THE EPIDERMAL CELL KINETIC RESPONSE TO ULTRAVIOLET B IRRADIATION COMBINES REGENERATIVE PROLIFERATION AND CARCINOGEN ASSOCIATED CELL CYCLE DELAY

The cell cycle traverse of epidermal basal cells 24 h after in vivo exposure of ultraviolet B (UVB) irradiation was studied by immunochemical staining of incorporated bromodeoxyuridine (BrdU) and bivariate BrdU/DNA flow cytometric analysis. The results were compared with the cell kinetic patterns following topical application of the skin carcinogen methylnitrosourea (MNU) as well as the skin irritant cantharidin. Hairless mice were injected intraperitoneally with BrdU 24 h after treatment of their back skin with either a minimal erythema dose of UVB, or a single application of MNU or cantharidin dissolved in acetone. The cell cycle traverse of the BrdU-labelled cohorts of epidermal basal cells were then followed for the subsequent 12 h. At 6 h after BrdU-injection, when all labelled cells in the control group as well as in the cantharidin group had left the S phase, the bivariate distributions of the UVB-exposed and the MNU group showed that BrdU-positive cells were still present in S phase. Hence, UVB irradiation, similar to the carcinogen MNU, prolonged the S phase duration in some of the basal cells. At 12 h after pulse labelling, however, BrdU-positive cells from UVB-exposed mice were re-entering S phase from G1 phase, indicating that UVB irradiation induced a shortening of the cell cycle time as well, similar to the response observed after cantharidin. The present data can not tell whether these cells also were delayed in S phase. Thus, the cell cycle traverse in hairless mouse epidermis 24 h after in vivo exposure to UVB seemed to be a combination of the cell kinetic effects following chemical skin carcinogens and skin irritants. UVB irradiation induced both a delay in transit time through S phase, probably due to DNA damage and subsequent repair, as well as a reduction in the total cell cycle time consistent with rapid regenerative proliferation.     

A relativistic 4-component general-order multi-reference coupled cluster method: initial implementation and application to HBr

We present the initial implementation of a determinant-based general-order coupled cluster method which fully accounts for relativistic effects within the four-component framework. The method opens the way for the treatment of multi-reference problems through a state-selective expansion of the model space. The evaluation of the coupled cluster vector function is carried out via relativistic configuration interaction expansions. The implementation is based on a large-scale configuration interaction technique, which may efficiently treat long determinant expansions of more than 10⁸ terms. We demonstrate the capabilities of the new method in calculations of complete potential energy curves of the HBr molecule. The inclusion of spin–orbit interaction and higher excitations than coupled cluster double excitations, either by multi-reference model spaces or the inclusion of full iterative triple excitations, lead to highly accurate results for spectral constants of HBr. An erratum to this article can be found at     

Combination of LC-ICP-MS, LC-MS and NMR for investigation of the oxidative degradation of selenomethionine

Selenomethionine (SeMet) was oxidized by heating an acidic solution with hydrogen peroxide. Samples were taken before and during the oxidation process. The oxidation products were separated by cation exchange chromatography followed by ICP-MS detection to identify the selenium containing compounds as well as electrospray ionization MS detection to determine the masses of the degradation products. Furthermore, the samples were analyzed by ⁷⁷Se-NMR. The first appearing degradation product was selenomethionine selenoxide, which was converted via the deaminated selenoxide to methane seleninic acid and selenite.     

The puromycin route to assess stereo- and regiochemical constraints on peptide bond formation in eukaryotic ribosomes

We synthesized a series of puromycin analogues to probe the chemical specificity of the ribosome in an intact eukaryotic translation system. These studies reveal that both d-enantiomers and beta-amino acid analogues can be incorporated into protein, and provide a quantitative means to rank natural and unnatural residues. Modeling of a d-amino acid analogue into the 50S ribosomal subunit indicates that steric clash may provide part of the chiral discrimination. The data presented provide one metric of the chiral and regiospecificity of mammalian ribosomes.     

In situ NMR spectroscopy of combustion

The first successful in situ studies of free combustion processes by one- and two-dimensional NMR spectroscopy are reported, and the feasibility of this concept is demonstrated. In this proof-of-principle work, methane combustion over a nanoporous material is investigated using hyperpolarized (hp)-xenon-129 NMR spectroscopy. Different inhomogeneous regions within the combustion cell are identified by the xenon chemical shift, and the gas exchange between these regions during combustion is revealed by two-dimensional exchange spectra (EXSY). The development of NMR spectroscopy as an analytical tool for combustion processes is of potential importance for catalyzed reactions within opaque media that are difficult to investigate by other techniques.     

Kinetics of Enzymatic High-Solid Hydrolysis of Lignocellulosic Biomass Studied by Calorimetry

Enzymatic hydrolysis of high-solid biomass (>10% w/w dry mass) has become increasingly important as a key step in the production of second-generation bioethanol. To this end, development of quantitative real-time assays is desirable both for empirical optimization and for detailed kinetic analysis. In the current work, we have investigated the application of isothermal calorimetry to study the kinetics of enzymatic hydrolysis of two substrates (pretreated corn stover and Avicel) at high-solid contents (up to 29% w/w). It was found that the calorimetric heat flow provided a true measure of the hydrolysis rate with a detection limit of about 500 pmol glucose s⁻¹. Hence, calorimetry is shown to be a highly sensitive real-time method, applicable for high solids, and independent on the complexity of the substrate. Dose–response experiments with a typical cellulase cocktail enabled a multidimensional analysis of the interrelationships of enzyme load and the rate, time, and extent of the reaction. The results suggest that the hydrolysis rate of pretreated corn stover is limited initially by available attack points on the substrate surface (<10% conversion) but becomes proportional to enzyme dosage (excess of attack points) at later stages (>10% conversion). This kinetic profile is interpreted as an increase in polymer end concentration (substrate for CBH) as the hydrolysis progresses, probably due to EG activity in the enzyme cocktail. Finally, irreversible enzyme inactivation did not appear to be the source of reduced hydrolysis rate over time.     

m-Phenylenedicarbene and m-Phenylenedinitrene Compounds Having Quintet Ground States

Higuchi¹, in his theoretical studies on hydrocarbons having high spin multiplicities, has suggested that compounds such as m-phenylenedlcarbene (I) and some of its substituted derivatives, e. g. m-phenylene-bis-phenylcarbene (II) should have ground states with four unpaired electrons, namely a quintet state. Recently, the zero-field (esr) splitting parameters (D and E) have been determined for I², II² and for m-phenylenedinitrene (III)². The results of the esr spectra are in agreement with quintet electronic ground states for these molecules^2,3.