On the basis of previous studies, an antioxidant enriched high yield potential genotype (Accession VA13) was selected for this investigation. This genotype was grown in pots of a rain shelter open field of Bangabandhu Sheikh Mujibur Rahman Agricultural University, Bangladesh (AEZ-28, 24° 23′ north latitude, 90° 08′ east longitude, 8.4 m.s.l.). The seeds were sown in plastic pots (15 cm in height and 40 cm length and 30 cm width) in a randomized complete block design (RCBD) with three replications. N: P₂O₅:K₂O were applied @92:48:60 kg/ha as a split dose. First, in pot soil, @46:48:60 kg ha 1 N: P₂O₅:K₂O and second, at 7 days after sowing (DAS) @46:0:0 kg/ha N: P₂O₅:K₂O. The genotype was grouped into three sets and subjected to four salinity stress treatments that are, 100 mM NaCl, 50 mM NaCl, 25 mM NaCl, and control or no saline water (NS). Pots were well irrigated with fresh water every day up to 10 days after sowing (DAS) of seeds for proper establishment and vigorous growth of seedlings. Imposition of salinity stress treatment was started at 11 DAS and continued up to 40 DAS (edible stage). Saline water (100 mM NaCl, 50 mM NaCl and 25 mM NaCl) and fresh water were applied to respective pots once a day. At 40 DAS the leaves of Amaranthus tricolor were harvested. All the parameters were measured in six samples.

At Moderate salinity stress (MSS) and Severe salinity stress (SSS) conditions, leaf color parameters and pigments, vitamins, phenolic acids, flavonoids and antioxidant capacity of A. tricolor leaves were very high compared to control condition. Hence, salt-stressed A. tricolor leaves had a good source of natural antioxidants compared to plant grown in normal cultivation practices.

The experiments were conducted under controlled conditions using plastic pots having lower diameter of 7.8 cm, upper diameter of 13.5 cm and 12 cm in height. The soil was collected from the top layer (0-20 cm) from the Botanical Garden of the university. Soil was air dried crushed and sieved through 2 mm filter autoclaved at 121 ℃ for 2 h. The soil was autoclaved to exclude soil pathogens and other microorganisms if any. The autoclaved soil was poured in pots and kept in the growth chamber. The pots were filled with 500 g uncontaminated soil and partially decayed compost (cow manure) (2:1) and was used as growing medium. The cow dung was added into the soil for better performance of earthworms. A subsample of the study soil before mixing with compost was analyzed for its physicochemical characteristics. The soil used for the experiment was sandy loam soil having pH 7.8 , EC (Electrical conductivity) (µS/cm) =184.25 , TDS (Total Dissolved Solids) (mg/kg) = 130 , N (Nitrogen) (mg/kg) = 103 , P (Phosphorus) (mg/kg) = 10.6 , K (Potassium) (mg/kg) = 0.343 , %OC = 0.894, Cd (mg/kg) = ND (not detected by AAS).The Cd treatment was given by using anhydrous CdCl₂ (Minimum assay: 95.0%) procured from Hi-Media laboratories. The CdCl₂ anhydrous was added to the soil to make different concentrations of Cd 0.50 mM, 0.75 mM, 1.00 mM, and 1.25 mM (i.e. 56 mg/Kg , 84 mg/Kg , 112 mg/Kg and 140 mg/Kg respectively). The various treatments given are as shown below:(1)C0 (Control): (Cadmium absence);(2)C1: (0.5 mM Cd);(3)C2: (0.75 mM Cd);(4)C3: (1.00 mM Cd);(5)C4: (1.25 mM Cd).Each Cd treatment was given in soils without as well as with earthworms (WTE = without, WE = with earthworms). Earthworms (3 earthworms per pot) were inoculated after seven days of Cd treatment and incubated for 7 d in soil with earthworms. The seeds after surface sterilization were sown in soil containing different concentration of Cd and earthworms in plastic pots. These pots were kept in seed germinator under controlled conditions i.e. 25 ℃ temperature and 16:8 h dark: light photoperiod (1700 lx) for 15 d. Seedlings were harvested after 15 d followed by washing with distilled water. The growth and biochemical analysis was done on these seedlings.

Increased Cd uptake in plants in presence of earthworms enhances the total antioxidative capacity, metal chelating compounds and content of other antioxidants in plants grown under metal polluted soils. Earthworms can improve plant growth by improving nutrient availability to plants through their vermicasting activity. Their role in modifying soil pH and increasing metal phytoavailability made their use ideal in phytoremediation of polluted soils. Increased uptake and accumulation of Cd in plants activates the antioxidative system of plants takes place by addition of earthworms to soil.

The gene expression for the key enzymes involved in organic acid metabolism was studied to understand the role of earthworms in organic acid metabolism in plants under Cd metal stress. It was observed that in comparison to control (C0) seedlings the expression of CS, SUCLG1, SDH and FH was enhanced 1.72, 1.58, 1.65 and 1.88 folds in seedlings given C4 treatment with 1.25 mM dose of Cd respectively . However, after supplementation of earthworms to Cd treated soils given C4 treatment resulted in further enhancement in expression of CS (2.53 fold), SUCLG1 (2.35 fold), SDH (2.13 fold) and FH (3.06 fold) .

Seeds of B. juncea (cv. RLC-1) were given pre-sowing treatment with 24-epibrassinolide (EBR) solutions (0 and 100 nM EBR/L) for 8 h. Petri-plates were lined with Whatman1 filter paper and were supplemented with different imidacloprid (IMI) concentrations (0, 150, 200, and 250 mg IMI/L). The EBR treated seeds were rinsed with distilled water and grown in Petri-plates supplemented with IMI solutions (three petri-plates for each treatment). The Petri-plates were kept in seed germinator (temperature = 25 ℃ , photoperiod = 16 h, light intensity = 175 µmol m ^-2 s^-1) and the seedlings were harvested 10 days after sowing for further analysis.

Seed soaking with 24-epibrassinolide recovers the impaired growth of B. juncea seedlings under imidacloprid stress by modulating the expression of genes encoding key enzymes including chlorophyllase, citrate synthase, succinyl Co-A ligase, succinate dehydrogenase, fumarate hydratase, malate synthase, phytoene synthase, chalcone synthase, and phenylalanine ammonialyase.

In the present study, as compared to control seedlings, the expression of gene CHLASE (encoding chlorophyllase) was observed to increase by 2.66-fold under IMI toxicity, but seed soaking with EBR significantly reduced the expression of CHLASE to 1.07-fold in the seedlings under IMI toxicity . Data analysis using two-way ANOVA and Tukey's HSD showed significant difference for CHLASE expression in B. juncea seedlings (FIMI p < 0.01, FEBR p < 0.01, FIMI * EBR p < 0.001). MLR analysis of the fold change in CHLASE expression also revealed the increased expression of gene with IMI toxicity and EBR application (positive betaIMI-value), whereas interaction between IMI and EBR was observed to be negative .Further, in comparison to control seedlings, the expression of PSY (encoding phytoene synthase) and CHS (encoding chalcone synthase) was significantly enhanced by 5.22 and 4.54-folds respectively in the seedlings raised from EBR treated as well as untreated seeds grown under IMI stress . Significant differences in expression PSY (FIMI p < 0.001, FEBR P<0.05) and CHS (FIMI * EBR p < 0.001) were observed after analyzing the data using two-way ANOVA and Tukey's HSD. MLR analysis of fold change in gene expression also revealed the role of EBR in modulation of gene expression of PSY and CHS. Concentrations of IMI as well as EBR were regressed positively on the fold change in gene expression of PSY and CHS, thus revealing enhanced expressions of these genes under both the treatments. Moreover, interaction between IMI and EBR was positive for PSY expression, whereas negative interaction was observed for the expression of CHS .In the present study, the expression of PAL was also observed to enhance significantly by 6.68-fold in the seedlings raised from EBR treated seeds and grown under IMI stress . After analyzing the data using two-way ANOVA and Tukey's HSD, significant difference in the expression of PAL was observed (FIMI p < 0.01, FEBR p < 0.01, FIMI * EBR P<0.05). MLR analysis of the fold change in gene expression also confirmed the role of EBR in increasing the PAL gene expression under IMI pesticide stress. Positive beta-regression coefficients were observed for IMI, EBR, and IMI * EBR .The expression of genes encoding the key enzymes involved in organic acid metabolism was also studied to understand the role of EBR in organic acid metabolism under IMI pesticide stress. It was observed that as compared to control seedlings, the expression of CS (encoding citrate synthase, 2.35-fold), SUCLG1 (encoding succinyl-Co-A ligase, 1.57-fold), SDH (encoding succinate dehydrogenase, 2.01-fold), FH (encoding fumarate hydratase, 1.57-fold), and MS (encoding malate synthase, 1.91-fold) were increased in B. juncea seedlings raised from untreated seeds and grown under IMI pesticide toxicity . However, seed soaking with 100 nM EBR and germinating them under IMI toxicity resulted in further enhancement in expression of CS (2.61-fold), SUCLGD1 (4.18-fold), SDH (2.55-fold), FH (3.73-fold), and MS (4.03-fold). Data analysis using two-way ANOVA and Tukey's HSD showed significant differences in the expression of CS (FEBR p < 0.01, FIMI * EBR p < 0.01), SUCLG1 (FEBR p < 0.001, FIMI * EBR P<0.05), SDH (FEBR p < 0.01), FH (FEBR p < 0.001), and MS (FEBR p < 0.001). MLR analysis showed that gene expression in seedlings under IMI stress as well as after the EBR seed treatment was increased as indicated by positive beta-regression coefficients. Whereas, negative interactions were noticed between IMI and EBR treatments for the expression of all genes studied related to organic acid metabolism.

Briefly, broccoli seeds (0.5 g per replication) were sanitized for 15 min in sodium hypochlorite (1.5%, v/v), rinsed with Milli-Q water and soaked with aeration overnight in darkness and at room temperature. After pouring off the soaking water, the seeds were spread evenly on standard 200 square cell plug trays (21.38 × 11.05 × 1.75) containing Canadian Sphagnum peat moss previously moistened. Sprouts were grown in a culture room with controlled temperature (25 ℃ ) and a photoperiod regime with cycles of 16 h light and 8 h darkness. Water (control) or a phytohormone solution were atomized every 12 h throughout the experiment.Six trays with broccoli sprouts seeds were prepared for this study, and were assigned for (A) Control (no UV or phytohormone application), (B) UVA treatment, (C) UVB treatment, (D) MJ treatment, (E) UVA + MJ treatment, and (F) UVB + MJ treatment.MJ treatments (D, E and F) were conducted based on Perez-Balibrea et al. with slight adjustments. Briefly, methyl jasmonate (MJ) was dissolved in 0.2% ethanol to obtain a 25 µM solution and applied every 12 h by exogenous spraying 65 mL of 25 µM MJ solution from sowing day until the end of the experiment (8th day after sowing). Due to the volatility of MJ and to avoid the fact that treatment to one tray may result in application to neighboring trays, the MJ solution was applied using physical separation. Control sprouts (A) and sprouts treated with UVA or UVB alone (B and C) were irrigated with the same frequency using 65 mL of Milli-Q water containing 0.04% ethanol.On the 7th day after sowing, UV (B and C) and UV + MJ (E and F) treatments were carried out in special UVA and UVB chambers based on Moreira-Rodriguez et al. with slight adjustments. Chambers used for treatments B and E were equipped with two 40 W UVA lamps (Sylvania F40W T12 BL350, Ledvance LLC., Wilmington, MA, USA), while chambers for treatments C and F consisted of two 40 W UVB lamps (Philips TL 40W/12 RS, Philips, Ljubljana, Slovenia). Trays with broccoli sprouts were placed 30 cm below the irradiation source. All UV treatments consisted of a single exposure for 120 min. The irradiation intensities were determined prior to the experiment as 9.47 and 7.16 W/m² for UVA and UVB, respectively, using a PMA 2200 radiometer equipped with PMA 2110 UVA and PMA 2106 UVB sensors (Solar Light, Glenside, PA, USA) measuring in the spectral range from 320-400 nm and 280-320 nm, respectively.After UV treatments, trays were returned to culture room and the proper irrigation with water or MJ solution continued for an additional (acclimatization) period of 24 h. Sprouts of all six trays were harvested at the 8th day after sowing, immediately flash-frozen in liquid nitrogen, placed at -80 ℃ , freeze-dried (Labconco, Kansas City, MO, USA), and then ground to a fine powder. Samples were stored at -80 ℃ until further analysis.

Simple pre-harvest treatments such as UV radiation, applied alone or in combination with exogenous MJ, can be used as an effective emerging technology that allows the accumulation of specific phytochemicals in broccoli sprouts. Furthermore, results demonstrated that the profile of glucosinolates accumulated in stressed broccoli sprouts could be tailored towards the over-production of most indole glucosinolates by applying 25 µM MJ alone or preferably in combination with a 120 min exposure to UVA or UVB radiation (9.47 and 7.16 W/m², respectively) 24 h prior harvest. Specifically, a synergistic effect in the accumulation of NGBS was achieved by combining UV and MJ stresses. On the other hand, the production of aliphatic or specific indole glucosinolates can be triggered by UVB supplementation alone. MJ treatments may be applied if an increase in gallic acid, its derivative GAH II, specific sinapic acid derivatives (e.g., 5-SQA) and ferulic acid derivatives (e.g., 1,2-diFG) is desired. However, such increases would be at the expense of the following compounds: GAH I, GTA, diGH, 3-O-H-K, 1-O-S-beta-d-g, sinapoyl malate, sinapic acid, K-3-O-S-so-7-O-g, 1,2-diSG, 1-S-2-FG, the majoritarian isomer of 1,2,2-triSG and 1,2-diS-1-FG; as they were significantly reduced after treatments with MJ. Application of UVA alone may be recommended to accumulate GAH I, 1-O-S-beta-d-g, sinapic acid, gallic acid, K-3-O-S-so-7-O-g, 1-S-2-FG and the second isomer of 1,2,2-triSG. Finally, a single 120 min exposure to UVA radiation should be applied to increase xanthophyll and chlorophyll content in broccoli sprouts.

AMF Inoculation in Pot : Saffron corms with horizontal diameters of 1.3 to 2.8 cm were sown in pots (4 L; 1 corm per pot) in the last ten days of August 2016. Pots were filled with sterile quartz sand (3 L per pot) on a layer of sterilized expanded clay (1 L per pot). Corms were treated with two inocula (MycAgro Lab, Breteniere, FR), one composed of a single fungus Rhizophagus intraradices (Ri) and one of R. intraradices and Funneliformis mosseae (Ri + Fm). Ten grams of each inoculum were placed under each corm in order to guarantee the contact between the inoculum and the roots and therefore to favor the symbiosis between AMF and roots. Saffron corms used as controls were not inoculated (AMF-). Corms were not treated against fungal pathogens. A randomized block design was used with a total of 48 pots displayed in two experimental plot units (24 pots per unit) and three treatments (8 pots per treatment). Cultivation lasted for one cycle (August 2016-April 2017) in a heated glasshouse of the Department of Agricultural Forest and Food Sciences (DISAFA) of the University of Torino (Italy, 45° 06′ 23.21″ N Lat, 7° 57′ 82.8″ E Long; 293 m a.s.l.), with an average temperature of 22 ℃ during the day and 16 ℃ in the night. Irrigation water (pH 7.4, EC 505 µS cm) was added weekly (250 mL per pot) with a drip system. The corms were fertilized by fertigation (VIGORFLOR, AL.FE. srl, MN, Italy) every two weeks starting from the emergence of the spate, in quantities of 1.5 g/L of water. No flowering occurred because of the small size of the corms.AMF Inoculation in Open Field : Saffron corms with horizontal diameters of 2.5 to 3.5 cm were planted in the last ten days of August 2016 in two Alpine experimental sites located in the municipality of Morgex (45° 45′ 35″ N; 7° 02′ 37.3″ E; 1000 m a.s.l.) and Saint Cristophe (45° 45′ 06″ N; 7° 20′ 37″ E; 700 m a.s.l.) in Italy and cultivation lasted for two cycles (2016-2017 and 2017-2018). Both sites were cultivated with saffron for at least the previous three years. Before starting the experiment both fields were milled. To assess the effects of AMF inocula on saffron cultivation and production, the same treatments used in the pot trial were applied (Ri, Ri + Fm or AMF-). A randomized block design was used, with three experimental plot units (blocks). Each plot unit consisted of 56 corms, planted in a 1.44 m² area (39 corms m^-2). Inter-row planting distance was of 7 cm, while between-row distance was 25 cm. Plots were separated from each other with at least 4 m distance. Before planting, 10 g of inoculum was placed under the corms to ensure contact between plant and the treatment. Irrigation was provided when needed and hand weeding control was conducted during cultivation, while no preplanting fertilization, tillage, or treatments against pathogens were applied. The two Alpine sites were characterized by semicontinental climate, with a long and cold winter . In general, both sites had a sandy-loam texture according to the USDA classification and similar chemical characteristics.

The inoculum composed by R. intraradices and F. mosseae was particularly effective in increasing flower production and saffron yield, while R. intraradices alone increased the content of some bioactive compounds-picrocrocin, quercitrin, crocin II-as well as antioxidant activity.

Seeds were treated with sulfuric acid (98%, 10 min) and then surface sterilized with 70% ethanol (v/v) for 1 min and sodium hypochlorite solution (10%, at 10 min). After sterilization, seeds were germinated on MS media (Murashige and Skoog, 1962) containing 7 g/L agar (Duchefa, Netherlands). Cultures were maintained under 16/8 h light/dark. Explants were taken from 4-week-old leaves for inoculation with bacteria strain.ATCC15834 strain of A. rhizogenes was supplied by microbial unit of the National Research Center for Genetic Engineering and Biotechnology, Tehran-Iran. Bacterial cells cultivated on LB (Luria-Bertani) culture medium (Bertani, 1952) on rotary shaker (at 26 ℃ and 180 rpm for 48 h) in the darkened state.The leaves were wounded and inoculated with bacterial suspension for 5 min and transferred to MS media containing 7 g/L agar in darkness at 25 ℃ . After 48 h treated Explants were cultured on the 1/2 MS media containing cefotaxime (500 mg/L) and indole-3-butyric acid (IBA) (2 mg/L). Hairy roots emerged at wounded sites, after 4-weeks of incubation, and then each hairy root line was isolated from explants tissue and was subcultured weekly in new media (1/2 MS hormone-free media) with appropriate antibiotic. The concentration of cefotaxime was decreased gradually and eliminated from the culture medium after 8 subcultures and axenic root cultures were obtained. Then hairy root lines were transferred to the 250 mL Erlenmeyer flasks containing 30 mL hormone- free 1/2 MS liquid medium and incubated on a rotary shaker (120 rpm) at 25 ℃ and subcultured every two week. Hairy root line, which showed sufficient growth in 1/2 MS liquid medium, was selected for further investigations.The genomic DNA was extracted from transformed hairy root lines and plant intact roots with CTAB method . Gene-specific primers from rol B were used for amplification of the 780-bp segment in PCR analysis. The primers sequences were, F:5'-ATGGATCCCAAATTGCTATTCCCCCACGA-3'and R:5'-TTAGGCTTCTTTCATTCGGTTTACTGCAGC-3'. Thirty-five PCR cycles were performed with 5 min initial denaturation at 94 ℃ , annealing steps at 60 ℃ for 80 s, extension at 72 ℃ for 90 s, and final extension step of 72 ℃ for 10 min. The amplimer were analyzed by 1% agarose gel electrophoresis.To investigate the effects of SiO₂ NPs, various concentrations (0, 25, 50, 100 and 200 mg/L) of this elicitor were added to the hairy roots culture medium (1/2 MS + 3% sucrose, pH = 5.7) at the end of log phase of growth stages (21-days-old cultures). Hairy roots were incubated with elicitor for 24 and 48 h of exposure time. Hairy roots were harvested 7 days after elicitation and dried on sterile filter paper to remove excess surface moisture and were weighed before freezing by liquid nitrogen and stored at -80℃ until used to measure growth, biochemical and phytochemical analysis.

The effect of silicon dioxide nanoparticles on production of phenolic compounds and expression rate of pal and ras genes involved in rosmarinic acid biosynthesis pathway has been investigated in D. kotschyi. SiO₂ nanoparticles, used as an abiotic elicitor in our study, has appropriate optical, electrical and catalysts properties and has many applications in various industries as well as agriculture. This study clearly suggested that, in the presence of this nanoparticle, induction, production and accumulation of valuable compounds and corresponding antioxidant activity increased in hairy roots.

According to the results, expression levels of the pal and ras genes were influenced by elicitor concentration and exposure time. The elicitation by SiO₂ NP of 100 mg/L after 48 h of exposure time dramatically increased pal expression compared to the control. Briefly, with increasing SiO₂ NP concentrations after 48 h of exposure time, the expression level of pal was also significantly induced . Similarly, ras expression was significantly raised at 48 h after treatment by increasing SiO₂ NP concentration and enhanced to the greatest extent in 50 mg/L concentration. After 24 h of exposure time, the minimum level of ras expression was observed in the 200 mg/L SiO₂ . Amplification products of real-time PCR were assessed with 1.8% agarose gel which was corresponded to the predicted size.

The experiment was conducted in a 'shade'-type greenhouse with 30% shade at the Instituto de Investigaciones Agropecuarias y Forestales (IIAF), Universidad Michoacana de San Nicolas de Hidalgo (UMSNH), Morelia, Michoacan, Mexico. Maximum and minimum temperatures in the greenhouse varied between 28 and 32 ℃ and between 8 and 18 ℃ respectively. Plants of the strawberry cultivar 'Aromas' were used that had previously been grown in a sterilised (95 ℃ water/steam, 40 min) substrate of coconut fibre/perlite (1:3 v/v) under greenhouse conditions. Before the experiment was established, the absence of AMF in the roots was verified by the ink and vinegar technique, modifying the duration of immersion in KOH and ink/vinegar solution (7 and 5 min respectively). Before planting, roots were disinfected by submerging them for 20 s in 20 g/L sodium hypochlorite solution and rinsing them in water. The inoculum was prepared with spores of Glomus intraradices cultivated in liquid medium (3.5 × 10⁶ spores/L, 90% viability; Premier Tech Biotechnologies Company, Quebec, Canada), which was diluted with fitagel (Sigma P-8169, Saint Louis, MO, USA) solution at 50 g/L to obtain a final concentration of about 5 × 10⁴ spores/L. The viability of spores was determined according to the method of An and Hendrix. Eighteen days after setting up the experiment, each plant received 2 mL of inoculum applied directly to the recently formed roots. One month later, after staining, the percentage of root colonisation was determined by the gridline intersect method. The experiment was organised as a full factorial, completely randomised design with two factors: inoculation (two levels: mycorrhizal and non-mycorrhizal plants) and N concentration in the nutrient solution (three levels: 3, 6 and 18 mmol/L). The six treatments were replicated four times, producing 24 experimental units with ten plants each. Every second day, all plants were irrigated up to substrate saturation. Nitrogen was supplied as NO and the cation/anion ratio was kept constant by varying the concentration of SO. When N was below 18 mmol/L, the cation concentrations were maintained as follows: K+, 3; Ca²⁺, 3.5; Mg²⁺, 1.5 mmol/L. They were increased in the 18 mmol/L N treatment: K+, 6.5; Ca²⁺, 7.5; Mg²⁺, 3.25 mmol/L. In all nutrient solutions the concentration of phosphorus (P) was 0.3 mmol/L. The other nutrients in the solutions were: H₃BO₃, 20; CuSO₄. 5H₂O, 0.5; Fe-EDTA (Ethylenediaminetetraacetic acid iron (III) sodium salt), 15; MnSO₄.H₂O, 12; (NH₄)₆Mo₇O₂₄ . 4H₂O, 0.05; ZnSO₄ . 7H₂O, 3 µmol/L. The pH was adjusted to 5.5 at every application date.

Mycorrhization did not modify the weight, diameter or length of strawberry fruits but had a negative effect on most colour parameters. Moreover, fruits of mycorrhizal plants had higher K and Cu concentrations and showed greater accumulation of most phenolic compounds.

Seeds were sterilized in 1% (v/v) sodium hypochloride (Sigma-Aldrich, USA) for 10 min, then drained and washed with distilled water until they reached neutral pH. They were placed in distilled water and soaked for 6 h at 25 ℃ . Seeds were dark germinated for 8 days in a growth chamber (SANYO MLR-350H) on Petri dishes (125 mm) lined with absorbent paper. Seedlings were watered with 5 ml of Milli-Q water daily. Sprout (8-day-old) samples were gently collected, weighed (fresh mass), rapidly frozen and kept in polyethylene bags at -20 ℃ . For each treatment, three replicates were performed.Elicitation conditions were selected in previous screening studies. For the experiments, temperature (4 ℃ and 40 ℃ - TC and TH, respectively), H₂O2 (20 mM and 200 mM - Ox1 and Ox2, respectively), mannitol (200 mM and 600 mM - Os1 and Os2, respectively) and NaCl (100 mM and 300 mM - S-Os1 and S-Os2, respectively) were selected as abiotic elicitors. All solutions were freshly prepared before each application. Mannitol (Os1, Os2), NaCl (S-O1, S-O2) and H₂O2 (Ox1) treatments were applied by watering daily (not soaking) 2-day-old sprouts with 5 ml of test solution. For Ox2 (200 mM H₂O2) treatment 2-day-old seedlings were only once watered with 5 ml of 200 mM H₂O2 and then cultivated under standard conditions. For temperature conditioning treatment, 2-day-old sprouts were incubated at 4 ℃ and 40 ℃ (TC and TH, respectively) for 1 h and then cultivated under standard conditions. Sprout (8-day-old) samples were gently collected, weighed (fresh mass), rapidly frozen and kept in polyethylene bags at -20 ℃ .

Application of abiotic elicitors (environmental shocks) was an effective method for improvement of sprout pro-health potential via an increase of phenolic contents and subsequent elevation of antioxidant potential. Innovative application of elicitors on 2-day-old sprouts (not seed) allowed the elimination of the unfavorable influence of the factors employed on germination yield and biomass production. Assuming that the optimal germination conditions are those which most effectively increase the antioxidant potential without any negative influence on biomass accumulation and nutritional quality the elicitation with 20 mM H₂O₂ for the future applications is recommended.

Different concentrations of TiO₂ NPs (0, 10, 20, 30, and 50) were prepared for hairy root treatments. 0.5 g of .S. officinalis hairy roots were transferred to 250 mL Erlenmeyer flasks containing 15 mL of liquid MS culture medium with three replicates. Then, they were placed in an incubator shaker at 110 rpm and 25 &#8451 in dark conditions. On the 22nd day, the liquid MS culture media containing different concentrations of nano titanium dioxide was added to Erlenmeyer flasks. 24 and 48 h after treatment, the hairy roots were taken out and transferred to the MS culture medium lacking elicitor.

The highest rate of total phenol (9.79 mg GLA/g FW) and total flavonoid contents (1.06 mg QE/g FW) were obtained in the treated hairy roots with 50 and 30 mg/L of nano elicitor in 24 and 48 h of treatments, respectively. The maximum level of most polyphenols, such as rosmarinic acid, cinnamic acid, and rutin, was produced in 24 h of treatment. The use of TiO₂ NP for 48 h with 50 mg/L concentration showed the highest production level of SO6 protein.