Tetrahydrogestrinone: discovery, synthesis, and detection in urine

Tetrahydrogestrinone (18a-homo-pregna-4,9,11-trien-17beta-ol-3-one or THG) was identified in the residue of a spent syringe that had allegedly contained an anabolic steroid undetectable by sport doping control urine tests. THG was synthesized by hydrogenation of gestrinone and characterized by mass spectrometry and NMR spectroscopy. We developed and evaluated sensitive and specific methods for rapid screening of urine samples by liquid chromatography/tandem mass spectrometry (LC/MS/MS) of underivatized THG (using transitions m/z 313 to 241 and 313 to 159) and gas chromatography/high-resolution mass spectrometry (GC/HRMS) analysis of the combination trimethylsilyl ether-oxime derivative of THG (using fragments m/z 240.14, 254.15, 267.16, and 294.19). A baboon administration study showed that THG is excreted in urine.     

Determination of the metabolites of gestrinone in human urine by high performance liquid chromatography, liquid chromatography/mass spectrometry and gas chromatography/mass spectrometry

Gestrinone was studied by high performance liquid chromatography (HPLC) for screening and by gas chromatography/mass spectrometry (GC/MS) for confirmation. When the chromatograms of blank, spiked urine and dosed urine were compared by HPLC, two unknown metabolites were found and these were excreted as the conjugated forms. Metabolites 1 and 2 were tested by LC/MS and LC/MS/MS and both had parent ions at m/z 325. The fragment ion of metabolite 1 was at m/z 263 and ions for metabolite 2 were m/z 307 [MH - H(2)O](+), 289, 279 and 241. LC/MS/MS of m/z 263 as the parent ion of metabolite 1 gave fragment ions at m/z 245 and 217, which were assumed to be [263 - H(2)O](+) and [235 - H(2)O](+), respectively. The trimethylsilyl (TMS)-enol-TMS ether derivative of gestrinone displayed three peaks in its GC/MS chromatogram, formed by tautomerism.     

Exact mass measurement of product ions for the structural confirmation and identification of unknown compounds using a quadrupole time-of-flight spectrometer: a simplified approach using combined tandem mass spectrometric functions

This article describes a simple method to perform lock mass corrected accurate mass measurements in tandem mass spectrometry (MS/MS) with a quadrupole time-of-flight (Q-TOF) mass spectrometer. The experimental approach consists of using the protonated molecule of a known compound, which is measured in a MS/MS function using low collision energy (no fragmentation), as mass calibrator. The unknown compound is acquired in MS/MS mode albeit using high collision energy. After the acquisition, the two MS/MS spectra of unknown and mass calibrator are combined, and the fragments of the unknown are lock mass corrected by using the protonated molecule of the mass calibrator. To prove this concept, 10 compounds were analyzed using this approach, the fragments interpreted and, where possible, related to structural data available in the literature. All the unequivocally assigned fragments were accurately mass measured with mass errors within appropriate limits, i.e. for m/z values <200 with a mass tolerance of 3 mDa while for m/z > 200 the mass tolerance is expressed as 10 ppm.     

Ion-trap tandem mass spectrometry-based analytical methodology for the determination of polychlorinated biphenyls in fish and shellfish. Performance comparison against electron-capture detection and high-resolution mass spectrometry detection

Optimization of the Varian Saturn 2200 ion-trap tandem mass spectrometry (IT-MS/MS) system and comparison of its data quality with two other detection methods [electron-capture detection (ECD) and high-resolution mass spectrometry (HRMS)] was pursued by measuring polychlorinated biphenyls (PCBs) levels in fish and shellfish samples. IT-MS/MS methodology provided limits of detection (LOD) comparable to those obtained by ECD but superior specificity for the detection of a selected number of 39 PCB native congeners and 9 (13)C-labelled PCB standards. The method detection limits (MDLs) established for IT-MS/MS ranged between 1.0 and 5.0 pg/g on a wet weight basis while those obtained by ECD and HRMS were 1.0-4.0 pg/g and 0.1-2.0 pg/g, respectively. Overall, the results obtained in the study demonstrate that gas chromatography (GC) combined with IT-MS/MS provide higher data quality than those achievable by GC-ECD. For this particular set of target analytes the specificity achievable with IT-MS/MS was comparable to that obtained by HRMS and both techniques provided comparable data in terms of accuracy and precision.     

Liquid chromatography/negative ion atmospheric pressure photoionization mass spectrometry: a highly sensitive method for the analysis of organic explosives

Gas chromatography/mass spectrometry (GC/MS) is applied to the analysis of volatile and thermally stable compounds, while liquid chromatography/atmospheric pressure chemical ionization mass spectrometry (LC/APCI‐MS) and liquid chromatography/electrospray ionization mass spectrometry (LC/ESI‐MS) are preferred for the analysis of compounds with solution acid‐base chemistry. Because organic explosives are compounds with low polarity and some of them are thermally labile, they have not been very well analyzed by GC/MS, LC/APCI‐MS and LC/ESI‐MS. Herein, we demonstrate liquid chromatography/negative ion atmospheric pressure photoionization mass spectrometry (LC/NI‐APPI‐MS) as a novel and highly sensitive method for their analysis. Using LC/NI‐APPI‐MS, limits of quantification (LOQs) of nitroaromatics and nitramines down to the middle pg range have been achieved in full MS scan mode, which are approximately one order to two orders magnitude lower than those previously reported using GC/MS or LC/APCI‐MS. The calibration dynamic ranges achieved by LC/NI‐APPI‐MS are also wider than those using GC/MS and LC/APCI‐MS. The reproducibility of LC/NI‐APPI‐MS is also very reliable, with the intraday and interday variabilities by coefficient of variation (CV) of 0.2–3.4% and 0.6–1.9% for 2,4,6‐trinitrotoluene (2,4,6‐TNT). Copyright © 2008 John Wiley & Sons, Ltd.     

Mass spectrometric characterization of toremifene metabolites in human urine by liquid chromatography-tandem mass spectrometry with different scan modes

The metabolism and excretion of toremifene were investigated in one healthy male volunteer after a single oral administration of 120 mg toremifene citrate. Different liquid chromatographic/tandem mass spectrometric (LC/MS/MS) scanning techniques were carried out for the characterization of the metabolites in human urine for doping control purposes. The potential characteristic fragmentation pathways of toremifene and its major metabolites were presented. An approach for the metabolism study of toremifene and its analogs by liquid chromatography-tandem mass spectrometry was established. Five different LC/MS/MS scanning methods based on precursor ion scan (precursor ion scan of m/z 72.2, 58.2, 44.2, 45.2, 88.2 relative to five metabolic pathways) in positive ion mode were assessed to recognize the metabolites. Based on product ion scan and precursor ion scan techniques, the metabolites were proposed to be identified as 4-hydroxy-toremifene (m/z 422.4), 4'-hydroxy-toremifene (m/z 422.4), α-hydroxy-toremifene (m/z 422.4), 3,4-dihydroxy-toremifene (m/z 404.2), toremifene acid (m/z 402.2), 3-hydroxy-4-methoxy-toremifene (m/z 456.2), dihydroxy-dehydro-toremifene (m/z 440.2), 3,4-dihydroxy-toremifene (m/z 438.2), N-demethyl-4-hydroxy-toremifene (m/z 408.3), N-demethyl-3-hydroxy-4-methoxy-toremifene (m/z 438.3). In addition, a new metabolite with a protonated molecule at m/z 390.3 was detected in all urine samples. The compound was identified by LC/MS/MS as N-demethyl-4,4'-dihydroxy-tamoxifene. The results indicated that 3,4-dihydroxy-toremifene (m/z 404.2), toremifene acid (m/z 402.2) and N-demethyl-4,4'-dihydroxy-tamoxifene (m/z 390.3) were major metabolites in human urine.     

False‐positive liquid chromatography/tandem mass spectrometric confirmation of sebuthylazine residues using the identification points system according to EU directive 2002/657/EC due to a biogenic insecticide in tarragon

In pesticide residue analysis using liquid chromatography/tandem mass spectrometry (LC/MS/MS) the confirmation of a sebuthylazine finding in a tarragon (Artemisia dranunculus) sample was demonstrated to be false positive. A coeluting interfering matrix compound produced product ions in MS/MS analysis, perfectly corresponding to the multiple reaction monitoring (MRM) of two sebuthylazine transitions. Using the EU directive 2002/657/EC which regulates the confirmation of suspected positive findings would have resulted in a false‐positive finding. A third LC/MS/MS transition with a deviant ion ratio and a gas chromatography (GC)/MS/MS analysis revealed the false‐positive results. With optimized high resolving ultra‐performance liquid chromatography (UPLC) conditions it was possible to separate spiked sebuthylazine from the interfering matrix compound. Using its exact mass and isotope ratios from LC/time‐of‐flight (TOF) MS measurements, the compound was identified as nepellitorine, a – not surprising – endogenous alkamide in tarragon (Arthemisia dranunculus). False‐positive results, especially in heavy matrix samples such as herbs, can be dealt with by further confirmatory analysis, e.g. a third transition, GC analysis if possible or more advantageous by an orthogonal criterion like exact mass. Copyright © 2009 John Wiley & Sons, Ltd.     

Study on the photostability of guaiazulene by high-performance liquid chromatography/mass spectrometry and gas chromatography/mass spectrometry

The photostability of guaiazulene (1,4-dimethyl-7-isopropylazulene; GA), a natural azulenic compound used in cosmetic and health-care products, as well as in pharmaceutical preparations, was investigated in solution (methanol, ethanol, acetonitrile), by different techniques: gas chromatography/mass spectrometry (GC/MS) and high-performance liquid chromatography combined with atmospheric pressure chemical ionization mass spectrometry and UV detection (LC/APCI-MS and HPLC/UV). A solar simulator (xenon-arc lamp) was used as UV-A radiation source. The study involved: monitoring compound decomposition, identifying products of photodegradation (PPs), assessing the role of oxygen and evaluating the kinetics of the process. Minor PPs are volatile compounds and were characterized by GC/MS, while oligomeric polyoxygenated compounds, tentatively characterized on the basis of MS and MS/MS spectra, were found to be the main photoproducts. The photodegradation was found to be enhanced by the presence of oxygen; nevertheless, determination of the singlet oxygen quantum yield for GA gave a lower value than that for the reference standard Rose Bengal. The obtained results and the developed stability-indicating methods (GC/MS and LC/MS) are of interest for stability studies and/or quality control purposes of GA as raw material or cosmetic products.     

N-linked oligosaccharide analysis of rat brain Thy-1 by liquid chromatography with graphitized carbon column/ion trap-Fourier transform ion cyclotron resonance mass spectrometry in positive and negative ion modes

We have previously described the site-specific glycosylation analysis of rat brain Thy-1 by LC/multistage tandem mass spectrometry (MS(n)) using proteinase-digested Thy-1. In the present study, detailed structures of oligosaccharides released from Thy-1 were elucidated by mass spectrometric oligosaccharide profiling using LC/MS with a graphitized carbon column (GCC-LC/MS). First, using model oligosaccharides, we improved the oligosaccharide profiling by ion trap mass spectrometry (IT-MS) coupled with Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS). Sequential scanning of a full MS(1) scan with FT-ICR-MS followed by data-dependent MS(n) with IT-MS in positive ion mode, and a subsequent full MS(1) scan with FT-ICR-MS followed by data-dependent MS(n) with IT-MS in negative ion mode enabled the monosaccharide composition analysis as well as profiling and sequencing of both neutral and acidic oligosaccharides in a single analysis. The improved oligosaccharide profiling was applied to elucidation of N-linked oligosaccharides from Thy-1 isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It was demonstrated that Thy-1 possesses a significant variety of N-linked oligosaccharides, including Lewis a/x, Lewis b/y, and disialylated structure as a partial structure. Our method could be applicable to analysis of a small abundance of glycoproteins, and could become a powerful tool for glycoproteomics.     

Analysis of biologically active compounds in water by ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry

A new method based on ultra-performance liquid chromatography (UPLC) quadrupole time-of-flight mass spectrometry ((Q-ToF)-MS) was developed for the analysis of 32 biologically active compounds including anti-inflammatories, analgesics, lipid regulators, psychiatric drugs, anti-ulcer agents, antibiotics, beta-blockers and phytoestrogens. This new method allows chromatographic analysis in 14 min, with instrumental detection limits from 2 to 84 pg, and limits of quantification ranging from 0.1 to 15 ng/L in tap water, and from 2 to 300 ng/L in wastewater. The potential of liquid chromatography with triple quadrupole mass spectrometry (LC/QqQ-MS) was compared with that of UPLC/(Q-ToF)-MS for the analysis of biologically active compounds in water samples. LC/Q-ToF provides accurate mass information and a significantly higher mass resolution than quadrupole analyzers. The available mass resolution of ToF instruments diminishes the problem of isobaric interferences and helps the analysis of trace compounds in complex samples. In this work UPLC/Q-ToF chromatograms were recorded containing full scan spectral data. The m/z values of analytes were extracted from the total ion chromatogram (TIC) and the accurate masses of the compounds were obtained. In addition, to increase the selectivity of ToF measurements a narrow accurate mass interval (20 m m/z units mass window) was used to reconstruct the chromatographic traces. However, regarding quantitative performance in terms of dynamic range and limits of detection (LODs), typical LODs achieved by QqQ instruments operating in multiple-reaction monitoring (MRM) mode ranged from 1 to 50 ng/L in wastewater, and the linear response for QqQ instruments generally covers three orders of magnitude. This is an important advantage over ToF instruments and one of the reasons why QqQ instruments are widely used in quantitative environmental analysis.     

Discovering metabolites of post-harvest fungicides in citrus with liquid chromatography/time-of-flight mass spectrometry and ion trap tandem mass spectrometry

In this study, we benefit from the combination of liquid chromatography (LC)/time-of-flight (TOF) MS accurate mass measurements to generate elemental compositions of ions and LC/ion trap multiple MS (MSn) providing complementary structural information, which is useful for the elucidation of unknown organic compounds at trace levels in complex food extracts. We have applied this approach to investigate different citrus fruits extracts, and we have identified two post-harvest fungicides (imazalil and prochloraz), the main degradation product of imazalil ([M + H]+, m/z 257) and a non-previously reported prochloraz degradation product ([M + H]+, m/z 282). The database-mediated identification of the parent compounds was based on the generated elemental composition obtained from accurate mass measurements and additional qualitative information from the high resolution chlorine isotopic clusters of both the protonated molecules (imazalil, [M + H]+ 297.0556, <0.1 ppm error, 2-Cl; prochloraz, [M + H]+ 376.0381, 1.9 ppm error, 3-Cl) and their characteristic fragments ions (imazalil: m/z 255 and 159; prochloraz: m/z 308 and 266). The correlation between the structural information provided by ion trap MS/MS fragmentation pathways of the parent species and the TOF accurate mass elemental composition data of the degradation products were the key to elucidate the structures of the degradation products of both post-harvest fungicides. Finally, where standards were not available (prochloraz), further confirmation was obtained by synthesizing the proposed degradation product by acid hydrolysis of the parent standard and confirmation by LC/TOF-MS.     

Pharmaceutical trace analysis in aqueous environmental matrices by liquid chromatography–ion trap tandem mass spectrometry

An analytical method based on solid-phase extraction followed by liquid chromatography tandem mass spectrometry with an ion trap analyser was developed and validated for the quantification of a series of pharmaceutical compounds with distinct physical–chemical characteristics in estuarine water samples. Method detection limits were between 0.03 and 16.4 ng/L. The sensitivity and the accuracy obtained associated with the inherent confirmatory potential of ion trap tandem mass spectrometry (IT-MS/MS) validates its success as an environmental analysis tool. Two MS/MS transitions were used to confirm compound identity. Almost all pharmaceuticals were detected at ng/L level in at least one sampling site of the Douro River estuary, Portugal.     

Ultra-performance liquid chromatography/tandem mass spectrometric quantification of structurally diverse drug mixtures using an ESI-APCI multimode ionization source

We report in this paper an ultra-performance liquid chromatography/tandem mass spectrometric (UPLC(R)/MS/MS) method utilizing an ESI-APCI multimode ionization source to quantify structurally diverse analytes. Eight commercial drugs were used as test compounds. Each LC injection was completed in 1 min using a UPLC system coupled with MS/MS multiple reaction monitoring (MRM) detection. Results from three separate sets of experiments are reported. In the first set of experiments, the eight test compounds were analyzed as a single mixture. The mass spectrometer was switching rapidly among four ionization modes (ESI+, ESI-, APCI-, and APCI+) during an LC run. Approximately 8-10 data points were collected across each LC peak. This was insufficient for a quantitative analysis. In the second set of experiments, four compounds were analyzed as a single mixture. The mass spectrometer was switching rapidly among four ionization modes during an LC run. Approximately 15 data points were obtained for each LC peak. Quantification results were obtained with a limit of detection (LOD) as low as 0.01 ng/mL. For the third set of experiments, the eight test compounds were analyzed as a batch. During each LC injection, a single compound was analyzed. The mass spectrometer was detecting at a particular ionization mode during each LC injection. More than 20 data points were obtained for each LC peak. Quantification results were also obtained. This single-compound analytical method was applied to a microsomal stability test. Compared with a typical HPLC method currently used for the microsomal stability test, the injection-to-injection cycle time was reduced to 1.5 min (UPLC method) from 3.5 min (HPLC method). The microsome stability results were comparable with those obtained by traditional HPLC/MS/MS.     

Investigation of the electrochemical oxidation products of zotepine and their fragmentation using on-line electrochemistry/electrospray ionization mass spectrometry

When zotepine, an antipsychotic drug, was electrochemically oxidized using electrospray ionization mass spectrometry (ESI-MS) coupled with a microflow electrolytic cell, [M + 16 + H]+ (m/z 348), [M-H]+ (m/z 330) and [M-14 + H]+ (m/z 318) were observed as electrochemical oxidation product ions (M represents the zotepine molecule). Although a major fragment ion that was derived from the dimethyl aminoethyl moiety was observed only at m/z 72 in the collision-induced dissociation (CID) spectrum of zotepine, new fragments such as m/z 315 and 286 ions could be generated in the CID spectrum by combining electrochemical oxidation and CID. Since these fragments were relatively specific with high ion strength, it was thought that they would be useful for developing a sensitive LC-MS/MS assay. The S-oxide and N-demethylated products were detected by electrolysis assuring that a portion of P450 metabolites of zotepine could be mimicked by the electrochemistry/electrospray ionization mass spectrometry (EC/ESI-MS) system.     

Stable isotope dilution method for the determination of guanidinoacetic acid by gas chromatography/mass spectrometry

For more than 30 years, guanidinoacetic acid (GAA), together with other guanidino compounds, has been proposed as an important marker for renal failure, in kidney transplantation, and for renal metabolism, especially for the metabolic activity of the renal proximal tubules. Since the discovery of the first patient with guanidinoacetic acid methyltransferase deficiency in 1994 by St?ckler et al. (Pediatr. Res. 1994; 36: 409), GAA has become of great interest for all laboratories involved in the diagnosis of metabolic diseases. In the literature there are several methods described for the determination of GAA, ranging from ion-exchange chromatography with post-column derivatisation, enzymatic methods, gas chromatography/mass spectrometry (GC/MS), to liquid chromatography/atmospheric pressure chemical ionisation mass spectrometry (LC/APCI-MS). Here a stable isotope dilution method for quantitative and accurate determination of GAA in urine, plasma, and cerebrospinal fluid is described. GAA is converted to the bis(trifluoromethyl)pyrimidine di(tert-butyldimethylsilyl) derivative by stepwise derivatisation with hexafluoroacetylacetone and N-(tert-butyldimethylsilyl)-N-methyltrifluoroacetamide (MTBSTFA). Analysis can be performed using a standard benchtop GC/MS system. For quantitative GAA determination with 1,2-(13)C-GAA as internal standard, selected ion monitoring is performed using m/z 460/462, with m/z 432/433 and 375/376 as qualifiers.     

乳制品中那他霉素的超高效液相色谱-串联质谱法快速测定

建立了超高效液相色谱-电喷雾串联质谱(UPLC-MS/MS)快速检测乳制品中那他霉素的方法。样品用甲醇提取,以甲醇-水为流动相经反相色谱柱分离后,采用多反应监测(MRM)负离子模式检测,定性离子对为m/z663.6/421.1和m/z663.6/439.1,其中m/z663.6/421.1用于外标法定量。空白样品及其加标实验结果表明:特征离子相对强度比值稳定,无基质干扰,结合保留时间可实现准确的定性定量;方法定量下限为50.0μg/kg;乳制品加标量为50~500μg/kg时,平均回收率为80%~91%,相对标准偏差(n=6)为2.7%~5.2%。方法简单、灵敏、稳定,可满足乳制品中那他霉素的快速检测与确证需要。     

Certification of drugs of abuse in a human serum standard reference material: SRM 1959

A new standard reference material (SRM) for drugs of abuse in human serum (SRM 1959) has been developed. This SRM is intended to be used as a control material for laboratories performing analysis of drugs of abuse in blood to evaluate the accuracy of their methods. SRM 1959 is a frozen human serum material fortified with seven compounds for which analyses are performed to determine evidence of illegal drug use: benzoylecgonine (BZE), methadone (METH), methamphetamine (MAMP), morphine (MOR), nordiazepam (NOR), phencyclidine (PCP), and 11-nor-Δ⁹-tetrahydrocannabinol-9-carboxylic acid (THC-9-COOH). Two independent methods involving isotope dilution (ID)-gas chromatography/mass spectrometry (GC/MS) and ID-liquid chromatography/mass spectrometry (LC/MS) were used for the value assignment. For THC-9-COOH, an additional measurement using LC/tandem mass spectrometry (LC/MS/MS) was also included. All methods used isotopically labeled compounds as internal standards and solid-phase extractions to isolate the analytes from the serum. The GC/MS methods used different clean-up procedures from those used for the LC/MS-based methods. Repeatability with within-set coefficients of variation (CVs) ranged from 0.5% to 4.3% for the GC/MS methods and from 0.2% to 1.2% for the LC/MS-based methods. Intermediate precision with between-set CVs for all the methods ranged from 0.1% to 1.1%. Agreement between the GC/MS and LC/MS methods ranged from 0.8% to 8.8%. The results from the methods were combined to obtain the certified concentrations and their expanded uncertainties.     

Characterisation of nicotine and related compounds using electrospray ionisation with ion trap mass spectrometry and with quadrupole time-of-flight mass spectrometry and their detection by liquid chromatography/electrospray ionisation mass spectrometry

Electrospray ionisation ion trap mass spectrometry (ESI-MS(n)) has been used to study the fragmentation patterns of nicotine and nine of its related compounds. From this study certain characteristic fragmentations are apparent with generally the pyrrolidine or piperidine ring being subject to chemical modifications. The structures of the product ions proposed for the ESI-MS(n) study have been supported by results from electrospray ionisation quadrupole time-of-flight mass spectrometry (ESI-QTOF-MS). Compounds with pyrrolidine and piperidine rings that possess an unsubstituted N atom have been shown to lose NH(3) at the MS(2) stage. Those compounds with N-methyl groups lose CH(3)NH(2) at the MS(2) stage. The loss of NH(3) or CH(3)NH(2) leaves the corresponding rings opened and this is followed by ring closure at the pyridine-2 carbon atom. Mono-N-oxides fragment in a similar way but the di-N-oxide can also fragment by cleavage of the bond between the pyridine and pyrrolidine rings. Cotinine also can undergo cleavage of this bond between the rings.This data therefore provides useful information on how substituents and the nature of the non-pyridine ring can affect the fragmentation patterns of nicotine and its related compounds. This information can be used in the characterisation of these compounds by liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) which results in the separation of nicotine and its related compounds with limits of detection (LODs) ranging from 15 to 105 ng/mL. The use of LC/ESI-MS to study nicotine-containing samples resulted in the simultaneous and unambiguous identification of seven of the compounds discussed in this paper: cotinine identified at retention time 12.5 min (with its [M+H](+) ion at m/z 177), nornicotine 16.0 min (m/z 149), anatabine 18.0 min (m/z 161), myosmine 18.5 min (m/z 147), anabasine 20.4 min (m/z 163), nicotine 22.2 min (m/z 163), and nicotyrine 31.4 min (m/z 159). For quality control of nicotine replacement therapy products, these nicotine impurities can be readily identified and determined at levels up to 0.3% for single impurities and up to 1.0% for total impurities.     

色谱与色谱/质谱法相结合分析热裂解汽油C9馏分

采用毛细管气相色谱-氢火焰离子化检测器（CGC-FID）和气相色谱-质谱法(GC/MS)分析了热裂解汽油C9 馏分的组成。实验使用PONA毛细管气相色谱柱(100 m×0.25 mm i.d.×0.5 μm),根据烃类化合物在PONA柱上的保留规律,以正构烷烃标样保留值作为碳数分布依据,定量分析了裂解汽油C9 馏分中烃类化合物的碳数分布和单体烃含量;用GC/MS联用技术和CGC保留值定性法相结合对裂解汽油C9 馏分中相对含量大于0.2%的39种化合物进行了定性。     

Characterization and identification of steroidal alkaloids in the Chinese herb Veratrum nigrum L. by high-performance liquid chromatography/electrospray ionization with multi-stage mass spectrometry

Electrospray ionization multi-stage mass spectrometry (ESI-MS(n)) was performed to study the fragmentation behaviour of seventeen steroidal alkaloids (4 protoverine-type alkaloids, 10 germine-type alkaloids and 3 zygadenin-type alkaloids) from the Chinese herb Veratrum nigrum L. The MS(n) spectra of the [M+H](+) ions for steroidal alkaloids provided a wealth of structural information on the substituted groups. In positive ion mode, the three types of alkaloids showed very different characteristic ions: m/z 436 or 418 for protoverine-type alkaloids; m/z 438, 420 or 402 for germine-type alkaloids; m/z 440 or 422 for zygadenin-type alkaloids. These fragments were used to deduce their mass spectral fragmentation mechanisms. Furthermore, the primary compounds in methanolic extracts of the herb of Veratrum nigrum L. were investigated by using liquid chromatography (LC)/ESI-MS(n). As a result, 21 steroidal alkaloids (5 protoverine-type alkaloids, 14 germine-type alkaloids and 2 zygadenin-type alkaloids) were selectively identified from 27 determined peaks. Eleven compounds were unambiguously identified by comparing with standard compounds and ten compounds were tentatively identified or deduced according to their MS(n) data. Two of these compounds (xingangermine and deacetyl xinganveratrine) were found to be novel steroidal alkaloids. In addition, the chemical structures of two pairs of steroidal alkaloid isomers were deduced by comparing their fragment ions. Given the important structural information of known and unknown steroidal alkaloids in crude herbal extracts, this study is useful for identifying these types of steroidal alkaloids in crude materials rapidly and selectively.