Chinchu Bose

Anacardic acid inhibits the catalytic activity of matrix metalloproteinase-2 and matrix metalloproteinase-9.

Cashew nut shell liquid (CNSL) has been used in traditional medicine for the treatment of a wide variety of pathophysiological conditions. To further define the mechanism of CNSL action, we investigated the effect of cashew nut shell extract (CNSE) on two matrix metalloproteinases, MMP-2/gelatinase A and MMP-9/gelatinase B, which are known to have critical roles in several disease states. We observed that the major constituent of CNSE, anacardic acid, markedly inhibited the gelatinase activity of 3T3-L1 cells. Our gelatin zymography studies on these two secreted gelatinases, present in the conditioned media from 3T3-L1 cells, established that anacardic acid directly inhibited the catalytic activities of both MMP-2 and MMP-9. Our docking studies suggested that anacardic acid binds into the MMP-2/9 active site, with the carboxylate group of anacardic acid chelating the catalytic zinc ion and forming a hydrogen bond to a key catalytic glutamate side chain and the C15 aliphatic group being accommodated within the relatively large S1′ pocket of these gelatinases. In agreement with the docking results, our fluorescence-based studies on the recombinant MMP-2 catalytic core domain demonstrated that anacardic acid directly inhibits substrate peptide cleavage in a dose-dependent manner, with an IC50 of 11.11 μM. In addition, our gelatinase zymography and fluorescence data confirmed that the cardol-cardanol mixture, salicylic acid, and aspirin, all of which lack key functional groups present in anacardic acid, are much weaker MMP-2/MMP-9 inhibitors. Our results provide the first evidence for inhibition of gelatinase catalytic activity by anacardic acid, providing a novel template for drug discovery and a molecular mechanism potentially involved in CNSL therapeutic action.

Gossypetin, a naturally occurring hexahydroxy flavone, ameliorates gamma radiation-mediated DNA damage

Abstract Purpose: To evaluate the protective effect of gossypetin (GTIN) against gamma (γ)-radiation-mediated DNA damage. Materials and methods: Increasing concentrations (10–150 μM) of GTIN were incubated with supercoiled DNA 1 h prior exposure to γ-radiation in the range of 5-Gy absorbed dose from Co60 γ source. To establish the effective protective concentration of GTIN, supercoiled DNA was pre-incubated with 50 μM of GTIN for 1 h followed by exposure of 5, 10 and 20 Gy doses of γ-radiation. Moreover, 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical, hydroxyl radical, nitric oxide (NO) scavenging, metal chelating activity and ferric reducing antioxidant power (FRAP) of GTIN were measured and compared with standards. The flowcytometric analysis and radiation-induced genomic DNA damage by comet assay were employed to estimate the level of intracellular reactive oxygen species (ROS) using isolated murine hepatocytes. Results: GTIN was able to effectively scavenge different free radicals in in vitro situations. It could significantly prevent radiation induced supercoiled and genomic DNA damage with reduced comet parameters. It also acted as a potent scavenger of the radiation induced ROS. Conclusions: GTIN ameliorated radiation-induced oxidative stress and DNA damage by its free-radical scavenging activity.

Antimicrobial activity of plumbagin, a naturally occurring naphthoquinone from Plumbago rosea, against Staphylococcus aureus and Candida albicans

Candida albicans and Staphylococcus aureus are opportunistic pathogens. Despite causing a number of independent infections, both pathogens can co-infect to cause urinary tract infections, skin infections, biofilm associated infections, sepsis and pneumonia. Infections of these two pathogens especially their biofilm associated infections are often difficult to treat using currently available anti-bacterial and anti-fungal agents. In order to identify a common anti-microbial agent which could confer a broad range of protection against their infections, we screened several phytochemicals and identified plumbagin (5-hydroxy-2-methyl-1,4-naphthoquinone), a phytochemical from Plumbago species as a potent antimicrobial agent against S. aureus and C. albicans, with a minimum inhibitory concentration of 5μg/ml. Antimicrobial activity of plumbagin was validated using an ex-vivo porcine skin model. For better understanding of the antimicrobial activity of plumbagin, a Drosophila melanogaster infection model was used, where D. melanogaster was infected using S. aureus and C. albicans, or with both organisms. The flys survival rate was dramatically increased when infected flies were treated using plumbagin. Further, plumbagin was effective in preventing and dispersing catheter associated biofilms formed by these pathogens. The overall results of this work provides evidence that plumbagin, possesses an excellent antimicrobial activity which should be explored further for the treatment of S. aureus and C. albicans infections.

Anacardic acid induces apoptosis-like cell death in the rice blast fungus Magnaporthe oryzae

Anacardic acid (6-pentadecylsalicylic acid), extracted from cashew nut shell liquid, is a natural phenolic lipid well known for its strong antibacterial, antioxidant, and anticancer activities. Its effect has been well studied in bacterial and mammalian systems but remains largely unexplored in fungi. The present study identifies antifungal, cytotoxic, and antioxidant activities of anacardic acid in the rice blast fungus Magnaporthe oryzae. It was found that anacardic acid causes inhibition of conidial germination and mycelial growth in this ascomycetous fungus. Phosphatidylserine externalization, chromatin condensation, DNA degradation, and loss of mitochondrial membrane potential suggest that growth inhibition of fungus is mainly caused by apoptosis-like cell death. Broad-spectrum caspase inhibitor Z-VAD-FMK treatment indicated that anacardic acid induces caspase-independent apoptosis in M. oryzae. Expression of a predicted ortholog of apoptosis-inducing factor (AIF) was upregulated during the process of apoptosis, suggesting the possibility of mitochondria dependent apoptosis via activation of apoptosis-inducing factor. Anacardic acid treatment leads to decrease in reactive oxygen species rather than increase in reactive oxygen species (ROS) accumulation normally observed during apoptosis, confirming the antioxidant properties of anacardic acid as suggested by earlier reports. Our study also shows that anacardic acid renders the fungus highly sensitive to DNA damaging agents like ethyl methanesulfonate (EMS). Treatment of rice leaves with anacardic acid prevents M. oryzae from infecting the plant without affecting the leaf, suggesting that anacardic acid can be an effective antifungal agent.

Anacardic acid inhibits gelatinases through the regulation of Spry2, MMP-14, EMMPRIN and RECK

Earlier studies from our laboratory have identified Anacardic acid (AA) as a potent inhibitor of gelatinases (MMP-2 and 9), which are over-expressed in a wide variety of cancers (Omanakuttan et al., 2012). Disruption of the finely tuned matrix metalloproteinase (MMP) activator/inhibitor balance plays a decisive role in determining the fate of the cell. The present study demonstrates for the first time, that in addition to regulating the expression as well as activity of gelatinases, AA also inhibits the expression of its endogenous activators like MMP-14 and Extracellular Matrix MetalloProteinase Inducer (EMMPRIN) and induces the expression of its endogenous inhibitor, REversion-inducing Cysteine-rich protein with Kazal motifs (RECK). In addition to modulating gelatinases, AA also inhibits the expression of various components of the Epidermal Growth Factor (EGF) pathway like EGF, Protein Kinase B (Akt) and Mitogen-activated protein kinases (MAPK). Furthermore, AA also activates the expression of Sprouty 2 (Spry2), a negative regulator of EGF pathway, and silencing Spry2 results in up-regulation of expression of gelatinases as well as MMP-14. The present study thus elucidates a novel mechanism of action of AA and provides a strong basis for utilizing this molecule as a template for cancer therapeutics.

Impregnation of Catheters with Anacardic Acid from Cashew Nut Shell Prevents Staphylococcus aureus Biofilm Development.

The effect of anacardic acid impregnation on catheter surfaces for the prevention of Staphylococcus aureus attachments and biofilm formations were evaluated.

Changing strategies in natural product research

T annual incidence of urolithiasis in industrialized regions is considered to be 1,500-2,000 cases per millions with reoccurrence rate of 75% in 20 years. There is no effective management therapy for renal calculi. Allopathic and herbal therapies havetheir inherent limitations and side-effects. Bergenia ligulata has been used since ancient time in many herbal compositions and it is major component of Cystone® (Himalaya, herbal healthcare) for treating kidney stones. The present work has been designed to study the anti-lithiatic potential of B. ligulata, isolation of the potent metabolite(s) and its mechanism of action. Commercially available dried rhizomes of B. ligulata were powdered and subjected to activity guided fractionation using in vitro calcium oxalate crystal growth inhibition assay. Further, rat hyperoxaluric model was used to assess anti-lithiatic ability in vivo. The isolated fractions showed anti-calcifying activity in vitro. The sequential isolation of the potent fraction led to the purification of the most active molecule. The metabolite was eventually characterized as bergenin employing LC-MS, NMR, FTIR and UV spectroscopy. Bergenin was found to be effective in reducing oxidative stressmarkers like malondialdehyde (187% in diseased vs. 20% in treated) and elevating reduced glutathione levels (-46% in diseased vs. -15% in treated). It exhibited anti-lithiatic activityas assessed by measuring the activity of lactate dehydrogenase and alkaline phosphatase in serum samples. The creatinine clearance was also normalized with bergenin treatment in rat hyperoxaluric model. The present study provides significant evidence in the effectiveness of bergenin in treating and managing renal calculi.D mellitus is a metabolic disorder constituting a major health concern today whose prevalence has continuously increased in the world. The aim of this study is to evaluate the anti-diabetic potential of methanolic extract of Hyoscyamus albus (HAMeOH) in diabetic rats. Hyoscyamus albus (Solanaceae) is an herbal medicine traditionally applied as a parasympatholytic and nervous sedative. The oral glucose tolerance test (OGTT) was carried out by administering glucose (2 g/kg, b.w), to non-diabetic rats treated with HAMeOH at oral doses 100 and 200 mg/kg, b.w and glibenclamide 5 mg/kg. Also, Streptozotocin-induced diabetic rats, these diabetic rats were administered (100 and 200 mg/Kg b.w ) and standard drugs glibenclamide was given to rats for 30 days. The oral administration of both doses of HAMeOH significantly reduced the levels of blood glucose and glycosylated hemoglobin in diabetic rats. Determination of plasma insulin levels revealed the insulin stimulating action of the leaves extract. It is concluded that HAMeOH have significant anti-diabetic activity.Grewia nervosa (Lour) Panighrahi, belonging to the family Malvaceae s.l. is widely distributed along the Western Ghats of India. Although it has been commonly used in traditional medicine, the medicinal properties have not been scientifically evaluated. Phytochemical analysis established the presence of phenolic compounds, tannins, alkaloids and saponins in leaves. The aqueous and methanol extracts from leaves and bark of G. nervosa were investigated for medicinal properties using in vitro assays. The methanol extract of leaves demonstrated 97.5% inhibition of α-amylase activity. Additionally, the methanol extract of leaves also demonstrated antioxidant activity (5.41± 0.23 mmol/g, dw) that was higher compared to aqueous extract (3.32±0.45 mmol/g, dw). Further the methanol extract of bark exhibited anti-lipoxygenase activity indicative of its potential to control inflammatory activity. These results suggest that Grewia nervosa would be a potential source for treatment of diabetes and its associated complications such as oxidative stress and inflammationC herbs (Asteraceae) are extensively used as food additives and in folk medicine. Anti-cancer, anti-human immunodeficiency virus type 1 (HIV-1), anti-inflammatory, antinociceptive and antiproliferative activities as well as antioxidant effects have been reported for Chrysanthemum species. We report the isolation and identification of flavonoids and new and known terpenoids from the endemic species, C. macrocarpum and C. deserticolum “guertoufa”, used in Algerian Sahara as tea drinks and in “couscous” and soups “Chorba”. Structures of the isolated compounds were established by 1-D and 2-D homo and hetero-nuclear NMR (1H, 13C, COSY, HSQC, HMBC, and NOESY), mass spectrometry, UV and comparison with literature data. C. deserticolum extracts were tested by four methods to identify the antioxidant activity namely, ABTS•+, DPPH• scavenging, CUPRAC and ferrous-ions chelating activity methods. The in vitro anticholinesterase activity was achieved by the use of the basic enzymes that occur in causing Alzheimer’s disease: acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). Anti-inflammatory, antinociceptive, antiproliferative and antioxidant activities of C. macrocarpum extracts and isolated compounds are also reported here.T practice of traditional medicine for the control of fertility in most parts of India is based on the uses of plant medicines for many years. The aim of the present study was to evaluate the post coital antifertility activity of different varieties of Curcuma longa and underlying mechanism thereof. The effects of n-hexane, chloroform and acetone fraction of hydroalcoholic extract of three different varieties i.e. Salem, Krishna and Rajapore of Curcuma longa rhizomes were studied at three different doses to evaluate their antifertility, early abortifacient, antiovulatory activities and underlying mechanism thereof. Acute toxicity and thin layer chromatographic studies of same were also carried out. All varieties found to have significant antifertility activity (p<0.01). The n-hexane fraction of all varieties showed anti-implantation activity at the dose of 150 mg/kg weight whereas chloroform and acetone fraction of all varieties exhibited 100% reduction in pregnancy at the dose of 300 mg/kg body weight. Antifertility activity of test drugs was found through its antizygotic mechanism whereas all test drugs devoid of any antiovulatory and early abortifacient activity at all doses.M charantia (MC) fruits have previously been reported to reduce blood glucose in laboratory animals and human subjects with diabetes. Increase in insulin secretion is one of the chief mechanisms of antidiabetic action of MC extracts or their purified molecules. In present study the effect of aqueous extracts of MC (AEMC) was studied on insulin secretion in isolated pancreatic islets from normal Wistar rats with an attempt to evaluate the mechanism of action. Islets were incubated in HBBS buffer containing 3.3 or 16.7mM glucose, and AEMC, diazoxide, nimodipine and calphostin C, alone and in combinations. Release of insulin in external media was measured by ELISA. Cytotoxicity studies, to assess the integrity of the islets cells, were carried out by trypan blue uptake and LDH release assay. Trypan blue gained access to 9.6 ± 1.2% cells and 8.3 ± 1.1% dead islet cells were observed in LDH release assay on AEMC exposure, suggesting that the extract was non-toxic at tested concentration. AEMC stimulated insulin secretion from the isolated islets at 3.3 and 16.7 mM glucose. The effect of AEMC was dose dependent. As loss of cell integrity was not observed on AEMC exposure, hence, alteration of membrane integrity as the possible mechanism of insulin release is ruled out by this study. Addition of dizoxide and nimodipine completely diminished glucose induced insulin secretion. AEMC induced insulin secretion at 16.7mM glucose was partially inhibited by dizoxide and nimodipine, however no reduction was observed at 3.3mM of glucose. No change in insulin secretion at basal level of 3.3 mM of glucose suggests that the phytochemicals of AEMC may not be binding to either KATP or Ca channels. Calphostin C significantly (p<0.01) reduced AEMC induced insulin production both at 3.3 mM and 16.7 mM. The finding suggests that PKC inducing activity of AEMC phytochemical/s may be responsible for its insulin secretagogues potential.Bioassay-guided fractionation of the CH2Cl2/MeOH extract of the Thai marine sponge Acanthodendrilla sp. resulted in the isolation of six bromotyrosine-derived alkaloids; aerothionin (1), homoaerothionin (2), 2-hydroxy-3,5-dibromo,4methoxyphenylacetamide (3), 2,4-cyclohexadiene-1-acetamide-3,5-dibromo-1,6-dihydroxy-4-methoxy (4), 11-oxoaerothionin (5), and 11,19-dideoxyfistularin (6) . The structures of the isolated compounds were identified on the basis of detailed spectroscopic analysis. The compounds were tested for the acetylcholinesterase-inhibiting activity, and 3 showed the best acetylcholinesterase-inhibiting activity (92.0% at 0.1 mg/mL).T hepatoprotective activity of methanolic extract of bark of Ficus bengalensis against paracetamol and CCl4 induced liver damage was investigated. Treatment of rats with paracetamol and CCl4 produced a significant increase in the levels of serum glutamate pyruvate transaminase SGPT, serum glutamate oxaloacetate transaminase SGOT, alkaline phosphatase ALP, total and direct bilirubin. Rats pretreated with methanolic extract of barks of F. bengalensis 100 and 250 mg/kg body weight p.o. exhibited rise in the levels of these enzymes but it was significantly less as compared to those treated with paracetamol or CCl4 alone. The results of methanolic extract of F. bengalensis were comparable with the standard hepatoprotective agent silymarin 100 mg/kg. Maximum hepatoprotective effect was found to be at the dose of 250 mg/kg body weight in case of CCl4 induced hepatic damage while 500 mg/kg body weight in case of paracetamol induced hepatic damage. Obtained data suggest that methanolic extract of F. bengalensis bark possesses a potential antihepatotoxic activity.The medicinal quality of plants has been known and exploited by man for centuries. A large number of modern drugs have been isolated from traditional herbal plants[1]. Numerous secondary metabolites obtained from plants, with previously unknown pharmacological activities, have been extensively investigated as a source of medicinal agents[1,2]. The acceptance of traditional medicine as an alternative form of health care and the development of microbial resistance to the available antibiotics led to investigation on the antimicrobial activity of medicinal plants. The increasing failure of chemotherapeutics and antibiotic resistance exhibited by pathogenic agents has led to the screening of several PEER REVIEW ABSTRACTT genus of Phoebe of family Lauraceae is found the most abundance in Borneo and the Malaysian Peninsular. Phoebe tavoyana is locally known as ‘medang rungkoi.’ The woods of Phoebe species have the commercial values usually for housebuilding. As a wood of a good type soft to moderately hard, light, slightly colored than the hardwood used for carving and sculpture, paneling for doors altars wardrobes, carriages and ceiling. Phytochemical study on the leaves of Phoebe tavoyana (Meissn.) H.K.F. from Chebar Besar Reserved Forest, Kuala Kangsar, Perak, Malaysia has resulted the isolation of four known aporphines; laurolitsine (1), roemerine (2), laetanine (3), boldine (4) and one morphinandienone type, sebiferine (5). The structures of alkaloids were determined by spectroscopic analysis. This paper reports the antiplasmodial activity of three alkaloids from the leaves of Phoebe tavoyana (Lauraceae). The results showed that (1), (2) and (5) have shown potent inhibitory activity against the growth of Plasmodium falciparum 3D7 clone, with IC50 1.49, 0.89 and 2.76μg/mL respectively. No previous phytochemical investigation has been performed on this plant.Natural polysaccharides have been widely used because of their biocompatibility and biodegradabilityproperties. An attempt has been made to explore tamarind seed xyloglucan (TSX), a glucosaminoglycan polysaccharide extracted from the kernels of seeds of Tamarindus indica Linn., family Fabaceae for bimodal(immediate and controlled)drug release of multilayer tablet. Chemically TSX powder is highly branched carbohydrate polymer. High drug holding capacity of this polysaccharide was investigated for bimodal release.An in-house extracted TSX polysaccharide was characterized for swelling index, flow property, viscosity and compatibility with drug. Multilayer tablet was comprised of immediate release layer of tramadol hydrochloride (an analgesic agent), followed by tri-layer. This tri-layer consisted of upper and lower barrier layers of TSX and middle layer of drug granular matrix. Multilayer tablets were compressed based on 3 2 factorial design consideringconcentrations of matrix and barrier TSX layers as independent variables. Immediate release layer released the drug within 90 min in acidic media, revealing the retarded action showed bypolysaccharide layer attached to this layer. Granules of matrix layer were prepared by wet granulation technology.Multiplayer tablet of TSX was evaluated for hardness, thickness and drug content. Dissolution test in presence of rat caecal content was found to control the drug release rate for more than 9h. Stability studies confirmed the stable formulation. Thus, this study suggested that inexpensiveand abundantly available natural TSX can act as a potential polymer for bimodal releaseof a multilayer tablet.