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Table of Contents : Pharmacognosy Magazine : 2010 - 6(21)


New free radical scavenging neolignans from fruits of Piper attenuatum


S Divya Reddy, Bandi Siva, B Poornima, D Anand Kumar, Ashok K Tiwari, U Ramesh, K Suresh Babu

Pharmacognosy Magazine 2015 11(42):235-241

Objective: The aim was to study and identify free radicals scavenging and antihyperglycemic principles in fruit of Piper attenuatum. Materials and Methods: Bioassay guided identification of extracts possessing potent free radical scavenging activity, and isolation of compounds was done. Chloroform extract of P. attenuatum possessing potent radical scavenging activity was also evaluated for antihyperglycemic activity following oral glucose tolerance test in rats. Results: Nine neolignans namely, denudatin B (1), iso-4', 5'- dimethoxy-3, 4-methylenedioxy-2'-oxo-Δ 3',5',8' -8.1'- lignan (2), lancifolin D (3), denudatin A (4), wallichinin (5), piperenone (6), lancifolin C (7), 2-oxo-piperol B (8), piperkadsin A (9) and a crotepoxide (10) was identified in Chloroform extract of P. attenuatum. Neolignans (1-9) displayed potent 2, 2'- azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) radical and piperkadsin A (9) also displayed 1, 1-diphenyl-2-picrylhydrazyl radical scavenging activity. Analysis of structure-activity relationship revealed that presence of furan ring and methoxy groups is an important criterion to influence 2, 2'-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) radical scavenging potentials. Chloroform extract of P. attenuatum fruit could not display antihyperglycemic activity following oral glucose tolerance test in rats. Conclusion: Neolignans present in P. attenuatum fruits are potent free radical scavengers and this is the first report identifying these compounds and activities in this fruit.

Anti-arthritic activity of Fu-Fang-Lu-Jiao-Shuang on collagen-induced arthritis in Balb/c mice and its underlying mechanisms


Yanyan Wang, Weiguang Sun, Laxia Chen, Xin Xu, Yunxia Wu, Jinwen Zhang, Yonghui Zhang

Pharmacognosy Magazine 2015 11(42):242-249

Background: Rheumatoid arthritis (RA) is a common, autoimmune disorder characterized by progressive multiple joint destruction, deformity, disability and premature death in most patients. Fu-Fang-Lu-Jiao-Shuang (FFLJS) is an effective traditional Chinese medicine, which has long been used clinically to treat RA patients. Objective: The objective of this study is aimed to evaluate the anti-rheumatic effects of FFLJS on collagen induced arthritis (CIA) model, as well as the underlying mechanisms, which have not previously been explored. Materials and Methods: CIA was induced by immunization with type II collagen (CII) in male Balb/c mice. The mice in the onset of arthritis were treated daily with FFLJS (125 or 500 mg/kg) or 1% carboxymethyl cellulose-Na for 28 days. Paw thickness and arthritic score were evaluated to confirm the anti-arthritic effect of FFLJS on CIA in mice. Levels of anti-CII antibody, proinflammatory cytokines interleukin-1 (IL-1) β, IL-17, and tumor necrosis factor-α (TNF-α) as well as prostaglandin E-2 (PGE-2) in serum and histological changes in the ankle joint were also analyzed. In addition, expressions of matrix metalloproteinases-1 (MMP-1), MMP-3 and tissue inhibitors of matrix metalloproteases-1 (TIMP-1) in synovial tissue were also detected to further study the molecular mechanism of the anti-arthritic effects of FFLJS. Results: During therapeutic treatment, FFLJS significantly reduced paw thickness and arthritic score in CIA mice, decreased the amounts of TNF-α, IL-1 β, IL-17, PGE-2 and anti-CII antibody in serum. In addition, FFLJS treatment could prevent the bone destruction by reducing the expression of MMP-1 and MMP-3, increasing the expression of TIMP-1 in synovial tissue of CIA mice. Conclusion: These findings offer the convincing evidence for the first time that the anti-rheumatic effects of FFLJS might be related to down-regulation of TNF-α, IL-1 β, IL-17 and PGE-2 levels for acute arthritis, and regulation of MMP-1, MMP-3 and TIMP-1 protein expression for chronic arthritis.

Shikonin induces apoptosis in the human gastric cancer cells HGC-27 through mitochondria-mediated pathway


Yue Hou, Jinghua Xu, Xia Liu, Xichun Xia, Ning Li, Xiuli Bi

Pharmacognosy Magazine 2015 11(42):250-256

Background: Gastric cancer (GC) is one of the most frequently occurring digestive tract cancers and fewer chemotherapeutic drugs for GC have shown promising results. In this study, we investigated the anti-tumor activity of shikonin, a natural compound isolated from the Chinese plant Lithospermum erythrorhizon, against the human GC cell line HGC-27. Materials and Methods: HGC-27 cells treated with shikonin at a concentration of 30μM or above showed significant growth inhibition compared to control cells. Shikonin-treated cells also underwent apoptosis as detected by flow cytometric analysis and microscopic examination of cellular morphology. Further investigation into the underlying mechanism of apoptosis by western blot showed that the shikonin promoted the activation of poly-(ADP-ribose)-polymerase, caspase-3 and caspase-9 following 24 h or 48 h of treatment time, as well as the activation of caspase-8, but only after 48 h of treatment time. Furthermore, the levels of mitochondrial membrane potential, B-cell lymphoma 2 (Bcl-2) and Bcl-extra large were reduced following shikonin treatment while the level of Bax was increased. In addition, shikonin also caused a signifi cant reduction of the protein Survivin, while having little effect on the expression on X-linked inhibitor of apoptosis protein. Conclusion: Taken together, these results showed that the shikonin exhibited its anti-tumor activity against HGC-27 cells through inhibiting cell growth and promoting apoptosis by targeting mitochondrial-related signaling pathway. Our finding may represent a positive step in finding a natural and effective compound that could be important implication for future development of chemotherapeutic and/or chemopreventive agent against GC.

The enriched fraction of Elephantopus scaber Triggers apoptosis and inhibits multi-drug resistance transporters in human epithelial cancer cells


Asmy Appadath Beeran, Naseer Maliyakkal, Chamallamudi Mallikarjuna Rao, Nayanabhirama Udupa

Pharmacognosy Magazine 2015 11(42):257-268

Background: Medicinal plants have played an important role in the development of clinically useful anticancer agents. Elephantopus scaber (Asteraceae) (ES) is widely used in Indian traditional system of medicine for the treatment of various ailments including cancer. Objective: To investigate anticancer effects of ES in human epithelial cancer cells. Materials and Methods: Cytotoxicity of ethanolic extract of ES (ES-ET) and its fractions, such as ES Petroleum ether fraction (ES-PET), ES Dichloromethane fraction (ES DCM), n Butyl alcohol fraction (ES-BT), and ES-Rest (ES-R) were assessed in human epithelial cancer cell lines using sulforhodamine B (SRB) assay. Acridine orange/ethidium bromide assay and Hoechst 33342 assays were used to gauge induction of apoptosis. Cell cycle analysis and micronuclei assay were used to assess cell cycle specific pharmacological effects and drug induced genotoxicty. Further, the ability of ES to inhibit multi drug resistant (MDR) transporters (ABC-B1 and ABC-G2) was determined by Rhodamine (Rho) and Mitoxantrone (MXR) efflux assays. Results: The enriched fraction of ES (ES DCM) possessed dose-dependent potent cytotoxicity in human epithelial cancer cells. Further, treatment of cancer cells (HeLa, A549, MCF-7, and Caco-2) with ES DCM showed hall mark properties of apoptosis (membrane blebbing, nuclear condensation etc.). Similarly, ES DCM caused enhanced sub G0 content and micronuclei formation indicating the induction of apoptosis and drug induced genotoxicity in cancer cells, respectively. Interestingly, ES DCM inhibited MDR transporters (ABC B1 and ABC G2) in cancer cells. Conclusion: The enriched fraction of ES imparted cytotoxic effects, triggered apoptosis, induced genotoxicity, and inhibited MDR transporters in human epithelial cancer cells. Thus, ES appears to be potential anticancer agent.

Methyl gallate isolated from Spondias pinnata exhibits anticancer activity against human glioblastoma by induction of apoptosis and sustained extracellular signal-regulated kinase 1/2 activation


Dipankar Chaudhuri, Nikhil Baban Ghate, Sudhir Shankar Singh, Nripendranath Mandal

Pharmacognosy Magazine 2015 11(42):269-276

Background: Spondias pinnata has been reported for its efficient anticancer effects, but the studies were mostly focused on its extract. Objective: Since its bioactive compounds are largely unknown, this study was designed to characterize the lead components present in it and their anticancer activity against human glioblastoma cell line (U87). Materials and Methods: Major compounds from the ethyl acetate fraction were isolated by column chromatography and their anticancer potentials against U87 cells were evaluated. Furthermore, flow cytometric and immunoblotting analyses were performed to demonstrate the mechanism of apoptosis inducing activity of methyl gallate (MG) against U87 cell line. Results: Four major compounds were isolated from the ethyl acetate fraction. Amongst these, two compounds showed promising activities and with the help of different spectroscopic methods they were identified as gallic acid and MG. Flow cytometric studies revealed that MG-induced apoptosis in U87 cells dose-dependently; the same was confirmed by activation of caspases through cleavage of endogenous substrate poly (adenosine diphosphate-ribose) polymerase. MG treatment also induced the expression of p53 and B-cell lymphoma-2-associated X and cleavage of BH3 interacting-domain with a concomitant decrease in B-cell lymphoma-2 expression. Moreover, MG-induced sustained phosphorylation of extracellular signal-regulated kinase (ERK1/2) in U87 cells with no change in the phosphorylation of other mitogen-activated protein kinases (c-Jun N-terminal of stress-activated protein kinases, p38). Conclusion: MG is a potent antioxidant and it induces sustained ERK1/2 activation and apoptosis in human glioblastoma U87, and provide a rationale for evaluation of MG for other brain carcinoma cell lines for the advancement of glioblastoma therapy.

Comparative evaluation of standardized alcoholic, hydroalcoholic, and aqueous extracts of Phyllanthus maderaspatensis Linn. against galactosamine-induced hepatopathy in albino rats


UK Ilyas, Deepshikha P Katare, Vidhu Aeri

Pharmacognosy Magazine 2015 11(42):277-282

Background: Phyllanthus maderaspatensis species (Euphorbiaceae) has been used in folk medicine of many countries as a remedy against several pathological conditions including jaundice and hepatitis. This study is an attempt to evaluate hepatoprotective activity of P. maderaspatensis against galactosamine-induced toxicity and also investigation of polyphenols in each extract. Materials and Methods: The extraction of P. maderaspatensis as per Ayurveda was simultaneously standardized and quantified for biochemical markers viz., polyphenols: Kaempferol, quercetin, catechin, rutin, and ellagic acid by high-performance thin layer chromatography. Hepatotoxicity was induced albino adult rats by intraperitoneal injection of galactosamine (400 mg/kg). The quantified aqueous, hydroalcoholic and alcoholic extract of P. maderaspatensis (200 and 400 mg/kg body weight/day) were compared for evaluation of hepatoprotective potential, which were assessed in terms of reduction in histological damage, change in serum enzymes such as aspartate amino transaminase, alanine amino transaminase and alkaline phosphatase and increase thiobarbituric acid reactive substances. Results and Discussion: The hydroalcoholic extract was found to contain comparatively high amount of kaempferol, quercetin, catechin, rutin, and ellagic acid which are responsible for hepatoprotection. Antioxidant parameters such as glutathione, catalase, and superoxide dismutase activity in liver tissues were restored toward the normalization more significantly by the hydroalcoholic extract when compared with other extracts. The biochemical observations were supplemented with histopathological examination. Conclusion: The hydroalcoholic extract standardized with respect to known biomarkers may be considered as a potent extract against hepatotoxicity.

Attenuation of reactive oxygen/nitrogen species with suppression of inducible nitric oxide synthase expression in RAW 264.7 macrophages by bark extract of Buchanania lanzan


Shailasree Sekhar, KK Sampath-Kumara, SR Niranjana, HS Prakash

Pharmacognosy Magazine 2015 11(42):283-291

Background: Oxidative stress is one of the most critical factors implicated in disease conditions. Buchanania lanzan Spr. (Anacardiaceae) bark powder preparation has been reported for treating an inflammatory condition in the Ayurvedic Pharmacopoeia of India. Objective: In the present study, we investigate the effect of the bark methanol extract (BLM) on reactive oxygen/nitrogen species (ROS/RNS), the expression of protein and mRNA of inducible nitric oxide synthase (iNOS) in RAW 264.7 macrophages stimulated with lipopolysaccharide (LPS) and sodium nitroprusside (SNP) to provide scientific validation of the above said medicinal property. Materials and Methods: The capacity to quench ROS and RNS was evaluated by 5-(and-6) chloromethyl-20,70-dichlorodihydrofluorescein diacetate acetyl ester fluorescence and nitrite estimations in LPS/SNP-stimulated macrophages respectively. The protein and transcript expression of iNOS was evaluated through Western Blot and reverse transcription-polymerase chain reaction (RT-PCR) analysis respectively. Results: Macrophages pretreated with BLM (>100 &#956;g/mL) for 24 h, stimulated with LPS for the last 18 h of experimental duration recorded a significantly (P < 0.05) reduced levels of ROS (3.45-fold) against LPS-stimulated conditions (5.7-fold). SNP-stimulation resulted in increased NO accumulation (17-fold) which was neutralized by BLM at >100 &#956;g/ml (1.6-fold) credited to a reduced protein and mRNA expression of iNOS as recorded by Western blots and RT-PCR results respectively. The reversed-phase liquid chromatography-diode array detection analysis identified the presence of 4-hydroxybenzoic acid, quercetin and p-coumaric acid (Rt values 5.444, 5.569 and 9.580 respectively). Conclusions: The potential of BLM inhibiting ROS/RNS production validates the medical use of bark, could find beneficial application under conditions of immune stimulation and/or bacterial infection.

Preconditioning with glycyrrhizic, ferulic, paeoniflorin, cinnamic prevents rat hearts from ischemia/reperfusion injury via endothelial nitric oxide pathway


Guo-Qiang Qian, Jingjing Ding, Xiaozhao Zhang, Xiaofeng Yin, Yuqin Gao, Guo-Ping Zhao

Pharmacognosy Magazine 2015 11(42):292-296

Objective: The objective was to investigate the endothelial nitric oxide synthase (eNOS/NO) pathway is involved or not in the protective effects of glycyrrhizic, ferulic, paeoniflorin, cinnamic (GFPC) in myocardial ischemia-reperfusion injury Sprague-Dawley rats. Materials and Methods: Ischemia-reperfusion (I/R) model was made by ligating the left anterior descending branch of the coronary artery for 30 min and releasing for 120 min, then the left ventricular apical was fixed and sliced, morphological changes of myocardial microvascular endothelial cell (MMVEC) was observed by electron microscopy, apoptosis index of MMVEC was observed by means of TUNEL, serum NO was tested by methods of nitrate reduction, lactate dehydrogenase (LDH), creatine kinase MB (CK-MB) was detected by automatic biochemical analyzer; Phosphorylated eNOS (PeNOS) and inducible NOS (iNOS) protein were measured by means of western blot. Results: In positive product control group, the serum levels of NO, LDH, CK-MB significantly increased (P < 0.05); MMVEC apoptosis was significantly decreased (P < 0.05); incidence of area at risk decreased significantly (P < 0.05); PeNOS protein increased (P < 0.05); iNOS protein decreased significantly (P < 0.05). Conclusion: Ischemic preconditioning of GFPC from GFPC plays a protective role in I/R heart through regulating the eNOS/NO signal pathway by increasing the PeNOS protein expression and decreasing the expression of iNOS protein.

Simultaneous determination three phytosterol compounds, campesterol, stigmasterol and daucosterol in Artemisia apiacea by high performance liquid chromatography-diode array ultraviolet/visible detector


Jiwoo Lee, Jin Bae Weon, Bo-Ra Yun, Min Rye Eom, Choong Je Ma

Pharmacognosy Magazine 2015 11(42):297-303

Background: Artemisia apiacea is a traditional herbal medicine using treatment of eczema and jaundice in Eastern Asia, including China, Korea, and Japan. Objective: An accurate and sensitive analysis method using high performance liquid chromatography-diode array ultraviolet/visible detector and liquid chromatography-mass spectrometry for the simultaneous determination of three phytosterol compounds, campesterol, stigmasterol and daucosterol in A. apiacea was established. Materials and Methods: The analytes were separated on a Shiseido C 18 column (5 &#956;m, 4.6 mm I.D. &#215;250 mm) with gradient elution of 0.1% trifluoroacetic acid and acetonitrile. The flow rate was 1 mL/min and detection wavelengths were set at 205 and 254 nm. Results: Validation of the method was performed to demonstrate its linearity, precision and accuracy. The calibration curves showed good linearity (R2 > 0.9994). The limits of detection and limits of quantification were within the ranges 0.55-7.07 &#956;g/mL and 1.67-21.44 &#956;g/mL, respectively. And, the relative standard deviations of intra- and inter-day precision were <2.93%. The recoveries were found to be in the range of 90.03-104.91%. Conclusion: The developed method has been successfully applied to the analysis for quality control of campesterol, stigmasterol and daucosterol in A. apiacea.

A new method for identification of natural, artificial and in vitro cultured Calculus bovis using high-performance liquid chromatography-mass spectrometry


Yonggang Liu, Peng Tan, Shanshan Liu, Hang Shi, Xin Feng, Qun Ma

Pharmacognosy Magazine 2015 11(42):304-310

Objective: Calculus bovis have been widely used in Chinese herbology for the treatment of hyperpyrexia, convulsions, and epilepsy. Nowadays, due to the limited source and high market price, the substitutes, artificial and in vitro cultured Calculus bovis, are getting more and more commonly used. The adulteration phenomenon is serious. Therefore, it is crucial to establish a fast and simple method in discriminating the natural, artificial and in vitro cultured Calculus bovis. Bile acids, one of the main active constituents, are taken as an important indicator for evaluating the quality of Calculus bovis and the substitutes. Several techniques have been built to analyze bile acids in Calculus bovis. Whereas, as bile acids are with poor ultraviolet absorbance and high structural similarity, effective technology for identification and quality control is still lacking. Methods: In this study, high-performance liquid chromatography (HPLC) coupled with tandem mass spectrometry (LC/MS/MS) was applied in the analysis of bile acids, which effectively identified natural, artificial and in vitro cultured Calculus bovis and provide a new method for their quality control. Results: Natural, artificial and in vitro cultured Calculus bovis were differentiated by bile acids analysis. A new compound with protonated molecule at m/z 405 was found, which we called 3&#945;, 12&#945;-dihydroxy-7-oxo-5&#945;-cholanic acid. This compound was discovered in in vitro cultured Calculus bovis, but almost not detected in natural and artificial Calculus bovis. A total of 13 constituents was identified. Among them, three bio-markers, including glycocholic acid, glycodeoxycholic acid and taurocholic acid (TCA) were detected in both natural and artificial Calculus bovis, but the density of TCA was different in two kinds of Calculus bovis. In addition, the characteristics of bile acids were illustrated. Conclusions: The HPLC coupled with tandem MS (LC/MS/MS) method was feasible, easy, rapid and accurate in identifying natural, artificial and in vitro cultured Calculus bovis.

Effects of Pogostemon cablin Blanco extract on hypoxia induced rabbit cardiomyocyte injury


Chi-Yeon Lim, Bu-Yeo Kim, Se-Hyun Lim, Su-In Cho

Pharmacognosy Magazine 2015 11(42):311-319

Background: Pogostemonis Herba, the dried aerial part of Pogostemon cablin Blanco, is a well-known materia medica in Asia that is widely used for syndrome of gastrointestinal dysfunctions. Objective: This study was undertaken to examine whether Pogostemon cablin extract (PCe) might have any beneficial effect on hypoxia induced rabbit cardiomyocyte injury. Materials and Methods: Isolated cardiomyocytes were divided into three groups and the changes of cell viability in cardiomyocytes of hypoxic and hypoxia/reoxygenation group were determined. The effect of PCe on reactive oxygen species (ROS) generation, intracellular formation of ROS was also measured by monitoring the 2',7'- dichlorofluorescein fluorescence. Results: PCe effectively protected the cells against both the hypoxia and reoxygenation induced injury, and the protective effect of PCe is not mediated by interaction with adenosine triphosphate-sensitive K + channels. In the presence of PCe, production of ROS under chemical hypoxia was remarkably reduced which suggests that PCe might exert its effect as a ROS scavenger. Conclusion: The present study provides clear evidence for the beneficial effect of PCe on cardiomyocyte injury during hypoxia or reoxygenation following prolonged hypoxia.

Quantitative analysis of the eight major compounds in the Samsoeum using a high-performance liquid chromatography coupled with diode array detection and electrospray ionization mass spectrometer


Jin Bae Weon, Hye Jin Yang, Bohyoung Lee, Jin Yeul Ma, Choong Je Ma

Pharmacognosy Magazine 2015 11(42):320-326

Background: Samsoeum was traditionally used for treatment of a respiratory disease. Objective: The simultaneous determination of eight major compounds, ginsenoside Rg3, caffeic acid, puerarin, costunolide, hesperidin, naringin, glycyrrhizin, and 6-gingerol in the Samsoeum using a high-performance liquid chromatography (HPLC) coupled with diode array detection (DAD) and an electrospray ionization mass spectrometer was developed for an accurate and reliable quality assessment. Materials and Methods: Eight compounds were qualitative identified based on their mass spectra and by comparing with standard compounds and quantitative analyzed by HPLC-DAD. Separation of eight compounds was carried out on a LUNA C 18 column (S-5 &#956;m, 4.6 mm i.d. &#215;250 mm) with gradient elution composed of acetonitrile and 0.1% trifluoroacetic acid. Results: The data showed good linearity (R2 > 0.9996). The limits of detection and the limits of quantification were <0.53 &#956;g and 1.62 &#956;g, respectively. Inter- and Intra-day precisions (expressed as relative standard deviation values) were within 1.94% and 1.91%, respectively. The recovery of the method was in the range of 94.24-107.90%. Conclusion: The established method is effective and could be applied to quality control of Samsoeum.

Sesamol, a lipid lowering agent, ameliorates aluminium chloride induced behavioral and biochemical alterations in rats


Jessy John, Madhavan Nampoothiri, Nitesh Kumar, Jayesh Mudgal, Gopalan Kutty Nampurath, Mallikarjuna Rao Chamallamudi

Pharmacognosy Magazine 2015 11(42):327-336

Background: Sesame oil from the seeds of Sesamum indicum Linn. (Pedaliaceae) has been used traditionally in Indian medical practice of Ayurveda in the treatment of central nervous system disorders and insomnia. A few published reports favor the anti-dementia effect of sesamol (SML), an active constituent of sesame oil. Objective: Thus, the present study was aimed to explore the anti-dementia effect and possible mechanism (s) of SML in aluminium chloride (AlCl 3 )-induced cognitive dysfunction model in rodents with special emphasis on memory centers viz., hippocampus and frontal cortex. Methods: Male Wistar rats were exposed to AlCl 3 (175 mg/kg p.o.) for 60 days. SML (10 and 20 mg/kg) and rivastigmine (1 mg/kg) were administered orally 45 min before administration of AlCl 3 for 60 days. Spatial memory was assessed using Morris water maze test. After 60 days of treatment animals were sacrificed, hippocampus and frontal cortex were collected and analyzed for acetylcholinesterase (AChE) activity, tumor necrosis factor (TNF-&#945;) level, antioxidant enzymes (Glutathione, catalase), lipid peroxidation, and nitrite level. The circulating triglycerides, total cholesterol, low-density lipoprotein (LDL) and high-density lipoprotein (HDL) levels were also analyzed. Results: SML significantly prevented behavioral impairments in aluminium-exposed rats. Treatment with SML reversed the increased cholesterol, triglycerides and LDL while raised the HDL levels. SML significantly corrected the effect of AlCl 3 on AChE activity. Further, SML reversed the elevated nitric oxide, TNF-&#945; and reduced antioxidant enzymes in hippocampus and frontal cortex. Conclusion: The present study suggests the neuro-protection by SML against cognitive dysfunction induced by environmental toxin (AlCl 3 ) in hippocampus and frontal cortex.

The influences of inorganic elements in soil on the development of famous - region Atractylodes lancea (Thunb.) DC


Weiwanqi Zhang, Zhen Ouyang, Ming Zhao, Yuan Wei, Huasheng Peng, Qiang Wang, Ling Guo

Pharmacognosy Magazine 2015 11(42):337-344

Background: Atractylodes lancea (Thunb.) DC., is an important medicinal plant in China. Recently, researches of A. Lancea were focused on chemical composition and genetics, only a few were concerned with soil factors. Objective: The aim was to discuss the relationship between geo-herbalism of A. Lancea (Thunb.) DC. and inorganic elements in soil. Materials and Methods: The contents of 15 kinds of inorganic elements in the rhizoma of A. Lancea (Thunb.) DC. and soils from various regions were determined with inductively coupled plasma-optical emission spectrometer and the data were analyzed with Statistical Package for the Social Sciences 20.0 software. Results: The contents of inorganic elements in rhizoma of A. Lancea and in soil with different geological background were different. The soils in the famous region contained high aluminum, iron, sodium and low sulfur content. The rhizoma of A. Lancea contained high aluminum, lithium, manganese and low iron, sulfur content. The famous-region crude drugs had a strong tendency to accumulate selenium, manganese. Ten characteristic elements of A. Lancea were K, Ca, S, Al, Li, Ti, Mn, Pb, Ni, SE. Conclusion: The contents of inorganic elements in rhizoma of A. Lancea showed a significant positive interrelationship with those in soil. It was identified that inorganic elements play an important role in forming authenticity A. Lancea (Thunb.) DC.

Ethyl acetate extract from marine sponge Hyattella cribriformis exhibit potent anticancer activity by promoting tubulin polymerization as evidenced mitotic arrest and induction of apoptosis


Pazhanimuthu Annamalai, Malini Thayman, Sowmiya Rajan, Lakshmi Sundaram Raman, Sankar Ramasubbu, Pachiappan Perumal

Pharmacognosy Magazine 2015 11(42):345-355

Background: Marine sponges are important sources of bioactive compounds. Objective: This study investigated the anticancer properties of Hyattella cribriformis ethyl acetate (EA) fraction in various cancer and normal cell lines. Materials and Methods: anticancer assay was carried out in 15 cell lines to evaluate the anticancer potential of the EA fraction. Impact on cell cycle distribution was determined using flow cytometry. The fraction was investigated for interfering microtubules assembly in both in vitro and cellular assay. Further studies were conducted to determine the fraction induced cell death (apoptosis) using calcein/propidium iodide dual staining, activated caspase-3 and phosphorylation of Bcl-2 protein at Ser70. DNA fragmentation assay was performed to confirm the apoptosis. Results: EA fraction exhibited potent inhibition of cancer cell growth and resulted in 50% growth inhibition (GI 50 ) of 0.27 &#956;g/mL in A673 cell line. Sarcoma (MG-63, Saos-2) and ovarian (SK-OV-3 and OVCAR-3) cancer cell lines also showed superior anticancer activity GI 50 of 1.0 &#956;g/mL. Colon and breast cancer cell lines exhibited moderate GI compare other cancer cell lines and normal human lung fibroblast showed GI 50 of 15.6 &#956;g/mL. EA fraction showed potent G2/M phase arrest in A673 cell line and induced apoptosis at 48 h exposure. EA fraction promoted microtubule polymerization in tubulin polymerization assay and increased level of polymerized tubulin in the HeLa cells. Fraction induced the activation of caspase-3 and phosphorylation of Bcl-2 anti-apoptotic protein. Fraction induced DNA fragmentation in HeLa cells as evidence of apoptosis. Conclusion: Marine sponge H. cribriformis EA fraction exhibited potent anticancer activity through tubulin polymerization and induction of apoptosis.

Identification of longevity, fertility and growth-promoting properties of pomegranate in Caenorhabditis elegans


Hasan Kiliçgün, Nazli Arda, Evren Önay Uçar

Pharmacognosy Magazine 2015 11(42):356-359

Background: Pomegranate (Punica granatum L.) is commonly consumed as fresh fruit and fruit juice. It is also used in the production of jam, wine, food coloring agent, and flavor enhancer. Objective: The aim of this study was to identify the possible longevity, fertility and growth promoting properties of different ethanolic extract concentrations of pomegranate in Caenorhabditis elegans, which is increasingly popular and has proven to be a very useful experimental model organism for aging studies as well as for testing antioxidants and other compounds for effects on longevity. Materials and Methods: In this study, five experimental groups (20, 10, 5, 2.5 and 1.25 mg pomegranate extract/mL and one control group) were used to determine the most effective dose of pomegranate in terms of longevity, fertility and growth parameters. Results: It was seen that, pomegranate extracts up to the concentration of 5 mg/mL, had the potential to promote for the longevity, formation of new generations, fertility of new generations and growth properties of C. elegans although higher concentrations significantly reduced these parameters. Conclusion: these findings indicated that pomegranate could be used as a supplement to enhance longevity, fertility and growth rate for the other living organisms and human beings, but the dose should be carefully adjusted to avoid adverse effects.

Simultaneous quantification of six alkaloid components from commercial stemonae radix by solid phase extraction-high-performance liquid chromatography coupled with evaporative light scattering detector


Rong-Rong Zhang, Dan-Yi Lu, Zhen-Ya Yang, Wen Zhao, Paul Pui-Hay But, Pang-Chui Shaw, Ren-Wang Jiang, Zhi-Guo Ma

Pharmacognosy Magazine 2015 11(42):360-367

Background: Stemonae radix has been applied in traditional Chinese medicine for centuries. Alkaloids are the main active ingredient in stemonae radix, so their composition and concentration levels are directly linked to clinic effects. Objective: The objective was to develop an analytical method with multiple markers for quality survey of commercial stemonae radix. Materials and Methods: A method for simultaneous determination of six compounds in commercial stemonae radix was performed using solid-phase extraction and high-performance liquid chromatography coupled with evaporative light scattering detector. The separation was carried out on an Agilent TC-C18 column with 0.1% acetonitrile solution of triethylamine aqueous solution and acetonitrile as the mobile phase under gradient elution within 70 min. The hierarchical clustering analysis (HCA) was successfully used to classify the samples in accordance with their chemical constituents. Results: Linearity (R 2 > 0.9990), intra- and inter-day precision (relative standard deviations <4%), limit of detection (0.011-0.086 &#956;g/mL), limit of quantification (0.033-0.259 &#956;g/mL) of the six alkaloids were determined, and the recoveries were between 96.6% and 103.7%. The method was successfully applied to analysis 36 batches of commercial stemonae radix. All the samples could be classified into five clusters by HCA. Conclusion: This article provides an accurate and simple analytical method for quality survey of commercial stemonae radix. Because of the significant chemical variations, careful selection of Stemona sources with obvious antitussive value but devoid of croomine followed by good agricultural practice and good manufacturing practice process is suggested.

Evaluation of extraction method on the chemical composition in Apeiba tibourbou Aubl's extracts


Frederico Severino Martins, Edemilson Cardoso da Conceição

Pharmacognosy Magazine 2015 11(42):368-373

Background: The extraction method of bioactive compounds is an important step in the manufacturing of herbal medicines, because secondary metabolites with therapeutic potential are usually found in small quantities in plant materials. Objective: Due the potential of Apeiba tibourbou Aubl, this study aimed to evaluate the impact of the extraction method on the quality of herbal extract and optimize the extraction of fatty acid, rosmarinic (Ra) and caffeic (Ca) acid from A. tibourbou. Materials and Methods: Determinations of residual moisture (Rm), proteins (Pt), lipids (Lp), total fiber (Tf), and carbohydrate (Cy) were performed in triplicate samples according assessment of antioxidant capacity. Extraction of fatty acids was carried out by two different methods: (i) By shoxlet and (ii) bligh and dyer. The optimized conditions were determined by surface response methodology (RSM), and the criterion of desirability was the maximum extraction of Ra and Ca. Results: The method of bligh and dyer was able to extraction more total Lp than the shoxlet. However, the extraction of fatty acid was different for the two methods. The optimized conditions to extract RA and Ca was calculated by RSM, 42&#176;C, 30% (alcohol degree) and 24 min, this conditions maximize simultaneously the extraction of Ca (0, 04%) and Ry (1.89), Conclusion: It was observed that the extraction method alters the chemical composition of extract, and it is possible to extract Ca and Ra from A. tibourbou's leaves using ultrasound-assisted extraction.

In vitro regeneration and Agrobacterium tumefaciens-mediated genetic transformation in asakura-sanshoo (Zanthoxylum piperitum (L.) DC. F. inerme Makino) an important medicinal plant


Xiaofang Zeng, Degang Zhao

Pharmacognosy Magazine 2015 11(42):374-380

Context: Asakura-sanshoo (Zanthoxylum piperitum [L.] DC. f. inerme Makino) is an important medicinal plant in East Asia. Transgenic technique could be applied to improve plant traits and analyze gene function. However, there is no report on regeneration and genetic transformation in Asakura-sanshoo. Aims: To establish a regeneration and Agrobacterium tumefaciens-mediated genetic transformation system in Asakura-sanshoo, which could be used for cultivar improvement and gene function analysis. Settings and Design: The various combinations of indole-3-butyric acid (IBA), 6-benzylaminopurine (BA) and naphthalene acetic acid (NAA) were explored for the optimal plant regeneration from petiole and stem of Asakura-sanshoo. The half-strength woody plant medium (WPM) with different concentrations of NAA and IBA was used to induce root. For genetic transformation, A. tumefaciens strain EHA-105 harboring the plasmid pBin-Ex-H-ipt which carries the isopentenyl transferase (ipt) gene, &#946;-glucuronidase (GUS) gene and kanamycin resistance gene neomycin phosphotransferase II (NPTII) were used. The transformation efficiency was detected by the kanamycin resistant frequency. Materials and Methods: Petioles and stems were obtained from the in vitro cultured Asakura-sanshoo. The petiole and stem segments were precultured for 3 days, and then inflected using the bacterium at the concentration of OD 600 0.5-0.8 for 10 min, followed by 3 days co-cultivation. Selection of the transgenic plants was carried out after 7 days the regeneration using gradient kanamycin at 30 mg/L and 50 mg/L, respectively. Successful transformed plants were confirmed by GUS histochemical assays, polymerase chain reaction (PCR), reverse transcription-PCR (RT-PCR), and Southern blotting analysis. Results: The highest shoots regeneration was obtained on WPM supplement with 0.5 mg/L BA and 0.2 mg/L NAA. The optimal rooting medium was half strength macro-element WPM. The kanamycin resistant frequency of petiole and stem was 24.66% and 25.93%, respectively. Thirty-five shoots in thousands adventitious buds were confirmed through GUS histochemical assays, PCR, RT-PCR, and Southern blotting. The regeneration shoot per explants elevated 5.85 fold compared with the wild-type plants. Conclusions: Individual transgenic Asakura-sanshoo lines were obtained. In this paper, it first revealed the expression of ipt gene significantly promoted the adventitious buds induction in Asakura-sanshoo as the same action as in other plants.

Effects of Valerianae Radix et Rhizoma extract on psychological stress in mice


Jeong Suk Kim, Jeong Deok Ahn, Su-In Cho

Pharmacognosy Magazine 2015 11(42):381-388

Background: The aim of this study was to identify the effects of Valerianae Radix et Rhizoma water extract (VRe) originated from Valeriana fauriei Briquet on reducing psychological stress (PS) on mice. Objective: Mice were put under PS with communication box method: Restraining mice and forcing to see other mice underfoot shock stress. Materials and Methods: Measurements on plasma corticosterone, noradrenaline and lipid peroxidation, and elevated plus-maze (EPM) tests were carried out to determine the effect of VRe administration on physiological and behavioral responses of mice. Results: VRe showed anxiolytic effects in plasma corticosterone, noradrenaline, and EPM transfer latency levels, but it did not show any significant effects on the other indicators. Conclusion: V. fauriei, which has been used as a natural anxiolytic drug, exerts positive effects in the communication box induced PS in mice.

Antiradical and antidiabetic properties of standardized extract of Sunderban mangrove Rhizophora mucronata


Tapas Kumar Sur, Alok Kumar Hazra, Dipankar Bhattacharyya, Avijit Hazra

Pharmacognosy Magazine 2015 11(42):389-394

Background: Mangroves have the ability to grow where no other vascular plants survive. Rhizophora mucronata is a true mangrove and traditionally used to treat diabetes and its allied complications. Objectives: In the present study, we standardized the 80% methanolic standardized extract of R. mucronata leaves (RH) and found out its antiradical and antidiabetic activities. Materials and Methods: The methanolic extract of R. mucronata leaves (RH) was standardized and quantified for phenolics, flavonoinds, gallic acid, quercetin, and coumarin. The reducing abilities and antiradical activities of RH were performed in vitro methods like, 1,1-diphenyl-2-picrylhydrazyl, nitric oxides, superoxides, hydroxyl, and ABTS (2,2'-azino-bis-3-ethyl benzthiazoline-6-sulphonic acid). Thereafter, RH was evaluated for it antidiabetic potentialities on streptozotocin (STZ)-induced type-2 diabetes. STZ (90 mg/kg, intraperitoneal) was administered to 2 days old pups to induce diabetes. RH was fed at doses of 50 and 100 mg/kg and glibenclamide (positive control) at 5 mg/kg, when the rats were 6 weeks old and continued for 10 weeks. Fasting glucose was monitored before and after the treatment. Further, lipid peroxides and reduced glutathione level were estimated on rat liver. Results: The results obtained from this study revealed RH possesses flavonoinds and also gallic acid, quercetin, and coumarin. Further, it has antiradical activities. It has also reduced blood glucose level in type-2 diabetic rats and reduced the formation of lipid peroxidation in liver. RH enhanced the level of glutathione in liver tissue. Conclusion: RH exhibits source of natural antioxidants and great potentialities as an antidiabetic agent by improving the hyperglycemia through its antiradical action.

Effect of co-administration of Angelicae gigantis radix and Lithospermi radix on rat hepatic injury induced by carbon tetrachloride


Chi-Yeon Lim, Bu-Yeo Kim, Se-Hyun Lim, Su-In Cho

Pharmacognosy Magazine 2015 11(42):395-403

Background: Co-administration of Angelicae gigantis radix (AGR) and Lithospermi radix (LR) has been commonly applied to patients to treat cardiac and hepatic disorders. Individual bioactivities of these herbal medicines have been widely investigated, but the hepatoprotective effects of co-treatment of AGR and LR have yet to be clarified. Objective: The present study investigated the protective effects of extracts of AGR and LR on carbon tetrachloride (CCl 4 ) induced hepatic injury. Materials and Methods: In this study, we measured the hepatoprotective activity of individual and co-treatment of the two herbal medicines on hepatic injury induced by CCl 4 by measuring different biochemical parameters such as serum aspartate aminotransaminase (AST) and serum alanine aminotransaminase (ALT). Microarray technology also used to compare ontological difference with individual and co-treatment of these two. Results: Combined treatment with AGR and LR (AGR + LR) decreased AST and ALT level in serum which demonstrate hepatoprotective effect of the therapy. When the effect of AGR and LR according to treatment conditions was measured, co-treatment showed the most prominent effect on hepatic injury by CCl 4 rather than individual treatment condition. We further defined gene set that could be the molecular target of herbal effect on hepatic injury by CCl 4 using bioinformatical analysis of interaction network. Highly recovered genes by treating AGR + LR play significant roles in response to hepatic injury induced by CCl 4 . Conclusion: Combined treatment with AGR and LR showed synergistic protective effects on the CCl 4 -induced rat hepatic tissue injury.

Quercetin induces human colon cancer cells apoptosis by inhibiting the nuclear factor-kappa B Pathway


Xiang-An Zhang, Shuangxi Zhang, Qing Yin, Jing Zhang

Pharmacognosy Magazine 2015 11(42):404-409

Quercetin can inhibit the growth of cancer cells with the ability to act as chemopreventers. Its cancer-preventive effect has been attributed to various mechanisms, including the induction of cell-cycle arrest and/or apoptosis as well as the antioxidant functions. Nuclear factor kappa-B (NF-kB) is a signaling pathway that controls transcriptional activation of genes important for tight regulation of many cellular processes and is aberrantly expressed in many types of cancer. Inhibitors of NF-kB pathway have shown potential anti-tumor activities. However, it is not fully elucidated in colon cancer. In this study, we demonstrate that quercetin induces apoptosis in human colon cancer CACO-2 and SW-620 cells through inhibiting NF-kB pathway, as well as down-regulation of B-cell lymphoma 2 and up-regulation of Bax, thus providing basis for clinical application of quercetin in colon cancer cases.

Bacopa monnieri promotes longevity in Caenorhabditis elegans under stress conditions


Suresh C Phulara, Virendra Shukla, Sudeep Tiwari, Rakesh Pandey

Pharmacognosy Magazine 2015 11(42):410-416

Background: Bacopa monnieri (L.) Pennell, commonly known as Brahmi is an important medicinal plant traditionally used as memory enhancer and antiepileptic agent. Objective: The present study investigated antioxidant and stress resistance potentials of B. monnieri aqueous extract (BMW) using Caenorhabditis elegans animal model system. Materials and Methods: The antioxidant activity of the BMW was measured using in vitro (DPPH, reducing power and total polyphenol content) and in vivo (DCF-DA assay) assays. The antistress potential of BMW (0.1, 0.01, and 0.001 mg/ml) was evaluated through thermal stress (37&#176;C) and oxidative stress (10 mM paraquat) using C. elegans. Quantification of the HSP-16.2 level was done using CL2070 transgenic worms. Results: Present study reveals that BMW possess in vitro and in vivo antioxidant activities. BMW significantly enhanced stress tolerance and increased the mean lifespan of worms during thermal and oxidative stress, although it did not extend lifespan at 20&#176;C and attenuated age dependent decline in physiological behaviors. Moreover, it was shown that BMW was able to up-regulate expression of stress associated gene hsp-16.2, which significantly (P < 0.001) extends the mean lifespan of worms under stress conditions. Conclusion: The study strongly suggests that BMW acts as an antistressor and potent reactive oxygen species scavenger which enhances the survival of the worms in different stress conditions.

Yin-Chen-Hao-Tang alleviates biliary obstructive cirrhosis in rats by inhibiting biliary epithelial cell proliferation and activation


Bing Wang, Ming-Yu Sun, Ai-Hua Long, Hong-Yan Cao, Shuang Ren, Yan-Qin Bian, Xiong Lu, Hong-Tu Gu, Cheng-Hai Liu, Ping Liu

Pharmacognosy Magazine 2015 11(42):417-425

Background: Yin-Chen-Hao-Tang (YCHT) consists of three aqueous extracts from Artemisia capillaris, Gardenia sp., and prepared Rheum rhabarbarum (rhubarb) (3:2:1). YCHT is characterized by its anti-inflammatory properties in liver regulation and relief of jaundice. We aimed to study the effects and mechanisms of action of YCHT on biliary obstructive cirrhosis. Materials and Methods: Secondary biliary fibrosis was induced in rats by bile duct ligation (BDL) and scission. One week after BDL, rats were randomly divided into a saline-treated BDL or YCHT-treated BDL group for 4 weeks. Liver function and hepatic hydroxyproline (Hyp) content were assessed. Types I and IV collagen (Col-IV), laminin, fibronectin, alpha smooth muscle actin (&#945;-SMA), and proliferating cell nuclear antigen protein and messenger ribonucleic acid (mRNA) expression were assessed with immunohistochemistry and real-time polymerase chain reaction. Results: In the YCHT-treated BDL group, serum total bilirubin, total bile acids, aspartate aminotransferase, alanine aminotransferase, and g-glutamyl transferase were lower than those in the sham-operated BDL group. The proliferation of bile ducts in hepatic tissues and the Hyp content and Col deposition were also significantly lower than those in control rats. In addition, &#945;-SMA and Col-IV staining was less obvious, and mRNA expression of Procol-&#945;1 (IV), platelet derived growth factor subunit B (PDGF)-B, connective tissue growth factor, and transforming growth factor-beta in proliferative biliary epithelial cells (BECs) in the YCHT-treated BDL group was significantly lower than those in controls. Conclusions: YCHT effectively reduces the formation of biliary obstructive cirrhosis mainly via inhibition of BEC proliferation by down-regulation of PDGF-B mRNA expression, inhibition of BEC profibrogenic paracrines, and the epithelial-mesenchymal transition pathological process.

In vivo antitumor potential of Ipomoea pes-caprae on melanoma cancer


Ashish Manigauha, MD Kharya, N Ganesh

Pharmacognosy Magazine 2015 11(42):426-433

Background: The incidence of skin cancers is rising gradually. The treatment of melanoma is also necessary to prevent the spread of cancer to other body organs. Scientific literatures have not documented any evidence of the antitumor potential of Ipomoea pes-caprae on melanoma. Aim of the Study: Explore in vivo antitumor potential of I. pes-caprae on melanoma cancer. Materials and Methods: Petroleum ether (60&#176;C-80&#176;C), methanolic and aqueous extracts, and swaras prepared from the whole herb of I. pes-caprae were assessed for their antitumor activity. The extracts and swaras at doses of 25 and 50 mg/kg b. wt. were administered intraperitoneal along with chemo and radiotherapy for 40 days for exploring antitumor activity against melanoma cancer (B16F10) in male C57BL mice. The results obtained from tumor volume, and histopathological studies were compared with the control and dacarbazine used as a standard. Results: Antitumor effect of I. pes-caprae extracts and swaras on melanoma cancer was found to be significant (P < 0.01) compared to normal control. The tumor volume inhibition against tumor-bearing mice, although differed from each other, was concentration dependent. Administration of plant extracts and swaras from the day 1 since tumor inducted. The induction of tumor was found delayed by 10-15 days and the tumor volume on the day 40 was similar to the Dacarbazine treatment used as a standard. Conclusion: The results obtained from the tumor volume and histopathological studies clearly revealed the antitumor potential of I. pes-caprae on melanoma cancer.