Contribution of liquid-phase and gas-phase ionization in extractive electrospray ionization mass spectrometry of primary amines

In this study, we investigated how binary mixtures of compounds influence each other's signal intensity in electrospray ionization (ESI), extractive electrospray ionization (EESI) and secondary electrospray ionization (SESI) experiments. The experiments were conducted using a series of homologous primary amines (from 1-butyl to 1- decylamine). In every experiment, two of the amines were present, and all 21 possible combinations were measured with EESI, ESI and SESI as ionization sources. Except for the volatility, which decreases with increasing molecular weight, the physico-chemical properties of the amines are very similar, so that the intensity ratio obtained in each experiment provides information about discrimination effects occurring during the ionization process. The results show that for the relatively volatile compounds investigated, the EESI ionization mechanism resembles the SESI-like gas-phase charge transfer more than ESI-like analyte ionization in solution. In addition, almost no discrimination effects were observed in the spectra obtained in EESI experiments. Quantitative EESI experiments with nonylamine as internal standard showed that EESI is capable of providing both more accurate and more precise results than SESI and ESI.     

Secondary Electrospray Ionization of Complex Vapor Mixtures. Theoretical and Experimental Approach

In secondary electrospray ionization (SESI) systems, gaseous analytes exposed to an electrospray plume become ionized after charge is transferred from the charging electrosprayed particles (the charging agent) to the vapor species. Currently available SESI models are valid for simplified systems having only one type of electrosprayed species, which ionizes only one single vapor species, and for the limit of low vapor concentration. More realistic models require considering other effects. Here we develop a theoretical model that accounts for the effects of high vapor concentration, saturation effects, interferences between different vapor species, and electrosprays producing different types of species from the liquid phase. In spite of the relatively high complexity of the problem, we find simple relations between the different ionic species concentrations that hold independently of the particular ion source configuration. Our model suggests that an ideal SESI system should use highly concentrated charging agents composed preferably of only one dominating species with low mobility. Experimental measurements with a MeOH-H₂O-NH₃ electrospray and a mixture of fatty acids and lactic acid served to test the theory, which gives good qualitative results. These results also suggest that the SESI ionization mechanism is primarily based on ions rather than on charged droplets.     

High mass resolution breath analysis using secondary electrospray ionization mass spectrometry assisted by an ion funnel

In this study, we used secondary electrospray ionization mass spectrometry assisted by an ion funnel (IF) operating at ambient pressure to find compounds in the mass range of 100–500 m/z in online breath fingerprinting experiments. In low‐resolution experiments conducted on an ion trap instrument, we found that pyridine is present in breath of individuals long after drinking coffee. In high‐resolution experiments conducted on a Fourier transform ion cyclotron resonance, we found more than 30 compounds in the mass range of 100–500 m/z in analogous online breath experiments. More than a third of these compounds have molecular weights above 200 Daltons and have not been mentioned in previous studies. In low‐resolution experiments as well as experiments without the IF, these compounds could not be detected. Copyright © 2012 John Wiley & Sons, Ltd.     

二次纳流电喷雾电离耦合超高分辨质谱检测人体呼出气中相对高分子量化合物

二次电喷雾电离源耦合超高分辨质谱(SESI-UHRMS)有望检出人体呼出气中分子量大于300的相对高分子量化合物,这些化合物的发现将有助于更准确地理解呼出气中挥发性有机化合物(VOCs)的来源、产生机制以及SESI源电离机理,更好地实现SESI-UHRMS的转化应用.本研究自组装nanoSESI源(尚无商业产品)耦合四极杆-静电场轨道阱质谱(最高质量分辨率1.2×10~5),考察了该装置对健康人体呼出气中分子量为300~500化合物的检出情况.结果表明,所搭建的nanoSESI-UHRMS装置检测人体呼出气的重现性好、灵敏度高,可检出数十种分子量为300~500的化合物.根据这些化合物在单次呼气过程中信号强度随时间变化的趋势,推断其来源分别为内源性和外源性;各化合物的元素组成主要包括C,H,N和O,环-双键当量(RDB)的均值为(4.5±3.1),表明检出的化合物可能为醛酮或不饱和脂肪酸,容易在SESI源中被电离.本研究初步验证了自组建nanoSESI-UHRMS检测人体呼出气中相对高分子量化合物的可行性,为后续进一步开展方法应用奠定了基础.     

Novel, fully automatic hydrophilic interaction/reversed-phase column-switching high-performance liquid chromatographic system for the complementary analysis of polar and apolar compounds in complex samples

In this study we developed and optimized a column-switching high-performance liquid chromatographic (HPLC) system for the complementary analysis of polar and apolar compounds in complex samples. The polar compounds are separated on a hydrophilic interaction chromatographic (HILIC) column; the part of the sample non-retained on HILIC is transferred into and separated on a reversed-phase (RP) column through an interface, thus preventing the loss of analytes eluting at dead time in common single-column mode. The signals are in turn recorded in the same chromatogram either by a UV or by a mass spectrometric detector. Applying a mixture of 25 standard compounds (12 hydrophilic and 13 hydrophobic) the system proves to be reliable, robust and easy-operating. The investigation of urine revealed an increase in the detected metabolite ion masses compared to the single-column HILIC-electrospray ionization (ESI)-MS analysis. Utilizing online the complementary selectivity of HILIC and RPLC in a fully automatic mode, this approach holds out the prospect to enlarge the number of detectable compounds in non-targeted "-omics" studies and sophisticated target-driven approaches by a single injection.     

A 31P NMR Study of the Stability of Carboxyifosfamide and Carboxycyclophosphamide,Two Metabolites of Ifosfamide and Cyclophosphamide

Abstract

Ifosfamide (IF) and cyclophosphamide (CP) are two phosphorated anticancer agents used in the treatment of solid tumours. Several phosphorated metabolites, among them carboxyifosfamide (CXIF) and carboxycyclophosphamide (CXCP), were detected and quantified by ³¹P NMR in urine from patients treated with IF or CP. In agreement with other authors [1], we observed a great inter-patient variability in the urinary excretion of CXIF in patients treated with IF [2]. This variability was attributed to a genetic polymorphism of aldehyde dehydrogenase, the enzyme responsible for the formation of CXCP or CXIF [1,3]. Since CXCP and CXIF are unstable, we thought that the inter-individual variability could also be due to a degradation during the storage of urine samples. A ³¹P NMR study of the stability of CXIF and CXCP in urine as a function of time, pH (7 and 5.5) and storage temperature (25°C, 8°C, ?20°C, ?80°C) demonstrated that (i) CXCP and CXIF are more stable at pH 7 than at pH 5.5, (ii) CXCP is more stable than CXIF at both pH, (iii) the degradation decreases with temperature but still occurs at ?20°C and even ?80°C. For an accurate quantification of these compounds, the storage of urine samples must be done at ?80°C immediately after collection and not exceed 1 month at pH 7 whereas, at pH 5.5, the assay must be carried out in the few days following the sampling. To identify the degradation products of CXCP and CXIF, the time course of hydrolysis (between pH 2 and 7) of these compounds was monitored by ³¹P NMR. The structure of each compound formed was determined by mass spectrometry and ¹H and ¹³C NMR after their isolation (except compound A too unstable to be isolated). The results are reported in the following schemes.     

基于QSAR模型的有机磷化合物毒性预测

随着有机磷化合物(OPs)的广泛应用,其在越来越多的环境介质中被检测出来。大多数OPs具有毒性,但人们缺乏快速且有效的预测手段来对毒性进行评估。本文将结合E-Dragon软件计算的分子描述符,采用不同的QSAR模型对36个OPs的毒性进行预测。文中采用后退法作为描述符筛选方法,以均方根误差(RMSE)作为评价标准,共找到14个对线性核函数支持向量机(SVM)模型贡献较大的描述符;在最终得到的SVM模型交叉验证结果中,计算值与实际值的相关系数为0. 913,均方根误差为0. 388;外部测试验证结果中,平均相对误差为9. 10%。此外,采用多元线性回归(MLR)、人工神经网络(ANN)以及偏最小二乘回归(PLS)模型对OPs的毒性进行预测,交叉验证结果显示,三个模型的计算值与实际值的相关系数分别为0. 878、0. 686与0. 620,没有SVM模型的预测能力好。因此采用线性核函数的SVM模型对OPs进行毒性预测是一个行之有效的方法。     

Development of a sheathless CE‐ESI‐MS interface

A sheath‐flow interface is the most common ionization technique in CE‐ESI‐MS. However, this interface dilutes the analytes with the sheath liquid and decreases the sensitivity. In this study, we developed a sheathless CE‐MS interface to improve sensitivity. The interface was fabricated by making a small crack approximately 2 cm from the end of a capillary column fixed on a plastic plate, and then covering the crack with a dialysis membrane to prevent metabolite loss during separation. A voltage for CE separation was applied between the capillary inlet and the buffer reservoir. Under optimum conditions, 52 cationic metabolite standards were separated and selectively detected using MS. With a pressure injection of 5 kPa for 15 s (ca. 1.4 nL), the detection limits for the tested compounds were between 0.06 and 1.7 μmol/L (S/N = 3). The method was applied to analysis of cationic metabolites extracted from a small number (12 000) of cancer cells, and the number of peaks detected was about 2.5 times higher than when using conventional sheath‐flow CE‐MS. Because the interface is easy to construct, it is cost‐effective and can be adapted to any commercially available capillaries. This method is a powerful new tool for highly sensitive CE‐MS‐based metabolomic analysis.     

Liquid chromatography and tandem mass spectrometry: a powerful approach for the sensitive and rapid multiclass determination of pesticides and transformation products in water

In this paper we will show the results of our research on the direct simultaneous determination of multi-class pesticides and transformation products with different polarities and acid-base properties by applying an on-line trace enrichment coupled to the chromatographic system supplied with electrospray interface (SPE-LC-MS/MS method). The specific chromatographic separation allows the correct determination of almost fifty compounds (37 pesticides and 10 transformation products) using very low sample volume and very little sample handling. Recoveries between 70-120% were obtained for all compounds in drinking and groundwater, meanwhile in surface water 44 compounds were correctly quantified. Relative standard deviations lower than 15% were obtained for all compounds. Even at the lowest concentration level tested (25 ng L(-1)) 40 compounds presented satisfactory recoveries and repeatability. The use of methanol as organic modifier and the increase of injection volume are also studied. The applicability of the developed method to a monitoring programme is demonstrated by applying it to the analysis of hundreds of samples.     

Molecular orientation of asphaltenes and PAH model compounds in Langmuir-Blodgett films using sum frequency generation spectroscopy

Asphaltenes are an important class of compounds in crude oil whose surface activity is important for establishing reservoir rock wettability which impacts reservoir drainage. While many phenomenological interfacial studies with crude oils and asphaltenes have been reported, there is very little known about the molecular level interactions between asphaltenes and mineral surfaces. In this study, we analyze Langmuir-Blodgett films of asphaltenes and related model compounds with sum frequency generation (SFG) vibrational spectroscopy. In SFG, the polarization of the input (vis, IR) and output (SFG) beams can be varied, which allows the orientation of different functional groups at the interface to be determined. SFG clearly indicates that asphaltene polycyclic aromatic hydrocarbons (PAHs) are highly oriented in the plane of the interface and that the peripheral alkanes are transverse to the interface. In contrast, model compounds with oxygen functionality have PAHs oriented transverse to the interface. Computational quantum chemistry is used to support corresponding band assignments, enabling robust determination of functional group orientations.     

Possibilities and Limitations of Fast Temperature Programming as a Route towards Fast GC

One possible way to speed up a gas chromatographic analysis is the application of fast temperature programming by using resistive heating techniques. With this heating technique programming rates up to 20° per second can be reached. A relative standard deviation of retention times better than 0.2% is obtained. Using fast temperature programming the analysis-times of a mineral oil sample, an industrial oligomer sample, and toxic compounds in diesel fuel have been reduced 5 to 20 times, compared to a standard temperature programmed analysis. In most cases resistive heating cannot be applied to reduce the analysis time of a complex sample. The use of fast temperature programming is preferable to the use of short columns and columns operated at above-optimum carrier gas velocities.     

Alkaline Hydrolysis of the Cytostatic Drug,Ifosfamide, and its N-Dechloroethylated Metabolites

Abstract

I fosfamide (IF) is an alkylating antitumor agent used in the treatment of solid tumors. Up to 50% of IF administered to patients undergoes an oxidative N-dealkylation reaction resulting in the loss or one, other or both chloroethyl side chain(s) to produce 2- or 3-dechloioethylIF (ZDCIF, 3DCIF) or 2,3-didechloroethyllF (DDCIF). The hydrolytic pathway of these four oxazaphosphorines has been studied earlier but only at acidic and neutral pHs^[l] In the present work, we monitored their time courses of hydrolysis at basic pHs using phosphorus-3 1 NMR. The structures of the compounds formed were determined by NMR (13C and 1H) and mass spectrometry. The results are reported in the following scheme.     

Probing of ferrocenylanilines on model micelle/reverse micelle membrane and their enhanced reactivity for reactive oxidants

Reactive oxygen species are formed in the human body but can be removed by suitable antioxidants. In this study we synthesized and characterized three ferrocene derivatives, 4‐ferrocenylaniline (pFA), 3‐ferrocenylaniline (mFA) and 3‐methyl‐4‐ferrocenylaniline (MeFA), having significant potential to be used as antioxidants. The synthesized compounds are insoluble in water, with the solubility of these compounds increasing in micelle solution. The micelle and reverse micelle solutions were considered as model membranes. The synthesized compounds were probed on the model membranes, made by sodium dioctylsulfosuccinate reverse micelle and tetradecyltrimethylammonium bromide micelle, using ¹H NMR spectroscopy. The ¹H NMR results indicated that these compounds are present in the polar region of the model membrane interface. Quantitative measurements showed that mFA has the greatest ability to penetrate into the micelle membrane among these compounds, and pFA is least penetrating in this respect. Solubilization of these compounds in aqueous micelle solution facilitates crystallization (of mFA) and enhances the antioxidant potential of these compounds. X‐ray crystal structure analysis revealed that mFA captures water molecules during crystallization in micelle solution. Their ability to act as antioxidants was evaluated, in dimethylsulfoxide (DMSO) and in micelle solution, using standard 1,1‐ diphenyl‐2‐picrylhydrazyl (DPPH) assay. It was found that their antioxidant potential is good in DMSO and that potential increases on the interface of the model membrane. The highest increase (by 19.6%) in the antioxidant potential, on the model membrane interface, was observed for mFA.     

Use of Solid Phase Microextraction (SPME) with Ion Mobility Spectrometry∗

Abstract

We report the use of solid phase microextraction (SPME) combined with ion mobility spectrometry (IMS) for sampling, screening and identification of organic compounds that are readily detected by IMS. This is a new SPME application. SPME has emerged recently as an excellent sample preparation technique for gas chromatography (GC) and high performance liquid chromatography (HPLC). We have found that SPME can be used very conveniently with IMS. An example of SPME-IMS is described using SPME headspace sampling at room temperature with 0.1 mL vials containing 1.0 microgram or less of either cocaine freebase or cocaine hydrochloride. This is followed by analysis using IMS. A hole, drilled in the IMS sample ticket holder, serves as the SPME-IMS interface.

     

The application of gas chromatography/atmospheric pressure chemical ionisation time‐of‐flight mass spectrometry to impurity identification in Pharmaceutical Development

Accurate mass measurement (used to determine elemental formulae) is an essential tool for impurity identification in pharmaceutical development for process understanding. Accurate mass liquid chromatography/mass spectrometry (LC/MS) is used widely for these types of analyses; however, there are still many occasions when gas chromatography (GC)/MS is the appropriate technique. Therefore, the provision of robust technology to provide accurate mass GC/MS (and GC/MS/MS) for this type of activity is essential. In this report we describe the optimisation and application of a newly available atmospheric pressure chemical ionisation (APCI) interface to couple GC to time‐of‐flight (TOF) MS. To fully test the potential of the new interface the APCI source conditions were optimised, using a number of standard compounds, with a variety of structures, as used in synthesis at AstraZeneca. These compounds were subsequently analysed by GC/APCI‐TOF MS. This study was carried out to evaluate the range of compounds that are amenable to analysis using this technique. The range of compounds that can be detected and characterised using the technique was found to be extremely broad and include apolar hydrocarbons such as toluene. Both protonated molecules ([M + H]⁺) and radical cations (M^+.) were observed in the mass spectra produced by APCI, along with additional ion signals such as [M + H + O]⁺. The technique has been successfully applied to the identification of impurities in reaction mixtures from organic synthesis in process development. A typical mass accuracy of 1–2 mm/zunits (m/z 80–500) was achieved allowing the reaction impurities to be identified based on their elemental formulae. These results clearly demonstrate the potential of the technique as a tool for problem solving and process understanding in pharmaceutical development. The reaction mixtures were also analysed by GC/electron ionisation (EI)‐MS and GC/chemical ionisation (CI)‐MS to understand the capability of GC/APCI‐MS relative to these two firmly established techniques. Copyright © 2010 John Wiley & Sons, Ltd.     

二维气相色谱技术分析汽油中的3种酯类化合物

建立了一种采用填充柱切割-反吹二维气相色谱分析汽油中酯类化合物（包括乙酸乙酯、乙酸仲丁酯、碳酸二甲酯）的方法。利用非极性填充预柱将汽油中沸点低于正辛烷的轻组分保留进入分析柱,重组分反吹放空,轻组分和酯类化合物经一个装填有强极性固定相的色谱柱分离分析。采用外标法定量,3种酯类化合物在50~50000 mg/L范围内线性关系良好,相关系数（r²）分别为0.99999、1.00000和0.99995,标准样品6次重复性测定的相对标准偏差（RSD）均小于1.0%,回收率在98.7%~107.9%之间,方法检出限（S/N=3）为5 mg/L。该方法不需要进行样品前处理,具有操作简单,准确高效的特点,是汽油中酯类化合物测定的理想分析方法。     

On-line detection of human skin vapors

Vapors released by the skin in the hand of one human subject are detected in real time by sampling them directly from the ambient gas surrounding the hand, ionizing them by secondary electrospray ionization (SESI, via contact with the charged cloud from an electrospray source), and analyzing them in a mass spectrometer with an atmospheric pressure source (API-MS). This gas-phase approach is complementary to alternative on-line surface ionization methods such as DESI and DART. A dominating peak of lactic acid and a complete series of saturated and singly unsaturated fatty acids (C₁₂ to C₁₈) are observed, in accordance with previous off-line studies by gas chromatography-mass spectrometry. Several other metabolites have been identified, including ketomonocarboxylic and hydroxymonocarboxylic acids.     

Targeting AMPK signalling pathway with natural medicines for atherosclerosis therapy: an integration of in silico screening and in vitro assay

An integration of virtual screening and kinase assay was reported to identify AMPK kinase inhibitors from various natural medicines.The activation of AMP-activated protein kinase (AMPK) signalling pathway plays a central role in the pathologic progression of atherosclerosis (AS). Targeting the AMPK is thus considered as a potential therapeutics to attenuate AS. Here, we report the establishment of a synthetic pipeline that integrates in silico virtual screening and in vitro kinase assay to discover new lead compounds of AMPK inhibitors. The screening is performed against a large-size pool of structurally diverse natural products, from which a number of compounds are inferred as promising candidates, and few of them are further tested in vitro by using a standard kinase assay protocol to determine their inhibitory potency against AMPK. With this scheme we successfully identify five potent AMPK inhibitors with IC₅₀ values at micromolar level. We also examine the structural basis and molecular mechanism of nonbonded interaction network across the modelled complex interface of AMPK kinase domain with a newly identified natural medicine.     

The potential of on-line flame photometric detection in microcolumn liquid chromatography

The potential of an interface for the on-line coupling of microcolumn liquid chromatography (LC)and a flame photometric detector (FPD) has been further investigated. With the micro-LC/FPD system, relatively high-molecular-weight polar compounds such as cyclic adenosine monophosphate, guanosine monophos- phate, glucose monophosphate, fructose monophosphate, and phytic acid were separated and selectively detected. In order to increase the sensitivity, on-line preconcentration with a microprecolumn inserted in the rotorof a Valcovalve has been applied. Preliminary results have shown that an injection volume of at least 500 1-11 water containing organophosphorus acids at a 5–50 ng/rnl concentration level is possible.     

Comparison of sample preparation methods for analysis of isoflavones in foodstuffs

Due to the lack of one universally applicable and commonly used reference method, sample preparation in isoflavone (IF) analysis has been performed by many different methods which renders comparison and quality assessment of published IF contents in foodstuffs difficult.In the present work, the impact of different experimental parameters on the IF concentrations determined in soybeans, tofu, soy drink and textured vegetable protein by different extraction and hydrolysis methods was assessed and IF contents obtained by optimized orthogonal methods were compared. Chromatographic analysis was performed by HPLC-UV-ESI-MS. If possible sources of error - which are also pointed out in this work - are avoided, IF contents obtained by extraction, acid-, base- and enzymatic hydrolysis are similar. However, these sample preparation methods differ in the amount of time, standard compounds and instruments required, ruggedness, and in their applicability to analysis of complex composite samples containing soy as minor ingredient. Enzymatic hydrolysis with Helix pomatia juice after extraction by sonication with first 50, then 80% aqueous acetonitrile in the presence of zinc sulfate heptahydrate and after adjustment to ≤10% organic solvent turned out to be the method of choice if only aglucone equivalent contents are required. The advantages of this method are short chromatographic run times, smallest danger of coelution, lowest achievable limits of quantitation and therefore best suitability for work-up of complex composite samples and that only aglucone standards are needed for quantitation.