High performance liquid chromatographic–mass spectrometric detection of giant fullerenes

Fullerene-rich soot generated by resistive heating of graphite has been gently extracted with toluene, in order to remove some C₆₀ and C₇₀ compounds, followed by extraction with boiling 1,2,4-trichlorobenzene at 214°C. After filtration and removal of the solvent, the residue was re-dissolved in dichloromethane and characterized by non-aqueous reversed phase liquid chromatography coupled on-line with atmospheric pressure chemical ionization mass spectrometry. Several novel fullerenes were detected, including C₇₂, C₈₀, C₈₆, and C₈₈ as well as other fullerenes up to C₁₀₈ and higher. The results indicate that chromatographic separation of large fullerene molecules can be achieved with low boiling point solvents and conventional liquid chromatographic techniques.     

超高效液相色谱-串联质谱法同时测定不同饲料中30种真菌毒素

采用QuEChERS前处理技术,建立了超高效液相色谱-串联质谱(UHPLC-MS/MS)检测不同饲料样品(预混料、浓缩料和配合料)中30种真菌毒素含量的分析方法。饲料样品经5 mL水和5 mL含1%(v/v)甲酸的乙腈溶液提取后,取上清液氮吹至近干,残渣经1 mL 5 mmol/L醋酸铵水溶液-乙腈(80∶20,v/v)复溶后,上机测定。采用基质匹配标准曲线结合同位素内标法进行定量分析。在低、中、高3个添加水平下,30种真菌毒素的平均加标回收率为72.0%~118.4%(n=5),30种真菌毒素在各自的线性范围内线性关系良好,相关系数(r 2)≥0.99,检出限(LOD,S/N=3)和定量限(LOQ,S/N=10)分别为0.7~20μg/L和2~50μg/L。该法简单、快速、实用性强,适用于预混料、浓缩料和配合料中30种真菌毒素的定量分析。     

Application of multiwalled carbon nanotubes as sorbents for the extraction of mycotoxins in water samples and infant milk formula prior to high performance liquid chromatography mass spectrometry analysis

In this work, a simple and environmental friendly methodology has been developed for the analysis of a group of six mycotoxins with estrogenic activity produced by Fusarium species (i.e. zearalanone, zearalenone, α‐zearalanol, β‐zearalanol, α‐zearalenol, and β‐zearalenol), using microdispersive SPE (μ‐dSPE) with multiwalled carbon nanotubes as sorbent. Separation, determination, and quantification were achieved by HPLC coupled to ion trap MS with an ESI interface. Parameters affecting the extraction efficiency of µ‐dSPE such as pH of the sample, amount of multiwalled carbon nanotubes, and type and volume of elution solvent, were studied and optimized. The methodology was validated for mineral, pond, and wastewater as well as for powdered infant milk using 17β‐estradiol‐2,4,16,16,17‐d₅ (17β‐E₂‐D₅) as internal standard, obtaining recoveries ranging from 85 to 120% for the three types of water samples and from 77 to 115% for powdered infant milk. RSD values were lower than 10%. The LOQs achieved were in the range 0.05–2.90 μg/L for water samples and 2.02–31.9 μg/L for powdered infant milk samples.     

高效液相色谱-串联质谱法同时测定饲料中11种霉菌毒素

建立了高效液相色谱-串联质谱(HPLC-MS/MS)同时检测饲料中11种霉菌毒素的分析方法。样品经乙腈提取,MycoSep 228多功能净化柱填料和PRIME HLB固相萃取柱净化;采用Agilent Zorbax SB-C18色谱柱(150 mm×2.1 mm,3.5μm)分离,以甲醇和5 mmol/L乙酸铵水溶液(含0.1%(v/v)甲酸)为流动相梯度洗脱;采用ESI源正、负离子同时扫描,多反应监测(MRM)模式测定,内标法定量。结果表明,11种目标物在各自的线性范围内线性关系良好,相关系数均大于0.99,定量限为2.0~50.0μg/kg。11种霉菌毒素在3个加标水平(1、2、5倍定量限)下的平均回收率为79.3%~101.6%,相对标准偏差(RSD)为5.9%~13.2%(n=5)。该法简单快速,净化效果好,灵敏度高,适用于饲料中11种霉菌毒素的分析。     

High-efficiency high performance liquid chromatographic analysis of red wine anthocyanins

The analysis of anthocyanins in natural products is of significant relevance in recent times due to the recognised health benefits associated with their consumption. In red grapes and wines in particular, anthocyanins are known to contribute important properties to the sensory (colour and taste), anti-oxidant- and ageing characteristics. However, the detailed investigation of the alteration of these compounds during wine ageing is hampered by the challenges associated with the separation of grape-derived anthocyanins and their derived products. High performance liquid chromatography (HPLC) is primarily used for this purpose, often in combination with mass spectrometric (MS) detection, although conventional HPLC methods provide incomplete resolution. We have previously demonstrated how on-column inter-conversion reactions are responsible for poor chromatographic efficiency in the HPLC analysis of anthocyanins, and how an increase in temperature and decrease in particle size may improve the chromatographic performance. In the current contribution an experimental configuration for the high efficiency analysis of anthocyanins is derived using the kinetic plot method (KPM). Further, it is shown how analysis under optimal conditions, in combination with MS detection, delivers much improved separation and identification of red wine anthocyanins and their derived products. This improved analytical performance holds promise for the in-depth investigation of these influential compounds in wine during ageing.     

Novel approaches in analysis of Fusarium mycotoxins in cereals employing ultra performance liquid chromatography coupled with high resolution mass spectrometry

Rapid, simple and cost-effective analytical methods with performance characteristics matching regulatory requirements are needed for effective control of occurrence of Fusarium toxins in cereals and cereal-based products to which they might be transferred during processing. Within this study, two alternative approaches enabling retrospective data analysis and identification of unknown signals in sample extracts have been implemented and validated for determination of 11 major Fusarium toxins. In both cases, ultra-high performance liquid chromatography (U-HPLC) coupled with high resolution mass spectrometry (HR MS) was employed. ¹³C isotopically labeled surrogates as well as matrix-matched standards were employed for quantification. As far as time of flight mass analyzer (TOF-MS) was a detection tool, the use of modified QuEChERS (quick easy cheap effective rugged and safe) sample preparation procedure, widely employed in multi-pesticides residue analysis, was shown as an optimal approach to obtain low detection limits. The second challenging alternative, enabling direct analysis of crude extract, was the use of mass analyzer based on Orbitrap technology. In addition to demonstration of full compliance of the new methods with Commission Regulation (EC) No. 401/2006, also their potential to be used for confirmatory purposes according to Commission Decision 2002/657/EC has been critically assessed.     

Improved liquid chromatographic determination of nine currently used (fluoro)quinolones with fluorescence and mass spectrometric detection for environmental samples

An HPLC method using C18-modified silica as stationary phase has been developed for environmental trace analysis of nine (fluoro)quinolones. Detection is done by fluorescence measurement or MS using the modes of SIM and selected reaction monitoring (SRM). Best separation is achieved with a gradient consisting of 50 mM formic acid and methanol, which is fully compatible with MS coupling. LOQs (S/N of 10) for fluorescence detection are between 10 and 60 microg/L, depending on the analyte. MS detection (SIM and SRM) yields LOQs that are better by a factor of at least an order of magnitude. Sample preconcentration and sample clean-up is accomplished by SPE (preconcentration factor of 1000), leading to LOQs in the low ng/L range. Recoveries of the preconcentration procedure are better than 80% for all analytes. The suitability for real samples has been demonstrated by analyzing surface waters, municipal waste waters, sewage treatment plant effluents, sewage sludge, and sediment taken from rivers and fish ponds. The method should also be useful for determination of residues of (fluoro)quinolones in food or other matrices. The degradation of the (fluoro)quinolones has been examined over 5 days in order to get information about the decomposition rate and the degradation products eventually occurring in the environment.     

Isolation and determination of paspalitrem-type tremorgenic mycotoxins using liquid chromatography with diode-array detection

A liquid chromatographic (LC) method is described for the isolation and determination of the tremorgenic mycotoxins paxilline (Penicillium paxilli NRRL 6110), paspaline, paspalinine and paspalicine (Claviceps paspali). Following a Soxhlet extraction of a mould-contaminated matrix using chloroform, the crude extract was partitioned between hexane and 80% aqueous methanol. The latter fraction, containing the desired toxin(s), was evaporated to dryness, the residue dissolved in methylene chloride and the solution analysed by liquid chromatography using a Supelcosil LC-Si column eluted with methylene chloride-diethyl ether (9 + 1, v/v). A mixture containing standards of these compounds was similarly analysed. All toxins were detected using a UV diode-array detector. The generated UV spectra and chromatographic data of the standard toxins were stored in a computer as a library and used to identify these toxins in a crude mixture. The purity of the separated peaks and the amount of toxin in the crude mixture were also determined. The toxins were isolated by selectively collecting the eluted peaks using a programmable fraction collector equipped with a peak level sensor. Further confirmation of compound identity was achieved by mass spectrometry using the direct inlet probe method. In comparison with methods used previously to isolate these toxins, the present technique is fast and allows the acquisition of complete UV spectral information and chromatographic data and the isolation of multiple toxins in a single chromatographic operation.     

Quantitation of hindered amine light stabilizers in plastic materials by high performance liquid chromatography and mass spectrometric detection using electrospray ionization and atmospheric pressure photoionization

In this work new high performance liquid chromatographic methods in combination with mass spectrometry have been developed for the quantitation of hindered amine light stabilizers (HALS) which are commonly used as monomeric and oligomeric species for stabilization of plastic materials. These analytes are difficult to separate under traditional reversed phase conditions. In the present study new silica-based pH stable reversed phases that had become available recently were investigated for HALS analysis, and turned out to be well suited employing mobile phases at a pH of around 11 adjusted by addition of ammonia. Detection was done by mass spectrometry employing both time-of-flight and triple quadrupole mass analyzers. The performance of electrospray ionization (ESI) as well as atmospheric pressure photoionization (APPI) was investigated and compared. Despite the high pH of the mobile phase, an excellent ionization could be obtained in the positive ion mode. ESI provided slightly lower limits of quantitation (on average a factor of 2) in comparison with APPI. The method allowed the quantitation of a range of different HALS down to 0.05–0.005% (depending on the HALS) in polymeric materials. Sample preparation consisted in dissolving the sample in toluene and precipitation of the polymer with acetone. The procedure can be routinely applied to aging tests of plastic materials in order to predict the lifetime of plastic components.     

Evaluation of different liquid chromatography-electrospray mass spectrometry systems for the analysis of heterocyclic amines

Three liquid chromatography-electrospray ionisation (LC-ESI) MS systems are evaluated for the analysis of heterocyclic amines (HAs). The electrospray sources and analysers (ion trap, single quadrupole and triple quadrupole) have been compared in terms of performance and quality parameters. In all cases, a C8 reversed-phase column and (acetic acid-ammonium acetate 30 mM pH 4.5)-acetonitrile (ACN) as mobile phase were used. Ionisation source parameters, post-column addition and working conditions for each acquisition mode (full scan, product ion scan, selected ion monitoring, and multiple reaction monitoring) were optimised for each instrument. The MS-MS spectra obtained with the ion trap and the triple quadrupole systems were very similar in both fragment ions and relative abundances, except for carbolines that showed adduct formation in the ion trap. Quality parameters were established and good precision (relative standard deviations (R.S.D.) < 12%) and very low limits of detection were obtained, mainly when using the triple quadrupole (< 9 pg injected). The content of HAs in a lyophilised beef extract was determined using the three instruments in order to compare their applicability for routine HAs analysis.     

HPLC,mass spectrometry,and LC/MS of gossypol and its derivatives

Conditions are reported for the reverse-phase HPLC analysis of gossypol and of some of its derivatives, including anhydrogossypol, gossypolone, gossypol hexaacetate, and Schiff's base derivatives. It is shown that field desorption (FD) and chemical ionization (CI) mass spectrometry have advantages over electron impact (EI) mass spectrometry for the characterization of these compounds and that combined high-performance liquid chromatography/mass spectrometry (LC/MS), especially when used in the CI mode, is particularly effective.     

Analysis of proteoglycans derived sulphated disaccharides by liquid chromatography/mass spectrometry

A method has been developed for the identification and quantitative determination of sulphated disaccharides derived from chondroitin sulphate (CS) and dermatan sulphate (DS) chains attached to proteoglycans (PGs). After digestion with Chondroitinase ABC, the pool of disaccharides can be directly separated by liquid chromatography on a porous graphitized carbon (PGC) column and identified by on-line electrospray mass spectrometry under negative ionization conditions. The relative intensities of the fragment ions obtained by MS/MS allow to distinguish the sulphate position. Calibration with standard disaccharides allows the quantification of the different isomers. The method showed good repeatability in terms of relative standard deviation (RSD < 2%) and linearity between 0.5 and 50 ng (total injected amount) for both 4- and 6-sulphated disaccharides. The limit of detection achieved in full scan mode was 0.1 ng. The methodology was applied to different types of biological samples obtained from patients suffering from chronic lung inflammation such as: lung tissue, bronchoalveolar lavage fluid (BALF), induced sputum and urine.     

高效液相色谱-串联质谱法检测牛奶中头孢洛宁残留

建立了牛奶中头孢洛宁残留检测的高效液相色谱-串联质谱方法。1 g牛奶经乙腈沉淀蛋白质后,上清液于37 ℃水浴下氮气吹干,用1 mL甲醇-0.1%甲酸水溶液（3：7,v/v）复溶,正己烷除脂净化后检测。流动相为乙腈和0.1%甲酸水溶液,梯度洗脱,经C₁₈色谱柱分离,采用多反应监测正离子模式对头孢洛宁进行定性定量分析。采用基质匹配法对牛奶中头孢洛宁的含量进行标准校正,在2~200 μg/L范围内,头孢洛宁质量浓度与其对应峰面积的线性关系良好,相关系数>0.999。牛奶中加标样品的检出限（按S/N≥3计）为0.5 μg/kg,定量限（S/N≥10计）为2 μg/kg。在定量限、1/2最高残留限量、最高残留限量、2倍最高残留限量添加水平下,牛奶中头孢洛宁的平均回收率为78.5%~86.2%,日内相对标准偏差为1.5%~6.2%,日间相对标准偏差为2.9%~5.6%。该方法可用于牛奶中头孢洛宁的残留检测。     

Ultra high performance liquid chromatography coupled to tandem mass spectrometry determination of lipid peroxidation biomarkers in newborn serum samples

Byproducts of arachidonic (AA) and docosahexaenoic acid (DHA) oxidation are highly relevant for the study of free radical associated conditions in the perinatal period. Plasma metabolites can provide the clinician with a snapshot of the oxidant status of patients before and after specific clinical interventions (e.g.: supplementation with oxygen). We describe a new andreliable ultra-performance liquid mass spectrometry method to determine F2-isoprostanes and other byproducts (isoprostanes, isofurans, neuroprostanes, neurofurans) in newborn serum samples. Cord blood samples were obtained from severely depressed newborn infants (Apgar score 1 min < 3; arterial cord pH < 7.00), and aliquoted for serum determination and stored at −80 °C. A UHPLC-MS/MS method was employed. It has a series of technical advantages: simple sample treatment; reduced sample volume (100 μL) which is essential for preterm neonates with low circulating blood volume, high throughput of sample analysis (96 samples in less than 24 h) and high selectivity for different isoprostanes isomers. Excellent sensitivity was achieved within limits of detection between 0.06 and 4.2 nmol L⁻¹, which renders this method suitable to monitoranalyte concentration in newborn samples. The method's precision was satisfactory; with coefficients of variation around 5–12% (intra-day) and 7–17% (inter-day). The reliability of the described method was assessed by analysis of spiked serum samples obtaining recoveries between 70% and 120%. The proposed method has rendered suitable for serum determination for newborn babies at risk of oxygen free radical associated conditions.     

Ultra-high capacity liquid chromatography chip/quadrupole time-of-flight mass spectrometry for pharmaceutical analysis

Nanoflow liquid chromatography/mass spectrometry (nano-LC/MS) has attracted increasing interest in virtue of high sensitivity, low sample consumption, and minimal matrix effect. In this work a HPLC-Chip/quadrupole time-of-flight (Q-TOF) MS device with a new ultra-high capacity small molecule chip (UHC-Chip) which features a 500 nL enrichment column and a 150 mm × 75 μm analytical column, was evaluated with a drug mixture covering a wide range of polarities. Excellent chromatographic precision with 0.1-0.5% RSD for retention time and 1.7-9.0% RSD for peak area, low limit of detection, good chip-to-chip reproducibility and linearity were obtained by using this UHC-Chip. Compared with the standard HPLC-Chip with 40 nL trapping column, the UHC-Chip showed higher enrichment capability and hence gave a higher response in signal detection. Additionally, 4-30 times increase in sensitivity was obtained compared with conventional LC/MS, which indicated that UHC-Chip/MS was a valuable tool for the quantitative analysis of low level impurities and degradation products in pharmaceuticals. Moreover, satisfactory results obtained from trace drug analysis of serum samples further proved its practicality and potential for use in drug testing and development.     

Liquid chromatography-tandem mass spectrometric analysis and regulatory issues of polar pesticides in natural and treated waters

Pesticides are among the most detected contaminants in the aquatic environment. This is mainly due to their use in agriculture and their physico-chemical properties that enable transportation and a persistent or pseudo-persistent existence in the water media. Several directives and guidelines set maximum levels of pesticides in water in order to protect the human and environmental health. A brief discussion of the existing directives and guidelines concerning pesticides in water is presented, e.g., the new regulatory framework for the Registration, Evaluation and Authorisation of Chemicals (REACH), and the Directive 91/414/EEC concerning the placing of plant protection products on the market. Up-to-date analytical tools to support the REACH program are of prime importance to ensure its complete implementation. Since liquid chromatography (LC) coupled to mass spectrometry (MS) is considered the most appropriate technique for determination of most modern pesticides in environmental waters, the most recent developments and applications in this field are discussed in detail in this review.     

高效液相色谱-串联质谱法同时测定贝类中22种农药残留

建立了高效液相色谱-三重四极杆质谱（HPLC-MS/MS）同时测定贝类中22种农药残留的分析方法。样品经含0.1%（v/v）甲酸的乙腈提取，N-丙基乙二胺（PSA）和石墨化碳黑（GCB）净化，然后采用ACE UltraCore 2.5 SuperC18柱（100 mm×2.1 mm，2.5 μm）分离，以甲醇-0.1%（v/v）甲酸水溶液为流动相梯度洗脱，流速为0.4 mL/min，柱温为35 ℃，然后以电喷雾电离（ESI）源，在多反应监测（MRM）、正离子模式下，采用三重四极杆质谱检测。22种农药在各自的线性范围内线性关系良好，相关系数均大于0.997，检出限为0.1~0.3 μg/kg，定量限为0.3~1.0 μg/kg。在3个添加水平下，22种农药的平均回收率为65.2%~109.4%，相对标准偏差为1.3%~15.2%（n=6）。该方法操作简单，快速，准确度高，灵敏度高，可用于贝类中22种农药残留的同时检测。