Indole-3-butyric acid and myo-inositol impacts on in vitro rooting of the cherry rootstocks CAB-6P and Gisela 6

The present study investigates the effects of indole-3-butyric acid (IBA) alone and in combination with myo-inositol on in vitro rooting and biochemical responses in the cherry rootstocks CAB-6P (Prunus cerasus L.) and Gisela 6 (Prunus cerasus × Prunus canescens). For the CAB-6P rootstock, the best results for root number (6.31), fresh mass (FM), dry mass (DM), and rooting percentage (100 %) were obtained on Murashige and Skoog (MS) medium with 2 mg dm^?3 IBA and maximum root length (30.57 mm) was obtained at 1 mg dm^?3 IBA. Myo-inositol suppressed the positive effects of IBA on root length. In the Gisela 6 explants, the inclusion of 2 mg dm^?3 IBA together with 0.5 mg dm^?3 of myo-inositol in the culture medium significantly enhanced root number (9.91) and root FM and DM. The root length was maximum in the combination of the lowest IBA and myo-inositol concentrations (0.5 mg dm^?3). The rooting percentage was the greatest (100 %) with the application of 1 mg dm^?3 IBA alone. In both explants, the application of IBA alone or in combination with myo-inositol resulted in a lower leaf proline content in comparison with the control (without growth regulators). The maximum leaf chlorophyll content was at 1 mg dm^?3 IBA in the CAB-6P whereas at 2 mg dm^?3 IBA and 1 mg dm^?3 myo-inositol in Gisela 6. Addition of myo-inositol mostly increased sugar content in comparison with control or IBA alone in both rootstocks.     

Effect of amino acids on rooting of apple dwarf rootstocksin vitro

The effect of twelve amino acids and lactalbumin hydrolysate in concentration of 200 mg 1^?1 on rooting of the dwarf apple rootstocks P 2 and P 60 was testedin vitro. Arginine, omithine, glutamic acid and glycine enhanced root number of the P 60 rootstock; proline and lactalbumin hydrolysate were neutral; and asparagine, tyrosine, methionine, cysteine and glutamine lowered the root number. Tyrosine, methionine, cysteine and glutamine reduced almost completely rooting of P 60. In the recalcitrant P 2 rootstock aspartic acid, glutamic acid and omithine significantly enhanced the number of roots and rooted shoots, arginine and tryptophan increased the root number only slightly, asparagine was neutral, and proline reduced the root number.     

Role of Amino Acids in Evolution of Ethylene and Methane, and Development of Microshoots in Cajanus Cajan

In pigeonpea (Cajanus cajan L.) shoot buds were induced when cotyledonary node explants were supplemented with benzylaminopurine (BAP; 2 mg dm^–3). When 0.1 mg dm^-3 BAP and 0.01 mg dm^-3 naphtahalene acetic acid were supplemented to the medium, the 34 – 35 % of induced buds developed to microshoots. By supplementing amino acids like proline, glutamine, asparagine and L-cysteine, shoot bud development to microshoots was enhanced at least by two fold. Amongst the amino acids proline gave maximum number of microshoots per explant. With increase in concentration of amino acids, fresh mass increased but microshoot number decreased. Also methane evolution was increased by addition of amino acids, and also in medium containing more of its nitrogen in the form of ammonia. Increased evolution of methane was accompanied by reduction in evolution of ethylene, and enhancement of efficiency of microshoot development.     

Proline Accumulation, Protein Pattern and Photosynthesis in Bacopa Monniera Regenerants Grown under NaCl Stress

Shoots of Bacopa monniera exhibited 100 % regeneration on Murashige and Skoog medium with 2 % sucrose, 0.2 mg dm⁻³ 1-naphthaleneacetic acid, 0.5 mg dm⁻³ 6-benzylaminopurine and 50 mg dm⁻³ glutamine. When the medium was supplied with various concentrations (5 - 15 g dm⁻³) of sodium chloride, proline content in regenerants was six times higher than in the control. With increasing NaCl concentration photosynthetic rate decreased and fresh mass and root length of regenerants declined. NaCl also induced formation of new proteins. This revised version was published online in July 2006 with corrections to the Cover Date.     

Metabolism of absorbed aspartate,asparagine, and arginine by rat small intestine in vivo

l-[U-¹⁴C]aspartate, l-[U-¹⁴C]asparagine, and l-[U-¹⁴C]arginine were administered luminally into isolated segments of rat jejunum in situ, and the radioactive products appearing in venous blood from the segment were identified and quantified, in a continuation of similar studies with l-glutamate and l-glutamine (Windmueller H.G. and Spaeth, A. E. (1975) Arch. Biochem. Biophys. 171, 662–672). Aspartate, administered alone (6 mm) or with 18 other amino acids plus glucose, was absorbed more rapidly than glutamate, but, as with glutamate, less than 1% was recovered intact in intestinal venous blood. More than 50% of aspartate carbon was recovered in CO₂, 24% in organic acids, mostly lactate, 12% in other amino acids (alanine, glutamate, proline, ornithine, and citrulline), and 10% in glucose, apparently the first demonstration of gluconeogenesis by intestine in vivo. In contrast to aspartate and glutamine, nearly all asparagine was absorbed intact, less than 1% being catabolized. About 4% of the absorbed dose was incorporated into the acid-insoluble fraction of intestine, as was the case with all the amino acids studied. In conventional or germ-free rats, only 60% of arginine was absorbed intact, while 33% was hydrolyzed to ornithine and urea. The urea and 38% of the ornithine were released into the blood; the remaining ornithine was metabolized further by intestine to citrulline, proline, glutamate, organic acids, and CO₂. Catabolism of several amino acids from the lumen plus glutamine from arterial blood may provide an important energy source in small intestine.     

Effect of natural amino acids on ethanol oxidation in isolated rat hepatocytes]

The effects of the various naturally occurring amino acids on ethanol oxidation in hepatocytes from starved rats was systematically studied. In order to minimize the non ADH pathways, the ethanol concentration used was 4 mmol/litre, the amino acids being added at the same concentration. In hepatocytes from fasted rats, alanine, arginine, asparagine, aspartate, citrulline, cysteine, glutamate, glutamine, glycine, histidine, hydroxyproline, ornithine and serine increase significantly ethanol consumption. The stimulatory effect of glutamine being much less pronounced than the asparagine one and proline being devoid of action, the influence of ammonium chloride addition on ethanol consumption in the presence of these amino acids was studied. Ammonium chloride determines an enhancement of ethanol oxidation in these conditions, the results showing no apparent correlation between intracellular glutamate concentration and ethanol oxidation rate, contrarily to previous data. In hepatocytes from fed rats, only alanine, asparagine, cysteine, glycine, hydroxyproline, ornithine and serine increase ethanol oxidation, although to a lesser extent than in cells from starved rats.     

Synthesis,binding ability,and cell cytotoxicity of fluorescent probes for l‐arginine detection based on naphthalene derivatives: Experiment and theory

Inspired by biological related parts, Schiff base derivatives and functional groups of chemical modification can provide efficient detection method of amino acids. Therefore, we have designed and prepared 4 compounds based on Schiff base derivatives involving ─NO₂, ─OH, and naphthyl group. Results indicated that compound 4 containing 2 nitro groups showed strong sensitivity and high selectivity for arginine (Arg) among normal 18 kinds of standard amino acids (alanine, valine, leucine, isoleucine, methionine, aspartic acid, glutamic acid, arginine, glycine, serine, asparagine, phenylalanine, histidine, tryptophan, proline, lysine, glutamine, and cysteine). Theoretical investigation also approved the strong binding ability of compound 4 for Arg. In addition, compound 4 displayed high combining ability of Arg and low cytotoxicity of MCF‐7 cell in the 0 to 150 μg mL^?1 of concentration range; it can be used for Arg in vivo detection of fluorescent probe.     

Regulation of RNA degradation in cultured rat hepatocytes: Effects of specific amino acids and insulin

The regulation of RNA degradation by specific amino acids and insulin was investigated in cultured rat hepatocytes from fed rats previously injected in vivo with [6-¹⁴C]orotic acid. The effects of three groups of amino acids were compared to those of a complete amino acid mixture. The first one consisted of the eight amino acids (leucine, proline, glutamine, histidine, phenylalanine, tyrosine, methionine, tryptophan) previously found to be particularly effective in the control of proteolysis. The two other groups were defined from our study with single additions of amino acids, one consisting of proline, asparagine, glutamine, alanine, phenylalanine, and leucine and the other including the latter group with serine, histidine, and tyrosine. The results showed that these three groups were able to strongly inhibit deprivation-induced RNA breakdown at one and ten times normal plasma concentrations but to a lower extent than the complete amino acid mixture. Six amino acids (proline, asparagine, glutamine, alanine, phenylalanine, leucine) inhibited individually RNA degradation by more than 20%. However, the deletions of proline, asparagine, glutamine, or alanine from the group of these six amino acids were not followed by a loss of inhibitory effect. On the contrary, an important loss of inhibition was observed when leucine and phenylalanine were deleted. Furthermore, only these two amino acids exhibited an additive inhibitory effect. Thus leucine and phenylalanine could be considered as important inhibitors of RNA breakdown in cultured rat hepatocytes. Finally, insulin which had no significant effect on RNA degradation in the absence of amino acids, was able to potentiate the inhibitory effect of different amino acid groups. © 1993 Wiley-Liss, Inc.     

Selenium treatment alters phytochemical and biochemical activity of in vitro-grown tissues and organs of Allium sativum L.

Garlic, an important flavoring agent and a medicinally useful plant, can take up selenium from its immediate surrounding medium and incorporate it at high concentrations into amino acids and phytochemicals. Selenium, supplied as 0.5, 1.0, 2.0, and 4.0?mg?L^?1 Na₂SeO₃ increased the amino acid, protein, proline, and alliin content of in vitro-grown callus, embryo, plantlet, leaf, and root tissues of Allium sativum L. The enhancement was significant at 2 and 4?mg?L^?1. Superoxide dismutase, catalase, and glutathione reductase activities increased in all in vitro-grown tissues and organs with increasing selenium concentrations, but enzyme activity was highest with 4?mg?L^?1 selenium.     

Intestinal metabolism of glutamine and glutamate from the lumen as compared to glutamine from blood.

Only a small fraction of the l-[U-¹⁴C]glutamate (2%) and the l-[U-¹⁴C]glutamine (34%) administered at a 6 mm concentration into the lumen of rat jejunal segments in situ was recovered unchanged in venous blood collected from the segments. The remaining ¹⁴C of both amino acids was recovered in the blood as CO₂ (60%), proline (5%), citrulline (4%), alanine (3%), ornithine (2%), and organic acids, mostly lactate (19%). The amide nitrogen of glutamine was recovered mostly as ammonia and the amino nitrogen of both amino acids predominantly in alanine. A nearly identical distribution of products was seen in previously published experiments in which rat intestine took up l-[U-¹⁴C]glutamine from arterial blood (Windmueller, H. G., and Spaeth, A. E. (1974) J. Biol. Chem., 249, 5070–5079). The results are therefore consistent with a single metabolic pool within mucosal cells for blood-derived and lumen-derived glutamine. When 6 mm glutamine was continuously perfused through the lumen, jejunal segments metabolized arterial and luminal glutamine at approximately equal rates (130–190 nmol min^?1 (g of tissue)^?1). The total combined rate was 1.7 times the rate of utilization of arterial glutamine alone in jejunal segments not absorbing glutamine. These results provide the first quantitative data on comparative metabolism by the intestine of substrates from the lumen and from blood. Rat intestine apparently metabolizes nearly all absorbed dietary glutamate and most glutamine in addition to circulating glutamine derived from other tissues.     

Somatic organogenesis and plant regeneration in <Emphasis Type="Italic">Ricinus communis</Emphasis>

An in vitro propagation system was developed for castor-bean (Ricinus communis L. cv. TMV 6) through cotyledon derived callus cultures. The impact of different concentrations of auxins, cytokinins, additives, amino acids and sugars were evaluated for callus induction and shoot proliferation. Green compact nodular organogenic callus was obtained on the medium fortified with Murashige and Skoog (MS) salts, B5 vitamins, 2.0 mg dm⁻³ 6-benzyladenine and 0.8 mg dm⁻³ α-naphthalene acetic acid (NAA). Multiple shoot proliferation from the callus cultures was achieved on the medium with MS salts, B5 vitamins, 2.5 mg dm⁻³ thidiazuron (TDZ), 0.4 mg dm⁻³ NAA and 15 mg dm⁻³ glutamine. During multiple shoot induction the phenolic secretion was controlled by the addition of 15 mg dm⁻³ polyvinylpyrolidone. The proliferated shoots were elongated on the medium comprising MS salts, B5 vitamins, 1.5 mg dm⁻³ TDZ and 0.3 mg dm⁻³ gibberellic acid. The elongated shoots were rooted on the medium containing MS salts, B5 vitamins, 0.3 mg dm⁻³ indole-3-butyric acid and 0.6 mg dm⁻³ silver nitrate. After root induction, the plants were hardened in earthen pots containing sand, soil and vermiculite.     

Glutamine limited fed-batch culture reduces the overflow metabolism of amino acids in myeloma cells

The murine myeloma cell line Sp 2/0-Ag 14 was cultured in an ordinary batch culture and in a glutamine limited fed-batch culture. In batch culture, the overflow metabolism of glutamine ends in excess production of ammonium and the amino acids alanine, proline, ornithine, asparagine, glutamate, serine and glycine. This pattern was dramatically changed in the fed-batch culture. Glutamine limitation halved the cellular ammonium production and reduced the ratio of NH₄ ⁺/glutamine. The excess production of alanine, proline and ornithine was reduced by a factor of 2–6 while asparagine was not produced at all. In contrary to the other amino acids glycine production was increased. These results are discussed in view of the different nature of glutamine metabolism in the mitochondrial compartment vs. the cytosolic. Furthermore, essential amino acids were used more efficiently in the fed-batch as judged by the increase in the cellular yield coefficients in the range of 1.3–2.6 times for seven of the 11 consumed ones. In all, this leads to a more efficient use of the energy sources glucose and glutamine as revealed by an increase in the cellular yield coefficient for glucose by 70% and for glutamine by 61%.     

Origin of Foliar Nitrogen and Changes in Free Amino-Acid Composition and Content of Leaves, Stubble, and Roots of Perennial Ryegrass During Re-growth after Defoliation

Nitrogen re-mobilization and changes in free amino acids werestudied as a function of time in leaves, stubble, and rootsduring ryegrass (Lolium perenne L.) re-growth. Experiments with¹⁵N labelling clearly showed that during the first days nearlyall the nitrogen in new leaves came from organic nitrogen re-mobilizedfrom roots and stubble. On the days of defoliation, stubblehad the highest content of free amino acids with 23 mg per gdry weight against 15 mg and 14 mg in leaves and roots, respectively.The major amino acids in leaves were asparagine (23% of totalcontent in free amino acids), aminobutyrate, serine, glutamine,and glutamate (between 7% and 15%) whereas in roots and stubblethe contribution of amides was high, especially asparagine (about50%). Re-growth after cutting was associated with a rapid increaseof the free amino acid content in leaves, with a progressivedecrease in roots while stubble content remained virtually unchanged.In leaves, asparagine increased from the first day of re-growth,while the aspartate level remained unchanged and glutamine increasedstrongly on the first day but decreased steadily during thenext few days of re-growth. Asparagine in stubble and rootschanged in opposite directions: in stubble it tended to increasewhereas in roots it clearly decreased. In contrast, stubbleand roots showed a similar decrease in glutamine. In these twoplant parts, as in leaves, aspartate remained at a low level.Results concerning free amino acids are discussed with referenceto nitrogen re-mobilization from source organs (stubble androots) to the sink organ (regrowing leaves). Key words: Lolium perenne L, re-growth, nitrogen, free amino acids, glutamine, asparagine     

Triacontanol-Induced Changes in the Growth, Photosynthetic Pigments, Cell Metabolites, Flowering and Yield of Green Gram

Seedlings of green gram [Vigna radiata (L.) Wilezek] cultivar KM-2 were sprayed with different concentrations of triacontanol (TRIA) (0, 0.5, 1.0, and 2.0 mg dm^-3) at 15 and 25 days after sowing. Foliar spray of 0.5 mg dm^-3 TRIA significantly promoted the plant height, fresh mass, and contents of chlorophylls, saccharides, starch, soluble proteins, amino acid and phenols. Leaf nitrate content was reduced by 0.5 and 1.0 mg dm^-3 TRIA with a corresponding increase in nitrate reductase activity. TRIA of 0.5 mg dm^-3 stimulated the onset of flowering, pod production and retention, but less number of pods and seeds per plant were observed at 2.0 mg dm^-3 treatment.     

Exogenous 24-epibrassinolide ameliorates high temperature-induced inhibition of growth and photosynthesis in Cucumis melo

This study was carried out to better understand the role of 24-epibrassinolide (EBR) in thermotolerance of melon (Cucumis melo L.). The melon seedlings were pretreated with various concentrations of EBR (0, 0.05, 0.1, 0.5, 1.0, and 1.5 mg dm^?3) as foliar spray and then exposed to a high temperature (HT) stress. Exogenous EBR (0.5–1.5 mg dm^?3) alleviated HT-caused growth suppression. In parallel, 1.0 mg dm^?3 EBR attenuated the decrease in chlorophyll content, net photosynthetic rate, stomatal conductance, maximum quantum efficiency of photosystem (PS) II, quantum yield of PS II, and photochemical quenching of chlorophyll a fluorescence in HT-stressed plants, and inhibited transpiration rate and non-photochemical quenching. Furthermore, exogenous EBR also significantly reduced the content of malondialdehyde (MDA) and increased the content of soluble proteins and free proline, and activities of antioxidant enzymes including superoxide dismutase, guaiacol peroxidase, catalase, and ascorbate peroxidase under the HT stress. The results show that protective effects of EBR against the HT stress in the melon seedlings were most likely mediated through the improvement of photosynthesis and the stimulation of antioxidant capacity.     

Synthesis of metabolism-resistant substrates for the transport system for cationic amino acids; their stimulation of the release of insulin and glucagon, and of the urinary loss of amino acids related to cystinuria

Several amino acids have been synthesized as model transport substrates building on the piperidine and cyclohexane rings. Only when the distal N atom is part of an unambiguously cationic structure are these compounds transported predominantly by the cationic amino acid system. These amino acids in labeled form are excreted rather slowly in unmodified state, very little ¹⁴CO₂ being released. Those which are unambiguously cationic (including also homoarginine) led to a greatly increased excretion of arginine, lysine, ornithine and citrulline. Those which might be expected to act as lysine analogs had little effect on the excretion of the basic amino acids, although the excretion of citrulline and the sum of glutamine plus asparagine was accelerated. Certain of the analogs intensified the excretion of citrulline in dissociation from effects on resorption of the basic amino acids, also in dissociation from effects on cystine resorption. These results indicate citrulline resorption does not occur principally by the same agency serving for the basic amino acids, nor by the agency serving for cystine, despite the observed interactions for resorption. The injection of either of three transport analogs for arginine into the rat leads to early increases in the circulating levels of immunologically reactive insulin and glucagon.     

Xylem to phloem transfer of solutes in fruiting shoots of legumes,studied by a phloem bleeding technique

Summary Comparisons were made of the levels of various solutes in xylem (tracheal) sap and fruit tip phloem sap of Lupinus albus (L.) and Spartium junceum (L.). Sucrose was present at high concentration (up to 220 mg ml^-1) in phloem but was absent from xylem whereas nitrate was detected in xylem (up to 0.14 mg ml^-1) but not in phloem. Total amino acids reached 0.5–2.5 mg ml^-1 (in xylem) versus 16–40 mg ml^-1 in phloem. Phloem: xylem concentration ratios for mineral nutrients (K, Na, Mg, Ca, Fe, Zn, Mn, Cu) spanned the range 0.7 to 20, the ratios generally reflecting an element's phloem mobility and its availability to the xylem from the roots.The accessibility of nitrate to xylem and phloem was studied in Lupinus. Increasing the nitrate supply to roots from 100 to 1000 mg NO₃–Nl^-1 increased nitrate spill over into xylem, but nitrate always failed to appear in phloem. However, phloem loading of small amounts of nitrate was induced by feeding 750 or 1000 mg NO₃–Nl^-1 directly to cut shoots via the transpiration stream. Transfer of reduced nitrogen to phloem was demonstrated by feeding ¹⁵NO₃ to shoots and recovering ¹⁵N-enriched amides and amino acids in phloem sap. Increased nitrate supply to roots led to increased amino acid levels in xylem and phloem but did not alter markedly the balance between individual amino acids.The fate of xylem-fed ¹⁴C-labelled asparagine, glutamine and aspartic acid and of photosynthetically fed ¹⁴CO₂ was studied in Spartium, with reference to phloem transport to seeds. Substantial fractions of the ¹⁴C of all sources appeared in non-amino compounds. [¹⁴C]asparagine passed largely in unchanged form to the phloem whereas the ¹⁴C from aspartic acid or glutamine appeared in phloem attached to other amino acids (e.g. asparagine and glutamic acid). Serine, asparagine and glutamine were the main amino compounds labelled in phloem sap after feeding ¹⁴CO₂. The wide distribution of ¹⁴C amongst free and bound amino acids of seeds suggested that extensive metabolism of phloem-borne solutes occurred in the fruits.     

Studies on the Nitrogen Metabolism in Ectomycorrhizae

Free and bound amino acids in the mycorrhizal and nonmycorrhizal root systems of Pinus nigra Arnold and Corylus avellna L. grown under semi-natural conditions were analyzed through automatic amino acid analyzers. Tuber brumale Vitt. and T. melanosporum Vitt. were the respective fungal symbionts. Arginine and citrulline were found to accumulate in large quantities in the free pool of the uninoculated P. nigra and C. avellana root systems respectively. In the mycorrhizal root systems these substances decreased in their levels with a parallel increase in the concentrations of glutamine and asparagine. Implications of these changes are discussed with reference to ectomycorrhizae. In general the majority of the identified free amino acids were found in larger concentrations in both the types of mycorrhizal root systems. Results obtained suggested that this may not be due to proteolysis but due to increased biosynthesis. Possible interrelationships in ectomycorrhizal root systems with reference to nitrogen metabolism are presented in the form of a scheme.     

Root xylem transport of amino acids in the root hemiparasitic shrub Olax phyllanthi (Labill) R.Br. (Olacaceaea) and its multiple hosts

The amino acid compositions of the root xylem saps of Olax phyllanthi and a range of its common hosts were examined in native coastal heath in Western Australia and in pot cultures of Olax reliant on single hosts. When hosts specializing in the xylem transport of one major solute (asparagine, glutamine, histidine, arginine or proline) were exploited, the endophytic tissue of haustoria and the xylem sap of Olax showed much lower proportions of this than of other solutes, suggesting pronounced metabolic transformation prior to xylem loading by the parasite. However, the xylem sap of Olax did partly reflect the compositions of its hosts; for example, djenkolic acid and pipecolic acid were present when Olax was parasitic on species of Acacia, and levels of citrulline and aspartic acid were higher than normal when it exploited hosts transporting large amounts of these compounds. Back-flow of S-ethenyl cysteine, a novel amino acid specific to Olax, was observed to another root hemiparasite (Exocarpos sparteus) in native habitat and to certain non-parasitic hosts in water-stressed pot cultures. Haustoria exhibited high levels of glutamine synthetase but showed appreciable in vivo nitrate reductase activity only when on hosts with high xylem levels of nitrate.     

Amino Acid Pools in Herbaceous Plants at the Wintering Stage and at the Beginning of Growth

Free amino acids in 40 herbaceous perennial plants were analyzedunder natural conditions. From the major amino acid contentat the wintering stage, the pools were separated into the followingfive types: 1) a group which accumulated arginine (20 plantsout of 40); 2) a group which accumulated arginine and proline(9 plants); 3) a group which accumulated glutamate and glutamine(3 plants); 4) a group which accumulated asparagine (4 plants);and 5) a group which accumulated proline (4 plants). Changesin the amino acid pools in the plants occurred under snow duringwintering for about five months. Particularly, asparagine wasno longer the major amino acid in the group which had accumulatedit in fall. There was a tendency for the glutamine content toincrease, suggesting that NH₃ is utilized for the synthesisof the amide. Also, the relative concentrations of almost allthe free amino acids increased several-fold, which was indicativeof the occurrence of biosynthetic processes of general aminoacids during wintering. As the mobile fractions of stored nitrogen,the amino acids appeared to contribute to the initial stageof rapid growth in early spring. (Received August 4, 1986; Accepted November 17, 1986)