Exogenous carbohydrate oxidation during ultraendurance exercise.

The purposes of this study were: 1) to obtain a measure of exogenous carbohydrate (CHO(Exo)) oxidation and plasma glucose kinetics during 5 h of exercise; and 2) to compare CHO(Exo) following the ingestion of a glucose solution (Glu) or a glucose + fructose solution (2:1 ratio, Glu+Fru) during ultraendurance exercise. Eight well-trained subjects exercised three times for 5 h at 58% maximum O2 consumption while ingesting either Glu or Glu+Fru (both delivering 1.5 g/min CHO) or water. The CHO used had a naturally high 13C enrichment, and five subjects received a primed continuous intravenous [6,6-2H2]glucose infusion. CHO(Exo) rates following the ingestion of Glu leveled off after 120 min and peaked at 1.24 +/- 0.04 g/min. The ingestion of Glu+Fru resulted in a significantly higher peak rate of CHO(Exo) (1.40 +/- 0.08 g/min), a faster rate of increase in CHO(Exo), and an increase in the percentage of CHO(Exo) oxidized (65-77%). However, the rate of appearance and disappearance of Glu continued to increase during exercise, with no differences between trials. These data suggest an important role for gluconeogenesis during the later stages of exercise. Following the ingestion of Glu+Fru, cadence (rpm) was maintained, and the perception of stomach fullness was reduced relative to Glu. The ingestion of Glu+Fru increases CHO(Exo) compared with the ingestion of Glu alone, potentially through the oxidation of CHO(Exo) in the liver or through the conversion to, and oxidation of, lactate.     

Exogenous utilization of 14C sugars and 14C proline as carbon source for lipid biosynthesis in the germinating Crotalaria juncea L. pollen

Abstract

Of the ¹⁴C labelled sugars (sucrose, glucose and fructose) and ¹⁴C proline, the incorporation of label into different lipid types was least from proline while sucrose was the preferred precursor over glucose or fructose. High incorporation into phosphatidyl inositol with ¹⁴C sucrose suggested that this phosphatide besides being a membrane component served possibly as the inositol storage component. High incorporation of label into phosphatidyl choline also suggested synthesis of new membranes during pollen tube growth.     

Radiometric detection of formate 14C and formaldehyde 14C oxidation in folic acid deficiency]

An ionization chamber method was used in vivo to demonstrate a delayed oxidation of [14C] formaldehyde and [14C] formate to 14CO2 in folic acid-deficient rats as compared to control rats or folic-acid-deficient rats treated by folic acid. Results obtained showed that oxidation of these two molecules required the presence of folic acid.     

Lactoperoxidase-catalyzed oxidation of [4-14C]estradiol

The conversion of [4-14C]estradiol to water-soluble products by lactoperoxidase (EC 1.11.1.7) in the presence of added or generated H2O2 was studied using albumin or tyrosine as acceptor. The enzyme was able to catalyze the oxidation and binding of estradiol to albumin even in the absence of 2,4-dichlorophenol at very low concentrations of hydrogen peroxide. Other systems in which H2O2 was replaced by oxygen and Mn2+, light-sensitized riboflavin or glutathione was also shown to be active in the conversion of estradiol to water-soluble products and the effect of inhibitors on these reactions was investigated. Possible mechanisms for the peroxidase-catalyzed formation of these estradio metabolites are discussed.     

Phyllotactic transitions in the vascular system of Populus deltoides bartr. as determined by 14C labeling

Populus deltoides seedlings progress through 2/5, 3/8, and 5/13 orders of phyllotaxis in attaining Plastochron Index 16 (PI 16). The manner in which the vascular system was reoriented during these phyllotactic transitions was determined by anatomical analysis of serial microsections, whereas the positions of the transitions were determined by ¹⁴C labeling. The midvein at the tip of leaves representing plants of different PI and leaves of different Leaf Plastochron Index (LPI) was fed ¹⁴CO₂ photosynthetically, and primordia LPI 0 through LPI-9 were dissected from the buds and analyzed for ¹⁴C. By combining the labeling data with the anatomical observations it was possible to reconstruct the vascular system of a plant of PI 16 and to locate the phyllotactic transitions. Both the anatomical and the labeling data showed a high degree of reproducibility among plants suggesting that the phyllotactic pattern to which the vascular system conforms may be programmed in the plant and transmitted acropetally through the developing leaves and procambial strands. The origin of new primordia and the concepts of orthostichy, ontogenetic helix, and Fibonacci sequence are discussed as they apply to the vascular system of P. deltoides.     

Successive 3H and 14C labeling of DNA in a stimulus-response system

Quiescent (G0) cells of the central zone region of the rat lens epithelium were recruited into the cell cycle by a wound stimulus. Cells were pulsed with labeled DNA precursor at several different times after the initiation of the DNA synthesis response to wounding and allowed to progress into the mitotic phase. Analysis of mitotic figures resulted in PLM (percentage labeled mitoses) curves that indicated a G2 duration of about 6 h. Double isotopic labeling ([3H]thymidine followed by [14C]thymidine) was utilized to demonstrate the completion of DNA synthesis in earliest responders. Cells completed DNA synthesis in less time (3-5 h) than reflected by the approximately 8-h widths of PLM curves. This discrepancy is attributed to the uptake and retention of labeled precursor by the stimulus-responsive cells while they are still in a pre-S phase condition. Based on a comparison of transit times through G2 and of labeling times to midpoint appearances of labeled mitotic figures, earlier responders do not appear to have faster rates of cell cycle progression than cells responding 2-4 h later. G2 transit time is also comparable for central zone lens cells responding to the relatively strong stimulus of wounding and for the nonperturbed cells previously studied in the germinative zone of the lens epithelium.     

Direct photoaffinity labeling of junctional sarcoplasmic reticulum with [14C]doxorubicin

Doxorubicin, an anticancer drug, induces Ca2+ release from the terminal cisternae (TC) of skeletal muscle (Zorzato, F., Salviati, G., Facchinetti, T., and Volpe, P. (1985) J. Biol. Chem. 260, 7349-7355). Long wave ultraviolet irradiation of a TC fraction with morphologically intact feet structures (Saito, A., Seiler, S., Chu, A., and Fleischer, S. (1984) J. Cell Biol. 99, 875-885) in the presence of [14C]doxorubicin, led to covalent photolabeling of two proteins that exhibited apparent Mr values of 350,000 and 170,000. Such proteins were found to be absent in a fraction of longitudinal sarcoplasmic reticulum but enriched in junctional face membranes obtained by Triton X-100 treatment of the TC fraction. Three additional proteins with Mr values of 80,000, 60,000, and 30,000 were also faintly labeled in the junctional face membrane fraction. On a molar basis the highest level of incorporation was found in the 170,000-Da protein, probably a Ca2+-binding protein (Campbell, K. P., MacLennan, D. H., and Jorgensen, A. O. (1983) J. Biol. Chem. 258, 11267-11273). A lower level of labeling was observed in the 350,000-Da protein, tentatively identified as a component of the feet structures (Cadwell, J. J. S., and Caswell, A. H. (1982) J. Cell Biol. 93, 543-550). Photolabeling of junctional TC proteins did not occur if a 10-50-fold excess cold doxorubicin was included in the assay medium, indicating that it was displaceable and specific, and if ultraviolet irradiation was omitted. Photolabeling was inhibited by caffeine or ruthenium red, i.e. by an activator and an inhibitor of Ca2+ release from TC, respectively. Furthermore, photolabeling was prevented by [ethylenebis(oxyethylenenitrilo)]tetraacetic acid suggesting that doxorubicin binding is Ca2+-dependent. Doxorubicin-binding proteins are constituents of the junctional sarcoplasmic reticulum and might be involved in modulating Ca2+ release from TC.     

In vitro labeling of beta-apolipoprotein with 3H or 14C and preliminary application to turnover studies.

3-H- or 14-C-labeled methyl groups were introduced into apolipoproteins of human and pig low density lipoproteins (LDL). 98% of the label was recovered in the apoprotein of radiomethylated LDL. Such methylated lipoprotein was compared with the corresponding unlabeled LDL with respect to its electrophoretic and immunochemical properties, and its behavior in the analytical ultracentrifuge. The data demonstrated that neither the human nor pig LDL underwent gross changes as a result of methylation. The applicability of radiomethylated pig LDL as a tracer for studying the turnover of LDL in pigs was examined. The results showed that the behavior of unscreened and screened 3-H-labeled LDL was similar. The LDL disappeared with an initial t1/2 of 1.1 hr and a later t1/2 of 30 hr. These values agreed well with those reported for radioiodinated LDL. The technique of radiomethylation of lipoprotein may afford an advantage over radioiodination as it may label peptides that do not have tyrosine.     

Estimation of rhizodeposition at field scale: upscaling of a 14C labeling study

Background and aims

Rhizodeposition of plants is the most uncertain component of the carbon (C) cycle. By existing approaches the amount of rhizodeposition can only roughly be estimated since its persistence in soil is very short compared to other organic C pools. We suggest an approach to quantify rhizodeposition at the field scale by assuming a constant ratio between rhizodeposited-C to root-C.

Methods

Maize plants were pulse-labeled with ¹⁴CO₂ under controlled conditions and the soil ¹⁴CO₂ efflux was separated into root and rhizomicrobial respiration. The latter and the ¹⁴C activity remaining in the soil corresponded to total rhizodeposition. By relating rhizodeposited-¹⁴C to root-¹⁴C a rhizodeposition-to-root ratio of 0.56 was calculated. This ratio was applied to the root biomass C measured in the field to estimate rhizodeposition under field conditions.

Results

Maize allocated 298 kg C ha^?1 as root-C and 166 kg C ha^?1 as rhizodeposited-C belowground, 50 % of which were recovered in the upper 10 cm. The fate of rhizodeposits was estimated based on the ¹⁴C data, which showed that 62 % of total rhizodeposition was mineralized within 16 days, 7 % and 0.3 % was incorporated into microbial biomass and DOC, respectively, and 31 % was recovered in the soil.

Conclusions

We conclude that the present approach allows for an improved estimation of total rhizodeposition, since it accounts not only for the fraction of rhizodeposits remaining in soil, but also for that decomposed by microorganisms and released from the soil as CO₂.     

14CO2 production is no adequate measure of [14C]fatty acid oxidation

Palmitate oxidation was comparatively assayed in various cell-free and cellular systems by 14CO2 production and by the sum of 14CO2 and 14C-labeled acid-soluble products. The 14CO2 production rate was dependent on incubation time and amount of tissue in contrast to the total oxidation rate. The 14CO2 contribution to the oxidation rate of [1-14C]palmitate varied with homogenates from 1% with rat liver to 28% with rat kidney and amounted to only 2-4% with human muscles. With cellular systems the 14CO2 contribution varied between 20% in human fibroblasts and 70% in rat muscles and myocytes. Addition of cofactors increased the oxidation rate, but decreased the 14CO2 contribution. Various conditions appeared also to influence to a different extent the 14CO2 production and the total oxidation rate with rat tissue homogenates and with rat muscle mitochondria. Incorporation of radioactivity from [1-14C]palmitate into protein was not detectable in cell-free systems and only 2-3% of the sum of 14CO2 and 14C-labeled acid-soluble products in cellular systems. Assay of 14CO2 and 14C-labeled acid-soluble products is a much more accurate and sensitive estimation of fatty acid oxidation than assay of only 14CO2.     

Exogenous sucrose utilization and starch biosynthesis among sweetpotato cultivars

Three sweetpotato cultivars were investigated for their starch content and amylose/amylopectin ratio. Ym starch contains 87.2% amylopectin and 12.8% amylose, when total starch was calculated as 100%. The Zm cultivar contains 33.6% amylopectin and 18.2% amylose, and its total starch was calculated as 51.8% of that of Ym. The Hm cultivar contains 39.1% amylopectin and 30.5% amylose, and its total starch was 69.6%. We analyzed the expression levels of starch and sucrose biosynthesis-related genes including AGPases a, b, and c; sucrose synthases I and II; starch synthase I; GBSS I; and SBEs I and II. All genes tested in this experiment were detected only in Ym, while several genes showed very faint or no expression in Zm and Hm. We also measured tissue-specific expression of these genes in whole plants of Ym. Most of the genes are expressed in the stem and roots of the plants. Expression profiles of starch synthesis-related genes of the sweetpotato leaves were investigated after supplementing the different concentrations of sucrose solution. All genes in Ym were clearly induced by sucrose, but the expression levels of some of these genes did not change in Zm and Hm. The total starch content of Ym, Zm, and Hm gradually increased over time on addition of 3%, 6%, and 9% sucrose concentrations. The greatest accumulation was observed in Ym at 48 h, and it was almost 2.24 times higher than that of the (0%) control, while Zm and Hm showed 1.76 and 1.91 times higher levels of starch, respectively. These results indicate that cooperative expression of all related genes is essential for starch biosynthesis from sucrose. This is the first report on different sucrose contents and the efficiency with which exogenous sucrose switches on gene expression of starch biosynthesis-related genes among cultivars.     

Effects of anionic polyacrylamide on maize growth: a short term 14C labeling study

Plant and Soil - Various anionic polyacrylamide polymers (PAMs) are frequently used to improve soil properties and reduce erosion. However, the effects of their application on plant growth remain...     

Reassessment of 14CO2 compartmentation and of [14C]formate oxidation in rat liver

Our previous report (Marsolais, C., Huot, S., David, F., Garneau, M., and Brunengraber, H. (1987) J. Biol. Chem. 262, 2604-2607) had concluded that a fraction of [14C]formate oxidation in liver occurs in the mitochondrion. This conclusion was based on the labeling patterns of urea and acetoacetate labeled via 14CO2 generated from [14C]formate and other [14C]substrates. We reassessed our interpretation in experiments conducted in (i) perifused mitochondria and (ii) isolated livers perfused with buffer containing [14C]formate, [14C]gluconolactone, 14CO2, or NaH13CO3, in the absence and presence of acetazolamide, an inhibitor of carbonic anhydrase. Our data show that the cytosolic pools of bicarbonate and CO2 are not in isotopic equilibrium when 14CO2 is generated in the cytosol or is supplied as NaH14CO3. We retract our earlier suggestion of a mitochondrial site of [14C]formate oxidation.