Fluorogenic derivatization reagents suitable for isolation and identification of cysteine-containing proteins utilizing high-performance liquid chromatography-tandem mass spectrometry

The fluorogenic derivatization reagents with a positive charge, 4-(dimethylaminoethylaminosulfonyl)-7-chloro-2,1,3-benzoxadiazole (DAABD-Cl) and 7-chloro-2,1,3-benzoxadiazole-4-sulfonylaminoethyltrimethylammonium chloride (TAABD-Cl), are proposed for use in proteomics studies. Following derivatization of protein mixtures with these reagents, a series of standard processes of isolation, digestion, and identification of the proteins were performed utilizing high-performance liquid chromatography-fluorescence detection and tandem mass spectrometry with the probability-based protein identification algorithm. Both DAABD and TAABD derivatives were detected fluorometrically at the femtomole level and showed more than 100-fold improvement in sensitivity compared to the underivatized original compounds with an electrospray ionization ion trap mass spectrometer analysis. The modification of the MASCOT database search system memorized with the fragment information of a DAABD-attached Cys residue allowed the identification of the proteolytic peptide fragments of the derivatized bovine serum albumin (BSA) with an estimated 38% sequence coverage of BSA. Utilizing DAABD-Cl as a derivatization reagent, identification of several proteins was also possible in a soluble extract of Caenorhabditis elegans (10 microg of protein). Consequently, for identification of proteins in the complex matrixes of proteins, DAABD-Cl could be a more appropriate reagent than ammonium 7-fluoro-2,1,3-benzoxadiazole-4-sulfonate as reported previously.     

Quantitative investigation into the nucleation mechanism of lead and silver films vacuum deposited onto SiO,carbon and mica substrates

The origin of the occurrence of nuclei was quantitatively investigated using a microbalance and an electron microscope. The completion time of preferred nucleation and the time variation of the number density of stable nuclei were expressed in terms of measurable quantities such as the sticking coefficient of the incident vapour beam and the surface diffusion distance of adatoms. An analysis of the experimental results showed that the sum of the calculated values of the completion time of preferred nucleation and the appearance time of nuclei ranged from 2 to 60 s, whereas the time at which nucleus saturation occurred was found to range from 100 to 6000 s. It was concluded that the observed increase in the density of nuclei is caused by random nucleation. The occurrence of nucleus saturation can also be explained on the basis of random nucleation and growth coalescence.     

Defective bone remodelling in osteoprotegerin-deficient mice

Previous studies have reported enhanced osteoclastogenesis, increased bone resorption and osteoporosis in osteoprotegerin (OPG)-deficient mice. In the present study, we show that the tibial epiphyses contain abundant, thin trabeculae lined with numerous osteoclasts and cuboidal osteoblasts. The increase in osteoblasts and osteoclasts was associated with a dramatic increase in calcein labelling of the mineralization fronts and replacement of much of the intertrabecular marrow with numerous alkaline phosphatase-positive preosteoblasts. Furthermore, the discrete, linear cement lines seen in wild-type mice were replaced by a randomly oriented meshwork of cement lines that were stained intensely for tartrate-resistant acid phosphatase and osteopontin in the OPG-/- mice. These indices of accelerated bone remodelling in mutant bone were associated with irregular trabecular surfaces, a disorganized collagen matrix interspersed with amorphous ground substance and numerous fissures between old and new bone. In total, these observations indicate that enhanced osteoclastic activity in OPG-/- epiphyses led to a coupled increase in osteoblast differentiation and activity and an increase in bone remodelling. The high bone turnover, disorganized matrix and impaired attachment of new to old bone in the cement lines in OPG-/- mice appear to cause bone fragility.     

Optical design of a hollow cube-corner retroreflector for a geosynchronous satellite

The optical characteristics of a single-element hollow cube-corner retroreflector for a geosynchronous satellite were numerically evaluated for laser ranging and laser long-path absorption measurements of atmospheric species. An optical design with spherical surfaces and tuned dihedral angles was considered to compensate for velocity aberrations. The parameters for the retroreflector were optimized with genetic algorithms for different retroreflector sizes and wavelengths (500 nm and 1, 3, and 10 mum). We found that 20-cm retroreflectors are sufficient for realistic measurements when the laser wavelength is 500 nm or 1 mum. However, a larger retroreflector is necessary to overcome the detector noise level at 3 and 10 mum.     

Adaptive impedance control of a variable stiffness actuator

This paper proposes an optimal impedance control method for a variable stiffness actuator (VSA), in which a variable stiffness mechanism and an actuator are aligned in series. First, we introduce a circuit expression of the robotic system and provide a unified framework to determine an optimal index of robots driven by VSAs, irrespective of the presence or absence of the environment. Next, we design a torque controller for a one-degree-of-freedom (DOF) robot and find the optimal condition of the stiffness in the VSA for a given task. Then, we design a stiffness control law for the VSA exploiting the intrinsic indivisible property between motion and passive impedance. This stiffness control law adaptively tunes the passive stiffness to minimize the energy consumption without defining any explicit desired impedance, which is usually required in impedance controllers. The stability of the closed loop system is proved using Lyapunov’s analysis. Simulations and experimental results validate the effectiveness of the proposed method and the robustness in response to parameter changes.     

Out-of-Plane Negative Magnetoresistance of Bi2Sr1.6La0.4CuO6+δ Single Crystals in the Underdoped Region

In order to investigate a relationship between the carrier concentration and negative magnetoresistance in the underdoped region, we have performed out-of-plane electrical resistivity measurements for underdoped Bi ₂ Sr _1.6 La _0.4 CuO ₆₊ single crystals. Giant negative magnetoresistance has been observed in the most abundant carrier sample with = 0.12 in our experiment. The negative magnetoresistance reduces with decreasing carrier concentration.     

Intake of different eicosapentaenoic acid-containing lipids and fatty acid pattern of plasma lipids in the rats

The ethyl ester of eicosapentaenoic acid (EPA) is the only pure EPA-containing lipid available in bulk for oral administration. However, there is doubt as to whether EPA ethyl ester can efficiently increase the plasma levels of EPA in comparison with the ability of other kinds of EPA-containing lipids to do so. Therefore, two other kinds of EPA-containing lipids were prepared to study the efficiency of oral administration of those lipids for increasing the EPA content in plasma phospholipids and cholesteryl esters. EPA-containing lipids which were investigated were [A], 1,2,3-trieicosapentaenoyl-glycerol, [B] 2-eicosapentaenoyl-phosphatidylcholine and [C] ethyl ester of EPA. An adjusted amount of lipids [A], [B] and [C] was administered to rats through a gastric tube for 4 days (the first experiment) or for 10 days (the second experiment), and the fatty acid composition of plasma phospholipids and cholesteryl esters was determined. In the first experiment, there were no significant differences in the efficiency for increasing EPA levels in either phospholipids or cholesteryl esters among the lipids. In the second experiment, the EPA levels of both plasma phospholipids and cholesteryl esters of rats administered ethyl ester of EPA were significantly higher than those of rats administered 2-eicosapentaenoyl-phosphatidylcholine. The EPA levels of the rats administered 1,2,3-trieicosapentaenoylglycerol were between the levels of the two groups mentioned above, but the differences in the EPA levels were not significant. Although an ethyl ester-type molecule is not a naturally occurring lipid, ethyl ester of EPA is equal to 1,2,3-trieicosapentaenoyl-glycerol and appears to be superior to 2-eicosapentaenoyl-phosphatidylcholine as to the efficiency for increasing EPA levels in total plasma phospholipids and plasma cholesteryl esters.     

Constitutive Effects of Lead on Aryl Hydrocarbon Receptor Gene Battery and Protection by β‐carotene and Ascorbic Acid in Human HepG2 Cells

Lead (Pb) is an environmental pollutant that can get entry into human body through contaminated foods, drinks, and inhaled air leading to severe biological consequences, and has been responsible for many deaths worldwide. The objectives of this study were 1st to investigate the modulatory effects of environmentally relevant concentrations of Pb on AhR gene battery, which is controlling xenobiotics metabolism. 2nd, trials to reduce Pb‐induced adverse effects were done using some phytochemicals like β‐carotene or ascorbic acid. Human hepatoma (HepG2) cell lines were exposed to a wide range of Pb concentrations varying from physiological to toxic levels (0 to 10 mg/L) for 24 h. High Pb concentrations (1 to 10 mg/L) significantly reduced phase I (CYP1A1 and 1A2) and phase II (UGT1A6 and NQO1) xenobiotic metabolizing enzyme mRNA expression in a mechanistic manner through the AhR regulation pathway. Additionally, these Pb concentrations induced oxidative stress in HepG2 cells in terms of production of reactive oxygen species (ROS) and induced heme oxygenase‐1 mRNA expression in a concentration‐dependent phenomenon. Coexposure of HepG2 cells to physiological concentrations of some micronutrients, like β‐carotene (10 μM) or ascorbic acid (0.1 mM), along with Pb (1 mg/L) for 24 h significantly reduced the levels of ROS production and recovered AhR mRNA expression into the normal levels. Thus, consumption of foods rich in these micronutrients may help to reduce the adverse effects of lead in areas with high levels of pollution.