Incorporation of 14C in the cyanobacterium Synechococcus PCC 6301 following salt stress

Synechococcus PCC 6301 synthesized sucrose as a compatible solute following hyperosmotic shock induced by NaCl. Initial rates of photosynthetic ¹⁴C incorporation were reduced following salt shock. Photosynthetic rates were comparable in cells enriched for glycogen (by growth in NO ₃ ^- -deficient medium) and cells grown in NO ₃ ^- -sufficient medium in the absence of osmotic shock. Incorporation of ¹⁴C was predominantly into the NaOH fraction and the residual acidic fraction in cells grown in NO ₃ ^- -sufficient medium, whereas incorporation was predominantly into the residual acidic fraction in cells grown in NO ₃ ^- -deficient medium. Following salt stress, ¹⁴C incorporation was initially into the ethanol-soluble fraction and the majority of tracer was recovered in sucrose. Carbon-14 was detected in sucrose in cells which had been enriched for [¹⁴C]glycogen prior to salt stress, inferring that glycogen can act as a carbon source for sucrose synthesis following salt stress. Changes in the specific activity of sucrose are consistent with an initial synthesis of sucrose from glycogen followed by synthesis of sucrose using newly fixed carbon, in response to salt stress.This work was supported by the Agricultural and Food Research Council.     

Partitioning of 14C-labeled photosynthate to allelochemicals and primary metabolites in source and sink leaves of aspen: evidence for secondary metabolite turnover

Theories on allelochemical concentrations in plants are often based upon the relative carbon costs and benefits of multiple metabolic fractions. Tests of these theories often rely on measuring metabolite concentrations, but frequently overlook priorities in carbon partitioning. We conducted a pulse-labeling experiment to follow the partitioning of ¹⁴CO₂-labeled photosynthate into ten metabolic pools representing growth and maintenance (amino acids, organic acids, lipids plus pigments, protein, residue), defense (phenolic glycosides, methanol:water and acetone-soluble tannins/phenolics), and transport and storage (sugars and starch) in source and importing sink leaves of quaking aspen (Populus tremuloides). The peak period of ¹⁴C incorporation into sink leaves occurred at 24 h. Within 48 h of labeling, the specific radioactivity (dpm/mg dry leaf weight) of phenolic glycosides declined by over one-third in source and sink leaves. In addition, the specific radioactivity in the tannin/phenolic fraction decreased by 53% and 28% in source and sink leaves, respectively. On a percent recovery basis, sink leaves partitioned 1.7 times as much labeled photosynthate into phenolic glycosides as source leaves at peak ¹⁴C incorporation. In contrast, source leaves partitioned 1.8 times as much ¹⁴C-labeled photosynthate into tannins/phenolics as importing sink leaves. At the end of the 7-day chase period, sink leaves retained 18%, 52%, and 30% of imported ¹⁴C photosynthate, and labeled source leaves retained 15%, 66%, and 19% of in situ photosynthate in metabolic fractions representing transport and storage, growth and maintenance, and defense, respectively. Analyses of the phenolic fractions showed that total phenolics were twice as great and condensed tannins were 1.7 times greater in sink than in source leaves. The concentration of total phenolics and condensed tannins did not change in source and sink leaves during the 7-day chase period. Received: 31 July 1998 / Accepted: 8 February 1999     

Influence of Etherel and Gibberellic Acid on Carbon Metabolism, Growth, and Essential Oil Accumulation in Spearmint (Mentha Spicata)

Changes in growth parameters and ¹⁴CO₂ and [U-¹⁴C]-sucrose incorporation into the primary metabolic pools and essential oil were investigated in leaves and stems of M. spicata treated with etherel and gibberellic acid (GA). Compared to the control, GA and etherel treatments induced significant phenotypic changes and a decrease in chlorophyll content, CO₂ exchange rate, and stomatal conductance. Treatment with etherel led to increased total incorporation of ¹⁴CO₂ into the leaves wheras total incorporation from ¹⁴C sucrose was decreased. When ¹⁴CO₂ was fed, the incorporation into the ethanol soluble fraction, sugars, organic acids, and essential oil was significantly higher in etherel treated leaves than in the control. However, [U-¹⁴C]-sucrose feeding led to decreased label incorporation in the ethanol-soluble fraction, sugars, organic acids, and essential oils compared to the control. When ¹⁴CO₂ was fed to GA treated leaves, label incorporation in ethanol-insoluble fraction, sugars, and oils was significantly higher than in the control. In contrast, when [U-¹⁴C]-sucrose was fed the incorporation in the ethanol soluble fraction, sugars, organic acids, and oil was significantly lower than in the control. Hence the hormone treatment induces a differential utilization of precursors for oil biosynthesis and accumulation and differences in partitioning of label between leaf and stem. Etherel and GA influence the partitioning of primary photosynthetic metabolites and thus modify plant growth and essential oil accumulation. This revised version was published online in June 2006 with corrections to the Cover Date.     

Chloroplasts, Kinetin and Protein Synthesis

The effect of kinetin on protein synthesis of isolated chloroplasts was investigated by following the incorporation of ¹⁴C-leucine into isolated chloroplasts from Nicotiana tabacum. The incorporation activity varied greatly during the year, being largest in the winter and smallest in the summer. Conversely, the relative effect of kinetin on the incorporation of ¹⁴C-leucine, whether applied as a pretreatment to the leaves or directly in the incubation medium, was largest in the summer and smallest or absent altogether in the winter. Kinetin did not prolong the net incorporation period, which lasted about 40 min, but only enhanced the initial rate of the reaction. Chloroplasts extracted from leaves that had been detached for 24 or 48 h displayed very little of their original, pre-aged incorporation activity and treating the leaves with kinetin did not, essentially, prevent this loss. It was concluded that the major effect of kinetin upon chloroplasts may be related primarily to an effect upon hydration and permeability of the chloroplast and its membranes, and not to an effect directly upon its machinery for protein synthesis.     

Short- and Long-term Effects of High Temperatures (47–49 °C) on Tobacco Leaves, in. Efflux and 32P Incorporation into Phospholipids

The efflux of ³⁶C and ¹⁴C metabolites from heat-treated tissue was studied. A two-minute heat treatment at 47.5°C increased the leakiness of the membranes. The heat effect may be reversed either by treatment with kinetin or may occur spontaneously with time. The spontaneous reversal is tem—perature dependent and resembles the recovery of CO₂ fixa—tion after heat treatment. The loss of membrane integrity is reflected in the rate of incorporation of ³²P into phospho—lipids.     

Effect of substrate salinity on the ability for protein synthesis in pea roots

The effect of salinity on incorporation of amino acids into root tip protein is apparently of dual nature: in presence of salts the uptake is depressed and the normal metabolic pathways are disturbed. If the roots were grown at high salt concentration, uptake and incorporation are affected even if they are carried out in the absence of salt. NaCl and Na₂SO₄ affect uptake, incorporation, and metabolism of ¹⁴C leucine in different ways. There are also preliminary indications that in pea roots grown at different types of salinity, different proteins may be synthesized. Kinetin was found to inhibit incorporation of amino acids into non stressed and Na₂SO₄ stressed roots, but promotes uptake and incorporation of amino acids into protein in NaCl stressed tissue. It seems that there are some pronounced differences between the effects of NaCl and Na₂SO₄ salinities on the metabolism of pea root tissue.     

Effect of Zn deficiency on net photosynthetic rate, 14C partitioning,and oil accumulation in leaves of peppermint

Changes in growth, CO₂ exchange rate, and distribution of photosynthetically fixed ¹⁴CO2 into the primary photosynthetic metabolic pool (sugars, amino acids and organic acids) and essential oil accumulation were determined in leaves (leaf positions 1-6 from apex) of developing peppermint grown in a solution culture at Zn concentrations of 0 and 0.05 g m-3. There was a significant decrease in ¹⁴C incorporation in total, ethanol-soluble and ethanol-insoluble fractions in Zn deficient plants at all leaf positions. 14C incorporated in essential oil and in sugars were significantly higher in leaf pairs 1 to 3 than in leaf pairs 4 to 6. ¹⁴C incorporation into amino acids and organic acids was higher in all leaf pairs in Zn deficient plants. Statistical analysis showed a positive significant association between Zn content of leaf and ¹⁴C incorporation into ethanol-soluble fraction and sugars and a negative correlation with ¹⁴C incorporation into amino acids and organic acids. Hence the content of sugars in leaves significantly influences essential oil accumulation under Zn stress. This revised version was published online in September 2006 with corrections to the Cover Date.     

The effects of levulinic Acid and 4,6-dioxoheptanoic Acid on the metabolism of etiolated and greening barley leaves

Application of levulinic acid (LA), a competitive inhibitor of δ-aminolevulinic acid (ALA) dehydratase, to greening plant tissues causes ALA to accumulate at the expense of chlorophyll. 4,6-Dioxoheptanoic acid (DA), which has been reported to be an effective inhibitor of this enzyme in animal systems, has a similar but more powerful effect on ALA and chlorophyll metabolism in greening leaves of Hordeum vulgare L. var. Larker. Both LA and DA also inhibit the uptake of [¹⁴C]amino acids into etiolated and greening barley leaves and reduce their incorporation into protein. Treatment of etiolated and greening leaves with these compounds results in the inhibition of ¹⁴CO₂ evolution from labeled precursors, including amino and organic acids. Inhibition of ¹⁴CO₂ evolution by these compounds is more effective in greening leaves than in etiolated leaves when [4-¹⁴C]ALA or [1-¹⁴C]glutamate are employed as precursors. Both LA and DA also inhibit the uptake and increase the incorporation of ³²Pi into organophosphorus by etiolated barley leaves. These results indicate that LA and DA have more far-reaching effects upon plant metabolism than was previously believed.     

Heat shock response ofChlorella protothecoides during greening

Heterotrophically grown cells ofChlorella protothecoides were transferred to autotrophic medium and allowed to green at 25°C. The protein synthetic activity of the greening cells measured in terms of incorporation of [³5S]-methionine showed a maximum around 20 h of greening and thereafter started declining. Similarly, an analysis of densitometric tracings of the fluorographic profile of the polypeptides associated with both total cellular fraction and membrane fractions during different hours of greening revealed that maximum number of polypeptides were getting labelled around 20 h of greening. At 20 h of greening, the cells were shifted to 40°C and the effect of heat shock on protein synthesis was studied. The heat shock treatment caused a definite decrease in the incorporation of [³⁵S]-methionine into proteins. Due to heat shock, the synthesis of total soluble proteins was affected much more than that of the thylakoid membrane bound proteins. When the cells were transferred back to 25°C after a brief period of heat shock at 40°C, there was a considerable recovery in the protein synthesis and this recovery was found to be significant in the case of soluble proteins, while there was no such definite recovery in the synthesis of thylakoid membrane bound proteins.     

The fate of labelled glucose molecules in the rat adipocyte: Dependence on glucose concentration

Isolated rat adipocytes were incubated with 15 nM [3-³H]glucose or 100 nM [U-¹⁴C]glucose with or without insulin and in the absence or presence of unlabelled glucose. Following a 2 h incubation with 15 nM [3-³H]glucose, about two thirds of the cell-associated ³H-labelled metabolic products were hydrophilic largely anionic intermediates and about one third was lipids. The equivalent values were 40 and 60%, respectively, when using 100 nM [U-¹⁴C]glucose. The only ¹⁴C-labelled metabolite escaping to the incubation medium was ¹⁴CO₂, which accounted for about 15% of the rate of metabolism. Therefore, the rate of incorporation of 100 nM [U-¹⁴C]glucose into the cell-associated metabolites was quite a good measure of its net influx rate. The conversion of the two tracers to the sum of the metabolic products in cells treated with a maximally stimulating insulin concentration remained constant with glucose concentrations up to about 100 μM and then decreased progressively. The incorporation of radioactivity into the different metabolites varied markedly over the glucose concentration range 0–100 μM, presumably due to the saturation of different metabolic pools at different glucose concentrations. This variation was much less in cells not stimulated with insulin. Consequently, the maximal effect of insulin on the incorporation of the tracers into a given metabolite (e.g., labelled lipids) varied over the entire glucose concentration range. In addition, the apparent sensitivity (ED₅₀) with respect to the incorporation into a given metabolite was also dependent on the glucose concentration.     

LEUCINE OXIDATION IN BRAIN SLICES AND NERVE ENDINGS1

—It is generally believed that leucine serves primarily as a precursor for protein synthesis in the central nervous system. However, leucine is also oxidized to CO₂ in brain. The present investigation compares leucine oxidation and incorporation into protein in brain slices and synaptosomes. In brain slices from adult rats, these processes were linear for 90min and ¹⁴CO₂ production from 0·1 mm -l -[l-¹⁴C]leucine was 23 times more rapid than incorporation into protein. The rate of oxidation increased further with greater leucine concentrations. Experiments with l -[U-¹⁴C]leucine suggested that all of the carbons from leucine were oxidized to CO₂ with very little incorporation into lipid. Oxidation of leucine also occurred in synaptosomes. In slices, leucine oxidation and incorporation into protein were inhibited by removal of glucose or Na⁺, or addition of ouabain. In synaptosomes, replacement of Na⁺ by choline also reduced leucine oxidation; and this effect did not appear to be due to inhibition of leucine transport. The rate of leucine oxidation did not change in brain slices prepared from fasted animals. Fasting, however, reduced the incorporation of leucine into protein in brain slices prepared from young but not from adult rats. These findings indicate that oxidation is the major metabolic fate of leucine in brain of fed and fasted animals.     

Relationship between leaf development and primary photosynthetic products in the C4 plant Portulaca oleracea L.

Summary The photosynthetic products of Portulaca oleracea differ greatly depending on leaf age and length of exposure to ¹⁴CO₂. Mature leaves of P. oleracea fix ¹⁴CO₂ primarily into organic and amino acids during a 10-s exposure period. Less than 2% of the ¹⁴CO₂ fixed appears in phosphorylated compounds. In contrast, incorporation into amino acids can account for over 60% of the total ¹⁴CO₂ fixed by young leaves in an equal time period, and incorporation into alanine alone can account for up to one half of this amount. Senescent leaves display a quantitative shift of primary products toward phosphorylated compounds with a concomitant reduction of the label residing in malate and asparate. About 8 times more phosphoglyceric acid is produced in senescent leaves than in mature leaves. The aspartate/ malate ratio is not constant and depends on the length of time the leaves are exposed to ¹⁴CO₂ and the age of the leaves under study. It appears as if the stage of leaf development is one of the most important factors determining the operation of a particular enzyme system in C₄ plants.     

蕾期土壤盐度降低后棉花叶片的生理功能恢复

试验于2011—2012年在江苏南京江苏省农业科学院经济作物研究所试验田进行,采用盆栽方法,以鲁棉研37号和苏棉22号为供试材料,设置土壤盐度降低试验(初始土壤含盐量为0.2%,棉花进入二叶期后每7d加入混合盐1次,每次增加0.1%,使土壤含盐量逐渐达到0.5%,蕾期进行盐度降低处理,使土壤含盐量降低到0.2%左右),研究蕾期土壤盐度降低后棉花叶片的生理代谢动态特征。结果表明:土壤盐度降低后,棉花叶片叶绿素(Chl)、类胡萝卜素(Car)含量和Chl/Car升高;净光合速率和气孔导度升高,且分别在土壤盐度降低后第14天和7天接近于低盐对照;土壤盐度降低后棉花叶片超氧化物歧化酶(SOD)和过氧化物酶(POD)活性升高,过氧化氢酶(CAT)活性和丙二醛(MDA)含量降低,MDA含量在土壤盐度降低后第14天接近于低盐对照;土壤盐度降低后棉花叶片中可溶性糖、游离氨基酸和脯氨酸含量降低,且接近于低盐对照。上述结果表明土壤盐度降低后,棉花叶片生理功能逐渐恢复,进而实现棉花生长发育的恢复补偿。棉花叶片生理功能在土壤盐度降低后的恢复能力存在品种间差异,鲁棉研37号较苏棉22号叶片生理功能表现出更强的恢复能力。     

Maintenance of High Photosynthetic Rates in Mesophyll Cells Isolated from Papaver somniferum

The establishment and maintenance of high rates of photosynthetic CO₂ incorporation in mesophyll cells of Papaver somniferum (opium poppy) depend on a regime of dark and light periods immediately following isolation, as well as carefully adjusted conditions of isolation. Analysis of the incorporation pattern of ¹⁴CO₂ by the isolated cells indicates an initial “stress-response” period of approximately 20 hours characterized by increased respiratory-type metabolism and diminished photosynthesis. Under the favorable regime, this period is followed by rapid recovery and the reinstatement of a metabolic state strikingly similar to that of intact leaves in which the initial rate of CO₂ incorporation is between 110 and 175 μmoles CO₂ fixed per mg chlorophyll per hour. The photosynthetic viability of these cells can be maintained for up to 80 hours.     

Glutamic Acid metabolism and the photorespiratory nitrogen cycle in wheat leaves: metabolic consequences of elevated ammonia concentrations and of blocking ammonia assimilation

The effects of methionine sulfoximine and ammonium chloride on [¹⁴C] glutamate metabolism in excised leaves of Triticum aestivum were investigated. Glutamine was the principal product derived from [U¹⁴C]glutamate in the light and in the absence of inhibitor or NH₄Cl. Other amino acids, organic acids, sugars, sugar phosphates, and CO₂ became slightly radioactive. Ammonium chloride (10 mm) increased formation of [¹⁴C] glutamine, aspartate, citrate, and malate but decreased incorporation into 2-oxoglutarate, alanine, and ¹⁴CO₂. Methionine sulfoximine (1 mm) suppressed glutamine synthesis, caused NH₃ to accumulate, increased metabolism of the added radioactive glutamate, decreased tissue levels of glutamate, and decreased incorporation of radioactivity into other amino acids. Methionine sulfoximine also caused most of the ¹⁴C from [U-¹⁴C]glutamate to be incorporated into malate and succinate, whereas most of the ¹⁴C from [1-¹⁴C]glutamate was metabolized to CO₂ and sugar phosphates. Thus, formation of radioactive organic acids in the presence of methionine sulfoximine does not take place indirectly through “dark” fixation of CO₂ released by degradation of glutamate when ammonia assimilation is blocked. When illuminated leaves supplied with [U-¹⁴C] glutamate without inhibitor or NH₄Cl were transferred to darkness, there was increased metabolism of the glutamate to glutamine, aspartate, succinate, malate, and ¹⁴CO₂. Darkening had little effect on the labeling pattern in leaves treated with methionine sulfoximine.     

Effects of light on terpene hydrocarbon synthesis in Pinus pinaster

The biosynthesis of terpene hydrocarbons has been investigated in maritime pine (Pinus pinaster Ait.) seedling primary leaves under light and darkness and with different precursors. Impossible in darkness, the synthesis of monoterpenes (mainly α- and β-pinene) is strongly activated by light. Only ¹⁴C-carbonate and ¹⁴C-acetate can be incorporated into monoterpenes. Activation by light is comparatively much more effective for seedling leaves previously cultivated under short days than in leaves from seedlings given long days. The spectral bands which are efficient for the synthesis of monoterpenes are located around 480 and 685 nm with ¹⁴C-carbonate and 480 and 630 nm with l-¹⁴C-acetate. Furthermore, this light activation does not occur if leaf pieces instead of whole leaves are used for the incorporation experiments. When 2-¹⁴C-mevalonic acid and 1-¹⁴C-isopentenyl pyrosphosphate are applied as precursors, no radioactivity is recorded in monoterpene hydrocarbons even after light exposures. In contrast, sesquiterpene hydrocarbons (caryophyllene and humulene) are easily synthesized under light or darkness in intact or fragmented leaves from the different precursors of photosynthetic or exogenous origin. From these results the compartmentalization in the synthesis of C₁₀ and C₁₅ hydrocarbons appears clear. There is a metabolic cooperation between the photosynthetic tissues and the specific site of elaboration of C₁₀ hydrocarbons, which site is located in the parts where the epithelial cells of resin ducts are functional. The synthesis of sesquiterpene hydrocarbons takes place in the whole leaf without activation by light.     

EBR预处理对盐胁迫下葡萄幼苗叶片抗氧化物质及酶活性的影响

以2年生葡萄(Vitis vinifera L.)酿酒品种赤霞珠扦插苗为材料,在水培条件下,分别用0、0.05、0.10和0.20mg/L 24-表油菜素内酯(EBR)预处理幼苗,然后进行50mmol/L NaCl胁迫,分别在胁迫6d和12d测定幼苗叶片中超氧阴离子(O_2~)、丙二醛(MDA)、抗氧化物质含量以及相关酶活性,探讨EBR预处理对葡萄幼苗耐盐性的影响。结果表明:与单独盐胁迫处理相比,不同浓度的EBR预处理使盐胁迫葡萄幼苗叶片O_2~和MDA含量显著降低,同时使其抗氧化物质抗坏血酸(AsA)、脱氢抗坏血酸(DHA)、还原型谷胱甘肽(GSH)和氧化型谷胱甘肽(GSSG)含量以及抗坏血酸过氧化物酶(APX)、谷胱甘肽还原酶(GR)、超氧化物歧化酶(SOD)活性显著升高;其中,0.10mg/L EBR预处理的表现最佳,在盐胁迫12d时,其葡萄叶O_2~和MDA含量比单独盐胁迫处理分别显著降低30.5%和22.0%,其叶片相应AsA和GSH的含量较单独盐胁迫处理分别显著提高82.8%和27.9%,且GR、APX和SOD活性分别显著提高7.2%、8.5%和24.0%。研究发现,在盐胁迫条件下,适宜浓度的外源BRs预处理能够显著降低葡萄叶片中活性氧含量,提高抗氧化物质含量和抗氧化酶活性,以促进AsA-GSH循环的快速有效运转,有效减轻植株的过氧化伤害,缓解盐胁迫对葡萄幼苗的伤害,提高葡萄的耐盐性。     

Dark Carbon Dioxide Fixation under Aerobic and Anaerobic Conditions in Maize Leaves after Preillumination in the Absence of Oxygen: Ribulose 1,5-Bisphosphate Can Serve as a Primary Acceptor of Carbon Dioxide

When dark ¹⁴CO₂ fixation in maize leaves was carried out under anaerobic conditions after preillumination in the absence of O₂, the ¹⁴C incorporation in aspartic acid was transient; its maximum level was very low compared with that of malic acid. The addition of 5% O₂ during the dark fixation period increased the total uptake of ¹⁴CO₂ and the ¹⁴C incorporation into aspartic acid.     

Effect of Tobacco Mosaic Virus Infection on Amino Acid Incorporation into Isolated Mitochondria from Tobacco Leaves

Mitochondria isolated from tobacco leaves incorporated ¹⁴C-leucine into the protein and the rate was enhanced by tobacco mosaic virus (TMV) infection as compared with noninfected level. In vitro amino acid incorporation by mitochondria required adenosine triphosphate (ATP), Mg²⁺, and KC1 and the energy sources from oxidative phosphorylation as well as from ATP-generating system. This incorporation was inhibited by ribonuclease (RNase), deoxyribonuclease (DNase), actinomycin D, mitomycin C, puromycin, and chloramphenicol added in the reaction medium. The pretreatment of the mitochondria with DNase and actinomycin D reduced the rate of incorporation. The mitochondria incorporated ³H-guanosine triphosphate (GTP) and this activity was blocked by actinomycin D. The presence in this system of 15,000 g supernatant cell sap fraction or bacterial contamination was carefully checked obtaining a negative result. The reaction product into which ^l4C-amino acids incorporated was solubilized by trypsin. The nature of the amino acid incorporating activity of isolated mitochondria obtained from TMV-infected tobacco leaves is discussed.     

Biosynthesis of gallic and ellagic acids with14C-labeled compounds inAcer andRhus leaves

The biosynthetic pathway for gallic and ellagic acids in young, mature and autumn leaves ofAcer buergerianum andRhus succedanea was examined by tracer experiments, and also by isotope competition, withd-shikimic acid-¹⁴C,l-phenylalanine-U-¹⁴C,l-phenyllactic acid-U-¹⁴C, gallic acid-G-¹⁴C and their unlabeled compounds. In young leaves of both plants, the incorporation rate of labeled shikimic acid into gallic acid was significantly higher than that of labeled phenylalanine, whereas in the mature and autumn leaves the latter was a good precursor rather than the former for the gallic acid biosynthesis. Therefore, two pathways for gallic acid formation, through β-oxidation of phenylpropanoid and through dehydrogenation of shikimic acid, could be operating inAcer andRhus leaves, and the preferential pathway is altered by leaf age. In both plants, the incorporation rate of labeled phenyllactic acid during a 24 hr metabolic period was almost the same as that of labeled phenylalanine. The incorporation ofd-skikimic acid-G-¹⁴C,l-phenylalanine-U-¹⁴C andl-phenyllactic acid-U-¹⁴C into ellagic acid was very similar to the case of the radioactive gallic acid formation. Furthermore, regardless of the presence of unlabeled shikimic acid and/or phenylalanine, incorporation of the radioactivity of labeled gallic acid into ellagic acid occurred at a very high rate, suggesting the reciprocal radical reaction of gallic acid for the ellagic acid formation. The incorporation of labeled compounds into ellagitanins was also examined and their biosynthesis discussed further.