Simultaneous determination of ginsenoside Rb1, naringin, ginsenoside Rb2 and oridonin in rat plasma by LC–MS/MS and its application to a pharmacokinetic study after oral administration of Weifuchun tablet

doi:10.1016/j.jchromb.2015.06.027

Journal of Chromatography B

Volume 1000, 1 September 2015, Pages 112-119

https://doi.org/10.1016/j.jchromb.2015.06.027 Get rights and content

Highlights

•
An analytical strategy via ESI source in a novel detection mode was developed.
•
Four compounds in rat plasma were simultaneously determined.
•
Comparisons in pharmacokinetic parameters were firstly reported.
•
Pharmacokinetic profile of Weifuchun tablet was firstly characterized.

Abstract

A sensitive, specific and rapid liquid chromatography–mass spectrometry (LC–MS) method was developed and validated for analysis of ginsenoside Rb₁, naringin, ginsenoside Rb₂ and oridonin in rat plasma using sulfamethoxazole as an internal standard (IS). Separation was conducted out on an Agilent Eclipse XDB C18 column with liner gradient elution using acetonitrile (A) and 0.1% aqueous acetic acid (B). A tandem mass spectrometric detection was conducted using multiple reaction monitoring (MRM) via an electrospray ionization (ESI) source. A novel multi-determination-periods program was executed to achieve a higher sensitivity by setting three scanning periods. All analytes exhibited good linearity within the concentration range (r > 0.9973). The lower limits of quantitation (LLOQ) of ginsenoside Rb₁, naringin, ginsenoside Rb₂ and oridonin were 2.64, 4.32, 2.32 and 1.56 ng/mL, respectively. Intra-day and inter-day precisions of the investigated components exhibited an RSD within 8.3%, and the accuracy (RE) ranged from −8.6% to 6.0% at all quality control levels. The developed method was successfully applied to a pharmacokinetic study of ginsenoside Rb₁, naringin, ginsenoside Rb₂ and oridonin in rats after oral administration of a Weifuchun tablet.

Introduction

The Weifuchun tablet, a well-known Chinese herbal preparation, is composed of three herbs: Red Ginseng, Isodon amethystoides and Fructus Aurantii. As recorded in China Pharmacopoeia [1], it has the following effects of spleen qi, promoting blood and detoxification. In clinics, it has been widely used for the treatment of early gastric cancer, chronic gastritis, precancerous lesions and post-operative adjuvant treatment of gastric cancer [2]. The chemical constituents of the Weifuchun tablet mainly include ginsenosides (including ginsenoside Rb₁ and Rb₂, etc.), diterpenoids (including oridonin and epinodosinol, etc.) and flavonoid glycosides (including naringin, etc.), etc [3].

Ginsenoside Rb₁ and Rb₂ are the characteristic constituents in Red Ginseng and they could partly represent the effects of the herb ginseng [4], [5]. Naringin is a flavonoid present in many citrus fruits and traditional Chinese medicines. Like most flavonoids, naringin has anti-inflammatory, anti-ulcer and antioxidant activities [6], [7], [8]. Diterpenoids from Isodon amethystoides, which includes oridonin and epinodosinol, etc., have been demonstrated as the major bioactive ingredients, which are usually chosen as chemical markers [9], [10], [11]. Hence, simultaneous determination of these compounds in rat plasma is necessary to characterize the pharmacokinetic profile of this drug. Ginsenoside Rb₁, naringin, ginsenoside Rb₂ and oridonin as the major constituents have been used as the markers for quality control of the Weifuchun tablet [12]. Earlier reported methods for the determination of some of these compounds in plasma samples consisted of the high-performance liquid chromatography coupled with ultraviolet (UV) detection (HPLC–UV) [13], [14], [15], liquid chromatography tandem–mass spectrometry (HPLC–MS/MS) [16], [17], ultraperformance liquid chromatography tandem mass spectrometry (UPLC–MS/MS) [18], [19] and ultra-fast liquid chromatography tandem mass spectrometry (UFLC–MS/MS) [20] methods. However, to our knowledge, there is no report on the simultaneous determination of ginsenoside Rb₁, naringin, ginsenoside Rb₂ and oridonin of the Weifuchun tablet in rat plasma for pharmacokinetic studies. Therefore, it was deemed necessary to develop a sensitive and reliable method for the determination of these analytes in bio-samples and to obtain the pharmacokinetic properties for further clinical application.

In the present study, a novel analytical strategy was employed and a sensitive and selective HPLC–ESI–MS method was developed and validated for the simultaneous determination of ginsenoside Rb₁, naringin, ginsenoside Rb₂ and oridonin in rat plasma. Using this method, the pharmacokinetic profiles of these four components were further investigated after a single oral administration of the Weifuchun tablet.

Section snippets

Chemicals and reagents

HPLC grade acetonitrile was purchased from J.T.Backer (Center Valley, USA) and HPLC grade formic acid was obtained from Dikma Technologies Inc. (Lake Forest, USA). The methanol used in sample preparation was of analytical grade and purchased from Yongda Chemical Reagent Co., Ltd. (Tianjin, China). Purified water was supplied by Wahaha Group Co., Ltd. (Hangzhou, China).

Ginsenoside Rb₁, naringin, ginsenoside Rb₂, oridonin and sulfamethoxazole were purchased from the National Institute for Food

Mass spectrometry

Ginsenoside Rb₁, naringin, ginsenoside Rb₂ and oridonin were initially characterized according to their mass spectra using syringe pump infusion analysis. Meanwhile, their precursor ions and product ions were ascertained for use in MRM. ESI was employed so that good sensitivity and fragmentation could be obtained. It was also found that all of the analytes and the IS could be ionized under positive and negative ESI conditions. Although the four analytes and IS could be ionized under positive or

Conclusions

The method in this study utilizes a novel determination strategy, enabling the application of positive and negative ESI–MS in a single run. A bioanalytical method involving an effective sample clean-up procedure was successfully applied to a pharmacokinetic study of the analytes after oral administration of the Weifuchun tablet extract for simultaneous determination of ginsenoside Rb₁, naringin, ginsenoside Rb₂ and oridonin in rat plasma for the first time. The proposed novel HPLC–MS/MS method

Acknowledgements

This work received financial support from the National Natural Science Foundation of China (81402894) and the Scientific and Technological Project of Hebei Province-CSPC Pharmaceutical Joint Research Fund, China (No. C2011206174). We also thank the School of Pharmacy of Hebei Medical University for providing instruments for this work.

References (22)

N. Tung et al.
Dammarane—type saponins from the flower buds of Panax ginseng and their effects on human leukemia cells
Bioorg. Med. Chem. Lett.
(2010)
Y.R. Jin et al.
Simultaneous quantification of 19 diterpenoids in Isodon amethystoides by high-performance liquid chromatography–electrospray ionization tandem mass spectrometry
J. Pharmaceut. Biomed. Anal.
(2010)
Q.F. Xu et al.
Pharmacokinetics and bioavailability of ginsenoside Rb₁ and Rg₁ from Panax notoginseng in rats
J. Ethnopharmacol.
(2003)
Y.H. Mei et al.
An HPLC method for determination of oridonin in rabbits using isopsoralen as an internal standard and its application to pharmacokinetic studies for oridonin-loaded nanoparticles
J. Chromatogr. B
(2008)
H.Y. Ji et al.
Simultaneous determination of ginsenoside Rb₁ and Rg₁ in human plasma by liquid chromatography–mass spectrometry
J. Pharmaceut. Biomed. Anal.
(2004)
L. Tong et al.
Simultaneous determination of naringin, hesperidin, neohesperidin, naringenin and hesperetin of Fractus aurantii extract in rat plasma by liquid chromatography–tandem mass spectrometry
J. Pharmaceut. Biomed. Anal.
(2012)
Q.L. Guo et al.
Brain distribution pharmacokinetics and integrated pharmacokineticsof panax notoginsenoside R₁, ginsenosides Rg₁, Rb₁, Re and Rd inrats after intranasal administration of panax notoginseng saponins assessed by UPLC/MS/MS
J. Chromatogr. B
(2014)
J. Wen et al.
UPLC–MS/MS determination of paeoniflorin, naringin, naringenin and glycyrrhetinic acid in rat plasma and its application to a pharmacokinetic study after oral administration of Si Ni San decoction
J. Pharmaceut. Biomed. Anal.
(2012)
H.Y. Zhu et al.
Simultaneous determination of two iri twoanthraquinones and four flavonoid glycosides of Zhi–Zi–Da–Huangdecoction in rat plasma by UFLC–MS/MS: Application to a comparative pharmacokinetic study in normal and cholestatic liver injury rats
J. Chromatogr. B
(2014)
T.Z. Fang et al.
A rapid LC/MS/MS quantitation assay for naringin and its two metabolites in rats plasma
J. Pharmaceut. Biomed. Anal.
(2006)

J. Zhao et al.

Determination of ginsenosides Rb₁, Rb₂, and Rb₃ in rat plasma by a rapid and sensitive liquid chromatography tandem mass spectrometry method: Application in a pharmacokinetic study

J. Pharmaceut. Biomed. Anal.

(2012)

Cited by (21)

Boronate affinity-based surface molecularly imprinted polymer microspheres using polyethyleneimine/dopamine coating for efficient selective recognition and separation of Ginsenoside Rb1
2024, Reactive and Functional Polymers
A novel approach combining boronate affinity with surface molecular imprinting was developed for the specific recognition and separation of ginsenoside Rb1. Firstly, the boronic acid-functionalized polymer microspheres were prepared as inner cores via suspension polymerization using methyl methacrylate, ethyleneglycol dimethacrylate, and 4-vinylphenyl boronic pinacol ester (4-VBPE) as polymerization monomers. Then, the Ginsenoside Rb1 (GRb1) template was covalently pre-anchored onto the surface of microspheres through boronate affinity binding after the hydrolysis of 4-VBPE. Finally, dopamine and polyethyleneimine were used as co-coating monomers to form a thickness-controllable imprinting layer onto the polymer microspheres via in-water self-copolymerization. The key factors affecting specific recognition, including the boronic acid content, adsorption media, and polymerization time were carefully optimized. The resulting molecularly imprinted microspheres (MIMs) exhibited strong surface hydrophilicity. In the aqueous media, the optimal adsorption capacity for the template reached 81.45 μmol g⁻¹, and the imprinting factor was 5.32. The specific evaluation showed that the MIMs had a much higher selectivity for GRb1 than its analogues, and the selectivity coefficients were all higher than 2.4. In addition, the MIMs can be applied as adsorbents for solid-phase extraction to selectively separate ginsenosides from American ginseng extract. The eluting recovery ratios of GRb1, GRg2, and GRd reach 92.39%, 88.90%, and 92.49%, respectively.
Citri Reticulatae Pericarpium (Chenpi): A multi-efficacy pericarp in treating cardiovascular diseases
2022, Biomedicine and Pharmacotherapy
Citation Excerpt :
The pharmacokinetics of all four main active ingredients in CRP (naringin (Table 1), naringenin (Table 2), hesperidin, and hesperetin) have been well investigated. Naringin can be promptly absorbed by oral administration (p.o.) and was detectable in rats in 15 min [31]. In studies with rabbits, oral naringenin was significantly more rapidly absorbed with a higher Cmax but a much shorter residence time compared to naringin [32].
Citri Reticulatae Pericarpium (CRP) has been utilized as a versatile medicinal herb with wide cardiovascular benefits in Asian nations for centuries. Accumulating evidence suggests that CRP and its components are effective in preventing cardiovascular diseases (CVDs) such as atherosclerosis, myocardial infarction, myocardial ischemia, arrhythmia, cardiac hypertrophy, heart failure, and hypertension. Studies show that the two most bioactive components of CRP are flavonoids and volatile oils. The cardiovascular protective effects of CRP have attracted considerable research interest due to its hypolipidemic, antiplatelet activity, antioxidant and anti-inflammatory effects. Hereby, we provide a rigorous and up-to-date overview of the cardiovascular protective properties and the potential molecular targets of CRP, and finally highlight the pharmacokinetics and the therapeutic potential of the main pharmacologically active components of CRP to treat CVDs.
Ginsenoside Rb2: A review of pharmacokinetics and pharmacological effects
2022, Journal of Ginseng Research
Ginsenoside Rb2 is an active protopanaxadiol-type saponin, widely existing in the stem and leave of ginseng. Rb2 has recently been the focus of studies for pharmaceutical properties. This paper provides an overview of the preclinical and clinical pharmacokinetics for Rb2, which exhibit poor absorption, rapid tissue distribution and slow excretion through urine. Pharmacological studies indicate a beneficial role of Rb2 in the prevention and treatment of diabetes, obesity, tumor, photoaging, virus infection and cardiovascular problems. The underlying mechanism is involved in an inhibition of oxidative stress, ROS generation, inflammation and apoptosis via regulation of various cellular signaling pathways and molecules, including AKT/SHP, MAPK, EGFR/SOX2, TGF-β1/Smad, SIRT1, GPR120/AMPK/HO-1 and NF-κB. This work would provide a new insight into the understanding and application of Rb2. However, its therapeutic effects have not been clinically evaluated. Further studies should be aimed at the clinical treatment of Rb2.
Polysaccharide deriving from Ophiopogonis Radix promotes metabolism of ginsenosides in the present of human gut microbiota based on UPLC-MS/MS assay
2019, Journal of Pharmaceutical and Biomedical Analysis
Citation Excerpt :
The sensitivity was evaluated by determination of LODs and LLOQs for the ginsenosides (Table 2). All ginsenosides showed a LLOQ lower than 0.48 μM (range of 0.20-0.48 μM), i.e. 44 pg per test calculated by injection volume of 2 μL, this value was comparable with previous studies for determination of ginsenosides in plasma samples [27,28] and in fermented broth [22]. The intra- and inter-assay performance was assessed by analyzing 3-level QC samples, representing the entire calibration range.
The combined usage of Ginseng Radix et Rhizoma (ginseng) and Ophiopogonis Radix is common in oriental countries for thousands of years. The major active constituents of ginseng are ginsenosides, and the conversion of ginsenosides to different metabolites by gut microbiota has been reported. However, the effect of Ophiopogonis Radix, especially its polysaccharides, on the metabolism of ginsenosides by gut microbiota is not known. Here, an in vitro metabolism of ginseng extract, or ginsenosides, in combination with or without Ophiopogon polysaccharide was conducted. A sensitive and reliable UPLC-MS/MS approach using multiple reaction monitoring (MRM) in positive ion mode was developed simultaneously to quantify 22 ginsenosides in the broth of gut microbiota. After fermentation with the microbiota, 15 ginsenosides were detected and quantified, including 6 primary ginsenosides, i.e. Rb₁, Rc, Rb₂, Rb₃, Rd and Re, and 9 metabolites, i.e. F₂, Rg₃, compound K, Rh₂, PPD, Rg₁, Rh₁, Rg₂ and PPT. The quantitative results therefore revealed the elimination of primary ginsenosides and the formation of their metabolites in time-dependent manners. Furthermore, Ophiopogon polysaccharide was shown to stimulate the metabolism of ginsenosides, triggered by gut microbiota. Our study can be extended to investigate the metabolism of different Panax species by gut microbiota when combining with other herbs.
Pharmacokinetic study of ginsenoside Re after vaginal administration in rabbits by UPLC-MS/MS determination
2018, Chinese Herbal Medicines
Citation Excerpt :
GRe was regarded to be an active component of ginseng extracts. Despite various well-known pharmacological effects (Gao, Wang, Luo, & Gao, 2017; Joo et al., 2010; Miao, Metcalfe, Hao, & March, 2002), pharmacokinetic behaviors of GRe in vaginal administration were lack of understanding (Bae, Shin, & Kim, 2005; Jin, Tian, Ma, Xu, & Du, 2015). Therefore, considering all these factors, we selected Xiaomi suppository which consists of GRe as a representative component of the medicine.
A selective and sensitive ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was employed to study the pharmacokinetics of ginsenoside Re (GRe) in rabbits after vaginal administration of Xiaomi suppository to evaluate the systemic exposure of the suppository for the local treatment.
Chromatographic separation was on an ACQUITY UPLC^® BEH C₁₈ column, and acetonitrile-0.1% formic acid was used as mobile phase in gradient elution. The plasma samples were deproteinized by acetonitrile, and the pharmacokinetic parameters were calculated by Winnonlin 6.4.
Calibration curve of GRe showed a good linearity over the concentration range from 5 ng/mL to 500 ng/mL (r = 0.9999). The low limit of quantification of 5 ng/mL could satisfy the experimental requirement. The intra-day and inter-day precision of GRe at three concentrations were less than 1.96%, and the average recoveries of GRe were more than 64.0%. The pharmacokinetic parameters for vaginal administration were as follows: T_max, 0.5 h; C_max, 20.88 ng/mL; AUC_0-t, 64.71 h · ng/mL and the residence time was 3.06 h. By using deconvolution calculation method, the cumulative absorption fraction of GRe was about 0.89%.
The systemic exposure of GRe was minimal after vaginal administration of Xiaomi suppository.
Identification and comparative oridonin metabolism in different species liver microsomes by using UPLC-Triple-TOF-MS/MS and PCA
2016, Analytical Biochemistry
Citation Excerpt :
ORI is expected to be examined as a new anti-tumor agent because of its strong inhibitory effect on multiple tumor cells and lower toxicity to humans [22–24]. Currently, ORI has been investigated in pharmacology and toxicology studies in the laboratory [25,26]. These data suggest that ORI can be soon metabolized in vivo, and the drug is mainly eliminated in the form of metabolites.
Oridonin (ORI) is an active natural ent-kaurene diterpenoid ingredient with notable anti-cancer and anti-inflammation activities. Currently, a strategy was developed to identify metabolites and to assess the metabolic profiles of ORI in vitro using ultra-high-performance liquid chromatography-Triple/time-of-flight mass spectrometry (UPLC-Triple-TOF-MS/MS). Meanwhile, the metabolism differences of ORI in the liver microsomes of four different species were investigated using a principal component analysis (PCA) based on the metabolite absolute peak area values as the variables. Based on the proposed methods, 27 metabolites were structurally characterized. The results indicate that ORI is universally metabolized in vitro, and the metabolic pathway mainly includes dehydration, hydroxylation, di-hydroxylation, hydrogenation, decarboxylation, and ketone formation. Overall, there are obvious inter-species differences in types and amounts of ORI metabolites in the four species. These results will provide basic data for future pharmacological and toxicological studies of ORI and for other ent-kauranes diterpenoids. Meanwhile, studying the ORI metabolic differences helps to select the proper animal model for further pharmacology and toxicological assessment.

View all citing articles on Scopus

View full text

Simultaneous determination of ginsenoside Rb1, naringin, ginsenoside Rb2 and oridonin in rat plasma by LC–MS/MS and its application to a pharmacokinetic study after oral administration of Weifuchun tablet

Highlights

Abstract

Introduction

Section snippets

Chemicals and reagents

Mass spectrometry

Conclusions

Acknowledgements

Bioorg. Med. Chem. Lett.

J. Pharmaceut. Biomed. Anal.

J. Ethnopharmacol.

J. Chromatogr. B

J. Pharmaceut. Biomed. Anal.

J. Pharmaceut. Biomed. Anal.

J. Chromatogr. B

J. Pharmaceut. Biomed. Anal.

J. Chromatogr. B

J. Pharmaceut. Biomed. Anal.

J. Pharmaceut. Biomed. Anal.

Simultaneous determination of ginsenoside Rb₁, naringin, ginsenoside Rb₂ and oridonin in rat plasma by LC–MS/MS and its application to a pharmacokinetic study after oral administration of Weifuchun tablet