Search

As the use of microbial inoculants in agriculture rises, it becomes important to understand how the environment may influence microbial ability to promote plant growth. This work examines whether there are light dependencies in the biological functions of Azospirillum brasilense, a commercialized prolific grass-root colonizer. Though classically defined as non-phototrophic, A. brasilense possesses photoreceptors that could perceive light conducted through its host’s roots. Here, we examined the light dependency of atmospheric biological nitrogen fixation (BNF) and auxin biosynthesis along with supporting processes including ATP biosynthesis, and iron and manganese uptake. Functional mutants of A. brasilense were studied in light and dark environments: HM053 (high BNF and auxin production), ipdC (capable of BNF, deficient in auxin production), and FP10 (capable of auxin production, deficient in BNF). HM053 exhibited the highest rate of nitrogenase activity with the greatest light dependency comparing iterations in light and dark environments. The ipdC mutant showed similar behavior with relatively lower nitrogenase activity observed, while FP10 did not show a light dependency. Auxin biosynthesis showed strong light dependencies in HM053 and FP10 strains, but not for ipdC. Ferrous iron is involved in BNF, and a light dependency was observed for microbial 59Fe2+ uptake in HM053 and ipdC, but not FP10. Surprisingly, a light dependency for 52Mn2+ uptake was only observed in ipdC. Finally, ATP biosynthesis was sensitive to light across all three mutants favoring blue light over red light compared to darkness with observed ATP levels in descending order for HM053 > ipdC > FP10.

Iron (Fe), an essential element for plant growth, is abundant in soil but with low bioavailability. Thus, plants developed specialized mechanisms to sequester the element. Beneficial microbes have recently become a favored method to promote plant growth through increased uptake of essential micronutrients, like Fe, yet little is known of their mechanisms of action. Functional mutants of the epiphytic bacterium Azospirillum brasilense, a prolific grass-root colonizer, were used to examine mechanisms for promoting iron uptake in Zea mays. Mutants included HM053, FP10, and ipdC, which have varying capacities for biological nitrogen fixation and production of the plant hormone auxin. Using radioactive iron-59 tracing and inductively coupled plasma mass spectrometry, we documented significant differences in host uptake of Fe2+/3+ correlating with mutant biological function. Radioactive carbon-11, administered to plants as 11CO2, provided insights into shifts in host usage of ‘new’ carbon resources in the presence of these beneficial microbes. Of the mutants examined, HM053 exhibited the greatest influence on host Fe uptake with increased plant allocation of 11C-resources to roots where they were transformed and exuded as 11C-acidic substrates to aid in Fe-chelation, and increased C-11 partitioning into citric acid, nicotianamine and histidine to aid in the in situ translocation of Fe once assimilated.

While photosynthates are partitioned by the relative strength of young developing leaves and roots as sinks for carbon-based resources, many plants also show a close relationship between partitioning, phyllotaxy and vascular connectivity, yielding sectorial patterns of allocation. We examined whether smoke influences phloem vascular sectoriality in a model plant, sunflower (Helianthis annuus L.). Using radioactive 11CO2 administered to single photosynthetically active source leaves, we examined the transport behavior and allocation patterns of 11C-photosynthates using gamma counting and autoradiography. Soil treated with liquid smoke caused significant reductions in phloem sectoriality involving young sink leaves and roots. The resulting increase in vascular connectivity could benefit young plant performance by allowing a more uniform allocation of nutrients and/or stress signal molecules at a critical time of their growth.One-Sentence SummarySmoke-exposed roots exhibit a significant reduction in phloem sectoriality involving carbon transport to young sink leaves and roots.

In agriculture, plant growth promoting bacteria (PGPB) are increasingly used for reducing environmental stress-related crop losses through mutualistic actions of these microorganisms, activating physiological and biochemical responses, building tolerances within their hosts. Here we report the use of radioactive carbon-11 (t

Herbaspirillum seropedicae, as an endophyte and prolific root colonizer of numerous cereal crops, occupies an important ecological niche in agriculture because of its ability to promote plant growth and potentially improve crop yield. More importantly, there exists the untapped potential to harness its ability, as a diazotroph, to fix atmospheric N2 as an alternative nitrogen resource to synthetic fertilizers. While mechanisms for plant growth promotion remain controversial, especially in cereal crops, one irrefutable fact is these microorganisms rely heavily on plant-borne carbon as their main energy source in support of their own growth and biological functions. Biological nitrogen fixation (BNF), a microbial function that is reliant on nitrogenase enzyme activity, is extremely sensitive to the localized nitrogen environment of the microorganism. However, whether internal root colonization can serve to shield the microorganisms and de-sensitize nitrogenase activity to changes in the soil nitrogen status remains unanswered. We used RAM10, a GFP-reporting strain of H. seropedicae, and administered radioactive 11CO2 tracer to intact 3-week-old maize leaves and followed 11C-photosynthates to sites within intact roots where actively fluorescing microbial colonies assimilated the tracer. We examined the influence of administering either 1 mM or 10 mM nitrate during plant growth on microbial demands for plant-borne 11C. Nitrogenase activity was also examined under the same growth conditions using the acetylene reduction assay. We found that plant growth under low nitrate resulted in higher nitrogenase activity as well as higher microbial demands for plant-borne carbon than plant growth under high nitrate. However, carbon availability was significantly diminished under low nitrate growth due to reduced host CO2 fixation and reduced allocation of carbon resources to the roots. This response of the host caused significant inhibition of microbial growth. In summary, internal root colonization did little to shield these endophytic microorganisms from the nitrogen environment.

Internal nitrogen (N) cycling is crucial to N use efficiency. For example, N may be remobilized from older, shaded leaves to young leaves near the apex that receive more direct sunlight, where the N can be used more effectively for photosynthesis. Yet our understanding of the mechanisms and regulation of N transport is limited. To identify relevant transporters in Arabidopsis, fifteen transporter knockout mutants were screened for defects in leaf N export using nitrogen-13 (13N) administered as 13NH3 gas to leaves. We found that three nitrate/peptide transporter family (NPF) genes were necessary for normal leaf N export under low N but not adequate soil N availability, including AtNPF7.1, which has not been previously characterized. High-throughput phenotyping revealed altered leaf area and chlorophyll fluorescence relative to wild-type plants. High AtNPF7.1 expression in flowers and large flower stalks of Atnpf7.1 mutants in low N suggests that AtNPF7.1 influences leaf N export via sink-to-source feedback, perhaps via a role in sensing plant internal N-status. We also identified previously unreported phenotypes for the mutants of the other two NPF transporters that indicate possible roles in N sensing networks.

Among the PGPB, the genus Azospirillum—with an emphasis on A. brasilense—is likely the most studied microorganism for mitigation of plant stress. Here, we report the investigation of functional mutants HM053, ipdC and FP10 of A. brasilense to understand how the biological functions of these microorganisms can affect host Zn uptake. HM053 is a Nif+ constitutively expressed strain that hyper-fixes N2 and produces high levels of the plant’s relevant hormone auxin. FP10 is a Nif- strain deficient in N2-fixation. ipdC is a strain that is deficient in auxin production but fixes N2. Zn uptake was measured in laboratory-based studies of 3-week-old plants using radioactive 65Zn2+ (t½ 244 days). Principal Component Analysis was applied to draw out correlations between microbial functions and host 65Zn2+ accumulation. Additionally, statistical correlations were made to our prior data on plant uptake of radioactive 59Fe3+ and 59Fe2+. These correlations showed that low microbial auxin-producing capacity resulted in the greatest accumulation of 65Zn. Just the opposite effect was noted for 59Fe where high microbial auxin-producing capacity resulted in the greatest accumulation of that tracer.

Herbaspirillum seropedicae is a rhizobacteria that occupies a specialized ecological niche in agriculture. As an endophyte and prolific grass root colonizer it has the potential to promote plant growth, enhancing crop yield in many cereal crops. While the mechanisms for plant growth promotion are controversial, the one irrefutable fact is these microorganisms rely heavily on plant-borne carbon as their main energy source in support of their biological functions. Unfortunately, the tools and technology enabling researchers to trace carbon exchange between plants and the microorganisms associating with them has been limiting. Here, we demonstrate that radioactive 11CO2 administered to intact maize leaves with translocation of 11C-photosynthates to roots can provide a ‘traceable’ source of carbon whose assimilation by microbial organisms can be quantified with enormous sensitivity. Fluorescence root imaging of RAM10, a green fluorescent protein (GFP) reporting strain of H. seropedicae, was used to identify regions of high microbial colonization. Microbes were mechanically removed from these regions via sonication in saline solution and extracts were subjected to fluorescence measurement and gamma counting to correlate carbon-11 atoms with numbers of colony forming units. The method has potential to translate to other microorganisms provided they possess an optical reporting trait.

An increasing global population of over 4.5 billion people drives increasing demand for calories—30% of which are satisfied by grain crops, such as maize. High-density farming practices have been implemented but tend to deplete the soil of essential elements resulting in lower nutritional value, notably iron, of cultivated crops. Low iron content in staple crops can contribute over time to severe, even fatal, micronutrient deficiencies. Enhancing grain iron content using post-harvest biofortification strategies can be costly. However, field inoculation using biologics like Azospirillum brasilense (HM053) can be a cost-effective alternative to improving crop nutritional value. Using ion chromatography with chemiluminescence detection, we have shown that maize seeds harvested from outdoor pot-grown plants possessed a four-fold higher iron content as ferrous iron (Fe+2) compared to non-inoculated plants. Seeds from A. brasilense HM053-inoculated plants also contained approximately 13 nmol of ferritin per ground dried weight of kernel compared to 3 nmol from non-inoculated plants. In addition, A. brasilense HM053 inoculation increased crop yield 30–50% relative to non-inoculated plants.

Boron (B) is an essential plant micronutrient. Deficiencies of B have drastic consequences on plant development leading to crop yield losses and reductions in root and shoot growth. Understanding the molecular and cellular consequences of B deficiency is challenging, partly because of the limited availability of B imaging techniques. In this report we demonstrate the efficacy of using 4-fluorophenylboronic acid (FPBA) as a B imaging agent, which is a derivative of the B deficiency mimic phenylboronic acid (PBA). We show that radioactively labelled [18F]FPBA (t½=110 m) accumulates at the root tip, the root elongation zone and at lateral root initiation sites in maize roots, and also translocates to the shoot where it accumulates along the leaf edges. Treatment of maize seedlings using FPBA and PBA causes a shortened primary root phenotype with absence of lateral roots in a dose-dependent manner. The primary root defects can be partially rescued by the addition of boric acid indicating that PBA can be used to induce B deficiency in maize and that radioactively labelled FPBA can be used to image sites of B demand on a tissue level.

Experiments designed to quantify the physiological and metabolic status of Arabidopsis thaliana seedlings across their photoperiod used wild-type (Col-0) and genotypes deficient in starch biosynthesis (adg1-1, pgm-1) and in starch degradation (sex1-1) in combination with radioactive 11CO2 tracer. Measurements of root allocation and root exudation of 11C-photosynthates were conducted at 1 and 8 h into a 10-h photoperiod. Additionally, bioassays conducted on leaflets provided a measure of ‘new’ carbon partitioning (as 11C) into structural and non-structural carbohydrates. As expected, starch biosynthesis increased across the photoperiod for Col-0 controls. This increase was correlated with systematic decreases in soluble sugar synthesis and sink allocation across the photoperiod. We showed that this was a causal effect, which could be eliminated in the adg1-1 and pgm-1 mutants that were deficient in starch biosynthesis. Furthermore, these genotypes exhibited systematically higher levels of soluble sugars and sink allocation relative to Col-0 and sex1-1, which resulted in very different root growth traits including (i) increased root area balanced by shorter primary roots; (ii) increased number of lateral roots; and (iii) decreased root gravitropism. These differences in growth traits were correlated with higher observed levels of root auxin in these genotypes and to lower starch levels in their root tips. In a broader context, we find that carbon storage in the aboveground tissues can strongly influence root growth and root architecture belowground.

The development of a convenient and rapid method to synthesize radiolabeled, enantiomerically pure amino acids (AAs) as potential positron emission tomography (PET) imaging agents for mapping various biochemical transformations in living organisms remains a challenge. This is especially true for the synthesis of carbon-11-labeled AAs given the short half-life of carbon-11 (11 C, t1/2 =20.4 min). A facile synthetic pathway to prepare enantiomerically pure 11 C-labeled l-asparagine was developed using a partially protected serine as a starting material with a four-step transformation providing a chiral five-membered cyclic sulfamidate as the radiolabeling precursor. Its structure and absolute configuration were confirmed by X-ray crystallography. Utilizing a [11 C]cyanide nucleophilic ring opening reaction followed by selective acidic hydrolysis and deprotection, enantiomerically pure l-[4-11 C]asparagine was synthesized. Further optimization of reaction parameters, including base, metal ion source, solvent, acid component, reaction temperature and reaction time, a reliable two-step method for synthesizing l-[4-11 C]asparagine was presented: within a 45±3 min (n=5, from end-of-bombardment), the desired enantiomerically pure product was synthesized with the initial nucleophilic cyanation yield of 69±4 % (n=5) and overall two-step radiochemical yield of 53±2 % (n=5) based on starting [11 C]HCN, and with radiochemical purity of 96±2 % (n=5).

A rapid, mild radiosynthesis of freebase [11C]nicotine was developed by the methylation of freebase nornicotine with [11C]methyl triflate in acetone (5 min, 45 °C). A basic (pH 10.5–11.0) HPLC system reproducibly yielded freebase [11C]nicotine as a well-defined single peak. The freebase [11C]nicotine was concentrated by solid phase extraction and formulated in 50 µL ethanol (370 MBq/50 µL) without evaporative loss suitable for a cigarette spiking study. A radiochemical yield of 60.4±4.7% (n=3), radiochemical purity ≥99.9% and specific activity of 648 GBq/µmol at EOB for 5 min beams were achieved.

Noninvasive techniques to measure phloem transport of carbon will be crucial to efforts to engineer improved crop yields, which are highly dependent on carbon partitioning. Phloem, which is buried in the interior of the plant, is highly sensitive to tissue damage. Here we describe nondestructive methods using carbon-11, fed to leaves as 11CO2, as a tracer to track export of recently fixed carbon from leaves, transport speed through the phloem, and distribution or partitioning throughout the plant.

In the struggle to survive herbivory by leaf-feeding insects, plants employ multiple strategies to defend themselves. One mechanism by which plants increase resistance is by intensifying their responsiveness in the production of certain defense agents to create a rapid response. Known as defense priming, this action can accelerate and amplify responses of metabolic pathways, providing plants with long-lasting resistance, especially when faced with waves of attack. In the work presented, short-lived radiotracers of carbon administered as 11CO2 and nitrogen administered as 13NH3 were applied in Nicotiana tabacum, to examine the temporal changes in &lsquo, new&rsquo, C/N utilization in the biosynthesis of key amino acids (AAs). Responses were induced by using topical application of the defense hormone jasmonic acid (JA). After a single treatment, metabolic partitioning of recently fixed carbon (designated &lsquo, carbon and reflected as 11C) increased through the shikimate pathway, giving rise to tyrosine, phenylalanine and tryptophan. Amplification in &lsquo, carbon fluxes preceded changes in the endogenous (12C) pools of these AAs. Testing after serial JA treatments revealed that fluxes of &lsquo, carbon were accelerated, amplified and sustained over time at this higher rate, suggesting a priming effect. Similar results were observed with recently assimilated nitrogen (designated &lsquo, nitrogen reflected as 13N) with its partitioning into serine, glycine and glutamine, which play important roles supporting the shikimate pathway and downstream secondary metabolism. Finally, X-ray fluorescence imaging revealed that levels of the element Mn, an important co-factor for enzyme regulation in the shikimate pathway, increased within JA treated tissues, suggesting a link between plant metal ion regulation and C/N metabolic priming.

The overarching challenges for agriculture and thus also for agricultural engineers are to contribute to long term food safety and security (including a fair access to food for everybody) as well as to sustainable primary food production. Agricultural engineers generally as well as (new) agricultural technologies (further jointly referred to as agricultural engineering) must comply with economical, ecological and social requirements and mitigate social unbalance in different parts of the world. For this, agricultural engineering must contribute to increase the level of agricultural mechanization esp. in developing countries. Furthermore, it is a core task of agricultural engineering to mitigate the impacts of climate change. The European association of agricultural equipment and machinery producers (CEMA) has defined "Producing More with Less" as a guiding principle for agricultural engineering1

Summary Nitrogen-fixing rhizobacteria can promote plant growth; however, it is controversial whether biological nitrogen fixation (BNF) from associative interaction contributes to growth promotion. The roots of Setaria viridis, a model C4 grass, were effectively colonized by bacterial inoculants resulting in a significant enhancement of growth. Nitrogen-13 tracer studies provided direct evidence for tracer uptake by the host plant and incorporation into protein. Indeed, plants showed robust growth under nitrogen-limiting conditions when inoculated with an ammonium-excreting strain of Azospirillum brasilense. 11C-labeling experiments showed that patterns in central carbon metabolism and resource allocation exhibited by nitrogen-starved plants were largely reversed by bacterial inoculation, such that they resembled plants grown under nitrogen-sufficient conditions. Adoption of S. viridis as a model should promote research into the mechanisms of associative nitrogen fixation with the ultimate goal of greater adoption of BNF for sustainable crop production.

A rapid method for the synthesis of carbon-11 radiolabeled indole was developed using a sub-nanomolar quantity of no-carrier-added [(11)C]cyanide as radio-precursor. Based upon a reported synthesis of 2-(2-nitrophenyl)acetonitrile (), a highly reactive substrate 2-nitrobenzyl bromide () was evaluated for nucleophilic [(11)C]cyanation. Additionally, related reaction conditions were explored with the goal of obtaining of highly reactive 2-(2-nitrophenyl)-[1-(11)C]acetonitrile () while inhibiting its rapid conversion to 2,3-bis(2-nitrophenyl)-[1-(11)C]propanenitrile (). Next, a RANEY® Nickel catalyzed reductive cyclization method was utilized for synthesizing the desired [2-(11)C]indole with hydrazinium monoformate as the active reducing agent. Extensive and iterative screening of basicity, temperature and stoichiometry was required to overcome the large stoichiometry bias that favored 2-nitrobenzylbromide () over [(11)C]cyanide, which both caused further alkylation of the desired nitrile and poisoned the RANEY® Nickel catalyst. The result is an efficient two-step, streamlined method to reliably synthesize [2-(11)C]indole with an entire radiochemical yield of 21 ± 2.2% (n = 5, ranging from 18-24%). The radiochemical purity of the final product was >98% and specific activity was 176 ± 24.8 GBq μmol(-1) (n = 5, ranging from 141-204 GBq μmol(-1)). The total radiosynthesis time including product purification by semi-preparative HPLC was 50-55 min from end of cyclotron bombardment.

An operationally friendly, two-step, one-pot process has been developed for the rapid synthesis of carbon-11 labeled indole-3-acetic acid ([11C]IAA or [11C]auxin). By replacing an aprotic polar solvent with tetraethylene glycol, nucleophilic [11C]cyanation and alkaline hydrolysis reactions were performed consecutively in a single pot without a time-consuming intermediate purification step. The entire production time for this updated procedure is 55 min, which dramatically simplifies the entire synthesis and reduces the starting radioactivity required for a whole plant imaging study.

Carbon-11 (β+ emitter, t 1/2 = 20.4 min) radiolabeled l-glutamine is a potentially useful molecular imaging agent that can be utilized with positron emission tomography for both human oncological diagnosis and plant imaging research. Based upon a previously reported [11C]cyanide end-capping labeling method, a systematic investigation of nucleophilic cyanation reactions and acidic hydrolysis reaction parameters, including base, metal ion source, phase transfer catalyst, solvent, reaction temperature and reaction time, was conducted. The result was a milder, more reliable, two-step method which provides l-[5-11C]-glutamine with a radiochemical yield of 63.8 ± 8.7 % (range from 51 to 74 %, n = 10) with >90 % radiochemical purity and >90 % enantiomeric purity. The total synthesis time was 40–50 min from the end of bombardment. In addition, an Fmoc derivatization method was developed to measure the specific activity of this radiotracer.

An improved production procedure and formulation method for the carbon-11 radiolabeled phytohormone, 3-indolyl-[l- 11 C]acetic acid ([ 11 C]IAA), was developed by modifying selected original reaction parameters. This updated procedure both doubled the yield (from 25.9±6.7% ( n =12) to 61.0±0.3% ( n =10)) and increased the concentration (0.2–0.4 GBq/0.15–0.3 mL), enabling us to provide the radiotracer [ 11 C]IAA suitable for in vivo phyto-PET-imaging studies. The specific activity was improved by more than a factor of three (26.7±5.6 GBq/µmol to 82.5±36.1 GBq/µmol). The total synthesis time for both production and formulation was 81.8±3.0 min ( n =10). In addition, a streamlined semi-remote controlled production system, containing five processing modules, was designed and built for routine [ 11 C]IAA production. This integrated system facilitated routine high radiation level production of [ 11 C]IAA while minimizing radiation exposure to the production chemists.

Upon attack by leaf herbivores, many plants reallocate photoassimilates below ground. However, little is known about how plants respond when the roots themselves come under attack. We investigated induced resource allocation in maize plants that are infested by the larvae Western corn rootworm Diabrotica virgifera virgifera. Using radioactive (11) CO(2), we demonstrate that root-attacked maize plants allocate more new (11) C carbon from source leaves to stems, but not to roots. Reduced meristematic activity and reduced invertase activity in attacked maize root systems are identified as possible drivers of this shoot reallocation response. The increased allocation of photoassimilates to stems is shown to be associated with a marked thickening of these tissues and increased growth of stem-borne crown roots. A strong quantitative correlation between stem thickness and root regrowth across different watering levels suggests that retaining photoassimilates in the shoots may help root-attacked plants to compensate for the loss of belowground tissues. Taken together, our results indicate that induced tolerance may be an important strategy of plants to withstand belowground attack. Furthermore, root herbivore-induced carbon reallocation needs to be taken into account when studying plant-mediated interactions between herbivores.

The full data set of the experiment NA31 at CERN has been used to search for the decay Ks→π0e+e-. The rate of this decay may be used to determine the magnitude of the state mixing contribution to the CP violating amplitude of the decay KL→π0e+e-. No candidate events for the decayKs→π0e+- were seen. This leads to a branching ratio upper limit of 1.1 × 10-6 at the 90% confidence level. © 1993.

A rapid, mild radiosynthesis of freebase [

Upon herbivory, plants can swiftly reallocate newly acquired resources to different tissues within a plant. Although the herbivore-induced movement of resources is apparent, the movement direction and the role of the remobilized resources are not well understood. Here, we used a two-pronged approach combining radioisotope and metabolomic techniques to shed light on whole-plant resource reallocation and changes in primary metabolism within the tomato, Solanum lycopersicum (L.) (Solanaceae), model in response to simulated herbivory by the specialist Manduca sexta (L.) (Lepidoptera: Sphingidae). Manduca sexta regurgitant applied to damaged leaves, but not mechanical damage alone, increased 11C-photosynthate allocation to roots but did not affect 11CO2 fixation and leaf export. Changes in primary metabolite concentrations occurred mostly in sink tissues (apex and roots) as well as in damaged leaves. Both damage treatments (with and without M. sexta regurgitant application) resulted in increased concentrations of primary metabolites relative to undamaged plants in the apex and decreased concentrations in the roots, but there were also extensive changes specific to each damage treatment. Mechanical damage alone led to changes consistent with water stress caused by tissue damage. Manduca sexta led to metabolite increases in the apex consistent with an increase in glucose breakdown, metabolite increases in damaged leaves consistent with starch degradation, and metabolite decreases in roots suggesting a high use of metabolites. A possible explanation for the observed patterns in the aboveground tissues might be an increase in carbohydrate degradation to support defense production in attacked leaves and vulnerable developing leaves, and/or subsequent remobilization to belowground tissues to support high carbohydrate demand for respiration, enhanced nutrient uptake, and storage.

Validamycin A was used to inhibit in vivo trehalase activity in tobacco enabling the study of subsequent changes in new C partitioning into cellulosic biomass and lignin precursors. After 12-h exposure to treatment, plants were pulse labeled using radioactive 11 CO 2 , and the partitioning of isotope was traced into [ 11 C]cellulose and [ 11 C]hemicellulose, as well as into [ 11 C]phenylalanine, the precursor for lignin. Over this time course of treatment, new carbon partitioning into hemicellulose and cellulose was increased, while new carbon partitioning into phenylalanine was decreased. This trend was accompanied by a decrease in phenylalanine ammonia-lyase activity. After 4 d of exposure to validamycin A, we also measured leaf protein content and key C and N metabolite pools. Extended treatment increased foliar cellulose and starch content, decreased sucrose, and total amino acid and nitrate content, and had no effect on total protein.

We examined the timeline by which methyl jasmonate (MeJA) reprograms new carbon partitioning into key metabolite pools. The radioactive isotope 11C (t1/2 20.4 min), administered to intact leaves of Nicotiana tabacum L. (cv Samsun) as 11CO2 gas enabled us to measure changes in new carbon partitioning into soluble sugar and amino acid pools of [11C]photosynthate. A 500 μM MeJA treatment resulted in a decrease in the [11C]soluble sugar pool and an increase in the [11C]amino acid pool after 4 h. This pattern was more pronounced 15 h after treatment. We also examined the timeline for 11C-partitioning into aromatic amino acid metabolites of the shikimate pathway. [11C]Tyrosine, [11C]phenylalanine and [11C]tryptophan were elevated 1.5-fold, 12-fold and 12-fold, respectively, relative to controls, 4 h after MeJA treatment, while endogeneous pools were unchanged. This suggests that only new carbon is utilized during early stages of defense induction. By 15 h, [11C]tyrosine and [11C]phenylalanine returned to baseline while [11C]tryptophan was elevated 30-fold, suggesting that MeJA exerts selective control over the shikimate pathway. Finally, we measured trans-cinnamic acid levels as a gauge of downstream phenolic metabolism. Levels were unchanged 4 h after MeJA treatment relative to controls, but were increased 2-fold by 15 h, indicating a lag in response of secondary metabolism.

Nitrogen-13 (t(1/2) 9.97 m), a radioactive isotope of nitrogen, offers unique opportunities to explore plant nitrogen utilization over short time periods. Here we describe a method for administering (13)N as gaseous (13)NH(3) to intact leaves of Nicotiana tabacum L. (cv Samsun), and measuring the labelled amino acids using radio high-performance liquid chromatography (HPLC) on tissue extract. We used this method to study the effects of defence induction on plant nitrogen utilization by applying treatments of methyl jasmonate (MeJA), a potent defence elicitor. MeJA caused a significant increase relative to controls in key [(13)N]amino acids, including serine, glycine and alanine by 4 h post-treatment, yet had no effect on (13)NH(3) incorporation, a process that is primarily under the control of the glutamine synthatase/glutamate synthase pathway (GS/GOGAT) in cellular photorespiration. We suggest that the reconfiguration of nitrogen metabolism may reflect induction of non-photorespiratory sources of nitrogen to better serve the plant's defences.

• Evidence is emerging to support the notion that in response to herbivory, plants undergo changes in their primary metabolism and are able to fine-tune the allocation of new and existing resources and temporarily direct them to storage organs. • We hypothesized that simulated herbivory increases the export of resources out of the affected tissues and increases allocation to roots. We used short-lived radioisotopes to study in vivo the dynamics of newly incorporated (11)CO(2) and (13)NH(3). Methyl jasmonate (MeJA), a known defense elicitor, was applied to the foliage of tomato plants and 4 h later we monitored leaf uptake, export and whole-plant allocation of [(11)C]photosynthate and [(13)N]amino acids. • There was a marginally significant decrease in the fixation of (11)CO(2), and an increase in the export of newly acquired carbon and nitrogen out of MeJA-treated leaves. The proportion of nitrogen allocated to roots increased, whereas the proportion of carbon did not change. • These results are in agreement with our hypotheses, showing a change in the allocation of resources after treatment with MeJA; this may reduce the chance of resources being lost to herbivores and act as a buffer to biotic stress by increasing the potential for plant regrowth and survival after the attack.

Here we explored the conditions for synthesizing [ 13 N]N 2 in a state that is suitable for the administration to plant root nodules enabling studies of nitrogen fixation. [ 13 N]N 2 was prepared batchwise, starting with [ 13 N]NO ― 3 from the 16 O(p,α) 13 N nuclear reaction on a liquid water target. [ 13 N]NO ― 3 was first reduced to [ 13 N]NH 3 using Devarda's alloy, and then the [ 13 N]NH 3 was oxidized to [ 13 N]N 2 by hypobromite using carrier-added NH 4 CI. The amounts of carrier NH 4 CI and hypobromite were varied to determine the effects these parameters had on the radiochemical yield, and on the radiotracer specific activity. As expected, increasing the amount of carrier NH 4 CI improved the radiochemical yield. Unexpectedly, increasing the amount of excess hypobromite from 1.6-fold to 6-fold molar equivalents (relative to NH 4 CI) improved the radiochemical yield and radiotracer specific activity under all conditions of carrier NH 4 Cl. As a comparison, we measured [ 13 N]N 2 specific activity derived from in-target production based on a 50 μA min irradiation driving the 14 N(p,pn) 13 N reaction on a gaseous N 2 target. The 'wet' radiochemistry approach afforded two advantages over the in-target approach with a ∼600-fold improvement in specific activity, and the ability to collect the tracer in a small volume of gas (∼20 mL at STP).

Cyclotron targets for the production of [ 18 F]-fluoride usually use a thin metal window to contain the 18 O-enriched water during irradiation. This window is activated by the proton beam, and undesired radioisotopes can enter the target water. A pre-packaged strong anion exchange resin is commonly used for [ 18 O]-water recovery. A two-column method has been developed which delivers >95% of the [ 18 F]-fluoride for radiosynthesis while rejecting >99.9% of the contaminants.

Cross-sections for the production of (181)Re, (182m)Re, (182g)Re, (183)Re, (184)Re, and (186)Re from proton bombardment of natural tungsten have been measured using the stacked foil technique for proton energies up to 17.6 MeV. Results are compared with the theoretical excitation functions as calculated by the EMPIRE II code (version 2.19) and experimental literature values. Results are in strong agreement with some of the previously reported literature as well at theoretical calculations for multiple reactions providing for more reliable estimates for the (186)W(p,n)(186)Re reaction.

In this research, we aim to directly measure the specific activity (SA) of the carbon-11 cyanide ([(11)C]CN¯) produced by our in-house built automated [(11)C]HCN production system and to identify the major sources of (12)C-cyanide ((12)CN¯). The [(11)C]CN¯ is produced from [(11)C]CO2, which is generated by the (14)N(p,α)(11)C nuclear reaction using a cyclotron. Direct measurement of cyanide concentrations was accomplished using a relatively inexpensive, and easy to use ion selective electrode (ISE) which offered an appropriate range of sensitivity for detecting mass. Multiple components of the [(11)C]HCN production system were isolated in order to determine their relative contributions to (12)CN¯ mass. It was determined that the system gases were responsible for approximately 30% of the mass, and that the molecular sieve/nickel furnace unit contributed approximately 70% of the mass. Beam on target (33µA for 1 and 10min) did not contribute significantly to the mass. Additionally, we compared the SA of our [(11)C]HCN precursor determined using the ISE to the SA of our current [(11)C]CN¯ derived radiotracers determined by HPLC to assure there was no significant difference between the two methods. These results are the first reported use of an ion selective electrode to determine the SA of no-carrier-added cyanide ion, and clearly show that it is a valuable, inexpensive and readily available tool suitable for this purpose.

BNL has recently purchased an EBCO TR-19 cyclotron for routine isotope production. A system has been built to recover oxygen-18 enriched water from the fluorine-18 target, and then transport the F-18 a distance of 50 m to a shielded dose splitter. The remotized system provides the operator with feedback on flow rates and radiation levels during processing. Recovery of the 2.6 mL of enriched water and transport of the F-18 to the radiochemistry labs takes under 10 min, with more than 80% of the activity arriving in the chemistry lab.

Monoclonal antibodies are capable of modulating drug metabolising enzymes resulting in unexpected plasma concentrations of a drug when given concomitantly. Therefore plasma concentration of paclitaxel (PTX) and its metabolites has been monitored in 10 patients with advanced breast cancer during treatment with PTX alone or combined with trastuzumab (TMAB, paired cross over design). Compared to the MONO regimen PTX peak plasma concentrations were about 25% lower in the TMAB schedule: cmax = 3294 +/- 1174 ng/ml (MONO: 4368 +/- 1887 ng/ml). TMAB also caused lower peak plasma concentration of the main metabolite 6-hydroxy PTX (248 +/- 89 ng/ml) compared to the MONO schedule (194 +/- 82 ng/ml). Cmax of the minor metabolites was distinctly below 100 ng/ml and consequently differed negligible in both schedules. The similar apparent formation rate of the metabolites in both schedules (range from 30 to 50 min) as well as identical tmax values (range 170-190 min) suggested that TMAB had no influence on PTX metabolism. In accordance to plasma concentrations, AUClast of PTX was lower in the MONO schedule (733 +/- 197 microg/ml*min, AUClast = 669 +/- 248 microg/ml*min for TMAB) but without significance. In summary no indices for an altered plasma disposition of PTX and its metabolites could be found when TMAB was given concomitantly.