中药复方“清开灵”注射液中胆酸类物质的液相色谱/质谱/质谱分析

采有液相色谱／质谱／质谱法,通过保留时间、分子量和二级质谱的信息对复方“清开灵”注射液中的胆酸、去氧胆酸和鹅去氧胆酸进行了定性;建立了内标法对复方中相对含量较大的胆酸的定量分析方法;结果表明胆酸在875ng/L-140μg／L范围内线性良好,线性相关系数R^2=0.9999;RSD=2.4%。该方法样品处理简单,选择性好,灵敏度高。对中药的质量控制具有重要的意义。     

Accurate mass measurements by Fourier transform mass spectrometry in the study of advanced glycation end products/peptides

The Maillard reaction occurring between sugars and amino groups is important in living systems. When amino groups belonging to protein chains are involved, the Maillard reaction has been invoked as responsible for protein cross-linking and the production of 'toxic' compounds. The reaction leads to the production of a heterogeneous group of substances, usually called advanced glycation end products (AGEs). Classical analytical approaches, such as spectroscopic (ultraviolet, fluorescence) and mass spectrometric (matrix-assisted laser desorption/ionization, liquid chromatography/electrospray ionization mass spectrometry) methods, have shown that the digestion mixture is highly complex. However, there are clear differences between the digestion mixtures of glycated and unglycated human serum albumin (HSA). In the former case, possible glycated peptides belonging to the AGE peptide class may be identified. Tandem mass spectrometric experiments on selected species seemed to be promising as regards structural information, but it was thought of interest to undertake the present investigation, based on liquid chromatography/electrospray ionization Fourier transform mass spectrometry, in order to obtain definitive results on their elemental composition. Using this approach, about 20 glycated peptides were detected and their possible structures were postulated by examining the known sequence of HSA.     

Mass spectrometry in the structural analysis of flavonoids

Flavonoids are very common and widespread secondary plant metabolites. They have a wide range of biological and physiological activities and serve as chemotaxonomic marker compounds. Therefore, they have been extensively investigated both in the past and during recent years. The interest in them is still increasing. In the search for new compounds, and also in quality control, there is a need to have reliable methodology for the analysis of flavonoids. Mass spectrometry can make an invaluable contribution because of its high sensitivity, possibilities of coupling with liquid chromatography and the availability of powerful tandem mass spectrometric techniques. A review of currently available mass spectrometric methodology used in the structure elucidation of flavonoids is presented. Sample preparation, liquid chromatographic/mass spectrometric analysis and tandem mass spectrometric procedures for the characterization of flavonoid aglycones, O-glycosides, C-glycosides and acylated glycosides are considered.     

液相色谱-串联质谱法同时测定血浆中白藜芦醇苷及其代谢产物

建立同时测定大鼠血浆中白藜芦醇苷及其代谢产物白藜芦醇的液相色谱-串联质谱方法。以Lichro-spher C18色谱柱为分析柱,乙腈-水为流动相,采用电喷雾离子源(ESI),以多反应监测(MRM)模式检测,内标法定量,用于定量分析的离子反应分别为m/z389/227(白藜芦醇苷)和m/z227/143(白藜芦醇)。血浆中的白藜芦醇苷及白藜芦醇用乙酸乙酯提取,N2吹干乙酸乙酯,残留物用甲醇溶解,注入LC/MS/MS系统进行检测。在选定的样品预处理、色谱及质谱条件下,白藜芦醇苷、白藜芦醇及内标物能够达到基线分离而且离子化效果好。用LC/MS/MS法检测大鼠血浆中的白藜芦醇苷及其代谢产物白藜芦醇,线性范围0.4~200μg/L,日内、日间精密度(RSD)均小于15%;检测血浆低、中、高3个浓度(1、20、100μg/L)白藜芦醇苷的回收率分别为106.2%、97.8%和91.6%;检测血浆低、中、高3个浓度(1、20、100μg/L)白藜芦醇的回收率分别为113.2%、103.6%和93.4%。本方法具有灵敏、准确、快速的特点,可用于白藜芦醇苷的药代动力学研究。     

Characterization of geometrid sex pheromones by electrospray ionization time-of-flight mass spectrometry

The utility of liquid chromatography combined with time-of-flight mass spectrometry (LC/TOFMS) was demonstrated for studies on chiral unsaturated epoxy compounds, sex pheromones produced mainly by female moths in the family Geometridae. By electrospray ionization (ESI), each synthetic epoxyalkadiene derived from (Z,Z,Z)-3,6,9-triene with a C(18)-C(23) straight chain showed three ion series, [M + NH(4)](+), [M + H](+) and [M - OH](+), with high resolution and good sensitivity, indicating its molecular formula. In addition to these, characteristic fragment ions at m/z M - 57 and M - 71 for the 3,4-epoxides and at m/z M - 123 and 123 for the 9,10-epoxides were detected, whereas the 6,7-epoxides did not produce fragment ions that reflected their structures. Monitoring these diagnostic ions during the LC/MS analysis of a gland extract, the natural sex pheromone of the mulberry looper was confirmed to be (Z,Z)-cis-9,10-epoxy-3,6-octadecadiene, which was separable from the other positional isomers on an ODS column. Furthermore, (Z,Z)-cis-3,4-epoxy-6,9-nonadecadiene secreted by the Japanese giant looper was analyzed with a chiral column, and the stereochemistry was determined directly.     

Mass spectrometry in the study of anthocyanins and their derivatives: differentiation of Vitis vinifera and hybrid grapes by liquid chromatography/electrospray ionization mass spectrometry and tandem mass spectrometry

A mass spectrometric-based procedure for anthocyanin profiling was set up to distinguish authentic Vitis vinifera from hybrid red grapevine cultivars. 3-O-Monoglucoside and the related acetyl-, p-coumaryl- and caffeoyl-monoglucoside anthocyanins occurred only in Vitis vinifera, whereas 3,5-O-diglucoside and the substituted acetyl-, p-coumaryl-, feruloyl- and caffeoyl-diglucoside anthocyanins were the additional pigments in hybrid grapevines. The procedure was applied expressly to identify red grape cultivars based on the anthocyanin chemo-type determination. In particular, a red grape cultivar, having 3,5-O-diglucoside anthocyanins and a novel class of anthocyanin monoglucosides, such as cyanidin-3-O-, cyanidin-3-O-(6-O-acetyl)- and cyanidin-3-O-(6-O-p-coumaryl)pentoside, was classified as hybrid. A second vine cultivar, characterized exclusively by 3-O-monoglucoside anthocyanins, was included among the Vitis vinifera species. Anthocyanin profiling by mass spectrometry could represent the core of a chemotaxonomic procedure for distinguishing American and European grapevines based on the identification of post-synthetic anthocyanidin modification.     

Quantification of Diaminomaleonitrile Using High Performance Liquid Chromatography with UV and Electrospray Ionization Mass Spectrometric Detection

Abstract

A method was developed to quantify diaminomaleonitrile (DAMN) in complex aqueous matrices using high performance liquid chromatography with UV and electrospray ionization mass spectrometric (ESI-MS) detection. The method is rapid (analytical times <10 min), able to quantify DAMN against complex backgrounds, and achieved with part per billion detection limits. Given the central role of DAMN in generating heterocycles of known biological significance, this study provides a rapid and sensitive method for quantifying DAMN in complex solutions.     

Quantitative determination of domperidone in human plasma by ultraperformance liquid chromatography with electrospray ionization tandem mass spectrometry

A simple and sensitive liquid chromatography-tandem mass spectrometry method was developed and validated for determining domperidone in human plasma. The analyte and internal standard (IS; mosapride) were isolated from plasma samples by protein precipitation with methanol (containing 0.1% formic acid). The chromatographic separation was performed on an Xterra MS C(18) Column (2.1 x 150 mm, 5.0 microm) with a gradient programme mobile phase consisting of 0.1% formic acid and acetonitrile at a flow rate of 0.30 mL/min. The total run time was 4.0 min. The analyses were carried out by multiple reaction monitoring using the parent-to-daughter combinations m/z 426 --> 175 and m/z 422 --> 198 (IS). The areas of peaks from the analyte and IS were used for quantification of domperidone. The method was validated according to the FDA guidelines on bioanalytical method validation. Validation results indicated that the lower limit of quantification was 0.2 ng/mL, and the assay exhibited a linear range of 0.2-60.0 ng/mL and gave a correlation coefficient (r(2)) of 0.999 or better. Quality control samples (0.4, 0.8, 15 and 50 ng/mL) in six replicates from three different analytical runs demonstrated an intra-assay precision (RSD) 4.43-6.26%, an inter-assay precision 5.25-7.45% and an overall accuracy (relative error) of <6.92%. The method can be applied to pharmacokinetic and bioequivalence studies of domperidone.     

乙酰螺旋霉素活性成分的分析

乙酰螺旋霉素是一种含有多个活性成分的抗生素药物，本文首先用电喷雾质谱法测定了混合物的组成，然后采用高效液相色谱和微乳毛细管电泳法分离了其中的活性成分，用液相色谱－质谱联用技术鉴定了液相色谱的流出峰。     

Relationships between structure,ionization profile and sensitivity of exogenous anabolic steroids under electrospray ionization and analysis in human urine using liquid chromatography–tandem mass spectrometry

The relationships between the ionization profile, sensitivity, and structures of 64 exogenous anabolic steroids (groups I–IV) was investigated under electrospray ionization (ESI) conditions. The target analytes were ionized as [M + H]⁺ or [M + H–nH₂O]⁺ in the positive mode, and these ions were used as precursor ions for selected reaction monitoring analysis. The collision energy and Q3 ions were optimized based on the sensitivity and selectivity. The limits of detection (LODs) were 0.05–20 ng/mL for the 64 steroids. The LODs for 38 compounds, 14 compounds and 12 compounds were in the range of 0.05–1, 2–5 and 10–20 ng/mL, respectively. Steroids including the conjugated keto‐functional group at C3 showed good proton affinity and stability, and generated the [M + H]⁺ ion as the most abundant precursor ion. In addition, the LODs of steroids using the [M + H]⁺ ion as the precursor ion were mostly distributed at low concentrations. In contrast, steroids containing conjugated/unconjugated hydroxyl functional groups at C3 generated [M + H ? H₂O]⁺ or [M + H ? 2H₂O]⁺ ions, and these steroids showed relatively high LODs owing to poor stability and multiple ion formation. An LC‐MS/MS method based on the present ionization profile was developed and validated for the determination of 78 steroids (groups I–V) in human urine. Copyright © 2015 John Wiley & Sons, Ltd.     

Effects of ion-pairing reagents on the electrospray signal suppression of sulphonated dyes and intermediates

High-performance liquid chromatography/mass spectrometry (HPLC/MS) analysis of anionic species such as sulphonic acid dyes and intermediates requires volatile ion-pairing mobile phase additives. Six di- and trialkylammonium acetates were compared with tetraalkylammonium salts and ammonium acetate in the concentration range 0-20 mmol l(-1) as mobile phase additives for HPLC/MS of polysulphonated compounds. The effects of the structure and concentration of the ion-pairing reagents on the electrospray response of mono-, di- and tetrasulphonic aromatic acids and acid dyes were studied in detail. Further, five different mass analysers and instrument geometries were compared. A higher signal decrease is observed with linear geometry instruments in comparison to orthogonal or even Z-spray geometry mass spectrometers. The concentration of mobile phase additives has a significant influence on the abundance ratios of multiply charged ions in the mass spectra of polysulphonated compounds. The competing ions of sulphonic acids may also cause significant signal suppression.     

Liquid chromatographic/electrospray ionization mass spectrometric identification of the oxidation end-products of metformin in aqueous solutions

Metformin is an antihyperglycemic drug that exhibits some antioxidant properties. HO*-induced oxidation of metformin was studied in aqueous solution, in both aerated and deaerated conditions. Gamma radiolysis of water was used to generate HO* free radicals, capable of initiating one-electron oxidation of metformin. Oxidation end-products were identified by direct infusion mass spectrometry (MS) and high-performance liquid chromatography/mass spectrometry (HPLC/MSn): for every product, structure elucidation was based on its mass (simple mass spectra confirmed by HPLC/MS). In addition, fragmentation spectra (MS2, MS3 and MS4) and the determination of deuterium-hydrogen exchange sites provided valuable information allowing the complete identification of some of the end-products. At low radiation dose, four products were identified as primary ones, since they result from the direct attack of HO* radicals on metformin. These primary oxidation end-products were identified respectively as hydroperoxide of metformin, covalent dimer of metformin, methylbiguanide and 2-amino-4-imino-5-methyl-1,3,5-triazine. At high radiation dose, seven other products were identified as secondary ones, resulting from the HO*-induced oxidation of the primary end-products. A reaction scheme was postulated for the interpretation of the results.     

高效液相色谱质谱法分析脂肪醇聚氧乙烯醚中的碳数及环氧乙烷加成数的分布

应用高效液相色谱法将脂肪醇聚氧乙烯醚按碳数和环氧乙烷加成数 (EO数 )分离后 ,在线联用电喷雾离子化质谱 (ESI MS) ,根据其一系列准分子离子峰判断烷基链长和环氧乙烷聚合度。解释了准分子离子峰强度失真的原因。     

Determination of ivermectin B(1a) in animal plasma by liquid chromatography combined with electrospray ionization mass spectrometry

A novel, sensitive and specific method for the quantitative determination of ivermectin B(1a) in animal plasma using liquid chromatography combined with positive electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) is presented. Abamectin was used as the internal standard. Extraction of the samples was performed with a deproteinization step using acetonitrile. Chromatographic separation was achieved on a Nucleosil ODS 5 microm column, using gradient elution with 0.2% (v/v) acetic acid in water and 0.2% (v/v) acetic acid in acetonitrile. The method was validated according to the requirements defined by the European Community. Calibration curves using plasma fortified between 1 and 100 ng ml(-1) showed a good linear correlation (r > or = 0.9989, goodness-of-fit coefficient < or =8.1%). The trueness at 2 and 25 ng ml(-1) (n = 6) was +4.2 and -17.1%, respectively. The trueness and between-run precision for the analysis of quality control samples at 25 ng ml(-1) was -4.0 and 11.0%, respectively (n = 16). The limit of quantification of the method was 1.0 ng ml(-1), for which the trueness and precision also fell within acceptable limits. Using a signal-to-noise ratio of 3 : 1, the limit of detection was calculated to be 0.2 ng ml(-1). The specificity was demonstrated with respect to ivermectin B(1b).The method was successfully used for the quantitative determination of ivermectin B(1a) in plasma samples from treated bovines, demonstrating the usefulness of the developed method for application in the field of pharmacokinetics.     

Characterization of the interglycosidic linkage in di-, tri-, tetra- and pentaglycosylated flavonoids and differentiation of positional isomers by liquid chromatography/electrospray ionization tandem mass spectrometry

The LC/UV-DAD/ESI-MSn negative fragmentation mode of 23 O-glycosylated flavonoids with two, three, four and five hexoses was studied. The results show that it is possible to differentiate the (1-->2) and (1-->6) interglucosidic linkages and also to discern between the flavonoid isomers with two glucoses (sophorosides, gentiobiosides and X,Y-diglucosides), three glucoses (sophorotriosides and X-sophoroside-Y-glucoside) and four glucoses (X-sophorotriosides-Y-glucoside and X-sophoroside-Y-sophoroside). In the characterization of the (1-->2) and (1-->6) interglycosidic linkages, the Y1- (-162 u) and Z1- (-180 u) ions play a relevant role. In the first case ions with high relative abundance (13-79%) are found, whereas in the other cases they are in very low abundance or absent. X,Y-di-O-glucoside flavonoids can be differentiated from the O-diglucoside flavonoids by the presence of Y1- (base peak) and Y0- (approximately 30%) ions and the absence of Z1- ions. Regarding flavonoids glycosylated with three glucoses, X-sophoroside-Y-glucoside flavonoids show the Y7(0-) (-162 u) ion as the only peak in MS2 events whereas in sophorotrioside flavonoids various ions due to intermediate fragmentations are observed. These ions are characteristic of a (1-->2) interglucosidic linkage. In MS2 experiments on flavonoids with four glucoses (X-sophorotrioside-Y-glucoside and X-sophoroside-Y-sophoroside), the base peak indicated the total loss of the sugar moieties in position 7. In addition, the characterization of the type of interglycosidic linkage in flavonoids glycosylated with five sugars can be achieved. On the other hand, in tetra- and pentaglycosylated flavonoids, the ions that characterize the (1-->2) interglucosidic linkage formed by intermediate fragmentation of the oligosacharide residues (sophorosides and sophorotriosides) are found in much higher relative abundance in MS3 than in MS2 experiments, where they are almost not detected.     

液相色谱/电喷雾质谱测定柑桔中的柠檬苦素类似物配糖体

 利用液相色谱 /电喷雾质谱联用技术对柚皮中的柠檬苦素类似物配糖体进行了定向分析 ,在柚皮的乙醇提取物中检测出奥巴叩酮配糖体和诺米林配糖体。利用制备液相色谱对这两种柠檬苦素类似物配糖体进行纯化 ,并用核磁共振对其结构进行鉴定。测定结果表明 ,液质联用法不需标样即可对柑桔中的柠檬苦素类似物配糖体进行定性分析。方法准确、快速、简便。     

Sensitive measurement of vinorelbine in dog plasma by liquid chromatography–electrospray ionization tandem mass spectrometry utilizing transitions from double‐charged precursor ions

A sensitive high‐performance liquid chromatography/tandem mass spectrometry (LC/MS/MS) method for measuring vinorelbine was developed. A 100 µL aliquot of plasma was spiked with deuterium‐labeled internal standard and subjected to solid‐phase extraction using an Oasis HLB μ‐elution plate. Two microliters of the extracted samples was directly injected into LC/MS/MS. Chromatographic separation was achieved on a Capcell Pak C18 UG column (2 × 75 mm) with a gradient elution of methanol (mobile phase B) against 0.05% formic acid in aqueous 10 mm ammonium formate (mobile phase A). The LC flow rate was set to 0.28 mL/min and the gradient (solvent B concentration) was processed from 40 to 90%. In mass spectrometric detection, observation of the reaction from a double‐charged precursor ion [M + 2H]²⁺ (m/z 390) to product ion m/z 122 provided very high sensitivity. The method was validated with a lower limit of detection of 0.2 ng/mL with 0.1 mL of plasma, and the method was used to determine the plasma pharmacokinetics of vinorelbine in dogs. Copyright © 2010 John Wiley & Sons, Ltd.     

离子色谱-质谱联用技术测定污泥样品中的痕量高氯酸盐

建立了离子色谱-质谱联用技术测定活性污泥样品中高氯酸盐的分析方法。以高容量、强亲水性的IonPacAS20(2mm)阴离子交换柱为分析柱,EGC在线产生等浓度KOH为淋洗液,淋洗液经抑制成水后将样品带入质谱检测。ESI-MS-MS以多元反应监测模式监控100.8/84.9、98.8/66.9、100.8/68.9和98.8/82.9离子对,以98.8/82.9离子对的峰面积进行定量。该方法对高氯酸盐的检出限(S/N=3)为0.01μg/L,高氯酸盐在0.05~100μg/L浓度范围内具有良好的线性,线性相关系数r=0.9988。0.2μg/L的标准溶液重复进样9次,高氯酸盐峰面积的相对标准偏差(RSD)为2.3%。运用该方法测定采自不同地区的活性污泥样品中的高氯酸盐,并对样品加标回收,得回收率在88.5%~102.2%之间。     

Sheath liquid interface for the coupling of normal-phase liquid chromatography with electrospray mass spectrometry and its application to the analysis of neoflavonoids

A novel interface that allows normal-phase liquid chromatography to be coupled with electrospray ionization (ESI) is reported. A make-up solution of 60 mM ammonium acetate in methanol, infused at a 5 microl min(-1) flow-rate at the tip of the electrospray probe, provides a sheath liquid which is poorly miscible with the chromatographic effluent, but promotes efficient ionization of the targeted analytes. Protonated molecules generated in the ESI source were subjected to tandem mass spectrometric experiments in a triple-quadrupole mass spectrometer. The main fragmentation reactions were characterized for each analyte and specific mass spectral transitions were used to acquire chromatographic data in the multiple reaction monitoring detection mode. Results obtained during optimization of the sheath liquid composition and flow-rate suggest that the electrospray process was mainly under the control of the make-up solution, and that it forms an external charged layer around a neutral chromatographic mobile phase core. This sheath liquid interface was implemented for the analysis of some neoflavonoid compounds and its performance was evaluated. Limits of detection were established for calophillolide, inophyllum B, inophyllum P and inophyllum C at 100, 25, 15 and 100 ng ml(-1), respectively.     

Profiles analysis of proanthocyanidins in the argun nut (Medemia argun—an ancient Egyptian palm) by LC–ESI–MS/MS

Medemia argun is an ancient palm rich in proanthocyanidins (PACs). These polyphenolic compounds are widely distributed in plants and are an integral part of the human diet. A sensitive high‐performance liquid chromatography and electrospray ionization mass spectrometry (HPLC–ESI–MS) method in the negative ion mode for sequencing these ubiquitous and highly beneficial antioxidants is described in order to profile different PACs in M. argun nuts. The analytical protocol based on tandem mass spectrometry was used to sequence dimers, trimers, tetramers and pentamers with different A‐type, B‐type and A/B‐type linkages. Diagnostic ions resulting from heterocyclic ring fission and retro‐Diels–Alder reaction of flavan‐3‐ol provided information on the hydroxylation pattern and the type of interflavan bond. The sequences were discovered through ions derived from quinone methide cleavage of the interflavan bond. The identification of PACs linkages through LC–MSⁿ eliminates a number of tedious separation steps. The method was successfully applied to give a view of PAC profile in M. argun nuts. M. argun nuts contained 636.88 mg/g PACs (as equivalent of (þ)‐catechin). The data obtained in our research show that M. argun is a rich source of hydrolyzable PACs. Copyright © 2014 John Wiley & Sons, Ltd.