固相萃取-超高压液相色谱-串联质谱测定水中19种抗生素

应用固相萃取(SPE)及液相色谱-串联质谱(LC-MS/MS)技术,建立了水中痕量(ng/L)四环素类、磺胺类、大环内酯类、喹诺酮类和β-内酰胺类5类共19种抗生素的同时定量检测方法。水样通过HLB萃取小柱富集后,以C18反相色谱柱为分析柱,乙腈-0.1%甲酸溶液为流动相,采用LC-MS/MS进行定量分析。选择电喷雾正电离源(ESI+),多反应监测模式(MRM),内标法定量。19种抗生素在0.5~1 000μg/L范围内均具有良好的线性关系,方法的定量下限(S/N=10,1 000倍浓缩)为0.1~0.5 ng/L。以纯水和河水(黄浦江水)作为基底,13C-咖啡因为内标物,加标质量浓度为20、100 ng/L时,抗生素的平均加标回收率分别为75%~125%和77%~132%,相对标准偏差(RSD)分别为1.7%~6.9%和0.9%~6.5%,表明所建立的测试方法准确可靠。研究结果表明,黄浦江水受到了抗生素污染,共检出15种抗生素,检出的四环素类、磺胺类、大环内酯类、喹诺酮类及β-内酰胺类抗生素污染质量浓度分别为13.0~56.9、12.2~103.4、53.8~84.8、3.1~26.2、16.5~181.6 ng/L。     

Targeting Bacterial Membranes: NMR Spectroscopy Characterization of Substrate Recognition and Binding Requirements of D‐Arabinose‐5‐Phosphate Isomerase

Lipopolysaccharide (LPS) is an essential component of the outer membrane of Gram‐negative bacteria and consists of three elements: lipid A, the core oligosaccharide, and the O‐antigen. The inner‐core region is highly conserved and contains at least one residue of 3‐deoxy‐D ‐manno‐octulosonate (Kdo). Arabinose‐5‐phosphate isomerase (API) is an aldo–keto isomerase catalyzing the reversible isomerization of D ‐ribulose‐5‐phosphate (Ru5P) to D ‐arabinose‐5‐phosphate (A5P), the first step of Kdo biosynthesis. By exploiting saturation transfer difference (STD) NMR spectroscopy, the structural requirements necessary for API substrate recognition and binding were identified, with the aim of designing new API inhibitors. In addition, simple experimental conditions for the STD experiments to perform a fast, robust, and efficient screening of small libraries of potential API inhibitors, allowing the identification of new potential leads, were set up. Due to the essential role of API enzymes in LPS biosynthesis and Gram‐negative bacteria survival, by exploiting these data, a new generation of potent antibacterial drugs could be developed.     

A switch-on fluorescence assay for bacterial β-lactamases with amyloid fibrils as fluorescence enhancer and visual tool

Herein is described the development of a novel switch-on fluorescence assay for detecting β-lactamases. The fluorescence assay comprises two components: solid beads coated with a β-lactam antibiotic, which is linked to an environment-sensitive fluorophore (dansylaminothiophenol, DTA), and amyloid fibrils of hen lysozyme (acting as fluorescence enhancer and visual tool). In the presence of the clinically significant TEM-1 β-lactamase, the DTA-antibiotic complex on the solid beads is hydrolyzed, thus releasing the DTA dye into solution. The DTA dye is only weakly fluorescent in solution but gives strong green fluorescence upon binding to lysozyme fibrils. These strongly fluorescent DTA-bound fibrils can be easily visualized by the naked eye upon illumination of the sample with a simple UV lamp. The fluorescence assay can detect TEM-1 at low concentration (0.01 nM). In contrast, no observable fluorescence appears when the fluorescence assay is performed on samples without the TEM-1 β-lactamase.     

脂肪醇聚醚产品及脂肪醇羟值的快速测定

以4-二甲氨基吡啶为催化剂,过量乙酸酐为乙酰化试剂,吡啶为溶剂,用电位滴定法快速测定脂肪醇聚醚产品及脂肪醇羟值,并对催化剂、反应条件、系统的适应性进行了探讨。用该方法对5种不同羟值的脂肪醇聚醚及脂肪醇样品进行了测定,测定结果的相对标准偏差小于0.5%(n=6),且测定结果与国标法测定结果相符合,相对误差在±1%之内。     

An amperometric penicillin biosensor with enhanced sensitivity based on co-immobilization of carbon nanotubes, hematein, and β-lactamase on glassy carbon electrode

An amperometric penicillin biosensor with enhanced sensitivity was successfully developed by co-immobilization of multi-walled carbon nanotubes (MWCNTs), hematein, and β-lactamase on glassy carbon electrode using a layer-by-layer assembly technique. Under catalysis of the immobilized enzyme, penicillin was hydrolyzed, decreasing the local pH. The pH change was monitored amperometrically with hematein as a pH-sensitive redox probe. MWCNTs were used as an electron transfer enhancer as well as an efficient immobilization matrix for the sensitivity enhancement. The effects of immobilization procedure, working potential, enzyme quantity, buffer concentration, and sample matrix were investigated. The biosensor offered a minimum detection limit of 50 nM (19 μg L⁻¹) for penicillin V, lower than those of the conventional pH change-based biosensors by more than two orders of magnitude. The electrode-to-electrode variation of the response sensitivity was 7.0% RSD.     

Fluoroquinolone antibiotics in environmental waters: Sample preparation and determination

The aim of this review is to provide a general overview on the analytical methods proposed in the last decade for trace fluoroquinolone (FQ) determination in environmental waters. A large number of studies have been developed on this topic in reason of the importance of their monitoring in the studies of environmental mobility and potential degradation pathways. Every step of the analysis has been carefully considered, with a particular attention to sample preparation, in relationship with the problems involved in the analysis of real matrices. The different strategies to minimise interference from organic matter and to achieve optimal sensitivity, especially important in those samples with lower FQ concentrations, were also highlighted. Results and progress in this field have been described and critically commented. Moreover, a worldwide overview on the presence of FQs in the environmental waters has been reported.     

Development and validation of an improved reversed‐phase liquid chromatographic method combined with pulsed electrochemical detection for the analysis of netilmicin

Netilmicin is one of the aminoglycoside antibiotics that lacks a strong UV absorbing chromophore. However, the application of pulsed electrochemical detection has been used successfully for the direct analysis of aminoglycoside antibiotics. This study describes an improved LC method combined with pulsed electrochemical detection for the analysis of netilmicin. Using a Zorbax SB C‐18 column (250 mm×4.6 mm id, 5 μm), isocratic elution was carried out with a mobile phase containing sodium sulfate (20 g/L), sodium octanesulfonate (0.3 g/L), THF (20 mL/L), and 0.2 M phosphate buffer pH 3.0 (50.0 mL/L). The robustness of the method was examined by means of an experimental design. The method proved to be sensitive, repeatable, linear, and robust. The method has also been used to analyze some commercial netilmicin samples.     

Highlighting the possible secondary interactions in the role of balhimycin and its analogues for enantiorecognition in capillary electrophoresis

It is believed that the enantiorecognition mechanism based on macrocyclic antibiotics involves multimodal interactions via hydrogen bonding, π–π interaction, steric hindrance, hydrophobic interaction and so on. A variety of enantiomeric N-benzoylated amino acids were separated using balhimycin (A) or its analogues bromobalhimycin (B) and dechlorobalhimycin (C) as chiral mobile phase additive using a CE method, which combined the partial filling technique with the dynamic coating technique and the co-EOF electrophoresis technique. The enantioresolution and the migration time were highly relevant to the structure of analytes, especially to the substitutions on the N-tagged benzoyl moiety of the amino acids. A steric effect and π–π interaction based mechanism is proposed in order to explain some observed enantioresolution differences between positional isomers. Notably dechlorobalhimycin exhibited the best enantioresolution for several N-benzoylated derivatives of leucine, which was rarely observed for N-dansylated amino acid derivatives. The hydrophobicity difference of the aglycone pocket among three chiral selectors was assumed to account for this behaviour.