The radium legacy: Contaminated land and the committed effective dose from the ingestion of radium contaminated materials.

The manufacture and use of radium in the early to mid-20th century within industrial, medicinal and recreational products have resulted in a large number of contaminated sites across a number of countries with notable examples in the USA and Europe. These sites, represent a significant number of unregulated sources of potential radiological exposure that have collectively and hitherto not been well characterised. In 2007, the Radioactive Contaminated Land (RCL) Regulations came into force in the UK, providing the statutory guidance for regulators to classify and deal with RCL. Here we report on results derived from digestion experiments to estimate committed effective dose, a key aspect of the RCL Regulations, from the ingestion of radium contaminated sources that can be found in the environment. This case study includes particles, clinker and artefacts that arise from past military activities on a site that was once an airfield at Dalgety Bay on the Firth of Forth, UK. Since 2011 the number of radium contaminated finds has increased by one order of magnitude on the foreshore areas of Dalgety Bay. The increase in finds may in large part be attributed to a change in monitoring practice. A subsample of sixty sources was selected, on the basis of their activity and dimensions, and subjected to digestion in simulated stomach and lower intestine solutions. The study demonstrated that more radium-226 ((226)Ra) and lead-210 ((210)Pb; driven by Polonium solubility) are dissolved from sources in artificial 'stomach' solutions compared with 'lower intestine' solutions. The combined 'gut' solubility for (226)Ra and apparent (210)Pb varied from less than 1% to up to 35% ICRP 72 conversion factors were used to convert the activities measured in solution to committed effective dose. A little over 10% of the sources tested dissolved sufficient radioactivity to result in 100mSv committed effective dose to an infant. Using the solubility of 35% as a worst case, minimum source activities necessary to deliver 100mSv to the full age range of users of the foreshore were estimated. All the estimated activities have been detected and recovered through routine monitoring.

Optimal mapping of terrestrial gamma dose rates using geological parent material and aerogeophysical survey data.

Regulatory authorities need ways to estimate natural terrestrial gamma radiation dose rates (nGy h⁻¹) across the landscape accurately, to assess its potential deleterious health effects. The primary method for estimating outdoor dose rate is to use an in situ detector supported 1 m above the ground, but such measurements are costly and cannot capture the landscape-scale variation in dose rates which are associated with changes in soil and parent material mineralogy. We investigate the potential for improving estimates of terrestrial gamma dose rates across Northern Ireland (13,542 km²) using measurements from 168 sites and two sources of ancillary data: (i) a map based on a simplified classification of soil parent material, and (ii) dose estimates from a national-scale, airborne radiometric survey. We used the linear mixed modelling framework in which the two ancillary variables were included in separate models as fixed effects, plus a correlation structure which captures the spatially correlated variance component. We used a cross-validation procedure to determine the magnitude of the prediction errors for the different models. We removed a random subset of 10 terrestrial measurements and formed the model from the remainder (n = 158), and then used the model to predict values at the other 10 sites. We repeated this procedure 50 times. The measurements of terrestrial dose vary between 1 and 103 (nGy h⁻¹). The median absolute model prediction errors (nGy h⁻¹) for the three models declined in the following order: no ancillary data (10.8) > simple geological classification (8.3) > airborne radiometric dose (5.4) as a single fixed effect. Estimates of airborne radiometric gamma dose rate can significantly improve the spatial prediction of terrestrial dose rate.

Observations of Fukushima fallout in Great Britain.

Following the Fukushima accident in March 2011, grass samples were collected from 42 sites around Great Britain during April 2011. Iodine-131 was measurable in grass samples across the country with activity concentrations ranging from 10 to 55 Bq kg(-1) dry matter. Concentrations were similar to those reported in other European countries. Rainwater and some foodstuffs were also analysed from a limited number of sites. Of these, (131)I was only detectable in sheep's milk (c. 2 Bq kg(-1)). Caesium-134, which can be attributed to releases from the Fukushima reactors, was detectable in six of the grass samples (4-8 Bq kg(-1) dry matter); (137)Cs was detected in a larger number of grass samples although previous release sources (atmospheric weapons test and the 1986 Chernobyl and 1957 Windscale accidents) are likely to have contributed to this.

Estimating sediment and caesium-137 fluxes in the Ribble Estuary through time-series airborne remote sensing.

High spatial and temporal resolution airborne imagery were acquired for the Ribble Estuary, North West England in 1997 and 2003, to assess the application of time-series airborne remote sensing to quantify total suspended sediment and radionuclide fluxes during a flood and ebb tide sequence. Concomitant measurements of suspended particulate matter (SPM) and water column turbidity were obtained during the time-series image acquisition for the flood and ebb tide sequence on the 17th July 2003 to verify the assumption of a vertically well mixed estuary and thus justifying the vertical extrapolation of spatially integrated estimate of surface SPM. The ¹³7Cs activity concentrations were calculated from a relatively stable relationship between SPM and ¹³7Cs for the Ribble Estuary. Total estuary wide budgets of sediment and ¹³7Cs were obtained by combining the image-derived estimates of surface SPM and ¹³7Cs with estimates of water volume from a two-dimensional hydrodynamic model (VERSE) developed for the Ribble Estuary. These indicate that around 10,000 tons of sediment and 2.72 GBq of ¹³7Cs were deposited over the tidal sequence monitored in July 2003. This compared favourably with bed height elevation change estimated from field work. An uncertainty analysis on the total sediment and ¹³7Cs flux yielded a total budget of the order of 40% on the final estimate. The results represent a novel approach to providing a spatially integrated estimate of the total net sediment and radionuclide flux in an intertidal environment over a flood and ebb tide sequence.

Exact mass measurement of polar organic molecules at low resolution using electrospray ionization and a quadrupole mass spectrometer.

Hitherto, exact mass measurement experiments have usually been performed using high-resolution mass spectrometry. However, under the right circumstances, measurements with comparable accuracy may be made at low resolution. Here we demonstrate the use of a low-resolution single-quadrupole mass spectrometer to accurately mass measure organic samples analyzed by electrospray ionization, using a variety of glycol polymers for internal calibration. Results are presented from 11 samples which yield molecular signals in the m/z range 190-750, including data for positive, negative, and multiply-charged sample and reference ions. Replicate determinations of the masses of 12 ions gave values within 4.5 ppm (1.1 millimass units, mmu) of their calculated values, with standard deviations no larger than 3 ppm (1.7 mmu). From a total of 88 individual 1 min measurements, 83 were within 5 ppm, and 87 within 2 mmu of the theoretical mass. The accuracy, precision, and sensitivity shown here are comparable to those achievable using high-resolution mass spectrometers, with the added benefits of simpler instrumentation and analytical technique afforded by the quadrupole mass analyzer.

Tracheal cytotoxin structural requirements for respiratory epithelial damage in pertussis.

The respiratory epithelial pathology of pertussis (whooping cough) can be reproduced by tracheal cytotoxin (TCT), a disaccharide-tetrapeptide released by Bordetella pertussis. TCT is a muramyl peptide, a class of peptidoglycan-derived compounds which have many biological activities including adjuvanticity, somnogenicity, pyrogenicity, and cytotoxicity. The structural requirements for muramyl peptides to produce some of these biological effects have been partially characterized. Using in vitro assays with respiratory epithelial cells and tissue, we have previously determined that the disaccharide moiety of TCT is not involved in toxicity and that the side-chain functional groups of diaminopimelic acid (A2pm) are crucial for toxicity. In this study, we determine the importance of every amino acid, functional group and chiral centre in the peptide portion of TCT. Although lactyl tetrapeptides are the most toxic of the TCT fragments, producing dose-response curves identical to TCT, the smallest analogues of TCT which are active in our assay are of the form X-gamma-(D)-Glu-meso-A2pm, where X may be an amino acid or a blocking group. Within this active substructure, main-chain chirality and all functional groups are essential for toxicity. This definition of the core region of TCT indicates that the TCT interaction site is unlike almost all other muramyl peptide interaction sites for which structure-activity data are available.

Improved detection limits for fast atom bombardment mass spectrometry: A study of time-dependent desorption using a model pulsed bombardment ionization method.

Certain sample preparations for fast atom bombardment (FAB) yield an intense but brief pulse of sample ions at the onset of ionization. A model system is used to study this phenomenon. This system utilizes a conventional source of a constant flux of fast atoms and a probe that permits mechanical movement of the sample stage. This is used to simulate the effect of pulsing the atom beam. Experiments with model samples and selected ion monitoring detection show that, following bombardment initiation, sample ions are preferentially desorbed with respect to ions from the FAB matrix. Exhibition of preferential sample desorption depends upon the analyte behaving as a surfactant in the selected matrix. When used in conjunction with an array detector that permits data collection in a time-resolved manner, this system allows collection of useful mass spectra with significantly enhanced sensitivity compared to normal bombardment. When applied to the undecapeptide eledoisin (sequence pyro-EPSKDAFIGLM-NH2, MW 1187.6 Da) this novel methodology allows an improvement in detection limit of at least three to four orders of magnitude over that observed when using conventional continuous FAB and a point detector.

Isolation and characterization of opioid peptides from rabbit cerebellum.

The rabbit cerebellum has been shown to contain significant quantities of opioid receptors consisting of both mu- and kappa-subtypes. To determine the nature of the endogenous opioid ligands in this tissue, extracts from rabbit cerebellum were separated by various chromatography techniques and fractions were assayed initially for opioid peptides with a radioimmunoassay capable of detecting all peptides with an amino-terminal Tyr-Gly-Gly-Phe sequence. This sequence is common to all mammalian opioid peptides and is critical for recognition by all known opioid receptors. Each of the three immunoreactive opioid peptide peaks detected was purified to homogeneity and subjected to amino acid composition and sequence analysis. One peak was analyzed further by mass spectrometry. This identified the major opioid peptides in the cerebellum as [Met5]enkephalin, [Leu5]enkephalin, and heptapeptide [Met5]enkephalyl-Arg6-Phe7. The comprehensiveness of this initial detection scheme in identifying biologically active opioid peptides was substantiated through subsequent analysis. Using specific radioimmunoassays for representative opioid peptides of the three opioid systems currently known, no other peptides of either the proenkephalin, proopiomelanocortin, or prodynorphin series were detected in any appreciable amounts. Collectively, these results are consistent with the position that rabbit cerebellar opioids are derived from proenkephalin. However, given that no appreciable quantities of either [Met5]enkephalyl-Arg6-Arg7-Val8-NH2 (metorphamide) or [Met5]enkephalyl-Arg6-Gly7-Leu8 were detected suggests that rabbit proenkephalin may have a slightly altered sequence and/or is differentially processed relative to other mammalian species studied.

Readily synthesized calibration compounds for quadrupole and magnetic instruments for use over the mass range to 2000 daltons.

Volatile mass calibration standards have been prepared by esterifying beta-cellobiose (4-beta-D-glucopyranosyl-D-glucose) with a mixture of heptafluorobutyric (HFB) anhydride and pentafluoropropionic (PFP) anhydride. The mixed esters produce spectra that are useful for mass spectrometer calibration with positive or negative ion methane chemical ionization and electron impact over the mass range 300-2000. The spectra contain prominent ions spaced at m/z 20 and m/z 50 intervals. Using the mixed ester with direct insertion probe introduction gives intense spectra that persist for tens of minutes. All signals above m/z 194 derived from these substances disappear rapidly upon withdrawal of the probe. The composition and exact masses are given for the positive and negative ion spectra of a mixed HFB/PFP ester of beta-cellobiose. Two other calibrants are described: one made from beta-cellobiose using a mixture of HFB, PFP and trifluoroacetic anhydrides, and another the HFB/PFP mixed ester of perseitol. These are examples of the flexibility of this approach with respect to mass range and ion composition.

Reconstruction of the immunogenic peptide RNase(43-56) by identification and transfer of the critical residues into an unrelated peptide backbone.

The involvement of each of the amino acid residues of the I-Ak-restricted T cell determinant RNase(43-56) was examined in detail using a series of peptides containing single amino acid substitutions. Four positions were identified as being essential for the formation of the determinant, Phe-46, Val-47, His-48, and Leu-51. When these four residues were substituted into the backbone of the unrelated peptide HA(130-144), a nonstimulatory peptide was obtained. The inclusion of an additional residue, Val-54, resulted in a chimeric peptide, RN/HA2, which was nearly as active as the native molecule. The peptide RN/HA2 was able to prime in vivo for RNase reactivity, confirming that these five residues contained all of the specificity of the RNase(43-56) determinant. The role of three of these critical residues was examined using both a functional competition assay and an in vivo priming assay. It was ascertained that the Phe-46 was directly involved in contacting the TCR, while the His-48 and Leu-51 were either involved in binding to the I-Ak molecule or in determining the conformation of the peptide. Thus, by critically evaluating the contribution of each of the amino acid residues in a T cell determinant, we were able to generate a chimeric peptide only containing 5 of 15 residues from the RNase(43-56) sequence that was functionally identical to the native RNase(43-56) molecule both in vitro and in vivo.

Primary structure of the peptidoglycan-derived tracheal cytotoxin of Bordetella pertussis.

The etiological agent of whooping cough, Bordetella pertussis, destroys the ciliated epithelial cells lining the large airways of infected individuals. This cytopathology can be reproduced in respiratory epithelium by tracheal cytotoxin (TCT), a small peptidoglycan-related molecule purified from the culture supernatant of growing B. pertussis organisms. Using fast atom bombardment mass spectrometry, we analyzed the positive- and negative-ion spectra of the purified, biologically active material and assigned a mass of 921 daltons to TCT. Analysis of fragment ions in these spectra as well as the spectra of the methyl ester and acetylated derivatives of TCT unambiguously defined the primary structure of TCT as N-acetylglucosaminyl-1,6-anhydro-N-acetylmuramylalanyl-gamma- glutamyldiaminopimelylalanine. TCT is therefore identical with the ciliostatic anhydropeptidoglycan monomer released by Neisseria gonorrhoeae and with the neurologically active slow-wave sleep-promoting factor FSu. These and other structurally related glycopeptides containing muramic acid thus form a family of molecules with remarkably diverse biological activities.

Myristic acid is the NH2-terminal blocking group of the 43-kDa protein of Torpedo nicotinic post-synaptic membranes.

The NH2-terminal blocking group of the 43-kDa peripheral membrane protein (43-kDa protein) of Torpedo post-synaptic membranes has been identified as myristic acid. To identify that blocking group pure 43-kDa protein was digested with trypsin and the blocked tryptic peptide was isolated by reverse phase HPLC. That peptide coeluted with and had the same amino acid composition as a synthetic peptide, myristoyl-Gly-Gln-Asp-Gln-Thr-Lys, the structure of the amino terminus predicted from the protein sequence deduced from a cDNA clone. The presence of myristate was confirmed by the precise molecular mass of the peptide, 886.5266, determined by fast atom bombardment mass spectroscopy.

T cell recognition of bovine ribonuclease. Self/non-self discrimination at the level of binding to the I-Ak molecule.

Bovine RNase A specific T-cell hybridomas were generated to study the recognition of foreign Ag by T lymphocytes. One hybrid, TS12, was shown to recognize RNase in association with I-Ak. This hybridoma required bovine RNase to be processed before recognition. The immunogenic determinant on the RNase molecule recognized by TS12 was localized to the tryptic fragment RNase(40-61). All of the stimulatory ability of this determinant was shown to be contained within the synthetic 14mer RNase(43-56). When this segment of bovine RNase was compared with the self murine sequence, only one amino acid difference was found, a substitution of a proline residue at position 50 for a serine residue. This substitution completely abolishes binding to the I-Ak molecule, as shown by both functional and direct binding assays. This finding shows that self/non-self discrimination not only occurs at the level of the T cell, but also can be caused by an inability of the self peptide to associate with a class II molecule.

Isolation of a component from commercial coomassie brilliant blue R-250 that stains rubrophilin and other proteins red on polyacrylamide gels.

Commercially available Coomassie Brilliant Blue R-250 (C.I. 42660) is a popular and useful dye that stains most proteins blue on polyacrylamide gels. Some proteins from brain (rubrophilin), collagens, histones and parotid gland proteins are distinctly red when stained with Coomassie Blue. Commonly used Coomassie Brilliant Blue R-250 preparations may contain more than 30 distinct colored and fluorescent components that can be separated on silica gel chromatographic columns. A specific component has been isolated on silica gel columns that stains rubrophilin and other proline-rich proteins a reddish color. Fast atom bombardment mass spectrometry of the isolated rubrophilin staining principle indicates a molecular weight of 634 as compared to 826 for the major dye in the original Coomassie Brilliant Blue R-250. Infrared spectrometry is consistent with a difference between the rubrophilin staining principle and Coomassie Brilliant Blue R-250 of a toluene sulfonic acid residue.

Methionine-enkephalin and thyrotropin-stimulating hormone are intimately related in the human anterior pituitary.

The tissue distribution and function of opioid peptides in humans is incompletely defined. We report here that, unlike that in other species, the human anterior pituitary gland contains high concentrations of methionine-enkephalin (met-enkephalin). The met-enkephalin immunoreactive material was isolated and identified as authentic met-enkephalin by fast atom bombardment-mass spectrometry and Edman degradation sequencing. The met-enkephalin was localized in a large subpopulation of TSH immunoreactive cells (thyrotrophs). No other proenkephalin-derived opioid peptides were found in the pituitary, and there was no overlap between proopiomelanocortin and met-enkephalin immunoreactive cells. These results suggest that the human anterior pituitary gland contains a novel met-enkephalin precursor and a possible role for met-enkephalin in regulating human thyroid function.

Isolation and characterization of beta-endorphin-(1-9) from human and rat pituitaries.

Using a radioimmunoassay specific for the carboxyl terminus of beta-endorphin-(1-9) large amounts of beta-endorphin-(1-9)-immunoreactive material was detected in the human pituitary. The major peak of immunoreactivity was purified and characterized by fast atom bombardment-mass spectrometry and Edman degradation sequencing as authentic beta-endorphin-(1-9). In the rat pituitary the highest concentration of beta-endorphin-(1-9) immunoreactivity was in the posterior neurointermediate lobe. This material was identified as N-acetyl beta-endorphin-(1-9) by multiple radioimmunoassays, gel chromatography, and reversed-phase high-performance liquid chromatography. Control experiments determined that beta-endorphin-(1-9) was not formed postmortem or during the extraction procedure. These studies suggest that single lysine residues, similar to single arginine residues, are potential sites of posttranslational processing.

Characterization of endorphins from the pituitary of the spiny dogfish Squalus acanthias.

Opioid-like immunoreactive material was extracted from the pituitary and brain of the Spiny Dogfish Shark Squalus acanthias. The immunoreactive material in the pituitary extracts was purified to apparent homogeneity by reverse phase high performance liquid chromatography and subsequently characterized by amino acid analysis, Edman degradation and fast atom bombardment mass spectrometry. The largest opioid-like peptide isolated contained 30 amino acids and showed 80 percent homology with salmon endorphin-II but less than 50 percent homology with human beta-endorphin. Three structural variants of this molecule were also characterized. These variants were shown to be shorter N-terminal fragments, two of which corresponded to cleavage products at the single basic residues arginine and lysine. Cleavage at a single lysine residue has not been reported for posttranslational processing of beta-endorphin in mammals and could represent a modification seen only in lower vertebrates. The remaining fragment corresponded to a loss of 3 residues from the C-terminus of the parent molecule. No alpha-N-acetylated peptides were detected. These results provide the first unequivocal confirmation of beta-endorphin in an elasmobranch and provide evidence of novel N-terminal variants of beta-endorphin.

Isolation and characterization of the inositol cyclic phosphate products of polyphosphoinositide cleavage by phospholipase C. Physiological effects in permeabilized platelets and Limulus photoreceptor cells.

Cleavage of the polyphosphoinositides, catalyzed by phospholipase C purified from ram seminal vesicles, produces phosphorylated inositols containing cyclic phosphate esters (Wilson, D. B., Bross, T. E., Sherman, W. R., Berger, R. A., and Majerus, P. W. (1985) Proc. Natl. Acad. Sci. U. S. A. 82, 4013-4017). In the present study we describe the isolation and characterization of inositol 1:2-cyclic 4-bisphosphate and inositol 1:2-cyclic 4,5-trisphosphate, the two cyclic phosphate products of phospholipase C catalyzed cleavage of phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate, respectively. We established the structures of these two cyclic compounds through 18O labeling of phosphate moieties, phosphomonoesterase digestion, and fast atom bombardment-mass spectrometry. We examined the physiological effects of these compounds in two systems: saponin-permeabilized platelets loaded with 45Ca2+ and intact Limulus photoreceptors. Both inositol 1:2-cyclic 4,5-trisphosphate and the noncyclic inositol 1,4,5-trisphosphate, but not inositol 1:2-cyclic 4-bisphosphate, release 45Ca2+ from permeabilized platelets in a concentration-dependent manner. Injection of inositol 1:2-cyclic 4,5-trisphosphate into Limulus ventral photoreceptor cells induces both a change in membrane conductance and a transient increase in intracellular calcium ion concentration similar to those induced by light. We injected inositol 1,4,5-trisphosphate and inositol 1:2-cyclic 4,5-trisphosphate into the same photoreceptor cell and found that the cyclic compound is approximately five times more potent than the noncyclic compound in stimulating a conductance change. We speculate that inositol 1:2-cyclic 4,5-trisphosphate may function as a second messenger in stimulated cells.

Fast atom bombardment (FAB) mass spectrometry: a mass spectral investigation of some of the insulins.

Mass measurements of the protonated molecules [M + H]+ of four insulins are presented. In addition, structurally significant fragment ions are observed in the mass spectrum and metastable scanning has been used to link these ions to the protonated molecule.