Magnetofection approach for the transformation of okra using green iron nanoparticles.

Climate change, pesticide resistance, and the need for developing new plant varieties have galvanized biotechnologists to find new solutions in order to produce transgenic plants. Over the last decade scientists are working on green metallic nanoparticles to develop DNA delivery systems for plants. In the current study, green Iron nanoparticles were synthesized using leaf extract of Camellia sinensis (green tea) and Iron Chloride (FeCl3), the characterization and Confirmation was done using UV-VIS Spectroscopy, FTIR, SEM, and TEM. Using these nanoparticles, a novel method of gene transformation in okra plants was developed, with a combination of different Magnetofection factors. Maximum gene transformation efficiency was observed at the DNA to Iron-nanoparticles ratio of 1:20, by rotation of mixture (Plasmid DNA, Iron-nanoparticles, and seed embryo) at 800 rpm for 5 h. Using this approach, the transformation of the GFP (green fluorescent protein) gene was successfully carried out in Abelmoschus esculentus (Okra plant). The DNA transformation was confirmed by observing the expression of transgene GFP via Laser Scanning Confocal Microscope (LSCM) and PCR. This method is highly economical, adaptable, genotype independent, eco-friendly, and time-saving as well. We infer that this approach can be a potential solution to combat the yield and immunity challenges of plants against pathogens.

The Inhibitory Effect of Polyphenon 60 from Green Tea on Melanin and Tyrosinase in Zebrafish and A375 Human Melanoma Cells.

Polyphenon 60 (PP60) from green tea has long been used as an antioxidant, anticancer, antimicrobial, and antimutagenic. Aim of the Study. To investigate tyrosinase inhibition-related kinetic mechanism and antimelanogenesis potential of PP60. Materials and Methods. The effect of PP60 on melanin and tyrosinase was evaluated in A375 melanoma cells and zebrafish embryos. The melanoma cells were treated with 20, 40, and 60 µg/mL of PP60, and tyrosinase expression was induced by using L-DOPA. The western blot method was used for the evaluation of tyrosinase expression. Cell lysates were prepared from treated and untreated cells for cellular tyrosinase and melanin quantification. Furthermore, zebrafish embryos were treated with 20, 40, and 60 µg/mL of PP60 and reference drug kojic acid for determination of depigmentation and melanin quantification. In vitro assays were also performed to examine the impact of PP60 on mushroom tyrosinase activity. To determine cytotoxicity, MTT was used against melanoma cell line A375. Results. PP60 showed good tyrosinase inhibitory activity with an IC50 value of 0.697 ± 0.021 µg/mL as compared to kojic acid a reference drug with an IC50 value of 2.486 ± 0.085 µg/mL. Kinetic analysis revealed its mixed type of inhibition against mushroom tyrosinase. In addition, western blot analysis showed that at 60 µg/mL dose of PP60 significantly reduced L-DOPA-induced tyrosinase expression in melanoma cells. PP60 significantly inhibits the cellular tyrosinase (p < 0.05) and reduces the melanin (p < 0.05) contents of melanoma cells. Furthermore, PP60 was found to be very potent in significantly reducing the zebrafish embryos' pigmentation (p < 0.05) and melanin (p < 0.05) content at the dose of 60 µg/mL. Conclusions. Our results demonstrate that PP60 has a strong potency to reduce pigmentation. It may be useful for the cosmetic industries to develop skin whitening agents with minimal toxic effects.

Light and scanning electron microscopic observation of palynological characteristics in spineless Astragalus L. (Fabaceae) and its taxonomic significance.

Palynological characterization is considered to be one of the significant taxonomic tools for the delimitation and identification of morphologically complicated taxa. Hence, the pollen morphology of 12 species of spineless Astragalus L. was examined using light and scanning electron microscopy. Studied pollen were small to medium, monad, prolate to per-prolate and tricolporate type in all studied taxa. The exine sculpturing varied from reticulate to microreticulate whereas colpus ornamentation ranged from scabrate to granulate. Furthermore, maximum polar and equatorial diameter was recorded in Astragalus leucocephalus Bunge. (45.00 µm) and A. pyrrhotrichus Boiss. (22.91 µm) while minimum in A. amherstianus Benth. ex Royle (28.75 µm) and A. amherstianus Benth. ex Royle (15.00 µm), respectively. Similarly, the ratio of polar to equatorial diameter was recorded maximum in A. ophiocarpus Boiss. (2.05). The width of colpi was larger in A. hamosus L. (1.29 µm) and smaller in A. ophiocarpus Boiss. (0.62 µm). We have also found the maximum value of mesocolpium in A. retamocarpus Boiss. (2.08 µm) while minimum in A. oxyglottis Steven ex M.Bieb. (1.87 µm). The quantitative pollen attributes helped in the development of pollen keys for the accurate and quick identification of the studied species. Furthermore, ordination and cluster analysis were performed for the differentiation of the investigated taxa at species level. Based on our results, we conclude that pollen features can be used for the delimitation and identification of the studied taxa. RESEARCH HIGHLIGHTS: Pollen micromorphology is a useful tool for classifying complicated taxa. The pollen micromorphology of 12 spineless species of Astragalus L. was studied using LM and SEM. The observed pollen characteristics aided in Astragalus L. serve for the identification and classification of taxa at specific level.

Implication of scanning electron microscopy and light microscopy for oil content determination and seed morphology of Verbenaceae.

Due to increase in energy catastrophe and accompanying environmental issues, the search for renewable and alternative energy resources is a fiercely disputed problem in the scientific world. Biodiesel has proven to be the finest sustainable alternative to gasoline. Because of their environment-friendly character and cost-effectiveness, nonedible oil-bearing seeds could be the possible resource for biodiesel production. Therefore, the present study uses scanning electron microscopy (SEM) to investigate and identify micromorphological features of six novel, nonedible oil-bearing seeds of Verbenaceae as potential biodiesel feed stocks. Selected seeds of Verbenaceae range in size from 1 to 5 mm in width and 1.8 to 6.4 mm in length, according to light microscopic studies. The oil content of seeds ranges from14 to 19%. Seed texture, color, shape, size, periclinal and anticlinal wall patterns, and cell outline are all more variable in SEM-mediated seed ultrastructure analyses. The findings of this work imply that SEM could be a useful method for revealing hidden micromorphological features in a variety of nonedible oil-producing seeds, which could aid to future investigation, proper identification, seed categorization, and authenticity. These seeds were hard to be identified by the native people due to their resemblance in seed color and shape, so there is a need to give a detailed description of seed characters for proper identification of these plants. This work will determine the new features of the seeds, which will be helpful in near future.

Implication of scanning electron microscopy as a tool for identification of novel, nonedible oil seeds for biodiesel production.

Biodiesel is a promising, bio-based, renewable, nontoxic, environment friendly, and alternative fuel for petroleum derived fuels which helps to reduce dependency on conventional fossil fuels. In this study, six novel, nonedible seed oil producing feedstock were explored for their potential for sustainable production of biodiesel. It is very important to correctly identify oil yielding plant species. Scanning electron microscopy (SEM) was used as reliable tool for authentic identification of oil yielding seeds. Macromorphological characters of seeds were studied with light microscopy (LM). Outcomes of LM of seeds exposed distinctive variation in seed size from 16.3 to 3.2 mm in length and 12.4 to 0.9 mm in width, shape varied from oval to triangular, and color from black to light brown. Oil content of nonedible seed ranged from 25 to 30% (w/w). Free fatty acid content of seed oil varied from 0.32 to 2.5 mg KOH/g. Moreover, ultra structural study of seeds via SEM showed variation in surface sculpturing, cell arrangement, cell shape, periclinal wall shape, margins, protuberances, and anticlinal wall shape. Surface sculpturing varied from rugged, reticulate, varrucose, papillate, and striate. Periclinal wall arrangements confirmed variation from rough, wavy, raised, depressed, smooth, and elevated whereas, anticlinal walls pattern showed variation from profuse undulating, smooth, raised, grooved, deep, curved, and depressed. It was concluded that SEM could be a latent and advanced technique in unveiling hidden micromorphological characters of nonedible oil yielding seeds which delivers valuable information to researchers and indigenous people for precise and authentic identification and recognition.

Pollen micromorphological analysis of tribe Acacieae (Mimosaceae) with LM and SEM techniques.

The morphology of pollen grains of the 06 species of tribe Acacieae (Mimosaceae) belonging to one genus were investigated using scanning electron microscopic (SEM) and light microscopic (LM) approach. Lactic acid acetolysis method was used for (LM) while non-acetolysis for (SEM). The species were examined for the both qualitative and quantitative palynological features. Qualitatively the pollen was studied for the shape, exine sculpturing, pollen shape, and pollen type. Oblate spheroidal shape of pollen was observed in Accacia modesta. Subprolate shape in Accaca nilotica. In Accacia farnesiana, prolate pollen was examined while prolate spheroidal in Acacia tortilis and spheroidal pollen grains were studied in Acacia catechu, respectively. Pollen exine sculpturing of the studied tribe possesses novel variation. Foveolate, reticulate, foveolate-psilate, faint reticulate, fossulate, perforate, and scrabate exine sculpturing were noticed. Colpi were absent in all species. Excessive variation was observed in both qualitative and quantitative character of pollen. SPSS software was use for the quantitative measurement of the pollen grains. The current study will be helpful for the identification of the problematic species and phylogenetic studies of family Mimosaceae.

Identification of green energy ranunculaceous flora of district Chitral, Northern Pakistan using pollen features through scanning electron microscopy.

Pollen micro-morphological features have proven to be helpful for the plant taxonomists in the identification and classification of plants. The utilization of this plantmayhelpfulin the areas of lignocellulosic conversion to biofuels and diversify application toward biomass. The current study was planned with the aim to evaluate the pollen features of complex Ranunculaceous flora of District Chitral, Northern Pakistan using both scanning electron microscopy (SEM) and Light Microscope (LM) for their taxonomic importance. Pollens of 18 Ranunculaceous species belonging to 6 genera were collected from different localities of the research area. SEM and LM were used to examine both qualitative and quantitative micro-morphological features. Sculptring of the sexine include; Scabrate, psilate, echinate, verrucate, perforate gemmate, and reticulate and so forth. Shape of the pollens was sub-spheroidal, spheroidal, prolate, subprolate and oblate and so forth. Type of pollen was ranged from mono to tricolpate and tricolporate. Quantitative characters include length/width of the pollen, colpus, exine thickness, and P/E ratio. Based on these micro-morphological features a taxonomic key was prepared for the fast and correct identification. RESEARCH HIGHLIGHT: Study of the pollen micro-morphological features of Ranunculaceous species by SEM and LM. Analysing both qualitative and quantitative characters of the pollens. Preparation of taxonomic key based on micro-morphological features for the correct and fast identification.

Green Synthesis of Silver Nanoparticles using Bidens Frondosa Extract and their Tyrosinase Activity.

Herbal nanoparticles gain lot of attention because of their pharmaceutical importance. The present study reports the eco-friendly synthesis, characterization and their tyrosinase activity of silver nanoparticles (AgNPs) using aqueous extract of Bidens frondosa. The appearance of brown color indicated the formation of B. frondosa AgNPs. The Formation of AgNPs was confirmed by UV-Vis spectroscopy, FTIR, FESEM and EDS analysis. The formation of herbal AgNPs of size ranging 20-70 nm further was assured by energy dispersive X-ray spectroscopy (EDS) and field emission scanning electron microscopy (FESEM). The mushroom tyrosinase inhibitory activity of synthesized AgNPs was evaluated. Nanoparticles were found to have significant higher tyrosinase inhibitory activity compared to control. The IC50 values of crude extract, AgNP and Kojic acid were found to be 9, 15, and 2.37 µg/mL, respectively. AgNPs of B. frondosa may be considered as potential candidate for the production of medical and cosmetic products.

Pharmacoinformatics exploration of polyphenol oxidases leading to novel inhibitors by virtual screening and molecular dynamic simulation study.

Polyphenol oxidases (PPOs)/tyrosinases are metal-dependent enzymes and known as important targets for melanogenesis. Although considerable attempts have been conducted to control the melanin-associated diseases by using various inhibitors. However, the exploration of the best anti-melanin inhibitor without side effect still remains a challenge in drug discovery. In present study, protein structure prediction, ligand-based pharmacophore modeling, virtual screening, molecular docking and dynamic simulation study were used to screen the strong novel inhibitor to cure melanogenesis. The 3D structures of PPO1 and PPO2 were built through homology modeling, while the 3D crystal structures of PPO3 and PPO4 were retrieved from PDB. Pharmacophore modeling was performed using LigandScout 3.1 software and top five models were selected to screen the libraries (2601 of Aurora and 727, 842 of ZINC). Top 10 hit compounds (C1-10) were short-listed having strong binding affinities for PPO1-4. Drug and synthetic accessibility (SA) scores along with absorption, distribution, metabolism, excretion and toxicity (ADMET) assessment were employed to scrutinize the best lead hit. C4 (name) hit showed the best predicted SA score (5.75), ADMET properties and drug-likeness behavior among the short-listed compounds. Furthermore, docking simulations were performed to check the binding affinity of C1-C10 compounds against target proteins (PPOs). The binding affinity values of complex between C4 and PPOs were higher than those of other complexes (-11.70, -12.1, -9.90 and -11.20kcal/mol with PPO1, PPO2, PPO3, or PPO4, respectively). From comparative docking energy and binding analyses, PPO2 may be considered as better target for melanogenesis than others. The potential binding modes of C4, C8 and C10 against PPO2 were explored using molecular dynamics simulations. The root mean square deviation and fluctuation (RMSD/RMSF) graphs results depict the significance of C4 over the other compounds. Overall, bioactivity and ligand efficiency profiles suggested that the proposed hit may be more effective inhibitors for melanogenesis.

Isolation, characterization, and in silico, in vitro and in vivo antiulcer studies of isoimperatorin crystallized from Ostericum koreanum.

CONTEXT: Ostericum koreanum (Maxim.) Kitagawa (Apiaceae) roots are traditionally used as an analgesic and antiulcer agent. However, the antiulcer potential of isoimperatorin isolated from O. koreanum has not yet been explored. AIM: To evaluate the antiulcer activity of isoimperatorin isolated from the roots of O. koreanum. MATERIALS AND METHODS: Isoimperatorin was isolated as cubic crystals by repeated column chromatography of the ethyl acetate fraction and structure was verified with 1H NMR, 13C NMR and high-resolution mass spectrometry (HRMS-FAB). The crystals obtained were analyzed with the single crystal X-ray method. The MTT assay was used to determine its cytotoxicity against chondrocytes at different concentrations (0.0-737.74 µM, 24 h). The in vivo antiulcer activity of isoimperatorin (40 mg/kg) was determined against ethanol-, indomethacin- and pyloric ligation-induced ulcers in Sprague-Dawley rats. Furthermore, the effect of isoimperatorin (0.0-737.74 µM, 24 h) on the expression of type II collagen in chondrocytes was determined using western blot method. The in vitro urease inhibitory activity of isoimperatorin (0-80 µM) and molecular docking was also performed against urease. RESULTS AND DISCUSSION: Isoimperatorin demonstrated significant inhibitory activity (IC50 36.43 µM) against urease as compared to the standard drug thiourea (IC50 33.57 µM) without cytotoxic effects. It provided 70.9%, 67.65% and 54.25% protection in ulcer models induced by ethanol, indomethacin and pyloric ligation, respectively. Isoimperatorin showed the highest expression level of type II collagen at 368.87 µM. The docking results confirmed strong binding affinity with the target protein. CONCLUSION: Isoimperatorin may be used to develop antiulcer drugs with decreased side effects.