Molecular and nanoparticle postcolumn reagents for assay of low-molecular-mass biothiols using high-performance liquid chromatography

Determination of low-molecular-mass (LMM) biothiols in biological matrixes is of importance in the studies of their related bio-processes and for the clinical diagnostics of a variety of diseases. Standard method for the assay of the small biothiols is in demand. Postcolumn techniques used in high-performance liquid chromatography (HPLC) allow automation of the derivatization step and, therefore, are suitable for standardization of HPLC analysis. This paper gives an overview of the existing reaction systems useful for the postcolumn assay of the LMM biothiol molecules in conjunction with HPLC. The postcolumn reagents are classified by the types of their reactions with thiol-containing compounds. The chemical reactivity and selectivity as well as the spectroscopic characteristics of the postcolumn reagents have been addressed. The emerging strategies of using nanoparticles as thiol-reactive reagents and their applications in postcolumn detection of the LMM biothiols have also been discussed in detail.     

Gene detection of staphylococcal enterotoxins in production strain of staphylococcin injection and superantigenic activity of rSEK and rSEQ

In China, staphylococcin injection has been commonly used in the combined treatment of cancer to enhance the systemic immune response and reduce the toxicities associated with chemotherapy and radiation therapy. It is claimed that the main active component in the injection is staphylococcal enterotoxin C2 (SEC2). To determine whether other serological types of staphylococcal enterotoxins (SEs) could also be present in the injection products, in this study, the distribution of se genes (from sea to see, from seg to seu) in the one and only production strain of Staphylococcus aureus from one manufacturing company was analyzed by PCR method. In addition, sek and seq genes were cloned from the strain and the corresponding recombinant proteins, rSEK and rSEQ, were expressed in Escherichia coli and purified by affinity chromatography and anion-exchange chromatography. The superantigenic properties of the two recombinant proteins were then measured by MTT method. The PCR results showed that seven se genes are harbored by the production strain. However, sec2 gene was not detected. The results of MTT assay showed that rSEK and rSEQ could elicit strong stimulatory effects on proliferation and cytotoxicity of murine splenocytes in vitro. Overall, the results in this study indicated that one or a plurality of the seven SEs may be present in the related products, and that the two recombinant SEs are promising candidates as immunomodulatory agents for cancer therapy.     

重金属污染土壤稳定/固化修复技术研究进展

修复重金属污染土壤一直是国际上的难点和热点研究课题.目前常用的污染场地修复技术主要包括挖掘、稳定/固化(solidification/stabilization,S/S)、化学淋洗、气提、热处理、生物修复等.本文在参考美国环境保护署(EPA)、英国环境署的S/S技术规范、国内外发明专利基础上,对S/S的概念、国内外发展现状及今后的发展方向进行了系统论述.固定化技术通过把污染物囊封入惰性基材中,或在污染物外面加上低渗透性的材料,来减少污染物暴露的淋滤面积以达到限制污染物迁移的目的.稳定化技术是从改变污染物的有效性出发,将污染物转化为不易溶解、迁移能力或毒性更小的形式.S/S技术包括:水泥固化、石灰火山灰固化、塑性材料包容固化、玻璃化技术、药剂稳定化.在稳定化技术中,加入药剂的目的是改变土壤的物理、化学性质,通过pH控制技术、氧化还原电势技术、沉淀技术、吸附技术、离子交换技术等改变重金属在土壤中的存在状态,从而降低其生物有效性和迁移性.本文还论述了S/S修复效果评价方法,并指出需加强S/S技术中的分子键合技术、土聚合物以及我国的S/S技术导则制定等工作.     

Quantification of CPT13 in rat plasma using LC-MS/MS for a pharmacokinetic study

A new, simple, sensitive and specific reversed-phase high performance liquid chromatographic (HPLC) method using tandem mass spectrometry detection was initially developed and validated for the analysis of 10-(2-pyrazolyl-ethoxy)-(20S)-camptothecin (CPT13) in rat plasma. Pretreatment of the sample obtained from plasma involved a single protein precipitation step with using acetonitrile containing 0.1% formic acid. An aliquot of 20 μl was injected into a C-18 column. The chromatographic separation was achieved using the mobile phase consisting of acetonitrile:water (35:65) at a flow rate of 1.0 mL/min. The total run time for each sample was 10 min, and camptothecin (CPT, IS) and CPT13 were well separated with retention times of 5.1 min and 5.6 min, respectively. Detection was performed using a triple quadrupole tandem mass spectrometer in multiple reaction monitoring (MRM) mode via an electrospray ionization (ESI) source. The calibration curve was linear (r2 = 0.9998) over the concentration range of 1-1000 ng/mL, with a LLOQ of 1 ng/mL for CPT13. The inter- and intra-day precision (%R.S.D.) were <2.58% and 6.28%, respectively, and the accuracies (%) were within the range of 97.34-110.67%. CPT13 in rat plasma was stable when stored at -20 °C or 4 °C for three freeze-thaw cycles, The method was employed for the first time during pharmacokinetic studies of CPT13 in rats following a single intravenous dose (0.1 mg/kg) and three different oral doses (50 mg/kg, 30 mg/kg, and 10 mg/kg). This fully validated method was successfully applied to a pharmacokinetic study of CPT13 in rats.     

Seroprevalence of pandemic (H1N1) 2009 in pregnant women in China: an observational study

Background

We investigated the seropositive rates and persistence of antibody against pandemic (H1N1) 2009 virus (pH1N1) in pregnant women and voluntary blood donors after the second wave of the pandemic in Nanjing, China.

Methodology/Principal Findings

Serum samples of unvaccinated pregnant women (n = 720) and voluntary blood donors (n = 320) were collected after the second wave of 2009 pandemic in Nanjing. All samples were tested against pH1N1 strain (A/California/7/2009) with hemagglutination inhibition assay. A significant decline in seropositive rates, from above 50% to about 20%, was observed in pregnant women and voluntary blood donors fifteen weeks after the second wave of the pandemic. A quarter of the samples were tested against a seasonal H1N1 strain (A/Brisbane/59/2007). The antibody titers against pH1N1 strain were found to correlate positively with those against seasonal H1N1 strain. The correlation was modest but statistically significant.

Conclusions and Significance

The high seropositive rates in both pregnant women and voluntary blood donors suggested that the pH1N1 virus had widely spread in these two populations. Immunity derived from natural infection seemed not to be persistent well.     

Characterization of five fungal endophytes producing Cajaninstilbene acid isolated from pigeon pea [Cajanus cajan (L.) Millsp

Five fungal endophytes (K4, K5, K6, K9, K14) producing Cajaninstilbene acid (CSA, 3-hydroxy-4-prenyl-5-methoxystilbene-2-carboxylic acid) were isolated from the roots of pigeon pea [Cajanus cajan (L.) Millsp.]. CSA is responsible for the prominent pharmacological activities in pigeon pea. The amount of CSA in culture solution varied among the five fungal endophytes. K4 produced the highest levels of CSA (1037.13 μg/L) among the endophytes tested after incubation for five days. Both morphological characteristics and molecular methods were used for species identification of fungal endophytes. The five endophytic isolates were characterized by analyzing the internal transcribed spacer (ITS) rRNA and β-tubulin genes. The K4, K5, K9 and K14 strains isolated from pigeon pea roots were found to be closely related to the species Fusarium oxysporum. K6 was identified as Neonectria macrodidym. The present study is the first report on the isolation and identification of fungal endophytes producing CSA in pigeon pea. The study also provides a scientific base for large scale production of CSA.     

A method for fabricating uni-dsDNA microarray chip for analyzing DNA-binding proteins

This paper describes an approach for preparing unimolecular double-stranded DNA (uni-dsDNA) microarray chip. In this method, the various target oligonucleotides containing a reverse complementary sequence at 5' end were firstly annealed to a same universal oligonucleotide with amino group at 5' end and immobilized on aldehyde-derivatized glass slide. An on-chip DNA polymerization reaction was then performed to elongate the universal oligonucleotides. After a denaturation and a followed intra-strand annealing, a hairpin structure was formed at the free 3' end of the immobilized oligonucleotides. Finally, another on-chip DNA polymerization was done to synthesize the uni-dsDNA microarray. Combining with a PCR amplification of chemically synthesized target oligonucleotides, this method was much cost-effective for production of the uni-dsDNA microarray. The uni-dsDNA microarray was verified applicable for detecting the presence and monitoring the DNA-binding activity of the sequence-specific DNA-binding proteins.     

High affinity iron(III) scavenging by a novel hexadentate 3-hydroxypyridin-4-one-based dendrimer: synthesis and characterization

The synthesis of a novel iron(III)-selective hydroxypyridinone hexadentate-terminated dendritic chelator based on a benzene tricarbonyl core polyamine dendrimer is described. The iron-chelating ability of the dendritic chelator was demonstrated by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and UV-vis spectroscopy. The physicochemical properties of the isolated hexadentate unit were also investigated. The dendrimer was found to possess an extremely high affinity for iron(III), namely logK=34.8, pFe3+=30.6.     

Determination of enantiomeric composition by negative-ion electrospray ionization-mass spectrometry using deprotonated N-(3,5-dinitrobenzoyl)amino acids as chiral selectors

The ability to use mixtures of deprotonated N-(3,5-dinitrobenzoyl)amino acids as chiral selectors for the determination of enantiomeric composition by electrospray ionization-mass spectrometry is demonstrated. For each experiment, two N-(3,5-dinitrobenzoyl)amino acids were chosen such that each would have opposite selectivity for the enantiomers of the analyte. Electrospray ionization-mass spectrometry, monitored in the negative ion mode, of solutions containing the two N-(3,5-dinitrobenzoyl)amino acids, sodium hydroxide, and the analyte, in a one-to-one mixture of methanol and water, afford peaks in the mass spectrum that correspond to the deprotonated 1:1 analyte-selector complexes. The ratio of the intensities of the complexes in the mass spectrum can be related to the enantiomeric composition of the analyte. Additionally, the sense and extent of chiral recognition is consistent with chromatographic observations, using chiral stationary phases derived from N-(3,5-dinitrobenzoyl)amino acids. Each analysis of enantiomeric composition requires less than 10 s to complete, indicating that this method has great potential for the development of fast-/high-throughput chiral analyses.     

Volume interpolation of CT images from tree trunks

Computerized tomography as a non-destructive scanning method to analyze wood structures has become an important technique in tree research. The possibility to reconstruct three-dimensional volumes based on a number of slices of two-dimensional data from CT scans is strongly dependent on the number of measured slices. Radial basis function methods can be successfully used to interpolate CT images with the aim of obtaining a satisfactory reconstruction of tree trunks. In contrast to standard interpolation techniques, our method takes into account that wood structures differ more in the radial than in the longitudinal direction. Therefore we obtain better interpolation results for wood structures.