Hybanthus Enneaspermus Research Articles

Valorization of the Endogenous Knowledge of Plants Used in the Treatment of Female Infertility in the District of Abomey-Calavi in Benin

The child is the foundation for the consolidation of a couple's home in the world. Thus, the absence of the fruit of the union (child) in the couple is subject to criticism and is a source of problems between the spouses. The first cause cited is female infertility. This study aims at solving the problems of female infertility through the knowledge and valorization of plants used for its treatment. The methodology used consisted of an ethnobotanical survey based on a pre-established questionnaire and complemented by individual semi-structured interviews with the target group. A total of 96 people were interviewed. Correspondence Factorial Analysis (CFA) using R software was used to establish the relationships between ethnic groups and their knowledge of female infertility. The calculation of ethnobotanical indices allowed the diversity and importance of plants to be assessed. The study population has specific knowledge about the symptoms of the causes of a woman's infertility. A diversity of 61 plant species belonging to 35 botanical families was identified. The most represented families are: Anonnaceae, Euphorbiaceae, Leguminoseae-Mimosoideae and Rubiaceae. Also, the plants: Xylopia aethiopica, Monodora myristica, Syzygium aromaticum, Allium cepa and Hybanthus enneaspermus are frequently used in recipes for the treatment of female infertility symptoms. This work constitutes a database for other studies on pharmacology, toxicology of useful plants and their valorisation.

Evaluation of the in Vivo Aphrodisiac Activity of a Cyclotide Extract from Hybanthus enneaspermus.

Hybanthus enneaspermus is an Indian folk medicinal herb that has been widely used as a libido enhancer. This plant belongs to the Violaceae plant family, which ubiquitously contains disulfide-rich cyclic peptides named cyclotides. Cyclotides are an expanding plant-derived peptide family with numerous interesting bioactivities, and their unusual stability against proteolysis has attracted much attention in drug design applications. Recently, H. enneaspermus has been reported to be a rich source of cyclotides, and hence, it was of interest to investigate whether cyclotides contribute to its aphrodisiac activity. In this study, we evaluated the in vivo aphrodisiac activity of the herbal powder, extract, and the most abundant cyclotide, hyen D, extracted from H. enneaspermus on rats in a single dose regimen. After dosing, the sexual behaviors of male rats were observed, recorded, analyzed, and compared with those of the vehicle group. The results show that the extract and hyen D significantly decreased the intromission latency of sexually naïve male rats and the extract improved a range of other measured sexual parameters. The results suggest that the extract could enhance libido as well as facilitate erectile function in male rats and that the cyclotide hyen D could contribute to the libido-enhancing activity of this ethnomedicinal herb.

Discovery and mechanistic studies of cytotoxic cyclotides from the medicinal herb Hybanthus enneaspermus

Cyclotides are plant-derived peptides characterized by an ∼30-amino acid-long cyclic backbone and a cystine knot motif. Cyclotides have diverse bioactivities, and their cytotoxicity has attracted significant attention for its potential anticancer applications. Hybanthus enneaspermus (Linn) F. Muell is a medicinal herb widely used in India as a libido enhancer, and a previous study has reported that it may contain cyclotides. In the current study, we isolated 11 novel cyclotides and 1 known cyclotide (cycloviolacin O2) from H. enneaspermus and used tandem MS to determine their amino acid sequences. We found that among these cyclotides, hyen C comprises a unique sequence in loops 1, 2, 3, 4, and 6 compared with known cyclotides. The most abundant cyclotide in this plant, hyen D, had anticancer activity comparable to that of cycloviolacin O2, one of the most cytotoxic known cyclotides. We also provide mechanistic insights into how these novel cyclotides interact with and permeabilize cell membranes. Results from surface plasmon resonance experiments revealed that hyen D, E, L, and M and cycloviolacin O2 preferentially interact with model lipid membranes that contain phospholipids with phosphatidyl-ethanolamine headgroups. The results of a lactate dehydrogenase assay indicated that exposure to these cyclotides compromises cell membrane integrity. Using live-cell imaging, we show that hyen D induces rapid membrane blebbing and cell necrosis. Cyclotide-membrane interactions correlated with the observed cytotoxicity, suggesting that membrane permeabilization and disintegration underpin cyclotide cytotoxicity. These findings broaden our knowledge on the indigenous Indian herb H. enneaspermus and have uncovered cyclotides with potential anticancer activity.

P0995EVALUATION OF ANTIDIABETIC, ANTIOXIDANT AND NEPHROPROTECTIVE ACTIVITY OF FLAVONOID RICH FRACTION OF HYBANTHUS ENNEASPERMUS IN DIABETIC RATS: IN-SILICO MOLECULAR DOCKING STUDIES FOR SOD, CATALASE, GPX, XANTHINE OXIDASE AND NF-KB

Abstract Background and Aims Diabetes mellitus is a chronic metabolic disease affected a wide range of population all over the world and leads to development of secondary complications. Enzymes play key role in the development of secondary complication. Method The flavonoid rich fraction was prepared using ethyl acetate. Oral glucose tolerance test normoglycemic effect of flavonoid rich fraction of Hybanthus enneaspermus (EHE) was evaluated at a dose of 45 and 90 mg/kg, p.o. Further effect of EHE on blood glucose, body weight, enzymes such as super oxide dismutase (SOD), catalase and thiobarbituric acid reactive substances (TBARS) and lipid parameters were estimated out at 45 and 90 mg/kg p.o. per day for 21 days in streptozotocin-induced type-2 diabetic rats. Molecular docking technique was also performed against Super Oxide Dismutase (SOD), Catalase, Glutathione Peroxidase (Gpx), xanthine oxidase and nuclear factor kappa B (NF-κB) for selected phytoconstituents. Results EHE contain significant amount of phenolic, tannin, flavonoid and saponin. EHE significantly decreased blood glucose level in diabetic rats. All the tested lipid parameters and enzyme level of SOD, Catalase and Gpx were significantly restored to the normal level in the EHE treated rats. Moreover, there was a significant increase in the body weight in the EHE treated rats. Docking study showed the best binding affinity and interaction of phytoconstituents with xanthine oxidase and NF-κB signified their role in kidney disorders. Conclusion EHE has very impressive profile as an antidiabetic, antioxidant and nephroprotective activity via enzymes modulation and it may prove to be effective treatment for kidney disorders in hyperglycemia.

Correction to: Enhanced l-Dopa production from elicited cell suspension culture of Hybanthus enneaspermus (L.) F. Muell

Correction to: Enhanced l-Dopa production from elicited cell suspension culture of Hybanthus enneaspermus (L.) F. Muell

Enhanced l-Dopa production from elicited cell suspension culture of Hybanthus enneaspermus (L.) F. Muell

Hybanthus enneaspermus is an important source for l-Dopa production. This study reports elicited l-Dopa production in cell suspension cultures of Hybanthus enneaspermus. Cell suspension culture was established using green friable calli from leaf explants cultured on MS medium containing 2.0 mg/l 2,4-D. Effects of different elicitors such as SA, YE, MeJA and AgNO3 on biomass accumulation and l-Dopa content were studied. Among the elicitor tested SA treated culture produced highest biomass and l-Dopa according to their exposure time and concentration. Maximum biomass of 15.5 ± 0.16 g FW, 4.05 ± 0.18 g DW and l-Dopa production of 8.88 mg/g DW were observed at 150 μM concentration of SA. This was 9.25-fold higher compared to that of the unelicited control culture. The results obtained in this study clearly show that the elicitation strategy is a promising method for biosynthesis of l-Dopa production by cell suspension cultures of H. enneaspermus.

Therapeutic potential of medicinal plants for the management of urinary tract infection: A systematic review.

Urinary tract infection (UTI) is one of the most severe public health problem affecting both sexes but females are more susceptible due to the differences in urogenital and reproductive anatomy, physiology and lifestyle. As in UTI, different parts of the urinary tract are affected and morbidity due to UTI is more common in women of all ages and older men. Due to multi-drug resistant strains and high recurrence rate, UTI has become a major socioeconomic burden. The current review article was aimed to describe the natural therapeutic strategies to manage and cure the UTI. For this purpose, different databases including Google Scholar, Cochrane database, and PubMed etc. were explored. Inclusion criteria were any research article investigating the current therapy of UTI. It was found that microbial infections including Escherichia coli, Enterococcus faecalis and Klebsiella species are the major causes of UTI with different signs and symptoms including painful urination or dysuria, hematuria, urinary urgency, burning micturition, frequent urination, nausea, and vomiting. Antibiotics like trimethoprim, sulfamethoxazole, quinolone etc. as the first choice of the drug are used worldwide. However, due to microbial resistance, several life-threatening side effects, repeated high doses, high cost and low efficacy of these antibiotics motivated the researchers to explore natural remedies for the treatment of UTI. Herbal medicines are effective to combat bacterial resistance with high efficacy, and easy availability with minimal or no side effects. For these reasons it has attained the attention of researchers wanting to explore the herbal treatment of UTI. Vaccinium macrocarpon, Tribulus terrestris, Trachyspermum copticum, Cinnamomum verum and Hybanthusenn easpermus are some common medicinal plants reported to have therapeutic potential for the management and cure of the UTI. Although herbal medicines have more potential over conventional medicine but more discoveries are required to explore the phytoconstituents and their mechanism of action responsible for the management and cure of UTI.

PHYTOCHEMICAL SCREENING AND ANTIMICROBIAL ACTIVITY OF MEDICINAL PLANT - Hybunthus enneaspermus L. F. Muell

Hybanthus enneaspermus (L.). F. Muell is a small suffrutescent perennial herb distributed in the trophical and subtropical regions of the world. It grows 15-30 cm in height with many different diffuse or ascending branches and is pubescent in nature. The plant is popularly known as Rathanapurus or Pursharathna (Sanskrit, Hindi) and Orithazhthmarai(Tamil). This herb is considered to extremely beneficial to men, hence it is called Rathanapurush. Traditionally the plant is used as an aphrodisiac, demulcent, tonic, diuretic, in urinary infections, diarrhea, leucorrhoea, dysuria and sterility. The plant kingdom shows many species of plant conataining substances of medicinal value which have yet to be identified. Phytochemical analysis of major phytoconstituents which are possessing many biological activities, hence this study creates a presence of different extracts alkaloids, flavanoids, carbohydrates etc., The medical plants Hybunthus enneaspermus was from in an around Rasipuram,Namakkl district. Preliminary photochemical studies were performed for the presence or absence of Alkaloida, Proteins and Amino acids, Flavnoids, Phenolic compounds, Carbohydates and Phytosterol. Petroleum ether, Chloroform, Methanol, Ethanol and Isopropyl alcohol extracts of Hybunthus enneaspermus were investigated for antibacterial activity was tested against pathogenic bacteria. The petroleum ether extract inhibited maximum zones of inhibition were observed in E.Coli. Petroleum ether, Chloroform, Methanol and Ethanol extracts of Hybunthus enneaspermus were investigated for antifungal activity. The Methanol and Ethanol extract inhibited maximum zone of inhibition were observed in Aspergillus niger and Aspergillus flavus. Keywords: Medicinal plant, Bacteria, Fungi, Phytochemical and Hybunthus enneaspermus L

Evaluation of in vitro anti-inflammatory potential of ethanolic extract of hybanthus enneaspermus

Evaluation of in vitro anti-inflammatory potential of ethanolic extract of hybanthus enneaspermus

Medicinal Plants Sold as Anti-Haemorrhagic in the Cotonou and Abomey-Calavi Markets (Benin)

Market herbalists are one of the primary uses of primary health care for people in developing countries. They contribute to the conservation of endogenous plants and knowledge. In order to identify plants with antihemorrhagic properties sold in markets in southern Benin, an ethnopharmacological survey was carried out among 34 herbalists in 17 markets in Cotonou and Abomey-Calavi. The method used is Triplet Purchase of Medicinal Recipes (ATRM). A total of 38 plant species in 24 families were identified. The most represented family is the Rubiaceae (13.16%). The most cited species are Cissampelos mucronata (12.96%), Hybanthus enneaspermus (9.26%) and Cassytha filiformis (8.02%). Considering the plants mentioned in single use, C. mucronata (37.5%), C. filiformis (12.5%) and N. laevis (10%) were the most cited species. The leafy stem (71%) is the most used part. Two methods of preparation are mainly used, maceration (45%) and decoction (55%). The extracts of these plants could be a source of Improved Traditional Medication (AHT) for the treatment of haemorrhages.

Pharmacological mechanisms involved in the analgesia induced by ethanol extract of Hybanthus enneaspermus leaves.

BackgroundHybanthus enneaspermus (HE) leaves are being used traditionally to relieve pain, and scientific studies have demonstrated their analgesic potential. This study attempted to elucidate the pharmacological mechanism(s) involved in the analgesic action of ethanol extract of H. enneaspermus leaves (EEHE).Materials and methodsForty-two male Wistar rats were separately randomized into seven groups (n=6 rats in each group) for tail immersion and formalin tests. Group I (control) received distilled water (10 mL/kg) while groups II and III received acetaminophen (the reference drug, 100 mg/kg ip) and EEHE (1000 mg/kg po), respectively. Groups IV–VII were pretreated with cimetidine (50 mg/kg ip), naloxone (5 mg/kg ip), propranolol (0.15 mg/kg ip), and prazosin (0.15 mg/kg ip), respectively, 1 hour before EEHE (1000 mg/kg po) treatment.ResultsThe EEHE-induced increase in tail-flick latency was reduced by blockade of histamine and adrenergic receptors but prevented by blockade of opiate receptor in the tail-flick test. However, the EEHE-induced decrease in paw licking time was prevented only by blockade of opiate receptor but unaffected by histamine and adrenergic receptors blockers.ConclusionThese findings suggest that the analgesic effect of EEHE in different pain types may involve different neural mechanisms and that the opioidergic pathway contributes more to EEHE-induced analgesia than the other pathways.

Evaluation of dietary Hybanthus enneaspermus (Linn F. Muell.) as a growth and haemato-immunological modulator in Labeo rohita

This study evaluated the effects of Hybanthus enneaspermus aqueous extract (HEE) as a feed supplement on growth, immune responses, haematological, and biochemical parameters of Labeo rohita and its susceptibility to Aeromonas hydrophila infection. Diets containing five variable concentrations of HEE (g kg−1) (0 [basal diet], 1.0 [H1], 2.0 [H2], 3.0 [H3], and 4.0 [H4]) were fed to fish (average weight: 17.2 ± 0.21 g) for 6 weeks. Growth parameters, immune parameters, and haemato-biochemical parameters were examined 6 weeks post-feeding. The results showed that growth parameters, such as final weight (39.47 ± 3.1 g) and specific growth rate (1.92 ± 0.2), were significantly higher in the H3 group than in the control. Immunological parameters such as serum lysozyme (24.73 ± 2.16 U mg−1), phagocytic activity (19.14± 0.26%), and respiratory burst activity (0.307 ± 0.005) were significantly higher in the H3 group, compared with the control. Among the haematological parameters examined, red blood cells, white blood cells, neutrophils, total serum protein, and albumin levels were significantly higher in the H3 group. However, HEE administration had no significant effect on haematocrit, haemoglobin, monocytes, or eosinophils levels. Blood biochemical parameters, including superoxide dismutase and alkaline phosphatase activities, were higher (P < 0.05) in the H3 group than in the control; however, the opposite result was found for aspartate transaminase, alanine transaminase, glucose, and cholesterol. Supplementation with 3 g kg−1 HEE increased the activities of intestinal digestive enzyme (amylase, protease, and lipase) in fish. Moreover, fish fed diets supplemented with 3 g kg−1 HEE exhibited the highest disease resistance against A. hydrophila infection. These results suggest that dietary administration of HEE (at 3 g kg−1) has several positive effects on growth, immune and haematological responses in L. rohita.

Somatic embryogenesis and in vitro flowering in Hybanthus enneaspermus (L.) F. Muell. – A rare multipotent herb

Present study reports the various factors affecting somatic embryogenesis and in vitro flowering in Hybanthus enneaspermus (L.) F. Muell. The effect of the salts strength of Murashige and Skoog's (MS) medium, concentration of sucrose and plant growth regulators were analyzed for the induction of direct somatic embryogenesis using nodal segments as explants. High frequency of somatic embryogenesis was reported on full strength MS medium (with 3% sucrose) and additives supplemented with 0.5 mg/L 6-benzylaminopurine (BAP) and 0.25 mg/L indole-3-acetic acid (IAA). Maximum somatic embryos (207.0 ± 4.2) were germinated on 1/2 strength MS medium augmented with 0.5 mg/L BAP. Microscopic studies revealed the typical developmental patterns in somatic embryogenesis from globular to heart-shaped and followed by bipolar torpedo-shaped somatic embryos from nodal explants. The plantlets raised from the somatic embryos resulted in flowering on full strength MS medium augmented with 1.0 mg/L each of BAP and Kinetin (Kin) + 0.5 mg/L IAA at 50 μmol/(m2·s) SFPD for 13 h/d photoperiod. About 92% plantlets were successfully acclimatized in the greenhouse. Field transferred plants exhibit normal flowering and fruit setting. The study could be exploited for large scale propagation of true to type plants as conservation strategies of this rare and endemic medicinal plant.

A pyridine and pyrrole coupled rhodamine derivative for Co(II) ion detection and its imaging application in plant tissues

A rhodamine derivative (3) incorporating 2-[(1H-pyrrol-2-ylmethyl)-(2-pyridinylmethyl) amino]- tripodal receptor in its architecture exhibited both fluorogenic and chromogenic signalling responses preferentially in presence of Co(II) ion. The observed absorption and fluorescence spectral enhancements along with a colour change in 3 on addition of Co(II) ion is associated with its complexation mediated structural transformation from spirolactam conformation to ring-opened amide derivative. The associated photophysical spectral modulations with Co(II) specificity, higher association constant corresponding to effective complexation, faster metal-probe kinetics, reversibility in ‘on-off’ signalling and a wider operation pH range endorses potential utility of 3 as chemosensing probe in practical applications such as detection of Co(II) ion uptake and accumulation in root and shoot tissues of Hybanthus enneaspermus plant species.

Factors affecting Agrobacterium-mediated transformation in Hybanthus enneaspermus (L.) F. Muell.

Agrobacterium tumefaciens-mediated transformation system was established for Hybanthus enneaspermus using leaf explants with the strain LBA4404 harbouring pCAMBIA 2301 carrying the nptII and gusA genes. Sensitivity of leaf explants to kanamycin was standardized (100 mg/l) for screening the transgenic plants. Transformation parameters (OD, virulence inducer, infection time, co-cultivation period, bactericidal antibiotics, etc.) influencing the gene transfer and integration were assessed in the present investigation. Fourteen-day pre-cultured explants were subjected with Agrobacterium strain LBA4404. Optimized parameters such as culture density of 0.5 OD600, infection time of 6 min, AS concentration of 150 µM with 3 days co-cultivation revealed maximum transformation efficiency based on GUS expression assay. The presence of gusA in transgenics was confirmed by polymerase chain reaction and Southern blotting analysis. The present transformation experiment yielded 20 shoots/explant with higher transformation efficiency (28 %). The protocol could be used to introduce genes for trait improvement as well as for altering metabolic pathway for secondary metabolites production.

Genetic stability and coumarin content of transformed rhizoclones and regenerated plants of a multi-medicinal herb, Hybanthus enneaspermus (L.) F. Muell.

Agrobacterium rhizogenes-transformed hairy root somaclones (rhizoclones) and regenerated shoots of a rare medicinal herb, Hybanthus enneaspermus (L.) F. Muell. were investigated as a source of coumarin. Transformed nature of rhizoclones was verified by PCR amplification of rolA-B and mas2 genes. Clonal fidelity among rhizoclones was revealed by RAPD analysis exhibiting DNA monomorphism. Of a total of 104 rhizoclones of A4 origin and 76 rhizoclones of 8196 origin, a sample of 16 PCR tested clones was selected for shoot regeneration including eight clones pertaining to each strain type on the basis of sustained root growth index. Opine gene expression was demonstrated in rhizoclones and regenerated shoots. Inter-rhizoclonal variations with respect to biomass proliferation and coumarin content were evident. HPLC-tested coumarin accumulation was ca. threefold higher in the superior rhizoclone of A4 origin (A4-HRL-2B7) compared to that in natural roots. Transformed leafy shoots derived from A4-HRL-2B7 had the maximum coumarin content (3.25 mg g−1 d.wt. extract) which was significantly higher than that of in natura aerial part samples. Genetic stability of selected fast-proliferating rhizoclones and their respective in vitro shoot regenerants in terms of RT-PCR based expression of rolB and rolC genes and coumarin content was sustained through 16 successive multiplication cycles. This investigation ushers the possibility of harnessing the genetically stable biosynthetic potential of hairy root clones and transformed shoot regenerants of H. enneaspermus in vitro towards sustainable production of coumarin and other medicinally important phytochemicals.

GC-MS analysis of bioactive components and biosynthesis of silver nanoparticles using Hybanthus enneaspermus at room temperature evaluation of their stability and its larvicidal activity.

Green synthesis of silver nanoparticles (AgNPs) using Hybanthus enneaspermus extract at room temperature that act as a reducing agent as well as capping agent has been investigated. The synthesized AgNPs were characterized by UV-visible spectroscopy, X-ray diffraction (XRD), Fourier transform infrared (FTIR), zeta potential, and dynamic light scattering (DLS) transmission electron microscopy (TEM) and energy-dispersive X-ray (EDX). The silver surface plasmon resonance was observed at 420 nm in the UV-visible spectrum. XRD peaks were observed at 2θ values in 38.20°, 44.40°, 64.60°, and 77.50° which are indexed as (111), (200), (220), and (311) bands of face-centered cubic (fcc) structures of silver. FTIR revealed the AgNPs were capped with plant compounds of alcohol, phenols, carbonyl, amines, and amide functional groups. TEM image shows that the particles were of spherical, hexagonal, and triangular in shape, and the size range was 16-26 nm. Further, DLS exhibits the average size of 25.2 nm and the zeta values were measured (-27.1 mV) which proves the stability of the AgNPs. The conversion of Ag(+) ions into Ag(0) was calculated using inductively coupled plasma atomic emission spectroscopy (ICP-MS) and was found to be 96 %. The biosynthesized AgNPs showed the larvicidal activity with the LC50 values of 17.24 and 13.12 mg/L against the fourth-instar larvae of Anopheles subpictus and Culex quinquefasciatus, respectively. The GC-MS analysis of the plant extract showed that 39 bioactive phytochemical compounds have been found to possess a wide range of activities, which may help in the protection against incurable diseases.

Afrohybanthus (Violaceae), a new genus for a distinctive and widely distributed Old World hybanthoid lineage

Recent traditional and molecular systematic studies of the violet family, Violaceae, have confirmed extensive polyphyly of the genus Hybanthus and substantial polyphyly in Rinorea as well. Phylogenetic analyses have proposed up to nine distinct hybanthoid clades. Broad studies of representative taxa within each clade have revealed coherent suites of macromorphological traits in foliage, flowers, fruits and seeds that easily discriminate the nine hybanthoid lineages from each other and from currently recognized genera in the family. Base chromosome numbers and biogeography also provide additional support for recognition of the hybanthoid clades as distinct segregate genera. Some hybanthoid clades have available older generic names, but one of the two Old World lineages, namely the Hybanthus enneaspermus group, is presently nameless. This clade, distinctive in its ellipsoid, pale yellow, often foveolate seeds, is the most diverse in the Paleotropics, with approximately 25 species distributed across Africa, Madagascar, southern Asia, northern Australia and the southwestern Pacific. The group is segregated here as Afrohybanthus gen. nov., with new combinations provided for existing names, all of which have thus far proven morphologically distinct and worthy of recognition at the rank of species. Imminent future studies will describe additional taxa in the new genus.

Comparative Study on Antioxidant Activity of Crude and Alkaloid Extracts of Hybanthus enneaspermus Linn) F. Mull

Free radical generation is directly associated with oxidation in foods and biological systems. Antioxidants are vital substances which possess the ability to protect the body from damage caused by free radical induced oxidative stress. The aim of the present study is to evaluate and compare the in vitro antioxidant and free radical scavenging capacity of crude and alkaloid extracts of Hybanthus enneaspermus. Antioxidants such as catalase, peroxidase, superoxide dismutase, vitamin C, vitamin E and lipid peroxidation assays were analyzed in both the extracts. The content of total tannins, phenols, flavonoids, carotenoids and alkaloids were estimated and radical scavenging activity was assayed using 1,1-diphenyl-2-picryl hydroxyl (DPPH), 2,2’ azinobis-3 ethylbenzothiozoline-6 sulfonic acid (ABTS+) cation decolourization test, superoxide radical, nitric oxide, hydroxyl radical (OH.), hydrogen peroxide (H2 O2), ferric reducing antioxidant power (FRAP) and 2 2 reducing power methods. The results confirm that the crude and alkaloid extracts of H. enneaspermus revealed high level of secondary metabolites and antioxidant activities. H. enneaspermus has significant antioxidant properties and could serve as a good source of natural antioxidant, which may be helpful in preventing the distructive process of various oxidative stress.

L-Dopa production and antioxidant activity in Hybanthus enneaspermus (L.) F. Muell regeneration.

Hybanthus enneaspermus is an ethanobotanical plant extensively used in Indian traditional medicine. Quick and efficient in vitro mass propagation of this plant species was established for commercial utilization from leaf and node explants using various concentrations and combinations of plant growth regulators and polyamines. The maximum number of multiple shoots per leaf explant (40 shoots) was achieved on MS medium supplemented with 20mg/l spermidine in combination with 4mg/l BA+1.5mg/l IAA after 8weeks of culture. The elongated shoots were rooted (16 roots/shoot) on MS medium with the best concentration of IBA (1.5mg/l) and in combination with 20mg/l putrescine after 5weeks of culture. The plants were successfully acclimatized (98%) in the sand: soil: vermiculite mixture (1:1:1 v/v/v) in the greenhouse. An increased antioxidant activity was recorded in vitroregenerated shoots when compared to in vitro-induced roots. L-Dopa content was recorded higher in leaves (8.31mg/g DW) followed by stem (6.22mg/g DW) and root (3.22mg/g DW) of leaf-derived plants than the field-grown parent plant after 5weeks. By adopting this protocol, the regenerated-plants could be used for drug production and pharmacology work with as an alternative to field-grown plants.