Circadian-based approach for improving physiological, phytochemical and chloroplast proteome in Spinacia oleracea under salinity stress and light emitting diodes.

Salt stress is a recognized annihilating abiotic stress that has a significant impact on agricultural and horticulture crop productivity. Plant development faces three distinct dangers as a result of salt stress: oxidative stress, osmotic stress, and ionic toxicity. It has been shown that plants can forecast diurnal patterns using the circadian clock; moreover, they can manage their defensive mechanism for the detoxification of reactive oxygen species (ROS). Circadian rhythmicity in gene expression assembles transcription and translation feedback networks to govern plant shape, physiology, cellular and molecular activities. Both external and internal variables influence the systemic rhythm via input routes. The Malav Jyoti (MJ) and Delhi Green (DG) genotypes of spinach (Spinacia oleracea) were grown in the plant growth chamber. The chamber had an optimized temperature of 25 °C and humidity of 65% containing light emitting diode (LED) having Red: Blue: white (one side) and White fluorescent (other side) under salinity stress. The samples were collected on the basis of 4 h intervals of circadian hours (0 h, 4 h, 8 h and 12 h) during Day-10 and Day-20 of salt treatments. Under salt stress, the circadian and light-emitting diode-based strategy had a substantial influence on spinach's anti-oxidative responses, stomatal movement, CO2 assimilation, PS-I and II efficiency, phytochrome pigment efficiency, and photosynthesis. Based on the findings of the free radical scavenging enzyme tests, the photoperiodic hours for the proteome analysis were set to 11 am and 3 pm on Day-20. When compared to white fluorescent, this study found that LED has the capacity to influence the entrainment cues of the circadian clock in the cultivation of salt-sensitive spinach genotypes. According to our findings, changing the cellular scavenging mechanism and chloroplast proteome has increased the survival rate of spinach genotypes under LED when compared to white fluorescent.

Effects of biofertilizers on the growth, leaf physiological indices and chlorophyll fluorescence response of spinach seedlings.

Chemcial fertilizer as the main strategy for improving the vegetable yields was excessively applied in recent years which led to progressively serious soil problems such as the soil acidification. According the situation, five different biofertilizer treatments [no fertilizer (CK), inoculations of Bacillus subtilis (Bs, T1), combination of Bs and Bacillus mucilaginosus (Bs+Bm, T2), Bs and Bacillus amyloliquefaciens (Bs+Ba, T3), and Bm+Ba (T4)] were conducted to investigate the effect of the growth, leaf physiological indices, and chlorophyll fluorescence of spinach seedlings in the growth chamber. The growth and physiological indices of the spinach seedlings attained a maximum under the T2 treatments. Under the T2 treatment, the ABS/RC (Absorption flux per RC), TR0/RC (Trapping flux per RC), and ET0/RC (Electron transport flux per RC) was significantly increased, while the DI0/RC [Dissipated energy flux per RC (at t = 0)] was decreased. The OJIP curve was improved under of the inoculations of fertilizers, and the increasing range was the largest under the T2 treatment. The leaf light response curve (LC) was also significantly increased under the T2 treatment. The plant growth characteristics [leaf length (LL), leaf weight (LW), plant height (PH)] were positively correlated with the J-I-P test chlorophyll fluorescence parameters [PIABS (Performance index for energy conservation from exciton to the reduction of intersystem electron acceptors), φP0 (Maximum quantum yield of primary photochemistry), φE0 (Quantum yield of electron transport), ψ0 (The probability that a trapped exciton moved an electron in electron transport chain further than QA-), TR0/RC, and ET0/RC] while negatively correlated with φD0 (Quantum yield of energy dissipation) and DI0/RC. The leaf physiological characteristics [SP (soluble protein concentrations), SC (soluble carbohydrate concentrations), Chl a (chlorophyll a), Chl b (chlorophyll b), Chl a+b, Chl a/b, and WP (water potential)] were positively correlated with the J-I-P test chlorophyll fluorescence parameters (PIABS, φP0, φE0, ψ0, ABS/RC, TR0/RC, and ET0/RC) while negatively correlated with φD0 and DI0/RC. These results indicated that the combination of Bs+Bm inoculations promoted the growth of the spinach and improved the adaptability of the vegetable to acid soil while Ba inoculation didn't have any effects to plants.

Co-application of copper nanoparticles and metal tolerant Bacillus sp. for improving growth of spinach plants in chromium contaminated soil.

Chromium (Cr) is classified as a toxic metal as it exerts harmful effects on plants and human life. Bacterial-assisted nano-phytoremediation is an emerging and environment friendly technique that can be used for the detoxification of such pollutants. In current study, pot experiment was conducted in which spinach plants were grown in soil containing chromium (0, 5, 10, 20 mgkg-1) and treated with selected strain of Bacillus sp. and Cu-O nanoparticle (CuONPs). Data related to plant's growth, physiological parameters, and biochemical tests was collected and analyzed using an appropriate statistical test. It was observed that under chromium stress, all plant's growth parameters were significantly enhanced in response to co-application of CuONPs and Bacillus sp. Similarly, higher levels of catalase, superoxide dismutase, malondialdehyde, and hydrogen peroxide were also observed. However, contents of anthocyanin, carotenoid, total chlorophyll, chlorophyll a & b, were lowered under chromium stress, which were raised in response to the combined application of CuONPs and Bacillus sp. Moreover, this co-application has significant positive effect on total soluble protein, free amino acid, and total phenolics. From this study, it was evident that combined application of Bacillus sp. and CuONP alleviated metal-induced toxicity in spinach plants. The findings from current study may provide new insights for agronomic research for the utilization of bacterial-assisted nano-phytoremediation of contaminated sites.

Changes in the light environment: Short-term responses of photosynthesis and metabolism in spinach.

Modification in the light environment can induce several changes even within a short time. In this article, light intensity and spectrum-dependent changes in photosynthetic and metabolic processes were investigated in spinach leaves. Short-term exposure of the youngest fully developed leaves provided an elevated CO2 assimilation capacity under red light compared with blue or white light, although the electron transport rate was lower. The stomatal opening was mainly stimulated by blue light. These spectrum-induced changes also depended on light intensity. When white light was used to activate the photosynthesis, the white light showed a similar light response to blue light regarding the electron transport processes and red light in terms of stomatal opening. In contrast, concerning CO2 assimilation characteristics, the white light resembled blue light at low and red light at high light intensities. These results indicate that the photosynthetic processes strongly interact with the light intensity and spectral composition. Furthermore, changes in spectral composition modified the primary metabolic processes as well. Red light induced the sugar accumulation, while more organic acids that belong to the respiration pathway were produced under blue and white lights. These changes occurred even within a short (30 min) time frame. These results also draw attention to the importance of the light environment used during the measurements of the photosynthetic activity of plants and/or sample collections.

Formulation of Zinc oxide/Gum acacia nanocomposite as a novel slow-release fertilizer for enhancing Zn uptake and growth performance of Spinacia oleracea L.

Zinc (Zn) deficiency has caused nutritional disorders in 17% of the world's population; thus, producing Zn-enriched plants as a dietary source is necessary. Recently, nanofertilizers have gained much attention as a substitute for conventional fertilizers; however, soil application of polymer-coated Zn-based nanofertilizer has not been explored much. The present study depicts the green synthesis of ZnO nanoparticles using Melia azedarach L. leaf extract, whose phytoconstituents have reducing abilities. The synthesized nanoparticles were combined with gum acacia (GA) to form a ZnOGA nanocomposite. The structural and morphological properties of ZnOGA were studied using XRD, FTIR, FESEM, and EDX. A pot experiment study was carried out with Spinacia oleracea L. at various doses (3, 5, and 10 mg/kg) of the synthesized ZnOGA to evaluate its effectiveness as a slow-release fertilizer and was compared with a commercial Zn fertilizer. The plant growth studies revealed a significant increase in the phyto-morphological traits of the plants fertilized with ZnOGA compared to commercial fertilizer. The plants also displayed significantly higher contents of protein (17-47%), phenols (25-60%), proline (82-94%), total soluble sugar (20-31%), DPPH activity (70-72%), and Zn uptake (91-106%). The doses of ZnOGA played an imperative role in determining the growth and productivity of the plant. Soil column studies showed that ZnOGA reduces Zn leaching by 52% compared to commercial Zn fertilizer. This study signifies the potential of ZnOGA to be applied as an eco-friendly and sustainable substitute for conventional Zn fertilizer minimizing Zn losses and Zn deficiency-related health problems in human populations.

Effect of organic photovoltaic and red-foil transmittance on yield, growth and photosynthesis of two spinach genotypes under field and greenhouse conditions.

The galloping rise in global population in recent years and the accompanying increase in food and energy demands has created land use crisis between food and energy production, and eventual loss of agricultural lands to the more lucrative photovoltaics (PV) energy production. This experiment was carried out to investigate the effect of organic photovoltaics (OPV) and red-foil (RF) transmittance on growth, yield, photosynthesis and SPAD value of spinach under greenhouse and field conditions. Three OPV levels (P0: control; P1: transmittance peak of 0.11 in blue light (BL) and 0.64 in red light (RL); P2: transmittance peak of 0.09 in BL and 0.11 in RL) and two spinach genotypes (bufflehead, eland) were combined in a 3 × 2 factorial arrangement in a completely randomized design with 4 replications in the greenhouse, while two RF levels (RF0: control; RF1: transmittance peak of 0.01 in BL and 0.89 in RL) and two spinach genotypes were combined in a 2 × 2 factorial in randomized complete block design with four replications in the field. Data were collected on growth, yield, photosynthesis and chlorophyll content. Analysis of variance (ANOVA) showed significant reduction in shoot weight and total biomass of spinach grown under very low light intensities as a function of the transmittance properties of the OPV cell used (P2). P1 competed comparably (p > 0.05) with control in most growth and yield traits measured. In addition, shoot to root distribution was higher in P1 than control. RF reduced shoot and total biomass production of spinach in the field due to its inability to transmit other spectra of light. OPV-RF transmittance did not affect plant height (PH), leaf number (LN), and SPAD value but leaf area (LA) was highest in P2. Photochemical energy conversion was higher in P1, P2 and RF1 in contrast to control due to lower levels of non-photochemical energy losses through the Y(NO) and Y(NPQ) pathways. Photo-irradiance curves showed that plants grown under reduced light (P2) did not efficiently manage excess light when exposed to high light intensities. Bufflehead genotype showed superior growth and yield traits than eland across OPV and RF levels. It is therefore recommended that OPV cells with transmittance properties greater than or equal to 11% in BL and 64% in RL be used in APV systems for improved photochemical and land use efficiency.

Evidence of increases of phytol and chlorophyllide by enzymatic dephytylation of chlorophylls in smoothie made from spinach leaves.

Phytol is a diterpene alcohol found abundantly in nature as the phytyl side chain of chlorophylls. Free form of phytol and its metabolites have been attracting attention because they have a potential to improve the lipid and glucose metabolism. On the other hand, phytol is unfavorable for those who suffering from Refsum's disease. However, there is little information on the phytol contents in leafy vegetables rich in chlorophylls. This study indicated that raw spinach leaves contain phytol of 0.4-1.5 mg/100 g fresh weight. Furthermore, crude enzyme extracted from the leaves showed the enzyme activities involved in dephytylation of chlorophyll derivatives and they were high at mild alkaline pH and around 45°C, and lowered at 55°C or above. Under the optimum pH and temperature for such enzymes determined in the model reaction using the crude enzyme, phytol content in the smoothie made from raw spinach leaves increased with an increase of chlorophyllide, another reaction product. Comparison between the increased amounts of phytol and chlorophyllide showed that the enzymatic dephytylation of chlorophylls was critically responsible for the increase of phytol in the smoothie. PRACTICAL APPLICATION: Phytol, which is released by the enzymes related to chlorophyll metabolism in plants, has been investigated because of its potential abilities to improve the lipid metabolism and blood glucose level. In contrast to such health benefits, they are known to be toxic for patients suffering from Refsum's disease. This research for the first time reports the phytol content in raw spinach leaves and that phytol can be increased in the smoothie made from spinach leaves by the action of endogenous enzymes on chlorophyll derivatives under a certain condition. These results help control phytol content in the smoothies.

Effect of Brazilian spinach (Alternanthera sissoo) pellet supplementation and dietary ratios on rumen characteristics, microorganisms, methane production, milk yield, and milk composition in dairy cows.

The aim of the previous research was to evaluate the effects of Brazilian spinach pellet (BSP) supplementation and dietary ratios on rumen characteristics, methane estimation, and milk production in dairy cows. Four crossbred Thai dairy cattle, with Holstein Friesian (HF) cows with a body weight of 442 ± 50 kg were assessed in a 2 × 2 factorial in a 4 × 4 Latin square design to obtain diets; factor A was the roughage (R) to concentrate (C) ratio at 40:60 and 30:70, and factor B was level of BSP supplantation at 2% and 6% of dry matter (basis) intake (DMI). R:C ratio and supplementation of BSP had no interaction effect on DMI and nutrient digestibility. On DM, organic matter (OM), crude protein (CP), and acid detergent fiber (ADF) intake, the R:C ratio increased (p < 0.05). The digestibility of OM improved (p < 0.05) when cows were fed a R:C ratio of 30:70. On pH, ammonia-nitrogen, protozoal population, and blood urea-nitrogen, there were no interactions between the R:C ratio and BSP supplementation. Increasing the BSP supplementation to 6% (p < 0.01) decreased the protozoal population. The R:C ratio of 30:70 increased total volatile fatty acid (VFA) and propionate (C3) concentrations while decreasing the acetate (C2) to C3 ratio and methane (CH4 ) estimation (p < 0.01). The average concentration of total VFA has increased by 114.46 mmol/L for 6% of BSP supplementation. Increased BSP supplementation increased the C3 concentration while decreasing the C2:C3 ratio and CH4 emissions (p < 0.05). The R:C ratio and BSP supplementation had no interaction effect on milk yield, 3.5% fat-corrected milk (FCM), or milk composition. The R:C ratio of 30:70 increased milk yield (p < 0.05) to the highest level of 12.18 kg/day. In conclusion, the diet containing a R:C ratio of 30:70 increased feed intake, milk yield, BUN, total VFA, and C3 concentration, and decreased the C2:C3 ratio and CH4 emission. BSP supplementation at 6% could increase TVFA and C3 concentrations while decreasing the protozoal population and CH4 estimation.

Changes in growth, physiology, and photosynthetic capacity of spinach (Spinacia oleracea L.) under different nitrate levels.

Nitrate content is an essential indicator of the quality of vegetables but can cause stress at high levels. This study aimed to elucidate the regulatory mechanisms of nitrate stress tolerance in spinach (Spinacia oleracea L.). We studied the effects of exogenous application of 15 (control), 50, 100, 150, 200, and 250 mM NO3- on spinach growth, physiology, and photosynthesis. The results showed that all the nitrate treatments inhibited the growth of the aerial parts of spinach compared to the control. In contrast, low nitrate levels (50 and 100 mM) promoted spinach root formation, but this effect was inhibited at high levels (150, 200, and 250 mM). Treatment with 150 mM NO3- significantly decreased the root growth vigor. Low nitrate levels increased the chlorophyll content in spinach leaves, whereas high levels had the opposite effect. High nitrate levels also weakened the net photosynthetic rate (Pn), the actual photochemical efficiency of PSII Y(II), and increased non-photochemical quenching (NPQ), reducing photosynthetic performance. Nitrate stress increased the activity of nitrate reductase (NR) and promoted the accumulation of nitrate in spinach leaves, exceeding the health-tolerance limit for nitrate in vegetables, highlighting the necessity of mitigating nitrate stress to ensure food safety. Starting with the 150 mM NO3- treatment, the proline and malondialdehyde content in spinach leaves and roots increased significantly as the nitrate levels increased. Treatment with 150 mM NO3- significantly increased soluble protein and flavonoid contents, while the activities of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX) were significantly reduced in leaves. However, spinach could resist nitrate stress by regulating the synthesis of osmoregulatory substances such as proline, thus showing some nitrate tolerance. These results provide insights into the physiological regulatory mechanisms of nitrate stress tolerance and its mitigation in spinach, an essential vegetable crop.

Selective accumulation of pharmaceutical residues from 6 different soils by plants: a comparative study on onion, radish, and spinach.

The accumulation of six pharmaceuticals of different therapeutic uses has been thoroughly investigated and compared between onion, spinach, and radish plants grown in six soil types. While neutral molecules (e.g., carbamazepine (CAR) and some of its metabolites) were efficiently accumulated and easily translocated to the plant leaves (onion > radish > spinach), the same for ionic (both anionic and cationic) molecules seems to be minor to moderate. The maximum accumulation of CAR crosses 38,000 (onion), 42,000 (radish), and 7000 (spinach) ng g-1 (dry weight) respectively, in which the most majority of them happened within the plant leaves. Among the metabolites, the accumulation of carbamazepine 10,11-epoxide (EPC - a primary CAR metabolite) was approximately 19,000 (onion), 7000 (radish), and 6000 (spinach) ng g-1 (dry weight) respectively. This trend was considerably similar even when all these pharmaceuticals applied together. The accumulation of most other molecules (e.g., citalopram, clindamycin, clindamycin sulfoxide, fexofenadine, irbesartan, and sulfamethoxazole) was restricted to plant roots, except for certain cases (e.g., clindamycin and clindamycin sulfoxide in onion leaves). Our results clearly demonstrated the potential role of this accumulation process on the entrance of pharmaceuticals/metabolites into the food chain, which eventually becomes a threat to associated living biota.

In situ monitoring of galactolipid digestion by infrared spectroscopy in both model micelles and spinach chloroplasts.

Galactolipids are the main lipids from plant photosynthetic membranes and they can be digested by pancreatic lipase related protein 2 (PLRP2), an enzyme found in the pancreatic secretion in many animal species. Here, we used transmission Fourier-transform infrared spectroscopy (FTIR) to monitor continuously the hydrolysis of galactolipids by PLRP2, in situ and in real time. The method was first developed with a model substrate, a synthetic monogalactosyl diacylglycerol with 8-carbon acyl chains (C8-MGDG), in the form of mixed micelles with a bile salt, sodium taurodeoxycholate (NaTDC). The concentrations of the residual substrate and reaction products (monogalactosylmonoglyceride, MGMG; monogalactosylglycerol, MGG; octanoic acid) were estimated from the carbonyl and carboxylate vibration bands after calibration with reference standards. The results were confirmed by thin layer chromatography analysis (TLC) and specific staining of galactosylated compounds with thymol and sulfuric acid. The method was then applied to the lipolysis of more complex substrates, a natural extract of MGDG with long acyl chains, micellized with NaTDC, and intact chloroplasts isolated from spinach leaves. After a calibration performed with α-linolenic acid, the main fatty acid (FA) found in plant galactolipids, FTIR allowed quantitative measurement of chloroplast lipolysis by PLRP2. A full release of FA from membrane galactolipids was observed, that was not dependent on the presence of bile salts. Nevertheless, the evolution of amide vibration band in FTIR spectra suggested the interaction of membrane proteins with NaTDC and lipolysis products.

Phytoextraction of nickel, lead, and chromium from contaminated soil using sunflower, marigold, and spinach: comparison of efficiency and fractionation study.

Heavy metals in soil pose a serious threat through their toxic effect on the human food chain. Phytoremediation is a clean and green potentially cost-effective technology in remediating the heavy metal-contaminated soil. However, the efficiency of phytoextraction is very often limited by low phytoavailability of heavy metals in soil, slow growth, and small biomass production of hyper-accumulator plants. To solve these issues, accumulator plant(s) with high biomass production and amendment(s) which can solubilize metals in soil is required for better phytoextraction. A pot experiment was conducted to assess the efficiency of phytoextraction of sunflower, marigold, and spinach as affected by the incorporation of Sesbania (solubilizer) and addition of gypsum (solubilizer) in nickel (Ni)-, lead (Pb)-, and chromium (Cr)-contaminated soil. A fractionation study was conducted to study the bioavailability of the heavy metals in contaminated soil after growing the accumulator plants and as affected by using soil amendments (Sesbania and gypsum). Results showed that marigold was the most efficient among the three accumulator plants in phytoextraction of the heavy metals in the contaminated soil. Both sunflower and marigold were able to reduce the bioavailability of the heavy metals in the post-harvest soil, which was reflected in their (heavy metals) lower concentration in subsequently grown paddy crop (straw). The fractionation study revealed that carbonate and organically bound fractions of the heavy metals control the bioavailability of the heavy metals in the experimental soil. Both Sesbania and gypsum were not effective in solubilizing the heavy metals in the experimental soil. Therefore, the possibility of using Sesbania and gypsum for solubilizing heavy metals in contaminated soil is ruled out.

Increasing ß-carotene bioavailability and bioactivity in spinach demonstrated using excipient nanoemulsions-especially those of long-chain triglycerides.

This study sought to improve the biological fate of ß-carotene obtained from spinach, using in vitro digestion, in situ single-pass intestinal perfusion, and in vivo approaches, to investigate the effects of excipient emulsions with medium- (MCT) and long-chain triglyceride (LCT) as a vehicle for improved health benefits of ß-carotene. Results showed that the bioavailability and bioactivity of ß-carotene were both significantly higher in the excipient emulsions relative to those without the emulsions. This was especially true when LCT was used as the vehicle. These results were confirmed by bioaccessibility, duodenal absorption, and in vivo absorption and metabolism. Furthermore, animal feeding studies revealed that LCT may have the potential to promote triglyceride and apo-B48 reconstitution and secretion. This suggested that LCT may facilitate the entry of carotenoids into circulation via the lymphatic pathway. These results highlight the importance of the optimization of excipient foods to improve the efficacy of lipophilic carotenoid.

Decellularized Spinach Biomaterials Support Physiologically Relevant Mechanical Cyclic Strain and Prompt a Stretch-Induced Cellular Response.

Recently, decellularized plant biomaterials have been explored for their use as tissue engineered substitutes. Herein, we expanded upon the investigation of the mechanical properties of these materials to explore their elasticity as many anatomical areas of the body require biomechanical dynamism. We first constructed a device to secure the scaffold and induce a strain within the physiological range of the normal human adult lung during breathing (12-20 movements/min; 10-20% elongation). Results showed that decellularized spinach leaves can support cyclic strain for 24 h and displayed heterogeneous local strain values (7.76-15.88%) as well as a Poisson's ratio (0.12) similar to that of mammalian lungs (10.67-19.67%; 0.01), as opposed to an incompressible homogeneous standard polymer (such as PDMS (10.85-12.71%; 0.4)). Imaging and mechanical testing showed that the vegetal scaffold exhibited strain hardening but maintained its structural architecture and water retention capacity, suggesting an unaltered porosity. Interestingly, we also showed that cells seeded on the scaffold can also sense the mechanical strain as demonstrated by a nuclear reorientation perpendicular to strain direction (63.3° compared to 41.2° for nonstretched cells), a nuclear location of YAP and increased expression of YAP target genes, a high cytoplasmic calcium level, and an elevated expression level of collagen genes (COL1A1, COL3A1, COL4A1, and COL6A) with an increased collagen secretion at the protein level. Taken together, these data demonstrated that decellularized plant leaf tissues have an inherent elastic property similar to that found in the mammalian system to which cells can sense and respond.

Effects of Novel Photosynthetic Inhibitor [CuL2]Br2 Complex on Photosystem II Activity in Spinach.

The effects of the novel [CuL2]Br2 complex (L = bis{4H-1,3,5-triazino [2,1-b]benzothiazole-2-amine,4-(2-imidazole)}copper(II) bromide complex) on the photosystem II (PSII) activity of PSII membranes isolated from spinach were studied. The absence of photosynthetic oxygen evolution by PSII membranes without artificial electron acceptors, but in the presence of [CuL2]Br2, has shown that it is not able to act as a PSII electron acceptor. In the presence of artificial electron acceptors, [CuL2]Br2 inhibits photosynthetic oxygen evolution. [CuL2]Br2 also suppresses the photoinduced changes of the PSII chlorophyll fluorescence yield (FV) related to the photoreduction of the primary quinone electron acceptor, QA. The inhibition of both characteristic PSII reactions depends on [CuL2]Br2 concentration. At all studied concentrations of [CuL2]Br2, the decrease in the FM level occurs exclusively due to a decrease in Fv. [CuL2]Br2 causes neither changes in the F0 level nor the retardation of the photoinduced rise in FM, which characterizes the efficiency of the electron supply from the donor-side components to QA through the PSII reaction center (RC). Artificial electron donors (sodium ascorbate, DPC, Mn2+) do not cancel the inhibitory effect of [CuL2]Br2. The dependences of the inhibitory efficiency of the studied reactions of PSII on [CuL2]Br2 complex concentration practically coincide. The inhibition constant Ki is about 16 µM, and logKi is 4.8. As [CuL2]Br2 does not change the aromatic amino acids' intrinsic fluorescence of the PSII protein components, it can be proposed that [CuL2]Br2 has no significant effect on the native state of PSII proteins. The results obtained in the present study are compared to the literature data concerning the inhibitory effects of PSII Cu(II) aqua ions and Cu(II)-organic complexes.

An oligonucleotide/oligosaccharide-binding-fold protein enhances the alternative splicing event producing thylakoid membrane-bound ascorbate peroxidase in Nicotiana tabacum.

The stromal and thylakoid membrane-bound ascorbate peroxidase isoforms are produced by the alternative splicing event of the 3'-terminal region of the APXII gene in spinach (Spinacia oleracea) and tobacco (Nicotiana tabacum), but not in Arabidopsis (Arabidopsis thaliana). However, all alternative splicing variants were detected in APXII gene-transformed Arabidopsis, indicating the occurrence of its regulatory mechanisms in Arabidopsis. The efficiency of this alternative splicing event in producing thylakoid membrane-bound ascorbate peroxidase mRNA is regulated by a splicing regulatory cis element, but trans splicing regulatory factor(s) for alternative splicing remain unclear. To identify this factor, we conducted a forward genetic screen using Arabidopsis in combination with a luciferase reporter system to evaluate the alternative splicing efficiency of thylakoid membrane-bound ascorbate peroxidase mRNA production. We isolated 9 mutant lines that showed low efficiency of the AS in producing thylakoid membrane-bound ascorbate peroxidase mRNA compared with that in the control plants. From one mutant [APXII alternative splicing inhibition (apsi1)], the causal gene responsible for the phenotype, AT5G38890 (oligonucleotide/oligosaccharide-binding-fold protein, APSI1), was identified. The levels of thylakoid membrane-bound ascorbate peroxidase mRNA from the transformed APXII gene decreased and increased in APSI1 knockout and APSI1-overexpressing plants, respectively. APSI1 was localized to the nucleus and specifically bound to the splicing regulatory cis element sequence. Tobacco plants that disrupted the closest homologs of APSI1 showed low levels of endogenous thylakoid membrane-bound ascorbate peroxidase mRNA. These results indicate that APSI1 is an enhancing component of the alternative splicing event of APXII.

Competitive interaction of Mn(II) and Fe(II) cations with the high-affinity Mn-binding site of the photosystem II: evolutionary aspect.

The evolutionary origin of the oxygen-evolving complex (OEC) in the photosystem II (PSII) is still unclear, as is the nature of electron source for the photosystem before the OEC had appeared. Johnson et al. (in PNAS 110:11238, 2013) speculated that Mn(II) cations were the source of electrons for transitional photosystems. However, Archean oceans also contained Fe(II) cations at concentrations comparable or higher than that of Mn(II). Fe(II) cations can bind to the high-affinity (НА) Mn-binding site in the OEC (Semin et al. in Biochemistry 41:5854, 2002). Now we have investigated the competitive interaction of Mn(II) and Fe(II) cations with the HA site in the Mn-depleted PSII membranes (PSII[-Mn]). Fe cations, oxidized under illumination, bind strongly to the HA site and, thus, prevent the interaction of Mn(II) with this site. If the Mn(II) and Fe(II) cations, at relatively equal concentration, are simultaneously present in the buffer, together with PSII(-Mn) membranes, there is competition between these two cations for the binding site, which manifests itself in partial inhibition of the Mn(II) oxidation and the blocking of the HA site by Fe(II) cations. If the concentration of Fe(II) cations is several times higher than the concentration of Mn(II), the HA site is completely blocked and the oxidation of Mn(II) cations is inhibited; under saturating light, the effectiveness of this inhibitory effect increases. This may be due to the generation of H2O2 on the acceptor side of the photosystem, which significantly accelerates the rate of the turnover reaction of Mn(II) on the HA site.

Silicon fertilization counteracts salinity-induced damages associated with changes in physio-biochemical modulations in spinach.

Plant growth and productivity are limited by the severe impact of salt stress on the fundamental physiological processes. Silicon (Si) supplementation is one of the promising techniques to improve the resilience of plants under salt stress. This study deals with the response of exogenous Si applications (0, 2, 4, and 6 mM) on growth, gaseous exchange, ion homeostasis and antioxidant enzyme activities in spinach grown under saline conditions (150 mM NaCl). Salinity stress markedly reduced the growth, physiological, biochemical, water availability, photosynthesis, enzymatic antioxidants, and ionic status in spinach leaves. Salt stress significantly enhanced leaf Na+ contents in spinach plants. Supplementary foliar application of Si (4 mM) alleviated salt toxicity, by modulating the physiological and photosynthetic attributes and decreasing electrolyte leakage, and activities of SOD, POD and CAT. Moreover, Si-induced mitigation of salt stress was due to the depreciation in Na+/K+ ratio, Na+ ion uptake at the surface of spinach roots, and translocation in plant tissues, thereby reducing the Na+ ion accumulation. Foliar applied Si (4 mM) ameliorates ionic toxicity by decreasing Na+ uptake. Overall, the results illustrate that foliar applied Si induced resistance against salinity stress in spinach by regulating the physiology, antioxidant metabolism, and ionic homeostasis. We advocate that exogenous Si supplementation is a practical approach that will allow spinach plants to recover from salt toxicity.

Spray-dried and freeze-dried protein-spinach particles; effect of drying technique and protein type on the bioaccessibility of carotenoids, chlorophylls, and phenolics.

The effects of protein carrier and drying technique on the concentration and bioaccessibility of lipophilic compounds (lutein, ß-carotene, chlorophylls a and b) and hydrophilic flavonoids in freeze-dried (FD) or spray-dried (SD) spinach juice and protein-spinach particles were investigated. Carotenoid and chlorophyll contents were highest in FD spinach juice without protein (147 and 1355 mg/100 g, respectively). For both SD and FD protein-spinach particles, SPI best protected carotenoids and chlorophylls (123 and 1160 mg/g, respectively), although the bioaccessibility of lipophilic compounds in WPI particles was higher than SPI particles (p < 0.05). For flavonoids, the drying technique was more important than the type of carrier, since FD particles had higher total flavonoids than SD. However, SD particles had higher bioaccessibility for most flavonoids (40-90 %) compared to FD (<20 %). The drying method and protein carrier can be designed to produce protein-spinach ingredients with desired concentration of compounds and bioaccessibility.

Dissipation Behavior and Acute Dietary Risk Assessment of Thiamethoxam and Its Metabolite Clothianidin on Spinach.

Thiamethoxam and its metabolite clothianidin residues pose a potential threat to human health. This study aims to investigate the residue behavior and acute dietary risk assessment of thiamethoxam and clothianidin on spinach. Thiamethoxam and clothianidin were extracted using a quick, easy, cheap, effective, rugged, safe (QuEChERS) method and analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). At spike levels from 0.01 to 5 mg kg−1, the average recoveries of both analytes were in the range of 94.5−105.5%, with relative standard deviations (RSDs) of 3.8−10.9%. The dissipation behavior of thiamethoxam followed first-order kinetics, with half-lives of ≤1.6 days. Clothianidin appeared readily as a plant metabolite with highest level exhibited during 3 to 5 days after application. Temperature and light may be two main factors for degradation of thiamethoxam. Besides, acute risk assessment of thiamethoxam and clothianidin was evaluated with risk quotients (RQs) <100%, which suggested a low health risk for all consumer groups of Chinese residents.