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World Congress on Biosensors 2014

World Congress on Biosensors 2014
Biosensors 2014

Friday, 29 June 2012


A new issue of this journal has just been published. To see abstracts of the papers it contains (with links through to the full papers) click here:
Talanta

Quantitative assessment of the contribution of high resolution mass spectrometric analysis to the reliability of compound confirmation

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Georgi Stoev, Yue Xuan, Milena Peycheva, Michaela Scigelova
Applications of high resolution mass spectrometry (HRMS) in food safety and residue analysis have increased remarkably over the last few years. The high resolution detection of ions reportedly enhances the assay selectivity but quantitative assessment of HRMS contribution to the assay selectivity has not yet been undertaken. We devised a method to assess the impact of instrument resolution on the probability that a spectral assignment to a given compound was made in error. The method allows for evaluating the quality of a spectral assignment based on resolution and the number of fragmentation stages. It thus provides a firm basis for comparing analytical methods performed on very different mass spectrometric instrumental platforms as well as in the context of the current regulatory framework.

Graphical abstract

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Graphical abstract Highlights

► High mass resolution impacts positively on the confidence of confirmation. ► Method quantifies the contribution of the resolution to assay selectivity. ► Method allows evaluation of regulatory framework identification criteria

Multiclass analysis of antibacterial residues in milk using RP-liquid chromatography with photodiode array and fluorescence detection and tandem mass spectrometer confirmation

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Isabela Maia Toaldo, Gabriel Zandonadi Gamba, Lidia Almeida Picinin, Gabriel Rubensam, Rodrigo Hoff, Marilde Bordignon-Luiz
A simplified procedure for simultaneous quantification of ceftiofur (CEF), fluoroquinolone (FQ) and sulfonamide (SA) antibacterials in bovine milk was developed. The reverse-phase liquid chromatography (RP-LC) multiclass method for analysis of eleven distinct compounds, from three antibacterial classes, was validated in line with Commission Decision 2002/657/EC. Confirmation of the analytes identities was performed by electrospray mass spectrometry detection. The analytes were extracted from milk matrix by liquid-liquid extraction with acidified ultrapure water and directly analyzed in the chromatograph. The SA compounds were pre-column derivatized with fluorescamine for fluorescence detection. The method provided good results regarding the analytical parameters of linearity, selectivity, sensitivity, precision, recovery, decision limit (CCα), detection capability (CCβ), limit of detection (LOD), limit of quantification (LOQ), stability and robustness. Analytes were extracted by liquid-liquid extraction in the fortified matrix and the compounds identity was confirmed by their precursor ion and fragments through tandem mass spectrometry analysis. Additionally, milk samples from two state capitals in the South Region of Brazil were analyzed by both the quantitative and confirmatory methods. The validation process showed correlation coefficients (r 2 ) greater than 0.98 for all the analytes, with recovery rates up to 98% for all the studied drugs. LOD and LOQ limits ranged from 8.0 to 20.0ngmL−1 and 10.0 to 32.0ngmL−1, demonstrating good specificity of the method. The intra-day and inter-day precisions for all the analytes were below or equal to 7.40 and 10.13, respectively. The studied antibacterials were not detected in milk samples. The developed method represents an efficient alternative for multi-residue analysis in milk, being suitable and especially viable for monitoring in developing countries.

Highlights

► Sulfonamides, fluoroquinolones and ceftiofur were determined in milk. ► Acidified water was efficient for extraction without further clean up procedure. ► Validation results showed great performance for the LC method. ► Analyzed samples from Brazil showed no detection.► A feasible alternative to monitor residues in milk was developed.

An insight into the adsorption and electrochemical processes occurring during the analysis of copper and lead in wines, using an electrochemical quartz crystal nanobalance.

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Alzira Yamasaki, João A.B.P. Oliveira, Armando C. Duarte, M.Teresa S.R. Gomes
Copper and lead in wine were quantified by anodic stripping voltammetry (ASV), performed onto the gold electrode of a piezoelectric quartz crystal. Both current or mass changes could be used as analytical signals, without a statistical difference in the results (α=0.05). However, the plot of mass vs. potential provided an in depth understanding of the electrochemical processes and allowed studying adsorption phenomena. Copper interaction with fructose is an example of a process which was not possible to ignore by observing the mass change on the gold electrode of the piezoelectric quartz crystal.

Determination of organophosphorus pesticides using dispersive liquid-liquid microextraction combined with reversed electrode polarity stacking mode - micellar electrokinetic chromatography

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Phimpha Soisungnoen, Rodjana Burakham, Supalax Srijaranai
A rapid and sensitive method using two preconcentration techniques, dispersive liquid-liquid microextraction (DLLME) followed by reversed electrode polarity stacking mode (REPSM) was developed for the analysis of five organophosphorus pesticides (OPPs) by micellar electrokinetic chromatography (MEKC). Parameters that affect the efficiency of the extraction in DLLME and preconcentration by REPSM, such as the kind and volume of the extraction and disperser solvents, salt addition, sample matrix and injection time were investigated and optimized. Under the optimum conditions, the enrichment factors were obtained in the range from 477 to 635. The linearity of the method for parathion, azinphos and fenitrithion was in the range of 20–1000ngmL−1, and for malathion and diazinon in the range of 50–1000ngmL−1, with correlation coefficients (r 2) ranging from 0.9931 to 0.9992. The limits of detecton (LODs) at a signal-to-noice ratio of 3 ranged from 3 to 15ngmL−1. The relative recoveries of five OPPs from water samples at spiking levels of 20 and 200ngmL−1 for parathion, azinphos and fenitrithion, and 50 and 500ngmL−1 for malathion and diazinon, were 69.5-103%. The proposed method provided high enrichment factors, good precision and accuracy with a short analysis time.

Analysis of renal stones by capillary isotachophoresis

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Zdeňka Jarolímová, Přemysl Lubal, Viktor Kanický
An analytical method for the determination of the composition of renal stones by capillary isotachophoresis with conductometric detection was developed. Using different leading/terminating electrolyte systems, the qualitative and quantitative analysis of organic compounds (urate, xanthate, oxalate) and inorganic ions (phosphate, Ca2+, Mg2+, Na+,, NH4 +) species commonly present in mixed renal stones in three separate steps can be carried out with limits of detection about 10μmol/L. The developed method was validated by the analysis of real samples and can be used for urinary calculi classification. In addition, it was verified that this method can also be employed for the determination of the above mentioned analytes in some other samples (bones, teeth) concerning apatite biominerals (fluoro-, carbonate-, chloro-apatite).

Highlights

► Cationic and anionic analysis by capillary isotachophoresis with conductivity detection. ► The robust and fast method was developed for analysis of biominerals. ► Application for qualitative/quantitative analysis of uric stones, bones and teeth.

Analysis of free fatty acids in Notopterygium forbesii Boiss by a novel HPLC method with fluorescence detection

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Shijuan Zhang, Jinmao You, Guoying Zhou, Chunli Li, Yourui Suo
A new labeling reagent for fatty acids, 1-(9H-carbazol-9-yl) propan-2-yl-methanesulfonate (CPMS), has been synthesized and successfully applied to the HPLC determination of fatty acids in traditional Chinese herb Notopterygium forbesii Boiss. The reaction of CPMS with fatty acids could proceed easily and quickly in the presence of K2CO3 catalyst within 30min. The derivatives exhibit excellent fluorescence property with excitation and emission wavelengths of 293nm and 360nm, respectively. The 34 derivatives of fatty acids were separated on a BDS C8 reversed-phase column with gradient elution. Good linear correlations were observed for all fatty acids with correlation coefficients of >0.996. The detection limits at a signal-to-noise ratio of 3 were in the range of 0.032–0.312μgg–1. Free fatty acids in the roots, stem, leaves and petioles of Notopterygium forbesii Boiss from different places was analyzed by the developed method. This is the first time that the fatty acids composition of Notopterygium forbesii Boiss has been reported. This method also shows powerful potential for the trace analysis of fatty acids or other carboxylic acids from complex samples.

Highlights

► A new labeling reagent for fatty acids has been synthesized. ► Fatty acids composition of Notopterygium forbesii Boiss was reported for the first time. ► Sensitivity was much higher than the often used GC methods.

Head- Space Voltammetry: A Novel Voltammetric Method For Volatile Organics and a Case Study For Phenol

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
K. Volkan Özdokur, Levent Pelit, Hasan Ertaş, Suna Timur, F. Nil Ertaş
Present paper describes the results of a novel method which combines the Headspace (HS) preconcentration of the analyte on the electrode prior to the voltammetric analysis. Thereafter, the method was called HS-Voltammetry. The performance of the method was tested upon using an electroactive and volatile molecule phenol molecule which gives an oxidation peak at conventional electrodes. In this study, a glassy carbon electrode was modified with polypyrrole by electropolymerization and then, the electrode was placed over the solution in a sealed vial heated gently on a hotplate with a stirrer for phenol determination. By controlling the thickness of polymeric coating and optimizing preconcentration parameters such as vial pH and temperature, stirring rate and exposure time, a very consistent (5.2% at 5.0×10−7 M) fraction of the analyte can be extracted during a predetermined time. The oxidation peak current at 0.8V depended linearly on the phenol concentration over a wide range (3 orders of magnitude). The detection limit was estimated as 7.0×10−8 M at 60°C (S/N=3) which is well below the limit set by the European Community for phenols in wastewaters (ca. 5×10−6 M). The effect of other phenolic compounds was also examined and it was shown that head space preconcentration eliminated the interference of nonvolatile phenolic acids studied. For volatile phenolic compounds, the selectivity can be maintained in cases when isolated peaks are obtained for each component. The proposed method has been applied successfully for the determination of phenol in artificial wastewater and recovery percentage was calculated as 93%.

Highlight

► Voltammetry was combined with the advantageous of headspace sampling. ► Detection limit for phenol was improved with HS accumulation. ► The interference of non-volatile components was eliminated by HS sampling. ► Phenol and 2,4-dichlorophenol can be determined simultaneously by this means

Urine Iodide Determination by Ion-Pair Reversed-Phase High Performance Liquid Chromatography and Pulsed Amperometric Detection

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Vo Thanh Phuong Nguyen, Virginie Piersoel, Tarik El Mahi
A sensitive and specific ion-pair reversed-phase high performance liquid chromatography (HPLC) method for urinary iodine analysis is described. This method is based on pulsed amperometric detection (PAD) using a silver working electrode (HPLC-PAD), which improves peak shape, electrode stability as well as linearity and reproducibility. A two-step extraction process consisting of solid phase extraction (SPE) and liquid-liquid extraction with dichloromethane was added in order to improve sample purification which is essential with the use of PAD. Treated samples were eluted on a C18 column, using a phosphate buffer containing ion-pairing reagent tetrabutylammonium and 5% MeOH. The calibration standard curves were linear up to 500µg/L and within-run and between–run coefficients of variation (CVs) were <6% with the quantification limit fixed at 6µg/L. Accuracy, expressed as recovery, ranged from 94 to 104%. Comparison with the Technicon AutoAnalyzer acid digestion (AA) method resulted in a high correlation (r=0.9916). Due to a low quantification limit and high sample throughput, the proposed technique appears suitable for both epidemiological and clinical follow-up studies.

Highlights

► We developed a HPLC-PAD method for determining iodide concentration in urine. ► Pulsed amperometric detection improves peak shape and electrode stability. ► Two extraction steps for sample purification are necessary. ► This method is sensitive, selective, precise and accurate. ► The method is effective for urinary iodine determination.

Simple and sensitive aptasensor based on quantum dot-coated silica nanospheres and the gold screen-printed electrode

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Yi Li, Liu Deng, Chunyan Deng, Zhou Nie, Minghui Yang, Shihui Si
A novel electrochemical aptasensor involving quantum dots-coated silica nanospheres (QDs/Si) and the screen-printed gold electrodes (SPGE) was developed for the detection of thrombin. The screen-printed electrode with several advantages including low cost, versatility, miniaturization, and mechanical regeneration after each measurement cycle were employed. On the other hand, the gold nanoparticles (AuNPs) were electrodeposited on the surface of SPGE to obtain the AuNPs/SPGE. And this sandwich format (Apt/thrombin/Apt−QDs/Si) was fixed on the AuNPs/SPGE to fabricate the electrochemical aptasensor. The bound CdTe QDs were dissolved in an acid-dissolution step and were detected by electrochemical stripping analysis. The proposed aptasensor has excellent performance such as high sensitivity, good selectivity and analytical application in real samples. The combination of nanoparticles with the screen-printed electrode is favorable for amplifying electrochemical signals, and useful for large-scale fabrication of the electrochemical aptasensors, which would lay a potential foundation for the development of the electrochemical aptasensor.

Highlights

► We fabricated the electrochemical aptasensor with simplicity and high sensitivity. ► Gold nanoparticles were electrodeposited onto the screen-printed electrode. ► The bound QDs were dissolved in an acid-dissolution step and were detected. ► Nanoparticles may be favorable for amplifying electrochemical signals. ► The screen-printed electrode was useful for the large fabrication of the aptasensor.

Preparation of a novel dual-function strong cation exchange/hydrophobic interaction chromatography stationary phase for protein separation

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Kailou Zhao, Li Yang, Xuejiao Wang, Quan Bai, Fan Yang, Fei Wang
We have explored a novel dual-function stationary phase which combines both strong cation exchange (SCX) and hydrophobic interaction chromatography (HIC) characteristics. The novel dual-function stationary phase is based on porous and spherical silica gel functionalized with a ligand containing sulfonic and benzyl groups capable of electrostatic and hydrophobic interaction functionalities, which displays HIC character in a high salt concentration, and IEC character in a low salt concentration in mobile phase employed. As a result, it can be employed to separate proteins with SCX and HIC modes, respectively. The resolution and selectivity of the dual-function stationary phase were evaluated under both HIC and SCX modes with standard proteins and can be comparable to that of conventional IEC and HIC columns. More than 96% of mass and bioactivity recoveries of proteins can be achieved in both HIC and SCX modes, respectively. The results indicated that the novel dual-function column could replace two individual SCX and HIC columns for protein separation. Mixed retention mechanism of proteins on this dual-function column based on stoichiometric displacement theory (SDT) in LC was investigated to find the optimal balance of the magnitude of electrostatic and hydrophobic interactions between protein and the ligand on the silica surface in order to obtain high resolution and selectivity for protein separation. In addition, the effects of the hydrophobicity of the ligand of the dual-function packings and pH of the mobile phase used on protein separation were also investigated in detail. The results show that the ligand with suitable hydrophobicity to match the electrostatic interaction is very important to prepare the dual-function stationary phase, and a better resolution and selectivity can be obtained at pH 6.5 in SCX mode. Therefore, the dual-function column can replace two individual SCX and HIC columns for protein separation and be used to set up two-dimensional liquid chromatography with a single column (2DLC-1C), which can also be employed to separate three kinds of active proteins completely, such as lysozyme, ovotransferrin and ovalbumin from egg white. The result is very important not only to the development of new 2DLC technology with a single column for proteomics, but also to recombinant protein drug production for saving column expense and simplifying the process in biotechnology.

Highlights

► We explored a bifunctional column with a ligand containing sulfo and benzyl groups. ► This column can provide two operation modes (HIC and SCX). ► High resolution and selectivity are obtained in both SCX and HIC modes, respectively. ► This column can replace two corresponding single mode (SCX and HIC mode) columns. ► Based on this bifunctional column, 2DLC was established using only a single column.

Simultaneous measurement of protein-bound 3-chlorotyrosine and homocitrulline by LC-MS/MS after hydrolysis assisted by microwave: application to the study of myeloperoxidase activity during hemodialysis

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Cédric Delporte, Thierry Franck, Caroline Noyon, Damien Dufour, Alexandre Rousseau, Philippe Madhoun, Jean-Marc Desmet, Didier Serteyn, Martine Raes, Joëlle Nortier, Michel Vanhaeverbeek, Nicole Moguilevsky, Jean Nève, Luc Vanhamme, Pierre Van Antwerpen, Karim Zouaoui Boudjeltia
A high degree of uremia is common in patients with end-stage renal disease and has been linked to the development of chronic inflammation and cardiovascular diseases. In conditions where transplantation is not possible, uremia can be reduced by hemodialysis although the repeated interventions have been implicated in loss of renal function, partially as a result of chronic inflammation and/or oxidative stress processes. In this context, it has been suggested that myeloperoxidase (MPO) can contribute to the oxidative stress during hemodialysis and to the cardiovascular risk. Protein damages due to MPO activity have never been assessed during hemodialysis although two of its reaction products, 3-chlorotyrosine and homocitrulline, are of interest. Indeed, the first one is a specific product of MPO activity and the formation of the second one could be catalyzed by MPO. In order to analyze these products in plasma proteins, a total hydrolysis method followed by liquid chromatography mass spectrometry analysis was developed. Different conditions of hydrolysis were tested and the optimized procedure was assessed for complete hydrolysis and artifactual chlorination. Finally, the method was used for analyzing 3-chlorotyrosine and homocitrulline in plasma proteins during a hemodialysis session in fifteen patients and data were related to measurements of MPO concentration and activity. Both increases in MPO activity and protein-bound 3-chlorotyrosine were observed, highlighting the involvement of MPO in oxidative stress during hemodialysis and further demonstrating the link between hemodialysis and cardiovascular diseases.

Highlights

► Simultaneous detection of protein-bound 3-chlorotyrosine and homocitrulline. ► Rapid protein acid hydrolysis assisted by microwave oven. ► Method applied to a clinical situation: patients undergoing hemodialysis. ► Myeloperoxidase activity increases during hemodialysis. ► 3-chlorotyrosine but not homocitrulline increases during hemodialysis

A Reaction Based Turn-On Type Fluorogenic and Chromogenic Probe for the Detection of Trace Amount of Nitrite in Water

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Vikash Kumar, Amrita Chatterjee, Mainak Banerjee
A turn-on fluorescent probe for the detection of nitrite ion in water is developed based on diazotization reaction of the amino group of the probe in an acidic solution (pH 1). The probe responds selectively to nitrite ion over various other anions with a turn-on type fluorogenic change from colorless to orange by the formation of rhodamine B via an analyte triggered fragmentation process. The fluorescence titration is complete within 1h with 1 equiv of nitrite ion. The probe is highly efficient, cost-effective and shows a detection limit of 4.6ppb.

Highlights

► A rhodamine-based fluorescent probe is developed for the detection of nitrite ions. ► The probe is highly selective to nitrite ions in presence of many other anions. ► The fluorogenic detection level of nitrite is found to be as low as 4.6ppb. ► The probe was successfully used for detection of NO2¯ level in several real samples.

Detection of DNA damage by using hairpin molecular beacon probes and graphene oxide

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Jie Zhou, Qian Lu, Ying Tong, Wei Wei, Songqin Liu
A hairpin molecular beacon tagged with carboxyfluorescein in combination with graphene oxide as a quencher reagent was used to detect the DNA damage by chemical reagents. The fluorescence of molecular beacon was quenched sharply by graphene oxide; while in the presence of its complementary DNA the quenching efficiency decreased because their hybridization prevented the strong adsorbability of molecular beacon on graphene oxide. If the complementary DNA was damaged by a chemical reagent and could not form intact duplex structure with molecular beacon, more molecular beacon would adsorb on graphene oxide increasing the quenching efficiency. Thus, damaged DNA could be detected based on different quenching efficiencies afforded by damaged and intact complementary DNA. The damage effects of chlorpyrifos-methyl and three metabolites of styrene such as mandelieaeids, phenylglyoxylieaeids and epoxystyrene on DNA were studied as models. The method for detection of DNA damage was reliable, rapid and simple compared to the biological methods.

Highlights

► DNA damage induced by several chemicals was detected by a hairpin MB based on FRET. ► The method was rapid, simple, reliable and sensitive. ► The method can be used to detect damaged DNA induced by other reagents or factors.

Speciation analysis of mercury in sediments using vortex-assisted liquid-liquid microextraction coupled to high-performance liquid chromatography-cold vapor atomic fluorescence spectrometry

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Geng Leng, Hui Yin, Shaobo Li, Yong Chen, Dezhong Dan
A simple and fast solvent microextraction method termed vortex-assisted liquid-liquid microextraction (VALLME) coupled with high-performance liquid chromatography-vapor generation atomic fluorescence spectrometry (HPLC-CVAFS) has been developed for the trace analysis of methylmercury (MeHg+), ethylmercury (EtHg+) and inorganic mercury (Hg2+) in sediment samples. Carbon tetrachloride was used as collecting solvent for the extraction of mercury species from sediment by a vortex-assisted extraction. In VALLME, 100μL 1% (m/v) L-Cysteine were used as extraction solvent and were injected into 4mL carbon tetrachloride. The extraction solvent dispersed into carbon tetrachloride under vigorously shaking by a vortex agitator. The fine droplets could extract mercury species within few minutes because of the shorter diffusion distance and larger specific surface area. After centrifugation, the floating extractant phase restored its initial single microdrop shape and was used for HPLC-CVAFS analysis. The parameters affecting the extraction efficiency of the proposed VALLME such as extraction solvent, vortex time, volumes of extraction solvent and salt addition etc. were investigated. Under the optimum conditions, linearity was found in the concentration range from 0.1 to 25ng g−1 for MeHg+, 0.2 to 65ng g−1 for EtHg+, and 0.1 to 30ng g−1 for Hg2+. Coefficients of determination (R2) ranged from 0.9938 to 0.9972. The limits of detection (LODs, signal-to-noise ratio (S/N)=3) were 0.028ng g−1 for MeHg+, 0.057ng g−1 for EtHg+, and 0.029ng g−1 for Hg2+. Reproducibility and recoveries were assessed by testing a series of 6 sediment samples, which were spiked with different concentration levels. Finally, the proposed method was successfully applied in analyses of real nature sediment samples. In this work, VALLME was applied to the extraction of mercury species in sediment samples for the first time. Using L-Cys as extraction solvent, the extraction process is sensitive and environmentally friendly and could be achieved within 3min.

Highlights

► VALLME was for the first time applied for the extraction of mercury species in sediment.► Using L-Cys as extraction solvent, the extraction is sensitive and environmentally friendly.► The extraction process is achieved within 3min.

Magnetic graphene nanosheets based electrochemiluminescence immunoassay of cancer biomarker using CdTe quantum dots coated silica nanospheres as labels

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Fang Liu, Yan Zhang, Shenguang Ge, Juanjuan Lu, Jinghua Yu, Xianrang Song, Su Liu
A highly sensitive electrochemiluminescence (ECL) immunosensor for the detection of prostate specific antigen (PSA) was designed using biofunctionalized magnetic graphene nanosheets (G@Fe3O4) as immunosensing probes and CdTe quantum dots coated silica nanospheres (Si/QDs) as signal amplification labels. In this work, a sandwich-type immunosensor was fabricated, which was assembled on the surface of indium tin oxide glass (ITO). The analyte was detected in a home-made flow injection ECL (FI-ECL) cell through the immunosensor. Owning to the signal amplification of G@Fe3O4 composite and Si/QDs, the ECL measurement showed a great increase in detection signals compared with the unamplified method. Under optimal conditions, a wide detection range (0.003–50ngmL−1) and low detection limit (0.72pgmL−1) were obtained through the sandwich-type immunosensor. The proposed strategy successfully demonstrated a reproducible, specific, and potent method that can be expanded to detect other proteins.

Highlights

► A sandwich-type electrochemluminence immunosensor was fabricated. ► Magnetic graphene nanosheets were synthesized as immunosensing probes. ► CdTe quantum dots coated silica nanospheres were used to amplify signals. ► A home-made injection electrochemluminence cell was used as reaction chamber.

Detection of Trace Levels of Lead in Aqueous Liquids Using Extractive Electrospray Ionization Tandem Mass Spectrometry

28 June 2012, 21:37:00
Publication year: 2012
Source:Talanta
Chunxiao Liu, Xinglei Zhang, Saijin Xiao, Bin Jia, Shasha Cui, Jianbo Shi, Ning Xu, Xi Xie, Haiwei Gu, Huanwen Chen
A sensitive approach, based on semi-quantitative measurement of the characteristic fragments in multi-stage extractive electrospray ionization mass spectrometry (EESI-MSn), was developed for fast detection of trace levels of lead in aqueous liquids including mineral water, lake water, tap water, energy drinks, soft drinks, beer, orange juice, and tea. A disodium ethylene-diamine-tetraacetic acid (EDTA) aqueous solution was electrosprayed to produce negatively charged primary ions which then intersected the neutral sample plume to generate anions of EDTA-Pb(II) complexes. The charged EDTA-Pb(II) complexes were characterized with multistage collision induced dissociation (CID) experiments. The limit of detection (LOD) using EESI-MS3 was estimated to be at the level of 10–13 g/mL for directly detecting lead in many of these samples. The linear dynamic range was higher than 2 orders of magnitude. A single sample analysis could be completed within 2min with reasonable semi-quantitative performance, e.g., relative standard deviations (RSDs) for deionized water were 4.6%-7.6% during 5 experimental runs (each of them had 10 repeated measurements). Coca-cola and Huiyuan orange juice, representative beverage samples with complex matrices, generated recovery rates of 91.5% and 129%, respectively. Our experimental data demonstrated that EESI-MS is a useful tool for the fast detection of lead in various solutions, and EESI-MS showed promises for fast screening of lead-contaminated aqueous liquid samples.

Highlights

► EESI can directly detect lead in complex samples. ► EESI can semi-quantitatively measure the lead concentration. ► EESI may have wide applications for food security and environmental science

SP Scientific and Praxair Expand Technology Agreement

SP Scientific, and Praxair Inc., headquartered in Danbury, CT, are pleased to announce the expansion of their collaborative relationship to commercialize Praxair's ControLyo™ Nucleation on Demand Technology.

The original agreement, signed in October 2010, gave SP Scientific the exclusive, global rights to commercialize the technology on Development Lyophilizers <1.0 m. The Praxair ControLyo™ Technology was implemented on SP Scientific's Lyostar 3 Freeze Dryer in December 2010. The expansion of the agreement allows SP Scientific to also equip its clinical, pilot and production dryers (Benchmark series and Hull Production Dryers) with the ControLyo™ Technology. Additionally, the agreement calls for the transfer of technology to allow SP Scientific to retrofit existing Pilot and Production units in the field, regardless of the original manufacturer.

"This is a major step in SP's strategy of technology leadership in lyophilization," states Chuck Grant, CEO for SP Scientific. "Given the ability to offer the technology from development through scale-up to production, SP Scientific is ideally positioned to be the supplier of choice for ControLyo™ Technology."

"Praxair is pleased to extend our existing agreement to cover the full range of lyophilizers offered to the industry by SP Scientific, " said Rich Jarrett, Global Director of Marketing and Business Development for Praxair. "Praxair's ControLyo™ Technology has seen quick industry adoption with SP Scientific's Lyostar 3 FreezeDryer and we look forward to providing the industry with nucleation control at all production levels."

ControLyo™ Technology offers many benefits to the pharmaceutical industry including: vial-to-vial uniformity, reduced cycle times, reduced protein aggregation and better conformance to the Food and Drug Administration's Process Analytical Technology (PAT) initiative.


Fluidigm Announces Unrestricted Sales into Prenatal Health and Non-Invasive Prenatal Diagnostics Fields of Research

Fluidigm Corporation has announced that it can now offer unrestricted sales of its digital PCR and other advanced technology to customers interested in pursuing research and product development into the prenatal health and non-invasive prenatal diagnostics fields, as well as other fields. This enhanced freedom to operate comes as a result of the ending of the collaboration agreement previously reported in the company's filings with the Securities and Exchange Commission between Fluidigm and Novartis Vaccines & Diagnostics, Inc.

Under the collaboration agreement, Fluidigm granted an exclusive option to its technology in specific areas of prenatal health and diagnostics, and Fluidigm could not sell its products or services in these fields, other than in some cases for research applications. This option has expired unexercised. Also, while the details of the collaboration remain confidential, Fluidigm can confirm that it successfully achieved all of its technical feasibility milestones in the first phase of the collaboration and, accordingly, received all milestone payments. The re-opening of previously restricted fields of use for Fluidigm products and access to significant product enhancements represent exciting opportunities for Fluidigm's customer base.

A number of Fluidigm's customers had expressed concerns over their ability to freely operate in the fields of prenatal health and non-invasive prenatal diagnostics in view of the collaboration agreement. Some potential customers chose to pursue alternative options because of this uncertainty. "With the termination of this agreement, highly valuable intellectual property rights in non-invasive prenatal diagnostics and digital PCR, which had been exclusively optioned under the agreement, now revert back to Fluidigm. We are open for business without restriction in all fields," said Gajus Worthington, Fluidigm President and Chief Executive Officer.

Fluidigm can now fully pursue all market opportunities with customers interested in researching and developing products in all fields, including prenatal health and non-invasive prenatal diagnostics. "We appreciate that there are significant market opportunities in these areas and there is substantial customer interest. We look forward to helping these customers achieve their research and development objectives," Worthington added.

With the termination of the collaboration, Fluidigm's customers may experience other, more direct benefits. Chief among them is unrestricted access to the company's prototype 200,000-chambered digital PCR chip with associated instrumentation and software developed under several on-going collaborative efforts. The company can now commercialize this much higher density chip, for which it has experimentally demonstrated both very high sensitivity and reproducibility.

Thursday, 28 June 2012

Article of the Future


Elsevier has announced the SciVerse ScienceDirect redesigned article page, with a new layout including a navigational pane and an optimized reading middle pane (see the video below).
The Article of the Future project- an ongoing initiative aiming to revolutionize the traditional format of the academic paper in regard to three key elements: presentation, content and context.
Given that only ten years ago paper copies of journals were the main way for publishers to deliver content, how do you think scientific content will be delivered (format, type of content, delivery system, etc) ten years from now? 
A $25 Amazon voucher is on offer for the best or most imaginative forecast. Post your replies below. (Deadline July 31st, 2012).
video

Caltech hosting joint ‘Etch Tech 2012’ Seminar and Workshop with Oxford Instruments in July 2012


The Kavli Nanoscience Institute (KNI) at Caltech, USA and Oxford Instruments are excited to announce their joint Seminar and Workshop entitled Etch Tech 2012, taking place 16th-17th July. The event is open to all those people working in industry and academia with an interest in recent progress in research and development, plus future trends in the fabrication and application of micro & nano structures and devices.

Topics include Advances in Deep RIE / MEMS; OpSIS, Single Digit Nanoscale Fabrication; Optical and Electrical Probes; Silicon Nanowires and Novel Etch Techniques; Diamond Etching and Nanoscale Applications of ALD.

Following an introduction by Oxford Instruments Managing Director Dan Ayres, there will be talks from invited guest speakers, senior KNI Caltech and Oxford Instruments Plasma Technology specialists.

Speakers from Caltech, include Professor Oskar Painter, Executive Officer for Applied Physics and Materials Science and Co-Director, Kavli Nanoscience Institute, Caltech;  Andrei Faraon Assistant Professor of Applied Physics and Materials Science; Rassul Karabalin, Senior Research Scientist, Condensed Matter Physics at Caltech;  Sameer Walavalkar, Postoctoral Scholar in Physics, Caltech. Guest speakers include Dr Deirdre Olynick, Nanofabrication Facility, Lawrence Berkeley National Lab and Professor Michael Hochberg, Electrical and Chemical Engineering, University of Delaware.

As a leading manufacturer of plasma processing systems, Oxford Instruments has Process Applications and Technology teams who are highly experienced and qualified, with many years experience. They will offer talks on various aspects of plasma etch and deposition processing, and on day two will be running tutorial sessions, allowing plenty of time for delegates to ask questions. Following this there will be tours of the Kavli Nanoscience Institute’s cleanroom facility.

Comments Prof Oskar Painter, Co-Director, Kavli Nanoscience Institute, Caltech, “Both Oxford Instruments Plasma Technology and the KNI aim to push the state-of-the-art beyond current capabilities in nanofabrication. The KNI has pursued aggressive acquisition of strategic instrumentation for advanced nanofabrication capabilities, including a number of Oxford Instruments systems that are successfully installed and producing great results for our researchers. Our multi-user laboratories and cleanrooms for nanostructure synthesis, fabrication, and characterization are available to users from academia, government and industry.”

There is no charge for this event but booking is essential: http://www.kni.caltech.edu/calendar/workshops/etch-tech-2012/index.html

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Wednesday, 27 June 2012

Just Published: Journal of Chromatography B


A new issue of this journal has just been published. To see abstracts of the papers it contains (with links through to the full papers) click here:
Selected papers from the latest issue:

Comparison of GC/MS and LC/MS methods for the analysis of propofol and its metabolites in urine

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Sun Young Lee, Na-Hyun Park, Eun-Kyung Jeong, Jae-Woo Wi, Chang-Ju Kim, Jin Young Kim, Moon Kyo In, Jongki Hong
Gas chromatography–mass spectrometry (GC/MS) and liquid chromatography–mass spectrometry (LC/MS) were compared for their capacity to metabolite identification, sensitivity, and speed of analysis for propofol and its metabolites in urine samples. Acidic hydrolysis, liquid–liquid extraction (LLE), and trimethylsilyl (TMS) derivatization procedures were applied for GC/MS analysis. The LC/MS analysis used a simple sample pretreatment based on centrifugation and dilution. Propofol and four metabolites were successfully analyzed by GC/MS following TMS derivatization. One compound, di-isopropanolphenol was tentatively characterized as a new metabolite observed for the first time in human urine. The TMS derivatization greatly improved the chromatographic properties and detection sensitivity, especially for hydroxylated metabolites. The lower limits of quantitation (LLOQ) of propofol were about 325 and 0.51ng/mL for the GC/MS scan mode and selected ion monitoring (SIM) mode, respectively. In addition, five conjugated propofol metabolites were successfully analyzed by LC–MS/MS in negative ion mode. The detection sensitivity for these conjugated metabolites could be greatly enhanced by the addition of triethylamine to the mobile phase without any loss of LC resolution capacity. The LLOQs of propofol-glucuronide (PG) were about 1.17 and 2.01ng/mL for the LC–MS-selected ion monitoring (SIM) and multiple reaction monitoring (MRM) mode, respectively. Both GC/MS and LC/MS methods sensitively detected nine metabolites of propofol and could be used to provide complementary data for the reasonable propofol metabolism study. Urinary excretion profiles for propofol and its metabolites following administration to human were suggested based on the total ion chromatograms obtained by GC/MS and LC/MS methods, respectively.

Highlights

► Comprehensive GC/MS and LC/MS methods were proposed for analysis of propofol and metabolites. ► Both methods provided excellent sensitivity and selectivity in urine samples. ► 2,6-ω-Di-isopropanol phenol as new metabolite was first observed in human. ► LLOQ was 0.51ng/mL for propofol and 2.01ng/mL for propofol-glucuronide.

Quantitative analysis of the opioid peptide DAMGO in rat plasma and microdialysis samples using liquid chromatography–tandem mass spectrometry

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Annika Lindqvist, Britt Jansson, Margareta Hammarlund-Udenaes
A liquid chromatography–electrospray ionization-tandem mass spectrometry (LC–ESI-MS/MS) method for the quantification of the opioid peptide DAMGO in rat plasma, as well as DAMGO and the microdialysis recovery calibrator [13C2,15N]-DAMGO in microdialysis samples, is described. The microdialysis samples consisted of 15μL Ringer solution containing 0.5% bovine serum albumin. Pretreatment of the samples involved protein precipitation with acetonitrile followed by dilution with 0.01% formic acid. The lower limits of quantification were 0.52ng/mL and 0.24ng/mL for DAMGO and [13C2,15N]-DAMGO respectively and the response was linear up to 5000 fold higher concentrations. The plasma samples (50μL) were precipitated with acetonitrile containing the isotope labeled analog [13C2,15N]-DAMGO as internal standard. The method was linear in the range of 11–110,000ng/mL. The separations were conducted on a HyPurity C18 column, 50×4.6mm, 3μm particle size, with a mobile phase consisting of acetonitrile, water and formic acid to the proportions of 17.5:82.5:0.01. Low energy collision dissociation tandem mass spectrometric (CID-MS/MS) analysis was carried out in the positive ion mode using multiple reaction monitoring (MRM) of the following mass transitions: m/z 514.2→453.2 for DAMGO and m/z 517.2→456.2 for [13C2,15N]-DAMGO. The intra-day precision and accuracy did not exceed 5.2% and 93–104% for both compounds and sample types described. The inter-day precision an accuracy were <6.8% and 95–105% respectively. The method described is simple, reproducible and suitable for the analysis of small sample volumes at low concentrations.

Highlights

► Analysis of DAMGO in microdialysis samples and plasma using LC–MS/MS is described. ► Plasma method was linear between 11 and 110,000ng/mL. Sample volume was 50μL. ► Microdialysis method was linear between 0.52 and 2600ng/mL. Sample volume was 15μL. ► The intra-day and inter-day precision and accuracy did not exceed 6.8% and 93–105%. ► The method was applied to determine the brain distribution of DAMGO in rat.

Screening, recognition and quantitation of salbutamol residues in ham sausages by molecularly imprinted solid phase extraction coupled with high-performance liquid chromatography–ultraviolet detection

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Hongyuan Yan, Ruiling Wang, Yehong Han, Suting Liu
A highly selective molecularly imprinted solid phase extraction (MISPE) coupled with liquid chromatography–ultraviolet detection was developed for the determination of salbutamol (SAL) in ham sausages. New molecularly imprinted polymers (MIPs) were synthesized with phenylephrine as dummy template and it revealed good affinity to SAL in methanol–acetonitrile system. Adsorption capacity of the MIPs was evaluated by dynamic adsorption experiments. The MIPs were used as SPE sorbent for the selective clean-up and pre-concentration of SAL in ham sausage samples. The results showed that the matrix compounds presented in ham sausage samples could be removed effectively and the recoveries of SAL at three spiked levels were ranged from 82.6 to 100.5% with the relative standard deviation (RSD) of less than 3.6%. This method is simple and sensitive, and is therefore an alternative tool to the existing methods for analyzing residual SAL in biological samples.

Highlights

► A simple and rapid MISPE–HPLC method for determination of salbutamol in ham sausages. ► The new MIPs were obtained by phenylephrine as dummy template in methanol–acetonitrile solution. ► Rapid screening of salbutamol in ham sausages, while interferences from milk matrix were eliminated. ► Extraction efficiency was markedly increased with reduced equilibrium time. ► This method improved the selectivity and eliminated the effect of template leakage on quantitative analysis.

Detection of Δ4-3-oxo-steroid 5β-reductase deficiency by LC–ESI-MS/MS measurement of urinary bile acids

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Akina Muto, Hajime Takei, Atsushi Unno, Tsuyoshi Murai, Takao Kurosawa, Shoujiro Ogawa, Takashi Iida, Shigeo Ikegawa, Jun Mori, Akira Ohtake, Takayuki Hoshina, Tatsuki Mizuochi, Akihiko Kimura, Alan F. Hofmann, Lee R. Hagey, Hiroshi Nittono
The synthesis of bile salts from cholesterol is a complex biochemical pathway involving at least 16 enzymes. Most inborn errors of bile acid biosynthesis result in excessive formation of intermediates and/or their metabolites that accumulate in blood and are excreted in part in urine. Early detection is important as oral therapy with bile acids results in improvement. In the past, these intermediates in bile acid biosynthesis have been detected in neonatal blood or urine by screening with FAB-MS followed by detailed characterization using GC–MS. Both methods have proved difficult to automate, and currently most laboratories screen candidate samples using LC–MS/MS. Here, we describe a new, simple and sensitive analytical method for the identification and characterization of 39 conjugated and unconjugated bile acids, including Δ4-3-oxo- and Δ4,6-3-oxo-bile acids (markers for Δ4-3-oxo-steroid 5β-reductase deficiency), using liquid chromatography-electrospray ionization tandem mass spectrometry (LC–ESI-MS/MS). In this procedure a concentrated, desalted urinary sample (diluted with ethanol) is injected directly into the LC–ESI-MS/MS, operated with ESI and in the negative ion mode; quantification is obtained by selected reaction monitoring (SRM). To evaluate the performance of our new method, we compared it to a validated method using GC–MS, in the analysis of urine from two patients with genetically confirmed Δ4-3-oxo-steroid 5β-reductase deficiency as well as a third patient with an elevated concentration of abnormal conjugated and unconjugated Δ4-3-oxo-bile acids. The Δ4-3-oxo-bile acids concentration recovered in three patients with 5β-reductase deficiency were 48.8, 58.9, and 49.4μmol/mmol creatinine, respectively by LC–ESI-MS/MS.

Highlights

► We analyze 39 bile acids including abnormal bile acids by LC–ESI-MS/MS. ► We analyze urine from patients with 5β-reductase deficiency by LC–ESI-MS/MS. ► The urine from those patients elevated concentration of Δ4-3-oxo-bile acids. ► The results obtained with LC–ESI-MS/MS agree quite well with those obtained by GC–MS.

Production, recovery and purification of a recombinant β-galactosidase by expanded bed anion exchange adsorption

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Valeria Boeris, Izabella Balce, Rami Reddy Vennapusa, Miguel Arévalo Rodríguez, Guillermo Picó, Marcelo Fernández Lahore
β-Galactosidase is a hydrolase enzyme that catalyzes the hydrolysis of β-galactosides into monosaccharides; its major application in the food industry is to reduce the content of lactose in lactic products. The aim of this work is to recover this enzyme from a cell lysate by adsorption onto Streamline-DEAE in an expanded bed, avoiding, as much as possible, biomass deposition onto the adsorbent matrix. So as to achieve less cell debris–matrix interaction, the adsorbent surface was covered with polyvinyl pyrrolidone. The enzyme showed to bind in the same extent to naked and covered Streamline-DEAE (65mg β-gal/g matrix) in batch mode in the absence of any biomass. The kinetics of the adsorption process was studied and no effect of the polyvinyl pyrrolidone covering was found. The optimal conditions for the recovery were achieved by using a lysate made of 40% wet weight of cells, a polyvinyl pyrrolidone-covered matrix/lysate ratio of 10% and carrying out the adsorption process in expanded bed with recirculation over 2h in 20mM phosphate buffer pH 7.4. The fraction recovered after the elution contained 65% of the initial amount of enzyme with a 12.6-fold increased specific activity with respect to the lysate. The polyvinyl pyrrolidone content in the eluate was determined and found negligible. The remarkable point of this work is that it was possible to partially purify the enzyme using a feedstock containing an unusually high biomass concentration in the presence of polyvinyl pyrrolidone onto weak anion exchangers.

Highlights

► β-Galactosidase production from recombinant Saccharomyces cerevisiae. ► Cell lysate and adsorption onto Streamline-DEAE in an expanded bed. ► Adsorption of target proteins from unclarified feedstock. ► Cell debris–matrix interaction, and the effect of polyvinyl pyrrolidone on the enzyme adsorption.

Simultaneous determination of asymmetric and symmetric dimethylarginine, l-monomethylarginine, l-arginine, and l-homoarginine in biological samples using stable isotope dilution liquid chromatography tandem mass spectrometry

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Mariska Davids, Eliane Swieringa, Fredrik Palm, Desirée E.C. Smith, Yvo M. Smulders, Peter G. Scheffer, Henk J. Blom, Tom Teerlink
Production of the endogenous vasodilator nitric oxide (NO) from l-arginine by NO synthase is modulated by l-homoarginine, l-monomethylargine (MMA), asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA). Here we report on a stable isotope dilution liquid chromatography tandem mass spectrometry (LC–MS/MS) method for simultaneous determination of these metabolites in plasma, cells and tissues. After addition of the internal standards (D7-ADMA, D4-l-homoarginine and 13C6-l-arginine), analytes were extracted from the samples using Waters Oasis MCX solid phase extraction cartridges. Butylated analytes were separated isocratically on a Waters XTerra MS C18 column (3.5μm, 3.9mm×100mm) using 600mg/L ammonium formate in water - acetonitrile (95.5:4.5, v/v) containing 0.1vol% formic acid, and subsequently measured on an AB Sciex API 3000 triple quadrupole mass spectrometer. Multiple reaction monitoring in positive mode was used for analyte quantification. Validation was performed in plasma. Calibration lines were linear (r 2 ≥0.9979) and lower limits of quantification in plasma were 0.4nM for ADMA and SDMA and 0.8nM for the other analytes. Accuracy (% bias) was <3% except for MMA (<7%), intra-assay precision (expressed as CV) was <3.5%, inter-assay precision <9.6%, and recovery 92.9–103.2% for all analytes. The method showed good correlation (r 2 ≥0.9125) with our previously validated HPLC-fluorescence method for measurement in plasma, and was implemented with good performance for measurement of tissue samples. Application of the method revealed the remarkably fast (i.e. within 60min) appearance in plasma of stable isotope-labeled ADMA, SDMA, and MMA during infusion of D3-methyl-1-13C-methionine in healthy volunteers.

Highlights

► The endogenous vasodilator nitric oxide (NO) is produced from arginine by NO synthase. ► Homoarginine and the methylated arginines MMA, ADMA, and SDMA modulate NO production. ► An LC–MS/MS method for the simultaneous measurement of these compounds was developed. ► Stable isotope labeled arginine, homoarginine and ADMA served as internal standards. ► The method was validated for plasma and is also suitable for analysis of tissue samples.

Three-phase hollow fiber liquid-phase microextraction of organophosphorous nerve agent degradation products from complex samples

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Charlotte Desoubries, Florence Chapuis-Hugon, Anne Bossée, Valérie Pichon
Degradation products of chemical warfare agents are considered as important environmental and biological markers of chemical attacks. Alkyl methylphosphonic acids (AMPAs), resulting from the fast hydrolysis of nerve agents, such as sarin and soman, and the methylphosphonic acid (MPA), final degradation product of AMPAs, were determined from complex matrices by using an emergent and miniaturized extraction technique, the hollow fiber liquid-phase microextraction (HF-LPME), before their analysis by liquid chromatography coupled to mass spectrometry (LC–MS). After studying different conditions of separation in the reversed phase LC–MS analysis, the sample treatment method was set up. The three-phase HF-LPME was carried out by using a porous polypropylene (PP) hollow fiber impregnated with 1-octanol that separates the donor and acceptor aqueous media. Various extraction parameters were evaluated such as the volume of the sample, the effect of the pH and the salt addition to the sample, the pH of the acceptor phase, the extraction temperature, the stirring speed of the sample, the immersion time in the organic solvent and the time of extraction. The optimum conditions were applied to the determination of MPA and five AMPAs in real samples, such as surface waters and urine. Compounds were extracted from a 3mL acidified sample into only 6μL of alkaline water without any other pretreatment of the complex matrices. Enrichment factors (EFs) higher than 170 were obtained for three less polar AMPAs. Limits of quantification (LOQs) in the 0.013–5.3ngmL−1 range were obtained after microextraction of AMPAs from river water and in the range of 0.056–4.8ngmL−1 from urine samples with RSD values between 1 and 9%.

Highlights

► High potential of HF-LPME applied to reduced size samples. ► High enrichment factor for the extraction of polar compounds from water and urine. ► Quantitative extraction of AMPAs from surface waters and urine samples. ► High sensitivity for the LC/MS analysis of AMPAs without any derivatization step.

Simultaneous determination of amoxicillin and prednisolone in bovine milk using ultra-high performance liquid chromatography tandem mass spectrometry

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Hui Li, Xi Xia, Yanan Xue, Shusheng Tang, Xilong Xiao, Jiancheng Li, Jianzhong Shen
A rapid and sensitive ultra-high performance liquid chromatography tandem mass spectrometric method was developed for simultaneous quantification of amoxicillin and prednisolone in bovine milk. In this method, amoxicillin, prednisolone and the internal standards penicillin G-d7 (for amoxicillin) and prednisolone-d6 were extracted from bovine milk using acetonitrile. The C18 solid phase extraction cartridges were selected for cleaning-up the extracts. The analytes were determined using a triple quadrupole mass spectrometry in positive electrospray ionization and multiple reaction monitoring mode. Calibration curves were linear over a concentration range of 2–1000μg/kg for the analytes. The mean recoveries were 89.2–92.3% for amoxicillin and 98.7–102.3% for prednisolone. Limits of detection were 0.5μg/kg for the analytes, and the limits of quantitation were 2μg/kg. Decision limit (CCα) and detection capability (CCβ) have also been estimated for each analyte. The method was validated according to the Commission Decision 2002/657/EC and successfully applied to the analysis of amoxicillin and prednisolone in real samples.

Highlights

► Amoxicillin and prednisolone were determined simultaneously using UPLC–MS/MS. ► Deuterated internal standards were applied to accurate quantification. ► LODs for two analytes were far below MRLs set by EU in developed method. ► Recoveries in raw milk were good with a simple sample preparation.

Development of a three-steps derivatization assay for the localization of double bond in monounsaturated monomers of poly-beta-hydroxyalkanoates by GC–MS

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Christelle Simon-Colin, Christelle Gouin, Pierre Lemechko, Nelly Kervarec, Jean Guezennec
A new gas chromatography–mass spectrometry (GC–MS) method for the localization of double bond in monounsaturated 3-hydroxyalkenoic acids monomers has been developed. A three steps derivation assay was used including a methanolysis, then acetylation and dimethyldisulfide (DMDS) addition to alkene groups. Electron impact GC–MS analysis of such derivatives offers characteristic fragments allowing the unambiguous determination of double bond position in side chain. This novel method is well-suited for the routine analysis of poly-beta-hydroxyalkanoates (PHAs), and was used to characterize monounsaturated monomers in both 3-hydroxyalkenoic acids standards as well as in mcl-PHAs and poly(3-hydroxyoctanoate-co-3-hydroxyundecenoate) (PHOU) produced by bacterial strain Pseudomonas guezennei from glucose or a mixture of sodium octanoate plus 10-undecenoic acid, respectively.

Highlights

► Localization of double bond in monounsaturated 3-hydroxyalkenoic acids by GC–MS. ► The new method implies methanolysis, then acetylation and thiomethylation. ► The method was successfully applied to bacterial poly-beta-hydroxyalcanoates.

Simultaneous clarification of Escherichia coli culture and purification of extracellularly produced penicillin G acylase using tangential flow filtration and anion-exchange membrane chromatography (TFF-AEMC)

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Valerie Orr, Jeno Scharer, Murray Moo-Young, C. Howie Honeyman, Drew Fenner, Lisa Crossley, Shing-Yi Suen, C. Perry Chou
Downstream purification often represents the most cost-intensive step in the manufacturing of recombinant proteins since conventional purification processes are lengthy, technically complicated, and time-consuming. To address this issue, herein we demonstrated the simultaneous clarification and purification of the extracellularly produced recombinant protein by Escherichia coli using an integrated system of tangential flow filtration and anion exchange membrane chromatography (TFF-AEMC). After cultivation in a bench-top bioreactor with 1L working volume using the developed host/vector system for high-level expression and effective secretion of recombinant penicillin G acylase (PAC), the whole culture broth was applied directly to the established system. One-step purification of recombinant PAC was achieved based on the dual nature of membrane chromatography (i.e. microfiltration-sized pores and anion-exchange chemistry) and cross-flow operations. Most contaminant proteins in the extracellular medium were captured by the anion-exchange membrane and cells remained in the retentate, whereas extracellular PAC was purified and collected in the filtrate. The batch time for both cultivation and purification was less than 24h and recombinant PAC with high purity (19U/mg), yield (72% recovery), and productivity (41mg of purified PAC per liter of culture) was obtained. Due to the nature of the non-selective protein secretion system and the versatility of ion-exchange membrane chromatography, the developed system can be widely applied for effective production and purification of recombinant proteins.

Highlights

► A novel protein separation system of TFF-AEMC was developed. ► The system was applied for effective culture clarification and protein purification. ► The batch time for combined cultivation and purification was less than 24h. ► Recombinant protein with a high purity and yield was obtained.

Determination of arachidonic acid by on-line solid-phase extraction HPLC with UV detection for screening of cytosolic phospholipase A2α inhibitors

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Walburga Hanekamp, Matthias Lehr
An on-line solid-phase extraction (SPE)–liquid chromatographic method with ultraviolet detection at 200nm for screening of inhibitors of cytosolic phospholipase A2α (cPLA2α) was developed and validated. cPLA2α was isolated from porcine platelets. Enzyme activity was determined by measuring the release of arachidonic acid from a phospholipid substrate using automated on-line sample clean up on a trap column followed by isocratic back-flush elution on a RP18 analytical column. While the use of a conventional RP18 column for trapping the analyte led to peak broadening only after a few runs due to pollution of the column by binding of components present in the enzyme preparation, the application of a turbulent flow column (TurboFlow Cyclone™) resulted in sharp peaks even after a plurality of injections. Interestingly, for sample introduction a turbulent flow of the mobile phase produced by high flow rates was not necessary to maintain good peak shapes. The same result could also be achieved applying low flow rates (0.5mL/min). Several known cPLA2α inhibitors were used to validate the test system.

Highlights

► Automated on-line solid-phase extraction–HPLC/UV method for cPLA2α inhibitor screening. ► Application of a turbulent flow column for trapping of the enzyme product arachidonic acid. ► A turbulent flow of the mobile phase is not necessary for effective separation of proteins.

Use of hollow fiber liquid phase microextraction and HPLC for extraction and determination of apigenin in human urine after consumption of Satureja sahendica Bornm.

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Mohammad Reza Hadjmohammadi, Mona Soltani, Vahid sharifi
The applicability of hollow fiber liquid phase microextraction (HF-LPME) was evaluated for extraction and preconcentration of apigenin prior to its determination by HPLC. Different parameters affecting the HF-LPME recovery such as nature of organic solvent, pH of donor and acceptor phases, extraction time, stirring speed, salt addition were optimized. Under optimum conditions (1-octanol as organic solvent, pH of the donor phase=3 and pH of acceptor phase=11.5, extraction time of 75min, stirring speed of 1000rpm) limit of detection (LOD) of 0.1ng/mL, linear range of 0.5–300ng/mL and correlation of determination (R 2) of 0.9956 were obtained. The relative intra and inter-day standard deviations (RSD%) based on five replicate measurement were 3.5% and 10.7% respectively. Enrichment factor of 315 and recovery 85% were achieved. Finally, the applicability of the proposed method was evaluated by extraction and determination of apigenin in urine sample after consumption of Satureja sahendica Bornm. which is a native medicinal plant from Iran. Concentration of apigenin in urine sample was found to be 6.20ng/mL.

Highlights

► A simple and sensitive method for determination of apigenin has been introduced. ► Hollow fiber liquid phase microextraction was used for extraction of apigenin. ► Trace amount of apigenin in biological sample was preconcentrated using this method. ► The HF-LPME method has excellent clean-up and high-preconcentration factor.

A liquid chromatography–tandem mass spectrometric method for quantification of curcumin-O-glucuronide and curcumin in human plasma

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Wei Chen, Patty Fan-Havard, Lisa D. Yee, Yu Cao, Gary D. Stoner, Kenneth K. Chan, Zhongfa Liu
Curcumin is a widely used herbal medicine for various human diseases including inflammation and cancer. The demonstration and optimization of curcumin's activities in the clinical setting, however, have been compromised by its poor bioavailability and the lack of analytic methods to monitor its absorption. In this paper, we report the first validated liquid chromatography–tandem mass spectrometric method for simultaneous quantification of curcumin and its major metabolite: curcumin-O-glucuronide (COG), in the linear range of 2.0–2000ng/mL in human plasma. The intra-day and inter-day accuracies of curcumin and COG in human plasma were in the range of 91.3–111.5% and 82.7–109.2% and their co-efficiency of variations were in the range of 3.5–12.7% and 3.1–11.3%, respectively. This method was capable of detecting only COG in human plasma samples from two healthy volunteers after an oral ingestion of curcumin.

Highlights

► First direct method to measure curcumin-O-glucuronide (COG) in human plasma. ► The LLOQ is 2ng/mL with CVs (<15%) and accuracy (85–115%). ► Capable of characterizing the pharmacokinetics of COG up to 24h in human.

Determination of phenylethanolamine A in animal hair, tissues and feeds by reversed phase liquid chromatography tandem mass spectrometry with QuEChERS

27 June 2012, 08:45:31
Publication year: 2012
Source:Journal of Chromatography B, Volume 900
Ming-Xia Zhang, Cun Li, Yin-Liang Wu
A simple, sensitive and reliable analytical method was developed for the determination of a new beta-agonist phenylethanolamine A in animal hair, tissues and animal feeds by ultra high performance liquid chromatography–positive electrospray ionization tandem mass spectrometry (UHPLC–ESI-MS/MS) with QuEChERS. Samples were extracted with acetonitrile/water (80:20, v/v). The extract was purified through QuEChERS method, then was dried with nitrogen and residues were redissolved in mobile phase for hair sample or directly diluted with 0.1% formic acid in water for other samples, and analyzed by LC–MS/MS on a Waters Acquity BEH C18 column with 0.1% formic acid in water/methanol as mobile phase with gradient elution. The samples were quantified using phenylethanolamine A-D3 as internal standards. The proposed method was validated according to the European Commission Decision 2002/657/EC determining specificity, decision limit (CCα), detection capability (CCβ), recovery, precision, linearity, robustness and stability. The CCα values ranged from 0.10 to 0.26μg/kg. The CCβ values ranged from 0.20 to 0.37μg/kg. The mean recoveries of 95.4–108.9% with intra-day CVs of 2.2–5.6% and inter-day CVs of 3.1–6.2% were obtained. The method is demonstrated to be suitable for the determination of phenylethanolamine A in animal hair, tissues and animal feeds. The total time required for the analysis of one sample except animal hair sample, including sample preparation, was about 25min.

Highlights

► A LC–MS–MS method has been developed to determine phenylethanolamine A residues. ► A simple pretreatment method based on QuEChERS has been developed. ► Animal hair, tissues and feeds were successfully analyzed using the method. ► The method was fast, reliable, convenient and sensitive.