Short-capillary electrophoresis with electrochemiluminescence detection using porous etched joint for fast analysis of lidocaine and ofloxacin

Fast analysis of ofloxacin and lidocaine, as bactericide and analgesic or anesthetics, is of clinic importance for understanding the patient's medical process. This paper presented a high throughput, simple analysis method of lidocaine and ofloxacin by capillary electrophoresis coupled with electrochemiluminescence (ECL) using porous etched joint. To shorten the analysis time and to improve the analytical performance, a capillary with 10 cm in length was used as the separation channel. The cyclic voltammograms of Ru(bpy)(3)(2+) with different capillary length at same field strength showed that the porous etched joint eliminated the effect of electrophoretic current on the ECL detection. Following micro total analysis systems (muTAS), some advantages of which this approach has, the fabrication of channel in chip was not needed. Compared with capillary electrophoresis with 40-cm-long capillary, the high sample throughput and low zone broadening may be the main advantage of the present system. Under optimal condition, the detection limits of lidocaine and ofloxacin based on peak height were 3.0 x 10(-8) and 5.0 x 10(-7) mot L-1 and a 60 h(-1) of sampling frequency was obtained.

High resolution mass spectrometric alveolar proteomics: Identification of surfactant protein SP-A and SP-D modifications in proteinosis and cystic fibrosis patients

In the present study, one- and two-dimensional gel electrophoresis combined with high resolution Fourier transform-ion cyclotron resonance mass spectrometry (FT-ICR MS) have been applied as powerful approaches for the proteome analysis of surfactant proteins SP-A and SP-D, including identification of structurally modified and truncation forms, in bronchoalveolar lavage fluid from patients with cystic fibrosis, chronic bronchitis and pulmonary alveolar proteinosis. Highly sensitive micro preparation techniques were developed for matrix-assisted laser desorption/ionization (MALDI) FT-ICR MS analysis which provided the identification of surfactant proteins at very low levels. Owing to the high resolution, FT-ICR MS was found to provide substantial advantages for the structural identification of surfactant proteins from complex biological matrices with high mass determination accuracy. Several protein bands corresponding to SP-A and SP-D were identified by MALDI-FT-ICR MS after electrophoretic separation by one- and two-dimensional gel electrophoresis, and provided the identification of structural modifications (hydroxy-proline) and degradation products.

Oriented nano-structured hydroxyapatite from the template

Natural bone is one kind of compounds consisting of hydroxyapatite (HAp) nano-rods, which are embedded in the template of collagen matrix in vivo with the same crystallographic organization. Herein HAp nano-rods precursors were synthesized via wet chemical method. Large-scale HAp nano-wires with the same crystallographic organization as the template of anodic aluminum oxide (AAO) were obtained by the electrophoretic deposition and the technology of the template. It provides a meaningful method to study and understand the information of biological molecules' mineralization process.

New technique for capillary electrophoresis directly coupled with end-column electrochemiluminescence detection

capillary electrophoresis (CE) is characterized. A 300 mum diameter Pt working electrode was used to directly couple with a 75 mum inner diameter separation capillary without an electric field decoupler. The hydrodynamic cyclic voltammogram (CV) of Ru(bpy)(3)(2+) showed that electrophoretic current did not affect the ECL reaction. The presence of high-voltage (HV) field only resulted in the shift of the ECL detection potential. The distance of capillary to electrode was an important parameter for optimizing detection performance as it determined the characteristics of mass transport toward the electrode and the actual concentration of Ru(bpy)(3)(2+) in the detection region. The optimum distance of capillary to electrode was decided by the inner diameter of the capillary, too. For a 75 mum capillary, the working electrode should be placed away from the capillary outlet at a distance within the range of 20-260 mum. The effects of pH value of ECL solution and molecular structure of analytes on peak height and theoretical plate numbers were discussed. Using the 75 mum capillary, under the optimum conditions, the method provided a linear range for tripropylamine (TPA) between 1 x 10(-10) and 1 X 10(-5) mol/L with correlation coefficient of 0.998. The detection limit (signal-to-noise ratio S/N = 3) was 5.0 x 10(-11) mol/L. The relative standard deviation in peak height for eight consecutive injections was 5.6%. By this new technique lidocaine spiked in a urine sample was determined. The method exhibited the linear range for lidocaine from 5.0 x 10(-8) to 1.0 X 10(-5) mol/L with correlation efficient of 0.998. The limit of detection (S/N = 3) was 2.0 x 10(-1) mol/L.

Characterization of proteolytic bacteria from the Aleutian deep-sea and their proteases

Six deep-sea proteolytic bacteria taken from Aleutian margin sediments were screened; one of them produced a cold-adapted neutral halophilic protease. These bacteria belong to Pseudoalteromonas spp., which were identified by the 16S rDNA sequence. Of the six proteases produced, two were neutral cold-adapted proteases that showed their optimal activity at pH 7-8 and at temperature close to 35 degrees C, and the other four were alkaline proteases that showed their optimal activity at pH 9 and at temperature of 40-45 degrees C. The neutral cold-adapted protease E1 showed its optimal activity at a sodium chloride concentration of 2 M, whereas the activity of the other five proteases decreased at elevated sodium chloride concentrations. Protease E1 was purified to electrophoretic homogeneity and its molecular mass was 34 kDa, as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The molecular weight of protease E1 was determined to be 32,411 Da by mass spectrometric analysis. Phenylmethyl sulfonylfluoride (PMSF) did not inhibit the activity of this protease, whereas it was partially inhibited by ethylenediaminetetra-acetic acid sodium salt (EDTA-Na). De novo amino acid sequencing proved protease E1 to be a novel protein.

In vitro selected peptides bind with thymidylate synthase mRNA and inhibit its translation

Thymidylate synthase (TS), an essential enzyme for catalyzing the biosynthesis of thymidylate, is a critical therapeutic target in cancer therapy. Recent studies have shown that TS functions as an RNA-binding protein by interacting with two different sequences on its own mRNA, thus, repressing translational efficiency. In this study, peptides binding TS RNA with high affinity were isolated using mRNA display from a large peptide library (>10(13) different sequences). The randomized library was subjected up to twelve rounds of in vitro selection and amplification. Comparing the amino acid composition of the selected peptides (12th round, R12) with those from the initial random library (round zero, R0), the basic and aromatic residues in the selected peptides were enriched significantly, suggesting that these peptide regions might be important in the peptide-TS mRNA interaction. Categorizing the amino acids at each random position based on their physicochemical properties and comparing the distributions with those of the initial random pool, an obvious basic charge characteristic was found at positions 1, 12, 17 and 18, suggesting that basic side chains participate in RNA binding. Secondary structure prediction showed that the selected peptides of R12 pool represented a helical propensity compared with R0 pool, and the regions were rich in basic residues. The electrophoretic gel mobility shift and in vitro translation assays showed that the peptides selected using mRNA display could bind TS RNA specifically and inhibit the translation of TS mRNA. Our results suggested that the identified peptides could be used as new TS inhibitors and developed to a novel class of anticancer agents.

Expression, purification, and characterization of recombinant Chinese shrimp crustin-like protein (CruFc) in Pichia pastoris

A crustin-like protein (CruFc) from Fenneropenaeus chinensis was expressed in Pichia pastoris and then purified to electrophoretic homogeneity on a Sephacryl S-100 column with a band corresponding to the expected one (13 kDa) shown by 15% SDS-PAGE. Western blot indicated that the rCruFc specifically reacted with polyclonal rabbit anti-Fenneropenaeus chinensis CruFc. Production in a 5 l bioreactor gave 237 mg rCruFc/l. Antimicrobial assay revealed that 4 mu M rCruFc inhibited growth of Staphylococcus aureus.

Zebrafish thymidylate synthase binds to its own mRNA in vitro and in vivo

Thymidylate synthase (TS), an essential enzyme for DNA de novo synthesis, is a critical therapeutic target in cancer therapy. Previous study has shown that TS was able to bind to its own mRNA in human and E.coli, resulting in translational repression. Zebrafish is the best animal model for vertebrate study. In order to study the regulatory mechanism of zebrafish TS, the enzyme were expressed in E. coli BL21 (DE3) and it was purified to homogeneity. Electrophoretic mobility shift assay (EMSA) was used to detect the interaction of zebrafish TS protein and its own TS transcript in vitro and the results showed that zebrafish TS could bound with its own mRNA specifically. Further study revealed that zebrafish TS was able to interact with its own mRNA in vivo using immunoprecipitation : RT-PCR technique. The results provide evidence that zebrafish may be developed as an useful model for studying the anti-metabolism agents.

Grain size distribution of quartz isolated from Chinese loess/paleosol

Grain size distribution of bulk loess-paleosol and quartz chemically extracted from the loess/paleosol shows that mean size of the bulk samples is always finer than that of the quartz, The original aeolian depositions have been modified to various degrees by post-depositional weathering and pedogenic processes. The grain size distribution of the isolated quartz should be close to that of the primary aeolian sediment because the chemical pretreatment excludes secondary produced minerals. Therefore, the grain size of the quartz may be considered to more clearly reflect the variations of winter monsoon intensity.

Effects of spark discharge on the anodic coatings on magnesium alloy

Spark discharge was the representative phenomenon of Micro-arc oxidation (MAO) method distinguished from other electrochemical oxidation methods. Under the spark discharge treatment, characteristics of the anodic layer were significantly changed. To investigate the influences of the spark discharge, a piece of magnesium alloy AZ91D specimen was partly treated by MAO method in alkaline silicate solution. And the microstructure, element distributions as well as the surface potential distributions of the specimen were studied by scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS) and scanning Kelvin probe (SKP) technique. As a result of intensive spark discharge treatment, porous external layer with dense internal layer were formed on Mg alloy surface. At the same time, the depositions of OH- and SiO32- ions were accelerated, which resulted in the enrichment of element oxygen and silicon at the spark discharge region. Moreover, due to the compact internal layer, the intensive spark discharge region exhibited more positive potentials with respect to other regions, which meant this region could restrain the ejection of electron and provide effective protection to the substrate. In addition, it was found that oxygen played a vital role in determining the intensity and size of sparks, and abundant oxygen resulted in intensive and larger sparks. (c) 2005 Elsevier B.V. All rights reserved.

Isolation and characterization of a novel variant of HMW glutenin subunit gene from the St genome of Pseudoroegneria stipifolia

The x- and y-type high molecular weight (HMW) glutenin subunits are conserved seed storage proteins in wheat and related species. Here we describe investigations on the HMW glutenin subunits from several Pseudoroegneria accessions. The electrophoretic mobilities of the HMW glutenin subunits from Pd. stipifolia, Pd tauri and Pd strigosa were much faster than those of orthologous wheat subunits, indicating that their protein size may be smaller than that of wheat subunits. The coding sequence of the Glu-1St1 subunit (encoded by the Pseudoroegneria stipifolia accession PI325181) was isolated, and found to represent the native open reading frame (ORF) by in vitro expression. The deduced amino acid sequence of Glu-1St1 matched with that determined from the native subunit by mass spectrometric analysis. The domain organization in Glu-1St1 showed high similarity with that of typical HMW glutenin subunits. However, Glu-1St1 exhibited several distinct characteristics. First, the length of its repetitive domain was substantially smaller than that of conventional subunits, which explains its much faster electrophoretic mobility in SDS-PAGE. Second, although the N-terminal domain of Glu-1St1 resembled that of y-type subunit, its C-terminal domain was more similar to that of x-type subunit. Third, the N- and C-terminat domains of Glu-1St1 shared conserved features with those of barley D-hordein, but the repeat motifs and the organization of its repetitive domain were more similar to those of HMW glutenin subunits than to D-hordein. We conclude that Glu-1St1 is a novel variant of HMW glutenin subunits. The analysis of Glu-1St1 may provide new insight into the evolution of HMW glutenin subunits in Triticeae species. (C) 2007 Elsevier Ltd. All rights reserved.

细粒沉积物多组分粒度特征及其成因机制研究

By using high-resolution laser grain size instrument Mastersizer 2000, the grain size distribution of windblown depositions (loess and sandy dunes), aqueous sediments (lake, river, riverside and foreshore sand), weathering crust, sloping materials and other fine-grain sediments are systemically measured. The multimodal characteristics of grain size distribution of these sediments are carefully studied. The standard patterns and their grain size characteristics of various sediments are systemically summarized. The discrepancies of multimodal distribution among windblown depositions, aqueous sediments and other sediments are concluded and the physical mechanisms of grain size multimodal distribution of various sediments are also discussed in this paper. The major conclusions are followed: 1. The multimodal characteristic of grain size distribution is a common feature in all sediments and results from properties of transportation medium, dynamic intensity, transportation manner and other factors. 2. The windblown depositions are controlled by aerodynamic forcing, resulting in that the median size of the predominant mode gradually decreases form sandy dunes to loess. Similarly, the aqueous sediments are impacted by dynamic forces of water currents and the median grain size of the predominant mode decreases gradually from river to lake sediments. Because the kinetic viscidity of air is lower than of water, the grain size of modes of windblown depositions is usually finer than that of corresponding modes of aqueous sediments. Typical characteristics of sediments grain size distribution of various sediments have been summarized in the paper: (1) Suspended particles which diameters are less than 75μm are dominant in loess and dust. There are three modes in loess’ grain size distribution: fine, median and coarse (the median size is <1μm、1-10μm、10-75μm, respectively). The coarse mode which percentage is larger than that of others is controlled by source distance and aerodynamic intensity of dust source areas. Some samples also have a saltation mode which median size is about 300-500μm. Our analysis demonstrates that the interaction of wind, atmospheric turbulence, and dust grain gravity along the dust transportation path results in a multimodal grain size distribution for suspended dust. Changes in the median sizes of the coarse and medium modes are related to variation in aerodynamic forcing (lift force related to vertical wind and turbulence) during dust entrainment in the source area and turbulence intensity in the depositional area. (2) There is a predominant coarse saltation mode in grain size distribution of sandy dunes, which median size is about 100-300μm and the content is larger than that of other modes. The grain size distribution curve is near axis symmetric as a standard logarithm normal function. There are some suspended particles in some samples of sandy dunes, which distribution of the fine part is similar to that of loess. Comparing with sandy samples of river sediments, the sorting property of sandy dunes is better than of river samples although both they are the saltation mode. Thus, the sorting property is a criterion to distinguish dune sands and river sands. (3) There are 5~6 modes (median size are <1μm, 1-10μm, 10-70μm, 70-150μm, 150-400μm, >400μm respectively) in grain size distribution of lacustrine sediments. The former 4 modes are suspensive and others are saltated. Lacustrine sediments can be divided into three types: lake shore facies, transitional facies and central lake facies. The grain size distributions of the three facies are distinctly different and, at the same time, the transition among three modes is also clear. In all these modes, the third mode is a criteria to identify the windblown deposition in the watershed. In lake shore sediments, suspended particles are dominant, a saltation mode sometimes occurs and the fourth mode is the most important mode. In the transitional facies, the percentage of the fourth mode decreases and that of the second mode increases from lake shore to central lake. In the central lake facies, the second mode is dominant. A higher content of the second mode indicates its position more close to the central lake. (4) The grain size distribution of river sediments is the most complex. It consist of suspension, saltation and rolling modes. In most situations, the percentage of the saltation mode is larger than that of other modes. The percentage of suspension modes of river sediments is more than of sandy dunes. The grain size distribution of river sediments indicates dynamic strength of river currents. If the fourth mode is dominant, the dynamic forcing of river is weaker, such as in river floodplain. If the five or sixth mode is dominant, the water dynamic forcing of rivers is strong. (5) Sediments can be changed by later forcing in different degree to form some complicated deposition types. In the paper, the grain size distribution of aqueous sediments of windblown deposition, windblown sediments of aqueous deposition, weathering crust and slope materials are discussed and analyzed. 3. The grain size distribution characteristics of different sediments are concluded: (1) Modal difference: Usually there are suspended and saltation modes in the windblown deposition. The third mode is dominant in loess dust and the fifth mode is predominant in sandy dunes. There are suspended, saltation and rolling particles in aqueous sediments. In lacustrine sediments, the second and fourth mode are predominant for central lake facies or lake shore facies, respectively. In river sediments, the fourth, or fifth, or sixth mode is predominant. Suspended modes: the grain size of suspended particles of windblown depositions usually is less than 75μm. The content of suspended particles is lower or none in sandy dunes. However, suspended particles of aqueous sediments may reach 150μm. Difference in grain size of suspended modes represents difference between transitional mediums and the strength of dynamic forcing. Saltation modes: the median size of saltation mode of sandy dunes fluctuates less than that of river sediments. (2) Loess dust and lacustrine sediment: Their suspended particles are clearly different. There is an obvious pit between the second and the third modes in grain size distribution of lacustrine sediments. The phenomenon doesn’t occur in loess dust. In lacustrine sediments, the second mode can be a dominant mode, such as central lake facies, and contents of the second and the third modes change reversely. However, the percentage of the third mode is always the highest in loess dust. (3) Dune Sand and fluvial sand: In these two depositions, the saltation particles are dominant and the median sizes of their saltation modes overlay in distribution range. The fifth mode of dune sand fluctuates is sorted better than that of fluvial sand. (4) Lacustrine and fluvial sediments: In lacustrine sediments, there are 5-6 modes and suspended particles can be predominant. The second mode is dominant in central lake facies and the third mode is dominant in lake shore facies. Saltation or roll modes occurred in central lake facies may indicate strong precipitation events. In fluvial sediments, saltation particles (or rolling particles) usually dominant. 4. A estimation model of lake depth is firstly established by using contents of the second, the third and the fourth modes. 5. The paleoenvironmental history of the eastern part of SongLiao basin is also discussed by analyzing the grain size distribution of Yushu loess-like sediments in Jilin. It was found that there is a tectonic movement before 40ka B.P. in SongLiao basin. After the movement, loess dust deposited in Yushu area as keerqin desert developed. In recent 2000 years, the climate became drier and more deserts activated in the eastern part of Song-Liao basin.

西部黄土高原中新世风尘沉积剖面的磁性地层学和沉积学研究

Observation of the Miocene aeolian dust deposits in Qin'an area provide a long-timescale, almost continuous geological record for understanding significant environmental changes, such as the environmental evolution of Asian monsoons and the inland Asian aridification, and make possible new explorations of global and regional environmental evolutions and other relevant studies. However, the QA-I and QA-II sections that are previously investigated are near to each other, and there is a lack in understanding the spatial characteristics of changes in paleoenvironment. Therefore, new sections of Miocene aeolian dust deposits are needed to complete the present spatial framework. Meanwhile, it is necessary to carry out in-depth studies, which are already belated, on the depositional characteristics of these Miocene aeolian dust deposits.This thesis choose the Miziwan section and Shaogou section that are west to Liupan Mountain for magnetostratigraphic studies, results of which are compared with the QA-I section with respect to the lithology, magnetic susceptibility and paleomagnetic polarity stratigraphy. Quartz-fraction grain size of 300 samples from the QA-I section was analyzed. In addition, 30 out of these samples were determined in terms of the quartz morphological characteristics by using scanning electronic microscopy. On the basis of these data and the correlation with the Xifeng sequence of aeolian dust deposits, the following 3 conclusions are drawn as below:The upper and the lower boundary age of the Miziwan section are 11.6 Ma B.Rand 18.5 Ma B.R, respectively. The Shaogou section spans from 15.1 Ma B.P, to20.8 Ma B.R The lower boundary age of both sections is no older than that of theQA-I section. The Miziwan and the Shaogou sections can be well correlated withthe QA-I section in terms of the field lithological features, magnetic susceptibilityand magnetic polarity stratigraphy.Sedimentological evidence of the QA-I quartz grain size and quartz morphologicalcharacteristics further supports the previous conclusion that the QA-Iloess-paleosol sequence is of aeolian origin, and also verifies the accretionalfeatures in paleosols of this sequence in terms of the grain-size composition.Modifications of the grain-size composition of the original dusts due to later-stageweathering and pedogenesis are common characteristic of Miocene and Quaternary aeolian dust depositions. The above-mentioned processes further rework and refine the bulk grain size, which is more evident in paleosol layers. 3, Like the hipparion Red-Earth and the Quaternary aeolian deposits, the Miocene loess deposits are transported by near-ground winter monsoons. However, the average wind intensity evidenced by the quartz median grain size of Miocene loess, together with the maximum wind intensity by the quartz maximum grain size, is weaker than those of the Quaternary.