Antioxidants from Callus Technology

Antioxidants are very important compounds that are very vital in human health and they have been proven to reduce the risk of diseases such as cancer in human health. Many researchers have used callus to produce antioxidant and most of them used different techniques to get reasonable amounts of antioxidants. The technique used determines the number of antioxidants that will be produced from any explants. Callus Technology involves the techniques of producing callus and metabolites in the presence of explants using different plant hormonal combination in media, different environmental culture condition (light, relative humidity and tempreture), use of elicitors and under a sterile conditions. Callus technology is very promising due to its ability to produce a larger quantity of metabolites (antioxidants) compare to the raw extract of its explants. The use of callus to produce antioxidants is very important and very useful in discovering new plants as a source of antioxidants. The use of callus technology was reviewed for production of antioxidant from the callus of the following plants: Sericostoma pauciflorum, Helicteres angustifolia L, Lepidium sativum L, Randia echinocarpa, Andrographis paniculata Nees, Citrullus colocynthis, Rauwolfia vomitoria Afzel, Decalepis hamiltonii, Bacopa monnieri (Linn.) and Isodon rugosus (Wall. Ex Benth). Callus technology can be utilized to produce antioxidants and other metabolites in industrial quantity. Most of the metabolites from plants have been found to have medicinal values or useful to mankind and antioxidant is one of them.

Inventorization of traditional ethnobotanical uses of wild plants of Dawarian and Ratti Gali areas of District Neelum, Azad Jammu and Kashmir Pakistan

Background An ethnobotanical expedition was conducted to document the traditional ethnobotanical (TEB) uses of wild flora of Dawarian and Ratti Gali villages of District Neelam, Azad Jammu and Kashmir (AJK) Pakistan. District Neelam has rich plant diversity and is hub of many endemic plant species while the study areas are not yet explored. The research area: Dawarian and Rati Gali (DRG) area is mountaineous terrain and villages are located on far and farther distances. DRG area has rich biocultural and plant diversity comprising of different ethnic groups of Kashmir state. The current research was aimed to explore and document traditional medicines (TEMs) and other domestic and commercial uses of wild plants. This study will assist to evaluate conservation and commercial worth of wild flora which can be potential candidate for drug discovery through ethnopharmacological analysis. Methods The current quantitative ethnobotanical research was carried out in 2018 by interviewing 150 indigenous informants (90 male and 60 female) of DRG area using questionnaire applying structured and semi structured interview methodology. Data analysis was analyzed by using quantitative ethnobotanical statistical tools such as fidelity level (FL), informant consensus factor (ICF), Spearman’s rank correlation (SRC) and data matrix ranking (DMR). Results The indigenous people of DRG area use wild plants in their daily life to cope life necessities i.e. food, vegetables, fodder, fuel, shelter, timber and herbal medicines. TEMs are primarily used to cure different infirmities like diabetics, asthma, dysentery, constipation, cold, fever, joint pain, wound healing, cancer, cardiovascular disorders, epilepsy, kidney infections and many types of skin diseases. Current study revealed the data of 103 wild plants species belonging to 46 plant families from selected areas of District Neelum, AJK. Results depicted that Asteraceae ranked 1st (12 plants spp). Among plant parts used leaf ranked 1st (18%), followed by seed (17%) and root (13%). While prevalent form recipe mode was decoction (20%), followed by powder (17%) and extract (14%) and fodder was highest (37%) EB use-form fodder, followed by food (32%) and fuel (17%). Quantitative ethnobotanical analysis (QEA) was carried to find the reliability and novelty of the study. Five plant species including Berberis lyceum (FL = 97.78%), Isodon rugosus (FL = 95.71%), Saussurea lappa (FL = 94.74%), Aconitum heterophyllum (FL = 92.71%) and Taxus baccata (91.58%) had shown high fidelity level which confirmed that these plants have high medicinal worth in study area. The highest value (0.94) of ICF was for diseases group “tuberculosis and leucorrhea”, followed by stomachache and flatulence (0.93), diabetics and blood pressure (0.92) and asthma and chest infections (0.88). For other uses fuel with ICF (0.83) ranked first and second was hedging and thatching (ICF = 0.82) where people use plants or their parts for construction. Spearman’s rank correlation (SRC) test indicated that number of TEB uses increases if number of species is increased. Jaccard index (JI) analysis depicted that 56.31% plants are being used as TEMs which are first time explored from the study area. While 26.21% plants are being used in different TEB uses which are different from past cited literature. These novel findings of research indicate that wild flora of the study area has great potential for novel drug discovery and provision of materialist services for the indigenous communities. Conclusion The present research revealed that TEMs uses of 58 plants are novel being first time reported from the study area (DRG) of District Neelam of AJK. The results showed that plants like Acer cappadocicum, Ajuga bracteosa and Swertia paniculata are used to cure diabetes, Viscum album, Viola canescens, Taxus baccata are used for cure of cancer, Isodon rugosus, Polygala chinensis are used in TEMs for treating cardiovascular disorders and Anaphalis triplinervis is used for epilepsy. Berberis lyceum, Ajuga bracteosa, Aconitum heterophyllum, Bistorta amplexicaule, Saussurea lapa and Jurinea dolomiaea are severely threatened and there is urgent need to do conservation measures for available of valuable MPs to the indigenous communities for life necessities and for future research. The current study will also be useful addition in ethnobotanical database, preservation of traditional culture and drug discovery and drug development through future ethnopharmacological research.

Molecular signature of nrDNA-ITS marker in Isodon rugosus (Lamiaceae)

Molecular features of nrDNA-ITS sequences in medicinal plant Isodon rugosus were assessed with universal ITS-1 and ITS-2 primers. ITS sequences of 46 Isodon species were defined for their respective ITS-1, 5.8S and ITS-2 regions through in-silico datamining and analysis. Only in 32 species, sequence of all the three defined regions was complete, while in the rest of 14 species was partial. Isodon rugosus revealed five genic regions, namely, 18S rRNA gene (partial sequence, > 1…..38), ITS-1 (complete sequence, > 39…..236), 5.8S rRNA gene (complete sequence, > 237…..400), ITS-2 (complete sequence, > 401…..607) and large subunit 26S rRNA gene (partial sequence, > 608…..672). The sequence regions were recorded with variable sites (VS), singleton (ST) and species-specific parsimonious-informative (PI) sites. The information was also collected in a phylogenetic tree obtained using the maximum likelihood (ML) method.

Molecular Autograph of Maturase-K Gene in Isodon rugosus (Lamiaceae)

Molecular autograph of trn/mat-K gene was used as one of the important candidate marker in addressing the questions of systematics and barcoding of medicinal plants. Features of trn/mat-K gene in Isodon rugosus (Lamiaceae) were assessed for the first time (NCBI GenBank Accession Number: MH939199.1). Sequence of 756 bp length was amplified by the universal matK primers (matK472F and matK1248R) in the cpDNA of the plant. It was reported to contain trnK gene (>1…..50; >746…..756), partial sequence; and matK gene (>51…..745), partial coding DNA sequence. Alignment search and analysis showed that only nine different Isodon species are currently available for matK sequences with repeated copy of submissions in GenBank. The matK sequences of I. rugosus was reported with 34.2 (G + C)% and 17 variable sites (VS), out of which seven were singleton (ST) and 10 sites were species-specific parsimoniously-informative (PI) that could be used to differentiate I. rugosus from other species as well as to authenticate the taxon. Phylogenetic analysis resulted into monophyletic clustering of I. rugosus near to the clade having I. coetsa by both maximum likelihood (ML) and maximum parsimony (MP) methods. Clades obtained in ML tree were more informative as compared to MP tree.

Bioactivity-guided isolation of rosmarinic acid as a principle bioactive compound from the butanol extract of Isodon rugosus against pea aphid, Acyrthosiphon pisum

Aphids are agricultural pest insects that transmit viruses and cause feeding damage on a global scale. Current pest control involving the excessive use of synthetic insecticides over decades has led to multiple forms of aphid resistance to most classes of insecticides. In nature, plants produce secondary metabolites during their interaction with insects and these metabolites can act as toxicants, antifeedants, anti-oviposition agents and deterrents towards the insects. In a previous study, we demonstrated that the butanol fraction from a crude methanolic extract of an important plant species, Isodon rugosus showed strong insecticidal activity against the pea aphid, Acyrthosiphon pisum. It was however not known as which compound was responsible for such activity. To further explore this finding, current study aimed to exploit a bioactivity-guided strategy to isolate and identify the active compound in the butanol fraction of I. rugosus. As such, reversed-phase flash chromatography, acidic extraction and different spectroscopic techniques were used to isolate and identify the new compound, rosmarinic acid as the bioactive compound in I. rugosus. Insecticidal activity of rosmarinic acid was carried out using standard protocols on A. pisum. The data was analyzed using qualitative and quantitative statistical approaches. Considering that a very low concentration of this compound (LC90 = 5.4 ppm) causes significant mortality in A. pisum within 24 h, rosmarinic acid could be exploited as a potent insecticide against this important pest insect. Furthermore, I. rugosus is already used for medicinal purposes and rosmarinic acid is known to reduce genotoxic effects induced by chemicals, hence it is expected to be safer compared to the current conventional pesticides. While this study highlights the potential of I. rugosus as a possible biopesticide source against A. pisum, it also provides the basis for further exploration and development of formulations for effective field application.