Sucrose efflux and export from the maize scutellum*

Abstract During incubation of maize scutellum slices in fructose, there was an efflux of sucrose. Efflux was constant for at least 4 h at fructose concentrations of 70 or 100 mol m^?3. Efflux was increased by EDTA, and decreased by Ca²⁺. Efflux was independent of pH after EDTA treatment, but increased from untreated slices when the pH was lowered from 7 to 4. Uranyl ion and PCMBS (p-chloro-mercuribenzenesulfonic acid) abolished sucrose uptake, but were only weak inhibitors of sucrose efflux. These results are consistent with efflux occurring by simple diffusion through aqueous pores, but they do not rule out facilitated diffusion. Rates of sucrose export from the scutellum to the root shoot axis were estimated from measurements of axis respiration and dry weight gain. Sucrose efflux from scutellum slices was only 14-22% of the export rate. Sucrose efflux from the whole scutellum was only 3-4% of the export rate. It is concluded that the observed efflux is from leaky cells and does not represent sucrose on the way to the phloem along a path that includes the apoplast. These results support the idea that the path for sucrose from parenchyma cell to sieve tube in the maize scutellum is entirely symplastic.     

Untersuchungen zur funktionellen Anatomie des Leitgewebesystems im Blatt von Pelargonium zonale

Summary The petiole of Pelargonium zonale is traversed by 17 bundles, whose arrangement and form are typical for this plant. The bundles of the petiole are connected with the conducting system of the axis and with the main nerves by a system of phloem anastomoses in the leaf base and in the junction between the petiole and the leaf blade (Fig. 2). The anatomical findings were confirmed and extended by a study of the translocation of K-fluorescein and ¹⁴C. It could be shown that the metaphloem of the central petiole bundle is composed of phloem subunits, each of which is connected with the phloem of one certain main nerve only (Fig. 4). Accordingly, if fluorescein or ¹⁴CO₂ is applied to one main nerve, the dye or ¹⁴C-material is translocated exclusively in a small phloem area of the central bundle. Autoradiograms of the petioles indicate that the ¹⁴C-labelled assimilates (sucrose, glucose, fructose and amino acids) are translocated exclusively in the phloem. A lateral movement of the labelled material within the petiole was not observed. The metaphloem of the central petiole bundle of Pelargonium zonale revealed a functional organization of phloem subunits.

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Teil einer Dissertation unter der wissenschaftlichen Leitung von Prof. Dr. J. Willenbrink.     

Photoassimilate translocation in the petiole of Cyclamen and Primula is independent of lateral retrieval

Phloem translocation of photoassimilates between source andsink is considered to be linked with active retrieval of sugarsleaked to the vascular apoplast. This hypothesis was evaluatedby studying photo-assimilate movement in petioles of intactplants of Cyclamen persicum and Primula obconica in the presenceof inhibitors affecting sucrose retrieval (PCMBS, CCCP). Inhibitorsolutions were applied by rinsing locally isolated petiole bundlesor by injection into the petioe parenchyma. PCMBS and CCCP reduced[¹⁴C]sucrose retrieval from the petiole apoplast by the vascularcells and altered the distribution pattern of ¹⁴C-photoassimilateswithin the petiole tissues. However, these treatments did notaffect translocation through the petiole phloem. Evidence isprovided that the reagents were present in the vascular apoplastsurrounding the translocating phloem. It was concluded thatassimilate movement in the petiole of Cyclamen and Primula wasindependent of apoplastic retrieval. Key words: Cyclamen, Primula, phloem, transport, path, sucrose, retrieval     

Sugar synthesis and phloem loading in Coleus blumei leaves

Sugar-synthesis and -transport patterns were analyzed in Coleus blumei Benth. leaves to determine where galactinol, raffinose, and stachyose are made and whether phloem loading includes an apoplastic (extracellular) step or occurs entirely within the symplast (plasmodesmata-connected cytoplasm). To clarify the sequence of steps leading to stachyose synthesis, a pulse (15 s) of ¹⁴CO₂ was given to attached leaves followed by a 5-s to 20-min chase: sucrose was rapidly labeled while galactinol, raffinose and stachyose were labeled more slowly and, within the first few minutes, to approximately the same degree. Leaf tissue was exposed to either ¹⁴CO₂ or [¹⁴C]glucose to identify the sites of synthesis of the different sugars. A 2-min exposure of peeled leaf tissue to [¹⁴C]glucose resulted in preferential labeling of the minor veins, as opposed to the mesophyll; galactinol, raffinose and stachyose were more heavily labeled than sucrose in these preparations. In contrast, when leaf tissue was exposed to ¹⁴CO₂ for 2 min for preferential labeling of the mesophyll, sucrose was more heavily labeled than galactinol, raffinose or stachyose. We conclude that sucrose is synthesized in mesophyll cells while galactinol, raffinose and stachyose are made in the minorvein phloem. Competition experiments were performed to test the possibility that phloem loading involves monosaccharide uptake from the apoplast. Two saturable monosaccharide carriers were identified, one for glucose, galactose and 3-O-methyl glucose, and the other for fructose. Washing the apoplast of peeled leaf pieces with buffer or saturating levels of 3-O-methyl glucose, after providing a pulse of ¹⁴CO₂, did not inhibit vein loading or change the composition of labeled sugars, and less than 0.5% of the assimilated label was recovered in the incubation medium. These and previous results (Turgeon and Gowan, 1991, Plant Physiol. 94, 1244–1249) indicate that the phloem loading pathway in Coleus is probably symplastic.Abbreviations 3-OMG 3-O-methyl glucose - PCMBS p-chloromercuribenzenesulfonic acid - SE-CCC sieve-element-companion-cell complex This research was supported by National Science Foundation Grant DCB-9104159, U.S. Department of Agriculture Competetive Grant 90000854, and Hatch funds.     

Phloem Unloading in Developing Leaves of Sugar Beet : I. Evidence for Pathway through the Symplast

Physiological and transport data are presented in support of a symplastic pathway of phloem unloading in importing leaves of Beta vulgaris L. (`Klein E multigerm'). The sulfhydryl reagent p-chloromercuribenzene sulfonic acid (PCMBS) at concentration of 10 millimolar inhibited uptake of exogenous [¹⁴C]sucrose by sink leaf tissue over sucrose concentrations of 0.1 to 5.0 millimolar. Inhibited uptake was 24% of controls. The same PCMBS treatment did not affect import of ¹⁴C-label into sink leaves during steady state labeling of a source leaf with ¹⁴CO₂. Lack of inhibition of import implies that sucrose did not pass through the free space during unloading. A passively transported xenobiotic sugar, l-[¹⁴C]glucose, imported by a sink leaf through the phloem, was evenly distributed throughout the leaf as seen by whole-leaf autoradiography. In contrast, l-[¹⁴C]glucose supplied to the apoplast through the cut petiole or into a vein of a sink leaf collected mainly in the vicinity of the major veins with little entering the mesophyll. These patterns are best explained by transport through the symplast from phloem to mesophyll.     

The extrafloral nectaries of cowpea (Vigna unguiculata (L.) Walp.) II. Nectar composition,origin of nectar solutes,and nectary functioning

Nectar was collected from the extrafloral nectaries of leaf stipels and inflorescence stalks, and phloem sap from cryopunctured fruits of cowpea plants. Daily sugar losses as nectar were equivalent to only 0.1–2% of the plant's current net photosynthate, and were maximal in the fourth week after anthesis. Sucrose:glucose:fructose weight ratios of nectar varied from 1.5:1:1 to 0.5:1:1, whereas over 95% of phloem-sap sugar was sucrose. [¹⁴C]Sucrose fed to leaves was translocated as such to nectaries, where it was partly inverted to [¹⁴C]glucose and [¹⁴C]fructose prior to or during nectar secretion. Invertase (EC 3.2.1.26) activity was demonstrated for inflorescence-stalk nectar but not stipel nectar. The nectar invertase was largely associated with secretory cells that are extruded into the nectar during nectary functioning, and was active only after osmotic disruption of these cells upon dilution of the nectar. The nectar invertase functioned optimally (phloem-sap sucrose as substrate) at pH 5.5, with a starting sucrose concentration of 15% (w/v). Stipel nectar was much lower in amino compounds relative to sugars (0.08–0.17 mg g^-1 total sugar) than inflorescence nectar (22–30 mg g^-1) or phloem sap (81–162 mg g^-1). The two classes of nectar and phloem sap also differed noticeably in their complements of organic acids. Xylem feeding to leaves of a range of ¹⁴C-labelled nitrogenous solutes resulted in these substrates and their metabolic products appearing in fruit-phloem sap and adjacent inflorescence-stalk nectar. ¹⁴C-labelled asparagine, valine and histidine transferred freely into phloem and appeared still largely as such in nectar. ¹⁴C-labelled glycine, serine, arginine and aspartic acid showed limited direct access to phloem and nectar, although labelled metabolic products were transferred and secreted. The ureide allantoin was present in phloem, but absent from both types of nectar. Models of nectary functioning are proposed.     

Retention of xenobiotics along the phloem path

Detached leaves of Cyclamen persicum Mill. can be used as a simple source-sink system. Phloem transport in the excised material was monitored by the noninvasive ¹¹C-technique. Assimilate movement stopped immediately when the petiole was cut off. However, within 20 min a recovery of transport was observed. The translocation rate in the detached leaf was only 13% of that in the intact plant. ¹⁴C-Xenobiotics and [³H]sucrose were injected into the upper petiole parenchyma (source). They moved downstream by a symplastic route. The stump of the petiole was inserted into a buffer solution containing ethylenediaminetetraacetic acid (sink). After 3 h, the distribution of sucrose and xenobiotics was determined in five subsequent segments of the petiole (path). The retention coefficient (r) was calculated from the ratio of radioactivity in the vascular bundle to that in the petiole parenchyma. The distribution along the vascular path was given by a geometric progression, whereas its constant was the transport coefficient (q). Values of r and q corresponded with the degree of phloem mobility and ambimobility. Four groups of compounds were classified: (i) acidic substances with log K_ow = — 2 to — 2.4 (K_ow is the partition coefficient octanol/water) at pH 8 (pH of sieve tube sap), retained by ion trapping and exhibiting small lateral efflux (q0.7; maleic hydrazide, dalapon); (ii) acidic substances with log K_ow = — 0.7 to — 0.8 at pH 8, retained by ion trapping and subjected to a moderate lateral efflux (0.7>q> 0.5; 2,4-dichlorophenoxyacetic acid, 2-methyl-4-chlorophenoxyacetic acid, bromoxynil); (iii) nonionised substances retained by optimum permeability, exhibiting a considerable lateral leakage (q<0.5; glyphosate, amitrole); (iv) substances without basipetal transport in the phloem (atrazine, diuron). Retention of sucrose corresponded quantitatively with that shown in group (i). This classification was also supported by results of uptake and efflux experiments using the isolated conducting tissue. Theoretical translocation profiles were calculated from the determined transport coefficients (q).Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - K_ow partition coefficient octanol/water - MCPA 2-methyl-4-chloro-phenoxyacetic acid - q transport coefficient in the vascular bundle - r retention coefficient in the vascular bundle The authors gratefully acknowledge the assistance of H. Fiedler and M. Neugebauer. We are particularly grateful to K. Dutschka, G. Hudepokl, and Dr. J. Knust for producing ¹¹CO₂.     

Carbon fluxes in mature peach leaves

The turnover and transport of sugars are described in peach (Prunus persica L. Batsch), a species exporting both sucrose and sorbitol. Apparent export rate was slower in peach leaves than in leaves of herbaceous species. Sorbitol was the major soluble end product of photosynthesis and the major soluble carbohydrate in the leaf (higher than sucrose). Carbon fluxes were described using ¹⁴C labeling, radioactivity loss curves, and compartmental analysis during the second half of the photoperiod when chemical steady state was reached for soluble carbohydrates. The measured specific radioactivity of sucrose was typical of a primary product. The delayed decrease in specific radioactivity of sorbitol indicated that part of it was secondarily synthesized. Sucrose is proposed to be the carbon source for the delayed synthesis of sorbitol in the light. The sorbitol to sucrose ratio was higher in the petiole than in the leaf tissues. In phloem sap, obtained using stylectomy of aphids and collected from the main stem between source leaves and apex, this ratio was lower than in the petiole, suggesting a preferential sorbitol demand by sinks.     

Long-term Metabolism of [14C]Sugars in Stored Potatoes

[¹⁴C]Sucrose, [¹⁴C]glucose and [¹⁴C]fructose were introducedinto potato tubers held at 10 °C and the redistributionof label chased over a 65 d period in storage. Respiratory losseswere identical in all treatments, as was the partitioning of¹⁴C between soluble and insoluble forms. Sucrose was the predominantlabelled sugar in the tubers after 20 h, regardless of the original[¹⁴C]sugar introduced, and was loaded and distributed throughoutthe tubers by the internal phloem system. After 20 h the proportionsof labelled sugars bore no relationship to those of the unlabelledendogenous sugars. However, with time the percentage of ¹⁴Cin sucrose fell while that in glucose increased and by 65 dthe proportions of the labelled sugars more closely resembledthe endogenous pools. Fructose represented a consistently lowproportion of both the labelled and unlabelled sugars. By 21d a considerable proportion of the soluble ¹⁴C had been convertedto starch (approx. 25% of the total tuber 14C), this value remainingrelatively constant for the remainder of the storage period.Sprouts which formed on the tubers contained up to 6% of thetotal tuber ¹⁴C but less than 0.2% of the tuber dry matter.It is suggested that the bulk of the translocated [¹⁴C]sucroseentered the symplast and exchanged slowly with the bulk of thesugars in the storage cell vacuoles. [¹⁴C]sugars, phloem loading, starch, potato tuber, Solunum tuberosum, cold storage     

The effect of season of growth on the chemical composition of cambial saps of Eucalyptus regnans trees

Summary Bark was stripped, at monthly intervals, from the stems of ten previously-unsampled trees of Eucalyptus regnans F. Muell. The exposed surfaces of inner phloem and outer xylem yielded phloem and cambial saps which were rapidly frozen. After freeze drying to determine the contents of water and dry-matter, the samples were extracted with 80% ethanol. The main components in this extract are low molecular weight carbohydrates and salts of inorganic acids. The carbohydrates comprise stachyose, raffinose, sucrose, galactinol, glucose, fructose, myo-inositol and galactose; sucrose is invariably the major component. The amounts of all components varied widely during the sampling period. Multiple regression analyses showed that season of growth has a significant effect on sucrose, glucose, fructose, total sugars and soluble dry-matter, maxima being recorded near the beginning of autumn and spring, and minima near the beginning of winter and summer; that oligosaccharide and myoinositol contents are significantly related to atmospheric temperature; and that rainfall has a significant effect on the hexose and total sugar contents, saps from the xylem surfaces being more affected than those from the phloem surfaces. The translocated photosynthates in E. regnans appear to be oligosaccharides of the raffinose family and sucrose. Significant negative correlations between oligosaccharides and both sucrose and myoinositol, and significant positive correlations between sucrose and both glucose and fructose, are consistent with enzymic hydrolysis and resynthesis of most di- and oligosaccharides. The biosynthetic demands of developing secondary tissues and/or the fluctuations in composition of sieve-tube assimilates appear to control the composition of the sugars in the saps. Oligosaccharides and sucrose may function as soluble reserve substances as well as translocated photosynthates. It is possible that myoinositolis a key component in the interconversion processes of the sugars; experiments with radioactive sugars tend to lend support to this contention, especially during winter conditions.     

Phloem loading in peach: Symplastic or apoplastic?

Sorbitol and sucrose are the two main soluble carbohydrates in mature peach leaves. Both are translocated in the phloem, in peach as in other rosaceous trees. The respective role of these two soluble carbohydrates in the leaf carbon budget, and their phloem loading pathway, remain poorly documented. Though many studies have been carried out on the compartmentation and export of sucrose in sucrose-transporting species, far less is known about sorbitol in species transporting both sucrose and sorbitol. Sorbitol and sucrose concentrations were measured in several tissues and in sap, in 2-month-old peach (Prunus persica L. Batsch) seedlings, i.e. leaf blade, leaf main vein, petiole, xylem sap collected using a pressure bomb, and phloem sap collected by aphid stylets. The sorbitol to sucrose molar ratio depended on the tissue or sap, the highest value (about 7) found in the leaf main vein. Sorbitol concentration in the phloem sap was about 560 mM, whereas that of sucrose was about 140 mM. The lowest sorbitol and sucrose concentrations were observed in xylem sap collected from the shoot. The volume of the leaf apoplast, estimated by infiltration with ³H-inulin, represented about 17% of the leaf blade water content. This volume was used to calculate a global intracellular concentration for each carbohydrate in the leaf blade. Following these simplifying assumptions, the calculated concentration gradient between the leaf's intracellular compartment and phloem sap is nil for sorbitol and could thus allow for the symplastic loading of the phloem of this alditol. However, infiltration of ¹⁴C-labelled source leaves with 2 mMp-chloromercuribenzenesulfonic acid (PC-MBS), a potent inhibitor of the sucrose carrier responsible for phloem loading in sucrose-transporting plants, had a significant effect on the exudation of both labelled sucrose and sorbitol from the phloem. Therefore, in peach, which is a putative symplastic loader according to minor vein anatomy and sorbitol concentration gradients, apoplastic loading may predominate.     

Translocation and accumulation of translocate in the sugar beet petiole

Accumulation of translocate during steady-state labeling of photosynthate was measured in the source leaf petioles of sugar beet (Beta vulgaris L. monogerm hybrid). During an 8-hr period, 2.7% of the translocate or 0.38 μg carbon/min was accumulated per cm petiole. Material was stored mainly as sucrose and as compounds insoluble in 80% ethanol. The minimum peak velocity of translocation approached an average of 54 cm/hr as the specific activity of the ¹⁴CO₂ pulse was progressively increased. The ratio of cross sectional area required for translocation to actual sieve tube area in the petiole was 1.2. A regression analysis of translocation rate versus sieve tube cross sectional area yielded a coefficient of 0.76. The specific mass transfer rate in the petiole was 1.4 g/hr cm² phloem or 4.8 g/hr cm² sieve tube. Histoautoradiographic studies indicated that translocation occurs through the area of phloem occupied by sieve tubes and companion cells while storage occurs in these cells plus cambium and phloem parenchyma cells. The ability of the petiole to act as a sink for translocate is consistent with the concept that storage along path tissue serves to buffer sucrose concentration in the translocate during periods of fluctuating assimilation.     

Endogenous Sugar Levels and Their Effects on Root Formation and Petiole Yellowing of Detached Mustard Cotyledons

Detached cotyledons of Sinapis alba rooted readily in water in petri dishes in the light. The addition of (6 × 10^?2M) galactose, mannose or 2-deoxy-D-glucose to the culture medium proved toxic to cotyledon growth. Of the other sugars tested that were not toxic, sucrose was the most inhibitory to root formation and increased petiolar yellowing to the greatest extent. Glucose was more inhibitory than fructose which in turn increased petiolar yellowing more than methyl-D-glucose. Sucrose, glucose, or fructose at 6 × 10^?2M in the culture medium gave rise to very substantial increases in the cotyledon petiole of reducing sugar and starch with smaller increases in sucrose. Methyl-D-glucose had much less effect on internal sugar levels. It was found that the higher the internal level of glucose the more rapid the rate and final extent of petiolar yellowing. In general, the degree of petiolar yellowing was inversely related to the ability of the cotyledon to root. Methyl-D-glucose differed from the other sugars in that it delayed and reduced root formation but had very little effect on petiolar yellowing.     

Autoradiographic and biochemical evidence for the systemic translocation of systemin in tomato plants

The movement of systemin, the 18-amino-acid polypeptide inducer of proteinase inhibitors in tomato (Lycopersicon esculentum L.) plants, was investigated in young tomato plants following the application of [¹⁴C]systemin to wounds on the surface of leaves. Wholeleaf autoradiographic analyses revealed that [¹⁴C]systemin was distributed throughout the wounded leaf within 30 min, and then during the next several hours was transported to the petiole, to the main stem, and to the upper leaves. The movement of [¹⁴C]systemin was similar to the movement of [¹⁴C]sucrose when applied to leaf wounds, except that sucrose was slightly more mobile than systemin. Analyses of the radioactivity in the petiole phloem exudates at intervals over a 5-h period following the application of [¹⁴C]systemin to a wound demonstrated that intact [¹⁴C]systemin was present in the phloem over the entire time, indicating that the polypeptide was either stable for long periods in the phloem or was being continually loaded into the phloem from the source leaf. The translocation pathway of systemin was also investigated at the cellular level, using light microscopy and autoradiography. Within 15 min after application of [³H]systemin to a wound on a terminal leaflet, it was found distributed throughout the wounded leaf and was primarily concentrated in the xylem and phloem tissues within the leaf veins. After 30 min, the radioactivity was found mainly associated with vascular strands of phloem tissue in the petiole and, at 90 min, label was found in the phloem of the main stem. Altogether, these and previous results support a role for systemin as a systemic wound signal in tomato plants.The authors acknowledge the Washington State University Electron Microscope Center and staff for their technical advice and collaboration. We also thank Greg Wichelns for growing our plants and Dr. Steven Doares for providing [³H]systemin. This research was supported in part by the Washington State College of Agriculture and Home Economics Project No. 1791 and National Science Foundation grants IBN 9117795 and IBN 9104542     

The kinetics of sucrose concentration in the phloem of individual vascular bundles of the Ricinus communis seedling measured by nuclear magnetic resonance microimaging

The sucrose concentration was measured at 70-min intervals in the phloem of individual bundles of the hypocotyl of Ricinus seedlings by ¹H nuclear magnetic resonance (NMR) spectroscopic imaging. The sucrose concentration stayed fairly constant in all bundles for more than 7 h if the cotyledons were embedded in the endosperm or excised and incubated in 100 mM sucrose. If, however, the sucrose solution was replaced by sucrose-free buffer solution, the sucrose levels in the phloem decreased with a kinetic depending on the seedling: in some cases there was a smooth decline, in some a decline followed by a slight recovery and in some cases a clear-cut oscillation. The sucrose concentration was often not identical in the phloem of the individual bundles. The oscillations were larger in the phloem at the apex of the hypocotyl than in the phloem at the base of the hypocotyl. Cutting the petiole of one cotyledon led to a decrease in sucrose not only in the four bundles directly connected to the severed petiole but in all eight bundles of the hypocotyl. Cutting the petiole and dividing the vascular ring at the cotyledonary node and at the root crown did not prevent the decline of sucrose in all eight bundles. Therefore, a functional equilibration of translocated solutes between the eight bundles may occur within the 1-h measuring interval by radial diffusion through the parenchyma of the hypocotyl. Received 4 July 1997 / Accepted: 4 October 1997     

Radiosynthesis of 6’-Deoxy-6’[18F]Fluorosucrose via Automated Synthesis and Its Utility to Study In Vivo Sucrose Transport in Maize (Zea mays) Leaves

Sugars produced from photosynthesis in leaves are transported through the phloem tissues within veins and delivered to non-photosynthetic organs, such as roots, stems, flowers, and seeds, to support their growth and/or storage of carbohydrates. However, because the phloem is located internally within the veins, it is difficult to access and to study the dynamics of sugar transport. Radioactive tracers have been extensively used to study vascular transport in plants and have provided great insights into transport dynamics. To better study sucrose partitioning in vivo, a novel radioactive analog of sucrose was synthesized through a completely chemical synthesis route by substituting fluorine-18 (half-life 110 min) at the 6’ position to generate 6’-deoxy-6’[¹⁸F]fluorosucrose (¹⁸FS). This radiotracer was then used to compare sucrose transport between wild-type maize plants and mutant plants lacking the Sucrose transporter1 (Sut1) gene, which has been shown to function in sucrose phloem loading. Our results demonstrate that ¹⁸FS is transported in vivo, with the wild-type plants showing a greater rate of transport down the leaf blade than the sut1 mutant plants. A similar transport pattern was also observed for universally labeled [U-¹⁴C]sucrose ([U-¹⁴C]suc). Our findings support the proposed sucrose phloem loading function of the Sut1 gene in maize, and additionally demonstrate that the ¹⁸FS analog is a valuable, new tool that offers imaging advantages over [U-¹⁴C]suc for studying phloem transport in plants.     

Carbon assimilation in carrot cells in liquid culture

Assimilation of carbohydrates by carrot (Daucus carota L. cv Danvers) cells in liquid culture was studied to delineate the major metabolic pathways used in transformation of external carbohydrates to UDP-glucose. The cells grown on either sucrose or glucose for several years proved equally capable of utilizing each of these sugars. Sucrose was rapidly hydrolyzed extracellularly to glucose and fructose, and glucose was preferentially taken up. Uptake of fructose was slower and delayed until glucose was nearly depleted from the medium. Concentrations of cellular sugars, mainly glucose and sucrose, increased during late logarithmic phase of growth and decreased during the plateau phase. Continuous labeling of the cells with d-[¹⁴C]glucose resulted in rapid accumulation of radioactivity in glucose-6-phosphate and UDP-glucose. Because there was virtually no uptake of sucrose, UDP-glucose was likely derived from glucose-1-phosphate in a reaction catalyzed by UDP-glucose pyrophosphorylase and not directly from sucrose. Concentrations of major nucleotides and nucleotide sugars were maximal during the early logarithmic phase of growth and decreased several-fold in the stationary phase. A modified `energy charge' for adenylates calculated with the omission of AMP decreased steadily from 0.9 to 0.8 during the course of culture cycle. An analogous uracil nucleotide ratio was considerably lower (0.85) during early culture, decreased to about 0.7 for the entire logarithmic phase, and returned to initial values as cells entered stationary phase. The uracil nucleotide ratio may provide a useful index to assess the coupling between the energy available in phosphoanhydride bond in adenine nucleotides and the demand for sugar for polysaccharide synthesis through uridine diphosphate-sugar pools.     

The effects of freeze-thaw cycles and leaching on the loss of soluble carbohydrates from leaf material of two subantarctic plants

Summary Leaves and litter of two phanerogams (Acaena magellanica (Lam.) Vahl and Poa flabellata (Lam.) Hook. f.) were collected in spring on the subantarctic island of South Georgia. Leaves immersed in water lost up to 80% of their total available soluble carbohydrates after 6–8 h. The loss of K⁺ and PO ₄ ^3- followed a similar pattern to that shown by the carbohydrates. Up to 9 daily freeze/thaw cycles gave no increase in metabolite loss for senescent leaves. GLC analysis showed sucrose to be the principal leachate from Acaena. Sucrose, glucose and fructose were the main leachates from Poa. A significant proportion of the soluble carbohydrates in standing dead leaves was trehalose. The relationship of such leachates to microbial decomposition is discussed.     

Compartmentation of soluble carbohydrates,of starch and of malate in motor organs (pulvini) and other parts of Phaseolus coccineus L. leaves

Quantitative histochemistry was used to investigate the tissue-specific compartmentation of soluble carbohydrates (sucrose, glucose, fructose), starch and malate in the laminar pulvinus, leaf blade and petiole of Phaselous coccineus L. at day and night positions of diurnal leaf movement. Total carbohydrate levels measured in a series of cross sections along individual pulvini of 24-d-old plants showed only small differences between the day and night positions of the respective leaf. In contrast, the level of malate changed during diurnal leaf movement, especially in the central part of a pulvinus. The levels of glucose and fructose in the pulvinus increased towards the transition zones between the pulvinus and lamina, and pulvinus and petiole, and this trend was even more pronounced for starch. By contrast, sucrose levels were highest in the pulvinus proper. The transverse compartmentation of metabolites was studied in distinct, approx. 0.5-mm-thick tissue slices from the central part of a pulvinus. These were dissected further into up to 14 distinct subsamples (bundle, bundle sheath, motor tissues, flanks). Irrespective of the position of the leaf (day or night), the central vascular core and the surrounding bundle sheath had high levels of sucrose (up to 500 mmol-(kg DW)^–1) and low levels of glucose and fructose (below 100 mmol-(kg DW)^–1), while in the cortex the situation was reversed. In the night position the level of sucrose decreased by approx. 30% in the bundle sheath and the central vascular core but not in the other sections. We thus suggest that because of the relatively small diurnal changes in their cortical pools, soluble sugars are not involved in the osmotic processes resulting in leaf movement. In contrast, pulvini from 14-d-old plants showed an interesting diurnal change in starch and malate pools in the outermost layer of the extensor. Here starch increased at night while the malate pool was lowered nearly stoichiometrically. Inverse pool sizes were found in the day position of the respective leaves. Although less significant, the opposite diurnal variation occurred in samples taken from the flexor region. We thus were able to locate areas of different carbohydrate activities in the laminar pulvinus of P. coccineus. The central vascular core, including the bundle sheath, is involved in temporary storage of photoassimilates, and the cortical regions are responsible for osmotically driven leaf movement. The results are discussed with respect to guard-cell physiology.Abbreviations CLP cut-leaf pulvini - ILP intact-leaf pulvini This work was supported by a grant from the Deutsche Forschungsgemeinschaft.