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International audience; We investigate here the potential of single step production of genetically engineeredmagnetosomes, bacterial biogenic iron-oxide nanoparticles embedded in a lipid vesicle, as a new tailorable magnetic resonance molecular imaging probe. We demonstrate in vitro the specific binding and the significant internalization into U87 cells of magnetosomes decorated with RGD peptide. After injection at the tail vein of glioblastoma-bearing mice, we evidence in the first 2 h the rapid accumulation of both unlabeled and functionalized magnetosomes inside the tumor by Enhanced Permeability and Retention effects. 24 h after the injection, a specific enhancement of the tumor contrast is observed on MR images only for RGD-labeled magnetosomes. Post mortem acquisition of histological data confirms MRI results with more magnetosomes found into the tumor treated with functionalized magnetosomes. This work establishes the first proof-of-concept of a successful bio-integrated production of molecular imaging probe for MRI.

Industrial applications of fungal compounds, coupled with the emergence of fungal threats to natural ecosystems and public health, have increased interest in filamentous fungi. Among all pathogenic fungi, Penicillium verrucosum is one of the most common mold-infecting stored cereals in temperate regions. However, it is estimated that 80% of fungal secondary metabolites remain unknown. To detect new P. verrucosum compounds, an untargeted metabolomic approach was applied to fungus grown on wheat grains labeled with stable isotopes: (i) natural grains (99% 12C); (ii) grains enriched with 97% of 13C; and (iii) grains enriched with 53% of 13C and 97% of 15N. Analyses performed by high-performance liquid chromatography coupled with high-resolution mass spectrometry (HPLC-HRMS) enabled the specific detection of fungal metabolites, and the unambiguous characterization of their chemical formulas. In this way, 98 secondary metabolites were detected and their chemical formulas were determined. Of these, only 18 identifications could be made based on databases, the literature and mass spectrometry fragmentation experiments, with the result that 80 were totally unknown. Molecular networks were generated to analyze these results, leading to the characterization by MSn experiments of a new fungisporin produced by P. verrucosum. More generally, this article provides precise mass spectrometric data about all these compounds for further studies of the Penicillium metabolome.

[FeFe]-hydrogenase-like genes encode [Fe 4 S 4 ]-containing proteins that are ubiquitous in eukaryotic cells. In humans, iron-only hydrogenase-like protein 1 (IOP1) represses hypoxia inducible factor-1α subunit (HIF1-α) at normal atmospheric partial O 2 pressure (normoxia, 21 kPa O 2 ). In yeasts, the narl mutant cannot grow at 21 kPa O 2 . but can develop at a lower O 2 pressure (2 kPa O 2 ). We show here that plant [FeFe]-hydrogenase-like GOLLUM genes are essential for plant development and cell cycle progression. The mutant phenotypes of these plants are seen in normoxic conditions, but not under conditions of mild hypoxia (5 kPa O 2 ). Transcriptomic and metabolomic experiments showed that the mutation enhances the expression of some hypoxia-induced genes under normal atmospheric O 2 conditions and changes the cellular content of metabolites related to energy metabolism. In conclusion, [FeFe]-hydrogenase-like proteins play a central role in eukaryotes including the adaptation of plants to the ambient O 2 partial pressure.

Mycotoxins are secondary metabolites of filamentous fungi which can cause a wide range of systemic effects. Human health effects of inhaled mycotoxins remain poorly documented, despite the large amounts present, associated with air-borne particles. Among these mycotoxins, sterigmatocystin is one of the most prevalent. Because its chemical structure is close to that of the aflatoxins, we studied its metabolism and its cellular consequences when in contact with the airway epithelium, using the mass spectral signature from the 10% 13C uniformly enriched sterigmatocystin. The metabolism was studied in vitro, using recombinant cytochrome P450s enzymes, and in porcine tracheal epithelial cell (PTEC) primary cultures at an air-liquid interface. The metabolites were analyzed by high-performance liquid chromatography coupled with tandem mass spectrometry detection. Expressed enzymes and PTECs were exposed to uniformly 13 C-enriched sterigmatocystin to confirm the relationship between sterigmatocystin and its metabolites because this isotopic cluster shape is conserved for all metabolites and their product ions. Incubation of sterigmatocystin with recombinant cytochrome P450 1A1 led to the formation of three metabolites identified as monohydroxysterigmatocystin, dihydroxysterigmatocystin and one glutathione adduct, the latter after the formation of a transient intermediate. In the PTEC cultures, sterigmatocystin metabolism resulted in a glucuro-conjugate. Two other products were detected, a sulfo-conjugate and a glucuro-conjugate of hydroxysterigmatocystin upon cytochrome P450 1A1 induction. This is the first study to report sterigmatocystin metabolism in airway epithelium, and it suggests that, contrary to the aflatoxins, sterigmatocystin is mainly detoxified into its conjugates and is unable to produce significant amounts of reactive metabolites in respiratory cells, at least in pigs. Copyright © 2011 John Wiley & Sons, Ltd.

International audience; Human health effects of inhaled mycotoxins remain poorly documented, despite the large amounts present in bioaerosols. Among these mycotoxins, sterigmatocystin is one of the most prevalent. Our aim was to study the metabolism and cellular consequences of sterigmatocystin once it is in contact with the airway epithelium. Metabolites were analyzed first in vitro, using recombinant P450 1A1, 1A2, 2A6, 2A13, and 3A4 enzymes, and subsequently in porcine tracheal epithelial cell (PTEC) primary cultures at an air liquid interface. Expressed enzymes and PTECs were exposed to sterigmatocystin, uniformly enriched with C-13 to confirm the relationship between sterigmatocystin and metabolites. Induction of the expression of xenobiotic-metabolizing enzymes upon sterigmatocystin exposure was examined by real-time quantitative real-time polymerase chain reaction. Incubation of 50 mu M sterigmatocystin with recombinant P450 1A1 led to the formation of three metabolites: monohydroxy-sterigmatocystin (M1), dihydroxy-sterigmatocystin (M2), and one glutathione adduct (M3), the latter after the formation of a transient epoxide. Recombinant P450 1A2 also led to M1 and M3. P450 3A4 led to only M3. In PTEC, 1 mu M sterigmatocystin metabolism resulted in a glucuro conjugate (M4) mainly excreted at the basal side of cells. If PTEC were treated with beta-naphthoflavone prior to sterigmatocystin incubation, two other products were detected, i.e., a sulfo conjugate (M5) and a glucoro conjugate (M6) of hydroxysterigmatocystin. Exposure of PTEC for 24 h to 1 mu M sterigmatocystin induced an 18-fold increase in the mRNA levels of P450 1A1, without significantly induced 7-ethoxyresorufin O-deethylation activity. These data suggest that sterigmatocystin is mainly detoxified and is unable to produce significant amounts of reactive epoxide metabolites in respiratory cells. However, sterigmatocystin increases the P450 1A1 mRNA levels with unknown long-term consequences. These in vitro results obtained in the porcine pulmonary tract need to be confirmed in human epithelial cells.

Due to their low concentrations in biological matrices, mycotoxin analyses often encounter detection and quantification problems, especially for toxicokinetic studies. We have developed a strategy to produce in a single process, several fungi secondary metabolites uniformly enriched with 13C, 15N stable isotopes in their 'natural' composition. This includes: (1) a plant culture in the presence of 10%, 50% or 100% 13CO2 as the only source of carbon, and in the presence or not of 10% 15N-enriched nitrogen salts – as expected wheat or maize uniformlyincorporate enriched isotopes into their bioproducts; (2) a subsequent solid culture of different filamentous fungi on plant biomass led to the production of a 'natural' mixture of isotopes-enriched mycotoxins – these compounds exhibit a characteristic isotopic cluster, which can be easily detected by mass spectrometry. As an example, we achieved 10% uniformly 13C-enriched zearalenone, deoxynivalenol and mycophenolic acid by growing Fusarium graminearum or Penicillium brevicompactum on 10% 13C enriched wheat seeds and 3 to 10% 13C, 15N uniformly enriched fumonisins from Fusarium verticillioides cultures on maize seeds or straw. These compounds were used for metabolism and transport studies in mammals either in vitro or in vivo and analysed by MS and MSn spectra of the isotopic cluster but also by 13C, 15N NMR. Moreover, such isotopic pattern enrichment can be used for quantitative evaluations of mycotoxins transport across mammalian biological membranes, alone or in their 'natural' conditions in the presence of other fungi secondary metabolites. Finally, we used such enriched compounds with high reliabilityin order to study zearalenone metabolism but these enriched compounds would also be used as internal standards to quantify zearalenone or fumonisins in contaminated food samples.

Cellulose and wheat straw degradation by Ruminococcus albus was monitored using NMR spectroscopy. In situ solid-state (13)C-cross-polarization magic angle spinning NMR was used to monitor the modification of the composition and structure of cellulose and (13)C-enriched wheat straw during the growth of the bacterium on these substrates. In cellulose, amorphous regions were not preferentially degraded relative to crystalline areas by R. albus. Cellulose and hemicelluloses were also degraded at the same rate in wheat straw. Liquid state two-dimensional NMR experiments were used to analyse in detail the sugars released in the culture medium, and the integration of NMR signals enabled their quantification at various times of culture. The results showed glucose and cellodextrin accumulation in the medium of cellulose cultures; the cellodextrins were mainly cellotriose and accumulated to up to 2 mm after 4 days. In the wheat straw cultures, xylose was the main soluble sugar detected (1.4 mm); arabinose and glucose were also found, together with some oligosaccharides liberated from hemicellulose hydrolysis, but to a much lesser extent. No cellodextrins were detected. The results indicate that this strain of R. albus is unable to use glucose, xylose and arabinose for growth, but utilizes efficiently xylooligosaccharides. R. albus 20 appears to be less efficient than Fibrobacter succinogenes S85 for the degradation of wheat straw.

The fast development of sensitive molecular diagnostic tools is currently paving the way for a personalized medicine. A new class of ultrasensitive magnetic resonance imaging (MRI) T₂-contrast agents based on magnetosomes, magnetite nanocrystals biomineralized by magnetotactic bacteria, is proposed here. The contrast agents can be injected into the blood circulation and detected in the picomolar range. Purified magnetosomes are water-dispersible and stable within physiological conditions and exhibit at 17.2 T a transverse relaxivity r₂ four times higher than commercial ferumoxide. The subsequent gain in sensitivity by T₂(*) -weighted imaging at 17.2 T of the mouse brain vasculature is evidenced in vivo after tail vein injection of magnetosomes representing a low dose of iron (20 μmoliron kg(-1)), whereas no such phenomenon with the same dose of ferumoxide is observed. Preclinical studies of human pathologies in animal models will benefit from the combination of high magnetic field MRI with sensitive, low dose, easy-to-produce biocompatible contrast agents derived from bacterial magnetosomes.

Carbon isotopes (*C) have been extensively used in man to describe oxidative vs nonoxidative disposal of an exogenous load of labeled carbohydrate (*C-CHO) at rest in various experimental situations. It is hypothesized that V*CO(2) reflects *C-CHO oxidation. However, when glycogen is synthesized through the indirect pathway (which is responsible for approximately 50% of glycogen storage), *C could be lost, diluted, and exchanged in the pyruvate-lactate pool, in the pool of tricarboxylic acid cycle intermediates, as well as at the entrance of the tricarboxylic acid cycle, and along the pathway of gluconeogenesis. This could result in a lower *C/C in the glycogen stored than in the CHO administered, in an increased production of *CO(2), and, respectively, in an overestimation and an underestimation of the oxidative and nonoxidative disposal of the CHO load. Results from the present experiment offer a support to this hypothesis. Over a 10-hour period after ingestion of a (13)C-pasta meal (313+/-10 g dry mass or 258+/-8 g of glucose) in 12 healthy subjects (6 men and 6 women), exogenous CHO oxidation computed from V(13)CO(2) (recovery factor, 0.54) significantly exceeded total CHO oxidation computed by indirect respiratory calorimetry corrected for urea excretion: 154.2+/-2.6 vs 133.5+/-3.2 g. In an additional study conducted in rats, (13)C/(12)C in glycogen stores was significantly approximately 50% lower than in the (13)C-CHO ingested, over a wide range of enrichment. These results suggest that because of dilution, loss, and exchange of *C in the indirect pathway of glycogen synthesis, the oxidative vs nonoxidative disposal of exogenous *C-CHO cannot be accurately tracked from V*CO(2).

Different transgenic tobacco lines down-regulated for either one or two enzymes of the monolignol pathway were compared for their lignin content and composition, and developmental patterns. The comparison concerned CCR and CAD down-regulated lines (homozygous or heterozygous for the transgene) and the hybrids resulting from the crossing of transgenic lines individually altered for CCR or CAD activities. Surprisingly, the crosses containing only one allele of each antisense transgene, exhibit a dramatic reduction of lignin content similar to the CCR down-regulated parent but, in contrast to this transgenic line, display a normal phenotype and only slight alterations of the shape of the vessels. Qualitatively the lignin of the double transformant displays characteristics more like the wild type control than either of the other transgenics. In the transgenics with a low lignin content, the transformations induced other biochemical changes involving polysaccharides, phenolic components of the cell wall and also soluble phenolics. These results show that the ectopic expression of a specific transgene may have a different impact depending on the genetic background and suggest that the two transgenes present in the crosses may operate synergistically to reduce the lignin content. In addition, these data confirm that plants with a severe reduction in lignin content may undergo normal development at least in controlled conditions.

Studying lignin-biosynthetic-pathway mutants and transgenics provides insights into plant responses to perturbations of the lignification system, and enhances our understanding of normal lignification. When enzymes late in the pathway are downregulated, significant changes in the composition and structure of lignin may result. NMR spectroscopy provides powerful diagnostic tools for elucidating structures in the difficult lignin polymer, hinting at the chemical and biochemical changes that have occurred. COMT (caffeic acid O-methyl transferase) downregulation in poplar results in the incorporation of 5-hydroxyconiferyl alcohol into lignins via typical radical coupling reactions, but post-coupling quinone methide internal trapping reactions produce novel benzodioxane units in the lignin. CAD (cinnamyl alcohol dehydrogenase) downregulation results in the incorporation of the hydroxycinnamyl aldehyde monolignol precursors intimately into the polymer. Sinapyl aldehyde cross-couples 8–O–4 with both guaiacyl and syringyl units in the growing polymer, whereas coniferyl aldehyde cross-couples 8–O–4 only with syringyl units, reflecting simple chemical cross-coupling propensities. The incorporation of hydroxycinnamyl aldehyde and 5-hydroxyconiferyl alcohol monomers indicates that these monolignol intermediates are secreted to the cell wall for lignification. The recognition that novel units can incorporate into lignins portends significantly expanded opportunities for engineering the composition and consequent properties of lignin for improved utilization of valuable plant resources.

Homologous antisense constructs were used to down-regulate tobacco cinnamyl-alcohol dehydrogenase (CAD; EC 1.1.1.195 ) and cinnamoyl-CoA reductase (CCR; EC 1.2.1.44 ) activities in the lignin monomer biosynthetic pathway. CCR converts activated cinnamic acids (hydroxycinnamoyl–SCoAs) to cinnamaldehydes; cinnamaldehydes are then reduced to cinnamyl alcohols by CAD. The transformations caused the incorporation of nontraditional components into the extractable tobacco lignins, as evidenced by NMR. Isolated lignin of antisense-CAD tobacco contained fewer coniferyl and sinapyl alcohol-derived units that were compensated for by elevated levels of benzaldehydes and cinnamaldehydes. Products from radical coupling of cinnamaldehydes, particularly sinapaldehyde, which were barely discernible in normal tobacco, were major components of the antisense-CAD tobacco lignin. Lignin content was reduced in antisense-CCR tobacco, which displayed a markedly reduced vigor. That lignin contained fewer coniferyl alcohol-derived units and significant levels of tyramine ferulate. Tyramine ferulate is a sink for the anticipated build-up of feruloyl–SCoA, and may be up-regulated in response to a deficit of coniferyl alcohol. Although it is not yet clear whether the modified lignins are true structural components of the cell wall, the findings provide further indications of the metabolic plasticity of plant lignification. An ability to produce lignin from alternative monomers would open new avenues for manipulation of lignin by genetic biotechnologies.

13 pages; International audience; The turnover of organic matter in soil depends on the activity of microbial decomposers. However, little is known about how modifications of the diversity of soil microbial communities induced by fresh organic matter (FOM) inputs can regulate carbon cycling. Here, we investigated the decomposition of two 13C labeled crop residues (wheat and alfalfa) and the dynamics of the genetic structure and taxonomic composition of the soil bacterial communities decomposing 13C labeled FOM and native unlabeled soil organic matter (SOM), respectively. It was achieved by combining the stable isotope probing method with molecular tools (DNA genotyping and pyrosequencing of 16S rDNA). Although a priming effect (PE) was always induced by residue addition, its intensity increased with the degradability of the plant residue. The input of both wheat and alfalfa residues induced a rapid dynamics of FOM-degrading communities, corresponding to the stimulation of bacterial phyla which have been previously described as copiotrophic organisms. However, the dynamics and the identity of the bacterial groups stimulated depended on the residue added, with Firmicutes dominating in the wheat treatment and Proteobacteria dominating in the alfalfa treatment after 3 days of incubation. In both treatments, SOMdegrading communities were dominated by Acidobacteria, Verrucomicrobia, and Gemmatimonadetes phyla which have been previously described as oligotrophic organisms. An early stimulation of SOM-degrading populations mainly belonging to Firmicutes and Bacteroidetes groups was observed in the alfalfa treatment whereas no change occurred in the wheat treatment. Our findings support the hypothesis that the succession of bacterial taxonomic groups occurring in SOM- and FOMdegrading communities during the degradation process may be an important driver of the PE, and consequently of carbon dynamics in soil.

International audience; Characterization of fungal secondary metabolomes has become a challenge due to the industrial applications of many of these molecules, and also due to the emergence of fungal threats to public health and natural ecosystems. Given that, the aim of the present study was to develop an untargeted method to analyze fungal secondary metabolomes by combining high-accuracy mass spectrometry and double isotopic labeling of fungal metabolomes. The strain NRRL 35693 of Aspergillus fumigatus, an important fungal pathogen, was grown on three wheat grain substrates: (1) naturally enriched grains (99% 12C), (2) grains enriched 96.8% with 13C, (3) grains enriched with 53.4% with 13C and 96.8% with 15N. Twenty-one secondary metabolites were unambiguously identified by high-performance liquid chromatography−high-resolution mass spectrometry (HPLC−HRMS) analysis. AntiBase 2012 was used to confirm the identity of these metabolites. Additionally, on the basis of tandem mass spectrometry (MSn) experiments, it was possible to identify for the first time the formula and the structure of fumigaclavine D, a new member of the fumigaclavines family. Post biosynthesis degradation of tryptoquivaline F by methanol was also identified during HPLC− HRMS analysis by the detection of a carbon atom of nonfungal origin. The interest of this method lies not only on the unambiguous determination of the exact chemical formulas of fungal secondary metabolites but also on the easy discrimination of nonfungal products. Validation of the method was thus successfully achieved in this study, and it can now be applied to other fungal metabolomes, offering great possibilities for the discovery of new drugs or toxins.

A Zea mays callus culture containing chlorophyll was established and grown photomixotrophically. Cell chloroplast structure, and pigment and soluble protein contents were examined. Expression of some key enzymes of C4 carbon metabolism was compared with that of etiolated (heterotrophic) and green photoautotrophic leaves. Chlorophyll content of the callus was 15–20% that of green leaves. Soluble protein content of callus was half that of leaf cells. Electron microscopic observations showed that green callus cells contained only typical granal chloroplasts. Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco, EC 4.1.1.38) activities in green callus were ca 30% those of green leaves but 2–3 times higher than in etiolated leaves. Quantitative enzyme protein determination, using antibodies specific to maize leaf Rubisco showed that the chloroplastic carboxylase represented about 7% of total soluble protein in green callus, in parallel to its low chlorophyll content. The specific activity of Rubisco in callus and leaves was unchanged. Phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) activity in green callus was about 20% that of green leaves and similar to that measured in etiolated leaves. Apparent Km (PEP) values (0.08 mM) for PEPC isolated from green callus and etiolated leaves were very different from values (0.5 mM) obtained with PEPC from green leaves. These kinetic characteristics together with the absence of inhibition by malate and activation by glucose-6-phosphate suggest that the properties of PEPC isolated from green callus and etiolated maize leaves are very similar to those of PEPPC from C3 plants. Using PEPC antibodies specific to green maize leaf enzyme, immunotitration of PEPC preparations containing identical enzyme units allowed complete precipitation of the green leaf enzyme with increasing antibody volumes. In contrast, 60–70% of the activity of PEPC from etiolated and green callus was inhibited, suggesting low affinity for the maize green leaf PEPC antiserum (typical C4 form). Ouchterlony double diffusion tests revealed only partial recognition of PEPC in green callus and etiolated leaves. NAD-malate dehydrogenase (NAD-MDH, EC 1.1.1.37) activity in callus was 2 and 3 times higher, respectively, than in etiolated and green leaves. NADP-malic enzyme (NADP-ME, EC 1.1.1.40) activity in callus cultures was much lower than in green leaves. All our data support the hypothesis that cultures of fully dedifferentiated chlorophyllous tissues of Zea mays possess a C3-like metabolism.

The effects of factors influencing sunflower protoplast isolation yield, plating efficiency (PE) and the early differentiation into embryoids (embryogenic capacity, EC) have been studied. Only hypocotyl-derived protoplasts divided. The variations of PE and EC in the various treatments did not seem to be linked to the protoplast yields. From statistical analysis of the data, we concluded that, the sunflower genotype, the age and height of seedlings, the part of hypocotyl used, the incubation time (from 6 to 16 hours) in enzymes of explants or of protoplasts alone, influenced PE but large variations were detected for EC. A comparison of the factors effecting EC suggested an origin, inside the hypocotyl, of cells able to give rise, after induction, to embryogenic protoplasts.

We investigated the paraffinic hydrocarbon composition of cell suspensions of Euphorbia characias L. grown in different regimes of carbon and energy supply. Heterotropic (H), photomixotrophic (PM) and photoautotrophic (PA) cell suspensions were compared to leaves. Alkane compositions varied greatly both quantitatively and qualitatively according to the mode of culture. PM and H cells had indentical gaussian-like hydrocarbon distributions, with maxima around C 27 or C 28 compounds, while PA cells and leaf hydrocarbons were enriched in odd carbon chain-length compounds, especially C 29 , C 31 and C 33 In long-term cultivated suspensions, total hydrocarbon amounts of PM and PA cells were about 5% and 6% that of leaf, respectively, and PA suspensions presented a major enrichment of all odd carbon chain-length compounds. Minor differences were observed between the growth phases of PM as well as PA cells. On the other hand, major differences appeared between these two modes of culture and were amplified along subcultures, indicating that the paraffinic hydrocarbon composition was directly related to the mode of culture. We discuss the possible origins of these alkane patterns.

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Wheat straw degradation by Fibrobacter succinogenes was monitored by nuclear magnetic resonance (NMR) spectroscopy and chemolytic methods to investigate the activity of an entire fibrolytic system on an intact complex substrate. In situ solid-state NMR with 13 C cross-polarization magic angle spinning was used to monitor the modification of the composition and structure of lignocellulosic fibers (of 13 C-enriched wheat straw) during the growth of bacteria on this substrate. There was no preferential degradation either of amorphous regions of cellulose versus crystalline regions or of cellulose versus hemicelluloses in wheat straw. This suggests either a simultaneous degradation of the amorphous and crystalline parts of cellulose and of cellulose and hemicelluloses by the enzymes or degradation at the surface at a molecular scale that cannot be detected by NMR. Liquid-state two-dimensional NMR experiments and chemolytic methods were used to analyze in detail the various sugars released into the culture medium. An integration of NMR signals enabled the quantification of oligosaccharides produced from wheat straw at various times of culture and showed the sequential activities of some of the fibrolytic enzymes of F. succinogenes S85 on wheat straw. In particular, acetylxylan esterase appeared to be more active than arabinofuranosidase, which was more active than α-glucuronidase. Finally, cellodextrins did not accumulate to a great extent in the culture medium.

International audience; BACKGROUND: Glucose absorption from starchy food has only been described with small amounts ingested ( approximately 20-75 g). OBJECTIVE: Our aim was to describe total plasma (Ra) and exogenous glucose (Ra(exo)) appearance, glucose release from the liver (HGP), and the metabolic response after ingestion of 5 g polished or parboiled rice/kg body mass. DESIGN: Gas exchange and urea excretion were monitored in 8 healthy subjects before (3.5 h) and after (8 h) ingestion of rice intrinsically labeled with (13)C; [6,6-(2)H(2)]glucose was infused for the measurement of Ra, Ra(exo), and HGP. RESULTS: Changes in plasma glucose, insulin, lactate, and free fatty acids and the increase in Ra(exo) and Ra ( approximately 200%) and the decrease in HGP ( approximately 90%) were not significantly different (P > 0.05) after ingestion of either rice. Glucose oxidation was not significantly different (111.6 +/- 8.2 compared with 89.0 +/- 11.3 g; P = 0.13), but fat oxidation was significantly lower (9.9 +/- 1.7 compared with 21.3 +/- 4.0 g; P < 0.05) after parboiled than after polished rice. The percentage of the glucose load that appeared in the circulation over 8 h was not significantly different after ingestion of polished (70.4 +/- 4.5%) or parboiled (63.8 +/- 2.0%) rice (P > 0.05). CONCLUSION: Although the starch in parboiled rice is less susceptible to digestion in vitro, exogenous glucose availability was not significantly different after ingestion of large amounts of polished or parboiled rice. Glucose absorption remains incomplete 8 h after ingestion of both types of rice.

Photoautotrophic, photomixotrophic and heterotrophic cell suspension cultures of Euphorbia characias were analysed for fatty acid composition of their neutral lipids. Contents in exponential and stationary phases of growth were compared to those of seeds, leaves and calli from which cell suspensions were derived. Quantitatively, on a dry weight basis, neutral lipid amounts varied in the different cells from 15 to 40% of that in the leaves (5.14 mg per g dry wt) but they were very low compared to that in the seeds (330 mg per g dry wt). The highest values were observed during the exponential phase of growth with photosynthetic cell suspensions. Qualitatively, fatty acid distribution was similar in photoautotrophic cell suspensions and leaves with a high content of linolenic acid. The linolenic acid/linoleic acid ratio decreased from photoautotrophic to heterotrophic cell suspensions. In seeds, the maximal amount of linoleic acid is reached while linolenic acid almost disappears. The desaturation of C18 fatty acids is closely linked with the development of the photosynthetic apparatus and with a low growth rate.

Pathways for hydroxycinnamyl aldehyde incorporation into lignins are revealed by examining transgenic plants deficient in cinnamyl alcohol dehydrogenase, the enzyme that converts hydroxycinnamyl aldehydes to the hydroxycinnamyl alcohol lignin monomers. In such plants the aldehydes incorporate into lignins via radical coupling reactions. As diagnostically revealed by long-range (13)C-(1)H correlative NMR, sinapyl aldehyde (3, 5-dimethoxy-4-hydroxy-cinnamaldehyde) 8-O-4-cross-couples with both guaiacyl (3-methoxy-4-hydroxyphenyl-propanoid) and syringyl (3, 5-dimethoxy-4-hydroxyphenyl-propanoid) units, whereas coniferyl aldehyde cross-couples only with syringyl units.

Les connaissances sur les risques lies a l'inhalation de mycotoxines restent limitees, alors que ces toxines, presentes au niveau des spores fongiques, peuvent atteindre directement les epitheliums respiratoires. Les enzymes du metabolisme des xeniobiotiques pulmonaires peuvent moduler la toxicite de ces mycotoxines en les detoxifiant ou en formant des metabolites plus reactifs. Notre objectif etait d'etudier in vitro le metabolisme pulmonaire de la sterigmatocystine et de la methoxy-sterigmatocystine, deux mycotoxines presentes dans les environnements contamines par les moisissures, et de predire les eventuelles consequences toxiques. L'etude du metabolisme en presence de cytochromes P450 exprimes dans des systemes heterologues, puis dans un modele de culture primaire de cellules epitheliales tracheales porcines, a montre que ces mycotoxines etaient oxydees, principalement par les cytochromes P450 1A, et conjuguees par glucucoridation et sulfo-conjugaison. Ces deux toxines semblent principalement detoxifiees au niveau pulmonaire. Si une activation metabolique de la sterigmatocystine est possible, celle-ci semble limitee

The chemical forms of inorganic carbon, CO2 or HCO3-, incorporated during photosynthesis in photoautotrophic Euphorbia characias cell suspension cultures were determined in experiments using 13CO2 and a mass spectrometry technique. From the equations of the CO2 hydration reaction, a kinetic model was first developed, and the effect of photosynthesis on the external CO2 concentration was simulated. It was predicted from this model that CO2 and HCO3- uptakes could be differentiated by recording only the CO2 variation rate in the external medium, successively in absence then in presence of an exogenous carbonic anhydrase activity. The results obtained with either CO2-grown or air-grown photoautotrophic cells were in good agreement with the model and demonstrated that CO2 was the sole species taken up during photosynthesis. In addition no accumulation of inorganic carbon within the cells was observed in the light. Similarly, in dark, CO2 was the only species released by respiration in the external medium.

Modifications of cytoplasmic pH in Acer pseudoplatanus L. cells cultivated in suspension have been induced by acid-loads and studied by using 31P nuclear magnetic resonance spectroscopy. The initial drop of cytoplasmic pH, observed in the first minutes of exposure to weak lipophilic acids, was followed by a slow recovery to reach a plateau phase with a pH value lower than the initial one. Conversely, removal of the acid led to a sharp increase of cytoplasmic pH with in most cases an overshoot toward more alkaline values than the initial one and a subsequent decrease to more acidic values. This shows that A. pseudoplatanus cells powerfully regulate their cytoplasmic pH both on the acid side of their normal pH, during the acid-load, and on the alkaline side, after removal of acid. Similar results were obtained with different types of acid-loads, i.e. treatments with propionic or benzoic acid or bubbling with CO2-enriched air. This indicates that the occurrence of pH regulation does not depend upon the method used to acid-load the cells. The time courses of cytoplasmic pH observed for A. pseudoplatanus and also Catharanthus roseus cells are similar to those recorded for animal cells but different from those described for other plant materials for which no recovery phase was observed. This can be explained by different balances between the initial rate of proton influx brought in by the acids, and the capacity of proton consumption by the regulatory mechanisms. The existence of the recovery phase offers a unique possibility to study the regulation of the cytoplasmic pH of plant cells, as it has been done in animal systems.

The vacuoles of higher plant cells have been shown to be the site for accumulation of ions and metabolites (Matile 1978; Leigh et al. 1979, Leigh 1983; Marty et al. 1980; Boudet et al. 1984). The mechanisms involved in the storage functions of vacuoles are not well known. However, the presence of ATPases on the tonoplast of a number of plant species has been demonstrated (d’Auzac 1977; Lin et al. 1977 ; Leigh and Walker 1980; Cretin 1982; Aoki and Nishida 1984). These ATPases are considered to be responsible for creating a proton electrochemical gradient across the tonoplast. A classical idea is that this gradient could be used for the transport of ions and metabolites and their accumulation in the vacuole. The proton gradient across the tonoplast can also be directly used for the accumulation of secondary plant products diffusing through membranes as neutral molecules and accumulated as cations. This is the case for some indole alkaloids (Renaudin and Guern 1982) and nicotine (Kurkdjian 1982).