Micropropagation of Hemidesmus indicus (L.) R. Br. through axillary bud culture

A procedure for rapid in vitro propagation of the aromatic and medicinal plant Hemidesmus indicus (L.) R.Br. (Family Asclepiadaceae) from nodal explants is described. The highest shoot multiplication rate of 8.2 ± 0.4 shoots/explant with a 95% frequency was achieved in S weeks culture period on Murashige and Skoog medium supplemented with 1.15 M kinetin and 0.054 M -naphthaleneacetic acid. Excised shoots were rooted on the same basal medium supplemented with 1.15 M kinetin and 7.35 M indole-3-butyric acid. Shoots derived from subcultures exhibited better rooting response than those from primary cultures. After a hardening phase of 2 weeks, there was a 70% transplantation success in the field.Abbreviations MS Murashige and Skoog (1962) medium - BA N⁶ benzyladenine; KN kinetin - NAA a-naphthaleneacetic acid - IBA indole-3-butyric acid - IAA indole-3-acetic acid     

Protective effect of Hemidesmus indicus R.Br. root extract against cisplatin-induced cytogenetic damage in mouse bone marrow cells

The aqueous extract of Hemidesmus indicus roots was investigated for its in vivo antigenotoxic effect against cisplatin-induced cytogenetic damage. Swiss albino mice were administered with various doses of the extract either singly (50, 100 and 200 mg/kg body weight) or as split doses (10, 20 and 40 mg/kg bw/day) for five consecutive days by oral gavage. As endpoints, chromosome aberrations, micronuclei in polychromatic erythrocytes, mitotic index and PCE/NCE ratio were estimated. The extract protected the bone marrow cells from cisplatin-induced genotoxicity in an inverse dose-dependent manner. However, the extract was cytotoxic at all doses. But, under split dose regime it conferred a higher level of genoprotection and was not cytotoxic at the lower two doses. The presence of saponins, tannins, phenols, terpenoids, flavonoids and coumarins in the crude extract could explain these effects.     

Impact of Hemidesmus indicus R.Br. extract on ethanol-mediated oxidative damage in rat kidney

Abstract

The antioxidant effect of the ethanolic extract of Hemidesmus indicus R.Br. root (EHI), an indigenous Ayurvedic medicinal plant in India, was studied in rats with ethanol-induced nephrotoxicity. Administering 5 g/kg body weight/day of ethanol for 60 days to male Wistar rats resulted in significantly elevated levels of serum urea, creatinine and uric acid as well as kidney thiobarbituric acid reactive substances (TBARS), lipid hydroperoxides (LOOH) and conjugated dienes (CD) as compared to those of the experimental control rats. Decreased levels of kidney superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), reduced glutathione (GSH), vitamin C and vitamin E were also observed on alcohol administration as compared with those of the experimental control rats. EHI was administered at a dose of 500 mg/kg body weight/day for the last 30 days of the experiment to rats with ethanol-induced kidney injury, which significantly decreased the levels of serum urea, uric acid and creatinine as well as kidney TBARS, LOOH and CD and significantly elevated the activities of SOD, CAT, GPx, GSH, vitamin C and vitamin E in kidney as compared to that of untreated ethanol-administered rats. Histopathological observations also correlated with the biochemical parameters. Thus, the data indicate that treatment with EHI offers protection against free radical-mediated oxidative stress in kidney of animals with ethanol-induced nephtrotoxicity.     

Impact of Hemidesmus indicus R.Br. extract on ethanol-mediated oxidative damage in rat kidney

The antioxidant effect of the ethanolic extract of Hemidesmus indicus R.Br. root (EHI), an indigenous Ayurvedic medicinal plant in India, was studied in rats with ethanol-induced nephrotoxicity. Administering 5 g/kg body weight/day of ethanol for 60 days to male Wistar rats resulted in significantly elevated levels of serum urea, creatinine and uric acid as well as kidney thiobarbituric acid reactive substances (TBARS), lipid hydroperoxides (LOOH) and conjugated dienes (CD) as compared to those of the experimental control rats. Decreased levels of kidney superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), reduced glutathione (GSH), vitamin C and vitamin E were also observed on alcohol administration as compared with those of the experimental control rats. EHI was administered at a dose of 500 mg/kg body weight/day for the last 30 days of the experiment to rats with ethanol-induced kidney injury, which significantly decreased the levels of serum urea, uric acid and creatinine as well as kidney TBARS, LOOH and CD and significantly elevated the activities of SOD, CAT, GPx, GSH, vitamin C and vitamin E in kidney as compared to that of untreated ethanol-administered rats. Histopathological observations also correlated with the biochemical parameters. Thus, the data indicate that treatment with EHI offers protection against free radical-mediated oxidative stress in kidney of animals with ethanol-induced nephrotoxicity.     

Indirect organogenesis from leaf explants of Hemidesmus indicus (L.) R. Br.: an important medicinal plant

Hemidesmus indicus (Asclepiadaceae) leaf explants were utilized for establishing culture in MS medium fortified with individual cytokinins, auxins, and their combinations. Optimum response (80%) was observed in N⁶-benzyladenine (BA, 20 μM) + indole-3-acetic acid (IAA, 1 μM) with 19.67 ± 0.81 shoots per explant. Roots were induced in ¼MS + indole-3-butyric acid (IBA, 20 μM).     

Hepatoprotective activity of Hemidesmus indicus R. br. in rats

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 roots of H. indicus (100-500 mg/kg body weight, po) 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 H. indicus 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. The results suggest that methanolic extract of H. indicus roots possesses a potential antihepatotoxic activity.     

Differential morphometric and micro-morpho-anatomical responses toward types of culture vessels used in micropropagation of Hemidesmus indicus (L.) R. Br.

Plant Cell, Tissue and Organ Culture (PCTOC) - Plantlets of Hemidesmus indicus (L.) R. Br. (Indian sarasaparilla) were developed in vitro in two different types of culture vessels and closure...     

In Vitro Clonal Propagation Through Bud Culture of Hemidesmus indicus (L) R Br: An Important Medicinal Herb

The present study involves in vitro propagation of Hemidesmus indicus (L) R Br through bud multiplication and subsequent plant regeneration. The buds multiplied to produce numerous shoots at variable rates in presence of a-naphthaleneacetic acid (NAA) and 6-benzylaminopurine (BAP) as well as NAA and kinetin. The best response in bud multiplication was obtained in Murashige and Skoog’s (MS) basal medium supplemented with 0.1 mg I^-1 NAA and 2.0 mg I^-1 BAP (7-8 shoots per explant) and the bud break time was only 4 days after inoculation. The multiplication rate was low when the buds were cultured in NAA and kinetin media and the shootlets regenerated were very thin, weak and elongated. The shoots regenerated were further cultured on MS and half strength MS basal media with variable levels of indole-3-butyric acid (IBA) for initiation of roots. Culture of shootlets for 34 weeks in one half strength of MS medium followed by culturing in the same medium with 1.5 mg 1-1 IBA induced highest production of roots (3-5 roots per shoot) within 2 weeks. Chromosome number stability with no detectable structural changes was observed in the regenerates. The rooted plants were successfully established in the soil with 85% survival rate.     

Correction to: Differential morphometric and micro-morpho-anatomical responses toward types of culture vessels used in micropropagation of Hemidesmus indicus (L.) R. Br.

Plant Cell, Tissue and Organ Culture (PCTOC) -     

In vitro propagation and conservation of Indian sarsaparilla, Hemidesmus indicus L. R. Br. through somatic embryogenesis and synthetic seed production

A protocol has been developed for achieving somatic embryogenesis from callus derived from nodal cuttings and production of synthetic seeds in Hemidesmus indicus L. R. Br. a highly traded ethnomedicinal plant. Proembryogenic, friable, light yellowish callus was induced from the basal cut end of the nodal cuttings on Murashige and Skoog (MS) medium supplemented with 3 μM indole-3-butyric acid (IBA). The highest rate of somatic embryogenesis (92 %) was observed when the callus was subcultured on half strength MS medium supplemented with 2 μM IBA. On induction medium somatic embryos were developed up to the torpedo stage. Further elongation and germination of somatic embryos were obtained in MS medium supplemented with 4 μM 6-benzylaminopurine (BA) in combination with 1.5 μM gibberellic acid (GA₃). Somatic embryos were collected and suspended in a matrix of MS medium containing sodium alginate (3 % W/V) dropped into 75 mM calcium chloride (CaCl₂·2H₂O) solution for the production of synthetic seeds and later transferred to MS medium for germination. The synthetic seeds were successfully germinated on medium even after 120 days of storage at 4 °C. The plantlets were eventually transferred to soil with 92 % success.     

Production of 2-hydroxy 4-methoxy benzaldehyde using root cultures of Hemidesmus indicus

Nodal explants of Hemidesmus indicus plants cultured in half strength Murashige and Skoog medium fortified with 2 mg indole-3-butyric acid l–1 in the dark produced 10–12 roots (1–2 cm) with minimal callusing in 10 days. Roots, 20 mg dry wt, cultured in Gamborg et al. medium supplemented with 2 mg IBA l–1, sucrose (4% w/v), pH (5.6) and agitation (70 rpm) in the dark yielded 550 mg root dry wt/flask and 0.18% 2-hydroxy 4-methoxy benzaldehyde in the roots after 30 days. The maintenance of normal morphology of the roots, consistent biomass and product yields over a 21-month period indicated the stability of the cultures. © Rapid Science Ltd. 1998     

Synthesis of nonembryonic synseeds in Hemidesmus indicus R. Br.: short term conservation,evaluation of phytochemicals and genetic fidelity of the regenerants

Plant Cell, Tissue and Organ Culture (PCTOC) - Non-embryogenic, synthetic seeds were formed by encapsulating the nodal segments (NS) of Hemidesmus indicus R. Br. in calcium alginate hydrogel...     

Antidiarrhoeal effects of methanolic root extract of Hemidesmus indicus (Indian sarsaparilla)--an in vitro and in vivo study

Methanolic extract of H. indicus root (MHI) was screened for its antimicrobial activity against S. typhimurium, E. coli and S. flexneri, in vitro and in experimentally induced diarrhoea in albino rats, in vivo. MHI had an anti enterobacteriae effect as evident from agar well diffusion method and decrease in CFU/ml in MHI treated LB broth culture. MHI inhibited the castor oil induced diarrhoea in rats as judged by a decrease in the amount of wet faeces in MHI-pretreated rats at a dose of 500-1500 mg/kg. The results indicated that MHI was more active than standard antidiarrhoeal drug, lomotil. Phytochemical tests revealed the main constituents as tannins, steroids, triterpenoids and carbohydrates. Present findings suggested that MHI might elicit an antidiarrhoeal effect by inhibition of intestinal motility and by its bacteriocidal activity.     

Nutlet morphology in Hemigenia R.Br. and Microcorys R.Br. (Lamiaceae)

Nutlets of Hemigenia R.Br. and Microcorys R.Br. were examined using SEM. Significant variation, mainly useful at the infrageneric level, was found in nutlet shape, nature of the attachment scar, nature of surface sculpturing, exocarp cell shape and sculpturing, and nature of the indumentum. Typical nutlets are ovoidal, strongly reticulate or rugose. The exocarp cells are isodiametric and convex to papillate. Also common are cylindrical nutlets, often with longitudinal ridging and papillate exocarp cells. Surface pitting and concave exocarp cells are rare. A cladistic analysis of nutlet characters suggests both Hemigenia and Microcorys are polyphyletic, and Microcorys paraphyletic with respect to Westringia Sm. Notwithstanding that, the infrageneric classification of Hemigenia was largely supported, while in Microcorys, there was support for sect. Hemigenioides, but sects Anisandra and Microcorys were not resolved as distinct.     

Growth and swainsonine production of Swainsona galegifolia (Andr.) R. Br. untransformed and transformed root cultures

Untransformed and transformed root cultures of Swainsona galegifollawere established for swainsonine production. Transformed rootsgrew faster and produced higher swainsonine levels (62.3 µgg^–1 DW) than untransformed roots (23.6 ,µg g^–1DW) or roots of intact plants (8.7 µg g^–1 DW). Transformationof a number of plant genotypes using A. rhizogenes strain LBA9402 showed that plant genotype Influences swainsonine levelin transformed roots but that a wide range of swainsonine levelscan be induced by separate transformation events in the samegenotype. Enhancement of swainsonine production was attemptedby treatment with sugars and induction of polyploid roots. Key words: Agrobacterium rhizogenes, root cultures, Swainsona galegifolia, swainsonine     

Analysis of the chemical composition of root essential oil from Indian sarsaparilla (Hemidesmus indicus) and its application as an ecofriendly insecticide and pharmacological agent

The Indian sarsaparilla (Hemidesmus indicus) is a commonly used plant in Indian traditional medicine of Ayurveda for the preparation of various non-alcoholic beverages. However, limited studies are available on the essential oil of H. indicus roots (HRO); therefore, the study evaluated the antioxidant, anti-inflammatory and antidiabetic activities of H. indicus root essential oil as well as insecticide potential against the common pests of stored food materials (Sitophilus oryzae, Callosobruchus maculatus and Tribolium castaneum). The repellant efficacy of HRO was found to be high against S. oryzae (8.21 ± 0.55 μg/mL). Likewise, the fumigant potential was also observed for HRO against these pests; the higher activities were observed against S. oryzae and C. maculatus (32.46 ± 1.42 and 35.18 ± 1.62 μg/L). Besides, the essential oil was also found to be active as a contact poison, however, against all the three pests, the toxicity was above 100 μg/mm³, being the highest against C. maculatus (122.8 ± 3.57 μg/mm³). To analyze the possible effect of the essential oil on grains, the different grains were allowed to germinate and compared to that of normal; thus, the non-toxic nature of HRO against the stored products is also confirmed. The essential oil shown to have DPPH hydrogen peroxide and ABTS radical scavenging, nitric oxide scavenging potential, and inhibition of lipoxgenase, alpha-amylase and alpha-glucosidase. Overall, the present study concludes that the H. indicus may be a suitable repellant and fumigant agent against different pests of stored products and a possible antioxidant, anti-inflammatory, and anti-diabetic agent.     

Hypolipidemic and antioxidant effects of Morus alba L. (Egyptian mulberry) root bark fractions supplementation in cholesterol-fed rats

The 70% alcohol extract of the Egyptian Morus alba L. root bark was fractionated over cellulose CC eluted with water, 50% methanol and finally with 100% methanol to yield 3 fractions (MRBF-1, MRBF-2 and MRBF-3), respectively. In continuation of chromatographic purification of 70% alcohol extract fractions of the Egyptian M. alba L. root bark, 4 compounds namely: mulberroside A, 5,7,2'-trihydroxyflavanone-4'-O-beta-D-glucoside and albanols A and B were isolated from MRBF-2 for the first time from the Egyptian plant. Experimentally induced atherosclerosis was produced by feeding rats a diet enriched in coconut oil (25% by weight) and cholesterol (2% by weight) for 21 days. Then, hypercholesterolemic rats were orally administered (MRBF-1, MRBF-2 and MRBF-3 fractions) in a dose of 500 mg kg(-1) day(-1) for 15 successive days, in order to evaluate their expected hypocholesterolemic activity. Lipid profile parameters such as plasma total cholesterol, LDL-C, VLDL-C, LDL:HDL ratio and triglycerides, as well as plasma and liver lipid peroxides and glutathione-S-transferase enzyme levels, serum paraoxonase enzyme level, LDL oxidation, LDL aggregation and LDL retention, were measured. Plasma and liver glutathione-S-transferase enzyme levels were unaffected in all studied groups. The results revealed that the administration of (MRBF-2 and/or MRBF-3) fractions resulted in alleviation of atherosclerotic state. Administration of MRBF-3 significantly retained plasma and liver peroxides towards their normal levels, and also, produced significant increase in resistance towards major atherogenic modifications; namely LDL oxidation, LDL aggregation and LDL retention by 44%, 30%, and 33%, respectively. Thus, it can be concluded that the consumption of MRBF-2 and (MRBF-3, in some extent) fractions of M. alba L. root bark 70% alcohol extract may act as a potent hypocholesterolemic nutrient and powerful antioxidant via the inhibition of LDL atherogenic modifications and lipid peroxides formation in hypercholesterolemic rats.     

Micropropagation of Hemidesmus indicus for cultivation and production of 2-hydroxy 4-methoxy benzaldehyde

Caulogenic responses of various explant types from 12-month-old plants of Hemidesmus indicus were tested. Second and third visible nodes (0.5 cm) from the apex and root segments (0.5 cm) were the most and least regenerative respectively, with the formation of 9.37 and 2.6 shoots in 4 weeks on half strength MS medium supplemented with 2.22 μM BA and 1.07 μM NAA and 4.44 μM BA and 2.69 μM NAA respectively. Caulogenic ability of the nodes decreased with increasing maturity. Shoot buds initiated upon the young nodes on day 10 developed into 7.2 cm long shoots within 4 weeks thereby making a shoot elongation phase unnecessary. Nodal explants of the in vitro raised shoots subcultured in the same medium produced 9.32 shoots of 7.1 cm length in 3–4 weeks, similar to those of the mature plant-derived nodes. Multiplication through subculture of the nodes up to 25 passages of 4 weeks each was achieved without decline. Shoot cultures were rooted in quarter salt strength MS medium containing 9.8 μM IBA and the rooted plants were hardened for establishment in pots at 96% rate. Four months after establishment, the micropropagated plants were stable and showed uniform morphological and growth characteristic. After 12 months of cultivation in the field, on an average micropropagated plant consisted of 4–5 shoots, 5–8 branches per shoot and increased root biomass (13.5 g) compared to the poor growth (single shoot and 2–3 branches) and root production (4.6 g) values obtained with plants raised from conventional rooted stem cuttings. The concentration of the root specific compound, 2-hydroxy 4-methoxy benzaldehyde per plant was 2–3 fold higher in micropropagated plants though on unit dry root biomass (0.12% per g dry wt) basis it remained the same between two sources of plants. This revised version was published online in June 2006 with corrections to the Cover Date.     

Water relations of the root hemiparasite Olax phyllanthi (Labill) R.Br. (Olacaceae) and its multiple hosts

Summary Water relations of the root hemiparasite Olax phyllanthi were compared with those of its major species of hosts in natural habitat in coastal heath near Denmark, SW Australia. Leaf water potentials of Olax during winter were 0.4 to 1.4 MPa lower (more negative) than those of all (29) non parasitic host species examined. During the dry summer months (January to March), shallow-rooted hosts developed water potentials up to 3 MPa lower than those of Olax, and were accordingly rated as no longer accessible as a source of water to the hemiparasite. By contrast, deep-rooted hosts, with access to the water table, showed water potentials less negative than Olax, and haustorial contacts retained with these apparently enabled continued extraction of water and nutrients throughout the summer. Three other species of root hemiparasites parasitized by Olax, but not themselves parasitizing Olax, showed leaf water potentials throughout the year very close to, and mostly slightly more negative than those of Olax. Nocturnal measurements of leaf water potential in winter (July and August) in soil at field capacity (water potential –0.006 MPa) showed maintenance of a 0.5–0.8 MPa potential difference between Olax and a range of common host species. By dawn most hosts had equilibrated with the water potential of the soil, whereas both exposed and bagged Olax plants recorded potentials of –0.8 MPa. Daytime rates of transpiration and photosynthesis of Olax were less than those of a range of common hosts, but water use efficiencies were not consistently different between hemiparasite and hosts. This was reflected in almost identical mean values for carbon isotope ratio (¹³C/¹²C) between Olax (mean value –27.0) and thirteen frequently exploited hosts ( value –27.1). The results are discussed in relation to published information on other angiosperm hemiparasites.     

Micropropagation of Telopea speciosissima R. Br. (Proteaceae). 2: Rhizogenesis and acclimatisation to ex vitro conditions

Conditions affecting rhizogenesis in vitro and ex vitro and subsequent acclimatisation of Telopea speciosissima (waratah) were investigated. Clonal selections were successfully rooted in vitro in agar, on filter paper bridges or using crushed quartz-sand, the last substrate resulting in superior growth of roots. The in vitro substrates were impregnated with half-strength MS, 7.5 gl^-1 sucrose and various concentrations of IBA. For the quartz-sand, an IBA concentration of 50 M was optimal, 70% of microcuttings were rooted. No plantlets rooted in vitro were acclimatised to ex vitro conditions (using mist, fog or humidity tent regimes). Microcuttings (25–45 mm in length) were rooted ex vitro in a fog humidity regime (droplet size <10 m) using an IBA powder dip (3 g IBA kg^-1). Neither a mist nor a humidity-tent regime was suitable for rooting of waratah microshoots ex vitro. A peat and perlite mixture was superior to crushed quartz-sand or potting mix for the rooting of microshoots; this appeared to be related to the air-filled porosity (>20%) of the mixture, measured after the medium was saturated and then drained for 24h. Plantlets must be left under the high humidity regime until shoot growth resumes (four to eight weeks) otherwise plant mortality increase significantly. In vitro-produced leaves abscised between eight and 12 weeks after transfer to ex vitro conditions, indicating that these structures did not acclimatise ex vitro.Abbreviations BA benzyladenine - GA₃ gibberellic acid - IBA indole-3-butyric acid - LSD least significant difference - MS Murashige and Skoog medium