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1.
《Journal of Fermentation and Bioengineering》1994,77(6):626-629
A new biosynthetic pathway, which can produce both vitamin B12 and large amounts of porphyrins from isopropanol, was identified in Arthrobacter hyalinus using carbon-13 stable isotope tracer techniques and carbon-13 nuclear magnetic resonance (13C-NMR) spectroscopy. Studies on the incorporation of [2-13C]isopropanol, [1- or 2-13C]sodium acetate, l-[1-13C]glutamate, and [1-, 2-, 3-, 4-, 5-13C]5-aminolevulinic acid into uroporphyrinogen III showed that isopropanol was metabolized into uroporphyrinogen III through acetyl CoA and that 5-aminolevulinic acid was produced from l-glutamic acid and not via Shemin's pathway. 相似文献
2.
John K. Meissen David A. Pirman Min Wan Emily Miller Aditi Jatkar Russell Miller Rick C. Steenwyk Matthew Blatnik 《Analytical biochemistry》2016
Metabolite stable isotope tracing is a powerful bioanalytical strategy that has the potential to unravel phenotypic markers of early pharmaceutical efficacy by monitoring enzymatic incorporation of carbon-13 atoms into targeted pathways over time. The practice of probing biological systems with carbon-13 labeled molecules using broad MS-based screens has been utilized for many years in academic laboratories but has had limited application in the pharmaceutical R&D environment. The goal of this work was to establish a LCMS analytical workflow that was capable of monitoring carbon-13 isotope changes in glycolysis, the TCA and urea cycles, and non-essential amino acid metabolism. This work applies a standardized protein precipitation with 80% cold methanol and two distinct reverse-phase ion-pair liquid chromatography methods coupled to either a positive- or negative-ion mode high-resolution accurate mass spectrometry screening method. The data herein combines thousands of single-point peak integrations into a novel metabolite network map as a visualization aid to probe and monitor stable isotope incorporation in murine hepatocytes using uniformly labeled 13C6 glucose, 13C3 lactate, and 13C5 glutamine. This work also demonstrates that nitrogen metabolism may have a large influence on the TCA cycle and gluconeogenic carbon fluxes in hepatocyte cell culture. 相似文献
3.
《Insect Biochemistry》1990,20(2):149-156
The precursors and directionality of synthesis of the methyl branched cuticular hydrocarbons and the female contact sex pheromone, 3,11-dimethyl-2-nonacosanone, of the German cockroach, Blattella germanica, were investigated by radiotracer and carbon-13 NMR techniques. The amino acids [G-3H]valine, [4,5-3H]isoleucine and [3,4-14C2]methionine labeled the hydrocarbon fraction in a manner indicating that the carbon skeletons of all three amino acids serve as the methyl branch group donor. The incorporation of [1,4-14C2]- and [2,3-14C2]succinates into the hydrocarbon and acylglycerol/polar lipid fractions indicated that succinate also served as a precursor to methylmalonyl-CoA. Carbon-13 NMR analyses showed that [1-13C]propionate labeled the carbon adjacent to the tertiary carbon, and, for the 3,x-dimethylalkanes, that carbon-4 and not carbon-2 was enriched. [1-13C]Acetate labeled carbon-2 of these hydrocarbons. This indicates that the methyl branching groups of the 3,x-dimethylalkanes were inserted early in the chain elongation process. [3,4,5-13C3]Valine labeled the methyl, tertiary and carbon adjacent to the tertiary carbon of the methyl branched alkanes. Thus, the methyl branched hydrocarbon was formed by the insertion of methylmalonyl units derived from propionate, isoleucine, valine, methionine and succinate early in chain elongation. 相似文献
4.
Summary. 6-N-carboxymethyllysine (CML), generated by the glycation and/or oxidation of lysine residues, has been measured in biological
materials and food products using techniques such as ELISA, HPLC with fluorescence detection and mass spectrometry methods.
Only limited information has been reported regarding the preparation of standards labeled with either deuterium, 13C or 15N atoms to be used as internal standards. In the present paper, a synthesis of carbon-13 labeled CML is described using l,2-13C2-glyoxylic acid and 2-N-acetyllysine as starting materials. The resulting labeled 2-N-acetyl-CML was purified by HPLC-UV as a dibutyl ester. After a deprotection step, the yield was evaluated to be 53% when
the reaction was conducted 17 h at 37°C. CML was extensively studied by 1H- and 13C-NMR and the fragments observed in the collision induced dissociation (CID) spectrum were also assigned. Finally, the standards
of CML and carbon-13 labeled CML were accurately quantified based on 1H-NMR and tandem MS using lysine as an internal reference. 相似文献
5.
Experiments were conducted to examine the development of photosynthetic carbon metabolism in Peperomia scandens, a tropical epiphyte. Leaves were sampled during a 10-day period when they were between 30 to 165 days old. P. scandens exhibits a C3 to CAM-cycling to CAM shift during maturation with the magnitude of CAM increasing with age. Initially, during both day and night, no significant CO2 uptake or diurnal acid flux was evident. C3 gas exchange was detected at 41 days of age with a gradual shift towards CAM gas exchange maximized thereafter. An acidity flux of 130 to 150 microequivalents per gram fresh weight was evident by 41 days. Between 40 and 90 days, the leaves shifted their CO2 uptake pattern from a daytime to a nighttime peak. After 90 days, the leaves remained in CAM. The δ13C values became progressively less negative as the leaves matured. In the 30-day-old leaves, the δ13C value was −21.1% while in the 165-day-old leaves the δ13C value was −18.3%. The time-dependent shift from C3 to CAM-cycling to CAM in P. scandens does not appear to result from changes in water, light, or temperature regimes since these variables were constant for all leaves sampled. 相似文献
6.
《Life sciences》1994,54(26):PL483-PL490
Ro 24-4736, a new platelet activating factor antagonist, is currently under preclinical and clinical development. The tissue distribution of the 14C-label in male rats following a single intravenous dose of 1.0 mg/kg of 14C-Ro 24-4736 indicated appreciable uptake by the liver, kidney, heart and gastrointestinal tract. Peak plasma and tissue concentrations were seen at 5 minutes after dosing except for the small intestine (4 hrs) and abdominal fat, stomach and large intestine (4 hrs). Thereafter, the 14C-label rapidly declined in all tissues. At 48 hours, only 3.5% of the dose was present in the tissues, and 6.1% in the lumen of the gastrointestinal tracts. The excretion of 14C was essentially completed; 94% of the administered 14C was excreted in the feces and 4.0% in the urine. Overall recoveries of the administered 14C label ranged from 96 to 116%. The purified major 14C-labelled component in the fecal extracts yielded essentially the same NMR spectrum as authentic Ro 24-4736 which accounted for 11% of the dose. In vitro incubations of Ro 24-4736 with rat liver 9S supernatant in an NADPH generating system produced two metabolites. NMR spectra indicated that one metabolite was hydroxylated at carbon-1 while the other one contained a hydroxyl at carbon-10 of the parent molecule. Interestingly, the sites of hydroxylation were at carbons C1, and C10 bearing the protons guarding the bay area of the phenanthrenoid ring, rather than carbons of the phenyl-methyl-thienotriazolodiazepine moiety. 相似文献
7.
[1-14C]acetate and [2-14C]acetate were incorporated into the β-diketones of barley spike epicuticular wax via the peduncle. Utilizing column chromatography with dry copper acetate, the β-diketones were isolated and the labeling pattern in the hentriacontan-14, 16-dione determined after its degradation. A modified iodoform procedure was used to give myristic and palmitic acids. Radio-gas chromatography was then performed on the products of chemical α-oxidation of the separated fatty acids. This procedure, in effect, gave the specific activity of every carbon atom of hentriacontan-14,16-dione except carbon-1 to carbon-5 (from myristic acid) and carbon-27 to carbon-31 (from palmitic acid) for each labeled substrate. The specific activity of carbon-15 was determined by an indirect method. On the basis of these data it is suggested that the hentriacontan-14,16-dione is synthesized from the carbon-31 end of the molecule by elongation as follows. C2 units are added, perhaps to a mixture of short chain precursors, to give a chain with 12 carbon atoms. This chain is then elongated to one with 16 carbon atoms so that the four added carbon atoms are uniformly labeled. Following this, the chain with 16 carbon atoms is elongated with C2 units to give the complete molecule. Possibly some change in mechanism occurs in this last elongation process when the chain is 22 carbon atoms long. Barley spike wax β-diketones contain about 2% nonacosan-13, 15-dione which seems to be synthesized in an analogous manner. 相似文献
8.
Autotrophy in maize husk leaves : evaluation using natural abundance of stable isotopes 总被引:2,自引:0,他引:2
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The natural abundance of carbon and hydrogen isotopic composition, expressed as a δ13C value of plant dry matter and cellulose in the hypsophylls (husk leaves) of maize (Zea mays L.) was measured and compared with that of leaves and cobs. The δ13C values of outer hypsophylls were usually 2 to 3%‰ more negative than leaves or other tissues, and became more negative with increasing chlorophyll content, indicating significant local C3 pathway fixation of CO2 in the outer hypsophylls. The δD values indicated a significant part of hypsophyll cellulose was derived from heterotrophic sources (sucrose from C4 photosynthesis in other tissues). Isotopic mass balance calculations allowed quantitative estimation of these carbon sources and, in the samples examined, about 16% of hypsophyll cellulose was derived from local C3 photosynthesis, about 62% from local C4 photosynthesis, and about 22% from sucrose imported from other leaves. 相似文献
9.
Victoria J. Stokes Michael D. Morecroft James I. L. Morison 《Trees - Structure and Function》2010,24(2):297-306
The seasonal trends in water use efficiency of sun and shade leaves of mature oak (Quercus robur) and sycamore (Acer pseudoplatanus) trees were assessed in the upper canopy of an English woodland. Intrinsic water use efficiency (net CO2 assimilation rate/leaf conductance, A/g) was measured by gas exchange and inferred from C isotope discrimination (δ13C) methods. Shade leaves had consistently lower δ13C than sun leaves (by 1–2‰), the difference being larger in sycamore. Buds had distinct sun and shade isotopic signatures
before bud break and received an influx of 13C-rich C before becoming net autotrophs. After leaf full expansion, δ13C declined by 1–2‰ gradually through the season, emphasising the importance of imported carbon in the interpretation of leaf
δ13C values in perennial species. There was no significant difference between the two species in the value of intrinsic water
use efficiency for either sun or shade leaves. For sun leaves, season-long A/g calculated from δ13C (72–78 μmol CO2 [mol H2O]−1) was 10–16% higher than that obtained from gas exchange and in situ estimates of leaf boundary layer conductance. For shade
leaves, the gas exchange–derived values were low, only 10–18% of the δ13C-derived values. This is ascribed to difficulties in obtaining a comprehensive sample of gas exchange measurements in the
rapidly changing light environment. 相似文献
10.
Effect of Varying CO(2) Partial Pressure on Photosynthesis and on Carbon Isotope Composition of Carbon-4 of Malate from the Crassulacean Acid Metabolism Plant Kalanchoë daigremontiana Hamet et Perr
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Intact leaves of Kalanchoë daigremontiana were exposed to CO2 partial pressures of 100, 300, and 1000 microbars. Malic acid was extracted, purified, and degraded in order to obtain isotopic composition of carbon-1 and carbon-4. From these data, it is possible to calculate the carbon isotope composition of newly fixed carbon in malate. In all three treatments, the isotopic composition of newly introduced carbon is the same as that of the CO2 source and is independent of CO2 partial pressures over the range tested. Comparison with numerical models described previously (O'Leary 1981 Phytochemistry 20: 553-567) indicates that we would expect carbon 4 of malate to be 4‰ more negative than source CO2 if diffusion is totally limiting or 7‰ more positive than source CO2 if carboxylation is totally limiting. Our results demonstrate that stomatal aperture adjusts to changing CO2 partial pressures and maintains the ratio of diffusion resistance to carboxylation resistance approximately constant. In this study, carboxylation and diffusion resistances balance so that essentially no fractionation occurs during malate synthesis. Gas exchange studies of the same leaves from which malate was extracted show that the extent of malate synthesis over the whole night is nearly independent of CO2 partial pressure, although there are small variations in CO2 uptake rate. Both the gas exchange and the isotope studies indicate that the ratio of external to internal CO2 partial pressure is the same in all three treatments. Inasmuch as a constant ratio will result in constant isotope fractionation, this observation may explain why plants in general have fairly invariable 13C contents, despite growing under a variety of environmental conditions. 相似文献
11.
We assessed the effects of doubling atmospheric CO2 concentration, [CO2], on C and N allocation within pedunculate oak plants (Quercus robur L.) grown in containers under optimal water supply. A short-term dual 13CO2 and 15NO3? labelling experiment was carried out when the plants had formed their third growing flush. The 22-week exposure to 700 μl l?1 [CO2] stimulated plant growth and biomass accumulation (+53% as compared with the 350 μl l?1 [CO2] treatment) but decreased the root/shoot biomass ratio (-23%) and specific leaf area (-18%). Moreover, there was an increase in net CO2 assimilation rate (+37% on a leaf dry weight basis; +71% on a leaf area basis), and a decrease in both above- and below-ground CO2 respiration rates (-32 and -26%, respectively, on a dry mass basis) under elevated [CO2]. 13C acquisition, expressed on a plant mass basis or on a plant leaf area basis, was also markedly stimulated under elevated [CO2] both after the 12-h 13CO2 pulse phase and after the 60-h chase phase. Plant N content was increased under elevated CO2 (+36%), but not enough to compensate for the increase in plant C content (+53%). Thus, the plant C/N ratio was increased (+13%) and plant N concentration was decreased (-11%). There was no effect of elevated [CO2] on fine root-specific 15N uptake (amount of recently assimilated 15N per unit fine root dry mass), suggesting that modifications of plant N pools were merely linked to root size and not to root function. N concentration was decreased in the leaves of the first and second growing flushes and in the coarse roots, whereas it was unaffected by [CO2] in the stem and in the actively growing organs (fine roots and leaves of the third growth flush). Furthermore, leaf N content per unit area was unaffected by [CO2]. These results are consistent with the short-term optimization of N distribution within the plants with respect to growth and photosynthesis. Such an optimization might be achieved at the expense of the N pools in storage compartments (coarse roots, leaves of the first and second growth flushes). After the 60-h 13C chase phase, leaves of the first and second growth flushes were almost completely depleted in recent 13C under ambient [CO2], whereas these leaves retained important amounts of recently assimilated 13C (carbohydrate reserves?) under elevated [CO2]. 相似文献
12.
CO2 transfer conductance, leaf structure and carbon isotope composition of Polygonum cuspidatum leaves from low and high altitudes 总被引:5,自引:4,他引:1
H. Kogami Y. T. Hanba T. Kibe I. Terashima & T. Masuzawa 《Plant, cell & environment》2001,24(5):529-538
Anatomy and some physiological characteristics of the leaves in Polygonum cuspidatum Sieb. et Zucc., a dioecious clonal herb, were compared between two populations, one from a lowland in Shizuoka City (10 m above sea level), and another from a highland on Mt. Fuji (2500 m above sea level). Leaf mass per area (LMA) of the highland plants was about twice that of the lowland plants. The greater leaf thickness, thicker mesophyll cell walls and higher mesophyll cell density in the highland leaves contributed to the larger LMA. Although mesophyll area exposed to intercellular airspaces was greater in the highland leaves than in the lowland leaves by 30%, the surface area of chloroplasts facing intercellular airspaces was similar between these leaves. CO2 transfer conductance inside the leaf (gi) of the highland leaves (0·75 μmol m?2 s?1 Pa?1) is the lowest recorded for herbaceous plants and was only 40% of that in the lowland leaves. On the other hand, the difference in stomatal conductance was small. δ13C values in the leaf dry matter were greater in the highland leaves by 4‰. These data and the estimation of CO2 partial pressures in the intercellular air spaces and in the chloroplast suggested that the greater dry matter δ13C in the highland leaves, indicative of lower long‐term ratio of the chloroplast stroma to the ambient CO2 partial pressures, would be mainly attributed to their lower gi. 相似文献
13.
Resuspension cultures of Gibberella fujikuroi, strain GF-1a, were shown to metabolise potassium [3′-13C] mevalonate to 13C-enriched C19-gibberellins, plus 13CO2 (derived from the loss of carbon-20). The formation of [13C]-gibberellins could be observed in vivo using 13C NMR; however that of 13CO2 could not. In contrast, removal of the mycelium and concentration of the filtrate at pH 12 enabled the 13CO2 produced to be observed using 13C NMR. During incubations of H14CO2Na with this fungus, complete conversion to other radioactive products was observed, and the significance of these results in the light of previous work is discussed. 相似文献
14.
Biogenic isoprene substantially affects atmospheric chemistry, but it is not known how or why many plants, especially trees, make isoprene. We fed 13CO2 to leaves of Quercus rubra and monitored the incorporation of 13C into isoprene by mass spectrometry. After feeding 13CO2 for 9 min we found all possible labelling patterns from completely unlabelled to fully labelled isoprene. By 18 min, 84% of the carbon atoms in isoprene were 13C. Labelling of the last 20% of the carbon atoms was much slower than labelling of the first 80%. The rate of labelling of isoprene was similar to that reported for phosphoglyceric acid indicating that there is a close linkage between the carbon source for isoprene synthesis and the photosynthetic carbon reduction pathway. 相似文献
15.
Distribution of carbon-14 assimilated by wheat awns 总被引:1,自引:0,他引:1
L. B. OLUGBEMI 《The Annals of applied biology》1978,90(1):111-114
The pattern of distribution of carbon assimilated by awns was investigated in two lines of Triticum aestivum. Single awns on basal florets of spikelets in the central part of the ear were dosed with 14CO2. Five days after dosing, 99% of the carbon-14 recovered was in the spikelet bearing the awn. Of the carbon-14 exported from the treated awn 57% went to the grain of the first floret, 1% to the second, 28% to the third and 7% to the fourth. 相似文献
16.
Application of tree leaves (C3 plants) on maize (Zea mays L.) (C4 plant) fields is an agroforestry management technology to restore or maintain soil fertility. The rate at which the tree
leaves decompose is crucial for the nutrient supply to the crop. We studied the in situ decomposition of Sesbania sesban (L.) Merr. leaves or C3 sugar for 4 – 8 days after application to a maize field in Kenya. By using the difference of around 10‰ in natural abundance
of 13C between the endogenous soil C (mainly C4) and the applied C (C3), we could calculate the contributions of the two C sources to soil respiration. The δ13C value of the basal respiration was from –15.9 to –16.7‰. The microbial response to the additions of leaves and sugar to
this tropical soil was immediate. Application of sesbania leaves gave an initial peak in respiration rates that lasted from
one to less than 6 days, after which it levelled off and remained about 2 – 3 times higher (230–270 mg C m-2 h-1) than the control respiration rates throughout the rest of the experiment (5 – 8 days). In the sugar treatment, there was
no initial peak in respiration rate. The respiration rate was 170 mg C m-2 h-1 after 4 days. At the end of the experiments, after 4–8 days, as much as 14–17% of the added C had been respired and about
60% of the total respiration was from the added sesbania leaves or C3 sugar. This non-destructive method allows repeated measurements of the actual rate of C mineralisation and facilitates decomposition
studies with high temporal resolution in the field.
This revised version was published online in August 2006 with corrections to the Cover Date. 相似文献
17.
《Phytochemistry》2012
In plants, jasmonic acid (JA) and its derivatives are thought to be involved in mobile forms of defense against biotic and abiotic stresses. In this study, the distal transport of JA-isoleucine (JA-Ile) that is synthesized de novo in response to leaf wounding in tomato (Solanum lycopersicum) plants was investigated. JA-[13C6]Ile was recovered in distal untreated leaves after wounded leaves were treated with [13C6]Ile. However, as [13C6]Ile was also recovered in the distal untreated leaves, whether JA-Ile was synthesized in the wounded or in the untreated leaves was unclear. Hence, stem exudates were analyzed to obtain more detailed information. When [13C6]Ile and [2H6]JA were applied separately into the wounds on two different leaves, JA-[13C6]Ile and [2H6]JA-Ile were detected in the stem exudates but [2H6]JA–[13C6]Ile was not, indicating that JA was conjugated with Ile in the wounded leaf and that the resulting JA-Ile was then transported into systemic tissues. The [2H3]JA-Ile that was applied exogenously to the wounded tissues reached distal untreated leaves within 10 min. Additionally, applying [2H3]JA-Ile to the wounded leaves at concentrations of 10 and 60 nmol/two leaves induced the accumulation of PIN II, LAP A, and JAZ3 mRNA in the distal untreated leaves of the spr2 mutant S. lycopersicum plants. These results demonstrate the transportation of de novo synthesized JA-Ile and suggest that JA-Ile may be a mobile signal. 相似文献
18.
Stable carbon isotope composition varies markedly between sun and shade leaves, with sun leaves being invariably more enriched (i.e., they contain more13C). Several hypotheses have emerged to explain this pattern, but controversy remains as to which mechanism is most general. We measured vertical gradients in stable carbon isotope composition (δ13C) in more than 200 trees of nine conifer species growing in mixed-species forests in the Northern Rocky Mountains, USA. For all species except western larch, δ13C decreased from top to bottom of the canopy. We found that δ13C was strongly correlated with nitrogen per unit leaf area (N
area), which is a measure of photosynthetic capacity. Usually weaker correlations were found between δ13C and leaf mass per area, nitrogen per unit leaf mass, height from the ground, or depth in the canopy, and these correlations were more variable between trees than for N
area. Gradients of δ13C (per meter canopy depth) were steeper in small trees than in tall trees, indicating that a recent explanation of δ13C gradients in terms of drought stress of upper canopy leaves is unlikely to apply in our study area. The strong relationship between N
area and δ13C here reported is consistent with the general finding that leaves or species with higher photosynthetic capacity tend to maintain lower CO2 concentrations inside leaves. We conclude that photosynthetic capacity is a strong determinant of δ13C in vertical canopy profiles, and must be accounted for when interpreting δ13C values in conifer forests. 相似文献
19.
The rate of photosynthesis (P
N) in leaves and pods as well as carbon isotope content in leaves, pod walls, and seeds was measured in well-watered (WW) and water-stressed (WS) chickpea plants. The P
N, on an area basis, was negligible in pods compared to leaves and was reduced by water stress (by 26%) only in leaves. WS pod walls and seeds discriminated less against 13CO2 than did the controls. This response was not observed for leaves as is usually the case. Pod walls and seeds discriminated less against 13CO2 than did leaves in both WW and WS plants. Measurement of carbon isotope composition in pods may be a more sensitive tool for assessing the impact of water stress on long-term assimilation than is the instantaneous measurement of gas exchange rates. 相似文献
20.
The binding of ethyl carbamate labelled with carbon-14 in the alkyl or carbonyl group, and of methyl, n-butyl and n-propyl carbamates labelled in the alkyl group, to the DNA of mouse liver, lung and kidney has been studied in male Crackenbush mice. Only ethyl carbamate bound to liver and kidney DNA to any significant extent.The binding of ethyl carbamate labelled with carbon-14 in the C1, C2 or the carbonyl position was examined and compared. The levels of binding of [1-14C]- and [2-14C]ethyl carbamate to liver DNA were not significantly different (328 ± 34 and 267 ± 24 dpm/mg DNA, respectively), but there was very little binding of the [carbonyl-14C]ethyl carbamate (26 ± 3 dpm/mg DNA). Furthermore, only 18% of the radioactivity was removed from the DNA labelled with the alkyl-labelled carbamates, whereas 65% of the radioactivity was removed from the DNA labelled with carbonyl-labelled ethyl carbamate on continuous ether extraction. It was concluded that the bound molecule does not contain the carbonyl carbon and is probably an ethyl group. 相似文献