Insolubility of Cr2O3 in Bioaccessibility Tests Points to Requirement for a New Human Oral Reference Dose for Trivalent Chromium

Bioaccessibility measurements have the potential to improve the accuracy of risk assessments and reduce the potential costs of remediation when they reveal that the solubility of chemicals in a matrix (e.g., soil) differs markedly from that in the critical toxicity study (i.e., the key study from which a toxicological or toxicity reference value is derived). We aimed to apply this approach to a brownfield site contaminated with chromium, and found that the speciation was Cr^III, using a combination of alkaline digestion/diphenylcarbazide complexation and X-ray absorption near edge structure analysis. The bioaccessibility of Cr₂O₃, the compound on which a reference dose for Cr^III is based, was substantially lower (<0.1%) than that of the Cr^III in the soils, which was a maximum of 9%, giving relative bioaccessibility values of 13,000% in soil. This shows that the reference dose is based on essentially an insoluble compound, and thus we suggest that other compounds be considered for toxicity testing and derivation of reference dose. Two possibilities are CrCl₃·6H₂O and KCr(SO₄)₂·12H₂O, which have been used for derivation of ecological toxicity reference values and are soluble at a range of dosing levels in our bioaccessibility tests.     

THE ABSORPTION OF P,P′-DDE BY LARVAE OF AGLAIS URTICAE (LEPIDOPTERA: NYMPHALIDAE)

Fifth instar larvae of Aglais urticae L. absorb single and double doses of 27.5 g of p,p-DDE at a constant rate 8–72 hours after dosing. There is little difference between the rates of absorption by the blood of larvae given doses on two sites, and the sum of the rates of absorption by the blood of larvae given single doses, so that the blood does not limit the rate at which apolar substances pass through the cuticle.

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Zusammenfassung Die Rate, mit welcher p,p-DDE durch die Kutikula des 5.Larvenstadiums von A. urticae eindringt, wurde durch direkte Messung der absorbierten Menge bestimmt: Verluste durch Stoffwechsel und Exkretion waren zu vernachlässigen. Die Raupen erhielten entweder einzelne Dosen von 27,5 g DDE in 1 l 1,4-Dioxan auf den Thorax oder das Abdomen, oder zwei Dosen, eine auf den Thorax und eine auf das Abdomen. Die Larven wurden 8, 24 und 72 Stunden nach der Applikation auf DDE untersucht. Die Aufnahmerate durch die Larven, die zwei Dosen erhalten hatten, war mit 0,6–1,1 ppm/Std ähnlich der Summe der Aufnahmeraten durch die Larven, denen einzelne Dosen auf Thorax oder Abdomen verabreicht worden waren, so daß die Aufnahmerate wahrscheinlich völlig durch Faktoren innerhalb der Kutikula begrenzt wird und die Absorptionskapazität des Blutes für diese apolare Substanz keinen entscheidenden Faktor darstellt.

     

Phylogeny of Greya (Lepidoptera: Prodoxidae), based on nucleotide sequence variation in mitochondrial cytochrome oxidase I and II: congruence with morphological data

The phylogeny of Greya Busck (Lepidoptera: Prodoxidae) was inferred from nucleotide sequence variation across a 765-bp region in the cytochrome oxidase I and II genes of the mitochondrial genome. Most parsimonious relationships of 25 haplotypes from 16 Greya species and two outgroup genera (Tetragma and Prodoxus) showed substantial congruence with the species relationships indicated by morphological variation. Differences between mitochondrial and morphological trees were found primarily in the positions of two species, G. variabilis and G. pectinifera, and in the branching order of the three major species groups in the genus. Conflicts between the data sets were examined by comparing levels of homoplasy in characters supporting alternative hypotheses. The phylogeny of Greya species suggests that host-plant association at the family level and larval feeding mode are conservative characters. Transition/transversion ratios estimated by reconstruction of nucleotide substitutions on the phylogeny had a range of 2.0-9.3, when different subsets of the phylogeny were used. The decline of this ratio with the increase in maximum sequence divergence among taxa indicates that transitions are masked by transversions along deeper internodes or long branches of the phylogeny. Among transitions, substitutions of A-->G and T-->C outnumbered their reciprocal substitutions by 2-6 times, presumably because of the approximately 4:1 (77%) A+T-bias in nucleotide base composition. Of all transversions, 73%-80% were A<-->T substitutions, 85% of which occurred at third positions of codons; these estimates did not decrease with an increase in maximum sequence divergence of taxa included in the analysis. The high frequency of A<-->T substitutions is either a reflection or an explanation of the 92% A+T bias at third codon positions.      

Validity of the tritiated thymidine method for estimating bacterial growth rates: measurement of isotope dilution during DNA synthesis.

The rate of tritiated thymidine incorporation into DNA was used to estimate bacterial growth rates in aquatic environments. To be accurate, the calculation of growth rates has to include a factor for the dilution of isotope before incorporation. The validity of an isotope dilution analysis to determine this factor was verified in experiments reported here with cultures of a marine bacterium growing in a chemostat. Growth rates calculated from data on chemostat dilution rates and cell density agreed well with rates calculated by tritiated thymidine incorporation into DNA and isotope dilution analysis. With sufficiently high concentrations of exogenous thymidine, de novo synthesis of deoxythymidine monophosphate was inhibited, thereby preventing the endogenous dilution of isotope. The thymidine technique was also shown to be useful for measuring growth rates of mixed suspensions of bacteria growing anaerobically. Thymidine was incorporated into the DNA of a range of marine pseudomonads that were investigated. Three species did not take up thymidine. The common marine cyanobacterium Synechococcus species did not incorporate thymidine into DNA.     

In vivo and in vitro inhibition of rat liver vitamin D3-25-hydroxylase activity by 19-hydroxy-10(S),19-dihydrovitamin D3

Rats treated with varying amounts of 19-hydroxy-10(S),19-dihydrovitamin D3 prior to administration of physiologic doses of vitamin D3 exhibit normal intestinal calcium transport but are unable to mobilize bone calcium. In contrast, 19-hydroxy-10(R),19-dihydrovitamin D3 had no inhibitory activity. Circulating serum levels of 25-hydroxy[3H]vitamin D3 and 1 alpha, 25-dihydroxy[3H]vitamin D3 are markedly suppressed but not totally eliminated in animals predosed with 19-hydroxy-10(S),19-dihydrovitamin D3 before [3H]vitamin D3. Hepatic 25-hydroxy[3H]vitamin D3 levels were approximately equal in both 19-hydroxy-10(S),19-dihydroviotamin D3 treated and untreated rats. However, the rate of conversion of [3H]vitamin D3 to 25-hydroxyvitamin D3 in vivo is greatly reduced in the treated rats. The inhibitory vitamin analogue was also show to block hepatic microsomal 25-hydroxylation in vitro. These results indicate that 19-hydroxy-10(S),19-dihydrovitamin D3 is a specific inhibitor for a hepatic microsomal vitamin D3-25-hydroxylase system.     

Cytochemical analysis of intracellular calcium distribution in the anterior pituitary of the rat

Summary In an attempt to assign morphologic identities to previously distinguished functional calcium compartments in the anterior pituitary of the rat, we employed the potassium pyroantimonate technique for cation localization. Tissues were incubated for In at 37°C in control medium; with 10mM theophylline; or with depolarizing amounts of potassium. Precipitate was quantified on photomicrographs of tissue prepared for electron microscopy with a Talos Systems Digitizer. The nature of the electron dense precipitate was dependent on the experimental state of the tissue. Treatment with 5 mM EGTA abolished the dense precipitate. Electron microprobe analysis also confirmed that calcium was the predominant cation in the observed precipitate. The most significant changes in precipitate deposition occurred along the plasma membrane, the limiting membrane of secretory granules and within mitochondria. Dense precipitate was present along the plasma membrane only in cells treated with potassium. Control tissue exhibited higher levels of precipitate associated with the limiting membrane of secretory granules than either theophylline-treated or potassium-treated tissue. Mitochondria contained more precipitate in potassium-treated tissue than in controls; the mitochondria of theophylline-treated tissue contained intermediate levels of precipitate. Addition of either theophylline or depolarizing amounts of potassium has been associated with hormone secretion in anterior pituitary tissue of normal rats. Kinetic studies in our laboratory indicate that intracellular calcium shifts occur. The pyroantimonate technique is useful in verifying morphologically the calcium compartments involved in shifts in intracellular calcium.