Compounds In Rat Plasma Research Articles

Pharmacokinetics of the material basis compounds of Tianshu capsule in treating migraine

Ethnopharmacological relevanceTianshu capsule (TSC) is a traditional Chinese medicine (TCM) preparation and has significant clinical effect on migraine. The composition characteristics of TSC formula are worth exploring for the treatment of migraine. Aim of the studyIdentify the compounds in vivo after oral administration of TSC, Gastrodin (GAS), ferulic acid, senkyunolide G and senkyunolide I in the blood of healthy and migraine rats were used as representative compounds for time-dependent processes investigation. And we focus on explaining the characteristics of TSC treatment on migraine from a pharmacokinetic perspective. Materials and methodsIn this study, Ultra-high performance liquid chromatography with Orbitrap tandem mass spectrometry (UPLC-Orbitrap-MS/MS) system were used to detect the compounds in rat plasma and brain after oral administration of TSC. A sensitive, selective and reliable ultra-performance liquid chromatography coupled with triple quadrupole mass spectrometry (UPLC-TQ-MS/MS) system method was established for the simultaneous quantitative analysis of multi-components and evaluate the pharmacokinetic behavior of four main compounds. ResultsA total of 46 compounds were significantly identified in vivo of rat, including 35 compounds in plasma, 7 compounds only in brain and 4 compounds both in plasma and brain. The quantitative method has been confirmed to be feasible for multi-components content determination, pharmacokinetic parameters indicated that the four compounds absorbed into blood rapidly, and senkyunolide I also cross the blood-brain barrier (BBB) quickly into brain. GAS has relatively high concentrations in plasma, and the parameters AUC0-t, CLz/F displayed significant differences between the normal and migraine rats. And the AUC0-t and Cmax of GAS and ferulic acid exhibited dose-dependent way. Conclusionswe added compounds for the pharmacokinetic study of TSC, providing powerful help for clinical medication in the treatment of migraine.

Development and validation of a liquid chromatography-tandem mass spectrometry method for simultaneous quantification of eight Xiakucao Oral liquid-related compounds in rat plasma and its application in pharmacokinetic study.

Xiakucao Oral Liquid (XKCOL) has been widely used for treating mammary gland hyperplasia and goiter in China. However, its pharmacokinetic data have been missing to date. To conduct its pharmacokinetic study, we established an LC-tandem mass spectrometry method for the simultaneous determination of eight XKCOL-related compounds in rat plasma. Liquid-liquid extraction was used for the sampling process. Chromatographic separation was performed on a Phenomenon Luna C18 column with a mobile phase of methanol and 2 mM ammonium acetate, using gradient elution at a flow rate of 0.8 mL/min. Detection was performed in the multiple reaction monitoring mode using negative electrospray ionization (ESI-) with optimized MS parameters. Endogenous substances and carryover did not interfere in the detection of analytes. The calibration curves showed a good linear relationship within the linear ranges. The intra- and inter-batch accuracy and precision were 94.8%-110.0% and ≤11.2%, respectively. There was no significant matrix effect and the recovery was reproducible. The dilution of samples did not affect the accuracy and precision. The solution and plasma samples were stable under the various test conditions. The major components of XKCOL absorbed into the blood were salvianic acid A and rosmarinic acid. They demonstrated linear kinetics over the dose range used in this study.

Determination of fraxetin, fraxin and dimethylfraxetin in rat plasma by UPLC-MS/MS

AbstractObjectiveThe levels of fraxetin, fraxin, and dimethylfraxetin in rat plasma to be measured using an ultra-performance liquid chromatography tandem mass-spectrometry (UPLC–MS/MS) technique and applied to their pharmacokinetics and bioavailability.MethodsThe protein precipitation technique was applied to the plasma preparation using acetonitrile and methanol (9:1, v/v). At a flow rate of 0.4 mL min−1, the elution time was 6 min. The mobile phase consisted of acetonitrile-water with 0.1% formic acid, and the chromatographic column was UPLC HSS T3 (50 mm × 2.1 mm, 1.8 μm). Quantitative analysis was conducted using multiple reaction monitoring (MRM) mode and detection was performed using electrospray ionization (ESI) positive ion mode. In each group, six rats were treated with fraxetin, fraxin, and dimethylfraxetin either orally (5 mg kg−1) or intravenously (1 mg kg−1).ResultsThe calibration curves showed good linearity in the range of 2–4,000 ng mL−1, where r was greater than 0.99. The bioavailability of dimethylfraxetin, fraxin, and fraxetin was determinated to be 19.7, 1.4, and 6.0%.ConclusionThe established UPLC-MS/MS method for determining the levels of these three compounds in rat plasma was successfully applied to the pharmacokinetics of dimethylfraxetin, fraxin, and fraxetin, and the bioavailability was calculated.

Pharmacokinetics of four tannin compounds from Sanguisorba officinalis L. before and after processing by ultra-high-performance liquid chromatography-tandem mass spectrometry.

Sanguisorba officinalis L. possesses detoxifying, analgesic, and hemostatic properties. After charred processing, S. officinalis exhibits significantly enhanced medicinal effects. Currently, most pharmacokinetic studies focus on the chemical constituents of unprocessed S. officinalis. There is limited research on the comparison of chemical constituents before and after processing. This study established a pharmacokinetic method using ultra-high-performance liquid chromatography-tandem mass spectroscopy (UHPLC-MS/MS) to simultaneously determine the levels of four tannin compounds in rat plasma. In negative ion mode, MS/MS detection was performed using an electrospray ionization source. Chromatographic separation was performed using WATERS ACQUITY HSS T3 column (2.1 × 100mm, 1.8µm) with a gradient elution of water and acetonitrile as the mobile phase. The pharmacokinetic results indicate that all four compounds reached peak concentrations within 2h, demonstrating rapid absorption into the bloodstream within the gastrointestinal tract. Notably, the absorption was generally faster in the charred compound of S. officinalis after processing. These four compounds exhibited slower elimination in rat plasma, while in S. officinalis charcoal, the compounds were eliminated more rapidly. The pharmacokinetic results have revealed the pharmacokinetic characteristics of the four analytes in rat plasma which provides valuable reference information for further investigating the in vivo absorption process of S. officinalis after processing.

A strategy of accuracy quantification by extending the concentration monitoring coverage based on online double collision energy of ultra-high performance liquid chromatography tandem mass spectrometry: The pharmacokinetics of Toddalia asiatica as a case study

To facilitate the safety, efficacy and rationality of clinical application of traditional Chinese medicines (TCMs), pharmacokinetic research played an indispensable role. The key challenge during pharmacokinetic investigation lied at the substantial fluctuation of compound concentrations in the plasma over the course of absorption. Taking the pharmacokinetics of six compounds after administration of Toddalia asiatica (TA) as an example, an efficient strategy was established by introducing the online double collision energy (ODCE) into the quantification process applying ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS). During the analytical program, double collision energy (DCE) was optimized to establish the dual calibration curve (DCC) with large concentration monitoring coverage (CMC) for meeting the wide content range of certain target compounds. Method validation test was performed in terms of linearity, precision, sensitivity, matrix effect, recovery, etc. The results displayed that the CMC of todarolactone with high exposure in plasma was extended from 1.25–2,500 ng/mL to 1.25–125,000 ng/mL. Furthermore, a rapid UHPLC-MS/MS method integrated with ODCE was successfully applied to the determination of six compounds in rat plasma, revealing an extremely high plasma concentration of todarolactone (16,662 ng/mL). This strategy could expand the range of quantification while retaining extraordinary sensitivity. Consequently, it could be a fit-for-purpose strategy to quantify compounds over a wide concentration range for in vivo process monitoring.

Systematic characterization of the chemical constituents in vitro and in vivo of Qianghuo by UPLC-Q-TOF-MS/MS

The Chinese herb Qianghuo is an antiphlogistic herb with many effects and complex components. In this study, the chemical composition of Qianghuo and its components in rat plasma after oral administration were investigated using ultra-high-performance liquid chromatography coupled with quadrupole-time-of-flight tandem mass spectrometry (UPLC-Q-TOF-MS/MS). The extracts, blank plasma, and plasma containing the drug were analyzed by mass spectrometry, and data collected in both positive and negative ion modes were analyzed using Masslynx software, and the structures were confirmed by combining the compound fragment ions and mass spectrometry cleavage pathways. A total of 62 in vitro chemical components were identified, including 27 coumarins, 18 organic acids, 5 amino acids, 5 glycosides, 2 flavonoids, 4 nucleotides, and 1 ester, which were summarized from the obtained compounds in terms of their possible cleavage patterns. Among the identified 31 compounds in rat plasma, 21 were prototypes, mostly coumarins, organic acids, and flavonoids, and 10 were metabolites, which were mainly generated via hydroxylation and methylation pathways. Based on these, this study provides a theoretical foundation for quality control and basic research on Qianghuo medicinal substances.

Identification of the absorbed constituents and metabolites of Huangqi Guizhi Wuwu decoction by ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry.

Huangqi Guizhi Wuwu decoction (HGWWD) is a widely used traditional Chinese medicine (TCM) preparation for the treatment of ischemic stroke and diabetes peripheral neuropathy. However, the material basis for the efficacy of HGWWD remains unclear. In this study, a rapid, sensitive and selective ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS) method was developed to separate and identify the absorbed components and metabolites of HGWWD in rat plasma after oral administration for the first time. By comparing the retention time, high-resolution mass spectrometry primary and secondary mass spectrometry data of blank plasma and drug-containing plasma, a total of 42 constituents, including 24 prototype compounds and 18 metabolites, were identified or tentatively characterized. The results indicated that monoterpenes, flavonoids, organic acids, amino acids, gingerols and alkaloids were main prototype compounds in rat plasma, and flavonoid-related metabolites, organic acid-related metabolites and gingerol-related metabolites were major metabolites. It is concluded the developed UHPLC-Q-TOF-MS method with high sensitivity and resolution is suitable for identifying and characterizing the absorbed components and metabolites of HGWWD, and the results will provide important data for further study on the relationship between the chemical constituents and pharmacological activities of HGWWD.

Studies on chemical profiling and pharmacokinetics of traditional Chinese medicine Formula Kang Shuai Lao Pian

Kang Shuai Lao Pian (KSLP) is a traditional Chinese medicine (TCM) preparation used to delay aging. However, due to the lack of research on the chemical composition and pharmacokinetic behavior of KSLP, its material basis and in vivo components with high exposure remain ambiguous. The UPLC/Q-Orbitrap-MS/MS was performed to identify the preliminary chemical profile of KSLP. A total of 138 compounds, including ginsenosides, phenylethanol glycosides, iridoids, alkaloids, ionones and others, were identified in accordance with their retention times, accurate masses and characteristic MS/MS fragment patterns. Moreover, considering the active components and characteristic components of KSLP, the extraction process of KSLP was optimized, and the quantitative analysis by UPLC/QQQ-MS/MS of 13 compounds in KSLP was established. The method was stable and sensitive, and could be used for the quality control of KSLP. Then, the pharmacokinetic study was carried out by further refining the components of KSLP. Besides, quantitative method for 6 compounds in rat plasma was validated and developed by UPLC/QQQ-MS/MS. The established approach was successfully applied to characterize the pharmacokinetic features of components in KSLP and it was found that the absorption and elimination of ginsenosides in KSLP was slow. Altogether, this study laid a solid foundation and provided theoretical guidance for further clarification of bioactive components of KSLP.

A pharmacokinetic study on twenty-one compounds in rat plasma by integrating UPLC-QQQ-MS/MS with GC-MS after oral administration of Suxiao Jiuxin pill

Suxiao Jiuxin pill (SX) was a commonly used traditional Chinese medical preparation for treating acute myocardial infarction. However, fewer studies have been reported for comprehensively investigating the pharmacokinetic of the non-volatile and volatile chemical constituents in SX. For current study, a comprehensive and sensitive quantitation of twenty-one compounds in rat plasma was developed and validated by integrating UPLC-QQQ-MS/MS with GC-MS after oral administration of SX. The pharmacokinetic study was performed under acceptable specificity, linearity, sensitivity, precision and accuracy, recovery, matrix effect and stability. Ultimately, borneol, isoborneol, senkyunolide A, senkyunolide Ⅰ, ligustilide, ferulic acid, neocnidilide, senkyunolide H and levistolide A were detected. Their ranges of Tmax, Cmax, T1/2z and AUC (0-∞) were 0.07 ∼ 0.33 h, 5.24 ∼ 683.35 μg/L, 0.29 ∼ 1.08 h and 6.55 ∼ 474.62 μg/L*h, respectively. This study would be meaningful in better revealing the therapeutic material basis of SX. Furthermore, it could provide reference on pharmacological effect of SX against acute myocardial infarction.

In vivo Metabolism of Nifurtimox and the Drug-Drug Interaction Potential Including its Major Metabolites.

Nifurtimox is an effective treatment for patients with Chagas disease, but knowledge of its biotransformation and excretion is limited. This study aimed to better understand the fate of oral nifurtimox in vivo. We investigated the exposure and excretion pathways of [14C]-labeled nifurtimox and its metabolites in rats. We then quantified the prominent metabolites and nifurtimox in the urine and plasma of patients receiving nifurtimox using LC-HRMS with reference standards and quantified these compounds in rat plasma after a single, high dose of nifurtimox. We also investigated potential drug-drug interactions (DDIs) of these compounds in vitro. In rats, orally administered nifurtimox was rapidly absorbed (tmax 0.5 h) and eliminated (t½ 1.4 h). Metabolism of nifurtimox yielded six predominant metabolites (M-1 to M-6) in urine and plasma, and the dose was excreted equally via the renal and fecal routes with only traces of unchanged nifurtimox detectable due to its instability in excreta. In patients with Chagas disease, only M-6 and M-4 achieved relevant exposure levels, and the total amount of excreted metabolites in urine was higher in fed versus fasted patients, consistent with the higher systemic exposure. For nifurtimox, M-6, and M-4, no potential perpetrator pharmacokinetic DDIs with the main cytochrome P- 450 enzymes and drug transporters were identified in vitro. This contemporary analysis of the complex metabolite profile and associated exposures emerging after oral dosing of nifurtimox in rats and humans, together with the expected low risk for clinically relevant DDIs, expands the understanding of this important anti-trypanosomal drug.

An integrated strategy by absorbed component characterization, pharmacokinetics, and activity evaluation for identification of potential nephroprotective substances in Zhu-Ling decoction.

An efficient strategy for the identification of potential nephroprotective substances in Zhu-Ling decoction has been established with the integration of absorbed components characterization, pharmacokinetics, and activity evaluation. A qualitative method was developed to characterize the chemical constituents absorbed components in vivo of Zhu-Ling decoction by using ultra-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry. A quantitative method was established and validated for the simultaneous determination of eight compounds in rat plasma by using ultra-performance liquid chromatography-triple quadruple tandem mass spectrometry. Finally, the nephroprotective activities of absorbed components with high exposure were assessed by cell survival rate, superoxide dismutase, and malondialdehyde activities in hydrogen peroxide-induced Vero cells. As a result, 111 compounds in Zhu-Ling decoction and 36 absorbed components were identified in rat plasma and urine, and poricoic acid A, poricoic acid B, alisol A, 16-oxo-alisol A, and dehydro-tumulosic acid had high exposure levels in rat plasma. Finally, poricoic acid B, poricoic acid A, 16-oxo-alisol A, and dehydro-tumulosic acid showed remarkable nephroprotective activity against Vero cells damage induced by hydrogen peroxide. Besides, superoxide dismutase and malondialdehyde activities were obviously regulated in hydrogen peroxide-induced Vero cells by treatment with the four compounds mentioned above. Therefore, these four compounds were considered to be effective substances of Zhu-Ling decoction due to their relatively high exposure in vivo and biological activity. This study provided a chemical basis for the action mechanism of Zhu-Ling decoction in the treatment of chronic kidney diseases.

Comparative Pharmacokinetic Study of 5 Active Ingredients after Oral Administration of Zuogui Pill in Osteoporotic Rats with Different Syndrome Types.

Zuogui Pill is a kidney-yin-tonifying formula in traditional Chinese medicine that is widely used to manage osteoporosis with kidney-yin-deficiency in China. Herein, an efficient and accurate high-performance liquid chromatography-tandem mass spectrometry method was developed to determine the concentrations of 5 bioactive compounds in rat plasma following oral administration of Zuogui Pill. Because drug absorption and distribution differ under physiological and pathological conditions, the established method was used to quantify blood components and dynamic change in osteoporotic rats with different syndrome types. Moreover, integrated pharmacokinetic study was conducted to describe the overall pharmacokinetic characteristics of traditional Chinese medicine. The results showed that the absorption, distribution, and metabolism of Zuogui Pill varied widely under different states. The bioavailability of most active components showed significant advantages in osteoporotic rats with kidney-yin-deficiency, which corresponds to the opinion that Zuogui Pill has the effect of nourishing kidney-yin. It is hoped that this finding could interpret the pharmacodynamic substances and mechanism of Zuogui Pill in the treatment of osteoporosis with kidney-yin-deficiency.

Simultaneous Determination of Three Compounds in Rat Plasma by UHPLC-QQQ-MS/MS and Its Application to Pharmacokinetics of Banxia Baizhu Tianma Tang.

A rapid and highly selective and sensitive ultra-high-performance liquid chromatography coupled with triple quadrupole mass spectrometry (UHPLC-QQQ-MS/MS) method was applied to simultaneously determine ephedrine, gastrodin, and liquiritin in rat plasma. The three analytes and vitexin-2″-O-rhamnoside (I.S.) were analyzed on a Waters Acquity UPLC C18 column (1.7 μm, 2.1 mm × 100 mm) at 30°C with gradient mobile phase consisting of 0.1% formic acid aqueous solution (A) and acetonitrile (B) after one-step direct protein precipitation with acetonitrile. The detection was performed by multiple reaction monitoring (MRM) mode via electrospray ionization (ESI) source in positive and negative ion modes. The product ions m/z 166.1⟶148.1, 285.1⟶123.1, 417.1⟶255.1, and 579.0⟶433.1 were used for determination of ephedrine, gastrodin, liquiritin, and I.S., respectively. The calibration curves of the three analytes were linear with r 2 greater than 0.994. The intra and interday precision RSD% was less than 11.5 and 13.4. The intra and interday precision RE% was between -10.4% and 9.33%. The average extraction recoveries of the three analytes were no less than 86.88 ± 1.08%. The developed and validated method was for the first time applied to the pharmacokinetics of three compounds in rat plasma after intragastric administration of Banxia Baizhu Tianma Tang.

Drug-polysaccharide/herb interactions and compatibility rationality of Sijunzi decoction based on comprehensive pharmacokinetic screening for multi-components in rats with spleen deficiency syndrome

Ethnopharmacological relevanceSijunzi decoction (SJZD) is composed of four herbs, namely Ginseng Radix et Rhizoma (RG, Panax ginseng C.A.Mey.), Atractylodes Macrocephalae Rhizoma (AM, Atractylodes macrocephala Koidz.), Poria (Poria cocos (Schw.) Wolf), and Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle (GRP, derived from Glycyrrhiza uralensis Fisch., Glycyrrhiza inflata Bat. or Glycyrrhiza glabra L.) based on the compatibility theory of traditional Chinese medicine (TCM), which is a classical formula for the treatment of spleen deficiency syndrome (SDS) in TCM. The polysaccharides and non-polysaccharides (NPSs) composition represented by flavonoids, saponins and terpenoids are the important pharmacodynamic material basis of SJZD. Aim of the studyThe aim of this study was to investigate the pharmacokinetic characteristics of SJZD in normal rats and SDS rats, and explore the potential interactions between NPSs and polysaccharides in SJZD, as well as the compatibility rationality of SJZD. Materials and methodsSDS model was established by oral administration of Radix Rhei (Rheum officinale Baill.) extract, loaded swimming, and intermittent fasting. A rapid, sensitive and reliable ultrafast liquid chromatography tandem mass spectrometry (UFLC–MS/MS) method was developed for the simultaneous analysis of fifteen representative compounds in rat plasma to investigate the differences in pharmacokinetics between normal and SDS rats. The SJZD-NPS samples were prepared by removing the polysaccharides of SJZD to explore the interactions between NPSs and polysaccharides of SJZD. According to the compatibility theory of TCM, four incomplete formulae of SJZD were obtained by randomly removing an herb (also called 'que fang' in TCM), and their pharmacokinetic differences were compared to elucidate the rationality of SJZD compatibility with oral administration to SDS rats. ResultsThe established UFLC–MS/MS method showed perfect performance in simultaneously analyzing fifteen compounds of SJZD in rat plasma. Compared with normal rats, the absorption efficiency of NPSs in SDS rats was lower, accompanied by the prolonged residence time (Cmax and AUC0-t reduced, while MRT0-t increased). Polysaccharides have the potential to enhance intestinal metabolism of glycosides among these components, thereby contributing to the circulating distribution of corresponding metabolites (e.g. aglycones). Furthermore, the compatibility of the four herbs in SJZD could alter their pharmacokinetic characteristics, and potentially improve the absorption of the effective components of RG and AM, which is in accordance with the principle that “monarch” and “minister” herbs play a major role in TCM. In detail, the improved absorption of ginsenosides was mainly regulated by GRP (the “guide” herb in SJZD), together with the effects of AM (“minister” herb) and Poria (“adjuvant” herb) on the pharmacokinetics of components in GRP, implying that herb-herb interactions existed in SJZD and demonstrated the compatibility rationality of SJZD potentially. ConclusionThis study laid a solid foundation for revealing the pharmacodynamic material basis and subsequent action mechanism of SJZD, as well as provided new insights into the compatibility of SJZD. The comprehensive pharmacokinetic approach adopted in the current research also provides a valuable strategy for TCM formulae research.

Qualitative Analysis and Quality Evaluation of Pimpinella thellungiana by Serum Pharmacochemistry, Network Pharmacology, and QAMS.

Pimpinella thellungiana H. Wolff (P. thellungiana) is widely used as a Chinese medicinal herb to treat coronary heart disease (CHD). However, the bioactive compounds of this herbal medicine have not been defined, and quality control is imperfect. To develop methods for qualitative analysis and improved quality evaluation of P. thellungiana. In this study, UHPLC-Q-Exactive Orbitrap MS, network pharmacology, and molecular docking approaches were applied to explore the quality markers of P. thellungiana. A quantitative analysis of multi-components via a single marker (QAMS) method was developed to simultaneously detect nine quality markers of P. thellungiana. Nine compounds were selected as quality markers for quality control of P. thellungiana: protocatechuic acid, neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, luteolin-7-O-glucuronide, 3,5-dicaffeoylquinic acid, 4,5-dicaffeoylquinic acid, apigenin-7-O-glucuronide, and 4,5-dicaffeoylquinic acid. The established QAMS method was validated and applied in 10 batches of P. thellungiana samples. These findings facilitate effective quality evaluation of P. thellungiana and suggest this approach could be applied to evaluate the quality of other herbal medicines. UHPLC-Q-Exactive Orbitrap MS was used to detect 36 compounds in P. thellungiana and 40 compounds in rat plasma after dosing with P. thellungiana. Quality markers were selected based on detection of compounds absorbed into rat plasma and network pharmacology analysis.

Pharmacokinetics and brain distribution studies of 6-hydroxykynurenic acid and its structural modified compounds.

6-Hydroxykynurenic acid (6-HKA) is an organic acid component in extracts of Ginkgo biloba leaves and acts as a major contributor to neurorestorative effects, while its oral bioavailability was low. Therefore, using prodrug method to improve the bioavailability and brain content of 6-HKA is significant. Three structural modified compounds of 6-HKA were synthesized, and ultra performance liquid chromatography-tandem mass spectrometry methods for quantification of these structural modified compounds in rat plasma and rat brain homogenate were established and comprehensively validated. The methods were effectively applied to investigate the effects of structural modification on apparent permeability coefficients in cells, the pharmacokinetics and the brain distribution in rats. The results illustrated that esterification can greatly improve the apparent permeability coefficient and bioavailability of 6-HKA. Comparing with direct oral administration of 6-HKA, the bioavailability of isopropyl ester was greatly improved (from 3.96 ± 1.45% to 41.8 ± 15.3%), and the contents of 6-HKA in rat brains (49.7 ± 9.2 ng/g brain) were significantly higher after oral administration. The bioavailability and the brain content of 6-HKA can be improved by the prodrug method. Among three structural modified compounds, isopropyl-esterified 6-HKA was the most promising treatment.

GC-MS Method for Quantification and Pharmacokinetic Study of Four Volatile Compounds in Rat Plasma after Oral Administration of Commiphora myrrh (Nees) Engl. Resin and In Vitro Cytotoxic Evaluation

A rapid, simple, and sensitive gas chromatography–tandem mass spectrometry (GC–MS) method was established and validated for simultaneous determination of four volatile compounds, namely curzerene, methoxyfuranodiene, β-elemene, and α-pinene in rat plasma samples after oral administration of the resin extract of Commiphora myrrh using limonene as an internal standard (IS). Liquid-liquid extraction using hexane and ethyl acetate (1:1) mixture as an extracting agent was used for the samples extraction procedure. The GC–MS system was operated under selective ion monitoring (SIM) mode using Perkin Elmer Elite 5MS column (30 m × 0.25 mm × 0.25 µm film thickness). Specificity, linearity, precision, accuracy, extraction recovery, and stability were used to validate the developed method. The assay showed good linearity (r2 ≥ 0.998), and the lowest limits of quantification (LLOQ) were 3.97–21.38 ng/mL for the four analytes. This assay was successfully applied to pharmacokinetic studies of the four volatile compounds in rat plasma. The antiproliferative activity of these volatile compounds was evaluated against lung carcinoma (A549) and colon (LoVo) cell lines, were each compound caused variable inhibition on cells proliferation and methoxyfuranodiene exerted the strong antiproliferative activity against both cell line according to IC50 values.

UHPLC-MS/MS Method for Quantifying Fangchinoline, Tetrandrine and Calycosin-7-O-β-D-Glucoside of Fangji Huangqi Decoction in Rat Plasma and Its Application to a Pharmacokinetic Study.

Fangji Huangqi Decoction is composed of Stephaniae Tetrandrae Radix, Astragli Radix, Atractylodis Macrocephalae Rhizoma and Glycyrrhizae Radix Et Rhizoma. It is a classic traditional Chinese medicine formula for the treatment of chronic glomerulonephritis in China. However, its pharmacokinetic characteristics in vivo are still unclear. In this study, a method for quantifying fangchinoline, tetrandrine and calycosin-7-O-β-D-glucoside, the main active constituents of Fangji Huangqi Decoction, in rat plasma by using ultrahigh-performance liquid chromatography-tandem mass spectrometry technique was developed. Plasma samples were processed with a deproteinization procedure using acetonitrile, followed by chromatographic separation on a Shim-pack XR-ODS C18 column using gradient elution of 0.1% aqueous formic acid and acetonitrile at 0.4mL/min. The analytes and internal standard, diphenhydramine hydrochloride, were detected using positive electrospray ionization in multiple reactions monitoring mode. The optimized mass transition ion-pairs (m/z) were 609.3/367.3 for fangchinoline, 623.3/174.3 for tetrandrine, 447.2/285.1 for calycosin-7-O-β-D-glucoside and 256.2/167.1 for diphenhydramine hydrochloride, respectively. The developed method was validated for intraday and interday precision and accuracy whose values fell in the acceptable limits. Recovery efficiency of all the analytes was found to be >90.5%. Matrix effect was found to be negligible. Stability results showed that the analytes were stable under all conditions. The validated method was successfully used for studying the pharmacokinetics of the three compounds in rat plasma after oral administration of Fangji Huangqi Decoction.

Simultaneous LC-MS/MS bioanalysis of alkaloids, terpenoids, and flavonoids in rat plasma through salting-out-assisted liquid-liquid extraction after oral administration of extract from Tetradium ruticarpum and Glycyrrhiza uralensis: a sample preparation strategy to broaden analyte coverage of herbal medicines.

Herbal medicines have historically been practiced in combinatorial way, which achieves therapeutic efficacy by integrative effects of multi-components. Thus, the accurate and precise measurement of multi bioactive components in matrices is inalienable to understanding the metabolism and disposition of herbal medicines. In this study, aiming to provide a strategy that improves analyte coverage, evaluation of six protocols employing sample pretreatment methods- protein precipitation (PPT), liquid-liquid extraction (LLE), sugaring-out-assisted liquid-liquid extraction (SULLE), and salting-out-assisted liquid-liquid extraction (SALLE)- was performed by LC-MS/MS using rat plasma and a mixture of alkaloid (evodiamine, rutaecarpine, dehydroevodiamine), terpenoid (limonin, rutaevin, obacunone), and flavonoid (liquiritin, isoliquiritin, liquiritigenin) standards isolated from Tetradium ruticarpum and Glycyrrhiza uralensis. These protocols were as follows: (1) PPT with methanol, (2) PPT with acetonitrile, (3) LLE with methyl tertiary-butyl ether-dichloromethane, (4) LLE with ethyl acetate-n-butanol, (5) SALLE with ammonium acetate, (6) SULLE with glucose. The results suggested that SALLE produced broader analyte coverage with satisfactory reproducibility, acceptable recovery, and low matrix interference. Then, sample preparation procedure of SALLE, chromatographic conditions, and mass spectrometric parameters were optimized, followed by method validation, showing that good sensitivity (LLOQ ≤1 ng mL-1), linearity (r ≥0.9933),precision (RSD≤ 14.45%), accuracy (89.54~110.87%), and stability could be achieved. Next, the developed method was applied successfully to determine the pharmacokinetic behavior of the nine compounds in rat plasma after intragastric administration with an extract from Tetradium ruticarpum and Glycyrrhiza uralensis (Wuzhuyu-Gancao pair). Based on an extensive review and experiments, a sample preparation procedure that matches with LC-MS/MS technique and can get wider analyte coverage was outlined. The developed SALLE method is rapid, reliable, and suitable for bioanalysis of analytes with diverse polarity, which was expected to be a promising strategy for the pharmacokinetic studies of herbal medicines. Graphical abstract.

Simultaneous determination of twenty-five compounds with anti-inflammatory activity in Spatholobi Caulis by using an optimized UFLC-MS/MS method: An application to pharmacokinetic study

As a kind of commonly used Traditional Chinese Medicine in clinical, Spatholobi Caulis (SPC) contains a wide variety of bioactive compounds, including protocatechuate (1), nicotinic acid (2), p-hydroxybenzoic acid (3), salicylic acid (4), 6,9-dihydroxy megastigma-4,7-dien-3-one (5), 8,9-dihydroxy megastigma-4,6-dien-3-one (6), daidzin (7), genistin (8), isolariciresinol (9), ononin (10), 4′,8-dimethoxy-7-O-β-d-glucopyranosyl isoflavone (11), 3′-methoxydaidzein (12), odoratin (13), spasuberol A (14), (+)-pinoresinol (15), 4-hydroxy-3-methoxy cinnamic acid methyl ester (16), (+)-epipinoresinol (17), calycosin (18), 8-O-methylretusin (19), formononetin sodium (20), formononetin (21), biochanin A (22), butesuperin A (23), homovanillyl-4-oxo-nonanoate (24) and (6aR,11aR)-maackiain (25). The pharmacokinetic characteristics of these twenty-five compounds in rat plasma were quantitatively and simultaneously studied using a fast, sensitive and precise ultra fast liquid chromatography combined with electrospray ionization triple quadrupole tandem mass spectrometry (UFLC-MS/MS) method after oral administration of aqueous extract of SPC to rats. The mobile phase consists of acetonitrile and 0.5 mM ammonium acetate in water, and these compounds were well separated at a gradient elution program with flow rate of 0.35 mL/min. Carbamazepine was employed as the internal standard (IS) and all samples were precipitated with MeOH-ACN (2:1, v/v). The analytical method has been proved to be good linearity (R2 ≥ 0.9957), precise, accurate, stable, recovery and matrix effect, which applicated becomingly to study the pharmacokinetic processes of these compounds in rat plasma. In addition, these twenty-five compounds exhibited anti-inflammatory activity on the inflammatory model of NO over production in RAW264.7 cells stimulated by lipopolysaccharide (LPS). Isoflavones, especially compounds 20-22 (The IC50 of which were 22.75 μM, 21.11 μM and 48.29 μM, respectively.) might be the important constituents for anti-inflammatory activity of SPC. This study provides reference values for the clinical application, in-depth study on new dosage forms and pharmacological activities of SPC.