Antibacterial, cytotoxicity and biodegradability studies of polycaprolactone nanofibers holding green synthesized Ag nanoparticles using atropa belladonna extract.

Atropa belladonna is one of the herbs used to treat wounds and prevent inflammation. This study provides a scientific assessment for the wound healing potential of biodegradable nanofibers containing Ag nanoparticles encapsulated with atropa belladonna extract (eAgNPs). Ultraviolet-visible (UV-vis) spectroscopy was used to observe the localized surface plasmon resonance (LSPR) band of AgNPs synthesized from atropa belladonna extract prepared under different conditions. Polycaprolactone (PCL) nanofibers with eAgNPs were fabricated using the electrospinning technique. The distribution of AgNPs and eAgNPs and the size of nanofibers were characterized with scanning and transmission electron microscopy (SEM, TEM) before and after degradation at the end of 18 weeks. Fourier transform infrared (FTIR) spectroscopy showed the surface interactivity between AgNPs, atropa belladonna extract and PCL nanofibers and also approved the modification of PCL nanofibers with eAgNPs. X-ray diffraction analysis (XRD) defined the formation of the crystalline AgNPs and appreciated the orientation of the nanofibers. Results of tension tests revealed that modification of PCL nanofibers with pure AgNPs and eAgNPs significantly increased strength and tensile modulus. Due to the hydrophobic nature of PCL, modification with pure AgNPs and eAgNPs slightly reduced its hydrophobicity. Biodegradation tests of PCL nanofibers with eAgNPs exhibited a higher degradation rate than neat PCL nanofibers. In vitro MTT results revealed that eAgNPs doped PCL samples have better cell viability than AgNPs doped and neat PCL nanofibers. Owing to their antibacterial properties, biodegradation rates, low cytotoxicity, mechanical and surface morphologic properties of AgNPs modified PCL nanofibers containing atropa belladonna are considered to have a great potential for skin regeneration.

Methyl-ß-cyclodextrin and coronatine as new elicitors of tropane alkaloid biosynthesis in Atropa acuminata and Atropa belladonna hairy root cultures.

Hyoscyamine (HYO) and scopolamine (SCO) are tropane alkaloids acting as anticholinergic factors on the parasympathetic nervous system in humans and are produced by Solanaceous plants. Two strains of Agrobacterium rhizogenes, A4 and LBA9402, were used to infect Atropa acuminata Royle ex Miers and Atropa belladonna L. leaf explants. A. acuminata was inoculated either by direct infection or sonicated-assisted A. rhizogenes-mediated transformation (SAAT) was performed. A. belladonna was inoculated with the A4 strain using a direct method. The selected hairy root lines of both species were elicited with 50 mM methyl-ß-cyclodextrin (ß-CD), 0.5 µM coronatine (Cor) or 50 mM ß-CD + 0.5 µM Cor on Day 14 of culture. The elicitor effect on growth and HYO and SCO content was analyzed after one (T1) and two (T2) weeks of treatment. In A. acuminata explants, the highest transformation percentage (T%) was obtained with strain A4 and the SAAT method (T%: 96.43). Cor significantly reduced the growth of A. acuminata hairy roots (fresh weight and dry weight [DW]: 2.52 and 0.3 g, respectively), whereas ß-CD increased their DW (0.4 g). Also, the combined ß-CD + Cor treatment had a positive significant effect on the DW of A. belladonna hairy roots (0.41 g). In A. acuminata hairy roots, the HYO level was lower under Cor treatment than in the control at both sampling times. In contrast, the SCO content was increased 10-fold by Cor elicitation at T1 compared to the control (10.95 mg g-1 DW) and was also positively affected by ß-CD + Cor. In A. belladonna hairy roots, all the elicitors had a negative effect on both HYO and SCO production. This report is the first assessment of the effect of ß-CD and Cor elicitors on tropane alkaloid production.

In vitro biological screening of a critically endangered medicinal plant, Atropa acuminata Royle Ex Lindl of north western Himalaya.

Atropa acuminata Royle Ex Lindl (Atropa acuminata) under tremendous threat of extinction in its natural habitat. However, the antimicrobial, antileishmanial and anticancer effects of the plant's extracts have not been reported yet. In the current study, an attempt has been made to evaluate the pharmacological potential of this plant's extracts against microbes, Leishmania and cancer. The roots, stems and leaves of Atropa acuminata were ground; then, seven different solvents were used alone and in different ratios to prepare crude extracts, which were screened for pharmacological effects. The aqueous, methanolic and ethanolic extracts of all parts carried a broad spectrum of anti-bacterial activities, while no significant activity was observed with combined solvents. Three types of cytotoxicity assays were performed, i.e., haemolytic, brine shrimp and protein kinase assays. The aqueous extract of all the parts showed significant haemolytic activity while n-hexane extracts of roots showed significant activity against brine shrimp. The acetone extracts strongly inhibited protein kinase while the methanolic extracts exhibited significant cytotoxic activity of roots and stem. The anti-leishmanial assays revealed that the methanolic extract of leaves and roots showed significant activity. These findings suggest that this plant could be a potential source of natural product based drugs.

Microclimatic variation in UV perception and related disparity in tropane and quinolizidine alkaloid composition of Atropa acuminata, Lupinus polyphyllus and Hyoscyamus niger.

The aim of current research was to evaluate the physiological adjustment in three medicinal herbs viz., Atropa acuminata, Lupinus polyphyllus and Hyoscyamus niger to the winter period characterised by intense UV flux in Kashmir valley across the North Western Himalaya. Quinolizidine (QA) and tropane alkaloid (TA) concentrations were analysed in these herbs thriving at two different altitudes via GC-MS and correlated by PCA analysis. This study investigated the hypothesis that UV reflectance and absorbance at low temperatures are directly related to disparity in alkaloid accumulation. Among QAs in L. polyphyllus, ammodendrine and lupanine accumulated at higher concentration and exhibited significant variation of 186.36% and 95.91% in ammodendrine and lupanine respectively in both sites. Tetrahydrohombifoline displayed non-significant variation of about 9.60% irrespective of sites. Among tropane alkaloid (TA), hyoscyamine was recorded as the most abundant constituent irrespective of the plant and site while apotropine accumulated in lesser quantity in A. acuminata than H. niger. However, apotropine demonstrated significant variation of 175% among both sites. The final concentration of quinolizidine (QA) and tropane alkaloid (TA) reflects the interplay between reflectance and absorbance of UV radiation response field. These findings suggest that spectral response of UV light contributes directly to alkaloid biosynthesis.

Modulation of T-helper cytokines and inflammatory mediators by Atropa accuminata. Royle in adjuvant induced arthritic tissues.

ETHNOPHARMACOLOGICAL RELEVANCE: Atropa acuminata has been widely used in traditional medicine against arthritis and several associated inflammatory disorders. AIM OF THE STUDY: The present study was undertaken to investigate the anti-arthritic activities of ethanolic extract of Atropa accuminata (AAEE) and to explore the probable mechanism of action. MATERIALS AND METHODS: The anti-arthritic activity of AAEE was evaluated within a dose range of 125-500 mg/kg b.w. in adjuvant induced-arthritis in male wistar rats. An array of pro-inflammatory mediators (PGE2 NO, IL-1ß and LTB4) and T-cell-mediated cytokines (IL-2, TNF-a, IFN-c, IL-4, IL-10, IL-12, IL-17, IL-6) were assayed in arthritic paw tissue homogenate of the treated animals. In addition the effects on arthritic lesions, changes in body weight; haematological (Hb, ESR, WBC and RBC) and biochemical parameters (SOD, GSH, GR) and the serum markers (CRP, RF) were also observed. RESULTS: Significant anti-arthritic activity was observed for AAEE in the polyarthiritis test both in the developing and developed phase of the disease. This was associated with dose dependant suppression of pro-inflammatory mediators (PGE2, NO, IL-1ß and LTB4)., Th1-Th17 cytokines (IL-2, TNF-α, IFN-γ, IL-12, IL-17, IL-6) and upregulation of Th2 cytokines (IL-4 and IL-10). AAEE was also observed to protect rats against the primary and secondary arthritic lesions, body weight changes and haematological perturbations. In addition, inhibitory effects of AAEE on biochemical parameters and the serum markers further confirmed that it reduced signs on chronic inflammatory responses. CONCLUSION: The present investigation therefore suggested that AAEE is a potent anti-arthritic agent. The multipronged attack on the inflammatory mediators and T-helper cytokines and strong potency of AAEE may have relevance for inhibition of the chronic inflammatory responses in arthritis.

Intra- and inter-laboratory validation of a dipstick immunoassay for the detection of tropane alkaloids hyoscyamine and scopolamine in animal feed.

Tropane alkaloids (TAs) are toxic secondary metabolites produced by plants of, inter alia, the genera Datura (thorn apple) and Atropa (deadly nightshade). The most relevant TAs are (-)-L-hyoscyamine and (-)-L-scopolamine, which act as antagonists of acetylcholine muscarinic receptors and can induce a variety of distinct toxic syndromes in mammals (anti-cholinergic poisoning). The European Union has regulated the presence of seeds of Datura sp. in animal feeds, specifying that the content should not exceed 1000 mg kg(-1) (Directive 2002/32/EC). For materials that have not been ground, visual screening methods are often used to comply with these regulations, but these cannot be used for ground materials and compound feeds. Immunological assays, preferably in dipstick format, can be a simple and cost-effective approach to monitor feedstuffs in an HACCP setting in control laboratories. So far no reports have been published on immunoassays that are capable of detecting both hyoscyamine and scopolamine with equal sensitivity and that can be used, preferably in dipstick format, for application as a fast screening tool in feed analysis. This study presents the results obtained for the in-house and inter-laboratory validation of a dipstick immunoassay for the detection of hyoscyamine and scopolamine in animal feed. The target level was set at 800 µg kg(-1) for the sum of both alkaloids. By using a representative set of compound feeds during validation and a robust study design, a reliable impression of the relevant characteristics of the assay could be obtained. The dipstick test displayed similar sensitivity towards the two alkaloids and it could be concluded that the test has a very low probability of producing a false-positive result at blank level or a false-negative result at target level. The assay can be used for monitoring of TAs in feedstuffs, but has also potential as a quick screening tool in food- or feed-related poisonings.

Multi-development-HPTLC method for quantitation of hyoscyamine, scopolamine and their biosynthetic precursors in selected solanaceae plants grown in natural conditions and as in vitro cultures.

INTRODUCTION: Hyoscyamine and scopolamine, anti-cholinergic agents widely used in medicine, are typically obtained from plants grown under natural conditions. Since field cultivation entails certain difficulties (changeable weather, pests, etc.), attempts have been made to develop a plant in vitro culture system as an alternative source for the production of these compounds. During experiments to locate the limiting steps in the biotechnological procedure, it is important to monitor not only the levels of the final products but also the changes in the concentration of their precursors. OBJECTIVE: To develop a HPTLC method for the separation and quantitation of the main tropane alkaloids hyoscyamine and scopolamine, their respective direct precursors littorine and anisodamine, and cuscohygrine, a product of a parallel biosynthetic pathway that shares a common precursor (N-methyl-∆(1) -pyrrolium cation) with tropane alkaloids. METHODS: Using alkaloid extracts from Atropa baetica hairy roots, different TLC chromatographic systems and developing procedures were investigated. RESULTS: Full separation of all compounds was obtained on HPTLC Si60 F254 plates preconditioned with mobile phase vapours (chloroform:methanol:acetone:25% ammonia ratios of 75:15:10:1.8, v/v/v/v). The chromatograms were developed twice (at distances of 4.0 and 3.0 cm) in a Camag twin trough chamber and visualised with Dragendorff's reagent. Densitometric detection (λ = 190 and 520 nm) was used for quantitative analyses of the different plant samples. CONCLUSION: This method can be recommended for quantitation of hyoscyamine, scopolamine, anisodamine, littorine and cuscohygrine in different plant material (field grown vs. in vitro cultures).

Atropa acuminata Royle Ex Lindl. blunts production of pro-inflammatory mediators eicosanoids., leukotrienes, cytokines in vitro and in vivo models of acute inflammatory responses.

ETHNOPHARMACOLOGICAL RELEVANCE: Atropa acuminata Royle Ex Lindl. has been widely used in folk medicine for several inflammatory disorders such as arthritis, asthma, conjunctivitis, encephalitis, pancreatitis, peritonitis, acute infections and neuroinflammatory disorders. AIM OF THE STUDY: Our aim was to evaluate Atropa acuminata for its anti-inflammatory properties and to delineate its possible mechanism of action on the modulation of the inflammatory mediators. MATERIALS AND METHODS: We investigated the inhibitory action of ethanolic extract of Atropa acuminata (AAEE) on production of NO, TNF-α and IL-1ß in lipopolysaccharide (LPS)-stimulated RAW264.7 cells and also assayed it for COX 1/2 and 5-LOX inhibitory activities. Next AAEE was tested in acute inflammatory animal models., carragenean induced rat paw edema, carragenean induce pleurisy in rats and vascular permeability in mice and the effects on NO, PGE2 and LTB4 production in the pleural fluid and paw exudates were evaluated. In addition the effects on leukocyte migration and exudation and vascular permeability were also observed. RESULTS: Our findings summarized novel anti-inflammatory mechanisms for Atropa acuminata based on dual in vitro cyclooxygenase 1/2/ and 5-Lipoxygenase inhibitory activities and also significant downregulation of nitric oxide (NO) and pro-inflammatory cytokin (TNF-α and Il-1 ß) release in LPS-stimulated RAW 246.7 macrophage cell line. In acute inflammatory models in vivo (carragenean induced edema, carragenean induced pleurisy in rats and vascular permeability in mice), AAEE exhibited an extensive diverse mechanism for anti-inflammatory properties. This was indicated on the basis of dose dependent suppression of multi targeted inflammatory mediators., NO, TNF-α and IL-1ß, eicosanoids., PGE2 and leukotrienes., LTB4 along with significantly decreased leucocyte migration, exudation and decreased vascular permeability. These effects were more potent and prolonged than traditional NSAIDS, thereby indicating fewer side effects. AAEE was found to be safe for long term administration, as confirmed by the results of acute toxicity studies and MTT assay. The complex mode of action of the herbs was attributed possibly due to the high polyphenolic, flavanol and flavonoid content present in the extracts as observed by means of quantitative screening for phytochemicals. CONCLUSION: Our study provides scientific evidence to support the traditional anti-inflammatory uses of Atropa acuminata and is probably due to inhibitory effects on multiple inflammatory mediators which indicates a promising potential for the development of a strong anti-inflammatory agent from this plant.

Nectar yeasts of two southern Spanish plants: the roles of immigration and physiological traits in community assembly.

Recent studies have shown that dense yeast populations often occurring in floral nectar are numerically dominated by a few species from the flower-insect interface specialized genus Metschnikowia, while generalist yeast species commonly occurring on leaf surfaces, soil, freshwater, and air were rarely isolated from nectar samples. This study was designed to understand the main factors responsible for the assembly of nectar yeast communities, by combining field experiments with laboratory tests characterizing the physiological abilities of all yeast species forming the pool of potential colonizers for two Spanish flowering plants (Digitalis obscura and Atropa baetica). Yeast frequency and species richness were assessed in external sources (bee glossae, air, plant phylloplane) as well as in pollinator rewards (pollen, nectar). Yeasts were most frequent in external sources (air, flower-visiting insects), less so in the proximate floral environment (phylloplane), and least in pollen and nectar. Nectar communities appeared to be considerably impoverished versions of those in insect glossae and phylloplane. Nectar, pollen, and insect yeast assemblages differed in physiological characteristics from those in other substrates. Nectarivorous Metschnikowia were not more resistant than other yeast species to plant secondary compounds and high sugar concentrations typical of nectar, but their higher growth rates may be decisive for their dominance in ephemeral nectar communities.

In vitro examination of potentized atropine sulfate dilutions on the contractility of the isolated rat ileum.

OBJECTIVES: Atropine sulphate, a competitive antagonist of acetylcholine (ACh) at muscarinic receptors, was first isolated from Atropa belladonna, one of the most used and best known homeopathic medicines. It has been suggested that high potencies of homeopathic atropine sulphate might have an influence on ACh-induced contraction of smooth muscles. The aim of the study was to determine the effects of homeopathic dilutions of atropine sulphate D6, D32, and D100 compared to the potentized diluents L6, L32, and L100 on ACh-induced contraction of isolated rat ileum. DESIGN: Forty-eight (48) ileal sections from 12 male Wistar rats were incubated in modified Krebs solutions, and the contractile activity responses to ACh obtained in the absence and presence of the test substances were recorded. Investigators and biometrician were completely blinded. RESULTS: No significant effects of atropine sulphate D6, D32, or D100 could be found (all p > 0.4 after Bonferoni-Holm correction) compared to the potentized diluents L6, L32, and L100, respectively. These figures did not change considerably even when strict a priori criteria were applied that define a measurement as valid and comparable. CONCLUSIONS: Our experiments could not replicate previous results on the effects of homeopathic atropine.

Cloning, characterization and analysis of expression profiles of a cDNA encoding a hyoscyamine 6beta-hydroxylase (H6H) from Atropa baetica Willk.

A new cDNA encoding hyoscyamine 6beta-hydroxylase (H6H, EC 1.14.11.11), a bifunctional enzyme catalyzing the last two steps in the biosynthesis of scopolamine, was isolated from Atropa baetica roots (GenBank accession no. EF442802). The full cDNA sequence showed an ORF of 1035bp, coding for a protein with 344 amino acid residues. Sequence analyses at the nucleotide level showed that this ORF shares high identity with other H6H from different plant species, such as Anisodus tanguticus and Hyoscyamus niger with 90% identity, and an almost total identity with A. belladonna (98%). Tissue expression analyses showed that the gene transcript was tissue dependent, appearing exclusively in roots, thus being the only biosynthetic site for the production of scopolamine. Furthermore, Southern hybridization experiments revealed that this gene was not part of a multigene family as appears in low copy number. Phylogenetic tree analysis indicated that A. baetica H6H had a very close relationship with A. belladonna and to a lesser extent with H. niger.

Effects of elicitors on tropane alkaloids and gene expression in Atropa baetica transgenic hairy roots.

Elicitation of transgenic Atropa baetica overexpressing the h6h gene with salicylic acid (SA), acetylsalicylic acid (ASA),or methyl jasmonate (MeJ) was conducted to boost tropane alkaloid yields. Scopolamine (1) amounts increased after treatment with ASA and MeJ, but not with SA. The highest enhancement of 1 was achieved with MeJ followed by ASA dissolved in EtOH. Transcriptomic analyses showed a direct relationship between content of 1 and gene expressions;the engineered h6h gene and other biosynthetic genes were stimulated. ASA dissolved in EtOH showed a high h6h gene expression, increasing 25-fold and 5-fold compared to controls; tr-I also displayed a 5-fold increase. The controls to which EtOH was added showed a 5-fold increase in h6h gene expression and 125-fold for pmt, demonstrating that EtOH also functioned as an enhancer of 1. MeJ was the best elicitor, displaying a 25-fold increase in h6h expression level, not affecting the expression of the other three genes analyzed, and it appears to possibly stimulate the phenylpropanoids branch of the tropane alkaloid pathway.

Enhanced secretory activity of Atropa belladonna hairy root culture over-expressing ADP-ribosylation factor gene.

Agrobacterium-mediated transformation of leaf tissues of Atropa belladonna with an adenosine 5'-diphosphate (ADP)-ribosylation factor gene of carrot, arf-001, was performed employing pBCR82 as an expression vector. This vector co-expresses rol gene cluster together with arf-001, and thus, the transformed host cells were obtained as hairy roots. Two cell lines of the transformed belladonna were established as the liquid cultures of hairy root tissues, and expression of arf-001 and accumulation of its product in the cells were confirmed by RT-PCR and Western blot analyses, respectively. A marked increase in extracellular protein concentrations was observed in the transformed belladonna root cultures as compared with the controls transformed with an empty vector. However, the secretion of the proteins of the transformants was markedly reduced in the presence of a physiological concentration of monensin. These results suggest that over-expression of arf-001 in belladonna results in the enhancement of secretory activity in the transformed cells.

Expression of tropane alkaloids in the hairy root culture of Atropa acuminata substantiated by DART mass spectrometric technique.

Agrobacterium rhizogenes-mediated 'hairy root' cultures were established in Atropa acuminata. The chemical profiling of the hairy roots was carried out by a new mass spectrometric technique, direct analysis in real time (DART). The intact hairy roots were directly analyzed by holding them in the gap between the DART ion source and mass spectrometer. Two alkaloids, atropine and scopolamine, were characterized. The structural confirmation of the two alkaloids was made through their accurate molecular formula determinations. This is the first report of establishing hairy roots in A. acuminata as well as application of the DART technique for the chemical profiling of its hairy roots.

Improvement of sexual destination in Atropa acuminata Royle (Solanaceae)--a critically endangered medicinal plant of Northwestern Himalaya.

Good seed set is no guarantee of absolute sexual destination in plants. Seed viability and seed vigour are crucial phases in the life cycle of every sexually reproducing plant. The present study was an attempt to improve the sexual destination-the germination and seedling survival of Atropa acuminata Royle (Solanaceae), an endemic and extremely restricted sub-alpine medicinal plant of North West Himalayas under ex situ conditions at (1580 m) with an aim to develop a successful germination protocol and agrotechnique in order to revegetate disturbed areas. Among various treatments given to the seeds, GA3, Scarification, warm water treatment and chilling at 4 degrees C for 90 days were found to be most effective with percentage germination of 73.3 +/- 18.80, 79.95 +/- 9.40, 66.6 +/- 6.6, 45 +/- 7.07 (X +/- SE), respectively. The results reveal that the seeds do not germinate unless specific environmental signals or events occur which trigger the genetic and hormonal response of the seeds thereby facilitating their germination. The diversity and the extent of the dormancy mechanisms encountered here suggest that under harsh conditions, natural selection may favour seeds with a genetic system for dormancy and delayed germination. A relation was observed between seed size/weight, % age germination and subsequent seedling survival. Seedling survival is also effected by specific habitat requirement and stiff intra and inter-specific competition particularly the whimsical behaviour of Sambucus wigthiana (an alien species which grows in the vicinity of Atropa) is beyond the ken of Atropa, adding fuel to the already burning candle apart from habitat fragmentation and herbivory.

Tailoring tropane alkaloid accumulation in transgenic hairy roots of Atropa baetica by over-expressing the gene encoding hyoscyamine 6beta-hydroxylase.

Atropa baetica hairy roots, over-expressing cDNA from Hyoscyamus niger encoding the gene for hyoscyamine 6beta-hydroxylase (H6H), were produced by Agrobacterium rhizogenes infection. The transgenic roots over-expressing h6h had an altered alkaloid profile in which hyoscyamine was entirely converted into scopolamine. In the best h6h clone, scopolamine accumulation increased 9-fold compared to plants, amounting to 5.6 mg g dry wt(-1), some of which was released into the liquid medium. Only negligible amounts of hyoscyamine were detected. In contrast, the gus control culture contained a much higher amount of hyoscyamine than scopolamine, mimicking the situation in the plant. At the molecular level, a higher conversion of hyoscyamine into scopolamine was related to a higher level of h6h mRNA; in some instances this was 5 - 10-fold higher.

Studies on tropane alkaloid extraction by volatile organic solvents: dichloromethane vs. chloroform.

In order to investigate the production of tropane alkaloids by hairy roots of Atropa baetica, transgenic for the gene h6h encoding the enzyme hyoscyamine 6beta-hydroxylase, solvent extraction with chloroform and with dichloromethane of the metabolites present in the liquid medium and in the root tissue was compared. The extraction of scopolamine from the liquid medium was equally effective with either solvent, giving maximum values of around 850 microg/flask. For the roots, three different extraction methods were employed: A, employing chloroform:methanol: (25%) ammonia (15:5:1) for initial extraction, followed by treatment with sulfuric acid and ammonia, and using chloroform for the final extraction and washes; B, as A but using dichloromethane for extraction and washes; and C, as B but substituting chloroform for dichloromethane in the extraction cocktail. Scopolamine was the most abundant metabolite (present in amounts of 3250-3525 microg/g dry weight) and presented similar extraction efficiencies with all of the extraction methods employed. The highest amounts of hyoscyamine and the intermediate 6beta-hydxoxyhyoscyamine were present on day 31 (800 and 975 microg/g dry weight, respectively) and no statistical differences between the three extraction methods employed were detected. This study confirms that, for the extraction of tropane alkaloids, dichloromethane can replace the commonly employed chloroform, the use of which incurs major health, security and regulation problems.

Highly oxygenated triterpenes from the roots of Atropa acuminata.

Two new oleanane triterpenes; 2alpha,3alpha,24-trihydroxyolean-12-ene-28,30-dioic acid ([structure: see text]) and 2alpha,3alpha,24,28-tetrahydroxyolean-12-ene ([structure: see text]) have been isolated from the roots of Atropa acuminata. Anti-oxidant p-hydroxyphenethyl trans-ferulate ([structure: see text]), beta-sitosterol-3-O-beta-D-glucopyranoside ([structure: see text]) and oleanolic acid ([structure: see text]) have also been reported for the first time from this species. The structures were determined by spectroscopic studies including 2D-NMR.