Acid effects on protein molecular weight determination

The acid effects of some proteins on measuring their molecular weights were studied using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry ( MALDI-TOF-MS) and electrospray ionization mass spectrometry (ESI-MS). It was found that the signal intensity was enhanced through adjusting the acid concentration in some protein samples. In this paper, both MALDI-MS and ESI-MS was applied to examine the molecular weights of several standard proteins. And the proper acid concentration was detected in these spectra. In the meanwhile, it demonstrates that some associations of proteins in solution can be preserved into the gas phase and observed by the "soft ionization" mass spectrometry.

Preparation and mass spectrometric study of egg yolk antibody (IgY) against rabies virus

Rabies virus was used as the antigen to immunize laying chickens. Anti-rabies virus immunoglobulin Y(IgY) was isolated from yolks of the eggs laid by these chickens using a two-step salt precipitation and one-step gel filtration protocol. The purified IgY was reduced with dithiothreitol, and heavy chains (HC) and light chains (LC) were obtained. In addition, the purified IgY was digested with pepsin and the fragment with specific antigen binding properties (Fab) was produced. Using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOFMS), the average molecular weights of IgY, HC, LC, and Fab were determined as 167 250, 65 105, 18 660, and 45,359 Da, respectively. IgY has two structural differences compared with mammalian IgGs. First, the molecular weight of the heavy chain of IgY is larger than that of its mammalian counterpart, while the molecular weight of the light chain of IgY is smaller. Second, upon pepsin digestion, anti-rabies virus IgY is degraded into Feb, in contrast to mammalian IgG, which has been reported to be degraded into F(ab')(2) under the same conditions. Copyright (C) 2001 John Wiley & Sons, Ltd.

Synthesis of cyclic precursors of poly(ether ether ketone)

A novel synthetic route to cyclic PEEK precursors is described. These new cyclic oligomers have been prepared from hydroquinone and N-phenyl(4,4'-difluorodiphenyl) ketimine. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis unambiguously confirmed the cyclic nature.

Synthesis and polymerization of some macrocyclic (arylene ether sulfone) containing cardo groups and macrocyclic (arylene ether ketone sulfone) oligomers

Some novel macrocyclic (arylene ether sulfone) containing cardo groups and (arylene ether ketone sulfone) oligomers have been synthesized in high yields by a nucleophilic aromatic substitution reaction of 4,4'-difluorophenylsulfone with bisphenols in the presence of anhydrous potassium carbonate under a pseudo-high-dilution condition. Detailed structural characterization of these oligomers by matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS), fast atom bombardment mass spectrometry (f.a.b.-m.s.), nuclear magnetic resonance spectrometry (n.m.r.) and single-crystal X-ray structure analysis confirms their cyclic nature, and the composition of the oligomeric mixtures is provided by g.p.c. analysis. Ring polymerization of cyclic oligomers 3a to a high molecular weight polymer with M-w of 59.1 k was achieved by heating at 290 degrees C for 40 min in the presence of a nucleophilic initiator. (C) 1998 Elsevier Science Ltd. All rights reserved.

Macrocyclic oligomeric arylene ether ketones: Synthesis and polymerization

Some novel macrocylic(arylene ether ketone)oligomers were synthesized in high yields by a nucleophilic aromatic substitution reaction of 4,4'-dinitrobenzophenone with bisphenols in the presence of anhydrous potassium carbonate under pseudo-high-dilution conditions. Detailed structural characterization of these oligomers by matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS), H-1 NMR and FT-IR confirmed their cyclic nature and the compositions of the oligomeric mixtures was indicated by GPC analysis. Ring-opening polymerization of cyclic oligomers 3a to a high molecular weight polymer with M-w of 52.3 and M-n of 17.2 k was achieved by heating at 280 degrees C for 40 min in the presence of a nucleophilic initiator.

Proteomic detection of changes in protein expression induced by cordycepin in human hepatocellular carcinoma BEL-7402 cells

The nucleoside analogue cordycepin (3'-deoxyodenosine, 3'-dA), one of the components of cordyceps militaris, has been shown to inhibit the growth of various tumor cells. However, the probable mechanism is still obscure. In this study, the inhibition of cell growth and changes in protein expression induced by cordycepin were investigated in BEL-7402 cells. Using the MTT assay and flow cytometry, we found that cordycepin inhibits cell viability and induces apoptosis in BEL 7402 cells. Additionally. the proteins were separated using two-dimensional polyacrylamide gel electrophoresis, and eight proteins were found to be significantly, affected by cordycepin compared to untreated control; among them, two were downregulated and six were upregulated. Of the eight proteins, six were identified with peptide mass fingerprinting using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) after in-gel trypsin digestion. These proteins are involved in various aspects of cellular metabolism. It is suggested that the effect of cordycepin on the growth of tumor cells is significantly related to the metabolism-associated protein expression induced by cordycepin. Copyright 2008 Prous Science, S.A.U. or its licensors. All rights reserved.

三聚氰胺的基质辅助激光解吸/电离飞行时间质谱研究

Oxidized carbon nanotubes are tested as the matrix for analysis of the melamine by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). Traditional MALDI matrix are not suit for analysis of the low molecular compounds due to the interference associated to the matrix clusters. Oxidized carbon nanotubes can transfer energy to the analyte under the laser irradiation, which makes analyte well ionized or desorbed. Moreover, the interference of the intrinsic matrix ions can be eliminated. Melamine as the a toxic additive which had been added in the milk powder, then it is necessary to establish a new method for detection of the melamine rapid and sensitive.

马鹿茸多肽的提取分离及基质辅助激光解吸飞行时间质谱分析

In this paper,active components of velvet antler polypeptide were extracted and separated.The molecular weight and purity of velvet antler polypeptide were determined by MALDI-TOFMS.The different influence factors such as matrix,sample concentration and laser energy were studied.This method is convenient and suitable for the identification of congener biochemical samples.

Comparative study of three short-chain neurotoxins from the venom of Naja kaouthia (Yunnan, China)

Three short-chain neurotoxins named NT-I, NT-II, and NT-III were purified from the venom of Naja kaouthia, a snake distributed throughout the south of Yunnan province, China, by a series of chromatographic steps, including an FPLC Resource S column. Their molecular weights, determined by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) MS, were 6952.19 Da, 6854.92 Da, and 6828.80 Da, respectively. NT-I consisted of 62 amino acid residues, and the other two consisted of 61 amino acid residues, including 8 cysteines. After hydrolysis by endoproteinase Glu-C, their primary sequences were determined. A test of their activities demonstrated that they effectively inhibited muscle contractions induced by electric stimulation. Furthermore, the extent of inhibition caused by NT-II and NT-III was less than that of NT-I. The IC(50)s were 0.04 mug/ml, 0.20 mug/ml, and 0.23 mug/ml for NT-I, NT-II, and NT-III, respectively. Compared with NT-II and NT-III, the higher activity of NT-I may be a result of the amino acid residue substitution Ile36 to Arg36.

Purification, characterization and primary structure of a chymotrypsin inhibitor from Naja atra venom

A chymotrypsin inhibitor, designated NA-CI, was isolated from the venom of the Chinese cobra Naja atra by three-step chromatography. It inhibited bovine (x-chymotrypsin with a K-i of 25 nM. The molecular mass of NA-CI was determined to be 6403.8 Da by matrix-assisted laser-desorption ionization time-of-flight (MALDI-TOF) analysis. The complete amino acid sequence was determined after digestion of S-carboxymethylated inhibitor with Staphylococcus aureus V8 protease and porcine trypsin. NA-CI was a single polypeptide chain composed of 57 amino acid residues. The main contact site with the protease (PI) has a Phe, showing the specificity of the inhibitor. NA-CI shared great similarity with the chymotrypsin inhibitor from Naja naja venom (identities = 89.5%) and other snake venom protease inhibitors. (C) 2003 Elsevier Inc. All rights reserved.

Protein Profiles in Zebrafish (Danio rerio) Embryos Exposed to Perfluorooctane Sulfonate

Perfluorooctane sulfonate (PFOS) is widely distributed and persistent in the environment and in wildlife, and it has the potential for developmental toxicity. However, the molecular mechanisms that lead to these toxic effects are not well known. In the present study, proteomic analysis has been performed to investigate the proteins that are differentially expressed in zebrafish embryos exposed to 0.5 mg/l PFOS until 192 h postfertilization. Two-dimensional electrophoresis coupled with mass spectrometry was employed to detect and identify the protein profiles. The analysis revealed that 69 proteins showed altered expression in the treatment group compared to the control group with either increase or decrease in expression levels (more than twofold difference). Of the 69 spots corresponding to the proteins with altered expression, 38 were selected and subjected to matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry (TOF/TOF) analysis; 18 proteins were identified in this analysis. These proteins can be categorized into diverse functional classes such as detoxification, energy metabolism, lipid transport/steroid metabolic process, cell structure, signal transduction, and apoptosis. Overall, proteomic analysis using zebrafish embryos serves as an in vivo model in environmental risk assessment and provides insight into the molecular events in PFOS-induced developmental toxicity.

Determination of odorous compounds in water by headspace solid-phase microextraction-gas chromatography-mass spectrometry

Three major odorous compounds are 2-methylisoborneol ( 2-MIB), geosmin and beta-cyclocitral, which in water were determined by coupling headspace solid-phase microextraction ( HS-SPME) with gas chromatography-mass spectrometry (GC-MS). The operating conditions of HS-SPME, such as fibre type, salt concentration, water temperature, stirring, absorption time and desorption time were studied and discussed.The highest absorption of the odorous compounds were obtained under the following operating conditions as the addition of 30% ( m/V) NaCl, stirring at 60 degrees C for 40 min, using 65 mu m polydimethyl siloxane/divinylbenzene coated fibre. After the odorous compounds had been absorbed in the fibre under the optimal conditions of HSSPME, they were desorbed at 250 degrees C and determined by GC-MS. The limits of detection for geosmin, beta-cyclocitral and 2-MIB in water were 1. 0, 1. 3, 1. 7 ng/L, and the relative standard deviations for them were 4. 9%, 8. 4%, 6. 2%,respectively. There were good linear correlation (the calibration coefficients were all above 0. 997) for the three odorous compounds in the range of 5 similar to 1000 ng/ L. Therefore, trace levels of the odorous compounds at ng/L in water could be quantified by the simple method with satisfactory result.

Synthesis and properties of novel polyimides derived from 2,2 ',3,3 '-benzophenonetetracarboxylic dianhydride

A new synthetic route to 2,2',3,3'-BTDA (where BTDA is benzophenonetetracarboxylic dianhydride), an isomer of 2,3',3',4'-BTDA and 3,3',4,4'-BTDA, is described. Single-crystal X-ray diffraction analysis of 2,2',3,3'-BTDA has shown that this dianhydride has a bent and noncoplanar structure. The polymerizations of 2,2',3,3'-BTDA with 4,4'-oxydianiline (ODA) and 4,4'-bis(4-aminophenoxy)benzene (TPEQ) have been investigated with a conventional two-step process. A trend of cyclic oligomers forming in the reaction of 2,2',3,3'-BTDA and ODA has been found and characterized with IR, NMR, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, and elemental analyses. Films based on 2,2',3,3'-BTDA/TPEQ can only be obtained from corresponding polyimide (PI) solutions prepared by chemical imidization because those from their polyamic acids by thermal imidization are brittle. PIS from 2,2',3,3'-BTDA have lower inherent viscosities and worse thermal and mechanical properties than the corresponding 2,3',3',4'-BTDA- and 3,3',4,4'-BTDA-based PIs. PIs from 2,2',3,3'-BTDA and 2,3',3',4'-BTDA are amorphous, whereas those from 3,3',4,4'-BTDA have some crystallinity, according to wide-angle X-ray diffraction.

Dibenzyl trithiocarbonate mediated reversible addition-fragmentation chain transfer polymerization of acrylonitrile

Reversible addition-fragmentation chain transfer polymerization has been successfully applied to polymerize acrylonitrile with dibenzyl trithiocarbonate as the chain-transfer agent. The key to success is ascribed to the improvement of the interchange frequency between dormant and active species through the reduction of the activation energy for the fragmentation of the intermediate. The influence of several experimental parameters, such as the molar ratio of the chain-transfer agent to the initiator [azobis(isobutyronitrile)], the molar ratio of the monomer to the chain-transfer agent, and the monomer concentration, on the polymerization kinetics and the molecular weight as well as the polydispersity has been investigated in detail. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and H-1 NMR analyses have confirmed the chain-end functionality of the resultant polymer.

Determination of ultra-trace concentrations of elements in high purity tellurium by inductively coupled plasma mass spectrometry after Fe(OH)(3) coprecipitation

In this work, a method was established for the determination of impurities in high purity tellurium by inductively coupled plasma mass spectrometry (ICP-MS) after Fe(OH)(3) coprecipitation. After comparison of coprecipitation ability and separation efficiency between Fe(OH), and Al(OH)(3), Fe(OH)(3) was chosen as the precipitate. A separation factor of 160 for 200 mg tellurium was obtained under conditions of pH 9 and 2 mg of Fe3(+). The 13 elements, such as Bi, Sn, Pb, In, Tl, Cd, Cu, Co, Ni, Zn, Ti, Be and Zr, could be almost completely coprecipitated under these conditions. In addition, Te memory effect imposed on the ICP-MS instrument was assessed, as well as Te matrix effect that caused the low recovery of Ga, As, Sb and V in real sample was discussed. Finally, the method was evaluated through recovery test and was applied to practical sample analysis, with detection limits of most of the elements being below 0.15 mug g(-1) and R.S.D. below or at approximately 10%, which indicated that this method could fully satisfy the requirements for analysis of 99.999% similar to 99.9999% high purity Te.