evaluated for their tyrosinase inhibitory effects. Among them, compounds 3, 7, 9, and 12 showed significant inhibitory effects, with inhibitory concentrations of 50% (IC50) values ranging from 8.61 to 63.5 µM.

Biotechnological production of eleutherosides: current state and perspectives.

Eleutherosides, the phenylpropanoid and lignan glycosides, are the active ingredients accumulated in the roots and stems of Eleutherococcus species and in Eleutherococcus senticosus in particular. Syringin (=eleutheroside B) and (-) syringaresinol-di-O-ß-D-glucoside (=eleutheroside E) appear as the most important bioactive compounds which are used as adaptogens, besides their abundant antidiabetic and anticancer properties. As the availability of "Eleuthero" is becoming increasingly limited because of its scanty natural distribution, the production of these compounds by biotechnological means has become an attractive alternative. In E. senticosus and other closely related species, Eleutherococcus sessiliflorus, Eleutherococcus chiisanensis, and Eleutherococcus koreanum, organogenic cultures have been induced for the production of eleutherosides. Bioreactor cultures have been established and various parameters, which influence on the accumulation of biomass and secondary metabolites, have been thoroughly investigated. Pilot-scale cultures have also been accomplished for the large-scale production of somatic embryos containing abundant amounts of eleutherosides. This review describes the biotechnological approaches and challenges for the production of eleutherosides.

Influence of gastrointestinal digestion and edible plant combination on oral bioavailability of triterpene saponins, using a biomimetic digestion and absorption system and determination by HPLC.

Saponins have many biological activities, but their overload could cause toxicity to the human body. Bionic gastrointestinal digestion and monolayer liposome extraction were used for oral bioavailability assessment of triterpene saponins (notoginsenoside R1, ginsenosides Rb1 and Rd1) in an edible herb (San-Chi) and its compound herbal medicine (Pien Tze Huang, PZH). The concentrations of affinity-monolayer liposome saponins in the chyme were determined by HPLC and used for oral bioavailability assessment. With the digestion of San-Chi and PZH from the stomach to the intestine, the release of saponins in their chyme was increased. The intestinal absorption ratios of N-R1, G-Rb1, G-Rd1, and total saponins from San-Chi were 86.57, 18.56, 73.30, and 40.20%, respectively, which were more than those from PZH (i.e., 19.56, 10.11, 30.11, and 16.08%). The oral bioavailability of saponins was controlled by saponin species, gastrointestinal digestion, and edible plants combination.

[Cloning of Eleutherococcus senticosus calmodulin gene and effect of endophytic fungus on expression amount of gene].

OBJECTIVE: To clone calmodulin (CaM) gene in Eleutherococcus senticosus, and study the effect of endophytic fungi on expression amount of CaM gene. METHOD: The CaM full length cDNA sequence was cloned by rapid amplification of cDNA ends (RACE). The gene was analyzed and corresponding structure and functions were predicted by the bioinformatics methods. The expression amount of CaM gene affected of endophytic fungus P116-1a, P116-1b, P1094 and P312-1 was detected by RT-PCR. RESULT: The full length of CaM cDNA was 856 bp containing an ORF of 450 bp that encoded a protein of 149 amino acids. The homologous of predicted protein was almost 100% with plants like Panax ginseng and Daucus carota. RT-PCR results showed that endophytic fungus improved CaM expression amount significantly (P<0.05). The highest expression amount of CaM occurred 90 d after reinoculated with endophytic fungi P1094, up to 2.96 times of the control. CONCLUSION: The CaM gene of E. senticosus was successfully cloned for the first time. The results demonstrated that endophytic fungus of E. senticosus improved CaM expression amount significantly.

[Effect of endophytic fungi on expression amount of key enzyme genes in saponins biosynthesis and Eleutherococcus senticosus saponins content].

OBJECTIVE: To analyze the effect of endophytic fungi on expression amount of key enzyme genes SS (squalene synthase gene), SE (squalene epoxidase gene) and bAS (beta-amyrin synthase gene) in saponin biosynthesis and saponins content in Eleutherococcus senticosus. METHOD: Wound method was used for back meeting the endophytic fungi to E. senticosus. With GAPDH as internal control gene, the expression of key enzyme genes was detected by real time PCR method. E. senticosus saponins content was measured by spectrophotometry method. RESULT: When wound method back meeting P116-1a and P116-1b after 30 d, the expression content of SS improved significantly (P < 0.05), however the back meeting of P109-4 and P312-1 didnt change the expression of SS. After that SS expression showed reduction-equality-reduction varying trend. Thirty days after back meeting P312-1, the expression content of SE improved significantly (P < 0.05). Ninty days after back meeting P116-1b and P312-1, the expression content of SE improved significantly to 130%,161%, respectively (P < 0.05). After 120 d, back meeting four endophytic fungi, the expression of SE were significantly higher than the control (P < 0.05). Back meeting four endophytic fungi form 60 d to 120 d, the expression of bAS was significantly higher than the control (P < 0.05). The back meeting four endophytic fungi improved E. senticosus saponins content significantly (P < 0.05). CONCLUSION: Endophytic fungi P116-1a, P116-1b, P1094 and P312-1 significantly effected the expression of key enzyme genes SS, SE and bAS and then affected E. senticosus saponins content. Among the genes, bAS was key target gene.

Simultaneous quantification of polyherbal formulations containing Rhodiola rosea L. and Eleutherococcus senticosus Maxim. using rapid resolution liquid chromatography (RRLC).

An RRLC method capable of simultaneous identification and rapid quantification of six biologically active compounds (salidroside, tyrosol, rosarin, rosavin, rosin, rosiridin) in Rhodiola rosea L. and two active compounds (eleutheroside B and eleutheroside E) in Eleutherococcus senticosus Maxim. was developed. The chromatographic analyses were performed on a reversed phase Phenomenex C18 (2)-HST column at 40°C with a neutral mobile phase (purified water and acetonitrile) gradient system at a flow rate of 1.0ml/min and UV detection at 205 and 220nm simultaneously. Baseline separation of eight active compounds was achieved within 8min. This developed method provides good linearity (R>0.9997), precision (RSD<1.99%) and recovery of the bioactive compounds. The RRLC method developed is capable of controlling the quality of R. rosea and E. senticosus raw herbs, commercial extracts, as well as polyherbal formulations containing R. rosea and E. senticosus as ingredients. This RRLC method is accurate and sensitive; in addition, it greatly increases sample analysis throughput with reduced analysis time, which is suitable for routine quality control analysis.

Short communication: Influence of pasteurization on the active compounds in medicinal plants to be used in dairy products.

Interest from the dairy industry in adding herbal drugs to milk and yogurt products raises the question of whether these plant materials can be pasteurized. Root material of Rhodiola rosea, Eleutherococcus senticosus, and Panax ginseng, all plants with adaptogenic activities, was pasteurized. The content of active compounds in the root material before and after pasteurization was quantified by HPLC analysis. The results show that the eleutherosides in E. senticosus, and to an extent the ginsenosides from P. ginseng, could withstand pasteurization, whereas salidroside and rosavin from R. rosea did not survive pasteurization. Thus, R. rosea is not suitable for products requiring pasteurization.

Extract from Acanthopanax senticosus harms (Siberian ginseng) activates NTS and SON/PVN in the rat brain.

The extract of the stem bark of Siberian ginseng, Acanthopanax senticosus Harms (ASH), is believed to play a body-coping role in stress through a brain noradrenergic mechanism. The present study was carried out to investigate the effect of ASH on the neuronal activation patterns of c-Fos expression in the rat brain. With ASH administration, c-Fos accumulated in both the supraoptic nuclei (SON) and paraventricular nuclei (PVN), which regulate stress response. Only the caudal regions in the nucleus of the solitary tract (NTS), a locus innervating both the SON and PVN, were activated. Such a neuro-anatomical pattern associated with ASH suggests the possible involvement of these stress-related brain loci.

Methyl jasmonate induced overproduction of eleutherosides in somatic embryos of Eleutherococcus senticosus cultured in bioreactors

This study was concentrated on the production of eleutherosides and chlorogenic acid in embryogenic suspension cultures of Eleutherococcus senticosus by exposing them to different concentrations (50-400 µM) of methyl jasmonate (MJ) during the culture period. In the bioreactor cultures, eleutheroside content increased significantly by elicitation of MJ, however, the fresh weight, dry weight and growth ratio of embryos was strongly inhibited by increasing MJ concentrations. The highest total eleutheroside (7.3 fold increment) and chlorogenic acid (3.9 fold increment) yield was obtained with 200 µM MJ treatment. There was 1.4, 3.4 and 14.9 fold increase in the eleutheroside B, E, and E1 production respectively with such elicitation treatment. These results suggest that MJ elicitation is beneficial for eleutheroside accumulation in the embryogenic cell suspension cultures.