Abhijit Bandyopadhyay

Efficient micropropagation and chlorocholine chloride induced stevioside production of Stevia rebaudiana Bertoni.

A promising method of micropropagation of Stevia rebaudiana Bertoni has been developed with an aim to increase the biomass, survivability of the plantlets and stevioside production, using chlorocholine chloride (CCC). Microshoots transferred to the MS medium containing different combinations CCC and IBA were found to be most effective in terms of growth pattern, hardening ability of the plantlets and stevioside content, compared to MS medium containing either IBA or CCC. Among other combinations tested, MS medium supplemented with 3 mg/l CCC and 3 mg/l IBA was found most effective in inducing significant changes like reduced shoot length, increased number of roots, higher leaf size, increased biomass and chlorophyll retaining capacity, higher survival percentage and most importantly the elevated stevioside content. Collectively, the major observations of this research indicate that application of CCC in micropropagation of S. rebaudiana Bertoni is a promising approach and has commercial prospects.

Elevated antioxidant potential of chlorocholine chloride-treated in vitro grown Stevia rebaudiana Bertoni

Medicinal plants contain a plethora of secondary metabolites, most of which are bioactive in nature. The role of a popular plant growth retardant CCC has been investigated to explore its impact on secondary metabolite production, particularly phenols and flavonoids from in vitro grown Stevia rebaudiana. CCC stimulated the production of total phenols and flavonoids in calli and leaves. Moreover, this elevated level of phenols and flavonoids was correlated with the antioxidant potential of the tissue extracts. Methanolic extracts from CCC-treated calli and leaves showed significant increment in antioxidant activity as determined by standard DPPH, ABTS, and hydroxyl radical scavenging assays. No significant antiproliferative effect of methanolic extracts from different tissue was noticed against THP-1 monocyte (ATCC-TIB202), Hela cell (ATCC-CCL2) lines endorses the issue of clinical safety of the extracts.

Phytoaccumulation of iron by callus tissue of Clerodendrum indicum (L)

Clerodendrum indicum (L.) is one of the important medicinal plants, originating in India. The plant is used as medicine for bronchitis, asthma and different immunological disease. Micro-propagation is a quick method, producing a huge number of plants. Apical leaf of Clerodendrum indicum (L.) was used as an explant for callogenesis. The combinations of different plant growth regulators (2,4-D, NAA, KN) in MS (Murashige and Skoog) medium produced two types of calli: one was friable and loose and the other was green, embryogenic and compact. Heavy metals cause pollution and health hazards. The plant absorbs heavy metals, which is very useful for controlling environmental pollution. Iron is a major nutrient for both plants and animals, but hyper accumulation of iron is injurious to health. Calli were used to study the optimum conditions of iron uptake in presence of different chemical environments. The different environments were created by using different chemical reagents such as (HCL, HNO3, H3PO4 and KNO3). Iron (III) was spiked with radioactive Fe-59 and dynamics of uptake was followed by measuring radioactivity by gamma-ray spectrometer. The best condition of iron uptake corresponded to 1 mol L−1 KNO3 after 4 hour of equilibration.

Stimulation of in vitro morphogenesis, antioxidant activity and over expression of kaurenoic acid 13-hydroxylase gene in Stevia rebaudiana Bertoni by chlorocholine chloride

Phytoconstituents from medicinal plants are considered as important source of raw materials of drugs for pharmaceutical industries. Biotechnology has become an inevitable approach in the area of research and development of medicinal plants for many decades. The present work has been carried out to ascertain the role of chlorocholine chloride (CCC) on in vitro morphogenesis, antioxidant activity and expression level of kaurenoic acid 13-hydroxylase (KA13H) gene in Stevia rebaudiana. To fulfill these purposes chlorocholine chloride was applied in the Murashige and Skoog (Physiol Plant 15(3):473–497, 1962) medium in combination with other plant growth regulators such as 1-naphthalene acetic acid, kinetin and thidiazuron. Chlorocholine chloride was found to contribute significant role on in vitro morphogenesis of S. rebaudiana as evidenced by the formation of embryogenic calli and increase in callusing and microshooting efficiency of explant, i.e., cotyledonary leaf. Moreover, antioxidant enzyme activity as well as ascorbic acid content of the calli and leaves was also stimulated after application of chlorocholine chloride. Q-PCR amplification using gene-specific primers revealed that CCC also promoted the expression level of KA13H gene in S. rebaudiana leaves. The overall study highlighted the promising role of chlorocholine chloride on regeneration efficiency of cotyledonary leaf, significant promotion in antioxidant potential and expression of KA13H gene in S. rebaudiana.

Chlorocholine chloride mediated resistance mechanism and protection against leaf spot disease of Stevia rebaudiana Bertoni

Investigations have been carried out to determine the influence of chlorocholine chloride on induction of the resistance mechanisms of Stevia rebaudiana against leaf spot disease, caused by Alternaria alternata. The paper also focuses an impact of chlorocholine chloride induced resistance on reduction of leaf spot disease. Chlorocholine chloride is attributed to its significant role in defence responses through augmentation of phenol and salicylic acid content as well as stimulation of phenylalanine ammonia lyase and peroxidase activity in S. rebaudiana following inoculation with A. alternata. Histochemical studies revealed that fungal invasion as well as infection process was appreciably reduced in chlorocholine chloride treated plants through peroxidase-H2O2 mediated strengthening of cell wall. The overall study highlights the significant role of chlorocholine chloride in induction of resistance in S. rebaudiana against A. alternata.