Inorganic carbon acquisition by aquatic photolithoatrophs of the Dighty Burn, Angus, U.K.: uses and limitations of natural abundance measurements of carbon isotopes.

The 13 C/12 C ratio (expressed as δ13 C) of benrhic photolithotrophs. in the Dighn Water (= Burn) were measured fur comparison with that of the potential inorganic carhun sources. CO2 and HCO3 - , in the Burn. The Burn water contains an average of 65.7 mmol m-3 CO2 with δ13 C of -14.7% and 1600 mmol m-3 HCO3 - with δ13 C of -4.%. δ13 C values of riparian vegetation were also measured as contributors, after respiration in the soil or the Burn, to the δ13 C of inorganic carbon in the Burn. The potential range of differences in 13 C/12C between dissolved CO2 and plant organic C is set by the intrinsic 13 c/12 C discrimination (α value) in CO2 fixation by Rubisco. Main results and conclusions are. as follows, (i) A literature survey suggests that there is no convincing evidence that the α, Values (rate constant for 12 CO2 fixation relative to that for 13 CO2 fixation by Rubisco in the absence of CO2 transport limitation) for the'lower plants'in the Burn (diatoms, green and red algae, mosses) are significantly different from the well-established αp values for the flowering plum enzyme. (ii) In confirmation of earlier work, the semi-erect 'streamer'gametophytes of the red alga Lemanea mamillosa and the moss Fontinalis antipyetica have δ13 C values which can only be interpreted in terms of diffusive CO2 entry with minimal limitation of photosynthesis by CO- diffusion, (iii) The serui-erect grren alga Cladophora glomerata and the flowering plant Ranunculus penicillatus ssp. pseudofluitons (formerly var. calcareus) are- both able to use HCO3 - . Their δ13 C values indicate that, if the HCO3 - -use system does not (as is likely) discriminate significantly between 13 C and 12 C, then a substantial fraction of the inorganic C made available to Rubisco must return to the medium, carrying 13 C-inorganic C not fixed by Rubisco. (iv) Two sets of δ13 C data from different hydrodynamic regimes distance from leading edge of a flat stone; different size of thalli) show that the attainable differences in situ in thickness of the diffusion boundary layer do not alter the fractional limitation of photosynthesis of Cladophora by external diffusion of inorganic C, considered with HCO3 use. (vi) The entrusting red alga Hildenbrandia rivularis has a δ13 C value suggestive of CO2 as the inorganic C source, but not entirely ruling nut HCO3 - . Marine species of both Hildenbrundia and Cladophora have δ13 C values which, even when corrected for source inorganic C δ13 C values, are 10%, more positive than the freshwater species. (vii) Mats of pennate diatoms were shown by pH-drift to by able to use HCO3 - ; the relatively high (i.e. not very negative) δ12 C value of these mats could relate to a relatively'non-leaky'HCO3 - aequisition mechanism and/or to limitation by external diffusion (e.g. through the mat).

Variations of the natural abundances of nitrogen and carbon isotopes in Triticum aestivum, with special reference to phloem and xylem exudates.

This work explored whether the natural abundances of carbon and nitrogen isotopes could be used to describe the movement of C and X within wheat plants; we also considered whether isotopic analyses of aphids or their honeydew would substitute for direct analysis of phloem exudate. The δ13 C of ears and roots (sinks) most closely matched those of the sugars + organic acids fraction (sources) in both growth stages; phloem δ13 C matched that of leaf blade sugars. Xylem exudate δ13 C matched no other putative (and measured) source in the ear-forming stage and matched that of whole roots and ears in the grain-filling stage. The δ15 N of grain and roots (sinks) resembled that of leaf amino acids (sources) in the ear-forming stage. In the gram-filling stage, ear δ15 N continued to resemble that of leaf amino acids, and δ15 N of roots most closely resembled that of whole leaves. In the grain-filling stage, phloem δ15 N fell between that of leaf blade amino acids and that of whole leaves and was 15 X-depleted relative to internal and external NO, -N. In both growth stages, xylem exudate δ15 N was less than that of soil NO3 - -N and more than that of residual soil N after mineral N extraction. The isotopic values are generally in agreement with data from other approaches, such as isotope labelling; they show NO3 - -N reduction in both shoots and roots of wheat and significant N recycling (root-shoot-phloem-root) and C movement. Aphids might serve as a substitute for isotopic analysis of phloem δ15 N. having the same value as their food source. Their excreta was 15 N-enriched relative to phloem.

Plant roots release phospholipid surfactants that modify the physical and chemical properties of soil.

• Plant root mucilages contain powerful surfactants that will alter the interaction of soil solids with water and ions, and the rates of microbial processes. • The lipid composition of maize, lupin and wheat root mucilages was analysed by thin layer chromatography and gas chromatography-mass spectrometry. A commercially available phosphatidylcholine (lecithin), chemically similar to the phospholipid surfactants identified in the mucilages, was then used to evaluate its effects on selected soil properties. • The lipids found in the mucilages were principally phosphatidylcholines, composed mainly of saturated fatty acids, in contrast to the lipids extracted from root tissues. In soil at low tension, lecithin reduced the water content at any particular tension by as much as 10 and 50% in soil and acid-washed sand, respectively. Lecithin decreased the amount of phosphate adsorption in soil and increased the phosphate concentration in solution by 10%. The surfactant also reduced net rates of ammonium consumption and nitrate production in soil. • These experiments provide the first evidence we are aware of that plant-released surfactants will significantly modify the biophysical environment of the rhizosphere.

Altritol synthesis by Notheia anomala.

The linear hexitol altritol has only been identified in six genera, all of them in the order Fucales of the brown algae. Five of these genera are closely related according to molecular phylogenetic and other data, while the sixth (Notheia) is an obligate epiphyte on two other altritol-containing genera with which it is symphanic. The possibility that Notheia obtains altritol from the algae on which it is epiphytic rather than by synthesizing altritol independently was investigated by supplying 13C-inorganic carbon in the light followed by mass spectrometric and nuclear magnetic resonance analysis. Notheia separated from the phorophyte Hormosira during exposure to 13C showed 13C enrichment in both altritol and mannitol, while the Hormosira only showed significant labelling of mannitol. These data show that altritol can be synthesized by Notheia, with implications for the number of gains and losses of the capacity to synthesize altritol in the evolution of the Fucales.

Identification of anatoxin-A in benthic cyanobacteria (blue-green algae) and in associated dog poisonings at Loch Insh, Scotland.

Dog deaths occurred in 1990 and 1991 after the animals drank water containing blooms of benthic cyanobacteria along the shoreline of Loch Insh, Scotland. Signs of poisoning in the affected animals and the high neurotoxicity of bloom extracts in laboratory bioassays indicated acute poisoning due to cyanobacterial neurotoxin(s). The neurotoxic blooms consisted largely of benthic Oscillatoria species which were also observed in the stomach contents of the poisoned dogs. Stomach contents were also neurotoxic in bioassays with the same signs of poisoning as the Oscillatoria blooms. The cyanobacterial alkaloid neurotoxin anatoxin-a was identified in bloom extracts and poisoned dog stomach contents by high-performance liquid chromatography and gas chromatography-mass spectrometry. A species of benthic Oscillatoria has been isolated from the neurotoxic bloom material and shown to produce anatoxin-a in laboratory culture. These findings are the first to associate anatoxin-a toxicoses with benthic, rather than planktonic, cyanobacteria. Procedures for anatoxin-a extraction and identification from the blooms and animal material are also detailed.

Fatty acids. Part 48. 13C nuclear magnetic resonance studies of acetylenic fatty acids.

The 13C NMR spectra of thirty-seven alkynoic acids (C8-C18) and ten alkadiynoic acids (C18 and C20) are reported and interpreted. The influence of COOH, COOCH3, CH3, and C identical to C groups on the chemical shifts of nearby carbon atoms is assessed. These influences are largely additive so that available spectra are readily interpreted and the spectra of new compounds of this type can be predicted. These preliminary results indicate that 13C NMR spectroscopy should be of considerable value in the structural identification of acetylenic compounds.

Fatty acids of the seed oils of alpineCruciferae and other alpine species.

A fewCruciferae seed oils do not contain erucic acid but are instead rich in linolenic acid. These oils are characteristic of alpine species, and the fatty acid composition of such oils may be an adaption to an alpine habitat.

Wild rabbit host and some parasites show trophic-level relationships for delta 13C and delta 15N: a first report.

We report the first isotopic study of an animal host-parasite system. Parasitic, intestinal nematodes, Graphidium strigosum and Passalurus ambiguus, were 15N-enriched relative to their host, the European rabbit Oryctolagus cuniculus, while parasitic cestodes, Cittataenia denticulata and Mosgovoyia pectinata, were 15N-depleted, suggesting different trophic relationships. Host embryos were more similar in their delta 13C and delta 15N values to maternal muscle than were any of the parasites. Coprophagy, the direct recycling of food by the rabbit eating its own faeces, did not lead to isotopic differences between stomach contents and faeces, suggesting that the major point for isotopic discrimination in lagomorph nitrogen metabolism is in the animal rather than in the gut. We conclude that bulk delta 13C and delta 15N can reveal valuable new information about host-parasite relationships, and these could be explored further at the biochemical level using compound-specific isotopic analyses.

Wild Rabbit Host and Some Parasites Show Trophic-Level Relationships for δ(13)C and δ(15)N: A First Report.

Abstract We report the first isotopic study of an animal host-parasite system. Parasitic, intestinal nematodes, Graphidium strigosum and Passalurus ambiguus, were (15)N-enriched relative to their host, the European rabbit Oryctolagus cuniculus, while parasitic cestodes, Cittataenia denticulata and Mosgovoyia pectinata, were (15)N-depleted, suggesting different trophic relationships. Host embryos were more similar in their δ(13)C and δ(15)N values to maternal muscle than were any of the parasites. Coprophagy, the direct recycling of food by the rabbit eating its own faeces, did not lead to isotopic differences between stomach contents and faeces, suggesting that the major point for isotopic discrimination in lagomorph nitrogen metabolism is in the animal rather than in the gut. We conclude that bulk δ(13)C and δ(15)N can reveal valuable new information about host-parasite relationships, and these could be explored further at the biochemical level using compound-specific isotopic analyses.

Turnover of carbon, nitrogen, and sulfur in bovine longissimus dorsi and psoas major muscles: Implications for isotopic authentication of meat.

Stable isotope ratio analysis of light elements (including C, N, and S) is a powerful tool for inferring the production and geographic origins of animals. The objectives of this research were to quantify experimentally the isotopic turnover of C, N, and S in bovine skeletal muscle (LM and psoas major) and to assess the implications of the turnover for meat authentication. The diets of groups (n = 10 each) of beef cattle were switched from a control diet containing barley and unlabelled urea to an experimental diet containing maize, (15)N-labeled urea, and seaweed for periods of up to 168 d preslaughter. The feeding of the experimental diet was clearly reflected by the delta(13)C, delta(15)N, and delta(34)S values of the LM and psoas major muscles, but isotopic equilibrium was not reached in either muscle for C, N, or S after 168 d of feeding the experimental diet. The slow turnover in skeletal muscle was reflected by the C and N half-lives of 151 and 157 d for LM and 134 and 145 d for psoas major, respectively, and by an S half-life of 219 d in LM. It is concluded that the turnover of light elements (C, N, and S) in bovine skeletal muscles is a slow process; therefore, skeletal muscles contain isotopic information on dietary inputs integrated over a long period of time (months to years).

Translocation of 15N indicates nitrogen recycling in the mat-forming lichen Cladonia portentosa.

Nitrogen translocation was measured in Cladonia portentosa during 2 yr growth in Scottish heathland. Translocation was predicted to occur if N is resorbed from senescent basal tissue and recycled within the thallus. (15)N was introduced into either the lower (TU thalli) or upper (TD thalli) 25 mm of 50-mm-long thalli as (15)N-NH(4) (+), (15)N-NO(3) (-) or (15)N-glycine. Labelled thalli were placed within intact lichen cushions, either upright (TU) or inverted (TD). Vertical distribution of label was quantified immediately following labelling and after 1 and 2 yr. Independently of the form of introduced label, (15)N migrated upwards in TU thalli, with new growth being a strong sink. Sink regions for (15)N during year 1 (including new growth) became sources of (15)N translocated to new growth in year 2. Upward migration into inverted bases was minimal in TD thalli, but was again marked in new growth that developed from inverted apices. Relocation of N to regions of growth could facilitate internal N recycling, a process postulated to explain the ecological success of mat-forming lichens.

Automated measurement of the concentration and 13C enrichment of carbon dioxide in breath and blood samples using the Finnigan MAT Breath Gas Analysis System.

The operation and performance of the Finnigan MAT automated Breath Gas Analysis System for the routine analysis of 13C breath tests taken in 20-ml Vacutainers is described. Up to four samples per hour can be analysed with a standard deviation of delta 13C of less than 0.05% being achieved over the range 10-50 mumol CO2. The equipment can also be used for the automatic measurement of the concentration and 13C enrichment of CO2 derived from carbonate or bicarbonate solutions, including blood and other physiological fluids.

Carbon and nitrogen stable isotope ratios in body tissue and mucus of feeding and fasting earthworms (Lumbricus festivus).

We used natural abundance stable isotope techniques to estimate carbon and nitrogen turnover rates in body tissue and mucus of earthworms. Isotope ratios of carbon (delta(13)C) and nitrogen (delta(15)N) were monitored simultaneously in body tissue and mucus for up to 101 days in feeding or fasting Lumbricus festivus kept in an artificial substrate. When the diet of the earthworms was switched from clover (C(3) plant, legume) to maize (C(4), non-legume), the new dietary delta(13)C signature manifested itself much more rapidly in the mucus than in the body tissue of the animals, causing a delta(13)C shift of about 4 per thousand in mucus and 1 per thousand in tissue after 13.5 days. Turnover of earthworm body tissue carbon, unlike that of mucus carbon, was described adequately by an exponential, single-pool model. Nitrogen turnover could not be assessed because the delta(15)N difference between sources was too small. Fasting for 56 days did not result in the expected whole-body (15)N or (13)C enrichment, but it caused a significant decrease in mucus and tissue C:N ratios and in the ratio (mucus C:N ratio):(tissue C:N ratio). We conclude that the separate analysis of body tissue and mucus has great potential for studying the ecophysiology, feeding ecology and role in elemental cycling of earthworms and other invertebrates.

Adequacy of food rations in soldiers during exercise in hot, day-time conditions assessed by doubly labelled water and energy balance methods.

The energy requirements of people doing physical work in hot climates are not clearly understood. In particular, we know little about the combined effects of heat stress and muscular work on energy requirements. During military exercises in the African bush, soldiers are supplied with standard rations, the adequacy of which is unknown. We have now assessed the adequacy of these food and water rations in 12 male Zimbabwean soldiers during 12 days of strenuous, heat-stress exercise in the field. We used two methods to measure energy expenditure: the double-labelled water method (DLW) and the energy balance method (i.e. comparing dietary energy with changes, if any, in body energy stores). Two groups were studied: one group (eight subjects) carried out field exercises; the control group consisted of four soldiers doing normal work. Mean daily energy expenditure as assessed by the DLW method was [mean (SE)] 23 (1.5) MJ x day(-1) for the field group and 14 (0.5) MJ x day(-1) for the control group (P < 0.001). By the energy balance method, daily energy expenditure was calculated to be 26 (0.7) MJ x day(-1) and 15.5 (0.4) MJ x day(-1) for the field group and control group, respectively. Body mass loss was 3 (0.1) kg [4.6 (0.3)% of body mass] for the field group, but the control group gained 1.1 (0.1) kg. Mean daily fluid intake was 11 (0.5) 1 x day(-1), suggesting that the standard ration supplied was inadequate. Body mass loss was caused by both the energy deficit and total body water loss. These results suggest strenuous work in hot, dry field conditions imposes extra energy requirements.

Automated measurement of 13C enrichment in carbon dioxide derived from submicromole quantities of L-(1-13C)-leucine.

The 13C enrichment of the carboxyl carbon of leucine was measured by isotope ratio mass spectrometry after conversion to CO2 by reaction with ninhydrin in a Vacutainer and cryogenic purification using the Finnigan MAT Breath Gas Analysis System designed for processing 13C breath test samples. The sources of error which arise with submicromole samples are examined and corrections provided for suboptimal mass spectrometer signals and contamination of the evolved CO2 with CO2 from the reaction medium. The main limitations to the accuracy and precision of the method are not instrumental but arise from the contamination with residual CO2 in the reaction medium, and this sets a lower limit of around 0.25 mumol leucine on the practical sample size. This is an improvement of about five-fold on the previous manual method of CO2 isolation.

Body water measurement in growth disorders: a comparison of bioelectrical impedance and skinfold thickness techniques with isotope dilution.

Total body water was estimated as part of the assessment of body composition in children with growth disorders, using the newly commercially available method of bioelectrical impedance. This was undertaken to compare the precision and accuracy of the results with those derived from skinfold thickness against measurement of stable isotopically labelled water (H2(18)O) dilution as a standard. The comparisons were carried out to see to what extent the impedance method could be applied with confidence to assessment of children with growth disorders. Total body water was derived from impedance (I) using an association with height (Ht2/I). Impedance and skinfold thickness estimates of total body water were equally precise when compared with values obtained from H2(18)O dilution (limits of agreement -1.9 to +1.3 and -1.7 to +2.0 kg respectively). The mean intraobserver coefficient of variation for repeat measurements of impedance was 0.9% compared with 4.6% for skinfold thickness with an interobserver coefficient of variation for impedance of 2.8%. Bioelectrical impedance estimation of body composition is likely to be of value in the growth clinic when expertise in measurement of skinfold thickness is limited or repeated measurements are to be undertaken by different observers.

Energy expenditure in type 2 diabetic patients on metformin and sulphonylurea therapy.

Insulin and sulphonylurea therapies have both been reported to cause weight gain in Type 2 diabetic patients whereas metformin does not have this adverse effect. The mechanism for this difference is unclear. We have investigated in a cross-over study the effect of sulphonylurea and metformin therapy on energy expenditure and body composition in 10 Type 2 diabetic patients (7 females, 3 males) of various weights (mean body mass index 33.4 (SD 7.6 kg m-2)). Free living total energy expenditure was measured over 14 days by the doubly labelled water method adjusted for urinary glucose energy losses and resting energy expenditure by ventilated hood indirect calorimetry. Overall, total energy expenditure (12.88 +/- 4.17 vs 13.1 +/- 3.69 MJ 24 h-1) and resting metabolic rate (7.30 +/- 1.75 vs 7.23 +/- 1.74 MJ 24 h-1) were similar on metformin and sulphonylurea therapy, respectively. When adjusted for differences in fat free mass, resting metabolic rate on sulphonylurea therapy was slightly but significantly lower (mean difference -5.5 kJ 24 h-1 kg-1, 95% CI -1.2, -9.9 kJ 24 h-1 kg-1, p < 0.05). Fat free mass also increased significantly by 1.3 kg (95% CI 0.4, 2.4 kg, p < 0.05) when on sulphonylurea therapy, thus compensating for the lower resting metabolic rate per kg fat free mass to leave overall resting metabolic rate unchanged compared to metformin therapy. We also investigated the effect of adding metformin to six Type 2 diabetic patients already on insulin. This did not lead to any measurable changes in any of the components of energy expenditure.(ABSTRACT TRUNCATED AT 250 WORDS)

Recovery of 13C in breath from infused NaH13CO3 increases during euglycaemic hyperinsulinaemia.

1. The effect of euglycaemic hyperinsulinaemia on the recovery of 13C in expired CO2 has been assessed in six normal subjects. Each was studied on three occasions: once with a 6 h primed constant infusion of NaH13CO3 combined with a euglycaemic hyperinsulinaemic clamp for the last 3 h (study 1), once with a 6 h primed constant infusion of NaH13CO3 alone (study 2) and once with a 6 h infusion of normal saline combined with a hyperinsulinaemic clamp for the last 3 h (study 3). Measurements of 13C enrichment of expired CO2 were made in the third and sixth hour of each infusion. 2. There was no significant increase in enrichment during study 3 (3 h 0.00047 +/- 0.00016 versus 6 h 0.00069 +/- 0.00028 atom per cent excess) with potato-starch-derived D-glucose used to maintain euglycaemia. 13C recovery increased in the sixth hour of both study 1 and 2 (study 1: 3 h 74.4 +/- 2.0 versus 6 h 85.5 +/- 2.6%, P < 0.01; study 2: 3 h 72.1 +/- 2.4 versus 6 h 81.7 +/- 1.4%, P < 0.01). There was no significant difference in recovery between studies 1 and 2. 3. These results suggest that increased recovery during a sequential euglycaemic clamp is predominantly time-dependent. Studies which use this technique to examine the effect of insulin on substrate oxidation should take this into account.