Pharmacokinetics of anthraquinones in rat plasma after oral administration of a rhubarb extract

A sensitive and specific LC‐MS/MS method was developed for simultaneous determination of aloe‐emodin, rhein, emodin, chrysophanol and physcion and their conjugates in rat plasma. The lower limit of quantitation of each anthraquinone was 0.020–0.040 µm . Intra‐day and inter‐day accuracies were 90.1–114.3% and the precisions were <14.6%. The matrix effects were 104.0–113.2%. The method was successfully applied to a pharmacokinetic study in rats receiving a rhubarb extract orally. The area under the concentration–time curve (AUC_0–t) and peak concentration (C_max) of free aloe‐emodin and emodin in rat plasma were much lower than those of rhein. The amounts of chrysophanol and physcion were too low to be continuously detected. After treating the plasma samples with β‐glucuronidases, each anthraquinone was detectable throughout the experimental period (36 h) and showed much higher plasma concentrations and AUC_0–t. The free/total ratios of aloe‐emodin, rhein and emodin were 6.5, 49.0 and 1.7% for C_max and 3.7, 32.5 and 1.1% for AUC_0–t, respectively. The dose‐normalized AUC_0–t and C_max of the total of each anthraquinone were in the same descending order: rhein > emodin > chrysophanol > physcion > aloe‐emodin. These findings reveal phase II conjugates as the dominant in vivo existing forms of rhubarb antharquinones and warrant a further study to evaluate their contribution to the herbal activity. Copyright © 2013 John Wiley & Sons, Ltd.     

Simultaneous determination of wogonin,oroxylin a,schisandrin, paeoniflorin and emodin in rat serum by HPLC–MS/MS and application to pharmacokinetic studies

Wogonin and oroxylin A in Scutellariae Radix, schisandrin in Chinensis Fructus, paeoniflorin in Moutan Cortex and emodin in Polygoni Cuspidate Rhizome et Radix are anti‐inflammatory active compounds. A method for simultaneous determination of the five compounds in rat was developed and validated using high‐performance liquid chromatography with tandem mass spectrometry (HPLC–MS/MS). The separation was performed on a Symmetry C₁₈ column (4.6 × 50 mm, 3.5 μm) with acetonitrile and 0.1% formic acid aqueous solution as the mobile phases. The detection was performed using multiple‐reaction monitoring with electrospray ionization source in positive–negative ion mode. The calibration curves showed good linearity (r ≥ 0.9955). The lower limit of quantification (LLOQ) was 5 ng/mL for wogonin and schisandrin, 10 ng/mL for oroxylin A and emodin, and 15 ng/mL for paeoniflorin, respectively. The relative standard deviations of intraday and interday precisions were <11.49 and 14.28%, respectively. The extraction recoveries and matrix effects were acceptable. The analytes were stable under the experiment conditions. The validated method has been successfully applied to pharmacokinetic studies of the five compounds in rats after oral administration of Hu‐gan‐kan‐kang‐yuan capsule. This paper would be a valuable reference for pharmacokinetic studies of Chinese medicine preparations containing the five compounds.     

HPLC–TOF‐MS and HPLC–MS/MS combined with multivariate analysis for the characterization and discrimination of phenolic profiles in nonfumigated and sulfur‐fumigated rhubarb

Sulfur fumigation has recently been used during the postharvest handling of rhubarb to reduce the drying duration and control pests. However, a few reports question the effect of sulfur fumigation on the bioactive components of rhubarb, which is crucial for the quality evaluation of the herbal medicine. The bottleneck limiting the study comes from the complex compounds that exist in herb samples with diverse structural features, wide concentration range and the difficulty to obtain all the reference standards. In this study, an integrated strategy based on the highly effective separation and analysis by liquid chromatography coupled with diode‐array detection and time‐of‐flight/triple‐quadruple tandem mass spectrometry combined with multivariate analysis was established. 68 phenolic compounds that exist in nonfumigated and sulfur‐fumigated herb samples of rhubarb were tentatively assigned based on their retention behavior, UV spectra, accurate molecular weight, and mass spectral fragments. Qualitative and semiquantitative comparison revealed a serious reduction of the majority of phenolic compounds in sulfur‐fumigated rhubarb. Furthermore, multivariate analysis was applied to holistically discriminate nonfumigated from sulfur‐fumigated rhubarb and explore the characteristic chemical markers. The established approach was specific and rapid for characterizing and screening sulfur‐fumigated rhubarb among commercial samples and could be applied for the quality assessment of other sulfur‐fumigated herbs.     

Application of a highly sensitive magnetic solid phase extraction for phytochemical compounds in medicinal plant and biological fluids by ultra‐high performance liquid chromatography coupled with quadrupole time‐of‐flight tandem mass spectrometry

A highly sensitive method using reduced graphene oxide with iron oxide (rGO/Fe₃O₄) as the sorbent in magnetic SPE has been developed for the purification of five anthraquinones (emodin, rhein, aloeemodin, physcion, and chrysophanol) in rhubarb and rat urine by ultra‐HPLC coupled with quadrupole TOF/MS. The extraction was accomplished by adding trace amount rGO/Fe₃O₄ suspension to 200 mL of aqueous mixture, and the excellent adsorption capacity of the nanoparticles was fully demonstrated in this procedure. Under the optimized conditions, the calibration curves were linear in the concentration range of 0.05–27.77 ng/mL with correlation coefficients varying from 0.9902 to 0.9978. The LODs ranged from 0.28 to 58.99 pg/mL. The experimental results indicated that the proposed method was feasible for the analysis of anthraquinones in rhubarb and urine samples.     

Use of liquid chromatography hybrid triple‐quadrupole mass spectrometry for the detection of emodin metabolites in rat bile and urine

Emodin is the representative form of rhubarb, which is widely used in traditional Chinese medicine for the treatment of purgative, anti‐inflammatory, antioxidative and antiviral, etc. Previous reports demonstrated that emodin glucuronide was the major metabolite in plasma. Owing to the extensive conjugation reactions of polyphenols, the aim of this study was to identify the metabolites of emodin in rat bile and urine. Neutral loss and precursor ion scan methods of triple‐quadrupole mass spectrometer revealed 13 conjugated metabolites in rat bile and 22 metabolites in rat urine, which included four phase I and 18 phase II metabolites. The major metabolites in rat biosamples were emodin glucuronoconjugates. Moreover, rhein monoglucuronide, chrysophanol monoglucuronide and rhein sulfate were proposed for the first time after oral administration of emodin. Overall, liquid chromatography hybrid triple‐quadrupole mass spectrometry analysis leads to the discovery of several novel emodin metabolites in rat bile and urine and underscores that conjugated with glucuronic acid is the main metabolic pathway.     

A rapid HPLC/ESI‐MS/MS method for quantitative analysis of isovalerylshikonin in rat plasma

A new, fast and sensitive high‐performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC/ESI‐MS/MS) method was developed and validated for isovalerylshikonin in rat plasma using emodin as internal standard (IS). The analyte was extracted from rat plasma with ethyl acetate, after 10% HCl treatment and protein precipitated by methanol. The compound was separated on an Ultimate? XB‐C₁₈ analytical column using a mobile phase of methanol–10 mM ammonium acetate in water–acetonitrile containing 0.05% formic acid (45 : 10 : 45, v/v/v) with isogradient elution. The analyte was detected in negative ion mode using multiple‐reaction monitoring. The method was validated and the specificity, linearity, lower limit of quantitation (LLOQ), precision, accuracy, recoveries and stability were determined. LLOQ was 9 ng/mL for isovalerylshikonin. Correlation coefficient (r) value for the linear range of the analyte was greater than 0.99. The intra‐day and inter‐day precision and accuracy were better than 8.52%. The relative and absolute recovery was above 86% and no matrix effects were observed for isovalerylshikonin. This validated method provides a modern, rapid and robust procedure for the pharmacokinetic study of the two compounds in rats after intravenous administration to rats (n = 4). Copyright © 2009 John Wiley & Sons, Ltd.     

Simultaneous quantification of anthraquinone glycosides,aglycones, and glucuronic acid metabolites in rat plasma and tissues after oral administration of raw and steamed rhubarb in blood stasis rats by UHPLC-MS/MS

Rhubarb is a widely used herbal medicine. To achieve different effects, rhubarb is usually steamed with rice wine (steamed rhubarb). This steaming treatment increases the blood-activating and stasis-removing effects of rhubarb. A specific and accurate ultra high performance liquid chromatography with tandem mass spectrometry method was established for simultaneous determination of anthraquinone glycosides, aglycones, and glucuronic acid metabolites in plasma and tissues after administration of raw rhubarb and steamed rhubarb in blood stasis rats. Chromatographic separation was performed on ACQUITY UPLC BEH Shield RP 18 column using the mobile phase consisting of water and acetonitrile both containing 0.1% formic acid. Satisfactory linearity, precision, accuracy, extraction recovery, and matrix effect have been achieved. From pharmacokinetic study, it showed that glucuronic acid metabolites were found abundantly in plasma as bioactive components. The lower area under concentration-time curve, maximum concentration, and higher apparent volume of distribution (P < 0.01), body clearance (P < 0.01) values in steamed rhubarb showed that most components of steamed rhubarb have lower bioavailability in plasma compared with raw rhubarb. But it found these components were mainly distributed in spleen and liver with large blood flow and perfusion rates. The pharmacokinetics and tissue distribution studies of anthraquinone components will provide helpful information for clinical application of steamed rhubarb and raw rhubarb.     

Profiling and analysis of multiple compounds in rhubarb decoction after processing by wine steaming using UHPLC–Q‐TOF‐MS coupled with multiple statistical strategies

Rhubarb is one of the most popular traditional Chinese medicines and has been used for thousands of years in many Asian countries. Prepared rhubarb is obtained by steaming raw rhubarb with glutinous rice wine until it turned black in appearance both inside and outside. After processing, the therapeutic effects of prepared rhubarb change a lot. To find out the exact compound changes of the chemical profile in a decoction of rhubarb after processing and to clarify the material basis of the changed therapeutic effects, an ultra‐high performance liquid chromatography with quadrupole time‐of‐flight mass spectrometry method coupled with automated data analysis software and statistical strategy was developed. As a result, 63 peaks in raw rhubarb and 54 peaks in prepared rhubarb were detected, and a total of 45 chemical compounds were identified. The analysis data were subjected to a principle component analysis and a t‐test. Based on the results, 16 peaks were found to be the main contributors to the significant difference (p < 0.05) between raw and prepared rhubarb. Compared with raw rhubarb, the content of 15 components in prepared rhubarb was lower, while only rhein (1,8‐dihydroxy‐3‐carboxy anthraquinone) showed a higher intensity.     

Solid-phase extraction and ultra high-performance liquid chromatography tandem mass spectrometry analysis of the gastrointestinal absorption of emodin in different digestive segments of rats

A rapid, simple and sensitive ultra high‐performance liquid chromatography (UHPLC‐MS/MS) method was established for determining the absorption amount of emodin in the five digestive segments of rat. Plasma samples were pre‐purified by solid‐phase extraction (SPE) with Oasis MAX cartridge. Separation of emodin and 1,8‐dihydroxyanthraquinone (internal standard) was performed on an Acquity BEH UHPLC C₁₈ column (100 mm×2.1 mm, 1.7 μm) with gradient elution of methanol and 0.1% formic acid aqueous solution. The LC/MS system was operated under multiple reaction monitoring mode using electrospray ionization (ESI) in negative ion mode. The results showed that this established method was valid and sensitive for emodin within 0.04–16.4 μg/mL, with low limits of detection and quantification of 0.6 ng/mL and 2.4 ng/mL, respectively and upper limit of quantification of 220.0 ng/mL. The intra‐ and interday variations were below 4.9% of RSD. The extraction recoveries were 98.9–106.1% with RSD of 1.9–3.2%. The plasma concentration‐time relationship showed that the absorption of emodin in stomach was faster than in intestine segments. The sequence of absorption amount was: ileum>jejunum>colon≈duodenum>stomach. Most of emodin was absorbed in ileum, and the absorption amount was increased with prolonged retention of drug form in intestine, especially in ileum and jejunum. The developed UHPLC‐ESI‐MS/MS method was appropriate for determining the in vivo absorption of emodin in other herbal medicines or preparations containing this compound.     

Evaluation of the influence of mirabilite on the absorption and pharmacokinetics of the ingredients in Dahuang‐mudan decoction by a validated UPLC/QTOF–MS/MS method

Dahuang‐mudan decoction (DMD) has been widely used for disease treatment in China for 1700 years. The formula consists of Rhubarb, moutan bark, Prunus persica, wax gourd kernel and mirabilite, which have been well studied by multidisciplinary approaches. However, the role of the mineral mirabilite in DMD is unclear. The objective of this study was to investigate the effects of mirabilite on the absorption and pharmacokinetics of the ingredients in DMD. The constituents were identified in DMD extract and the plasma of mirabilite–DMD (MDMD, 50 g kg^?1) treated rats and nonmirabilite–DMD (NMDMD, 50 g kg^?1) treated rats. The plasma was also used to investigate the effects of mirabilite on the pharmacokinetics of active ingredients in DMD using a new validated UPLC–MS/MS method. The results showed that 63 compounds were identified in the extract of DMD, 27 and 22 of which were found in the plasmas of MDMD‐ and NMDMD‐treated rats, respectively. Furthermore, the results of a pharmacokinetic study suggested that mirabilite influenced the absorption of the five constituents by decreasing the absorption of emodin and rhein while increasing the absorption of aloe‐emodin, paeoniflorin and amygdalin; the pharmacokinetic parameters, including the T_max, C_max, AUC_0–t, MRT_0–t, CLz and t_1/2 of five constituents, significantly changed in MDMD‐treated rats compared with the NMDMD. The method validation for selectivity, precision, accuracy, matrix effect, recovery and stability met the acceptance criteria. These findings uncover the roles of mirabilite in DMD and demonstrate the application of scientific principles to the study of DMD in human health care.     

Extraction of Anthraquinones from Rhubarb by a Molecularly Imprinted–Matrix Solid-Phase Dispersion Method with HPLC Detection

A simple method based on matrix solid-phase dispersion for selective extraction of anthraquinones from rhubarb samples was developed using a molecularly imprinted polymer as sorbent. The molecularly imprinted polymer was prepared using emodin as the template molecule, methacrylic acid as the functional monomer, and ethylene glycol dimethacrylate as the cross-linking agent. The polymer was characterized by scanning electron microscopy and Fourier-transform infrared spectrometry. Isothermal adsorption and dynamic adsorption experiments were performed. The best extraction conditions for anthraquinones were obtained at a ratio of molecularly imprinted polymer to sample of 1:1, a dispersion time of 5 minutes, with 5% aqueous methanol as the washing solvent, and an elution solvent of methanol-acetic acid (99:1, v/v). Once the matrix solid-phase dispersion process was optimized, the extract was reacted with 8% hydrochloric acid for hydrolysis. The anthraquinones extracted from rhubarb were determined by liquid chromatography. The detection limits of chrysophanol, emodin, physcion, and aloe-emodin were 0.23, 0.24, 0.28, and 0.27 µg mL^?1, respectively. The proposed method was compared with the method in Chinese pharmacopoeia, and the results show that the extraction yield of anthraquinones obtained by molecularly imprinted polymer–matrix solid-phase dispersion method was higher. Moreover, the proposed method is faster and simpler and can achieve extraction and purification in the same system.     

Simultaneous quantification of multiple active components from Xiexin decoction in rat plasma by LC‐ESI‐MS/MS: application in pharmacokinetics

A sensitive and specific liquid chromatography–electrospray ionization–tandem mass spectrometric (LC‐ESI‐MS/MS) method was developed and validated to simultaneously quantify 11 active compounds (coptisine, jatrorrhizine, berberine, palmatine, baicalin, baicalein, wogonoside, wogonin, rhein, emodin and aloeemodin) from Xiexin decoction (XXD) in rat plasma. Plasma samples extracted by a single‐step protein precipitation procedure were separated using the gradient mode on a Dikma ODS‐C₁₈ column. Selected reaction monitoring scanning was employed for quantification with switching electrospray ion source polarity between positive and negative modes in a single run. Calibration curves offered satisfactory linearity (r > 0.995) at linear range of 0.47–60 ng/mL for coptisine, jatrorrhizine, berberine and palmatine, 15–1930 ng/mL for baicalin, 20–2560 ng/mL for baicalein, 14–1790 ng/mL for wogonoside, 0.57–72.8 ng/mL for wogonin, 10–1280 ng/mL for rhein, 0.6–76.8 ng/mL for emodin and 3.0–384 ng/mL for aloeemodin. The intra‐ and interday precisions were less than 10.2% in terms of relative standard deviation (RSD), and the accuracies were within ±10.84% in terms of relative error (RE). It was successfully applied to the evaluation of pharmacokinetics after single oral doses of XXD were administered to rats. Copyright © 2010 John Wiley & Sons, Ltd.     

UPLC‐ESI/MS determination of 17 active constituents in two categorized formulas of traditional Chinese medicine,Sanhuang Xiexin Tang and Fuzi Xiexin Tang: application in comparing the differences in decoctions and macerations

A rapid and sensitive UPLC‐ESI/MS method was established and validated to determine 17 active constituents (aconitine, hypaconitine, mesaconitine, benzoylaconine, benzoylhypaconine, benzoylmesaconine, berberine, palmatine, jatrorrhizine, coptisine, baicalein, wogonin, baicalin, wogonoside, emodin, aloe‐emodin and rhein) in Sanhuang Xiexin Tang (SXT) and Fuzi Xiexin Tang (FXT), which are two classic compound recipes from Xiexin Tang categorized formulas in traditional Chinese medicien. The separation was performed on a UPLC BEH C₁₈ column gradient eluted using acetonitrile and 0.1% formic acid as mobile phase. ESI/MS was operated in positive ([M + H]⁺) in selected ion recording mode for analysis of alkaloids and flavones, while in negative ([M ? H]^?) selected ion recording mode for anthraquinones. All of the 17 constituents exhibited good linearity in a relatively wide concentration ranges with the lowest limits of detection of 0.38 ng/mL. All of the relative standard deviation values of intra‐ and inter‐precisions and stabilities of 17 constituents were within 5%. The method was successfully applied to determine 17 active constituents in decoctions and macerations of SXT and FXT. The results indicated that different preparative methods resulted in significant diversity in concentrations of the 17 analytes. Herb–herb interaction appeared between aconitum alkaloids in Aconiti Lateralis Radix Preparata and another three herbs. Copyright © 2013 John Wiley & Sons, Ltd.     

Screening of bioactive compounds in Rhizoma Polygoni Cuspidati with hepatocyte membranes by HPLC and LC–MS

A new method employing HPLC, LC–MS, and hepatocyte membranes for the screening of bioactive compounds in traditional Chinese medicines (TCMs) has been proposed. We hypothesized that exposure of the TCM extracts to hepatocyte membranes should decrease the concentration of membrane‐permeable compounds in the solution. Using this approach, the permeability of the compounds in Rhizoma Polygoni Cuspidati was investigated. By comparing chromatograms of samples prepared both before and after interaction with hepatocyte membranes, seven permeable compounds of Rhizoma Polygoni Cuspidati were identified. Additionally, it was found that piceid, resveratrol, emodin‐8‐β‐d‐glucoside, physcion‐8‐β‐d‐glucoside, aloe‐emodin, emodin, and physcion combined specifically with hepatocyte membranes, which might indicate a useful approach for revealing the antiatherosclerotic effects of Rhizoma Polygoni Cuspidati. Therefore, the proposed method could be a good approach to predict the potential bioactivities of multiple compounds in TCMs simultaneously. Based on the significance of these results, this method could be a novel approach for identifying potentially bioactive components in other TCMs.     

Profiling the metabolic differences of anthraquinone derivatives using liquid chromatography/tandem mass spectrometry with data‐dependent acquisition

This work concentrates on the in vitro metabolism of anthraquinone derivatives from rhubarb. Different metabolic reactions for anthraquinone derivatives were dependent on the substituent group in the skeleton. Monohydroxylation in the exocyclic methyl group was the major metabolic modification for emodin. Aloe‐emodin containing a hydroxymethyl group was mainly metabolized through a methylation reaction. For rhein, both hydrogenation and methyl substitution on the benzene ring were the major metabolic reactions. Hydroxyl substitution on the benzene ring was the predominant metabolic reaction for chrysophanol. For physcion, demethylation of the exocyclic methoxy group was the most important metabolic reaction. Copyright © 2009 John Wiley & Sons, Ltd.     

Simultaneous determination of piperaquine and its N‐oxidated metabolite in rat plasma using LC‐MS/MS

A sensitive and efficient liquid chromatography tandem mass spectrometry method was developed and validated for the simultaneous determination of piperaquine (PQ) and its N ‐oxidated metabolite (PQ‐M) in plasma. A simple protein precipitation procedure was used for sample preparation. Adequate chromatographic retention was achieved on a C₁₈ column under gradient elution with acetonitrile and 2 mm aqueous ammonium acetate containing 0.15% formic acid and 0.05% trifluoroacetic acid. A triple‐quadrupole mass spectrometer equipped with an electrospray source was set up in the positive ion mode and multiple reaction monitoring mode. The method was linear in the range of 2.0–400.0 ng/mL for PQ and 1.0–50.0 ng/mL for PQ‐M with suitable accuracy, precision and extraction recovery. The lower limits of detection (LLOD) were established at 0.4 and 0.2 ng/mL for PQ and PQ‐M, respectively, using 40 μL of plasma sample. The matrix effect was negligible under the current conditions. No effect was found for co‐administrated artemisinin drugs or hemolysis on the quantification of PQ and PQ‐M. Stability testing showed that two analytes remained stable under all relevant analytical conditions. The validated method was successfully applied to a pharmacokinetic study performed in rats after a single oral administration of PQ (60 mg/kg).     

A DFT study on the radical scavenging activity of hydroxyanthraquinone derivatives in rhubarb

The structural and electronic properties of hydroxyanthraquinone derivatives in rhubarb, namely, chrysophanol, emodin, physcion, aloe‐emodin, rhein, and their radicals were investigated at density functional level. The bifurcated hydrogen bond property of the studied structures was investigated using the atoms in molecules theory. It turned out that the presence of the dihydroxy functionality increases the radical stability through hydrogen bonds formation and favors hydrogen atom abstraction. Bond dissociation energy and ionization potential were also determined to know if the radical scavenging activity of these compounds proceeds via an H‐atom or an electron‐transfer mechanism. © 2010 Wiley Periodicals, Inc. Int J Quantum Chem, 2011     

Preparation of magnetic metal-organic framework MIL-101(Fe) and its application in the extraction of anthraquinones in rhubarb

In this work, a magnetic octahedral metal-organic framework (Fe₃O₄@NH₂-MIL-101(Fe)) was synthesized for the magnetic solid-phase extraction of three anthraquinones, including aloe-emodin, emodin, and physcion, in rhubarb. The Fe₃O₄@NH₂-MIL-101(Fe) exhibits a high specific surface area of 259.2 m²/g with an average pore size of 6.0 nm and high magnetic responsivity of 23.4 emu/g, which may be used as an adsorbent for rapid preconcentration and separation of target analytes. The main parameters for magnetic solid-phase extraction of anthraquinones, including the amount of adsorbent, extraction time, extraction temperature, extraction pH, elution solvent, and elution time, were systematically optimized. The whole extraction process requires a very low amount of adsorbent and a small volume of the sample. Besides, under the optimized conditions, the method shows satisfactory spiked recovery for anthraquinones in the range of 93.3–109.1% and the limits of detection are 1.7–3.4 ng/mL. The relative standard deviations for intra- and inter-day precision are 0.2–1.3% and 0.2–0.6%, respectively. The experimental results indicate that the developed method is feasible for the analysis of anthraquinones in rhubarb.     

An improved LC‐MS/MS method for simultaneous determination of 1,5‐dicaffeoylquinic acid and its active metabolites in human plasma and its application to a pharmacokinetic study in patients

In this study, a sensitive, selective and reproducible liquid chromatography–tandem mass spectrometry method for the simultaneous determination of 1,5‐dicaffeoylquinic acid (1,5‐DCQA) and its active metabolites, 1‐caffeoyl‐5‐feruoylquinic acid and 1,5‐O‐diferuoylquinic acid, in human plasma, using puerarin as internal standard, was developed and validated. Analytes were extracted from plasma samples by liquid–liquid extraction with ethyl acetate, separated on a C₁₈ reversed‐phase column with water containing 5 mM ammonium acetate and acetonitrile as the mobile phase and detected by electrospray ionization mass spectrometry in negative selected reaction monitoring mode. The accuracy and precision of the method were acceptable and linearity was good over the range 1–200 ng/mL for each analyte. In addition, the selectivity, extraction recovery and matrix effect were satisfactory too. The validated LC‐MS/MS method was successfully applied to phase II clinical pharmacokinetic study of 1,5‐DCQA in patients. Copyright © 2010 John Wiley & Sons, Ltd.     

Qualitative and quantitative analysis of major constituents from Dazhu Hongjingtian capsule by UPLC/Q‐TOF‐MS/MS combined with UPLC/QQQ‐MS/MS

In this work, a sensitive and efficient method was established and validated for qualitative and quantitative analysis of major bioactive constituents in Dazhu Hongjingtian capsule by liquid chromatography tandem mass spectrometry. A total of 32 compounds were tentatively identified using ultra‐performance liquid chromatography coupled with quadrupole time‐of‐flight mass spectrometry. Furthermore, 12 constituents, namely gallic acid, 3,4‐dihydroxybenzoic acid, salidroside, p‐ coumaric acid‐4‐O ‐β ‐d ‐glucopyranoside, bergeninum, 4‐hydroxybenzoic acid, 4‐hydroxyphenylacetic acid, syringate, 6′′‐O ‐galloylsalidroside, rhodiosin, rhodionin and kaempferol‐7‐O ‐α ‐l ‐rhamnoside, were simultaneously quantified by the developed ultra‐performance liquid chromatography coupled with a triple quadrupole mass spectrometry method in 9 min. All of them were analyzed on an Agilent ZorBax SB‐C₁₈ column (3.0 × 100 mm, 1.8 μm) with linear gradient elution of methanol–0.1% formic acid water. The proposed method was applied to analyze three batches of samples with acceptable linearity (R , 0.9979–0.9997), precision (RSD, 1.3–4.7%), repeatability (RSD, 1.7–4.9%), stability (RSD, 2.2–4.9%) and recovery (RSD, 0.6–4.4%) of the 12 compounds. As a result, the analytical method possessing high throughput and sensitivity is suitable for the quality control of Dazhu Hongjingtian capsule.