Comparative study of in vitro, ex vitro and in vivo grown plants of Arnica montana – polyphenols and free radical scavenging activity

Arnica montana L. is an endangered species rich in sesquiterpene lactones, phenolic acids and flavonoids with high pharmaceutical value. The polyphenolic content and free radical scavenging activity of plants that had passed all stages of cultivation: micropropagation and rooting (in vitro), adaptation in greenhouse (ex vitro) and mountain conditions (in vivo) were evaluated. Four surface flavonoid aglycones [scutellarein 6-methyl ether (hispidulin), scutellarein 6,4’-dimethyl ether (pectolinarigenin), 6-OH luteolin 6-methyl ether and kempferol-6-methyl ether] were detected in the acetone exudates of the studied samples by means of thin layer chromatography.No differences in the accumulation of surface flavonoids were found among the tested leaf extracts of in vitro, ex vitro and in vivo samples. However, the extracts from the flowers were richer in surface flavonoids than extracts from the leaves. The methanol extracts of the samples from ex vitro and in vivo grown A. montana plants had significantly higher radical scavenging activity and polyphenolic content than the extracts of in vitro samples. The observed differences in the contents of these biologically active compounds were related to different growth conditions and stages of plant development. The biotechnological method of A. montana established holds promise for the future production of antioxidants

Multiplication and Conservation of Threatened Medicinal Plant Arnica montana L. by in vitro Techniques

An efficient and reproducible in vitro protocol for mass production of the threatened medicinal plant Arnica montana L. (Asteraceae) was developed. The effectiveness of various combinations of plant growth regulators on A. montana clonal multiplication was assessed, using seedlings stems as initial explants. Among 12 tested nutrient media, the optimum one (MS supplemented with 1.0 mg/l BAP and 0.1 mg/l IAA) increased the organogenesis frequency up to 95% in the best origin, with mean number of shoots per explant 4.25 for 5 weeks. Sub-cultivations on this medium every 4 weeks led to increase of the propagation rate as in the fifth subculture the average number of shoots per explant reached 12.32+-0.82. Rooting of uniform in vitro shoots was 100% successful on half strength MS medium supplemented with 0.5 mg/l IBA. The ex vitro adapted plants showed 90% survival, and were further acclimatized to two mountain ex situ collections. Plants looked healthy and true-to- type and began to bloom in the second or the third year. In addition, a successful protocol for slow-growth storage of in vitro A. montana cultures was elaborated, after testing 8 media with mannitol or sorbitol. The medium 1/2 MS containing 3% sorbitol and 2% sucrose was chosen as the best one, efficiently retarding the growth of the in vitro plantlets, thus allowing 6-month maintenance without sub-cultivation. The developed in vitro protocols could be of great value for commercial propagation and sustainable conservation of this threatened medicinal plant

Enzyme and Non-Enzyme Antioxidant Activity of Micro Propagated Cape Gooseberry (Physalis peruviana L.)

Several studies have represented data on in vitro clonal multiplication of Physalis peruviana L. using different nutrient media supplemented with diverse growth regulators, but there is still scarce information on the changes in the antioxidant potential of the plants subjected to micropropagation. The present research investigates the effect of various plant growth regula-tors on the antioxidant activity of micropropagated P. peruviana L. The seedling explants of P. peruviana (L.) were cultured on Murashige and Skoog (MS) medium supplemented with different concentrations of the studied citokines, 6-benzylaminopurine (BAP), zeatin and thidiazuron (TDZ).The micro-shoots produced normal roots within two weeks of cultivation on half-strength MS medium supplemented with the auxins indole-3-butyric acid (IBA) or indole-3-acetic acid (IAA) at concentrations 0.1 mg L-1 and 0.5 mg L-1, respectively. The most efficient plant rooting was achieved with plants grown on half-strength MS medium with 0.1 mg L-1 IBA (number of shoots per explant 12.1 with a shoot height 5.2 cm) as well as 0.1 mg L-1 IAA (number of shoots per explant 10.4 with a shoot height 7.3 cm) after four weeks of cultivation. The reported micropropagation protocol here is characterized by a rapid proliferation and facilitated rooting of the microshoots. The multiple plants were successfully adapted showing 100% survival rate after two months of ex vitro growth. The changes of en-zyme and non-enzyme antioxidant activity were found to depend on the concentration of the used cytokinins. At the lower concentration of 0.5 mg L-1 of studied cytikinins the antioxidant capacity was characterized by a higher concentration of metabolites with antioxidant poten-tial. The higher cytokinin concentration of 0.1 mg L-1 used in the micropropagation procedure resulted in higher antioxidant enzyme activity. The influence of different growth regulators on the antioxidant potential of micropropagated P. peruviana L. plantlets is discussed

Enzyme and Non-Enzyme Antioxidant Activity of Micro Propagated Cape Gooseberry (Physalis peruviana L.)

Several studies have represented data on in vitro clonal multiplication of Physalis peruviana L. using different nutrient media supplemented with diverse growth regulators, but there is still scarce information on the changes in the antioxidant potential of the plants subjected to micropropagation. The present research investigates the effect of various plant growth regula-tors on the antioxidant activity of micropropagated P. peruviana L. The seedling explants of P. peruviana (L.) were cultured on Murashige and Skoog (MS) medium supplemented with different concentrations of the studied citokines, 6-benzylaminopurine (BAP), zeatin and thidiazuron (TDZ).The micro-shoots produced normal roots within two weeks of cultivation on half-strength MS medium supplemented with the auxins indole-3-butyric acid (IBA) or indole-3-acetic acid (IAA) at concentrations 0.1 mg L-1 and 0.5 mg L-1, respectively. The most efficient plant rooting was achieved with plants grown on half-strength MS medium with 0.1 mg L-1 IBA (number of shoots per explant 12.1 with a shoot height 5.2 cm) as well as 0.1 mg L-1 IAA (number of shoots per explant 10.4 with a shoot height 7.3 cm) after four weeks of cultivation. The reported micropropagation protocol here is characterized by a rapid proliferation and facilitated rooting of the microshoots. The multiple plants were successfully adapted showing 100% survival rate after two months of ex vitro growth. The changes of en-zyme and non-enzyme antioxidant activity were found to depend on the concentration of the used cytokinins. At the lower concentration of 0.5 mg L-1 of studied cytikinins the antioxidant capacity was characterized by a higher concentration of metabolites with antioxidant poten-tial. The higher cytokinin concentration of 0.1 mg L-1 used in the micropropagation procedure resulted in higher antioxidant enzyme activity. The influence of different growth regulators on the antioxidant potential of micropropagated P. peruviana L. plantlets is discussed

INDIRECT SHOOT ORGANOGENESIS OF EGGPLANT (SOLANUM MELONGENA L.)

A protocol for indirect shoot organogenesis of Solanum melongena ‘Larga Negra’ and ‘Black Beauty’ was established using hypocotyl and cotyledon derived calluses. The maximum morphogenic callus induction was observed from cultured cotyledons of 30-days old seedlings on Murashige and Skoog (MS) medium containing 2.0 mg/l α-naphthalene acetic acid and 0.5 mg/l 6-benzylaminopurine. The highest percentage of shoot regeneration and the highest mean number of shoots/callus were obtained on hormone-free MS medium. In terms of callus induction and subsequent plant regeneration, cotyledon explants were more responsive than hypocotyl explants. Regenerated shoots (2-3 cm) were rooted on MS hormone-free medium or medium containing 0.1 mg/l indole-3-butyric acid. About 90% of regenerated plantlets survived under field conditions after hardening in the glasshouse. Several somaclones exhibiting useful variation would to be proposed as initial plant material for eggplant breeding programs

Multiplication and Conservation of Threatened Medicinal Plant Arnica montana L. by in vitro Techniques

An efficient and reproducible in vitro protocol for mass production of the threatened medicinal plant Arnica montana L. (Asteraceae) was developed. The effectiveness of various combinations of plant growth regulators on A. montana clonal multiplication was assessed, using seedlings stems as initial explants. Among 12 tested nutrient media, the optimum one (MS supplemented with 1.0 mg/l BAP and 0.1 mg/l IAA) increased the organogenesis frequency up to 95% in the best origin, with mean number of shoots per explant 4.25 for 5 weeks. Sub-cultivations on this medium every 4 weeks led to increase of the propagation rate as in the fifth subculture the average number of shoots per explant reached 12.32+-0.82. Rooting of uniform in vitro shoots was 100% successful on half strength MS medium supplemented with 0.5 mg/l IBA. The ex vitro adapted plants showed 90% survival, and were further acclimatized to two mountain ex situ collections. Plants looked healthy and true-to- type and began to bloom in the second or the third year. In addition, a successful protocol for slow-growth storage of in vitro A. montana cultures was elaborated, after testing 8 media with mannitol or sorbitol. The medium 1/2 MS containing 3% sorbitol and 2% sucrose was chosen as the best one, efficiently retarding the growth of the in vitro plantlets, thus allowing 6-month maintenance without sub-cultivation. The developed in vitro protocols could be of great value for commercial propagation and sustainable conservation of this threatened medicinal plant