A study of the analytical behaviour of selected psycho-active drugs using liquid chromatography, ion trap mass spectrometry, gas chromatography and polarography and the construction of an appropriate database for drug characterisation

This paper provides analytical chemical information on a range of psycho-active drugs. This analytical chemical information on liquid chromatography-electrospray ionisation-mass spectrometry (HPLC-ESI-MS), ion trap mass spectrometry (ESI-MSⁿ), gas chromatography-flame ionisation detection (GLC-FID) and polarographic behaviour is then incorporated into a database which is of use in drug characterisation. Application is found in the determination of selected drug compounds in hair samples.     

Rapid determination of Cs and precise Cs/Cs atomic ratio in environmental samples by single-column chromatography coupled to triple-quadrupole inductively coupled plasma-mass spectrometry

For source identification, measurement of ¹³⁵Cs/¹³⁷Cs atomic ratio not only provides information apart from the detection of ¹³⁴Cs and ¹³⁷Cs, but it can also overcome the application limit that measurement of the ¹³⁴Cs/¹³⁷Cs ratio has due to the short half-life of ¹³⁴Cs (2.06 y). With the recent advancement of ICP-MS, it is necessary to improve the corresponding separation method for rapid and precise ¹³⁵Cs/¹³⁷Cs atomic ratio analysis. A novel separation and purification technique was developed for the new generation of triple-quadrupole inductively coupled plasma-mass spectrometry (ICP-MS/MS). The simple chemical separation, incorporating ammonium molybdophosphate selective adsorption of Cs and subsequent single cation-exchange chromatography, removes the majority of isobaric and polyatomic interference elements. Subsequently, the ICP-MS/MS removes residual interference elements and eliminates the peak tailing effect of stable ¹³³Cs, at m/z 134, 135, and 137. The developed analytical method was successfully applied to measure ¹³⁵Cs/¹³⁷Cs atomic ratios and ¹³⁵Cs activities in environmental samples (soil and sediment) for radiocesium source identification.     

A study of the analytical behaviour of selected synthetic and naturally occurring quinolines using electrospray ionisation ion trap mass spectrometry, liquid chromatography and gas chromatography and the construction of an appropriate database for quinoline characterisation

Mass spectral fragmentation of quinoline alkaloids of significance in plants has been investigated using electrospray ionisation ion trap mass spectrometry (ESI-MSⁿ) with a view to characterisation of molecules of unknown structure isolated from these natural sources. This investigation has led to the generation of an appropriate database incorporating data from ESI-MSⁿ and also from gas liquid chromatography (GLC) and liquid chromatography (HPLC) for these low molecular mass quinolines. This has been put to practical application in the identification of quinoline alkaloids in a plant extract. Thus, an acid extraction of the leaves of Choisya ternata containing such tertiary alkaloids was analysed by liquid chromatography-electrospray ionisation mass spectrometry (HPLC-ESI-MS) and the resulting behaviour of the quinolines was compared with that of the quinoline alkaloids in the database.     

Identification of polyoxypregnane glycosides from the stems of Marsdenia tenacissima by high-performance liquid chromatography/tandem mass spectrometry

A facile method based on high-performance liquid chromatography coupled with electrospray ionization tandem mass spectrometry (HPLC/(+)ESI-MSⁿ) has been established for the analysis of polyoxypregnane glycosides in the stems of Marsdenia tenacissima. The data reveals the ability of MSⁿ in the structural elucidation of polyoxypregnane glycosides including the nature of the polyoxypregnane core, the kinds of the substituents and the types of sugar residues. Offline Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) is also performed to assign accurate elemental compositions. In this study, eighteen polyoxypregnane glycosides have been investigated. Among these components, five compounds are unambiguously identified as Marsdenoside K, Tencissoside A, B, C and D; two compounds are established as novel compounds based on mass spectral data; and the other eleven compound's structures are tentatively proposed. Furthermore, breakdown curves are constructed to distinguish five pairs of isomers among these eighteen compounds. As far as our knowledge, this is the first report on identification of polyoxypregnane glycosides in the stems of M. tenacissima by HPLC/ESI-MSⁿ directly, which could save time and material consuming efforts in traditional phytochemistry analyses.     

Site-specific C content by quantitative isotopic C Nuclear Magnetic Resonance spectrometry: A pilot inter-laboratory study

Isotopic ¹³C NMR spectrometry, which is able to measure intra-molecular ¹³C composition, is of emerging demand because of the new information provided by the ¹³C site-specific content of a given molecule. A systematic evaluation of instrumental behaviour is of importance to envisage isotopic ¹³C NMR as a routine tool. This paper describes the first collaborative study of intra-molecular ¹³C composition by NMR. The main goals of the ring test were to establish intra- and inter-variability of the spectrometer response. Eight instruments with different configuration were retained for the exercise on the basis of a qualification test. Reproducibility at the natural abundance of isotopic ¹³C NMR was then assessed on vanillin from three different origins associated with specific δ¹³C_i profiles. The standard deviation was, on average, between 0.9 and 1.2‰ for intra-variability. The highest standard deviation for inter-variability was 2.1‰. This is significantly higher than the internal precision but could be considered good in respect of a first ring test on a new analytical method. The standard deviation of δ¹³C_i in vanillin was not homogeneous over the eight carbons, with no trend either for the carbon position or for the configuration of the spectrometer. However, since the repeatability for each instrument was satisfactory, correction factors for each carbon in vanillin could be calculated to harmonize the results.     

Analysis of palytoxin-like in Ostreopsis cultures by liquid chromatography with precolumn derivatization and fluorescence detection

A rapid liquid-chromatography (LC) method is presented which uses fluorescence detection (FLD) for palytoxin analogues analysis in benthic dinoflagellates of the genus Ostreopsis. The amino-acidic reagent 6-aminoquinolyl-N-hydroxisuccinimidyl carbamate (AccQ) was used for fluorescence labelling followed by LC-FLD.The efficacy of the method is exemplified by comparison of the results of the quantification obtained by LC-FLD and the hemolytic assay performed for palytoxins for which a highly significant linear correlation was achieved (r² = 0.9118). The derivatized palytoxin analogues were determined in the range of 0.75-25 ng.The proposed method was successfully applied to the determination and quantification of palytoxin analogues in 14 samples from different strains of Ostreopsis from different locations (Western Mediterranean Sea, Canary Islands, Madeira Islands and Southern coasts of Brazil). To confirm the chemical structure of the toxins, samples were also analyzed by liquid chromatography coupled with mass spectrometry (LC-MS) with a system that has a poorer sensitivity when compared with LC-FLD detection and the hemolytic assay. The successful use of this method with dinoflagellates is a good indicator of suitability for other types of marine samples.     

Screening and identification of multi-component in Qingkailing injection using combination of liquid chromatography/time-of-flight mass spectrometry and liquid chromatography/ion trap mass spectrometry

An approach for screening and identification of multi-component in complex traditional Chinese medicine systems with a combinative LC/MS (MSⁿ) technique was described in this paper. The chemical profile of Qingkailing injection, a well-known traditional Chinese formula in China, was studied using the established method as for an application. Benefit from combining the accurate mass measurement of LC/TOF-MS to generate elemental compositions and the complementary multilevel structural information provided by LC/ion trap MSⁿ, 33 components in Qingkailing injection were identified in all. The three isomers of chlorogenic acid, isochlorogenic acid and neochlorogenic acid which are derived from Flos Lonicerae, one of the medicinal materials in Qingkailing, were differentiated by verifying their MS³ fragmentation data. All the components identified were surveyed and classified according to their medicinal materials derivation. This study is expected to provide an effective and reliable pattern for comprehensive and systematic characterization of the complex traditional Chinese medicine systems.     

Studies on the Components of Crude and Processed Fructus Corni by ESI-MS

The components of crude and processed Fructus Corni were investigated by means of electrospray ionization-tandem mass spectrometry（ESI-MSn） technique in the negative ion mode. Compared with those of crude Fructus Corni, the chemical components of the processed Fructus Corni were changed both in quality and in quantity. From the ESI-MS spectra of the crude and processed Fructus Corni, six peaks were selected to establish the characte-ristic ESI-MS peaks. Several factors in the processing procedure were examined. The experimental results demonstrate that the chemical reactions that occurred in the processing procedure can be used for the elucidation of the processed mechanism of Fructus Corni, which is regularly affected by the processing conditions. The present article provides both the chemistry evidence for the understanding of the processing procedure of Fructus Corni and the specific methodology for the research of the processing procedure and quality identification of traditional Chinese medicine.     

Crystalline O,O′-di-sec-butyl and O,O′-diethyl dithiophosphate platinum(II) complexes: Synthesis, C and P CP/MAS NMR, single crystal X-ray diffraction studies and thermal behaviour

Crystalline bis(O,O′-di-sec-butyldithiophosphato)platinum(II) was prepared and studied by means of ¹³C, ³¹P CP/MAS NMR spectroscopy and single-crystal X-ray diffraction. The unit cell of the platinum(II) compound is comprised of one centrosymmetric mononuclear molecule [Pt{S₂P(O-sec-C₄H₉)₂}₂], in which the dithiophosphate groups display structural equivalence in both ³¹P NMR and XRD data. A pair of the dithiophosphate ligands exhibit the same S,S′-bidentate chelating structural function and form two planar four-membered chelate rings, [PtS₂P], in this molecule. The planar configuration of the [PtS₄] chromophore in structure 1 is governed by the dsp²-hybrid state of platinum(II). The structural states of the dithiophosphate groups in two different samples of complex 1 (one crystallised from ethanol and the other one precipitated from an aqueous solution) are all characterised by almost rhombic ³¹P chemical shift tensors. The observed essential dispersion of the ³¹P NMR chemical shift is caused by a coexistence of six optical isomers of molecule 1. The thermal behaviour of this compound was studied by means of simultaneous thermal analysis (a combination of TG and DSC) under an argon atmosphere. The thermal behaviour shows that the mass of 1 is lost in three steps, involving successive thermal decompositions of the organic and inorganic parts of this compound with platinum(II) dithio-meta-phosphate and reduced metallic platinum as the intermediate and the final products, respectively.     

Investigation of Symphytum cordatum alkaloids by liquid-liquid partitioning, thin-layer chromatography and liquid chromatography-ion-trap mass spectrometry

From the alkalised crude extract of Symphytum cordatum (L.) W.K. roots, pyrrolizidine alkaloids (PAs) were extracted as free tertiary bases and polar N-oxides in a merely one-step liquid-liquid partitioning (LLP) in separation funnel and subsequently pre-fractionated by preparative multiple-development (MD) thin-layer chromatography (TLC) on silica gel plates. In this way three alkaloid fractions of different polarities and retention on silica gel plates were obtained as: the most polar N-oxides of the highest retention, the tertiary bases of medium retention, and diesterified N-oxides of the lowest retention. The former fraction was reduced into free bases by sodium hydrosulfite and purified by LLP on Extrelut-NT3 cartridge. It was further analysed together with the two other fractions by high-performance liquid chromatography (HPLC)-ion-trap mass spectrometry with atmospheric pressure chemical ionization (APCI) interface on XTerra C₁₈ column using a gradient elution. Based on MSⁿ spectra, 18 various alkaloids have been tentatively determined for the first time in this plant as the following types of structure: echimidine-N-oxide (three diasteroisomers), 7-sarracinyl-9-viridiflorylretronecine (two diasteroisomers), echimidine (two diasteroisomers), lycopsamine (two diasteroisomers), dihydroechinatine-N-oxide, dihydroheliospathuline-N-oxide, lycopsamine-N-oxide (three diasteroisomers), 7-acetyllycopsamine-N-oxide, symphytine-N-oxide (two diasteroisomers) and 2″,3″-epoxyechiumine-N-oxide.     

The use of fast protein liquid chromatography with ICP-OES and ES-MS-MS detection for the determination of various forms of aluminium in the roots of Chinese cabbage

The distribution of aluminium (Al) species was investigated in the roots of Al-tolerant Chinese cabbage (Brassica rapa L. ssp. pekinensis) by employing fast protein liquid chromatography (FPLC) with inductively coupled plasma optical emission spectrometry (ICP-OES) and electrospray tandem mass spectrometry (ES-MS-MS) detection. The cabbage was exposed to a nutrient solution that contained 10 μg cm⁻³ of Al³⁺. The results demonstrated that after 24 h of exposure, Al was quantitatively taken up by the cabbage and was distributed in different parts of the plant. 36 ± 6% of total Al was located in the roots, while the remaining 64 ± 10% was transferred to the leaves. It was found that in the roots Al was partially present in the root sap (15.5%), while the majority (84.5%) was accumulated in its apoplasmic compartments. It was further demonstrated that the proportion of Al that entered the symplasm formed a complex with organic acid. Speciation analysis by FPLC with ICP-OES detection and ES-MS-MS identification of the binding ligand indicated that Al-citrate complex was the prevailing species in the root sap.The results of the present study showed that both immobilization of Al in the apoplasmic compartments of the roots and transformation of Al³⁺ to Al-citrate are most likely responsible for the tolerance of Chinese cabbage (B. rapa L. ssp. pekinensis) to the toxic effects of Al³⁺.     

Identification of C-21 steroidal glycosides from the roots of Cynanchum chekiangense by high-performance liquid chromatography/tandem mass spectrometry

High-performance liquid chromatography coupled with electrospray tandem mass spectrometry (HPLC/ESI-MS/MS) was used to identify C-21 steroidal glycosides with immunological activities in roots of Cynanchum chekiangense. In the MS/MS spectra, fragmentation reactions of the [M + Na]⁺ were recorded to provide structural information about the glycosyl and aglycone moieties. To further confirm the fragments structures, off-line Fourier transform ion cyclotron resonance tandem mass spectrometry (FT-ICR-MS/MS) was also performed. In the study, four known steroidal glycosides cynascyroside C, chekiangensosides A and B, glaucoside H, and four novel steroidal glycosides chekiangensosides C, D, E and chekiangensoside A isomer were identified based on mass spectral data, NMR spectral data and standards. This is the first report on identifying steroidal glycosides in roots of C. chekiangense by HPLC/ESI-MS/MS directly, which could save time and material consuming efforts in traditional phytochemistry analysis.     

Studies on principal components and antioxidant activity of different Radix Astragali samples using high-performance liquid chromatography/electrospray ionization multiple-stage tandem mass spectrometry

The principal components, isoflavonoids and astragalosides, in the extract of Radix Astragali were detected by a high-performance liquid chromatography couple to electrospray ionization ion trap multiple-stage tandem mass spectrometry (HPLC-ESI-IT-MSⁿ) method. By comparing the retention time (t_R) of HPLC, the ESI-MSⁿ data and the structures of analyzed compounds with the data of reference compounds and in the literature, 17 isoflavonoids and 12 astragalosides have been identified or tentatively deduced. By virtue of the extracted ion chromatogram (EIC) mode, simultaneous determination of isoflavonoids and astragalosides could be achieved when the different components formed overlapped peaks. And this method has been utilized to analyze the constituents in extracts of Radix Astragali from Helong City and of different growth years. Then the antioxidant activity of different samples has been successfully investigated by HPLC-ESI-MS method in multiple selected ion monitoring (MIM) mode, applying the spin trapping technology, and the Ferric Reducing Antioxidant Power (FRAP) assay was applied to support the result. The correlations of the isoflavonoids and astragalosides components and the antioxidant activities of Radix Astragali were summarized. The present paper demonstrates that HPLC-ESI-MSⁿ is a powerful method for the characterization of the principal components and evaluation of the antioxidant activity of Chinese medicinal herbs.     

Study of essential element accumulation in the leaves of a Cu-tolerant plant Elsholtzia splendens after Cu treatment by imaging laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS)

Laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) was used for the quantitative imaging of Cu and other essential elements (such as K, Mg, Mn, P, S and B) in the leaves of a Cu-tolerant plant Elsholtzia splendens treated with the enriched ⁶⁵Cu isotope tracer (isotope abundance of 89.2%). The leaves (newly formed, fully grown and oldest) were scanned directly with a focused Nd:YAG laser in the laser ablation chamber. The ablated material was transported with argon as carrier gas to a quadrupole-based ICP-MS (ICP-QMS), and the ion intensities of ⁶⁵Cu⁺, ³⁹K⁺, ²⁴Mg⁺, ⁵⁵Mn⁺, ³¹P⁺, ³⁴S⁺ and ¹¹B⁺ were measured by ICP-QMS to study the accumulation of Cu and other elements of interest. Standard reference material NIST SRM 1515 Apple Leaves doped with known concentrations of analytes (from 0.1 to 2000 mg L⁻¹) was measured together with the samples by LA-ICP-MS and was used for the quantification of the analytical data. Notable accumulation of Cu in the newly formed leaves was clearly identified by imaging LA-ICP-MS. The increased isotope ratios of ⁶⁵Cu/⁶³Cu measured by LA-ICP-MS demonstrated the path of Cu uptake and accumulation via the petiole and main veins in the leaves. Cu stress-induced accumulation of K, Mg, Mn, P and S in the newly formed leaves was observed, while B was not significantly affected. In the present study, the concentrations of K, Mg, Mn, P and S were not obviously changed in the fully grown leaves after short-term treatment. Along with the treatment, a visible decrease of K and P was found in the oldest leaves, while other elements were not influenced by Cu stress.     

Hydrophobic interaction chromatography coupled with atomic fluorescence spectrometric detection Effect of the denaturation on the determination of thiolic proteins

Hydrophobic interaction chromatography coupled online with chemical vapour atomic fluorescence spectrometry (HIC-CVGAFS) has been optimized for the analysis of thiolic proteins in denaturing conditions. Proteins are pre-column simultaneously denatured and derivatized in phosphate buffer solution containing 8.0 mol dm⁻³ urea and p-hydroxymercurybenzoate (PHMB) and the derivatized denatured proteins are separated on a silica HIC Eichrom Propyl column in the presence of 8.0 M urea in the mobile phase. Post-column online reaction of derivatized denatured proteins with bromine, generated in situ by KBr/KBrO₃ in HCl medium, allowed the fast conversion of the uncomplexed PHMB and of the PHMB bound to proteins to inorganic mercury also in presence of urea. Hg²⁺, present in solution as Hg²⁺-urea complex, is selectively detected by AFS in a Ar/H₂ miniaturized flame after sodium borohydride reduction to Hg. Under optimized conditions, online bromine treatment gives a 100±2% recovery of both free and protein-complexed PHMB. Denatured glyceraldehyde-3-phosphate dehydrogenase, aldolase, lactate dehydrogenase, trioso phosphate isomerase and β-lactoglobulin have been examined. As the sensitivity and limit of detection of proteins in the HIC-CVGAFS apparatus depends on number of SH groups reacting with PHMB, the denaturation process, which increases the number of PHMB-reactive thiolic groups in proteins, improves the analytical performances of the described system in protein analysis. The detection limit for the denatured proteins examined was found in the range of 10⁻¹⁰-10⁻¹² mol dm⁻³, depending on the considered protein, with linear calibration curves spanning over four decades of concentration.     

Interactions and stability of silver nanoparticles in the aqueous phase: Influence of natural organic matter (NOM) and ionic strength

The rapid development of nanotechnology and the related production and application of nanosized materials such as engineered nanoparticles (ENP) inevitably lead to the emission of these products into environmental systems. So far, little is known about the occurrence and the behaviour of ENP in environmental aquatic systems. In this contribution, the influence of natural organic matter (NOM) and ionic strength on the stability and the interactions of silver nanoparticles (n-Ag) in aqueous suspensions was investigated using UV–vis spectroscopy and asymmetrical flow field-flow fractionation (AF⁴) coupled with UV–vis detection and mass spectrometry (ICP-MS). n-Ag particles were synthesized by chemical reduction of AgNO₃ with NaBH₄ in the liquid phase at different NOM concentrations. It could be observed that the destabilization effect of increasing ionic strength on n-Ag suspensions was significantly decreased in the presence of NOM, leading to a more stable n-Ag particle suspension. The results indicate that this behaviour is due to the adsorption of NOM molecules onto the surface of n-Ag particles (“coating”) and the resulting steric stabilization of the particle suspension. The application of AF⁴ coupled with highly sensitive detectors turned out to be a powerful method to follow the aggregation of n-Ag particle suspensions at different physical–chemical conditions and to get meaningful information on their chemical composition and particle size distributions. The method described will also open the door to obtain reliable data on the occurrence and the behaviour of other ENP in environmental aquatic systems.     

Correlation between the synthetic origin of methamphetamine samples and their 15N and 13C stable isotope ratios

The active ingredient of ecstasy, N-methyl-3,4-methyldioxyphenylisopropylamine (MDMA) can be manufactured by a number of easy routes from simple precursors. We have synthesised 45 samples of MDMA following the five most common routes using N-precursors from 12 different origins and three different precursors for the aromatic moiety. The ¹³C and ¹⁵N contents of both the precursors and the MDMA samples derived therefrom were measured by isotope ratio mass spectrometry coupled to an elemental analyser (EA-IRMS). We show that within-pathway correlation between the ¹⁵N content of the precursor and that of the derived MDMA can be strong but that no general pattern of correlation can be defined. Rather, it is evident that the δ¹⁵N values of MDMA are strongly influenced by a combination of the δ¹⁵N values of the source of nitrogen used, the route by which the MDMA is synthesised, and the experimental conditions employed. Multivariate analysis (PCA) based on the δ¹⁵N values of the synthetic MDMA and of the δ¹⁵N and δ¹³C values of the N-precursors leads to good discrimination between the majority of the reaction conditions tested.     

Qualitative and quantitative analysis of polycyclic polyprenylated acylphloroglucinols from Garcinia species using ultra performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry

Polycyclic polyprenylated acylphloroglucinols (PPAPs) are a group of natural products isolated from different Garcinia species with a wide range of important biological activities. In this study, an ultra performance liquid chromatography (UPLC) coupled to photodiode-array detection and quadrupole time-of-flight mass spectrometry (Q-TOF) method was developed to characterize 16 PPAPs in 10 Garcinia species. In source dissociation techniques based on cone voltage fragmentation were used to fragment the deprotonated molecules and multiple mass spectrometry (MS/MS) using ramping collision energy were used to further break down the resulting product ions. The resulting characteristic fragment ions were generated by cleavage of C1-C5 bond and C7-C8 bond through concerted pericyclic reaction, which is especially valuable for differentiating three types of PPAPs isomers. As such, two new PPAPs isomers present in minor amount in the extracts of Garcinia oblongifolia were tentatively characterized by comparing their tandem mass spectra to the known ones. In addition, an UPLC-Q-TOF-MS method was validated for the quantitative determination of PPAPs. The method exhibited limits of detection from 2.7 to 21.4 ng mL⁻¹ and intra-day and inter-day variations were less than 3.7% and the recovery was in the range of 89-107% with RSD less than 9.0%. This UPLC-Q-TOF-MS method has successfully been applied to quantify 16 PPAPs in 32 samples of 10 Garcinia species, which were found to be a rich source of PPAPs.     

Lipid characterization of individual porcine oocytes by dual mode DESI-MS and data fusion

The development of sensitive measurements to analyze individual cells is of relevance to elucidate specialized roles or metabolic functions of each cell under physiological and pathological conditions. Lipids play multiple and critical roles in cellular functions and the application of analytical methods in the lipidomics area is of increasing interest. In this work, in vitro maturation of porcine oocytes was studied. Two independent sources of chemical information (represented by mass spectra in the positive and negative ion modes) from single oocytes (immature oocytes, 24-h and 44-h in vitro matured oocytes) were acquired by using desorption electrospray ionization-mass spectrometry (DESI-MS). Low and mid-level data fusion strategies are presented with the aim of better exploring the large amount of chemical information contained in the two mass spectrometric lipid profiles. Data were explored by principal component analysis (PCA) within the two multi-block approaches to include information on free fatty acids, phospholipids, cholesterol-related molecules, di- and triacylglycerols. After data fusion, clearer differences among immature and in vitro matured porcine oocytes were observed, which provide novel information regarding lipid metabolism throughout oocyte maturation. In particular, changes in TAG composition, as well as increase in fatty acid metabolism and membrane complexity were evidenced during the in vitro maturation process. This information can assist the improvement of in vitro embryo production for porcine species.     

Simultaneous quantification of three isoflavonoid glycosides in rabbit plasma after oral administration of Astragalus mongholicus extract by high-performance liquid chromatography coupled with electrospray ionization tandem mass spectrometry

A selective and sensitive liquid chromatography coupled with triple stage quadruple tandem mass spectrometry (HPLC/TSQ-MS/MS) was developed and validated for simultaneous quantification of calycosin-7-O-β-d-glycoside (CCSG), formononetin-7-O-β-d-glycoside (Ononin) and (6R,10R)-9,10-dimethoxypterocarpan-3-O-β-d-glycoside (DPG) in rabbit plasma. Plasma samples were extracted with solid-phase extraction (SPE), separated on an Inertsil ODS-3 column and detected by tandem mass spectrometry with electrospray ionization (ESI) interface in positive selective reaction monitoring (SRM) mode. 3,7,8-Trimethoxy-xanthone-1-O-primaverose was used as internal standard (IS) for quantitative measurement. For each analyte, one major product ion was chosen and used for screening of it. Calibration curves were generated over the range of 2-1000 ng mL⁻¹ with the correlation coefficients greater than 0.99 by using a weighted (1/χ) least squares linear regression. The method had the lower limit quantification of 0.15, 0.21 and 0.19 for CCSG, Ononin and DPG, respectively, with precision less than 20%. The intra- and inter-day precisions ranged from 2.48 to 6.38% and 4.81 to 11.78% (R.S.D.%), respectively. This assay is suitable for determining the above three trace glycosides in rabbit plasma simultaneously and thus investigating the pharmacokinetics of glycosides from Astragalus mongholicus extract in rabbits.