Exposure to single-walled carbon nanotubes differentially affect in vitro germination, biochemical and antioxidant properties of Thymus daenensis celak. seedlings.

Carbon nanomaterials such as single-walled carbon nanotubes (SWCNTs) offer a new possibility for phyto-nanotechnology and biotechnology to improve the quality and quantity of secondary metabolites in vitro. The current study aimed to determine the SWCNTs effects on Thyme (Thymus daenensis celak.) seed germination. The seedlings were further assessed in terms of morphological and phytochemical properties. Sterile seeds were cultured in vitro and treated with various concentrations of SWCNTs. Biochemical analyses were designed on seedling sample extracts for measuring antioxidant activities (AA), total flavonoids (TFC) and phenolic contents, and the main enzymes involved in oxidative reactions under experimental treatments. The results indicated that an increase in SWCNTs concentration can enhance the total percentage of seed germination. The improvement was observed in samples that received SWCNTs levels of up to 125 µg ml-1, even though seedling height and biomass accumulation decreased. Seedling growth parameters in the control samples were higher than those of grown in SWCNT-fortified media. This may have happened because of more oxidative damage as well as a rise in POD and PPO activities in tissues. Additionally, secondary metabolites and relevant enzyme activities showed that maximum amounts of TPC, TFC, AA and the highest PAL enzyme activity were detected in samples exposed to 62.5 µg ml-1 SWCNTs. Our findings reveal that SWCNTs in a concentration-dependent manner has different effects on T. daenensis morphological and phytochemical properties. Microscopic images analysis revealed that SWCNTs pierce cell walls, enter the plant cells and agglomerate in the cellular cytoplasm and cell walls. The findings provide insights into the regulatory mechanisms of SWCNTs on T. daenensis growth, germination and secondary metabolites production.

Melatonin strongly enhances the Agrobacterium- mediated transformation of carnation in nitrogen-depleted media.

With the rising demand for new cultivars of carnation, efficient transformation protocols are needed to enable the bioengineering of new traits. Here, we established a novel and efficient Agrobacterium-mediated transformation system using callus as the target explant for four commercial carnation cultivars. Leaf-derived calli of all cultivars were inoculated with Agrobacterium tumefaciens strain LBA4404 containing the plasmid pCAMBIA 2301 harboring genes for ß-glucuronidase (uidA) and neomycin phosphotransferase (nptII). Polymerase chain reaction (PCR) and histochemical assays confirmed the presence of uidA and ß-glucuronidase (GUS), respectively in transgenic shoots. The effect on transformation efficiency of medium composition and the presence of antioxidants during inoculation and co-cultivation was investigated. The transformation efficiency was increased in Murashige and Skoog (MS) medium lacking KNO3 and NH4NO3, and also in MS medium lacking macro and micro elements and Fe to 5% and 3.1% respectively, compared to 0.6% in full-strength medium. Transformation efficiency was increased dramatically to 24.4% across all carnation cultivars by the addition of 2 mg/l melatonin to nitrogen-depleted MS medium. Shoot regeneration was also doubled in this treatment. The establishment of this efficient and reliable transformation protocol can advance the development of novel carnation cultivars through molecular breeding approaches.

ZnO nanoparticles efficiently enhance drought tolerance in Dracocephalum kotschyi through altering physiological, biochemical and elemental contents.

Using nanofertilizers in certain concentrations can be a novel method to alleviate drought stress effects in plants as a global climate problem. We aimed to determine the impacts of zinc nanoparticles (ZnO-N) and zinc sulfate (ZnSO4) fertilizers on the improvement of drought tolerance in Dracocephalum kotschyi as a medicinal-ornamental plant. Plants were treated with three doses of ZnO-N and ZnSO4 (0, 10, and 20 mg/l) under two levels of drought stress [50% and 100% field capacity (FC)]. Relative water content (RWC), electrolyte conductivity (EC), chlorophyll, sugar, proline, protein, superoxide dismutase (SOD), polyphenol oxidase (PPO) and, guaiacol peroxidase (GPO) were measured. Moreover, the concentration of some elements interacting with Zn was reported using the SEM-EDX method. Results indicated that foliar fertilization of D. kotschyi under drought stress with ZnO-N decreased EC, while ZnSO4 application was less effective. Moreover, sugar and proline content as well as activity of SOD and GPO (and to some extent PPO) in treated plants by 50% FC, increased under the influence of ZnO-N. ZnSO4 application could increase chlorophyll and protein content and PPO activity in this plant under drought stress. Based on the results, ZnO-N and then ZnSO4 improved the drought tolerance of D. kotschyi through their positive effects on physiological and biochemical attributes changing the concentration of Zn, P, Cu, and Fe. Accordingly, due to the increased sugar and proline content and also antioxidant enzyme activity (SOD, GPO, and to some extent PPO) on enhancing drought tolerance in this plant, ZnO-N fertilization is advisable.

Chitosan nanoparticles improve physiological and biochemical responses of Salvia abrotanoides (Kar.) under drought stress.

BACKGROUND: The use of organic nanoparticles to improve drought resistance and water demand characteristics in plants seems to be a promising eco-friendly strategy for water resource management in arid and semi-arid areas. This study aimed to investigate the effect of chitosan nanoparticles (CNPs) (0, 30, 60 and 90 ppm) on some physiological, biochemical, and anatomical responses of Salvia abrotanoides under multiple irrigation regimes (30% (severe), 50% (medium) and 100% (control) field capacity). RESULTS: The results showed that drought stress decreases almost all biochemical parameters. However, foliar application of CNPs mitigated the effects caused by drought stress. This elicitor decreased electrolyte conductivity (35%), but improved relative water content (12.65%), total chlorophyll (63%), carotenoids (68%), phenol (23.1%), flavonoid (36.4%), soluble sugar (58%), proline (49%), protein (45.2%) in S. abrotanoides plants compared to the control (CNPs = 0). Furthermore, the activity of antioxidant enzymes superoxide dismutase (86%), polyphenol oxidase (72.8%), and guaiacol peroxidase (75.7%) were enhanced after CNPs treatment to reduce the effects of water deficit. Also, the CNPs led to an increase in stomatal density (5.2 and 6.6%) while decreasing stomatal aperture size (50 and 25%) and semi-closed stomata (26 and 53%) in leaves. CONCLUSION: The findings show that CNPs not only can considerably reduce water requirement of S. abrotanoides but also are able to enhance the drought tolerance ability of this plant particularly in drought-prone areas.

A Comparative Study on Essential Oil Profile and Antioxidant Activity in Different Parts of Three Ferulago Species.

In the present study, the phytochemical features of the essential oils from leaves, flowers, and fruits in three Ferulago species including F. angulata, F. carduchorum, and F. contracta was investigated. The essential oils were extracted using the hydrodistillation method and analyzed by GC-FID and GC/MS. The essential oil content varied between 0.46 to 2.65 % (v/w) among the various parts of the studied species. Monoterpene hydrocarbons (leaves: 34.80-78.76 %, flowers: 88.67-94.26 %, fruits: 89.54-94.21 %) constituted the main fraction of three Ferulago species. The major compounds of the essential oil among the species were α-pinene (15.01-22.16 %), ß-phellandrene (2.48-14.73 %), α-phellandrene (0.52-13.8 %), and germacrene B (0.11-13.28 %) in leaves; (Z)-ß-ocimene (38.46-47.21 %), α-pinene (10.25-18.32 %), and α-phellandrene (5.07-9.44 %) in flowers; (Z)-ß-ocimene (10.21-41.19 %), α-phellandrene (7.51-31.89 %), α-pinene (8.96-17.71 %), ß-phellandrene (7.24-17.44 %), terpinolene (2.90-7.77 %), and δ-3-carene (1.57-7.66 %) in fruits. The classical antioxidant activity of the essential oils was assessed using the DPPH method. The fruit essential oils of the studied species induced a significant level of antioxidant activity compared to ascorbic acid (EC50 =50.24 µg/mL). The highest level of antioxidant activity was found in F. carduchorum (EC50 =68.75 µg/mL) fruits essential oil. Ferulago species, therefore, have the potential to be utilized as natural antioxidants in the pharmaceutical, food or cosmetic industries owing to their high level of antioxidant activity.

Comparative assessment of proliferation and immunomodulatory potential of Hypericum perforatum plant and callus extracts on mesenchymal stem cells derived adipose tissue from multiple sclerosis patients.

BACKGROUND: Mesenchymal stem cells-derived adipose tissue (AT-MSCs) are recognized for the treatment of inflammatory diseases including multiple sclerosis (MS). Hypericum perforatum (HP) is an anti-inflammatory pharmaceutical plant with bioactive compounds. Plant tissue culture is a technique to improve desired pharmacological potential. The aim of this study was to compare the anti-inflammatory and proliferative effects of callus with field-growing plant extracts of HP on AT-MSCs derived from MS patients. MATERIALS AND METHODS: AT-MSCs were isolated and characterized. HP callus was prepared and exposure to light spectrum (blue, red, blue-red, and control). Total phenols, flavonoids, and hypericin of HP callus and plant extracts were measured. The effects of HP extracts concentrations on proliferation were evaluated by MTT assay. Co-culture of AT-MSCs: PBMCs were challenged by HP plant and callus extracts, and Tregs percentage was assessed by flow cytometry. RESULTS: Identification of MSCs was performed. Data showed that blue light could stimulate total phenols, flavonoids, and hypericin. MTT test demonstrated that plant extract in concentrations (0.03, 1.2, 2.5 and 10 µg/ml) and HP callus extract in 10 µg/ml significantly increased. Both HP extracts lead to an increase in Tregs percentage in all concentrations. In particular, a comparison between HP plant and callus extracts revealed that Tregs enhanced 3-fold more than control groups in the concentration of 10 µg/ml callus. CONCLUSIONS: High concentrations of HP extracts showed effectiveness on AT-MSCs proliferation and immunomodulatory properties with a certain consequence in callus extract. HP extracts may be considered as supplementary treatments for the patients who receiving MSCs transplantation.

Organic and inorganic elicitors enhance in vitro regeneration of Rosa canina.

BACKGROUND: Rosa canina is one of the most popular rose species which is widely used as the rootstock for the propagation of rose cultivars. The purpose of the present study is to improve the in vitro propagation efficiency of this valuable plant species using various growth stimulants in a proliferation medium. In this study, in vitro-derived axillary buds of R. canina were inoculated in Vander Salm (VS) medium supplemented with varying levels of organic or inorganic elicitors including casein hydrolysate (200, 400, and 600 mg/l), glutamic acid (2, 4, 8, and 12 mg/l), proline (500, 1000, 1500, and 2000 mg/l), and silver nitrate (25, 50, 75, and 100 mg/l), separately. Benzyl amino purine (BAP) as well as naphthalin acetic acid (NAA) were added to all media at a constant rate to promote shoot proliferation. RESULTS: The results indicated that the supplementation of casein hydrolysate to the VS medium markedly stimulated shoot regeneration by 173% in comparison to control. Shoot proliferation was also positively influenced by glutamic acid at all levels, however, at a lesser extent compared to casein hydrolysate. Silver nitrate at 100 mg/l induced the longest shoots (2.52 ± 0.248 cm) and maximum leaf number (8.90 ± 0.276) among all treatments. Although it did not encourage efficient shoot regeneration, the highest quality shoots with maximum growth vigor were observed in this treatment. CONCLUSION: In this study, the promising role of casein hydrolysate in combination with plant growth regulators has been emphasized for the improved efficiency of R. canina regeneration protocol. Moreover, the addition of silver nitrate to the culture medium seems vital for enhancing the quality of regenerated shoots. The results of this study could be beneficial either for the further pharmaceutical or biochemical investigations of R. canina or commercial purposes for mass propagation of this specimen.

Multi-walled carbon nanotubes stimulate growth, redox reactions and biosynthesis of antioxidant metabolites in Thymus daenensis celak. in vitro.

This research was aimed at determining the effects of multi-walled carbon nanotubes (MWCNTs) on seed germination, seedling growth parameters and secondary metabolite (SM) production of Thymus daenensis in vitro. Seeds were aseptically cultured in Murashige and Skoog medium (MS) with various concentrations of MWCNTs (0, 125, 250, 500, 1000 and 2000 µg ml-1). Seed germination and morphological changes in seedlings were measured. The measurements were aimed at quantifying the total phenolic contents (TPC) and flavonoids (TFC), antioxidant activities and the activity of polyphenol oxidase (PPO), l-phenylalanine ammonia-lyase (PAL), dehydrogenase (DHA) and peroxidase enzyme (POD) of the seedling extract. Seedling biomass and seedling height grew significantly as the MWCNTs level increased. The biomass and height peaked at 250 µg ml-1 (0.41 ± 0.01 gr FW, 5.99 ± 0.55 cm) and then rapidly decreased to 0.040 ± 0.1 gr FW and 1.42 ± 0.24 cm in response to 1000 µg ml-1, 30 days after the treatment. Additionally, SM and the analyses of enzyme activity revealed that the highest amounts of TPC (6.70 ± 0.06 mg GAE g-1 DW), TFC (8.19 ± 0.01 mg QUE g-1 DW), antioxidant activities (73.88 ± 0.47%) and maximum PAL activity (1.25 ± 0.08 mM cm g-1 FW) were detected in plants grown on MS media fortified with 250 µg ml-1 MWCNTs. The results reveal that MWCNTs in low doses (250 µg ml-1) can encourage the production of biomass, elicit more SM from seedlings and enhance the biosynthesis of antioxidants. TEM images showed that MWCNTs could cross the plant cell wall and enter the cellular cytoplasm.