Full-scan Mass Spectra Research Articles

Bioinert UHPLC system improves sensitivity and peak shapes for ionic metabolites.

The analysis of ionic compounds by liquid chromatography is challenging due to the interaction of analytes with the metal surface of the instrument and the column, leading to poor peak shape and decreased sensitivity. The use of bioinert materials in the chromatographic system minimizes these unrequired interactions. In this work, the ultrahigh-performance liquid chromatography (UHPLC) with bioinert components was connected to a high-resolution mass spectrometer to develop a method for untargeted metabolomic analysis. 81 standards of metabolites were used for the development and optimization of the method. In comparison to the conventional chromatographic system, the application of bioinert technology resulted in significantly improved peak shapes and increased sensitivity, especially for metabolites containing phosphate groups. The calibration curves were constructed for the evaluation of the method performance, showing a wide dynamic range, low limit of detection, and linear regression coefficients higher than 0.99 for all standards. The optimized method was applied to the analysis of NIST SRM 1950 human plasma, which allowed the detection of 156 metabolites and polar lipids based on the combination of mass accuracy in the full-scan mass spectra in both polarity modes, characteristic fragment ions in MS/MS, and logical chromatographic behavior leading to the high confidence level of annotation/identification. We have demonstrated an improvement in the peak shapes and sensitivity of ionic metabolites using bioinert technology, which indicates the potential for the analysis of other ionic compounds, e.g., molecules containing phosphate groups.

A SIFT Study of Reactions of Positive and Negative Ions With Polyfluoroalkyl (PFAS) Molecules in Dry and Humid Nitrogen at 393 K.

Data are required for SIFT-MS analysis of perfluoroalkyl and polyfluoroalkyl substances (PFAS), which are persistent in the environment and cause adverse health effects. Specifically, the rate coefficients and product ion branching ratios of the reactions of H3O+, NO+, O2 +•, O-•, OH-, O2 -•, NO2 - and NO3 - with PFAS vapours are needed. The dual polarity SIFT-MS instrument (Voice200) was used to generate these eight reagent ions and inject them into the flow tube with N2 carrier gas at a temperature of 393 K. Vapours of pentafluoropropionic acid, heptafluorobutyric acid, nonafluoro-1-hexanol, perfluoro-2-methyl-2-pentene, perfluorohexanoic acid, perfluoro(2-methyl-3-oxahexanoic) acid, tridecafluoro-1-octanol and nonafluorobutane-1-sulfonic acid were introduced in dry and humid air. Full-scan mass spectra were collected for all reagents at variable PFAS concentrations and analysed numerically. Rate coefficients were determined for 64 reactions, for which 55 positive and 71 negative product ions were identified. The branching ratios for the primary reaction channels were extracted from the data, and the secondary chemistry with H2O molecules was qualitatively assessed. The thermochemical data were calculated for the H3O+ reactions using density functional theory (DFT). An important observation is that secondary reactions with water molecules remove the positive product ions, making them unsuitable for practical SIFT-MS analysis of PFAS vapours. In contrast, most negative reaction product ions are not significantly affected by humidity and are thus preferred for the SIFT-MS analyses of PFAS substances in various gaseous matrices.

Quantitative data of up to thirty sterols in vegetable oils and fats

Sterols are known for a plethora of 250 different structures. Between 5 and 10% of them usually occur with varying abundance ratios (~ four orders of magnitude) and total amounts (0.4–1000 mg/100 g oil) in samples. Yet, quantitative data are mostly restricted to the few major sterols which are available as reference standards. Here, we developed a gas chromatography with mass spectrometry method operated in selected ion monitoring mode (GC/MS-SIM) that enabled the quantitation of 30 (silylated) sterols although only ten were available as reference standards. This could be obtained by studying the full-scan mass spectra of these ten sterol standards and 20 additional sterols measured in seven oils. In the next step, sterols were assigned to different groups. Values for quantification were then selected on the premise that response factors were constant within a sterol group. The deviation of the response factors within one sterol group was frequently below ± 10% and otherwise about ± 11–12%. Using mean response factors for all sterols, the novel GC/MS-SIM quantification method was superior to GC/FID which was exemplarily applied to two oils. Between eight and 21 of the 30 studied sterols and pentacyclic triterpenols were detected and quantified in 18 vegetable oils and two vegetable fats. The much higher number of sterols that could be quantified resulted in higher sterol amounts and the method and data may be useful for food authentication.

Drosophilin A methyl ether (DAME) and other chlorinated dimethoxybenzenes in fungi and forest litter from Sweden

Fungi and substrates undergoing fungal decomposition were collected from forests in northern and southern Sweden and analyzed for chlorinated dimethoxybenzenes (DMBs). Specimens were fungi fruiting bodies, rotting wood, forest litter and underlying humus. Targeted compounds were DAME (1,2,4,5-tetrachloro-3,6-DMB) and related fungal secondary metabolites. A screening procedure was developed which involved soaking the specimens in ethyl acetate followed by analysis by capillary gas chromatography – mass spectrometry with mass selective detection (GC-MSD). DAME was the most frequently found (62% of 47 specimens) and often the most abundant target compound, with range and mean ± SD concentrations of <0.0017–3.81 and 0.21 ± 0.63 mg kg−1 ww. Based on log-log correlations of partition coefficients of hydrophobic compounds between fungal biomass/water (KD) and octanol/water (KOW), five species of fungi are suggested to produce DAME de novo versus bioaccumulation from forest runoff water. Full-scan mass spectra of some high-concentration specimens indicated the presence of a Cl2DMB and a Cl3DMB, which could not be identified further due to lack of standards, and drosophilin A (DA = 2,3,5,6-tetrachloro-4-methoxyphenol), the precursor to DAME. Tetrachloroveratrole (TeCV = 1,2,3,4-tetrachloro-5,6-DMB) was found in only a few specimens. This study supports our hypothesis of fungi as a source of DAME in terrestrial runoff and indicates that other chlorinated secondary metabolites are present. DAME is widely distributed globally, and it would be good to have a better understanding of its sources and pathways as a marker of terrestrial organochlorines and their availability for bioaccumulation.

Comprehensive micropollutant characterization of wastewater during Covid-19 crisis in 2020: Suspect screening and environmental risk prioritization strategy

Micropollutants monitoring in wastewater can serve as a picture of what is consuming society and how it can impact the aquatic environment. In this work, a suspect screening approach was used to detect the known and unknown contaminants in wastewater samples collected from two wastewater treatment plants (WWTPs) located in the Basque Country (Crispijana in Alava, and Galindo in Vizcaya) during two weekly sampling campaigns, which included the months from April to July 2020, part of the confinement period caused by COVID-19. To that aim, high-resolution mass spectrometry was used to collect full-scan data-dependent tandem mass spectra from the water samples using a suspect database containing >40,000 chemical substances. The presence of > 80 contaminants was confirmed (level 1) and quantified in both WWTP samples, while at least 47 compounds were tentatively identified (2a). Among the contaminants of concern, an increase in the occurrence of some compounds used for COVID-19 disease treatment, such as lopinavir and hydroxychloroquine, was observed during the lockdown. A prioritization strategy for environmental risk assessment was carried out considering only the compounds quantified in the effluents of Crispijana and Galindo WWTPs. The compounds were scored based on the removal efficiency, estimated persistency, bioconcentration factor, mobility, toxicity potential and frequency of detection in the samples. With this approach, 33 compounds (e.g. amantadine, clozapine or lopinavir) were found to be considered key contaminants in the analyzed samples based on their concentration, occurrence and potential toxicity. Additionally, antimicrobial (RQ-AR) and antiviral (EDRP) risk of certain compounds was evaluated, where ciprofloxacin and fluconazole represented medium risk for antibiotic resistance (1 > RQ-AR > 0.1) in the aquatic ecosystems. Regarding mixture toxicity, the computed sum of toxic unit values of the different effluents (> 1) suggest that interactions between the compounds need to be considered for future environmental risk assessments.

Analysis of the Fragmentation Pathways for the Collision-Induced Dissociation of Protonated Cyclophosphamide: A Mass Spectrometry and Quantum Mechanical Study.

Cyclophosphamide is a well-known anticancer agent acting by means of DNA alkylation. Associated with its tumor selectivity, it also possesses a wide spectrum of toxicities. As the requirement of metabolic activation before cyclophosphamide exerts either its therapeutic or toxic effects is well recognized, research aiming at elucidating the pathways that lead to the activation of this drug is of key importance. This has created the necessity for developing an effective analytical method for detecting cyclophosphamide and its breakdown products. In this paper, an Acquity TQ tandem quadrupole mass spectrometer equipped with electrospray ionization in positive-ion mode was employed for detecting cyclophosphamide in its protonated form. The full-scan mass spectrum of cyclophosphamide shows two ion clusters displaying the characteristic isotopic pattern of two chlorine atoms and assigned as sodiated cyclophosphamide, [CP + Na]+, and protonated cyclophosphamide, [CP + H]+ or PCP. With the aid of quantum mechanical DFT calculation, free energy differences in the gas phase among PCP protomers were computed with respect to the most stable protomer being protonated on the 2-oxide oxygen of the 1,3,2-oxazaphosphorine-2-oxide ring. In addition, the interconversion mechanisms among the different protomers were also proposed by intercepting the corresponding transition states in the gas phase. Collision-induced dissociation (CID) of PCP generated six characteristic product ions. Fragmentation mechanisms were proposed and supported by computation. The calculated energy barriers for all of the located transition states were found to be accessible under the reported experimental conditions.

Rapid screening procedures for a variety of complex forensic samples using laser diode thermal desorption (LDTD) coupled to different mass spectrometers.

The applications shared in this paper demonstrate the wide variety of samples that can be analyzed when Laser Diode Thermal Desorption (LDTD) is interfaced with a high-resolution mass spectrometer and show the speed at which high quality data can be generated from complex matrices. Samples are solvent extracted and spotted in a 96-well plate. In the case of biological fluids, hydrolysis followed by solid-phase extraction is required. The solvent in the 96-well plate is evaporated followed by mass spectrometric (MS) analysis with atmospheric pressure chemical ionization. Where applicable, the instrument is operated in data-dependent mode, with a full-scan mass spectrum followed by MS/MS spectra of the top 10 ions with a total runtime of 0.4 min. Four applications (MAAQ and Tear Gas, twelve rodenticides, seven explosives, and 40 drugs of abuse) are reported in this paper. MAAQ, tear gas, and rodenticides were identified by full-scan, followed by MS/MS experiments at levels of 125 μg/L, 125 μg/L, and 500 μg/L, respectively. Explosives were all identified at 102 μg/L by full-scan experiments. The drugs of abuse were identified by multiple reaction monitoring (MRM) experiments at defined cutoff levels from 2 to 1000 μg/L. Interfacing LDTD with a mass spectrometer allows for rapid screening of a wide range of samples, with either minimal or complex sample preparation. Using a high-resolution mass spectrometer with the combination to perform full-scan and MS/MS experiments adds a high level of specificity.

Chemical synthesis and characterization of single-chain C18-chloroparaffin materials with defined degrees of chlorination

Technical chlorinated paraffins (CPs) are produced via radical chlorination of n-alkane feedstocks with different carbon chain-lengths (∼C10–C30). Short-chain CPs (SCCPs, C10–C13) are classified as persistent organic pollutants (POPs) under the Stockholm Convention. This regulation has induced a shift to use longer-chain CPs as substitutes. Consequently, medium-chain (MCCPs, C14–C17) and long-chain (LCCPs, C>17) CPs have become dominant homologues in recent environmental samples. However, no suitable LCCP-standard materials are available. Herein, we report on the chemical synthesis of single-chain C18-CP-materials, starting with a pure n-alkane and sulfuryl chloride (SO2Cl2). Fractionation of the crude product by normal-phase liquid-chromatography and pooling of suitable fractions yielded in four C18-CP-materials with different chlorination degrees (mCl,EA = 39–52%). In addition, polar side-products, tentatively identified as sulfite-, sulfate- and bis-sulfate-diesters, were separated from CPs. The new single-chain materials were characterized by LC-MS, 1H-NMR and EA. LC-MS provided Relative retention times for different C18-CP homologues and side-products. Mathematical deconvolution of full-scan mass spectra revealed the presence of chloroparaffins (57–93%) and chloroolefins (COs, 7–26%) in the four single-chain C18-CP-materials. Homologue distributions and chlorination degrees were deduced for CPs and COs. 1H-NMR revealed chemical shift ranges of mono-chlorinated (δ = 3.2–5.3 ppm) and non-chlorinated (δ = 1.0–3.2 ppm) hydrocarbon moieties. The synthesized C18-single-chain standard materials and respective spectroscopic data are useful to identify and quantify LCCPs in various materials and environmental samples. CP- and CO-distributions resemble the ones of existing SCCP and MCCP reference materials and technical mixtures. Furthermore, these materials now allow specific studies on the environmental fate and the transformation of long-chain chloroparaffins and chloroolefins.

Metabolic profiles of human brain parenchyma and glioma for rapid tissue diagnosis by targeted desorption electrospray ionization mass spectrometry.

Desorption electrospray ionization mass spectrometry (DESI-MS) is well suited for intraoperative tissue analysis since it requires little sample preparation and offers rapid and sensitive molecular diagnostics. Currently, intraoperative assessment of the tumor cell percentage of glioma biopsies can be made by measuring a single metabolite, N-acetylaspartate (NAA). The inclusion of additional biomarkers will likely improve the accuracy when distinguishing brain parenchyma from glioma by DESI-MS. To explore this possibility, mass spectra were recorded for extracts from 32 unmodified human brain samples with known pathology. Statistical analysis of data obtained from full-scan and multiple reaction monitoring (MRM) profiles identified discriminatory metabolites, namely gamma-aminobutyric acid (GABA), creatine, glutamic acid, carnitine, and hexane-1,2,3,4,5,6-hexol (abbreviated as hexol), as well as the established biomarker NAA. Brain parenchyma was readily differentiated from glioma based on these metabolites as measured both in full-scan mass spectra and by the intensities of their characteristic MRM transitions. New DESI-MS methods (5min acquisition using full scans and MS/MS), developed to measure ion abundance ratios among these metabolites, were tested using smears of 29 brain samples. Ion abundance ratios based on signals for GABA, creatine, carnitine, and hexol all had sensitivities > 90%, specificities > 80%, and accuracies > 85%. Prospectively, the implementation of diagnostic ion abundance ratios should strengthen the discriminatory power of individual biomarkers and enhance method robustness against signal fluctuations, resulting in an improved DESI-MS method of glioma diagnosis.

Retention dependences support highly confident identification of lipid species in human plasma by reversed-phase UHPLC/MS.

Reversed-phase ultrahigh-performance liquid chromatography-mass spectrometry (RP-UHPLC/MS) method was developed with the aim to unambiguously identify a large number of lipid species from multiple lipid classes in human plasma. The optimized RP-UHPLC/MS method employed the C18 column with sub-2-μm particles with the total run time of 25 min. The chromatographic resolution was investigated with 42 standards from 18 lipid classes. The UHPLC system was coupled to high-resolution quadrupole-time-of-flight (QTOF) mass analyzer using electrospray ionization (ESI) measuring full-scan and tandem mass spectra (MS/MS) in positive- and negative-ion modes with high mass accuracy. Our identification approach was based on m/z values measured with mass accuracy within 5 ppm tolerance in the full-scan mode, characteristic fragment ions in MS/MS, and regularity in chromatographic retention dependences for individual lipid species, which provides the highest level of confidence for reported identifications of lipid species including regioisomeric and other isobaric forms. The graphs of dependences of retention times on the carbon number or on the number of double bond(s) in fatty acyl chains were constructed to support the identification of lipid species in homologous lipid series. Our list of identified lipid species is also compared with previous publications investigating human blood samples by various MS-based approaches. In total, we have reported more than 500 lipid species representing 26 polar and nonpolar lipid classes detected in NIST Standard reference material 1950 human plasma.

Urine metabolomics analysis based on ultra performance liquid chromatography-high resolution mass spectrometry combined with osmolality calibration sample concentration variability

尿液是代谢组学研究中主要关注的体液样本之一。尿液样本中的代谢物浓度受饮食、疾病等因素影响变异较大,这极大阻碍了高质量组学数据的获取和可靠生物标志物的鉴定。研究为克服尿液样本的浓度变异性,在原始数据采集前,根据样本渗透压的大小,针对性地调整进样量或者稀释样本,从而确保代谢组学分析样本的渗透压与进样量的乘积相当,再经超高效液相色谱-高分辨质谱技术(UPLC-HRMS)分析,采用总离子丰度或总有用峰面积(MSTUS)对数据集进行归一化处理。研究利用临床样本及其梯度稀释的溶液,对该方法与现有研究普遍使用的方法进行了比较,随后通过先天性肾积水患者及健康志愿者的尿液样本做了进一步的方法学验证。数据集经校正后,峰面积RSD<30%的提取峰数量增加,主成分分析结果较校正前有更高的组内聚集和组间分群效应,正交偏最小二乘判别分析的统计模型更不易过拟合。与肌酐比较,渗透压值与质谱信号间呈现了更好的线性关系。以上结果表明,数据采集前通过样本渗透压进行校正,能有效消除因样本本身代谢物浓度变化引起的组内差异,提高方法的重复性和统计模型的可靠度。以渗透压为基准的校正策略,比肌酐校正法适用范围更广,结果也更准确。研究可对后续各类来源的尿液代谢组学研究提供数据归一化的指导和参考。

Metal-ion-assisted structural and anomeric analysis of Amadori compounds by electrospray ionization mass spectrometry.

The Maillard reaction plays an important role in food, physiology and traditional Chinese medicine, and its primary reaction products are formed through Amadori rearrangement by reducing sugars and amino acids. The analysis of the characteristic fragmentation and of the glycosidic bond configuration of Amadori compounds will promote their fast discovery and identification by mass spectrometry. Four Amadori compounds that reduce disaccharides and proline/tryptophan were used to investigate the fragmentation mechanisms via tandem mass spectrometry (MS/MS) with different alkali metal ion adducts. Cu2+ could be used to distinguish glycosidic bond configurations of the reducing disaccharides in the full-scan mass spectra. Quantum calculations were also conducted for a single Amadori compound with Cu2+ for analysis of the most optimized configurations and binding energies of metal complexes. MS/MS analysis of Amadori-alkali metal complexes revealed that the radius of the alkali metal ions had profound effects on the degree of fragmentation of such compounds, among which lithium-cationized ions produced the most extensive fragmentation. Amadori compounds with different glycosidic bonds formed differently proportioned metal complexes with Cu2+ , and the complexity of the copper complexes containing tryptophan moieties was higher than that of those containing proline moieties in the mass spectra. Quantum calculations showed that Amadori compounds with β-configurations can form more binding sites with Cu2+ than those with α-configurations, thus making the metal complex with a single ligand more stable. In addition, the chelation of tryptophan with copper ions increased the coordination binding energy, which showed that α-configured Amadori compounds were readily able to form multi-ligand copper complexes. Metal-ion-assisted analysis provides crucial information for structural and anomeric analysis of Amadori compounds by electrospray ionization mass spectrometry. Elucidation of binding sites and binding energies by quantum calculations has significantly improved the knowledge of metal complexes in the gas phase and provides background information for determining the glycosidic configuration of Amadori isomers.

Determination of sildenafil citrate adulterated in a dietary supplement capsule by LC/MS/MS

A liquid chromatography/tandem mass spectrometry (LC/MS/MS) method was used to identify and quantify an adulterated substance, in a dietary supplement capsule. The capsule is claimed to be an extract of animal organs and traditional Chinese herbs and indicated for enhancing sexual activity. At first, the sample was extracted with 50% v/v methanol in water and the extract was injected directly into the LC/MS/MS with no separation and a full scan of positive ion eletrospray (ESI +) analysis was performed. The full scan mass spectrum was compared with the NLFD3/LM library created by Division 3rd of National Laboratories of Foods and Drugs for LC/MS/MS analysis. A sildenafil specific mass spectrum was matched indicating that the capsule may have been adulterated with sildenafil, the active ingredient of VIAGRA®. Furthermore, sildenafil was confirmed by carrying out a daughter ion scan and a parent ion scan. The daughter ion spectrum was compared with the NLFD3/LM library. A negative ion eletrospray (ESI -) analysis was performed for confirming the citrate salt in the capsule. The results indicated that the adulterated ingredient was sildenafil citrate. For quantitative analysis of the sildenafil citrate, multiple reaction monitoring (MRM) was performed. Pirenzepine 2HCL was used as the internal standard. A series of standards solution and sample solution with internal standard were analyzed by LC/ MS/MS. The analytical condition is as following: column: Cosmosil 5C18-AR, 4.6 x 150 mm; mobile phase:methanol:acetonitrile: 1% acetic acid (25:17:58); capillary voltage: 3 kV;cone voltage: 80 V; collision energy: 25 eV; source temperature: 120°C; desolvation temperature: 350°C. The detection limit was 40 ng/mL. An excellent calibration curve of sildenafil citrate was obtained with the γ 2 correlation coefficient of 0.9977. The RSDs of interday and intraday were between 7.56∼7.75% and 9.09∼11.25%, respectively. The analytical strategy and method is suitable for identification and measurement of sildenafil citrate adulterated in capsule. Amount of sildenafil citrate in the capsule was determined as 42.8 mg/capsule.

Quantification of Colistin in Plasma by Liquid Chromatography-Tandem Mass Spectrometry: Application to a Pharmacokinetic Study

Colistin is a polymixin antibiotic (polymixin E) that is produced by Bacillus colistinus bacteria. The aim of the present study was to develop and validate a method to quantify colistin levels in plasma using high performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) technique and then apply it in experimental animals (rats) to investigate the pharmacokinetic profile of colistin in this species. Polymyxin B was used as an internal standard (IS) and the quantitation was carried out using ESI + interface and employing multiple reaction monitoring (MRM) mode. A mobile phase consisting of acetonitrile:water:formic acid (30:70:0.1%; v/v/v) was employed and Zorbax eclipse plus C18 (1.8 µm, 2.1 mm i.d. x 50 mm) was the optimal column for this method and utilized at a flow rate of 0.2 mL/min. The full scan mass spectra of precursor/product ions of colistin A were at m/z 585.5 > 100.8, for colistin B at m/z 578.8 > 101 and for the IS at m/z 602.8 > 101. The lower limit of quantification (LLOQ) was 0.5 µg/mL. The method demonstrated acceptable intra-run and inter-run precision and accuracy for both colistin A and colistin B. Colistin was stable when assessed for long-term stability, freeze-thaw stability and autosampler stability. However, it was not stable when stored at room temperature. The matrix effect evaluation showed minimal or no effect. Incurred sample reanalysis findings were within acceptable ranges (<20% of the nominal concentration). The pharmacokinetic parameters of colistin were investigated in rats using the present method. The developed method for colistin demonstrates that it is rapid, sensitive, specific, accurate, precise, and reliable.

Quantitative analysis of pesticide residues in tea by gas chromatography–tandem mass spectrometry with atmospheric pressure chemical ionization

In this study, gas chromatography–tandem mass spectrometry (GC–MS/MS) using an atmospheric pressure chemical ionization (APCI) source was applied for the quantitative analysis of pesticide residues in tea. To determine the optimum ionization conditions for multiresidue analysis, the full-scan mass spectra and peak intensities of pesticides were compared in the presence and absence of water as a modifier. When water was added as a modifier in the ion source, most of the target compounds formed [M+H]+ ions and exhibited enhanced intensities. However, compounds consisting of only carbon, hydrogen, and chlorine, such as aldrin, γ-hexachlorocyclohexane, and p,p′-dichlorodiphenyldichloroethane, typically formed M+· or fragment ions, whose intensities were significantly decreased by the addition of water. GC–MS/MS methods using APCI (without modifier addition) and electron ionization (EI) were validated for 16 pesticides in tea at spiking levels of 0.01 and 0.1 mg/kg. Unlike EI, signal suppression was observed for most compounds at a spiking level of 0.01 mg/kg using APCI; however, dilution of the samples minimized this effect. Using APCI, the trueness of the target compounds ranged from 77% to 121% at both spiking levels, except for pyrethrins owing to matrix effects, with relative standard deviations of less than 14%. For most compounds, these results were comparable with those obtained using EI. However, because the use of APCI limited fragmentation, this ionization technique offered significantly higher sensitivity and specificity than EI. Using APCI, linear calibration curves with coefficients of determination greater than 0.998 were obtained in the range of 0.0005–0.5 μg/mL for all compounds. These findings indicated that GC–MS/MS with APCI is applicable for the routine monitoring of pesticide residues, even in complex samples such as tea.

Accurate identification of dimers from α-pinene oxidation using high-resolution collision-induced dissociation mass spectrometry.

Interest in mass spectrometry of highly oxidized dimers from α-pinene oxidation has increased in the atmospheric chemistry field. Here, we apply high-resolution collision-induced dissociation mass spectrometry (HR-CID-MS) with an atmospheric pressure ionization source to investigate in detail how α-pinene-derived dimers are detected and identified by MS. The resulting HR-CID spectra and specific fragmentation patterns suggest that a large fraction of dimer ions detected in full-scan mass spectra can be hydrogen-bonded artifact clusters and the residual small fraction includes covalently bonded actual dimers. We also show how individual fractions of the artifact clusters and actual dimers are calculated using the HR-CID spectra.

Unexpected cytosine-AuCl4- interaction under electrospray ionization mass spectrometry conditions-Formation of cytosine-Au(I) complexes.

The interaction of cytosine with AuCl4-, under electrospray ionization mass spectrometric conditions, is discussed. On the basis of respective full scan mass spectra and product ion spectra, obtained in positive and negative ion mode, it has been deduced that cytosine is very prone to form Au(I)-containing complexes. The complexes may be formed in the gas phase by decomposition of Au(III)-containing complexes and also in the electrospray ionization source as a result of the occurrence of redox process. It has also been found that the interaction of cytosine with Au+ is stronger than that with Cu+ or Ag+, although taking into account the electrostatic attraction, it is not expected.

Untargeted Metabolomic Profile for the Detection of Prostate Carcinoma-Preliminary Results from PARAFAC2 and PLS-DA Models.

Prostate-specific antigen (PSA) is the main biomarker for the screening of prostate cancer (PCa), which has a high sensibility (higher than 80%) that is negatively offset by its poor specificity (only 30%, with the European cut-off of 4 ng/mL). This generates a large number of useless biopsies, involving both risks for the patients and costs for the national healthcare systems. Consequently, efforts were recently made to discover new biomarkers useful for PCa screening, including our proposal of interpreting a multi-parametric urinary steroidal profile with multivariate statistics. This approach has been expanded to investigate new alleged biomarkers by the application of untargeted urinary metabolomics. Urine samples from 91 patients (43 affected by PCa; 48 by benign hyperplasia) were deconjugated, extracted in both basic and acidic conditions, derivatized with different reagents, and analyzed with different gas chromatographic columns. Three-dimensional data were obtained from full-scan electron impact mass spectra. The PARADISe software, coupled with NIST libraries, was employed for the computation of PARAFAC2 models, the extraction of the significative components (alleged biomarkers), and the generation of a semiquantitative dataset. After variables selection, a partial least squares–discriminant analysis classification model was built, yielding promising performances. The selected biomarkers need further validation, possibly involving, yet again, a targeted approach.

Localization of malonyl and acetyl on substituted saikosaponins according to the full-scan mass spectra and the fragmentation of sodium-adduct ions in the positive mode.

Discriminating between aglycone-substituted and saccharide-substituted saikosaponins by liquid chromatography/tandem mass spectrometry (LC/MSn ) is a long-standing issue that is still to be resolved. It is necessary to characterize the two types of substituted saikosaponins taking into consideration the potential significant difference in their bioactivity. Taking the substituents malonyl and acetyl as examples, we developed a MS strategy to discriminate between the aglycone-substituted and saccharide-substituted saikosaponins through comparing their Y0 - nH2 O (n = 1-2) ions from the protonated molecules in the full-scan mass spectra and their B ions in the MS2 spectra of sodium-adduct molecules in the positive mode. The deprotonated molecules of the aglycone-substituted saikosaponins presented similar fragmentation patterns to those of saccharide-substituted ones in the negative mode, which could not discriminate whether the substitutes were located on the aglycone or the saccharide. In contrast, the Y0 - nH2 O (n = 1-2) ions containing or no substituent were observed respectively in the mass fragmentation of the protonated molecules of aglycone-substituted or saccharide-substituted saikosaponins in the positive mode. In addition, the B ions containing or no substituent were observed respectively in the mass fragmentation of the sodium-adduct molecules of the saccharide-substituted or aglycone-substituted saikosaponins in the positive mode. Two aglycone-malonylated saikosaponins were reported for the first time. Whether the substituents were located on the aglycone or the saccharide could be determined according to the Y0 - nH2 O (n = 1-2) ions from the protonated molecules in the full-scan mass spectra and the B ions in the MS2 spectra of sodium-adduct molecules in the positive mode. Our results have updated the mass fragmentation patterns of substituted saikosaponins, which is helpful for the quality control of pharmaceutical preparations containing saikosaponins. More importantly, this MS strategy should be able to be extended to characterize other substituted saponins of bioactive significance in future studies.

Biotransformation of short-chain chlorinated paraffins (SCCPs) with LinA2: A HCH and HBCD converting bacterial dehydrohalogenase

Short-chain chlorinated paraffins (SCCPs) are polyhalogenated hydrocarbons as are hexachlorocyclohexanes (HCHs) and hexabromocyclododecanes (HBCDs). They all have been classified as persistent organic pollutants (POPs) under the UN Stockholm Convention. Per se such compounds are transformed slowly in the environment, transported over long distances and accumulate in biota. Several Sphingomonadacea strains isolated from HCH dump sites have evolved to express enzymes that can transform HCHs and HBCDs. We hypothesized that LinA2, a dehydrohalogenase expressed in such bacteria, may also transform CPs to chlorinated olefins (COs). Three mixtures of penta- to deca-chlorinated undecanes (C11), dodecanes (C12) and tridecanes (C13) were exposed to LinA2. High-resolution full-scan mass spectra (R∼8′000) of CPs and COs were obtained applying a soft ionization method, enhancing chloride-adduct [M+Cl]- formation. A mathematical deconvolution procedure was used to separate interfering spectra to verify that LinA2 indeed catalyzed the conversion of CPs to COs. About 20–40% of the material was transformed in 24 h, about 50–70% was converted in 200 h. A bimodal first-order kinetic model could describe transformations of reactive and persistent CPs. Under the given conditions reactive CPs (τ1/2 = 1.4–6.9 h) were converted 30 to 190-times faster than the persistent ones (τ1/2 = 150–260 h). Proportions of persistent isomers (pp) varied from 60 to 80%. Lower chlorinated homologues contained higher proportions of persistent isomers. In conclusion, SCCP mixtures contain both, material that is readily converted by LinA2, and persistent material that is not or only slowly transformed.