A matrix-assisted laser desorption/ionization tandem mass spectrometry method for direct screening of small molecule mixtures against an aminoglycoside kinase

Aminoglycoside phosphotransferase 3′IIIa (APH3′IIIa) is a bacterial enzyme involved in antibiotic resistance through phosphorylation of aminoglycosides, which can potentially be overcome by co-administration of an APH3′IIIa inhibitor with the antibiotic. Current assay methods for discovery of APH3′IIIa inhibitors suffer from low specificity and high false positive/negative hit rates. Here, we describe a method for screening APH3′IIIa inhibitors based on direct detection of kanamycin A phosphorylation using MALDI-MS/MS, which is more rapid than conventional assays and does not require secondary assays or sample cleanup. The MALDI-MS/MS assay operates at an ionic strength of 45 mM and co-factors can be utilized at near-physiological levels for optimal enzyme activity. Detection via MALDI-MS/MS allowed for improved reproducibility when compared to ESI-MS/MS. Furthermore, the use of MS/MS provided better signal-to-noise ratios relative to MS alone on the MALDI instrument. The assay was validated via generation of Z′-factors, with values of 0.78 and 0.56 in the absence and presence of 0.2% DMSO, respectively. The assay was used to screen a kinase directed library of >200 compounds, assayed as 21 mixtures of 10 compounds each. Five novel synthetic inhibitors were identified following mixture deconvolution. Inhibition constants were obtained for the aforementioned inhibitors using the MALDI-MS/MS assay, revealing several low to mid micromolar “hits”, and highlighting the quantitative nature of the assay.     

(BiO)2CO3-Bi-TiO2复合纳米纤维制备及其光催化降解抗生素

以静电纺丝制备的TiO₂纳米纤维为基质,葡萄糖为还原剂,在不同的酸碱环境中,采用一步水热法可控合成了异质结型Bi-TiO₂、(BiO)₂CO₃-TiO₂和(BiO)₂CO₃-Bi-TiO₂复合纳米纤维光催化剂。通过X射线粉末衍射(XRD)、X射线光电子能谱(XPS)、扫描电子显微镜(SEM)、透射电子显微镜(TEM)、紫外-可见漫反射光谱(UV-Vis DRS)和光致发光光谱(PL)等对样品进行表征。以洛美沙星、环丙沙星和诺氟沙星为目标污染物,研究了TiO₂及其复合纳米纤维的光催化降解性能,并探究其降解反应机理。结果表明,(BiO)₂CO₃-Bi-TiO₂光催化活性最高,模拟太阳光照60 min,对诺氟沙星、洛美沙星和环丙沙星的降解率分别达到93.2%、97.5%和100%。     

Der [15N]Methacetin-Leberfunktionstest zum Nachweis von Arzneimittelnebenwirkungen. Antibiotika-Therapie mit Amplicillin und mit Gefotaxime bei Neugeborenen

Abstract

The effect of soil temperature and moisture on plant growth and mineralisation of organic residues was investigated using ¹⁵N-labelled soybean residues and temperature-controlled tanks in the glasshouse. Treatments were arranged in a factorial design with: three soil temperatures (20, 26 and 30°C), two soil moisture regimes (8% (–800 Kpa) or 12% (–100 Kpa)), soybean residues added (enriched at 1.82 atom % ¹⁵N excess) or no residues; and either sown with ryegrass or not sown. Pots were sampled six weeks after planting and ¹⁵N-enrichment and δ¹³C of the plant and soil fractions were determined. Soil inorganic N was also periodically measured.

Available inorganic N increased significantly with addition of residues and generally decreased with increasing temperature. Plant dry matter decreased significantly with increase in soil temperature and increased with increasing moisture. Root-to-shoot ratio declined with increased temperature and moisture. Percentage nitrogen derived from residues (%Ndfr) increased linearly with increased temperature and moisture. Δ¹³C decreased linearly with increasing temperature and decreasing moisture status. There was a significant correlation between transpiration and dry matter production, but there was no correlation between water use efficiency and Δ¹³C.

The results suggest that C: N ratio of the root material effects the root turnover and in turn the water supply capacity of the root system.     

基于紫外LED阵列光催化降解抗生素的研究

抗生素的大量使用对生态环境造成巨大的影响,光催化技术具有操作简单且无二次污染等特点被广泛应用于污染物的降解。在光催化降解抗生素过程中,光源对其降解效率至关重要,与传统的汞灯催化光源相比,紫外LED技术具有更高的能源效率及更低的功耗,使光催化工艺发生了巨大的变化。首先建立基于紫外LED阵列的光催化平台,采用光栅光谱仪和紫外照度计对LED阵列光源光谱特性及装置内光场分布进行测量分析。结果显示紫外LED光源波长介于265～295 nm之间,其主波长为275 nm,由于光场叠加效果,光照强度随着装置径向位置距离的增大而明显增大,装置轴向位置光照强度分布较为均匀;其次通过三维超景深显微镜、UV-Vis光谱测量技术对P25型光催化剂的粒子结构进行表征分析,同时使用半导体求导公式对TiO₂粉末进行禁带分析,结果显示TiO₂为球形,由于空气中相对湿度过大,水在TiO₂微粒表面的润湿性加强了微粒间的粘附力,因此有团聚现象产生,其禁带宽度为3.1 eV;最后以紫外LED阵列和高压汞灯为催化光源,P25型TiO₂为催化剂分别对甲基橙、磺胺类抗生素进行光催化降解,使用紫外-可见光分光光度计测量降解过程中的吸收光谱曲线,进而对抗生素降解率进行分析。结果表明,甲基橙和磺胺二甲嘧啶在紫外LED阵列为光源条件下均能够被降解,分别经过160和240 min的催化降解过程后,降解率分别达到70%和36%,符合一级动力学方程,经计算LED阵列光源与汞灯对甲基橙的降解动力学常数分别为-0.007 5和-0.113 5 min-1,对磺胺二甲嘧啶的降解动力学常数分别为-0.001 9和-0.019 4 min-1。因此对甲基橙和磺胺二甲嘧啶进行降解时,汞灯降解速率高于紫外LED阵列;由于紫外LED阵列和汞灯系统在催化降解污染物过程中功率和其与反应器中轴线距离不同,对两种光源的抗生素降解效率建立评价方法,即对紫外LED和汞灯以单位功率为标准进行距离降解效率分析,对于甲基橙,汞灯在单位功率下的距离降解效率高于紫外LED,但对于抗生素,紫外LED阵列的距离降解效率明显高于汞灯。依据以上各类光谱分析和应用结果,紫外LED阵列是一种有竞争力的光催化应用替代光源,此技术的广泛应用为抗生素的降解提供新途径。     

Shorter puromycin analog synthesis by means of an efficient Staudinger-Vilarrasa coupling

An efficient Staudinger-Vilarrasa coupling generates amides from azides and 1-hydroxybenzotriazole esters of amino- or hydroxy acid derivatives in very high isolated yields and purity. New puromycin analogs, mostly putative biosynthetic intermediates, were synthesized in nine steps from adenosine.     

新型絮凝剂F－717对抗生素M－90发酵液预处理研究

本文设计了九种有机、无机絮凝剂对新抗Ｍ—９０发酵液预处理的方法研究，探索了以新型絮凝剂Ｆ－７１７与ＺｎＳＯ４、ＡｌＣｌ３等各种絮凝剂的加入量对发酵液滤速，光密度与效价的关系。     

Cross-metathesis approach to a (2E,4E)-dienoic acid intermediate for the synthesis of elaiolide

A (2E,4E)-7-hydroxy-2,4-dienoic acid, previously employed as a key intermediate for the total synthesis of the macrodiolide antibiotic elaiolide, was prepared stereoselectively and concisely from (S)-2-methyl-3-trityloxypropanal by a three-step sequence consisting of Brown’s asymmetric crotylboration, olefin cross-metathesis, and alkaline treatment. Ethyl 3-pivaloyloxy-4-pentenoate was used as a masked dienoate in the cross-metathesis step.     

Food safety evaluation: Detection and confirmation of chloramphenicol in milk by high performance liquid chromatography-tandem mass spectrometry

A simple and rapid procedure for extraction of chloramphenicol (CAP) in milk and analysis by high-performance liquid chromatography coupled with quadrupole mass spectrometry in tandem was developed. The method consisted of one step of liquid-liquid extraction using ethyl acetate and acidified water (10 mmol L⁻¹ formic acid) and HPLC-MS/MS detection. CAP-D5 was used as internal standard. The method was validated according to Commission Decision 2002/657/EC. The calibration curves were linear, with typical r² values higher than 0.98. Absolute recovery of CAP from milk proved to be more than 95%, however CAP-D5 absolute recovery was 75%. The method was accurate and reproducible, being successfully applied to the monitoring of CAP in milk samples obtained from the Brazilian market. Decision limit (CCα) was 0.05 ng mL⁻¹ and detection capability (CCβ) was 0.09 ng mL⁻¹.     

Feasibility of biofilm production capacity by Levilactobacillus brevis isolated from motal cheese and evaluation of biofilm resistance produced in vitro and in yogurt

Traditional dairy products are a unique source which have been considered for the extraction of indigenous probiotic strains in recent years. In this study, biofilm formation power of Levilactobacillus brevis that isolated from Mutal traditional cheese were investigated. Survival was assessed during 21 days of storage time and at the presence of residues antibiotics as well as gastrointestinal conditions. The results showed after 120 min of treatment in high acidic conditions (pH 2.0), the survival rate decreased only 0.75 log CFU/mL in biofilm formed. The antibiotic susceptibility evaluated of probiotic to enrofloxacin, sulfadimidine, tetracycline, and oxytetracycline showed reducing the bacterial population in the biofilm form only 2.6 log. probiotic strain that isolated from indigenous dairy sources showed excellent resistance in the biofilm state. Therefore, extracting strong probiotic strains from indigenous resources, it can significantly improve functional products and fermentory engineering.     

The use of NIR as a multi-parametric in situ monitoring technique in filamentous fermentation systems

The use of Fourier transform near infrared (FT-NIR) spectroscopy for simultaneous determination of multiple properties in an active pharmaceutical ingredient (API) fermentation process is described, together with procedures for developing accurate NIR calibrations with a performance independent of scale and the specific bioreactor used. Measurements were made in situ, by insertion of transflection probes into pilot and industrial bioreactors providing direct contact with the fermentation culture media. The ultimate goal was to establish methods for real time process monitoring aimed at enhanced process supervision, fault detection diagnosis and control of bioreactors. The in situ acquired spectra were related to lab results of samples taken from the reactors during the course of the manufacturing process. Suitable spectral wavenumber regions were selected and calibration models based on partial least squares (PLS) were developed. The root mean square errors of prediction for API content, viscosity, nitrogen source and carbon source concentration were all within acceptable ranges as compared to the off-line lab measurements, respectively, 0.03% (w/w), 150 cp, 0.01% (w/w), and 0.4% (w/w).