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