万古霉素在食源性致病菌检测中的应用

食源性致病菌污染是引起食品安全的主要因素之一,对人类健康构成了严重威胁。万古霉素作为一种对革兰氏阳性菌具有特异性识别功能的小分子,能够修饰在磁性纳米粒子表面,使粒子获得靶向目标菌的能力,结合其他检测方法,可以实现分离检测革兰氏阳性菌的目的。本文对万古霉素功能化的磁珠在革兰氏阳性菌分离检测的研究进展进行了介绍,分析了万古霉素不同修饰方式和检测方法对其检测效果的影响,并对各种方法的检测结果进行了分析比较,最后,提出了万古霉素在食源性致病菌检测中存在的问题,并对其未来应用进行了展望。     

Analysis of delta-L-alpha-aminoadipyl-L-cysteinyl-D-valine by ion chromatography and pulsed amperometric detection

A novel high-performance liquid chromatography (HPLC) method is presented for the detection and trace level determination of the tripeptide delta-L-alpha-aminoadipyl-L-cysteinyl-D-valine (ACV). The tripeptide, an intermediate in penicillin production, is derived from fungal fermentation. The technique relies on ion-exchange separation of the tripeptide on an anion-exchange column followed by detection by reduction on a gold electrode using pulsed amperometry. The sensitivity of direct determination of ACV is increased by employing pulsed amperometric detection (PAD) over direct ultraviolet detection. Choice of the working electrode and optimisation of electrode potentials was based on cyclic voltammograms recorded for the tripeptide in the mobile phase. A linear regression equation was obtained over the range 0-100 micrograms ml-1. The detection limit in fermentation broths was found to be 0.1 micrograms ml-1 whereas in buffer the detection limit was found to be 10 ng ml-1. A good correlation coefficient was observed when ACV concentrations determined by ion chromatography-PAD were compared with measurements obtained by pre-column derivatisation with fluoromethylorthochloroformate followed by HPLC separation on a reversed-phase C18 silica column with UV detection. The procedure has been applied to the measurement of natural levels of ACV in fermentation broths of selected strains of Aspergillus nidulans and Penicillin chrysogenum.     

Integrated capillary electrophoresis amperometric detection microchip with replaceable microdisk working electrode. II. Influence of channel cross-sectional area on the separation and detection of dopamine and catechol

The interference of separation high voltage with the electrochemical detection is a major challenge to the microchip capillary electrophoresis-electrochemical detection systems with end-channel detection mode. Using dopamine and catechol as model analytes, the influences of channel cross-sectional area and channel-to-electrode distance on the high-voltage interference, accordingly on the separation and detection performances of the microchip capillary electrophoresis-electrochemical detection system were investigated. With the increase of the channel cross-sectional area from 312 through 450-615 microm2, the apparent half-wave potentials of hydrodynamic voltammetry for dopamine at the field strength of 288 V/cm shifted positively from 285 through 330-400 mV. By using a chip with the smallest channel cross-section (312 microm2 with top width of 37.3 microm and depth of 8.9 microm) the residual high-voltage field in the detection cell was small, so that detection was conducted at a channel-to-electrode distance of 20 microm to achieve better performances of separation and detection.     

基于氧化还原反应界面可视化定量检测辣根过氧化物酶

该文建立了一种可视化的、基于氧化还原反应界面移动距离定量检测辣根过氧化物酶(HRP)的方法。比较了隐色结晶紫显色体系和3,3′,5,5′-四甲基联苯胺显色体系对HRP的显色效率,并构建了基于3,3′,5,5′-四甲基联苯胺显色体系的氧化还原反应电泳滴定模型。同时,文中还设计了适用于该模型的小型化、便携式滴定检测芯片,并对滴定通道凝胶中组分进行了优化。结果表明,界面移动距离与HRP浓度存在对数线性关系,检测灵敏度可达0.002 mg/L,且可在10 min内完成HRP的裸眼检测。该方法不需要配备信号读取装置,用户只需要读取有色界面移动的距离即可实现对待测物的可视化定量检测,对于即时检测具有潜在的应用价值。     

基于智能手机数字比色法的有机磷农残快速检测技术研究

提出了基于智能手机实现有机磷农残的快速定量检测方法,分析了智能手机的性能对检测结果的影响;并利用LED面光源解决了因照明条件的变化所引起的采集图片颜色不均匀的问题,提高了检测的准确度。实验将LED面光源垂直放置于样品上方,智能手机置于样品正前方对有机磷农药样品进行图片采集,检测结果与紫外可见分光光度计所测结果一致,二者具有良好的线性相关度。     

Simultaneous HPLC determination of phenylurea herbicides and their corresponding anilines in water after on-line preconcentration

On-line preconcentration on a short C₁₈ column, prior to HPLC with UV and electrochemical detection, has been used for determination of some phenylurea herbicides and their possible degradation products, substituted anilines, in water samples. With electrochemical detection the detection limit at a signal-to-noise ratio of 3 was 5 ppt for 4-chloroaniline and 4-bromoaniline and 7 ppt for 3,4-dichloroaniline; with UV detection the detection limit was ca 300 ppt for all analytes.     

Comparison of high-selectivity gas chromatographic methods, including column switching, for the determination of felodipine in plasma

The selectivity required for the determination of low concentrations of felodipine in plasma was achieved by either mass-selective detection, optimization of stationary phase selectivity or column-switching gas chromatography (GC) with a dual-oven chromatograph, the latter two with electron-capture detection. The three approaches were evaluated in terms of selectivity, detectability, precision and suitability for routine applications with automated injection. Using mass-selective detection, the detectability in plasma samples was limited by the performance of the mass spectrometer. The detection limit (signal-to-noise ratio = 3) was 4.7 pmol (1.8 pg) of felodipine. Pre-concentration of extracts permitted quantitation in plasma down to 0.2 nmol/1. Using electron-capture detection, the detectability was determined by the selectivity and bleeding characteristics of the columns. For single-column separation, a 35% phenyl phase was selected. The detection limit was 3.0 fmol (1.2 pg). The limit of quantitation in plasma was 1 nmol/1. In column-switching GC, bleeding products from the first column will separate on the second column and may interfere in separations for trace analysis. Bleeding products from a 50% phenyl phase (DB-17) were characterised by GC-mass spectrometry. With a dual-column system, employing a DB-17 (50% phenyl) column for selective introduction on to a CP-Sil 5 (0% phenyl) column, the signal-to-noise ratio was limited by the low-bleeding second column, provided that the bleeding products from the first column were adequately separated from felodipine. The detection limit in this instance was significantly lower 0.35 fmol (0.13 pg). Direct injection of plasma extracts permitted quantitation down to 0.4 nmol/l. All three methods were well suited for use with autosamplers.     

Light-scattering and turbidimetric detection of silica colloids in size-exclusion chromatography

Silica colloids were separated by size-exclusion chromatography and monitored by fluorimetric and UV detection. In the former means of detection, silica colloids were visualized by light-scattering. The signal intensity based on the light scattering increased with increasing size of the silica colloids. The maximum intensity was observed at excitation wavelengths around 270–290nm. In UV detection, silica colloids were visualized based on turbidimetry, and the signal intensity also increased with increasing size of the silica colloids and with decreasing detection wavelength. The signal intensities for both light-scattering and turbidimetric detection were a linear function of the concentration of the silica colloids. The detection limit at S/N = 3 for 78-nm colloids was 0.06 ppm for light-scattering detection whereas the LOD was 2.3 ppm for UV detection. Effects of mobile phase conditions and flow rate on resolution and peak shape were examined. Use of phosphate buffer allowed the separation of silica colloids of different sizes in size-exclusion chromatography.     

Early detection of viral DNA in breast cancer using fingered aluminium interdigitated electrode modified by Streptavidin-biotin tetravalent complex

Human Mammary Tumor Virus (HMTV) or Mouse Mammary Tumor Virus holds similarity as an endogenous onco-retrovirus belongs to retroviridae family, predominantly infects the epithelial cell of human as well as mouse. With the recognition of nano-biosensor in nanotechnology, ideal interdigitated electrode (IDE) was genuinely performed for HMTV detection. Aluminium enriched IDE (AlIDE) was fabricated for high performance detection with a cost-effective photolithography technique. In this research, (3-glycidyloxypropyl) trimethoxysilane refined platform was selected to detect the conductivity with HMTV target DNA interaction on the designed AlIDE. Strong binding affinity of streptavidin-biotin with target DNA enhanced the sensitivity by empowering higher number of HMTV probe and target complementation on sensing surface. Furthermore, the target DNA was immobilized on probe modified AlIDE and a quantitative value of 100 aM attained as a lowest detection. A linear with dose-dependent duplex formation was shown with the regression coefficient value of 0.964. Negative control has shown insignificant detection at 10 pM, which justifies the higher fold discrimination with specificity. The excellence of AlIDE performance in detection of HMTV may pave the way for more verification on other diseases.     

Detection of α‐Methylacyl‐CoA Racemase in Serum and Urine Using a Highly Sensitive Electrochemical Immunodetector

The detection of α‐methylacyl‐CoA racemase (AMACR), a novel biomarker for prostate cancer, is demonstrated in serum and urine using a novel immuno‐detection method. The detection system consists of a three‐electrode conventional electrochemical cell modified with a gating electrode for applying a gating voltage V_G to the immune complex immobilized on the working electrode to provide signal amplification. The detection system is realized by integrating gating electrodes with screen‐printed electrodes. This detection method does not require involved sample preparation procedures. The detection was demonstrated in serum and urine samples on the nanogram/mL level with V_G equal to 0.6 V. Detection in serum was also performed on the picogram/mL level with a limit of 100 picogram/mL with V_G=0.6 V being a necessary condition.     

Electrochemical probe DNA design in PCR amplicon sequence for the optimum detection of microbiological diseases

Direct electrochemical genosensor was developed for the detection of a probe sequence relative position in a PCR amplicon for the optimum detection of bacterial and microbiological diseases, in this study. The genosensor relies on a label-free electrochemical detection. The amino-linked inosine modified (guanine-free) coequal capture probes which were chosen from different parts of a PCR amplicon, immobilized on to disposable pencil graphite electrodes (PGE) by electrostatically and covalently. As a model case Hepatitis B virus (HBV) genome amplicon was used for the detection and specification. Hybridization was occurred after surface coverage with denatured amplicons. After hybridization, optimum probe sequence position was identified by using the differences between the responses of guanine oxidation signals. The results of this study might have a great convenience for the microbiological diseases detection applications such as DNA micro arrays.     

HPLC/UV-spectrometric and electrochemical detection of lignin-decomposition products in alcoholic beverages

Summary The determination of compounds responsible for the flavour of alcoholic beverages, which were separated by HPLC-RP-chromatography, was performed with a combination of UV-diode-array and electrochemical detection. The recording and comparing of diode-array spectra on the rising and descending flanks of each peak was very suitable for identifying peak impurities. The selectivity of the detection depends strongly on the detection wavelength, detection potential and electrode material. For higher sensitivity the electrochemical detection is favoured. The lowest detection limit is 30 pg (absolute) for syringic acid.     

An indicator-displacement assay for naked-eye detection and quantification of histidine in human urine

A simple and efficient colorimetric method for the naked-eye detection and quantification of histidine in biological fluids was developed based on an indicator-displacement assay (IDA) and the Ni(2+)-histidine affinity pair. In this IDA approach, a commercially available dye, murexide, was used as the indicator and the selective detection of histidine was achieved based on the competition between indicator and histidine for the binding with Ni(2+). The competition of histidine with murexide for Ni(2+) resulted in an obvious color change of the solution from yellow to purple, and the permitted naked-eye detection of trace histidine. The developed bioassay allows the rapid, sensitive and selective detection of histidine in urine samples, and does not need complicated sample pretreatment. The detection limit was 0.4 μM with a linear range from 2 to 30 μM. The relative standard deviation for 11 replicate detections of 8 μM histidine was 2.0%. The developed sensor was successfully applied to the determination of histidine in human urine samples with recoveries from 97 to 105%.     

Determination of midazolam in rabbit plasma by GC and LC following nasal and ocular administration

GC with nitrogen phosphorus detection and HPLC with UV detection were used to determine midazolam (MDZ) levels in rabbit plasma following ocular and nasal administration. For GC with nitrogen phosphorus detection, the analyte was extracted from the plasma using a three‐step liquid–liquid extraction including extraction with an isopropanol/butyl chloride mixture in an alkaline solution, followed by extractions with 1 M HCl, and finally with an alkaline solution of butyl chloride. The recovery of MDZ was dependent on the sample alkalization time prior to the final extraction. The procedure increased the recovery of MDZ up to 99.6%. Improved sample preparation led to a significant increase in the sensitivity of the determination by GC with nitrogen phosphorus detection. The achieved detection limit was 0.34 ng/mL, which is ten times lower than that obtained using HPLC with UV detection. The small plasma volume was another advantage of the GC with nitrogen phosphorus detection method (200 μL per assay). Both administration routes of the anesthetic (nasal and ocular) resulted in comparable plasma MDZ levels. Kinetic simulation of the MDZ plasma was performed for both administration routes.     

Pulsed amperometric detection of carbohydrates on an electrophoretic microchip

The first report of pulsed amperometric detection (PAD) on an electrophoretic microchip is presented. A hybrid poly-(dimethylsiloxane)/glass device was coupled with a platinum working electrode for the electrochemical detection of glucose, maltose, and xylose. Under optimized detection conditions, glucose was found to respond linearly from 20 to 500 microM with a measured detection limit of 20 microM. The coupling of PAD with a microchip provides a straightforward approach to the analysis of a wide range of carbohydrates using microfluidics.     

Cyclic chronopotentiometric determination of sugars at Au and Pt microelectrodes in flowing solutions

The main advantage of the application of cyclic chronopotentiometry (CCP) in end-column CE detection arises from the fact that the detection parameters and the magnitude of the analytical signal are (in contrast with other electrochemical detection methods) independent of the ohmic polarization of the solution caused by the separation current at the detection end of the capillary. CCP was used to determine sugars on platinum and gold microelectrodes after separation by CE. The results obtained with a gold microelectrode were better. Subsequently this detection method was used for quantitative determination of sugars in honeys and for their authentication.     

Analysis of baclofen by capillary electrophoresis with laser-induced fluorescence detection

A new analytical method for baclofen (4-amino-3-p-chlorophenylbutyric acid) based on capillary electrophoretic separation and laser-induced fluorescence detection has been developed. Naphthalene-2,3-dicarboxaldehyde was used for precolumn derivatization of the non-fluorescent drug. Optimal separation and detection were obtained with an electrophoretic buffer of 50 mM sodium borate (pH 9.5) and a He-Cd laser (excitation at 442 nm, emission at 500 nm). Linearity (r > or = 0.99) over three orders of magnitude was generally obtained and the concentration limit of detection was in the nanomolar level. Coupled with a simple cleanup procedure, the method was successfully applied to the analysis of baclofen in human plasma. Recovery of spiked baclofen in plasma was 98%. The relative standard deviation values on peak size and migration time were 7.9% and 0.4%, respectively. The limit of detection of baclofen in plasma was 10 ng/ml.     

Benefits and Pitfalls of HPLC Coupled to Diode-Array,Charged Aerosol,and Coulometric Detections: Effect of Detection on Screening of Bioactive Compounds in Apples

The new screening method for rapid evaluation of major phenolic compounds in apples has been developed. Suitability of coupling HPLC/UHPLC separation with the diode-array detection and universal charged aerosol detection with respect to the presence of interfering substances was tested. Characteristics of both detection techniques were compared and method linearity, limits of detection and quantitation, and selectivity of them determined. Student t-test based on slopes of calibration plots was applied for the detailed comparison. The diode-array detection provided the best results regarding sensitivity and selectivity of the developed method in terms of evaluation of phenolics profiles. The response of the charged aerosol detector was negatively affected by co-eluting substances during rapid-screening analyses. Coulometric detection was used for advanced characterization of extracts in terms of antioxidant content and strength to obtain more complex information concerning sample composition. This detection also allowed evaluation of unidentified compounds with antioxidant activity. HPLC/UHPLC separation using a combination of diode-array and coulometric detectors thus represented the best approach enabling quick, yet complex characterization of bioactive compounds in apples.     

The use of polyethyleneimine for fabrication of potentiometric cholinesterase biosensors

Potentiometric biosensors based on butyrylcholinesterase are developed by co-reticulation of the enzyme with glutaraldehyde on an electropolymerized polyethyleneimine film at the electrode surface. The BuChE-electrode was tested as biochemical sensor for detection of an organophosphorus pesticide, trichlorfon in liquid, the detection being based on the enzyme inhibition. The enzyme electrode showed a detection limit for trichlorfon below 10⁻⁷ M.     

电场耦合法芯片电泳柱上电导检测特性研究

通过电场耦合作用对毛细管电泳柱上直流电导检测原理进行了分析, 利用带有双T结构检测池的玻璃和聚合物芯片对其特性进行了研究. 在电泳分离过程中, 电泳高电场通过检测通道耦合到出口的检测电极上并产生可被检测的电势差. 当导电性与支持电解质不同的组分谱带通过分离通道上的检测池时造成该电位差的变化, 该变化与溶液电导率、检测池长度和电场强度直接相关. 检测信号与基线相除得到的信号只和检测池中溶液的物理特征参数(电导率及电阻)有关. 用自制高阻抗信号转换系统可使芯片电泳电场强度提高到450 V/cm以上, 以1 mmol/L Tris-HCl为支持电解质, 在12 s内实现了K＋和Na＋的高重现性分离检测, 检出限达到5 μmol/L, 线性范围达两个数量级(10 μmol/L~2 mmol/L).