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Quinoa ( Chenopodium quinoa Willd), an ancient Andean seed crop, exhibits exceptional nutritional properties and resistance to abiotic stress. The species' tolerance to heavy metals has, however, not yet been investigated nor its ability to take up and translocate chromium (Cr). This study aimed to investigate the metabolic adjustments occurring upon exposure of quinoa to several concentrations (0.01–5 mM) of CrCl 3 . Young hydroponically grown plants were used to evaluate Cr uptake, growth, oxidative stress, and other biochemical parameters three and/or seven days after treatment. Leaves accumulated the lowest amounts of Cr, while roots and stems accumulated the most at low and at high metal concentrations, respectively. Fresh weight and photosynthetic pigments were reduced only by the higher Cr(III) doses. Substantially increased lipid peroxidation, hydrogen peroxide, and proline levels were observed only with 5 mM Cr(III). Except for a significant decrease at day 7 with 5 mM Cr(III), total polyphenols and flavonoids maintained control levels in Cr(III)-treated plants, whereas antioxidant activity increased in a dose-dependent manner. Maximum polyamine accumulation was observed in 1 mM CrCl 3 -treated plants. Even though α- and γ-tocopherols also showed enhanced levels only with the 1 mM concentration, tyrosine aminotransferase (TAT, EC 2.6.1.5) activity increased under Cr(III) treatment in a dose- and time-dependent manner. Taken together, results suggest that polyamines, tocopherols, and TAT activity could contribute to tolerance to 1 mM Cr(III), but not to the highest concentration that, instead, generated oxidative stress. [ABSTRACT FROM AUTHOR]

Quinoa (Chenopodium quinoa Willd) is an ancient Andean seed-producing crop well known for its exceptional nutritional properties and resistance to adverse environmental conditions, such as salinity and drought. Seed storage proteins, amino acid composition, and bioactive compounds play a crucial role in determining the nutritional value of quinoa. Seeds harvested from three Chilean landraces of quinoa, one belonging to the salares ecotype (R49) and two to the coastal-lowlands ecotype, VI-1 and Villarrica (VR), exposed to two levels of salinity (100 and 300 mM NaCl) were used to conduct a sequential extraction of storage proteins in order to obtain fractions enriched in albumins/globulins, 11S globulin and in prolamin-like proteins. The composition of the resulting protein fractions was analyzed by one- and two-dimensional polyacrylamide gel electrophoresis. Results confirmed a high polymorphism in seed storage proteins; the two most representative genotype-specific bands of the albumin/globulin fraction were the 30- and 32-kDa bands, while the 11S globulin showed genotype-specific polymorphism for the 40- and 42-kDa bands. Spot analysis by mass spectrometry followed by in silico analyses were conducted to identify the proteins whose expression changed most significantly in response to salinity in VR. Proteins belonging to several functional categories (i.e., stress protein, metabolism, and storage) were affected by salinity. Other nutritional and functional properties, namely amino acid profiles, total polyphenol (TPC) and flavonoid (TFC) contents, and antioxidant activity (AA) of protein extracts were also analyzed. With the exception of Ala and Met in R49, all amino acids derived from protein hydrolysis were diminished in seeds from salt-treated plants, especially in landrace VI-1. By contrast, several free amino acids were unchanged or increased by salinity in R49 as compared with VR and VI-1, suggesting a greater tolerance in the salares landrace. VR had the highest TPC and AA under non-saline conditions. Salinity increased TPC in all three landraces, with the strongest increase occurring in R49, and enhanced radical scavenging capacity in R49 and VR. Overall, results show that salinity deeply altered the seed proteome and amino acid profiles and, in general, increased the concentration of bioactive molecules and AA of protein extracts in a genotype-dependent manner. [ABSTRACT FROM AUTHOR]

Poplar is a suitable species for phytoremediation, able to tolerate high concentrations of heavy metals (HMs). Arbuscular mycorrhizal fungi (AMF) form symbiotic associations with the roots of most land plants; they improve nutrient uptake and enhance phytoextraction of HMs while alleviating stress in the host plant. This review summarizes previous results from field and greenhouse studies conducted by us and dealing with this topic. In a field trial on a highly Zn- and Cu-contaminated site, differences in plant survival and growth were observed among 168 clones originating from natural populations of Populus alba L. and Populus nigra L. from northern Italy. After two and a half years from planting, the density, activity and metabolic versatility of the culturable fraction of the soil bacteria in the HM-polluted field was higher in the soil close to where larger poplar plants were growing, in spite of comparable HM concentrations recorded in these soils. One well-performing clone of P. alba (AL35), which accumulated a higher concentration of both metals and had high foliar polyamine (PA) levels, was used for further investigation. In a greenhouse study, AL35 cuttings pre-inoculated with AMF (Glomus mosseae or Glomus intraradices) and then transferred to pots containing soil, collected from the HM-polluted site, displayed growth comparable to that of controls grown on unpolluted soil, in spite of higher Cu and Zn accumulation. Such plants also showed an overall up-regulation of metallothionein (MT) and PA biosynthetic genes, together with increased PA levels. A genome-wide transcriptomic (cDNA- AFLP) analysis allowed the identification of a number of genes, mostly belonging to stress-related functional categories of defense and secondary metabolism, that were differentially regulated in mycorrhizal vs. non mycorrhizal plants. A proteomic analysis revealed that, depending on sampling time, changes in protein profiles were differentially affected by AMF and/or HMs. It is concluded that soil-borne microorganisms affect plant performance on HMpolluted soil. In particular, mycorrhizal plants exhibited increased capacity for phytostabilization of HMs, together with improved growth. Their greater stress tolerance may derive from the protective role of PAs, and from the strong modulation in the expression profiles of stress-related genes and proteins. [ABSTRACT FROM AUTHOR]

In the present study, the effects of epibrassinolide (EBL) on indole-3-acetic acid (IAA), abscisic acid (ABA) and polyamine (PA) tissue concentrations and antioxidant potential of 7-day-old Raphanus sativus L. cv. 'Pusa chetki' seedlings grown under Cu stress were investigated. EBL treatment alone or in combination with Cu enhanced free and bound IAA titers when compared with the metal alone. Modest increases in free and bound ABA contents were observed for EBL treatment alone. However, the combination of EBL with Cu caused major increases in both forms of ABA, over Cu alone. Among the PAs analyzed, only putrescine and cadaverine concentrations were enhanced by EBL treatment alone. By contrast, a significant decline in putrescine and spermine contents was found in seedlings treated with EBL plus Cu. EBL treatments alone or in combination with Cu enhanced activities of guaiacol peroxidase (EC1.11.1.7), catalase (EC 1.11.1.6), superoxide dismutase (EC 1.15.1.1) and glutathione reductase (EC 1.6.4.2) and protein contents in comparison with metal and control treatments. A major decrease in malondialdehyde content was also recorded for EBL treatments with or without Cu. An increase in phytochelatin content was also observed in seedlings treated with EBL alone or in combination with Cu. Major improvement in radical scavenging activities, as attested by the antioxidant activity assay using DPPH (1,1-diphenylpicrylhydrazyl), and elevated deoxyribose and reducing powers, along with increased contents of ascorbic acid, total phenols and proline, also suggest a major influence of EBL application in mitigating copper-induced oxidative stress in radish seedlings. [ABSTRACT FROM AUTHOR]

Abstract: The effects of a high concentration of zinc on two registered clones of poplar (Populus alba Villafranca and Populus nigra Jean Pourtet), inoculated or not with two arbuscular mycorrhizal fungi (Glomus mosseae or Glomus intraradices) before transplanting them into polluted soil, were investigated, with special regard to the extent of root colonization by the fungi, plant growth, metal accumulation in the different plant organs, and leaf polyamine concentration. Zinc accumulation was lower in Jean Pourtet than in Villafranca poplars, and it was mainly translocated to the leaves; the metal inhibited mycorrhizal colonization, compromised plant growth, and, in Villafranca, altered the putrescine profile in the leaves. Most of these effects were reversed or reduced in plants pre-inoculated with G. mosseae. Results indicate that poplars are suitable for phytoremediation purposes, confirming that mycorrhizal fungi can be useful for phytoremediation, and underscore the importance of appropriate combinations of plant genotypes and fungal symbionts. [Copyright &y& Elsevier]

An in vitro model system of micropropagated shoots of the commercial clone ‘Villafranca’ of Populus alba (L.) was used to investigate polyamine (PA) biosynthesis and accumulation in response to high concentrations of zinc (Zn) and copper (Cu). Based on leaf symptoms, rate of adventitious root formation and ethylene production, 0.5–1 mM Zn was transiently toxic while 2–4 mM Zn concentrations were increasingly toxic. Free and conjugated putrescine and spermidine accumulated in a dose–response manner and proportionally to toxicity. The expression profiles of the PA biosynthetic genes were analysed by reverse transcriptase polymerase chain reaction in plants exposed to Zn. In both leaves and stems, PaADC and PaODC transcript levels were early enhanced by all Zn concentrations, while those of PaSAMDC were not. In adventitious roots, free and conjugated PA levels also rose, although their composition and reciprocal ratios were different from those of leaves and no changes in transcript levels of PA biosynthetic genes were detected. Several Cu concentrations (5–500 μ M) were tested in shoots showing severe leaf toxicity, strongly impaired adventitious root formation and high ethylene production. No changes in PA levels were detected until end of culture. The different timing of the PA response to Zn and Cu, despite their rapid uptake and translocation already at 24 h, is discussed in relation to the extent of leaf toxicity. [ABSTRACT FROM AUTHOR]

• The time course of ethylene biosynthesis and perception was investigated in ripening peach fruit ( Prunus persica) following treatments with the polyamines putrescine (Pu) and spermidine (Sd), and with aminoethoxyvinylglycine (AVG). • Fruit treatments were performed in planta. Ethylene production was measured by gas chromatography, and polyamine content by high-performance liquid chromatography; expression analyses were performed by Northern blot or real-time polymerase chain reaction. • Differential increases in the endogenous polyamine pool in the epicarp and mesocarp were induced by treatments; in both cases, ethylene production, fruit softening and abscission were greatly inhibited. The rise in 1-aminocyclopropane-1-carboxylate oxidase ( PpACO1) mRNA was counteracted and delayed in polyamine-treated fruit, whereas transcript abundance of ethylene receptors PpETR1 (ethylene receptor 1) and PpERS1 (ethylene sensor 1) was enhanced at harvest. Transcript abundance of arginine decarboxylase (ADC) and S-adenosylmethionine decarboxylase (SAMDC) was transiently reduced in both the epicarp and mesocarp. AVG, here taken as a positive control, exerted highly comparable effects to those of Pu and Sd. • Thus, in peach fruit, increasing the endogenous polyamine pool in the epicarp or in the mesocarp strongly interfered, both at a biochemical and at a biomolecular level, with the temporal evolution of the ripening syndrome. [ABSTRACT FROM AUTHOR]

In several in vitro systems, treatment with methyl jasmonate (MJ) stimulates polyamine biosynthesis as well as accumulation of acid-soluble and -insoluble conjugated polyamines. This effect is attributed to changes in gene expression and enzyme activity of arginine decarboxylase (ADC), ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase (SAMDC). In the present study, we used tobacco (Nicotiana tabacum L. cv. Samsun) leaf discs to investigate the interaction between this MJ-induced response and those triggered by indole-3-acetic acid (IAA), N6-benzyladenine (BA) and ethylene. Our results indicate that MJ-induced accumulation of conjugated polyamines is further stimulated by auxin and counteracted by BA. The MJ-induced stimulation of ODC and SAMDC, but not ADC, mRNA levels was diminished by IAA±BA, whereas the corresponding enzyme activities were further enhanced in the presence of hormones. MJ enhanced ethylene production only when combined with hormones, and this trend was reflected in 1-aminocyclopropane-1-carboxylate oxidase transcript levels. Results suggest that the induction by MJ of conjugated polyamine levels, and of the polyamine biosynthetic genes and activities are differentially modulated by hormones, and that ethylene does not seem to be directly involved in this response. [Copyright &y& Elsevier]

Production of acid-soluble conjugated di- and polyamines, like that of other secondary metabolites, is enhanced by exposure to methyl jasmonate (MJ). We investigated this metabolic response, and activities of enzymes involved in putrescine (Put) and tropane alkaloid biosynthesis, in root cultures of Hyoscyamus muticus and compared it with that of callus cultures. In root cultures, free Put and N-methylputrescine (mPut) increased upon treatment with MJ, whereas in callus cultures mPut levels were not affected. Differently from roots, conjugated amines were scarce or absent in callus cultures, and accumulated only transiently upon treatment with MJ. Arginine decarboxylase, ornithine decarboxylase and diamine oxidase activities in root cultures were strongly stimulated by treatment with MJ, but were inhibited in callus cultures. Exposure to MJ also enhanced putrescine N-methyltransferase activity in root cultures more than in callus cultures. These results are discussed in relation to the different capacity for tropane alkaloid production in the two culture systems. [Copyright &y& Elsevier]

The effect of various concentrations of aminoethoxyvinylglycine (AVG; 0.32 and 1.28 mM), an ethylene biosynthesis inhibitor, and of the polyamines putrescine (10 mM), spermidine (0.1, 1 and 5 mM) and spermine (2 mM) on peach (Prunus persica L. Batsch cv. Redhaven) fruit ripening was evaluated under field conditions. Treatments were performed 19 (polyamines) and 8 (AVG) days before harvest. Fruit growth (diameter, fresh and dry weight), flesh firmness, soluble solids content and ethylene emission were determined on treated and untreated (controls) fruits. Moreover, endogenous polyamine content and S-adenosylmethionine decarboxylase (SAMDC, EC 4.1.1.21) activity were determined to check for a possible competition between polyamines and ethylene for their common precursor S-adenosylmethionine (SAM). Both treatments strongly inhibited ethylene emission and delayed flesh softening. On a biochemical level, AVG and exogenous polyamines both reduced the free-to-conjugate ratio of endogenous polyamines, and transiently altered SAMDC activity. The possible use of these compounds to control fruit ripening is discussed also in the light of their rejuvenating effect on peach fruits. [ABSTRACT FROM AUTHOR]

The nutraceutical interest in quinoa (Chenopodium quinoa Willd.) seeds is associated with the presence of macronutrients, micronutrients, minerals, vitamins, and polyphenols. In particular, polyphenols contribute to the health-promoting effects of this food crop, and their levels are influenced by environmental conditions. Production of quinoa is recently being explored in temperate climate areas, including Italy. The aim of this research was to assess the profile of bioactive compounds in seeds of two quinoa varieties, Regalona-Baer and Titicaca, grown in northern Italy, compared to that of seeds of those varieties grown in Chile and Denmark, respectively. High-performance liquid chromatography-diode array detector (HPLC-DAD) analysis of phenolic acid and flavonoid profiles, both in their free and soluble conjugated forms, showed that the main differences between Regalona grown in Chile and Italy were for the free vanillic acid and daidzein contents, while the two Titicaca samples mainly differed in quercetin derivative levels. The total phenolic index was comparable in Titicaca and Regalona, and only a slight decrease in this parameter was found in seeds of the two varieties grown in Italy. The in vitro antioxidant activity of seed extracts, evaluated by means of three different assays, indicated that it correlated with flavonol (quercetin derivative) levels. In conclusion, the results indicate that, although environmental conditions alter the polyphenolic profile and biological activities, it is possible to grow good-quality quinoa in northern Italy. [ABSTRACT FROM AUTHOR]

In indoor vertical farms, energy consumption represents a bottleneck for both a system's affordability and environmental footprint. Although switching frequency (sf) represents a crucial factor in determining the efficacy of light emitting diodes (LED) lighting systems in converting electricity into light, the impact of sf is still underexplored. The aim of this work was to investigate the effect of LEDs sf on the productive and qualitative responses of lettuce (Lactuca sativa L.), also considering the resource use efficiency. Plants were grown for 14 days under red and blue LEDs (215 μmol m−2 s−1 and 16/8 h light/dark, with a red:blue ratio of 3) characterized by two different sf for the blue diode, namely high sf (850 kHz) and low sf (293 kHz). A fluorescent light (same light intensity and photoperiod) was included. LED sf did not alter plant morphological parameters, including fresh or dry biomass, leaf number, leaf area, or water use efficiency. A low sf increased the energy use efficiency (EUE) by 40% as compared to high sf. The latter enhanced the leaf antioxidant capacity, as a consequence of increased concentrations of caftaric and chicoric acids, isoquercetin, and luteolin, consistent with the upregulation of a few genes related to the biosynthetic pathway of phenolic compounds (4C3H and DFR). The study highlights that different sf may significantly affect the EUE as well as crop nutritional properties. [ABSTRACT FROM AUTHOR]

Previous results indicated that in shoot-forming tobacco thin layers the putative inhibitor of S-adenosylmethionine decarboxylase (SAMDC, EC 4.1.1.21) activity methylglyoxal(bisguanylhydrazone) (MGBG) inhibited meristemoid/primordia formation while enhancing conjugated polyamine accumulation and ethylene biosynthesis, thus showing an inverse relationship between the two phenomena (Scaramagli et al. 1999). In order to better understand how MGBG causes polyamine overproduction and by using the same model system, we further examined the effect of 0.5 mM MGBG on the incorporation of labelled methionine into spermidine and spermine, on SAMDC transcript levels and activity and on putrescine oxidising (DAO) activity at different times in culture. MGBG-induced ethylene accumulation was also evaluated by gas chromatography (GC). In shoot-forming controls (grown on a medium supplemented with IAA+BA) label was mainly incorporated into free spermidine (82–86% of the total amount incorporated into polyamines) while only 6–12% of radioactivity was recovered in free spermine. Some label was also found in putrescine. In the presence of MGBG, spermidine biosynthetic capacity was enhanced throughout the culture by 28–45% compared with controls. The same occurred for spermine and putrescine. MGBG-enhanced spermidine synthesis was supported by increased SAMDC activity (days 1–2) and higher levels of SAMDC message (days 1–7). On the contrary, the effect of the drug on putrescine oxidation was inhibitory starting from day 2. These results, together with the previous ones, suggest that MGBG, in fact, does not behave as a SAMDC inhibitor in shoot-forming tobacco thin layers and that its negative effect on organised growth is not due to SAMDC inhibition/polyamine depletion. [ABSTRACT FROM AUTHOR]

The optimal assay conditions and the trend with time in culture (28 days) of arginine decarboxylase (ADC; EC 4.1.1.19), ornithine decarboxylase (ODC; EC 4.1.1.17) and diamine oxidas (DAO; EC 1.4.3.6) activities in habituated (H) and normal (N) auxin- and cytokinin-requiring sugar beet callus were compared. Although the response to variations in buffer pH and EDTA and pyridoxal phosphate (PLP) concentrations varied for ADC and ODC activities between the two callus types, pH 8.3, 50 μM PLP and 5 mM EDTA were generally optimal or near-optimal for both H and N callus. In most cased the addition of ornithine or arginine in the ADC and ODC assays, respectively, given to block the interconversion between the two substrates, resulted in lower 14CO2recovery, DAO activity was very differently affected in H and N callus by the presence of polyvinylpyrrolidone in the extraction buffer. However, in both cases, this activity increased with time in culture. ADC activity was always predominant in both cell lines and always higher in N callus. In the latter, ADC activity rose sharply between days 14 and 21 and then leveled off while in H callus it increased steadily from day 14 onwards. ODC activity was also higher in N callus and peaked sharply on day 21 while in H callus it was not detectable in the second half of the culture period. In both cell lines this activity was low or nil on day 28, 3,4[14]-methionine incorporation into ethylene and polyamines was also compared in N and H callus. In the latter, ethylene synthesis was lower and [14C]-spermidine formation higher than in N callus. This is in accord with the significantly higher spermidine titres found in H callus. [ABSTRACT FROM AUTHOR]

An attempt was made to identify some of the hormonal factors that control adventitious root formation in our Prunus avium micropropagation system in order to improve rooting in difficult-to-root genotypes. Changes in endogenous contents of free polyamines were determined at intervals during auxin-induced rooting of shoot cultures. Accumulation of putrescine and spermidine peaked between days 9 and 11. Spermine was only present in traces. Exogenously supplied putrescine or spermine (50-500 μM), in the presence of optimal or suboptimal levels of indolebutyric acid (IBA), had no effect on rooting percentage or rooting density, except for spermine at 500 μM. At this external concentration spermine caused a substantial accumulation in both free spermine and putrescine. The use of several inhibitors of polyamine biosynthesis, namely α-difluoromethylornithine (DFMO), &alpha-difluoromethylarginine (DFMA), dicyclohexylammonium sulphate (DCHA) and methylglyoxal-bis-guanyl-hydrazone (MGBG) alone or in combination in the 0.1 to 5 mM range, resulted in an inhibition of rooting that was partially reversed by the addition of the corresponding polyamine. Cellular polyamine levels were significantly reduced by DFMO and DFMA but not by DCHA and MGBG. Laveled putrescine incorporation into spermidine increased somewhat in the presence of the ethylene synthesis inhibitor aminoethoxyvinylglycine (AVG). A system based on [3,4-14C]methionine incorporation was used to measure ethylene synthesis by the in vitro cultured shoots. Label incorporation was drastically reduced by 10 μM AVG and increased 3.5-fold in the presence of 50 μM IBA with respect to controls (no IBA). Labeled methionine incorporation into spermidine increased to some extent when ethylene synthesis was inhibited by AVG. Adding the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) to the rooting medium significantly inhibited rooting percentage; AVG caused the formation of a greater number of roots per shoot but delayed their growth. Supplying the shoots with both compounds resulted in an intermediate rooting response, in which both rooting percentage and root density were affected. These results indicate that polyamines may play a significant role at least in some stages of root formation. The polyamine and ethylene biosynthetic pathways seem to be competitive but under our conditions, the enhancement of one pathway when the other was inhibited, was not dramatic. Although IBA promoted ethylene synthesis, AVG, which drastically reduced it, also promoted root formation. Thus, the auxin effect on root induction cannot be directly related to its ability to enhance ethylene synthesis. [ABSTRACT FROM AUTHOR]

The effect of 1 mM dicyclohexylamine (DCHA) on the synthesis of spermidine and spermine was examined in excised cotyledons of radiata pine (Pinus radiata D. Don) cultured under shoot-forming (with cytokinin) and non-shoot-forming (minus cytokinin) conditions by incubation with [14C]-putrescine. In control cotyledons incorporation into spermidine showed a peak at day 2 in the presence and at day 5 in the absence of N6-benzyldenine (BA). DCHA-treated cotyledons gave the same labeling pattern, both in the presence and absence of benzyladenine, with a much smaller peak at day 2. The incorporation into spermidine and spermine was insignificant at day 5 and later. The total radioactivity in the trichloroacetic acid supernatant indicated that precursor uptake was strongly reduced by the drug. In addition, the percentage label found in the benzene phase and combined in the 3 polyamines was lower in DCHA-treated cotyledons. Thus, treatment with DCHA not only inhibited the conversion from putrescine to spermidine and spermine, but also reduced its conversion to other benzene-extractable compounds. S-Adenosylmethionine decarboxylase (SAMDC, EC 4.1.1.50) activity, which furnishes the propylamine group to spermidine and spermine synthases (EC 2.5.1.16 and EC 2.5.1.-), was not significantly affected by DCHA and appeared to be independent of the spermidine and spermine synthase reactions, suggesting that spermine synthesis decreased as a result of substrate depletion. The correlation between morphological development and polyamine biosynthesis is discussed.

Excised cotyledons of radiata pine (Pinus radiata D. Don) were cultured in the presence or absence of benzyladenine and two concentrations of dicyclohexylamine, a potent inhibitor of spermidine synthesis in animals and bacteria. Cellular levels of the drug and of putrescine, spermidine and spermine were determined by dansylation followed by thin layer chromatography. The rate of uptake of the drug was rapid during the first 6 h of incubation and then diminished; nevertheless, its cellular content increased with time in culture and uptake was greater if more was present in the medium. Cotyledons cultured on benzyiadenine-containing media accumulated more dicyclohexylamine than on benzyladenine-free media and this resulted in toxic side-effects. The drug had no significant effect on the elongation growth occurring, in the absence of the hormone. In cotyledons treated with the drug, spermidine levels fell to zero after a 5- to 10-day exposure. Putrescine increased transiently at 24 h and then declined significantly. Spermine levels also declined to 11% of control values, by day 5 and remained low throughout. Except for a marked decrease in all three polyamines during the first 24 h of culture, all control cotyledons maintained significantly higher polyamine levels than dicyclohexylamine-treated ones. [ABSTRACT FROM AUTHOR]