Nitrogen Nutrition and Xylem Transport of Nitrogen in Ureide-producing Grain Legumes

Xylem sap composition was examined in nodulated and nonnodulated cowpea (Vigna unguiculata [L.] Walp.) plants receiving a range of levels of NO₃ and in eight other ureide-forming legumes utilizing NO₃ or N₂ as sole source of nitrogen. A ¹⁵N dilution technique determined the proportions of plant nitrogen derived from N₂ in the nodulated cowpeas fed NO₃. Xylem sap composition of NO₃-fed, nodulated cowpea varied predictably with the relative extents to which N₂ and NO₃ were being utilized. The ratios of asparagine to glutamine (N/N) and of NO₃ to ureide (N/N) in xylem sap increased with increasing dependence on NO₃ whereas per cent of xylem nitrogen as ureide and the ratio of ureide plus glutamine to asparagine plus NO₃ (N/N) in xylem sap increased with increasing dependence on N₂ fixation. The amounts of NO₃ and ureides stored in leaflets, stems plus petioles, and roots of cowpea varied in a complex manner with level of NO₃ and the presence or absence of N₂ fixation. All species showed higher proportions of organic nitrogen as ureide and several-fold lower ratios of asparagine to glutamine in their xylem sap when relying on N₂ than when utilizing NO₃. In nodulated (minus nitrate) cowpea and mung bean (Vigna radiata [L.] Wilczek) the percentage of xylem nitrogen as ureide remained constant during growth but the ratio of asparagine to glutamine varied considerably. The biochemical significance of the above differences in xylem sap composition was discussed.     

Transport of nitrogen in the xylem of soybean plants

Experiments were conducted to characterize the distribution of N compounds in the xylem sap of nodulated and nonnodulated soybean plants through development and to determine the effects of exogenous N on the distribution of N compounds in the xylem. Xylem sap was collected from nodulated and nonnodulated greenhouse-grown soybean plants (Glycine max [L.] Merr. “Ransom”) from the vegetative phase to the pod-filling phase. The sum of the nitrogen in the amino acid, nitrate, ureide (allantoic acid and allantoin), and ammonium fractions of the sap from both types of plants agreed closely with total N as assayed by a Kjeldahl technique. Sap from nodulated plants supplied with N-free nutrient solution contained seasonal averages of 78 and 20% of the total N as ureide-N and amino acid-N, respectively. Sap from nonnodulated plants supplied with a 20 millimolar KNO₃ nutrient solution contained seasonal averages of 6, 36, and 58% of total N as ureide-N, amino acid-N, and nitrate-N, respectively. Allantoic acid was the predominant ureide in the xylem sap and asparagine was the predominant amino acid. When well nodulated plants were supplied with 20 millimolar KNO₃, beginning at 65 days, C₂H₂ reduction (N₂ fixation) decreased relative to nontreated plants and there was a concomitant decrease in the ureide content of the sap. A positive correlation (r = 0.89) was found between the ureide levels in xylem sap and nodule dry weights when either exogenous nitrate-N or urea-N was supplied at 10 and 20 millimolar concentrations to inoculated plants. The results demonstrate that ureides play a dominant role in N transport in nodulated soybeans and that the synthesis of ureides is largely dependent upon nodulation and N₂ fixation.     

Modeling C and N transport to developing soybean fruits

Xylem sap and phloem exudates from detached leaves and fruit tips were collected and analyzed during early pod-fill in nodulated soybeans (Glycine max (L.) Merr. cv Wilkin) grown without (−N) and with (+N) NH₄NO₃. Ureides were the predominant from (91%) of N transported in the xylem of −N plants, while amides (45%) and nitrate (23%) accounted for most of the N in the xylem of +N plants. Amino acids (44%) and ureides (36%) were the major N forms exported in phloem from leaves in −N plants, but amides (63%) were most important in +N plants. Based on the composition of fruit tip phloem, ureides (55% and 33%) and amides (26% and 47%) accounted for the majority of N imported by fruits of −N and +N plants, respectively.

C:N weight ratios were lowest in xylem exudate (1.37 and 1.32), highest in petiole phloem (24.5 and 26.0), and intermediate in fruit tip exudate (12.6 and 12.1) for the −N and +N treatments, respectively. The ratios were combined with data on fruit growth and respiration to construct a model of C and N transport to developing fruits. The model indicates xylem to phloem transfer provides 35% to 52% of fruit N. Results suggest the phloem entering fruits oversupplies their N requirement so that 13% of the N imported is exported from fruit in the xylem.

     

Economy of Carbon and Nitrogen in Nodulated and Nonnodulated (NO(3)-grown) Cowpea [Vigna unguiculata (L.) Walp.

The response of non-nodulated cowpea (Vigna unguiculata (L.) Walp. cv Caloona) to a wide range of NO₃ levels in the rooting medium was studied 40 days after sowing by in vitro assays of plant organs for NO₃ reductase (EC 1.6.6.1) and analyses of root bleeding (xylem) sap for nitrogenous solutes. Plants fed 1, 5, 10, 20, and 40 millimolar NO₃ showed, respectively, 64, 92, 94, and 91% of their total reductase activity in shoots and 34, 30, 66, 62, and 58% of the total N of their xylem sap as NO₃. These data, and the absence in the plants of significant pools of stored NO₃, indicated that shoots were major organs of NO₃ assimilation, especially at levels of NO₃ (10 to 40 millimolar) that maintained plant growth at near maximum rates. Partitioning and utilization of C and N were studied in nodulated, minus NO₃ plants and non-nodulated plants fed 10 or 20 millimolar NO₃, the levels of NO₃ which gave rates of growth and N assimilation closest to those of the symbiotic plants. The conversion of the C of net photosynthate to dry matter was similar in nodulated plants (67%) and NO₃-grown plants (64%), but greater proportions of photosynthate were translocated to below ground parts of nodulated plants (37%) than of NO₃-fed plants (23 to 26%). Greater photosynthate consumption by nodulated roots was associated with proportionately greater root growth and respiration and 2-fold greater export of C in xylem than in the NO₃-fed plants. Theoretical considerations suggest that the elevated CO₂ output of nodulated roots was due not only to CO₂ loss associated with nodule function, but also to a much greater nonassimilatory component of respiration in the supporting root of the nodulated plant compared to roots of the NO₃-fed plants. Data are compared with previously published information from other legumes.     

Economy of Carbon and Nitrogen in a Nodulated and Nonnodulated (NO(3)-grown) Legume

Partitioning and utilization of assimilated C and N were compared in nonnodulated, NO₃-fed and nodulated, N₂-fed plants of white lupin (Lupinus albus L.). The NO₃ regime used (5 millimolar NO₃) promoted closely similar rates of growth and N assimilation as in the symbiotic plants. Over 90% of the N absorbed by the NO₃-fed plants was judged to be reduced in roots. Empirically based models of C and N flow demonstrated that patterns of incorporation of C and N into dry matter and exchange of C and N among plant parts were essentially similar in the two forms of nutrition. NO₃-fed and N₂-fed plants transported similar types and proportions of organic solutes in xylem and phloem. Withdrawal of NO₃ supply from NO₃-fed plants led to substantial changes in assimilate partitioning, particularly in increased translocation of N from shoot to root. Nodulated plants showed a lower (57%) conversion of C or net photosynthate to dry matter than did NO₃-fed plants (69%), and their stems were only half as effective as those of NO₃-fed plants in xylem to phloem transfer of N supplied from the root. Below-ground parts of symbiotic plants consumed a larger share (58%) of the plants' net photosynthate than did NO₃-fed roots (50%), thus reflecting a higher CO₂ loss per unit of N assimilated (10.2 milligrams C/milligram N) by the nodulated root than by the root of the NO₃-fed plant (8.1 milligrams C/milligram N). Theoretical considerations indicated that the greater CO₂ output of the nodulated root involved a slightly greater expenditure for N₂ than for NO₃ assimilation, a small extra cost due to growth and maintenance of nodule tissue, and a considerably greater nonassimilatory component of respiration in root tissue of the symbiotic plant than in the root of the NO₃-fed plant.     

Transport of organic solutes in Phloem and xylem of a nodulated legume

Collections of xylem exudate of root stumps or detached nodules, and of phloem bleeding sap from stems, petioles, and fruits were made from variously aged plants of Lupinus albus L. relying on nodules for their N supply. Sucrose was the major organic solute of phloem, asparagine, glutamine, serine, aspartic acid, valine, lysine, isoleucine, and leucine, the principal N solutes of both xylem and phloem. Xylem sap exhibited higher relative proportions of asparagine, glutamine and aspartic acid than phloem sap, but lower proportions of other amino acids. Phloem sap of petioles was less concentrated in asparagine and glutamine but richer in sucrose than was phloem sap of stem and fruit, suggesting that sucrose was unloaded from phloem and amides added to phloem as translocate passed through stems to sinks of the plant. Evidence was obtained of loading of histidine, lysine, threonine, serine, leucine and valine onto phloem of stems but the amounts involved were small compared with amides. Analyses of petiole phloem sap from different age groups of leaves indicated ontogenetic changes and effects of position on a shoot on relative rates of export of sucrose and N solutes. Diurnal fluctuations were demonstrated in relative rates of loading of sucrose and N solutes onto phloem of leaves. Daily variations in the ability of stem tissue to load N onto phloem streams were of lesser amplitude than, or out of phase with fluctuations in translocation of N from leaves. Data were related to recent information on C and N transport in the species.     

Spontaneous Phloem bleeding from cryopunctured fruits of a ureide-producing legume

The vasculature of the dorsal suture of cowpea (Vigna unguiculata [L.] Walp) fruits bled a sugar-rich exudate when punctured with a fine needle previously cooled in liquid N₂. Bleeding continued for many days at rates equivalent to 10% of the estimated current sugar intake of the fruit. A phloem origin for the exudate was suggested from its high levels (0.4-0.8 millimoles per milliliter) of sugar (98% of this as sucrose) and its high K⁺ content and high ratio of Mg²⁺ to Ca²⁺. Fruit cryopuncture sap became labeled with ¹⁴C following feeding of [¹⁴C]urea to leaves or adjacent walls of the fruit, of ¹⁴CO₂ to the pod gas space, and of [¹⁴C] asparagine or [¹⁴C]allantoin to leaflets or cut shoots through the xylem. Rates of translocation of ¹⁴C-assimilates from a fed leaf to the puncture site on a subtended fruit were 21 to 38 centimeters per hour. Analysis of ¹⁴C distribution in phloem sap suggested that [¹⁴C]allantoin was metabolized to a greater extent in its passage to the fruit than was [¹⁴C] asparagine. Amino acid:ureide:nitrate ratios (nitrogen weight basis) of NO₃-fed, non-nodulated plants were 20:2:78 in root bleeding xylem sap versus 90:10:0.1 for fruit phloem sap, suggesting that the shoot utilized NO₃-nitrogen to synthesize amino acids prior to phloem transfer of nitrogen to the fruit. Feeding of ¹⁵NO₃ to roots substantiated this conclusion. The amino acid:ureide ratio (nitrogen weight basis) of root xylem sap of symbiotic plants was 23:77 versus 89:11 for corresponding fruit phloem sap indicating intense metabolic transfer of ureide-nitrogen to amino acids by vegetative parts of the plant.     

Cytokinins in the Phloem Sap of White Lupin (Lupinus albus L.)

Cytokinin-like activity in samples of xylem and phloem sap collected from field-grown plants of white lupin (Lupinus albus L.) over a period of 9 to 24 weeks after sowing was measured using the soybean hypocotyl callus bioassay following paper chromatographic separation. The phloem sap was collected from shallow incisions made at the base of the stem, the base of the inflorescence (e.g. stem top), the petioles, and the base and tip of the fruit. Xylem sap was collected as root exudate from the stump of plants severed a few centimeters above ground level. Concentration of cytokinin-like substances was highest in phloem sap collected from the base of the inflorescence and showed an increase over the entire sampling period (from week 10 [61 nanogram zeatin equivalents] to week 24 [407 nanogram zeatin equivalents]). Concentrations in the xylem sap and in the other phloem saps were generally lower. Relatively high concentrations of cytokinin-like substances in petiole phloem sap (70 to 130 nanogram zeatin equivalents per milliliter) coincided in time with high concentrations in sap from the base of the inflorescence (see above). Concentrations in sap (phloem or xylem) from the base of the stem were very much lower. This finding is consistent with movement of cytokinins from leaves into the developing inflorescence and fruit, rather than direct input to the fruit from xylem sap. However, an earlier movement of cytokinins from roots into leaves via the xylem cannot be ruled out. Sap collected at an 18-week harvest was additionally separated by sequential C₁₈ reversed-phase high performance liquid chromatography → NH₂ normal phase high performance liquid chromatography, bioassayed, and then analyzed by electron impact gas chromatography-mass spectrometry. Identification of zeatin riboside and dihydrozeatin as two of the major cytokinins in combined sap samples was accomplished by gas chromatography-mass spectrometry-selected ion monitoring.     

Effects of n(2) deficiency on transport and partitioning of C and N in a nodulated legume

Nodulated root systems of white lupin (Lupinus albus L. cv Ultra: Rhizobium strain WU425) were exposed to Ar:O₂ (80:20, v/v) or Ar:N₂:O₂ (70:10:20, v/v/v) and C and N partitioning were examined over a 9- or 10-day period in comparison with control plants with nodulated roots retained in air. Accumulation of N ceased in plants exposed to Ar:O₂ or was much reduced in plants exposed to Ar:N₂:O₂, but net C assimilation rates and profiles of C utilization remained similar to those of control N₂-fixing plants. There was, however, a proportional reduction in CO₂ evolution from nodulated roots of the Ar:O₂ treatment. Xylem N levels fell rapidly after application of Ar:O₂. C:N ratios of phloem sap of petioles and of stem base rose during the first day of Ar:O₂ treatment and then fell progressively back to levels close to that of control plants as leaf reserves of N became available for loading of phloem. Stem top phloem sap increased progressively in C:N ratio throughout Ar:O₂ treatment, presumably due to increasing shortage of xylem derived N for xylem to phloem exchange. Reexposure of Ar:O₂-treated nodulated root systems to air prompted a rapid recovery of N₂ fixation and restoration of plant N status. Rates of N₂ fixation in plants whose roots were exposed to a range of N₂ concentrations indicated an apparent K_m of 10% N₂ for the attached intact white lupin nodule.     

Transport, metabolism, and redistribution of xylem-borne amino acids in developing pea shoots

Amino acid metabolism and transport was investigated in the leaves of 3-week-old nonnodulated seedlings of Pisum sativum L. Xylem sap entering the shoot contained nitrate (about 5 millimolar), and amino compounds (11 millimolar) of which 70% was asparagine plus glutamine; aspartate and homoserine were also present. Mature leaves showed stable nitrogen levels and incoming nitrogen was redistributed to growing leaves. Younger leaves, still enclosed in the stipules, showed negligible rates of transpiration, suggesting that most of their nitrogen must arrive in the phloem.     

Studies on Genetic Male-Sterile Soybeans : IV. Effect of Male Sterility and Source of Nitrogen Nutrition on Accumulation, Partitioning, and Transport of Nitrogen

Soybean (Glycine max [L.] Merr.) germplasm, isogenic except for loci controlling male sterility (ms₁) and nodulation (rj₁), was used to investigate the effects of reproductive tissue development and source of nitrogen nutrition on accumulation, transport, and partitioning of nitrogen in a greenhouse experiment. Nodulated plants were supplied nitrogen-free nutrient solution, and nonnodulated plants were supplied nutrient solution containing 20 millimolar KNO₃. Plants were sampled from flowering until maturity (77 to 147 days after transplanting).

Accumulation rates of nitrogen in whole plants during reproductive growth were not significantly different among the four plant types. Nitrogen accumulation in the sterile, nonnodulated plants, however, ceased 2 weeks earlier than in fertile, nonnodulated or fertile and sterile, nodulated plants. This early cessation in nitrogen accumulation resulted in sterile, nonnodulated plants accumulating significantly less whole plant nitrogen by 133 days after transplanting (DAT) than fertile, nonnodulated plants. Thus, changing the site of nitrogen assimilation from nodules (N₂-fixing plants) to roots and leaves (NO₃-fed plants) resulted in similar whole-plant nitrogen accumulation rates in fertile and sterile plants, despite the absence of seed in the latter.

Leaflet and stem plus petiole tissues of both types of sterile plants had significantly higher nitrogen concentrations after 119 DAT than both types of fertile plants. Significantly higher concentrations and exudation rates of nonureide, reduced-nitrogen in xylem sap of sterile than of fertile plants after 105 DAT were observed. These latter results indicated possible cycling of nonureide, reduced-nitrogen from the downward phloem translocation stream to the upward xylem translocation stream in roots of sterile plants. Collectively, these results suggest a lack of sinks for nitrogen utilization in the shoots of sterile plants. Hence, comparison of nitrogen accumulation rates for sterile and fertile plants does not provide a definitive test of the hypothesis that reproductive tissue development limits photosynthate availability for support of N₂ fixation and nitrate assimilation in determinate soybeans.

Nitrogen assimilation during reproductive growth met a larger proportion of the reproductive-tissue nitrogen requirement of nitrate-dependent plants (73%) than of N₂-fixing plants (63%). Hence, vegetative-tissue nitrogen mobilization to reproductive tissue was a more prominent process in N₂-fixing than in nitrate-dependent plants. N₂-fixing plants partitioned nitrogen to reproductive tissue more efficiently than nitrate-dependent plants as the reproductive tissues of the former and latter contained 65 and 55%, respectively, of the whole-plant nitrogen at the time that nitrogen accumulation in reproductive parts had ceased (133 DAT).

     

Distribution of NO3− reduction between roots and shoots of peachtree seedlings as affected by NO3− uptake rate

The distribution of NO₃^? reduction between roots and shoots was studied in hydro-ponically-grown peach-tree seedlings (Prunus persica L.) during recovery from N starvation. Uptake, translocation and reduction of NO₃^?, together with transport through xylem and phloem of the newly reduced N were estimated, using ¹⁵N labellings, in intact plants supplied for 90 h with 0.5 mM NH₄⁺ and 0.5, 1.5 or 10 mM NO₃^?. Xylem transport of NO₃^? was further investigated by xylem sap analysis in a similar experiment. The roots were the main site of NO₃^? reduction at all 3 levels of NO₃^? nutrition. However, the contribution of the shoots to the whole plant NO₃^? reduction increased with increasing external NO₃^? availability. This contribution was estimated to be 20, 23 and 42% of the total assimilation at 0.5, 1.5 and 10 mM NO₃^?, respectively. Both ¹⁵N results and xylem sap analysis confirmed that this trend was due to an enhancement of NO₃^? translocation from roots to shoots. It is proposed that the lack of NO₃^? export to the shoots at low NO₃^? uptake rate resulted from a competition between NO₃^? reduction in the root epidermis/cortex and NO₃^? diffusion to the stele. On the other hand, net xylem transport of newly reduced N was very efficient since ca 70% of the amino acids synthesized in the roots were translocated to the shoots, regardless of the level of NO₃^? nutrition. This net xylem transport by far exceeded the net downward phloem transport of the reduced N assimilated in shoots. As a consequence, the reduced N resulting from NO₃^? assimilation, principally occurring in the roots, was mainly incorporated in the shoots.     

Isotopic composition and concentration of total nitrogen and nitrate in xylem sap under near steady-state hydroponics

After root uptake, nitrate is effluxed back to the medium, assimilated locally, or translocated to shoots. Rooted black cottonwood (Populus trichocarpa) scions were supplied with a NO₃⁻-based (0.5 mM) nutrient medium of known isotopic composition (δ¹⁵N), and xylem sap was collected by pressure bombing. To establish a sampling protocol, sap was collected from lower and upper stem sections at 0.1–0.2 MPa above the balancing pressure, and after increasing the pressure by a further 0.5 MPa. Xylem sap from upper stem sections was partially diluted at higher pressure. Further analysis was restricted to sap obtained from intact shoots at low pressure. Total-, NO₃⁻-N and, by difference, organic-N concentrations ranged from 6.1–11.0, 1.2–2.4, and 4.6–9.4 mM, while discrimination relative to the nutrient medium was −6.3 to 0.5‰, −23.3 to −11.5‰ and − 1.3 to 4.9‰, respectively. There was diurnal variation in δ¹⁵N of total- and organic-N, but not NO₃⁻. The difference in δ¹⁵N between xylem NO₃⁻ and organic-N suggests that discrimination by nitrate reductase is near 25.1 ± 1.6‰. When this value was used in an isotope mass balance model, the predicted xylem sap NO₃⁻-N to total-N ratio closely matched direct measurement.     

Relative Content of NO(3) and Reduced N in Xylem Exudate as an Indicator of Root Reduction of Concurrently Absorbed NO(3)

It is unclear if the relative content of NO₃⁻ and reduced N in xylem exudate provides an accurate estimate of the percentage reduction of concurrently absorbed NO₃⁻ in the root. Experiments were conducted to determine whether NO₃⁻ and reduced N in xylem exudate of vegetative, nonnodulated soybean plants (Glycine max [L.] Merr., `Ransom') originated from exogenous recently absorbed ¹⁵NO₃⁻ or from endogenous ¹⁴N pools. Plants either were decapitated and exposed to ¹⁵NO₃⁻ solutions for 2 hours or were decapitated for the final 20 minutes of a 50-minute exposure to ¹⁵NO₃⁻ in the dark and in the light. Considerable amounts of ¹⁴NO₃⁻ and reduced ¹⁴N were transported into the xylem, but almost all of the ¹⁵N was present as ¹⁵NO₃⁻. Dissimilar changes in transport of ¹⁴NO₃⁻, reduced ¹⁴N and ¹⁵NO₃⁻ during the 2 hours of sap collection resulted in large variability over time in the percentage of total N in the exudate which was reduced N. Over a 20-minute period the rate of ¹⁵N transport into the xylem of decapitated plants was only 21 to 36% of the ¹⁵N delivered to the shoot of intact plants. Based on the proportion of total ¹⁵N which was found as reduced ¹⁵N in exudate and in intact plants in the dark, it was estimated that 5 to 17% of concurrently absorbed ¹⁵NO₃⁻ was reduced in the root. This was much less than the 38 to 59% which would have been predicted from the relative content of total NO₃⁻ and total reduced N in the xylem exudate.     

Temporal Patterns of Uptake, Flow and Utilization of Nitrate, Reduced Nitrogen and Carbon in a Leaf of Salt-Treated Castor Bean (Ricinus communis L.)

Changes in net photosynthesis, respiration, transpiration andcontents of total C, NO₃-N and reduced N were followed throughoutthe life of leaf 6 of nitrate-dependent plants of castor beanexposed to moderate salinity stress (71 mol m^–3 NaCl).Salt treatment was applied for measuring mineral flows in aparallel study (Jeschke and Pate, 1991b). Concurrent measurementswere made of solute composition and C: N molar ratios and concentrationsof reduced N and collected NO₃-N in phloem sap bleeding fromshallow incisions in the top and at the base of petioles andin xylem exudates from flaps of proximal leaf midribs followingpressurization of the root system. The resulting data were usedto construct empirical models of the respective economies ofC, total N, NO₃ and reduced N for a sequence of defined phasesof leaf life. Water use efficiency increased 3-fold from emergenceto a maximum of 1·5 mmol CO₂ mol^–1 H₂O before decliningto 0·5 mmol CO₂ mol^–1 H₂O at senescence. Xylemmolar ratios of C:N varied from 1·2–2·8,with nitrate always a smaller component than reduced N. Phloemsap C:N increased from 10–40 with leaf expansion and wasthen maintained in the range of 40–50 until falling steeplyto 20 at leaf senescence. Nitrate comprised less than 1% oftotal N in all phloem sap samples. The models of C uptake, flow,and utilization showed a major role of phloem import and thenincreasingly of laminar photosynthesis in providing C for leafgrowth. The carbon budget was thereafter characterized by ratesof phloem export closely matched to net rates of CO₂ fixationby the lamina. Corresponding data for total N depicted an earlymajor role of both xylem and phloem import, but the eventualdominance of xylem import as the N source for leaf growth. Cyclingof N by xylem to phloem exchange commenced before the leaf hadachieved maximum N content, and was the major contributor tophloem export until leaf senescence when mobilized N providedmost exported N. The nitrate economy of the leaf was characterizedby early establishment of tissue pools of the ion in the petioleand to a lesser extent in the lamina, continued high rates ofnitrate reduction in the lamina but negligible assimilationin the petiole, and a release through xylem of previously accumulatedNO₃ from petiole to lamina. Related data for reduced N illustratedthe much greater importance of this form of N than nitrate intransport, storage and cycling of N at all stages of leaf andpetiole life. Xylem to phloem interchanges of reduced N in petiolewere minimal in comparison with cycling through the lamina.The ratio of CO₂ reduction to NO₃ reduction in the lamina wasat first low (57 mol mol^–1) increasing to a peak valueof 294 during mature leaf functioning before declining to 190during the presenescence phase of leaf development. This patternreflected age-related effects on water use efficiency, changesin NO₃ levels in the xylem stream entering the lamina, and therelatively low photosynthetic performances of very young andsenescent laminae. Key words: Ricinus communis, leaf development, phloem transport, xylem transport, carbon, nitrogen, nitrate, reduced nitrogen, nitrate reduction, partitioning     

Uptake and Utilization of Xylem-borne Amino Compounds by Shoot Organs of a Legume

Amino compounds representative of the major N solutes of xylem sap were pulse-fed (10 to 20 minutes) singly in ¹⁴C-labeled form to cut transpiring shoots of white lupin (Lupinus albus L.). ¹⁴C distribution was studied by autoradiography and radioassays of phloem sap, leaflet tissues, and shoot parts harvested at intervals after labeling. Primary distribution of N by xylem was simulated using a 20-minute labeling pulse followed by a 30-minute chase in unlabeled xylem sap. Shoots fed ¹⁴C-labeled asparagine, glutamine, valine, serine, or arginine showed intense labeling of leaflet veins and marked retention (35 to 78%) of ¹⁴C by stem + petioles. Shoots fed ¹⁴C-labeled aspartic acid or glutamic acid showed heaviest ¹⁴C accumulation in interveinal regions of leaflets and low uptake (11 to 20%) of ¹⁴C by stem + petioles. Departing leaf traces were major sites of uptake of all amino compounds, and the implications of this were evaluated. Fruits acquired only 1 to 5% of the fed label directly from xylem, but more than doubled their intake during the period 30 to 160 minutes after feeding through receipt of ¹⁴C transferred from xylem to phloem in stem and leaves. ¹⁴C-Labeled asparagine and valine transferred directly from xylem to phloem, but the ¹⁴C of ¹⁴C-labeled aspartic acid and arginine appeared in phloem mainly as metabolic products of the fed compound. The labeling of the soluble pool of leaflets reflected these differences. The significance of heterogeneity in distribution and metabolism of xylem amino compounds in the shoot was discussed.     

Enhancement of Phloem Exudation from Fraxinus uhdei Wenz. (Evergreen Ash) using Ethylenediaminetetraacetic Acid

Ethylenediaminetetraacetic acid (EDTA) enhanced the exudation of ¹⁴C-labeled assimilates from excised leaflets and whole plant specimens of Fraxinus uhdei Wenz. A 2 millimolar EDTA concentration was found to be most effective in promoting exudation from excised leaflets, while 10 millimolar EDTA was most effective in whole plants experiments. Exudation rate reached a maximum after 24 hours in both experiments. The continuous presence of EDTA throughout the treatment period was required for maximum exudation from excised leaflets. Stachyose, raffinose, verbascose, and sucrose were the principal compounds found to occur in exudate samples. These compounds are typically transported in sieve elements of various Fraxinus species suggesting the exudate was of phloem origin. Electron microscope studies of petiolule sieve plate pores from excised leaflets showed substantially less callose appearing after treatment with EDTA than after H₂O treatment. It is suggested that EDTA enhances phloem exudation by inhibiting or reducing callose formation in sieve plate pores. The exudation enhancement technique described for whole plant specimens is suggested as a useful means of collecting phloem sap and studying translocation in woody plants.     

Diurnal Patterns of Transport and Accumulation of Minerals in Fruiting Plants of Lupinus angustifolius L.

Fluctuations in mineral elements id xylem (tracheal) sap, fruitphloem sap, leaflets and dmloping fruits were studied in a fieldpopulation of Lupinus angustifolius L. by three-hourly samplingover a 39 h period. Elements usually reached maximum contentsor concentrations at or near noon, minimum levels during thenight. Amplitudes of diurnal fluctuations in minerals lay withinthe range ±4–33 per cent of the mean content ofleaflets, and ±17–157 per cent of the mean concentrationsin xylem and phloem sap. Most minerals elements fluctuatcd inphase with daily changes in sugar level of phloem sap and drymatter and carbohydrate fluctuations of leaflets, suggestinga coupling of translocation of photosynthate and minerals fromthe leaflets. Rates of import of minerals by shoots wereestimatedfrom shoot transpiration and mineral concentrations in trachealsap. Average day time rates of import of most elements were12–25 times those at night. Translocation of minerals,nitrogen and carbon to fruits also exhibited diurnal periodicity,average rates of import king three to seven times higher inthe day than at night. A model of transport based on the carbonand water economy of the fruit suggested that P, K, Fe, Zn,Mn and Cu were imported predominantly by phloem. Estimates ofvascular import accounted for 87–104 per cent of the fruit'sactual increment of these elements. Na and Ca were gauged tobe imported mainly by xylem, Mg almost equally by xylem andphloem. However, large discrepancies existed for these threeelements between estimated vascular import and actual intakeby the fruit. Lupinus angustifolius L., mineral transport, accumulation, fruits, xylem sap, phloem sap, transpiration     

How aphids decide what is good for them: experiments to test aphid feeding behaviour on Tanacetum vulgare (L.) using different nitrogen regimes

Leaf-chewing herbivores select food with a protein/carbohydrate ratio of 0.8–1.5, whereas phloem sap, which aphids feed on, has a ratio of ~0.1. Enhanced N fertilization increases the amino acid concentration in phloem sap and elevates the N/C ratio. The study examines: (1) whether aphids select between plants of different N nutrition, (2) whether feeding time correlates with the amino acid composition of phloem sap, and (3) at which stage of probing aphids identify the quality of the plant. Uroleucon tanaceti (Mordvilko) and Macrosiphoniella tanacetaria (Kaltenbach), specialist aphids feeding on tansy (Tanacetum vulgare L.), were reared on this host plant grown essentially hydroponically (in Vermiculite) in the greenhouse on 1, 3, 6, or 12 mM NH₄NO₃. One and 3 mM NH₄NO₃ corresponds to the situation found in natural tansy stands. Aphid stylet penetration was monitored by electrical penetration graphs whilst phloem sap was sampled by stylectomy. Both aphid species settled 2–3 times more frequently on plants fertilized with 6 or 12 mM NH₄NO₃. The phloem sap of these plants contained up to threefold higher amino acid concentrations, without a change in the proportion of essential amino acids. No time differences were observed before stylet penetration of plant tissue. After the first symplast contact, most aphids penetrated further, except M. tanacetaria on low-N plants, where 50% withdrew the stylet after the first probing. The duration of phloem feeding was 2–3 times longer in N-rich plants and the time spent in individual sieve tubes was up to tenfold longer. Aphids identified the nutritional quality of the host plant mainly by the amino acid concentration of phloem sap, not by leaf surface cues nor the proportion of essential amino acids. However, U. tanaceti infestation increased the percentage of methionine plus tryptophan in phloem tenfold, thus manipulating the plants nutritional quality, and causing premature leaf senescence.     

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.