Comparative Enzymic Studies of Sucrose Metabolism in the Taproots and Fibrous Roots of Beta vulgaris L

Comparative enzymic studies of sugar beet (Beta vulgaris L.) taproots and fibrous roots revealed differences in invertase (EC 3.2.1.26) and sucrose synthetase (EC 2.4.1.13) activity. Invertase activity of the two root forms differs with respect to specific activity, pH optimum, and enzyme solubility. Acid invertase (pH 4.5) in the taproot was restricted to the peripheral meristematic tissue which produces cells for both taproot and fibrous root growth. This finding supports the hypothesis that the enzyme regulates sucrose partitioning between the taproot and fibrous roots. A distinct alkaline invertase (pH 8.0) was detected in sucrose storage tissues of the taproot.     

Sucrose translocation and storage in the sugar beet

Several physiological processes were studied during sugar beet root development to determine the cellular events that are temporally correlated with sucrose storage. The prestorage stage was characterized by a marked increase in root fresh weight and a low sucrose to glucose ratio. Carbon derived from ¹⁴C-sucrose accumulation was partitioned into protein and structural carbohydrate fractions and their amino acid, organic acid, and hexose precursors. The immature root contained high soluble acid invertase activity (V_max 20 micromoles per hour per milligram protein; K_m 2 to 3 millimolar) which disappeared prior to sucrose storage. Sucrose storage was characterized by carbon derived from ¹⁴C-sucrose uptake being partitioned into the sucrose fraction with little evidence of further metabolism. The onset of storage was accompanied by the appearance of sucrose synthetase activity (V_max 12 micromoles per hour per milligram protein; K_m 7 millimolar). Neither sucrose phosphate synthetase nor alkaline invertase activities were detected during beet development. Intact sugar beet plants (containing a 100-gram beet) exported 70% of the translocate to the beet, greater than 90% of which was retained as sucrose with little subsequent conversions.     

Sucrose utilization and mineral nutrient uptake during hairy root growth of red beet (Beta vulgaris L.) in liquid culture

Information concerning the sugar status of plant cells is of greatimportance during all stages of the plant life cycle. The aim of this work wasto study primary carbohydrate metabolism in hairy roots of red beet. Growth ofhairy roots of red beet in vitro and changes in concentration of major nutrientsand sugar in the media were measured over a growth cycle of 16 days. We havealso determined the levels of key enzymes in the pathways of sucrose metabolism.Sucrose concentration decreased as hairy root growth proceeded while no changein glucose and fructose levels in the medium was found during the first 3 daysindicating that external sucrose is preferably taken to the cell before it ishydrolyzed by extracellular invertase. The increase in glucose and fructoselevels in the media after 5 days of culture indicates extracellular hydrolysisof sucrose which was further supported by the activity of acid invertaseobserved during that time in the culture medium. The uptake of mineral nutrientsby hairy root of red beet was monitored continuously during the culture cycle.The preferential use of NH₄ ⁺ overNO₃ ^– at the beginning of the culture andacidification of culture media were the two most notable results concerningnitrogen nutrition during hairy root growth of red beet.     

Light/Dark profiles of sucrose phosphate synthase, sucrose synthase, and Acid invertase in leaves of sugar beets

The activity of sucrose phosphate synthase, sucrose synthase, and acid invertase was monitored in 1- to 2-month-old sugar beet (Beta vulgaris L.) leaves. Sugar beet leaves achieve full laminar length in 13 days. Therefore, leaves were harvested at 2-day intervals for 15 days. Sucrose phosphate synthase activity was not detectable for 6 days in the dark-grown leaves. Once activity was measurable, sucrose phosphate synthase activity never exceeded half that observed in the light-grown leaves. After 8 days in the dark, leaves which were illuminated for 30 minutes showed no significant change in sucrose phosphate synthase activity. Leaves illuminated for 24 hours after 8 days in darkness, however, recovered sucrose phosphate synthase activity to 80% of that of normally grown leaves. Sucrose synthase and acid invertase activity in the light-grown leaves both increased for the first 7 days and then decreased as the leaves matured. In contrast, the activity of sucrose synthase oscillated throughout the growth period in the dark-grown leaves. Acid invertase activity in the dark-grown leaves seemed to be the same as the activity found in the light-grown leaves.     

The development of invertase activity in slices of the root of Beta vulgaris L. washed under aseptic conditions

1. When disks of root tissue from sugar or red beet (Beta vulgaris L.) are washed in running aerated tap water the sucrose contained in them disappears and glucose and fructose are formed. 2. Invertase activity in the disks has been measured by a polarimetric method. Freshly cut tissue has a very low activity, but a considerable increase occurs during the first 3–4 days of washing, the final activity being sufficient to hydrolyse the sucrose contained in the disk within a few hours. 3. Disks of red beet have been cut and shaken in water under aseptic conditions. Sucrose breakdown and invertase development still took place. Microbial contamination is therefore not responsible. 4. Trisaccharides that appear in sugar-beet disks during the washing process have been isolated and identified; their formation also suggests that a higher-plant invertase is acting. 5. The significance of these results is discussed in relation to protein synthesis in washed storage-tissue slices, and the occurrence of high invertase activity in growing plant cells.     

蔗糖酶活力与甜菜块根贮藏质量的关系

可溶性酸性蔗糖酶是决定甜菜块根贮藏质量的关键酶。贮藏期间其活力的提高是由于蛋白质重新合成所致。不良的贮藏条件使块根汁液pH降低,膜透性增加,这两种因素与可溶性酸性蔗糖酶活力成正相关,与贮藏质量成负相关。     

Cyanide-and rotenone-resistant respiration in mitochondria of sugar beet taproots during plant growth and development

Effects of cyanide and rotenone were examined on respiration (oxygen uptake) in mitochondria isolated from sugar beet (Beta vulgaris L.) taproots at various stages of plant growth and development. In mitochondria from growing and cool-stored taproots, the ability of cyanide-resistant, salicylhydroxamic acid-sensitive alternative oxidase (AO) to oxidize malate, succinate, and other substrates of tricarboxylic acid cycle (TCA) was low and constituted less than 10% compared to predominant activity of the cytochrome oxidase pathway during State 3 respiration. Artificial aging of storage tissue (2-day incubation of tissue sections under high humidity at 20°C) substantially activated AO, but the highest capacity (V _alt) of this pathway of mitochondrial oxidation was only observed in the presence of pyruvate and a reducing agent dithiothreitol. At the same time, mitochondria from growing taproots exhibited high rates of rotenone-resistant respiration, and these rates gradually declined during plant growth and development. The slowest rates of this respiration were observed during oxidation of NAD-dependent TCA substrates in mitochondria from dormant storage organ. The results are discussed in relation to significance of alternative electron transport pathways during growth and storage of sugar beet taproots.     

Immobilized Sclerotinia sclerotiorum invertase to produce invert sugar syrup from industrial beet molasses by product

The fungus Sclerotinia sclerotiorum produces invertase activity during cultivation on many agroindustrial residues. The molasses induced invertase was purified by DEAE-cellulose chromatography. The molecular mass of the purified enzyme was estimated at 48 kDa. Optimal temperature was determined at 60 °C and thermal stability up to 65 °C. The enzyme was stable between pH 2.0 and 8.0; optimum pH was about 5.5. Apparent K_m and V_max for sucrose were estimated to be respectively 5.8 mM and 0.11 μmol/min. The invertase was activated by β-mercaptoethanol. Free enzyme exhibited 80 % of its original activity after two month’s storage at 4 °C and 50 % after 1 week at 25 °C. In order to investigate an industrial application, the enzyme was immobilized on alginate and examined for invert sugar production by molasses hydrolysis in a continuous bioreactor. The yield of immobilized invertase was about 78 % and the activity yield was 59 %. Interestingly the immobilized enzyme hydrolyzed beet molasses consuming nearly all sucrose. It retained all of its initial activity after being used for 4 cycles and about 65 % at the sixth cycle. Regarding productivity; 20 g/l of molasses by-product gave the best invert sugar production 46.21 g/day/100 g substrate related to optimal sucrose conversion of 41.6 %.     

Effect of pollination on carbohydrate metabolism,in young fruits of Citrullus lanatus and Capsicum annuum

During a 9 day period after anthesis the concentration of reducing sugars showed a 6-fold increase in fruits of Citrullus lanatus, and a 2-fold increase in those of Capsicum annuum. These increases were associated with acid invertase, the specific activity of which was high in ovaries at anthesis and which increased 5-fold in watermelon and 1.5-fold in pepper during the same period. Sucrose synthase apparently plays only a minor role in sucrose hydrolysis. Changes in sugar concentrations and both acid invertase and sucrose synthase activities were similar in fruits developed both after pollination or hormone (NAA) treatment of ovaries. In non-pollinated ovaries of watermelon there was also an increase in invertase activity up to 6 days after anthesis which paralleled the increase in activity in seeded and parthenocarpic fruits. However, there was no increase in either reducing sugars or sucrose, indicating that sucrose is not imported into non-pollinated ovaries. Utilisation of reserve starch may help prolong the life of non-pollinated ovaries for up to one week after anthesis.     

Saturation and Utilization of Nitrate Pools in Pea and Sugar Beet Leaves

The critical periods in the saturation of pea and sugar beet leaves with nitrate absorbed by roots were discriminated. In peas, during the first 14 h, all nitrate penetrating leaf cells was concentrated in the cytosol (metabolic pool). During the second period (14–62 h), nitrate began to flow into the vacuole (storage pool), and the filling of the metabolic pool continued. Metabolic pool was saturated by the end of this period (62 h). During the third period (62–110 h), further nitrate accumulation in the cell occurred because of expanding of the storage pool. Its saturation (similarly as total cell saturation) commenced 86 h after the start of nitrate uptake. In sugar beet leaves, both metabolic and storage nitrate pools were saturated by the end of the first period (14 h), and the sizes of these pools did not change during the second period (14–86 h). When pea plants were transferred to the nitrate-free medium, nitrate efflux began from the storage pool until its complete exhausting after 3 days. In sugar beet leaves, nitrate was still present in the storage pool 4 days after plant transfer to the nitrate-free medium. In both crops, nitrate export from the storage pool was aimed at the maintenance of the optimum nitrate concentration in the metabolic pool and, thus, at the maintenance of nitrate reductase activity. A functional diversity of nitrate compartmentation in the cells of various plant species is discussed.     

Changes in Assimilated C Distribution and Soluble Acid Invertase Activity of Zinnia elegans Induced by Uniconazol, an Inhibitor of Gibberellin Biosynthesis

The physiological aspects involved in the Uniconazol-induced morphological changes in Zinnia elegans Jacq. cv Red Sun were clarified biochemically by determining the distribution of assimilated ¹³C as well as the soluble acid invertase activity. The application of Uniconazol, (E)-1-(4-chlorophenyl)-4,4-dimethyl-2-(1,2,4-triazol-1-yl)-1-pentane-3-ol, reduced the growth of stems and leaves without affecting the roots. In addition, the translocation of assimilated ¹³C from leaf to other organs was inhibited, with the stem being more restricted than the root. These changes were matched by a corresponding decrease in the specific activity of soluble acid invertase. Subsequent treatment of GA₃ counteracted these effects of Uniconazol. Moreover, the total and reducing sugar content was closely correlated with the soluble acid invertase activity in the stem. It is concluded that the reduction in invertase activity of stem is a biochemical manifestation of the retardation of stem growth induced by Uniconazol.     

Effect of Vanadium on Growth, Chemical Composition, and Metabolic Processes of Mature Sugar Beet (Beta vulgaris L.) Plants

As measured 7, 14, and 21 days after the application of 10⁻² M vanadyl sulfate solution to the foliage of 4.5-month-old sugar beet plants, significantly less growth of the leaves and an increase in the sucrose content of the storage root resulted. Accompanying these alterations were a higher rate of carbon dioxide fixation, a lower rate of respiration, and a decreased rate of nitrate reductase, glutamic-pyruvic transaminase, phosphatase, and invertase activity. The enzymes of sucrose synthesis, sucrose synthetase, sucrose phosphate synthetase and uridine diphosphate glucose-pyrophosphorylase were stimulated. The content of reducing sugar, nitrite N, amino acids and protein was less, and that of nitrate N was greater in the vanadium-treated plants. In the majority of cases the greatest magnitude of change occurred during the first 7 days following treatment. The changes in growth and chemical composition are believed to be closely related to the stimulation or inhibition of the various enzymes by vanadyl sulfate.     

Response of sugar beet plants to ultraviolet-B (280–320 nm) radiation and Cercospom leaf spot disease

Sugar beet ( Beta vulgaris L.) plants injected with Cercospora beticota Sace. as well as non-infected plants were grown under visible light with or without ultraviolet-B (UV-B, 280–320 nm) radiation for 40 days. An interaction between UV-B radiation and Cercospora leaf spot disease was observed, resulting in a large reduction in leaf chlorophyll content, dry weight of leaf laminae, petioles and storage roots. Lipid peraxidation in leaves also increased the most under the combined treatments. This was also true for ultraweak luminescence from both adaxial and abaxial leaf surfaces. However, no correlation between lipid peroxidation and ultraweak luminescence was observed. Ultraviolet-B radiation given alone appeared to have either a stimulating effect, giving an increase in dry weight of laminae and reducing lipid peroxidation, or no effect. This lack of effect was seen in the absence of change in dry weight of storage roots and chlorophyll content relative to controls. The :study demonstrated a harmful interaction between UV-B radiation and Cercospom leaf spot disease on sugar beet.     

Suppression of Rhizoctonia solani diseases of sugar beet by antagonistic and plant growth-promoting yeasts

AIMS: Isolates of Candida valida, Rhodotorula glutinis and Trichosporon asahii from the rhizosphere of sugar beet in Egypt were examined for their ability to colonize roots, to promote plant growth and to protect sugar beet from Rhizoctonia solani AG-2-2 diseases, under glasshouse conditions. METHODS AND RESULTS: Root colonization abilities of the three yeast species were tested using the root colonization plate assay and the sand-tube method. In the root colonization plate assay, C. valida and T. asahii colonized 95% of roots after 6 days, whilst Rhod. glutinis colonized 90% of roots after 8 days. Root-colonization abilities of the three yeast species tested by the sand-tube method showed that roots and soils attached to roots of sugar beet seedlings were colonized to different degrees. Population densities showed that the three yeast species were found at all depths of the rhizosphere soil adhering to taproots up to 10 cm, but population densities were significantly (P < 0.05) greater in the first 4 cm of the root system compared with other root depths. The three yeast species, applied individually or in combination, significantly (P < 0.05) promoted plant growth and reduced damping off, crown and root rots of sugar beet in glasshouse trials. The combination of the three yeasts (which were not inhibitory to each other) resulted in significantly (P < 0.05) better biocontrol of diseases and plant growth promotion than plants exposed to individual species. CONCLUSIONS: Isolates of C. valida, Rhod. glutinis and T. asahii were capable of colonizing sugar beet roots, promoting growth of sugar beet and protecting the seedlings and mature plants from R. solani diseases. This is the first successful attempt to use yeasts as biocontrol agents against R. solani which causes root diseases. SIGNIFICANCE AND IMPACT OF THE STUDY: Yeasts were shown to provide significant protection to sugar beet roots against R. solani, a serious soil-borne root pathogen. Yeasts also have the potential to be used as biological fertilizers.     

Evidence for the presence of a sucrose carrier in immature sugar beet tap roots

The objectives of this work were to determine the path of phloem unloading and if a sucrose carrier was present in young sugar beet (Beta vulgaris L.) taproots. The approach was to exploit the characteristics of the sucrose analog, 1'-fluorosucrose (F-sucrose) which is a poor substrate for acid invertase but is a substrate for sucrose synthase. Ten millimolar each of [³H]sucrose and [¹⁴C]F-sucrose were applied in a 1:1 ratio to an abraded region of an attached leaf for 6 hours. [¹⁴C]F-sucrose was translocated and accumulated in the roots at a higher rate than [³H]sucrose. This was due to [³H]sucrose hydrolysis along the translocation path. Presence of [³H]hexose and [¹⁴C]F-sucrose in the root apoplast suggested apoplastic sucrose unloading with its subsequent hydrolysis. Labeled F-sucrose uptake by root tissue discs exhibited biphasic kinetics and was inhibited by unlabeled sucrose, indicating that immature roots have the ability for carrier-mediated sucrose transport from the apoplast. Collectively, in vivo and in vitro data indicate that despite sucrose hydrolysis by the wall-bound invertase, sucrose hydrolysis is not entirely essential for sugar accumulation in this tissue.     

Role of calonyctin on free sugars in relation to starch accumulation in developing sweet potatoes

Calonyctin, a natural plant growth regulator extracted from the leaves of Calonyction aculeatum (L.) House, can promote crop growth and increase crop yield. The specific reasons for this response are unknown. This study was conducted to determine the effect of calonyctin treatment on the free sugars of sweet potato [Ipomoea batatas (L.) Lam.] as related to starch accumulation. The sweet potatoes were grown in the field in 1992, treated by foliar spray with Calonyctin concentrations of 0 (control) and 0.1 activity unit (CTSP) at 20 days after planting (DAP) at the rate of 190 liters of diluted solution/ha., and sampled periodically to determine free sugars. The response of sweet potato to calonyctin was first detected at 40 days after treatment (on 60 DAP). Data indicated that calonyctin treatment significantly increased starch synthesis in storage roots, decreased the fluctuation tendency of total sugar level during the growth period, and kept the sugar level relatively constant with a gradual rise regardless of variations in weather. The level of the reducing sugars in CTSP leaves was higher at 60 and 160 DAP and lower at 100, 120, and 140 DAP. During rainy days (100 DAP), the reducing sugars in CTSP storage roots remained at a lower level when those in controls reached high levels. The sucrose content in CTSP leaves was 40–138% greater than that in controls except at 80 and 120 DAP, and the ratio of sucrose to total nonreducing sugars remained at 100% in CTSP leaves even on rainy and cool days and above 96% in CTSP storage roots except on cool days (140 and 160 DAP), suggesting that calonyctin treatment promoted the synthesis and transfer of sucrose and supplied abundant sugar precursors for starch accumulation in storage roots.Abbreviations DAP days after planting - CTSP calonyctin-treated sweet potato with 0.1 activity unit     

Biosynthesis, translocation, and accumulation of betaine in sugar beet and its progenitors in relation to salinity

Like other halophytic chenopods, sugar beet (Beta vulgaris L.) can accumulate high betaine levels in shoots and roots. N,N,N-trimethylglycine impedes sucrose crystallization and so lowers beet quality. The objective of this research was to examine the genetic variability and physiological significance of betaine accumulation in sugar beet and its relatives. Three cultivated genotypes of B. vulgaris and two genotypes of the wild progenitor B. maritima L. were grown with and without gradual salinization (final NaCl concentration = 150 millimolar). At 6 weeks old, all five genotypes had moderately high betaine levels in shoots and roots when unsalinized (averages for all genotypes: shoots = 108 micromoles per gram dry weight; roots = 99 micromoles per gram dry weight). Salinization raised betaine levels of shoots and roots 2- to 3-fold, but did not greatly depress shoot or root growth. The genotype WB-167—an annual B. maritima type—always had approximately 40% lower betaine levels in roots than the other four genotypes, although the betaine levels in the shoots were not atypically low.

The site and pathway of betaine synthesis were investigated in young, salinized sugar beet plants by: (a) supplying 1 micromole [¹⁴C]ethanolamine to young leaf blades or to the taproot sink of intact plants; (b) supplying tracer [¹⁴C]formate to discs of leaf, hypocotyl, and taproot tissues in darkness. Conversion of both ¹⁴C precursors to betaine was active only in leaf tissue. Very little ¹⁴C appeared in the phospholipid phosphatidylcholine before betaine was heavily labeled; this was in marked contrast to the labeling patterns in salinized barley. Phosphorylcholine was a prominent early ¹⁴C metabolite of both [¹⁴C]ethanolamine and [¹⁴C]formate in all tissues of sugar beet. Betaine translocation was examined in young plants of sugar beet and WB-167 by applying tracer [methyl-¹⁴C]betaine to a young expanded leaf and determining the distribution of ¹⁴C after 3 days. In all cases, extensive ¹⁴C translocation to young leaves and taproot sink occurred; neither in the fed leaf nor in sink organs were any ¹⁴C metabolites of betaine detected.