Field evaluations of the VDmax approach for substantiation of a 25 kGy sterilization dose and its application to other preselected doses

The International and European standards for radiation sterilization require evidence of the effectiveness of a minimum sterilization dose of 25 kGy but do not provide detailed guidance on how this evidence can be generated. An approach, designated VD_max, has recently been described and computer evaluated to provide safe and unambiguous substantiation of a 25 kGy sterilization dose. The approach has been further developed into a practical method, which has been subjected to field evaluations at three manufacturing facilities which produce different types of medical devices. The three facilities each used a different overall evaluation strategy: Facility A used VD_max for quarterly dose audits; Facility B compared VD_max and Method 1 in side-by-side parallel experiments; and Facility C, a new facility at start-up, used VD_max for initial substantiation of 25 kGy and subsequent quarterly dose audits. A common element at all three facilities was the use of 10 product units for irradiation in the verification dose experiment.

The field evaluations of the VD_max method were successful at all three facilities; they included many different types of medical devices/product families with a wide range of average bioburden and sample item portion values used in the verification dose experiments. Overall, around 500 verification dose experiments were performed and no failures were observed. In the side-by-side parallel experiments, the outcomes of the VD_max experiments were consistent with the outcomes observed with Method 1.

The VD_max approach has been extended to sterilization doses >25 and <25 kGy; verification doses have been derived for sterilization doses of 15, 20, 30, and 35 kGy. Widespread application of the VD_max method for doses other than 25 kGy must await controlled field evaluations and the development of appropriate specifications/standards.     

Retention of alkali,alkaline earth and transition metals on an itaconic acid cation-exchange column. Eluent pH,ionic strength and temperature effects upon selectivity

The unusual selectivity of a methylene succinic (itaconic) acid modified polymeric column was investigated for the separation of alkali, alkaline earth, transition and heavy metals employing non-chelating inorganic eluents. The retention of selected metal ions on the column was investigated with simple HNO3 eluents and eluents prepared from KNO3 and KCl salts of varying pH (adjusted using HNO3). From these studies both the effect of eluent ionic strength and pH upon retention was evaluated for the itaconic acid stationary phase. The results obtained showed that despite slow exchange kinetics causing poor efficiencies, acceptable baseline separations of selected alkaline earth and transitions could be obtained under optimum conditions (the baseline separation of Mg(II), Ca(II), Mn(II), Cd(II), Zn(II) and Co(II) was possible using a 15 mM KNO3-5 mM KCl eluent at pH 3.50 in under 25 min). The use of an simple ionic strength step gradient was shown that facilitated the addition of Pb(II) to the above group of metal ions. An investigation into the effect of temperature upon peak efficiency and retention showed increased column temperature could be used to improve the resolution of closely eluting metal ions such as Ca(II) and Sr(II) and Ca(II) and Mn(II).     

Sensitive measurement of radiation trapping in cold-atom clouds by intensity correlation detection

We present experimental evidence that the intensity correlations of light scattered from a cold-atom cloud are sensitive to the presence of small amounts of radiation trapping in an atomic sample of density 6 x 10(8)/cm3, with an optical depth (for a resonant light beam) of 0.4. This density and optical depth are approximately an order of magnitude less than the density and on-resonance optical depth at which effects of multiple scattering in cold-atom clouds have been previously observed [Phys. Rev. Lett. 64, 408 (1990)].     

Characterization of the restricted rotation of the dimethyl groups in chemically N-terminal 13C-labeled antifreeze glycoproteins: a temperature-dependent study in water to ice through the supercooled state

Site-specific chemical modification, especially with isotopically enriched groups, allows one to study the structure and dynamics of proteins for which uniform enrichment is difficult. When the N-terminal alanine in antifreeze glycoprotein (AFGP) is replaced with an N,N-dimethyl alanine the methyl groups show signatures of slow rotation about the C-N bond. In order to separate the local dynamics of the N-terminus from the overall protein dynamics, we present a complete characterization of this dynamics. Temperature-dependent nuclear magnetic-resonance experiments from room temperature to subzero temperatures, including the supercooled state and in the presence of ice, are presented. Quantum chemical calculations are also performed on a localized N-terminus of the AFGP. Our results show that in the solution state at room temperature and in the super cooled regime, the dimethyl groups undergo a slow, restricted rotation with an unequal distribution of population between two major conformations. At lower temperatures in the presence of ice, the dynamics become much more complex due to freezing out of several conformational states. Based on these results, we conclude that the segmental dynamics of the N-terminus are local to the first residue and do not affect the overall dynamics of the protein.     

A two-dimensional adsorption kinetic model for thermal hysteresis activity in antifreeze proteins

Antifreeze proteins (AFPs) and antifreeze glycoproteins (AFGPs), collectively abbreviated as AF(G)Ps, are synthesized by various organisms to enable their cells to survive in subzero environments. Although the AF(G)Ps are markedly diverse in structure, they all function by adsorbing to the surface of embryonic ice crystals to inhibit their growth. This adsorption results in a freezing temperature depression without an appreciable change in the melting temperature. The difference between the melting and freezing temperatures, termed thermal hysteresis (TH), is used to detect and quantify the antifreeze activity. Insights from crystallographic structures of a number of AFPs have led to a good understanding of the ice-protein interaction features. Computational studies have focused either on verifying a specific model of AFP-ice interaction or on understanding the protein-induced changes in the ice crystal morphology. In order to explain the origin of TH, we propose a novel two-dimensional adsorption kinetic model between AFPs and ice crystal surfaces. The validity of the model has been demonstrated by reproducing the TH curve on two different beta-helical AFPs upon increasing the protein concentration. In particular, this model is able to accommodate the change in the TH behavior observed experimentally when the size of the AFPs is increased systematically. Our results suggest that in addition to the specificity of the AFPs for the ice, the coverage of the AFPs on the ice surface is an equally necessary condition for their TH activity.     

First observational tests of eternal inflation

The eternal inflation scenario predicts that our observable Universe resides inside a single bubble embedded in a vast inflating multiverse. We present the first observational tests of eternal inflation, performing a search for cosmological signatures of collisions with other bubble universes in cosmic microwave background data from the WMAP satellite. We conclude that the WMAP 7-year data do not warrant augmenting the cold dark matter model with a cosmological constant with bubble collisions, constraining the average number of detectable bubble collisions on the full sky N(s) < 1.6 at 68% C.L. Data from the Planck satellite can be used to more definitively test the bubble-collision hypothesis.     

Unexpected Self‐Sorting Self‐Assembly Formation of a [4:4] Sulfate:Ligand Cage from a Preorganized Tripodal Urea Ligand

The design and synthesis of tripodal ligands 1 – 3 based upon the N‐methyl‐1,3,5‐benzenetricarboxamide platform appended with three aryl urea arms is reported. This ligand platform gives rise to highly preorganized structures and is ideally suited for binding SO₄^2? and H₂PO₄^? ions through multiple hydrogen‐bonding interactions. The solid‐state crystal structures of 1 – 3 with SO₄^2? show the encapsulation of a single anion within a cage structure, whereas the crystal structure of 1 with H₂PO₄^? showed that two anions are encapsulated. We further demonstrate that ligand 4 , based on the same platform but consisting of two bis‐urea moieties and a single ammonium moiety, also recognizes SO₄^2? to form a self‐assembled capsule with [4:4] SO₄^2?: 4 stoichiometry in which the anions are clustered within a cavity formed by the four ligands. This is the first example of a self‐sorting self‐assembled capsule where four tetrahedrally arranged SO₄^2? ions are embedded within a hydrophobic cavity.     

Enantiomer ratio of MK-0767 in humans and nonclinical species

MK-0767, (+/-)-5-[(2,4-dioxothiazolidin-5-yl)methyl]-2-methoxy-N-[[(4-trifluoromethyl)phenyl]methyl]benzamide, is a thiazolidinedione-containing dual peroxisome proliferator-activated receptor (PPAR) alpha/gamma agonist that has been studied as a potential treatment for patients with type 2 diabetes. MK-0767 contains a chiral center at the C-5 position of the thiazolidinedione ring and was being developed as the racemate, due to the rapid interconversion of its enantiomers in biological samples. In the present work the in vitro and in vivo concentration ratios of the (+)-(R) to (-)-(S) enantiomers of MK-0767 were determined in plasma from humans (in vitro only) and nonclinical species used in the toxicological evaluation of rac-MK-0767, namely CD-1 mice, Sprague-Dawley rats, beagle dogs, New Zealand white rabbits, and rhesus monkeys. The R/S ratio was determined by chiral liquid chromatography/tandem mass spectrometry. Species differences were observed in the in vitro and in vivo enantiomeric ratios, as well as differences between in vitro and in vivo in some species. The in vitro R/S ratio was similar in dogs and humans (approximately 1.5-1.7). In rats and monkeys, the ratio was approximately unity, both in vitro and in vivo. In mice, the ratio was higher in vitro (approximately 1) than in vivo (approximately 0.6), while in rabbits it was higher in vivo (approximately 1) than in vitro (approximately 0.5). These results suggested that differential binding of the MK-0767 enantiomers to plasma and tissue proteins and other macromolecules may be affecting the R/S ratio both in vitro and in vivo, since in protein-free systems MK-0767 exists as the racemate.