Salt stress amelioration and nutrient strengthening in spinach (Spinacia oleracea L.) via biochar amendment and zinc fortification: seed priming versus foliar application.

Soil salinity is a major nutritional challenge with poor agriculture production characterized by high sodium (Na+) ions in the soil. Zinc oxide nanoparticles (ZnO NPs) and biochar have received attention as a sustainable strategy to reduce biotic and abiotic stress. However, there is a lack of information regarding the incorporation of ZnO NPs with biochar to ameliorate the salinity stress (0, 50,100 mM). Therefore, the current study aimed to investigate the potentials of ZnO NPs application (priming and foliar) alone and with a combination of biochar on the growth and nutrient availability of spinach plants under salinity stress. Results demonstrated that salinity stress at a higher rate (100 mM) showed maximum growth retardation by inducing oxidative stress, resulted in reduced photosynthetic rate and nutrient availability. ZnO NPs (priming and foliar) alone enhanced growth, chlorophyll contents and gas exchange parameters by improving the antioxidant enzymes activity of spinach under salinity stress. While, a significant and more pronounced effect was observed at combined treatments of ZnO NPs with biochar amendment. More importantly, ZnO NPs foliar application with biochar significantly reduced the Na+ contents in root 57.69%, and leaves 61.27% of spinach as compared to the respective control. Furthermore, higher nutrient contents were also found at the combined treatment of ZnO NPs foliar application with biochar. Overall, ZnO NPs combined application with biochar proved to be an efficient and sustainable strategy to alleviate salinity stress and improve crop nutritional quality under salinity stress. We inferred that ZnO NPs foliar application with a combination of biochar is more effectual in improving crop nutritional status and salinity mitigation than priming treatments with a combination of biochar.

Impact of nitrogen fertilizer type and application rate on growth, nitrate accumulation, and postharvest quality of spinach.

Background: A balanced supply of nitrogen is essential for spinach, supporting both optimal growth and appropriate nitrate (NO3 -) levels for improved storage quality. Thus, choosing the correct nitrogen fertilizer type and application rate is key for successful spinach cultivation. This study investigated the effects of different nitrogen (N) fertilizer type and application rates on the growth, nitrate content, and storage quality of spinach plants. Methods: Four fertilizer types were applied at five N doses (25, 50, 200, and 400 mg N kg-1) to plants grown in plastic pots at a greenhouse. The fertilizer types used in the experiment were ammonium sulphate (AS), slow-release ammonium sulphate (SRAS), calcium nitrate (CN), and yeast residue (YR). Spinach parameters like Soil Plant Analysis Development (SPAD) values (chlorophyll content), plant height, and fresh weight were measured. Nitrate content in leaves was analyzed after storage periods simulating post-harvest handling (0, 5, and 10 days). Results: The application of nitrogen fertilizer significantly influenced spinach growth parameters and nitrate content. The YRx400 treatment yielded the largest leaves (10.3 ± 0.5 cm long, 5.3 ± 0.2 cm wide). SPAD values increased with higher N doses for AS, SRAS, and CN fertilizers, with AS×400 (58.1 ± 0.8) and SRAS×400 (62.0 ± 5.8) reaching the highest values. YR treatments showed a moderate SPAD increase. Fresh weight response depended on fertilizer type, N dose, and storage period. While fresh weight increased in all fertilizers till 200 mg kg-1 dose, a decrease was observed at the highest dose for AS and CN. SRAS exhibited a more gradual increase in fresh weight with increasing nitrogen dose, without the negative impact seen at the highest dose in AS and CN. Nitrate content in spinach leaves varied by fertilizer type, dose, and storage day. CNx400 resulted in the highest NO3 - content (4,395 mg kg-1) at harvest (Day 0), exceeding the European Union's safety limit. This level decreased over 10 days of storage but remained above the limit for CN on Days 0 and 5. SRAS and YR fertilizers generally had lower NO3 - concentrations throughout the experiment. Storage at +4 °C significantly affected NO3 - content. While levels remained relatively stable during the first 5 days, a substantial decrease was observed by Day 10 for all fertilizers and doses, providing insights into the spinach's nitrate content over a 10-day storage period. Conclusion: For rapid early growth and potentially higher yields, AS may be suitable at moderate doses (200 mg kg-1). SRAS offers a more balanced approach, promoting sustained growth while potentially reducing NO3 - accumulation compared to AS. Yeast residue, with its slow nitrogen release and consistently low NO3 - levels, could be a viable option for organic spinach production.

Enhancing spinach (Spinacia oleracea L.) resilience in pesticide-contaminated soil: Role of pesticide-tolerant Ciceribacter azotifigens and Serratia marcescens in root architecture, leaf gas exchange attributes and antioxidant response restoration.

This study unveils the detoxification potential of insecticide-tolerant plant beneficial bacteria (PBB), i.e., Ciceribacter azotifigens SF1 and Serratia marcescens SRB1, in spinach treated with fipronil (FIP), profenofos (PF) and chlorantraniliprole (CLP) insecticides. Increasing insecticide doses (25-400 µg kg-1 soil) significantly curtailed germination attributes and growth of spinach cultivated at both bench-scale and in greenhouse experiments. Profenofos at 400 µg kg-1 exhibited maximum inhibitory effects and reduced germination by 55%; root and shoot length by 78% and 81%, respectively; dry matter accumulation in roots and shoots by 79% and 62%, respectively; leaf number by 87% and leaf area by 56%. Insecticide application caused morphological distortion in root tips/surfaces, increased levels of oxidative stress, and cell death in spinach. Application of insecticide-tolerant SF1 and SRB1 strains relieved insecticide pressure resulting in overall improvement in growth and physiology of spinach grown under insecticide stress. Ciceribacter azotifigens improved germination rate (10%); root biomass (53%); shoot biomass (25%); leaf area (10%); Chl-a (45%), Chl-b (36%) and carotenoid (48%) contents of spinach at 25 µg CLP kg-1 soil. PBB inoculation reinvigorated the stressed spinach and modulated the synthesis of phytochemicals, proline, malondialdehyde (MDA), superoxide anions (O2•-), and hydrogen peroxide (H2O2). Scanning electron microscopy (SEM) revealed recovery in root tip morphology and stomatal openings on abaxial leaf surfaces of PBB-inoculated spinach grown with insecticides. Ciceribacter azotifigens inoculation significantly increased intrinsic water use efficiency, transpiration rate, vapor pressure deficit, intracellular CO2 concentration, photosynthetic rate, and stomatal conductance in spinach exposed to 25 µg FIP kg-1. Also, C. azotifigens and S. marcescens modulated the antioxidant defense systems of insecticide-treated spinach. Bacterial strains were strongly colonized to root surfaces of insecticide-stressed spinach seedlings as revealed under SEM. The identification of insecticide-tolerant PBBs such as C. azotifigens and S. marcescens hold the potential for alleviating abiotic stress to spinach, thereby fostering enhanced and safe production within polluted agroecosystems.

Exogenous curcumin mitigates As stress in spinach plants: A biochemical and metabolomics investigation.

The spinach (S. oleracea L.) was used as a model plant to investigate As toxicity on physio-biochemical processes, exploring the potential mitigation effect of curcumin (Cur) applied exogenously at three concentrations (1, 10, and 20 µM Cur). The employment of Cur significantly mitigated As-induced stress in spinach photosynthetic performance (Fv/Fm, Fo/Fm, and Fv/Fo). Moreover, the co-incubation of Cur with As improved physiological processes mainly associated with plant water systems affected by As stress by recovering the leaf's relative water content (RWC) and osmotic potential (ψπ) nearly to the control level and increasing the transpiration rate (E; 39-59%), stomatal conductivity (gs; 86-116%), and carbon assimilation rate (A; 84-121%) compared to As stressed plants. The beneficial effect of Cur in coping with As-induced stress was also assessed at the plant's oxidative level by reducing oxidative stress biomarkers (H2O2 and MDA) and increasing non-enzymatic antioxidant capacity. Untargeted metabolomics analysis was adopted to investigate the main processes affected by As and Cur application. A multifactorial ANOVA discrimination model (AMOPLS-DA) and canonical correlation analysis (rCCA) were employed to identify relevant metabolic changes and biomarkers associated with Cur and As treatments. The results highlighted that Cur significantly determined the accumulation of glucosinolates, phenolic compounds, and an increase in glutathione redox cycle activities, suggesting an overall elicitation of plant secondary metabolisms. Specifically, the correlation analysis reported a strong and positive correlation between (+)-dihydrokaempferol, L-phenylalanine (precursor of phenolic compounds), and serotonin-related metabolites with antioxidant activities (ABTS and DPPH), suggesting the involvement of Cur application in promoting a cross-talk between ROS signaling and phytohormones, especially melatonin and serotonin, working coordinately to alleviate As-induced oxidative stress. The modulation of plant metabolism was also observed at the level of amino acids, fatty acids, and secondary metabolites synthesis, including N-containing compounds, terpenes, and phenylpropanoids to cooperate with As-induced stress response.

The mechanism behind tenuazonic acid-mediated inhibition of plant plasma membrane H+-ATPase and plant growth.

The increasing prevalence of herbicide-resistant weeds has led to a search for new herbicides that target plant growth processes differing from those targeted by current herbicides. In recent years, some studies have explored the use of natural compounds from microorganisms as potential new herbicides. We previously demonstrated that tenuazonic acid (TeA) from the phytopathogenic fungus Stemphylium loti inhibits the plant plasma membrane (PM) H+-ATPase, representing a new target for herbicides. In this study, we further investigated the mechanism by which TeA inhibits PM H+-ATPase and the effect of the toxin on plant growth using Arabidopsis thaliana. We also studied the biochemical effects of TeA on the PM H+-ATPases from spinach (Spinacia oleracea) and A. thaliana (AHA2) by examining PM H+-ATPase activity under different conditions and in different mutants. Treatment with 200 µM TeA-induced cell necrosis in larger plants and treatment with 10 µM TeA almost completely inhibited cell elongation and root growth in seedlings. We show that the isoleucine backbone of TeA is essential for inhibiting the ATPase activity of the PM H+-ATPase. Additionally, this inhibition depends on the C-terminal domain of AHA2, and TeA binding to PM H+-ATPase requires the Regulatory Region I of the C-terminal domain in AHA2. TeA likely has a higher binding affinity toward PM H+-ATPase than the phytotoxin fusicoccin. Finally, our findings show that TeA retains the H+-ATPase in an inhibited state, suggesting that it could act as a lead compound for creating new herbicides targeting the PM H+-ATPase.

Effect of zinc nanoparticles seed priming and foliar application on the growth and physio-biochemical indices of spinach (Spinacia oleracea L.) under salt stress.

Salt stress is the major risk to the seed germination and plant growth via affecting physiological and biochemical activities in plants. Zinc nanoparticles (ZnNPs) are emerged as a key agent in regulating the tolerance mechanism in plants under environmental stresses. However, the tolerance mechanisms which are regulated by ZnNPs in plants are still not fully understood. Therefore, the observation was planned to explore the role of ZnNPs (applied as priming and foliar) in reducing the harmful influence of sodium chloride (NaCl) stress on the development of spinach (Spinacia oleracea L.) plants. Varying concentrations of ZnNPs (0.1%, 0.2% & 0.3%) were employed to the spinach as seed priming and foliar, under control as well as salt stress environment. The alleviation of stress was observed in ZnNPs-applied spinach plants grown under salt stress, with a reduced rise in the concentration hydrogen peroxide, melondialdehyde and anthocyanin contents. A clear decline in soluble proteins, chlorophyll contents, ascorbic acid, sugars, and total phenolic contents was observed in stressed conditions. Exogenous ZnNPs suppressed the NaCl generated reduction in biochemical traits, and progress of spinach plants. However, ZnNPs spray at 0.3% followed by priming was the most prominent treatment in the accumulation of osmolytes and the production of antioxidant molecules in plants.

Photosynthesis-Inhibiting Activity of N-(Disubstituted-phenyl)-3-hydroxynaphthalene-2-carboxamides.

A set of twenty-four 3-hydroxynaphthalene-2-carboxanilides, disubstituted on the anilide ring by combinations of methoxy/methyl/fluoro/chloro/bromo and ditrifluoromethyl groups at different positions, was prepared. The compounds were tested for their ability to inhibit photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. N-(3,5-Difluorophenyl)-, N-(3,5-dimethylphenyl)-, N-(2,5-difluorophenyl)- and N-(2,5-dimethylphenyl)-3-hydroxynaphthalene-2-carboxamides showed the highest PET-inhibiting activity (IC50 ~ 10 µM) within the series. These compounds were able to inhibit PET in photosystem II. It has been found that PET-inhibiting activity strongly depends on the position of the individual substituents on the anilide ring and on the lipophilicity of the compounds. The electron-withdrawing properties of the substituents contribute towards the PET activity of these compounds.

Singlet oxygen damages the function of Photosystem II in isolated thylakoids and in the green alga Chlorella sorokiniana.

Singlet oxygen (1O2) is an important damaging agent, which is produced during illumination by the interaction of the triplet excited state pigment molecules with molecular oxygen. In cells of photosynthetic organisms 1O2 is formed primarily in chlorophyll containing complexes, and damages pigments, lipids, proteins and other cellular constituents in their environment. A useful approach to study the physiological role of 1O2 is the utilization of external photosensitizers. In the present study, we employed a multiwell plate-based screening method in combination with chlorophyll fluorescence imaging to characterize the effect of externally produced 1O2 on the photosynthetic activity of isolated thylakoid membranes and intact Chlorella sorokiniana cells. The results show that the external 1O2 produced by the photosensitization reactions of Rose Bengal damages Photosystem II both in isolated thylakoid membranes and in intact cells in a concentration dependent manner indicating that 1O2 plays a significant role in photodamage of Photosystem II.

Photophysics of DFHBI bound to RNA aptamer Baby Spinach.

The discovery of the GFP-type dye DFHBI that becomes fluorescent upon binding to an RNA aptamer, termed Spinach, led to the development of a variety of fluorogenic RNA systems that enable genetic encoding of living cells. In view of increasing interest in small RNA aptamers and the scarcity of their photophysical characterisation, this paper is a model study on Baby Spinach, a truncated Spinach aptamer with half its sequence. Fluorescence and fluorescence excitation spectra of DFHBI complexes of Spinach and Baby Spinach are known to be similar. Surprisingly, a significant divergence between absorption and fluorescence excitation spectra of the DFHBI/RNA complex was observed on conditions of saturation at large excess of RNA over DFHBI. Since absorption spectra were not reported for any Spinach-type aptamer, this effect is new. Quantitative modelling of the absorption spectrum based on competing dark and fluorescent binding sites could explain it. However, following reasoning of fluorescence lifetimes of bound DFHBI, femtosecond-fluorescence lifetime profiles would be more supportive of the notion that the abnormal absorption spectrum is largely caused by trans-isomers formed  within the cis-bound DFHBI/RNA complex. Independent of the origin, the unexpected discrepancy between absorption and fluorescence excitation spectra allows for easily accessed screening and insight into the efficiency of a fluorogenic dye/RNA system.

Effect of the suspension of Ag-incorporated TiO2 nanoparticles (Ag-TiO2 NPs) on certain growth, physiology and phytotoxicity parameters in spinach seedlings.

In this work, the effect of the inoculation of silver-incorporated titanium dioxide nanoparticles (Ag-TiO2 NPs) in spinach seeds was evaluated on certain growth, physiology and phytotoxicity parameters of the plants. This is an important crop for human consumption with high nutritional value due to their low calorie and fat content, providing various vitamins and minerals, especially iron. These NPs were obtained by means of the sol-gel method and heat treatment; the resulting powder material was characterized using X-ray diffraction and scanning electron microscopy and the influence of these NPs on plants was measured by estimating the germination rate, monitoring morphological parameters and evaluating phytotoxicity. The photosynthetic activity of the spinach plants was estimated through the quantification of the Ratio of Oxygen Evolution (ROE) by the photoacoustic technique. Samples of TiO2 powder with particle size between 9 and 43 nm were used to quantify the germination rate, which served to determine a narrower size range between 7 and 26 nm in the experiments with Ag-TiO2 NPs; the presence of Ag in TiO2 powder samples was confirmed by energy-dispersive X-ray spectroscopy. The analysis of variance showed that the dependent variable (plant growth) could be affected by the evaluated factors (concentration and size) with significant differences. The statistical trend indicated that the application of the Ag-TiO2 NPs suspension of lowest concentration and smallest particle size could be a promoting agent of the growth and development of these plants. The inoculation with NPs of 8.3 nm size and lowest concentration was related to the highest average ROE value, 24.6 ± 0.2%, while the control group was 20.2 ± 0.2%. The positive effect of the Ag-TiO2 NPs treatment could be associated to the generation of reactive oxygen species, antimicrobial activity, increased biochemical attributes, enzymatic activity or improvements in water absorption.

Role of sulfur in combating arsenic stress through upregulation of important proteins, and in-silico analysis to study the interaction between phosphate transporter (PHO1), arsenic and phosphate in spinach.

An adequate amount of Sulfur (S) is essential for proper plant growth and defence against abiotic stresses including metals and metalloids. Arsenic (As) contamination is increasing in agricultural soils rapidly due to anthropogenic activities. Sulfur deficiency and arsenic stress could be more harmful than these individual stresses alone. To understand the impact of S-deficiency and arsenic (31 ppm Na3AsO4 of soil) on ecophysiology, growth, inorganic phosphate level, and proteomic profile of spinach, the present study was conducted. Interaction of arsenic with phosphate transporters, phytochelatins, and glutathione was also analyzed in silico. Comparative 2D MS/MS proteomics helped in the identification of important proteins which might be the key players under S-deficiency and As stress. Upregulation and downregulation of 36 and 21 proteins under As stress; 19 and 36 proteins under S-deficiency; 38 and 31 proteins under combined stress, respectively was observed. A total, 87 proteins subjected to identification via MS/MS ion search were found to be associated with important plant functions. PHO1 abundance was highly influenced by As stress; hence an in-silico homology modeling based molecular docking was performed which indicated high interaction between PHO1 and As/phosphate. Varied proximity of arsenic with phosphate transporters, phytochelatin, and glutathione revealed these components as a potential target of As toxicity/detoxification in Spinach, reflecting sulfur as an important criterion for arsenic tolerance.

Binary mixture of nanoparticles in sewage sludge: Impact on spinach growth.

Present study carried out pot experiments and evaluated effects of single and binary mixture of nanoparticles (exposed via sludge as soil conditioner) on spinach plant. Exposure of Ag2O nanoparticles (NPs) (1 and 10 mg/kg soil-sludge) did not show significant reduction in plant as compared to control. On the other hand, TiO2 NPs (exposed as single and in binary mixture) resulted in significant increase in root length (29% and 37%) and fresh weight (60% and 48%) at highest exposure concentration. Total chlorophyll content decreased for Ag2O and binary mixture (7% and 4%, respectively) and increased for TiO2 (5%) at 10 mg/kg soil-sludge. The toxic interaction between Ag2O and TiO2 NPs was additive at both exposure concentrations. Ag2O NPs had higher tendency of root surface adsorption than TiO2 NPs. Metal content in spinach leaves at highest exposure concentration was Ag: 2.6 ± 0.55 mg/g plant biomass(for Ag2O NPs) and 1.02 ± 0.32 mg/g plant biomass (for Ag2O + TiO2 NPs) and for Ti: 1.12 ± 0.78 (for TiO2 NPs) mg/g plant biomass and 0.58 ± 0.41 mg/g (for Ag2O + TiO2 NPs). The inadvertent ingestion of NPs- contaminated spinach resulted in projected daily intake (DI) of Ag and Ti for different age-mass classes (child to adult) exceeding the oral reference dose for toxicity during oral ingestion. In conclusion, we report no acute toxicity of single and binary mixture of NPs to spinach but significant accumulation of Ag and Ti metals in spinach leaves. There are high chances that ingestion of spinach grown in such environment might lead to human health risks.

Combined effects of ultrasound and aqueous chlorine dioxide treatments on nitrate content during storage and postharvest storage quality of spinach (Spinacia oleracea L.).

The objectives of this study were to optimize the condition of ultrasonic treatment combined with aqueous chlorine dioxide (ClO2) on nitrate content of spinach by response surface methodology (RSM), and determine the effectiveness of ultrasound (US) and ClO2 alone and in combination, on spinach postharvest quality during 7 days' storage period. The optimal treatment parameters obtained were ultrasonic power (300 W), ClO2 concentration (50 ppm), treatment time (4 min). The combined treatments significantly reduced the nitrate content and maintained better storage quality in terms of total soluble solids (TSS) and ascorbic acid content compared with the individual treatment or untreated. For Chlorophyll content, the combined treatment was significantly higher than the control and ClO2 treatment, but lower than ultrasonic treatment. The results demonstrated that US combined with ClO2 are promising alternatives for the reduction of nitrate content, as well as preserving the quality of stored leafy vegetables.

Potential role of compost mixed biochar with rhizobacteria in mitigating lead toxicity in spinach.

Consumption of heavy metals, especially lead (Pb) contaminated food is a serious threat to human health. Higher Pb uptake by the plant affects the quality, growth and yield of crops. However, inoculation of plant growth-promoting rhizobacteria (PGPR) along with a mixture of organic amendments and biochar could be an effective way to overcome the problem of Pb toxicity. That's why current pot experiment was conducted to investigate the effect of compost mixed biochar (CB) and ACC deaminase producing PGPR on growth and yield of spinach plants under artificially induced Pb toxicity. Six different treatments i.e., control, Alcaligenes faecalis (PGPR1), Bacillus amyloliquefaciens (PGPR2), compost + biochar (CB), PGPR1 + CB and PGPR2 + CB were applied under 250 mg Pb kg-1 soil. Results showed that inoculation of PGPRs (Alcaligenes faecalis and Bacillus amyloliquefaciens) alone and along with CB significantly enhanced root fresh (47%) and dry weight (31%), potassium concentration (11%) in the spinach plant. Whereas, CB + Bacillus amyloliquefaciens significantly decreased (43%) the concentration of Pb in the spinach root over control. In conclusion, CB + Bacillus amyloliquefaciens has the potential to mitigate the Pb induced toxicity in the spinach. The obtained result can be further used in the planning and execution of rhizobacteria and compost mixed biochar-based soil amendment.

Comparative analysis of tissue-specific transcriptomic responses to nitrogen stress in spinach (Spinacia oleracea).

Nitrogen (N) is critical to the growth and productivity of crops. To understand the molecular mechanisms influenced by N stress, we used RNA-Sequencing (RNA-Seq) to analyze differentially expressed genes (DEGs) in root and leaf tissues of spinach. N stress negatively influenced photosynthesis, biomass accumulation, amino acid profiles, and partitioning of N across tissues. RNA-seq analysis revealed that N stress caused most transcriptomic changes in roots, identifying 1,346 DEGs. High-affinity nitrate transporters (NRT2.1, NRT2.5) and glutamine amidotransferase (GAT1) genes were strongly induced in roots in response to N deplete and replete conditions, respectively. GO and KEGG analyses revealed that the functions associated with metabolic pathways and nutrient reservoir activity were enriched due to N stress. Whereas KEGG pathway enrichment analysis indicated the upregulation of DEGs associated with DNA replication, pyrimidine, and purine metabolism in the presence of high N in leaf tissue. A subset of transcription factors comprising bHLH, MYB, WRKY, and AP2/ERF family members was over-represented in both tissues in response to N perturbation. Interesting DEGs associated with N uptake, amino acid metabolism, hormonal pathway, carbon metabolism, along with transcription factors, were highlighted. The results provide valuable information about the underlying molecular processes in response to N stress in spinach and; could serve as a resource for functional analysis of candidate genes/pathways and enhancement of nitrogen use efficiency.

Biochar counteracts nitrification inhibitor DMPP-mediated negative effect on spinach (Spinacia oleracea L.) growth.

The use of nitrification inhibitors (NIs) such as 3,4-dimethylpyrazole phosphate (DMPP) has been suggested to diminish agricultural soil nitrate (NO3-) loss and increase nitrogen (N) use efficiency (NUE). However, the yield of ammonium (NH4+)-sensitive plants such as spinach (Spinacia oleracea L.) may be adversely affected by the application of NIs at high N levels and, on the other hand, the efficiency of the NIs may also be affected by soil amendments such as biochar. These two issues are still not adequately addressed. The aim of this study was to evaluate the effect of different N levels including DMPP or not in a calcareous soil with and without amendment of wheat straw biochar on spinach yield, NUE, nitrate concentration of spinach leaf, activity of enzymes nitrate reductase (NR) and nitrite reductase (NiR), and soil ammonium (NH4+) and NO3- concentration under greenhouse conditions. This experiment was carried out with different N rates factor at seven levels (un-fertilized, N0; fertilized with 50 mg N kg-1 soil, N50; fertilized with 75 mg N kg-1 soil, N75; fertilized with 100 mg N kg-1 soil, N100; fertilized with N50 + DMPP; fertilized with N75 + DMPP; and fertilized with N100 + DMPP) and biochar (BC) factor at two levels (0, 0%BC; and 2% (w/w), 2%BC) with six replications over a 56-day cultivation period of spinach. Results showed that the application of DMPP had no significant effect on the yield of spinach plant at low and medium levels of N (50 and 75 mg N kg-1 soil), but decreased the yield of this plant at the higher level of N (100 mg N kg-1 soil). However, application of BC decreased the negative effect of DMPP on spinach yield as the yield in spinach plants fertilized with N75 + DMPP and N100 + DMPP significantly increased. Both application of DMPP and addition of BC to soil decreased leaf NO3- concentration by 29.2% and 16.3% compared to control, respectively. Biochar compared to control decreased NR activity by 46.3%. With increasing N rate, NR and NiR activities increased, but DMPP decreased the activities of both enzymes. Biochar reduced the efficiency of DMPP as soil NH4+ concentration was higher in the treatments containing DMPP without BC at 56 days after planting. Biochar and DMPP could increase the quality of spinach plant through decreasing the leaf NO3- concentration. In general, wheat straw biochar counteracted DMPP-mediated negative effect on growth of spinach plant at high level of N by decreasing the efficiency of this inhibitor. These results provide the useful information for managing the application rate of N fertilizers including DMPP in biochar-amended soil.

Risk assessment and biophysiochemical responses of spinach to foliar application of lead oxide nanoparticles: A multivariate analysis.

Despite extensive research progress in the recent past, the data regarding foliar uptake of heavy metals, associated biophysiochemical changes inside plants and possible health hazards are limited. This study determined the effect of foliar application of lead oxide nanoparticles (PbO-NPs) on lead (Pb) accumulation, physiological and biochemical changes inside spinach plants and associated health risks. A green method was used to prepare PbO-NPs using coconut water. Scanning electron microscopy (SEM) showed the preparation of smooth, unwrinkled, granular and spherical PbO-NPs. Spinach leaves were exposed via foliar application to three concentrations of PbO-NPs (0, 10 and 50 mg/plant). Foliar PbO-NPs application resulted in a significant accumulation of Pb in leaves (42.25 µg g-1), with limited translocation towards root tissues (4.46 µg g-1). This revealed that spinach can accumulate considerable amount of Pb via foliar uptake. Lead accumulation inside spinach caused a significant decrease in pigment contents (38%) and dry weight (67%). After foliar uptake, Pb caused several-fold increase in the activities of catalase and peroxidase. However, foliar PbO-NPs did not induce significant changes in H2O2 production, lipid peroxidation and superoxide dismutase activity. Application of PbO-NPs (50 mg/plant) showed possible health risks (non-carcinogenic) due to ingesting Pb-contaminated leaves of spinach. It is proposed that atmospheric contamination and foliar deposition of metal-PM can seriously affect vegetable growth and can provoke health issues due to ingestion of metal-enriched vegetables. Therefore, atmospheric levels of heavy metals need to be monitored on a regular basis to avoid their food chain contamination and possible human exposure.

Physiological response of spinach to toxic heavy metal stress.

This study was carried out to investigate the concentration of two heavy metals, i.e., mercury (Hg) and arsenic (As) in soil and plant. Spinach (Spinacia oleracea L.) was used as a test vegetable in a pot experiment. Five spiked concentrations of both the metals along with sewage water were used as treatments. The analyses of the metals were determined in two cuttings. The results showed significant effect of treatments on the concentration of the two metals in soil and plant. The concentrations of As recorded were higher in 1st spinach cutting and reduced in the second harvest. However, comparing the two metal concentrations, it was found that As was absorbed greater as compared with Hg. Analyzing the plant growth parameter, it was found that metal stress has significantly influenced the plant growth. In sewage water pots, As was significantly higher than Hg. The transfer factor from soil to plant showed higher As in plants at lower concentration, but at higher As levels, the transfer rate declined, while Hg showed it was completely inverse. Positive correlation was found between soil applied metal concentration and plant uptake. It may be concluded from the above results that spinach is a good accumulator of heavy metals and has shown significant result of both As and Hg accumulation in plant. The concentration increased with the increasing concentration in soil.

Metal(loid) induced toxicity and defense mechanisms in Spinacia oleracea L.: Ecological hazard and Prospects for phytoremediation.

A pot study was conducted to assess the phytoremediation potential of Spinach plants along with their physiological and biochemical response when grown in soil contaminated with heavy metal(loid)s (HMs). Plants were grown under different doses of Pb, Cd and As; and their metal(loid) accumulation efficiency was studied upon harvest; expressed in terms of bioabsorption coefficient (BAC), bioconcentration factor (BCF) and translocation factor (TF). Results showed significant (p ≤ 0.05) difference in physiological and biochemical mechanisms of plants as detected through decrease in concentration of cellular constituents (pigments, carbohydrates, total nitrogen content); and increase in antioxidants (both enzymatic and non-enzymatic). Despite of accumulating high amount of HMs in tissues, no visible signs of toxicity were seen; and hence the efficient survival and defense mechanism shown by spinach plants conclude that they are a viable option to be used for phytoremediation of sites contaminated with Cd and Pb. Since the content of Cd and Pb in edible part was higher than safe limits prescribed by USEPA, the present investigation also highlights the ecological hazards that may result upon cultivation of spinach in contaminated soil for agricultural purpose; or its accidental exposure to food chain when grown for phytoremediation.

Bioactivity of Methoxylated and Methylated 1-Hydroxynaphthalene-2-Carboxanilides: Comparative Molecular Surface Analysis.

A series of twenty-six methoxylated and methylated N-aryl-1-hydroxynaphthalene- 2-carboxanilides was prepared and characterized as potential anti-invasive agents. The molecular structure of N-(2,5-dimethylphenyl)-1-hydroxynaphthalene-2-carboxamide as a model compound was determined by single-crystal X-ray diffraction. All the analysed compounds were tested against the reference strain Staphylococcus aureus and three clinical isolates of methicillin-resistant S. aureus as well as against Mycobacterium tuberculosis and M. kansasii. In addition, the inhibitory profile of photosynthetic electron transport in spinach (Spinacia oleracea L.) chloroplasts was specified. In vitro cytotoxicity of the most effective compounds was tested on the human monocytic leukaemia THP-1 cell line. The activities of N-(3,5-dimethylphenyl)-, N-(3-fluoro-5-methoxy-phenyl)- and N-(3,5-dimethoxyphenyl)-1-hydroxynaphthalene-2-carbox- amide were comparable with or even better than the commonly used standards ampicillin and isoniazid. All promising compounds did not show any cytotoxic effect at the concentration >30 µM. Moreover, an in silico evaluation of clogP features was performed for the entire set of the carboxamides using a range of software lipophilicity predictors, and cross-comparison with the experimentally determined lipophilicity (log k), in consensus lipophilicity estimation, was conducted as well. Principal component analysis was employed to illustrate noticeable variations with respect to the molecular lipophilicity (theoretical/experimental) and rule-of-five violations. Additionally, ligand-oriented studies for the assessment of the three-dimensional quantitative structure-activity relationship profile were carried out with the comparative molecular surface analysis to determine electron and/or steric factors that potentially contribute to the biological activities of the investigated compounds.