The influence of essential oils from ZhaLi NuSi Prescription on the pharmacokinetics of its non-volatile components in normal rats.

Hui Medicine ZhaLi NuSi Prescription (ZLNS) is described in "Hui Hui Prescription," and it has been used to treat cerebral infarction in Hui Region, China. In this study, a rapid and reliable ultra-performance liquid chromatography coupled with mass spectrometry (UPLC-MS/MS) method was established and applied to simultaneously determine geniposidic acid, oxypaeoniflorin, hydroxysafflor yellow A, caffeic acid, magnoflorine, paeoniflorin, ferulic acid, ß-ecdysterone, icariin, rhein, and baohuoside I in rat plasma. The pharmacokinetic parameters of these components and the influence of essential oils (EOs) on them were investigated in normal rats. The results showed that the pharmacokinetic parameters (AUC0 - t , AUC0 - ∞ , t1/2 , tmax , cmax ) of the aforementioned compounds were significantly changed after co-administering with ZLNS EO. The AUC values of oxypaeoniflorin, paeoniflorin, ferulic acid, and baohuoside I with EOs were decreased significantly. This is the first report for the comparative pharmacokinetic study of ZLNS bioactive components in normal rats, which may provide the basis for drug interaction study in vivo and insight into their clinical applications.

Pharmacokinetic-pharmacodynamic modeling analysis and anti-inflammatory effect of Wangbi capsule in the treatment of adjuvant-induced arthritis.

Clinically, Wangbi Capsule (WBC) is widely used in the treatment of Rheumatoid arthritis (RA) because of its remarkable therapeutic effect. To reveal the mechanism, a pharmacokinetic-pharmacodynamic (PK-PD) model was developed for the first time to assess the relationship between time-concentration (dose)-effect. Freund's Complete Adjuvant was used to induce the adjuvant-induced arthritis model. Multi-indices were used to evaluate the therapeutic effect and an S-Imax PK-PD model was established based on the concentrations of osthole, 5-O-methylvisamminoside, cimifugin, albiflorin, paeoniflorin and icariin and the levels of interleukin-1ß and prostaglandin E2 using a two-compartment PK model together with a PD model with an effect-site compartment. The results suggest that WBC can treat RA by regulating the levels of prostaglandin E2 and interleukin-1ß. For the PK-PD model, the parameters indicated that WBC had a large safety margin and all six bioactive ingredients of WBC have therapeutic effects on RA. Among them icariin, osthole and 5-O-methylvisamminoside may be the main effective substances. This study provided a scientific basis for further study of population pharmacokinetics / population pharmacodynamics (PPK/PPD), to develop a reasonable administration plan and improve individualized drug therapy.

Intranasal administration of perillyl alcohol-loaded nanoemulsion and pharmacokinetic study of its metabolite perillic acid in plasma and brain of rats using ultra-performance liquid chromatography/tandem mass spectrometry.

Perillyl alcohol (POH) is a monocyclic terpene that has strong antitumor activity. Brain tumors are particularly difficult to treat with therapeutic agents, and clinical trials have shown their low tolerance through oral administration. We proposed the entrapment of POH into an oil-in-water chitosan nanoemulsion aiming its intranasal administration for brain targeting. An ultra-performance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS) method was developed and validated for the quantitation of total metabolite perillic acid (PA) in plasma and brain of rats. The rat samples containing the metabolite were treated by liquid-liquid extraction with acetonitrile. The mobile phase was 0.1% formic acid in water (solvent A) and 0.1% formic acid in methanol (solvent B), at a flow rate of 0.3 mL min-1 in gradient elution. The chromatography was run for 10 min, and analytical curves were built in acetonitrile, plasma, and brain. The PA was detected in positive ion mode with multiple reaction monitoring. The method has shown high selectivity, sensitivity, and throughput. The low quantification limits of 162, 178, and 121 ng mL-1 for acetonitrile, brain, and plasma, respectively, indicate a good detectability of the method. The repeatability and precision observed were within the limits recommended in the literature. The accuracy of the method was verified through high recovery rates (106-118%). The validated method was successfully applied to the pharmacokinetic study of the metabolite PA after the intranasal administration of free or POH-loaded nanoemulsion in rats. The results showed that chitosan nanoemulsion improved the plasma and brain bioavailability of POH, representing a promising alternative to free POH treatment.

UPLC-MS/MS simultaneous determination of seven active ingredients of Yaobitong capsule in rat plasma and its integrated pharmacokinetic application.

A reliable and sensitive UPLC-MS/MS method was first established and validated for the simultaneous determination of seven active ingredients of Yaobitong capsule in rat plasma: ginsenoside Rg1, ginsenoside Rb1, osthole, tetrahydropalmatine, paeoniflorin, albiflorin, and ferulic acid. And this method was further applied for the integrated pharmacokinetic study of Yaobitong capsule in rats after oral administration. Plasma samples (100 µL) were precipitated with 300 µL of methanol using carbamazepine as internal standard. Chromatographic separation was achieved using an Aquity UPLC BEH C18 column (100 × 2.1 mm, 1.7 µm), with the mobile phase consisting of 0.1% formic acid and acetonitrile. The method was validated using a good linear relationship (r ≥ 0.991), and the lower limit of quantification of the analytes ranged from 0.5 to 40 ng/mL. In the integrated pharmacokinetic study, the weight coefficient was calculated by the ratio of AUC0-∞ of each component to the total AUC0-∞ of the seven active ingredients. The integrated pharmacokinetic parameters Cmax , Tmax , and t1/2 were 81.54 ± 9.62 ng/mL, 1.00 ± 0.21 h, and 3.26 ± 1.14 h, respectively. The integration of pharmacokinetic parameters showed a shorter t1/2 because of fully considering the contribution of the characteristics of each active ingredient to the overall pharmacokinetics.

Simultaneous determination of ferulic acid, paeoniflorin, and albiflorin in rat plasma by ultra-high performance liquid chromatography with tandem mass spectrometry: Application to a pharmacokinetic study of Danggui-Shaoyao-San.

A rapid, selective, and sensitive ultra-high performance liquid chromatography-tandem mass spectrometry method was developed for simultaneous determination of ferulic acid, paeoniflorin, and albiflorin, the major active constituents of Danggui-Shaoyao-San, in rat plasma using geniposide as the internal standard. The plasma samples were processed by protein precipitation with acetonitrile, and then separated on a Shim-Pack XR-ODS C18 column (75 mm × 3.0 mm, 2.2 µm) using gradient elution program with a mobile phase consisting of 0.1% aqueous formic acid and acetonitrile at a flow rate of 0.4 mL/min. The detection was achieved on a 3200 QTRAP mass spectrometer equipped with electrospray ionization source in negative ionization mode. Quantification was performed using multiple reaction monitoring mode by monitoring the fragmentation of m/z 192.9→134.0 for ferulic acid, m/z 525.0→120.9 for paeoniflorin, m/z 525.2→121.0 for albiflorin, and m/z 433.1→225.1 for the internal standard, respectively. The calibration curve was linear in the range of 5-2500 ng/mL for all the three analytes (r ≥ 0.9972) with the lower limit of quantitation of 5 ng/mL. The intraday and interday precisions were below 12.1% for all the analytes in terms of relative standard deviation, and the accuracy was within ±11.5% in terms of relative error. The extraction recovery, matrix effect and stability were satisfactory in rat plasma. The validated method was successfully applied to a pharmacokinetic study of ferulic acid, paeoniflorin, and albiflorin after oral administration of Danggui-Shaoyao-San to rats.

Simultaneous measurement of perillyl alcohol and its metabolite perillic acid in plasma and lung after inhalational administration in Wistar rats.

The inhalational administration of drugs is a practical and non-invasive approach with the potential to reduce side effects and with a quick onset of therapeutic activity. Perillyl alcohol (POH) is a monoterpene with antitumor activity that currently is undergoing clinical evaluation as an inhalational anticancer agent. A detection method was developed that will be applicable to pharmacokinetic studies of not only POH, but also its longer-lived main metabolite, perillic acid (PA), in lung tissue and plasma after inhalational delivery. The anticancer activity of POH was investigated in vitro with the use of various lung cancer cell lines. Toxicity was established by a standard MTT assay, and apoptosis markers were analyzed by Western blot. For the detection of POH and PA in lungs and plasma, albino Wistar rats were used that were exposed to POH inhalation. Tissues were subjected to chromatographic separation on an Agilent Zorbax Eclipse XDB C18 column, followed by detection of absorption in the ultraviolet (UV) range. In vitro, POH exerted cytotoxic activity against six different lung tumor cell lines, and apoptotic cell death was indicated by induction of active caspase 3 and cleavage of poly (ADP-ribose) polymerase 1 (PARP1). These results demonstrate that inhalational delivery of POH results in effective biodistribution and metabolism of POH in the systemic circulation. In addition, our study introduces a simple, rapid HPLC-UV method with high accuracy for simultaneous detection of POH and its metabolite PA in plasma, and for sensitive detection of PA in lung tissue, which should prove useful for applications in clinical studies.

The profiling and identification of the absorbed constituents and metabolites of Naoshuantong capsule in mice biofluids and brain by ultra- fast liquid chromatography coupled with quadrupole-time-of-flight tandem mass spectrometry.

Naoshuantong capsule (NSTC) is an oral traditional Chinese medicine formula used widely in the clinic for ischemic stroke. The absorbed ingredients and metabolites of NSTC have never been reported before. In this study, a method incorporating rapid resolution liquid chromatography with quadrupole-time-of-flight mass spectrometry (UPLC-Q-TOF/MS) was used to identify absorbed ingredients and metabolites after oral administration of NSTC. A total of 15 constituents were detected and identified as prototypes of NSTC. 109 metabolites related to catechin, gallic acid, paeoniflorin, chlorogenic acid, protocatechuate, typhaneoside, ß-elemene, calycosin were identified in serum, urine and brain. 19 metabolites of typhaneoside, 3 metabolites of ß-elemene, 12 metabolites of calycosin were reported for the first time. This is the first time to explore the absorption and metabolism of NSTC. The work will provide helpful information for further research of the mechanism and application of NSTC.

Effects of glycyrrhizin on the pharmacokinetics of paeoniflorin in rats and its potential mechanism.

Context: Paeoniflorin is reported to possess numerous pharmacological activities. Paeoniflorin and glycyrrhizin are always used together for the treatment of disease in China clinics; however, the drug-drug interaction between glycyrrhizin and paeoniflorin is still unknown. Objective: This study investigates the effects of glycyrrhizin on the pharmacokinetics of paeoniflorin in rats. Materials and methods: The pharmacokinetics of orally administered paeoniflorin (20 mg/kg) with or without glycyrrhizin pre-treatment (at a dose of 100 mg/kg/day for 7 days) were investigated in male Sprague-Dawley rats using LC-MS/MS. Additionally, Caco-2 cell transwell model and rat liver microsome incubation experiments were also conducted to investigate its potential mechanism. Results: The results showed that when the rats were pre-treated with glycyrrhizin, the Cmax of paeoniflorin decreased from 59.57 ± 10.24 to 45.36 ± 8.61 ng/mL, and AUC0-inf also decreased from 282.02 ± 35.06 to 202.29 ± 28.28 µg·h/L. The t1/2 value of paeoniflorin decreased from 8.48 ± 2.01 to 5.88 ± 1.15 h (p < 0.05). The Caco-2 cell transwell experiments indicated that glycyrrhizin could increase the efflux ratio of paeoniflorin from 2.71 to 3.52, and the rat liver microsome incubation experiments showed that glycyrrhizin could significantly increase its intrinsic clearance rate from 53.7 ± 4.6 to 85.6 ± 7.1 µL/min/mg protein. Conclusions: These results indicated that glycyrrhizin could affect the pharmacokinetics of paeoniflorin, and it might work through decreasing the absorption of paeoniflorin by inducing the activity of P-gp or through increasing the clearance rate in rat liver by inducing the activity of CYP450 enzyme.

A network pharmacology integrated pharmacokinetics strategy for uncovering pharmacological mechanism of compounds absorbed into the blood of Dan-Lou tablet on coronary heart disease.

ETHNOPHARMACOLOGICAL RELEVANCE: Dan-Lou tablet (DLT) is developed from the traditional Chinese medicine (TCM) formula Gualou Xiebai Baijiu Tang which has been used for at least 2000 years in China. DLT has been widely used in clinical practice to treat cardiovascular diseases. AIM OF THE STUDY: This study aimed to uncover the pharmacological mechanism of the compounds absorbed into the blood of Dan-Lou tablet (DLT) on coronary heart disease (CHD) using a network pharmacology integrated pharmacokinetics strategy. MATERIALS AND METHODS: A rapid and sensitive method was developed for the simultaneous determination of the six compounds (puerarin, formononetin, calycosin, paeoniflorin, cryptotanshinone and tanshinone IIA) in rat plasma by liquid chromatography tandem mass spectrometry (LC-MS/MS). Then, the pharmacology network was established based on the relationship between five compounds absorbed into the blood targets (puerarin, formononetin, calycosin, cryptotanshinone and tanshinone IIA) and CHD targets. RESULTS: The intra-and inter-day precision were less than 11% and the accuracy ranged from 88.2% to 112%, which demonstrated that the LC-MS/MS method could be used to evaluate the pharmacokinetic feature of the six compounds in rats after oral administration of DLT. The pathway enrichment analysis revealed that the significant bioprocess networks of DLT on CHD were positive regulation of estradiol secretion, negative regulation of transcription from RNA polymerase II promoter, lipopolysaccharide-mediated signaling pathway and cytokine activity. CONCLUSION: The proposed network pharmacology integrated pharmacokinetics strategy provides a combination method to explore the therapeutic mechanism of the compounds absorbed into the blood of multi-component drugs on a systematic level.

Comparative pharmacokinetics and metabolites study of seven major bioactive components of Shaoyao-Gancao decoction in normal and polycystic ovary syndrome rats by ultra high pressure liquid chromatography with tandem mass spectrometry.

A simple and sensitive liquid chromatography with tandem mass spectrometry method was developed for simultaneous quantification of paeoniflorin, albiflorin, oxypaeoniflorin, liquiritin, liquiritigenin, glycyrrhetinic acid, and glycyrrhizin in rat plasma after oral administration of Shaoyao-Gancao decoction, which is traditionally used in the treatment of polycystic ovary syndrome. The plasma samples were pretreated with methanol as precipitant. The method exhibited good linearity (correlation coefficient (R2 ) > 0.99) with lower quantification limits of 0.595-4.69 ng/mL for all analytes. Intra- and interbatch precision, accuracy, recovery, and stability of the method were all within accepted criteria. The results showed that the pharmacokinetic behaviors of the seven compounds were altered in the pathological status of polycystic ovary syndrome. Furthermore, a total of 36 metabolites were structurally identified based on their accurate masses and fragment ions. The major metabolic pathway involves phase I metabolic reactions (such as hydroxylation), phase II metabolic reactions (such as sulfation and glucuronidation conjugation) as well as the combined multiple-step metabolism. This study is the first report on the pharmacokinetic and metabolic information of Shaoyao-Gancao decoction in both normal and model rats, which would provide scientific evidences for the bioactive chemical basis of herbal medicines and also promote the clinical application of Shaoyao-Gancao decoction for treating polycystic ovary syndrome.

Simultaneous determination of calycosin-7-O-ß-D-glucoside, cinnamic acid, paeoniflorin and albiflorin in rat plasma by UHPLC-MS/MS and its application to a pharmacokinetic study of Huangqi Guizhi Wuwu Decoction.

Huangqi Guizhi Wuwu Decoction (HGWWD), consisting of Radix Astragali, Cinnamomi Ramulus, Paeoniae Radix Alba, Zingiberis Rhizoma Recens and Jujubae Fructus, is a widely used Traditional Chinese Medicine (TCM) formula for the treatment of human blood impediment in China for nearly 2000 years. In order to make good and rational use of this formula in the future, a rapid, sensitive and robust ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed for simultaneous determination of calycosin-7-O-ß-D-glucoside, cinnamic acid, paeoniflorin and albiflorin, the main active constituents of HGWWD, in rat plasma using geniposide as internal standard (IS). The plasma samples were extracted by protein precipitation with acetonitrile and separated on a Shim-pack XR-ODS C18 column (75 mm × 3.0 mm, 2.2 µm) using gradient elution with a mobile phase consisting of water (containing 0.1% formic acid) and acetonitrile at a flow rate of 0.4 mL/min. Mass spectrometric detection was performed on 3200 QTRAP mass spectrometry equipped with electrospray ionization source in negative ionization mode. Quantification was performed using multiple reaction monitoring (MRM) by monitoring the fragmentation of m/z 491.1→282.9 for calycosin-7-O-ß-D-glucoside, m/z147.0→103.1 for cinnamic acid, m/z 525.0→120.9 for paeoniflorin, m/z 525.2→121.0 for albiflorin and m/z 433.1→225.1 for IS, respectively. The method was well validated in terms of linearity, precision, accuracy, recovery, matrix effect and stability. All calibration curves had good linearity (r>0.9977) over the concentration range from 0.1-50 ng/mL for calycosin-7-O-ß-D-glycoside, 50-25000 ng/mL for cinnamic acid, 5-2500 ng/mL for paeoniflorin and albiflorin. The intra-day and inter-day precisions (relative standard deviation) were within 11.8%, the accuracy (relative error) ranged from -9.4% to 9.1%, and the lower limit of quantification (LLOQ) were 0.1, 50, 5, 5 ng/mL for calycosin-7-O-ß-D-glucoside, cinnamic acid, paeoniflorin and albiflorin, respectively. Extraction recovery, matrix effect and stability were satisfactory in rat plasma. The validated method was successfully applied to a pharmacokinetic study of calycosin-7-O-ß-D-glucoside, cinnamic acid, paeoniflorin and albiflorin after oral administration of HGWWD to rats.

Methotrexate improves the anti-arthritic effects of Paeoniflorin-6'-O-benzene sulfonate by enhancing its pharmacokinetic properties in adjuvant-induced arthritis rats.

Paeoniflorin-6'-O-benzene sulfonate (CP-25), is a novel derivative of paeoniflorin (Pae) with improved anti-arthritic effects. The aim of this study was to evaluate the therapeutic effects and pharmacokinetics of CP-25 when co-administered with MTX in adjuvant-induced arthritis (AA) rats. AA rats were randomly divided into 6 groups and treated as follows: TGP (50 mg/kg/day, ig), CP-25 (50 mg/kg/day, ig), MTX (0.5 mg/kg, 3 times/week, ig), TGP (50 mg/kg/day, ig) + MTX (0.5 mg/kg, 3 times/week, ig) and CP-25 (50 mg/kg/day, ig) + MTX (0.5 mg/kg, 3 times/week, ig) from days 14 - 35 after induction. Clinical, biochemical, and histological scores were used to assess the severity of arthritis while novel ultra-performance liquid chromatography mass spectrometry (UPLC-MS/MS) was used to detect plasma concentration of CP-25. Co-administration of CP-25 and MTX significantly reduced clinical signs of inflammation, suppressed the serum production of IL-17 compared to TGP (50 mg/kg/day, ig) and MTX (0.5 mg/kg, 3 times/week, ig). Pharmacokinetics studies showed increased AUC0-t, Cmax, and t1/2 of CP-25 while V/F and CL/F decreased when co-administered with MTX. We observed an increase concentration and longer exposure with decreased clearance of CP-25 when combined with MTX, that should be further explored for clinical RA therapy.

Established UPLC-MS/MS procedure for multicomponent quantitative analysis in rat plasma: A contrastive pharmacokinetics study of Qiangshen tablet in normal and kidney yang deficiency syndrome models.

Qiangshen tablet, an important prescription consisting of 14 kinds of Chinese herbal medicines, has been used for decades to treat kidney yang deficiency syndrome (KYDS) in China. Qiangshen tablet has been recorded in ChP (2015 edition) and possesses the effect of strengthening yang, invigorating qi and tonifying kidneys. In this research, a simple, reliable and specific method was established for simultaneous determination of stachydrine, psoralen, isopsoralen, morroniside, paeoniflorin and loganin in normal and KYDS rat plasma after intragastric administration of a Qiangshen tablet suspension by UPLC-MS/MS. Protein precipitation (PP) by acetonitrile and liquid-liquid extraction (LLE) by ethyl acetate - n-butanol (1: 1, v/v) were used for pretreatment of plasma samples. Chromatographic separation of two IS (Internal Standard) and six analytes was achieved using an ACQUITY UPLC® BEH C18 column (2.1 × 100 mm, 1.7 µm). The mobile phase consisted of 0.1% formic acid aqueous solution (solvent A) and acetonitrile (solvent B) with a gradient scheme. Multiple reaction monitoring (MRM) mode with positive and negative ion source switching was applied to perform the mass spectrometric analyses. This method has been validated with good linearity (r ≥ 0.9942) and acceptable precision and accuracy (RSD ≤ 11%, RE from -4.8% to 7.7%). The mean recovery values of the analytes and IS were all ≥68.28%, and the matrix effects ranged from 94.4% to 101.7%. The stability of the IS and analytes was measured throughout the experiment. The results showed significant differences between the pharmacokinetic traits of the analytes in the normal and KYDS groups, suggesting that pharmacokinetic procedures involving these analytes could be modified in cases of KYDS.

Preparation, physicochemical characterization and pharmacokinetics of paeoniflorin-phospholipid complex.

In order to enhance lipophilicity and oral bioavailability of paeoniflorin (PF), this study developed paeoniflorin-phospholipid complex (PF-PLC) by solvent-evaporation method. The optimum preparation technology of PF-PLC was screened by the combination of single factor and orthogonal experiment. The physicochemical properties of PF-PLC were evaluated via differential scanning calorimetry (DSC), Fourier-transform infrared spectroscopy (FTIR), X-ray powder diffraction (XRD) and oil-water partition coefficient study (lgP). The result of FTIR spectra indicated that there was some strong hydrogen bond interaction and good compatibility between the phospholipid molecule and PF in the complex. DSC and XRD structure analysis showed that PF was in form of amorphous structure in PF-PLC, and lgP of PF-PLC was enhanced, suggesting that the lipophilicity of PF-PLC was higher than that of PF. In vitro release of PF-PLC showed slower release than PF solution with its cumulative release rate of 93.81% at 24 h compared to 93.43% of PF at 1.5 h. In pharmacokinetic experiments, the AUC and Cmax of the PF-PLC were 1.97-fold and 2.5-fold higher than PF solution. These results suggested that PF-PLC could enhance lipophilicity and oral bioavailability of PF and provide a promising delivery system for the application of PF.

Absorption and efflux characteristics of CP-25 in plasma and peripheral blood mononuclear cells of rats by UPLC-MS/MS.

Paeoniflorin-6'-O-benzene sulfonate (code: CP-25), is an active monomer obtained by modifying the structure of paeoniflorin (Pae). CP-25 can alleviate the course of adjuvant arthritis (AA) rats by regulating immune inflammatory response and reducing bone damage. In addition, our research has found that immune cells are important target cells for its anti-inflammatory and immunomodulatory effects. Therefore, it is of great significance to study the pharmacokinetics of CP-25 in immune cells. The aim of this study was to investigate the absorption and efflux of CP-25 in plasma and peripheral blood mononuclear cells (PBMCs) of rats. We established a sensitive ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) to rapidly determine CP-25 in plasma and PBMC of rat. We found that the transport amount of CP-25 in PBMC gradually decreased with the increase of time, and reached equilibrium after 1 h. Moreover, there is a certain correlation between the concentration of CP-25 in plasma and the concentration of CP-25 in PBMC. In addition, we used several transporter inhibitors to study their effects on the efflux of CP-25 in PBMC. The efflux of CP-25 in PBMC increased with the increase of time in the first 30 min, and the efflux of CP-25 decreased gradually after 30 min. Furthermore, after multiple administration of 50 mg/kg in rats, concentration of CP-25 in PBMC is similar to the change of concentration of CP-25 in plasma. Our results suggest that CP-25 may enter PBMC by passive transport, and P-glycoprotein (P-gp), multidrug resistance-associated protein 2 (MRP2) and breast cancer resistance protein (BCRP) may be involved in the efflux of CP-25 in PBMC. This research provides a basis and guidance for further study of the clinical application of CP-25.

Simultaneous UPLC-MS/MS determination of four components of Socheongryong-tang tablet in human plasma: Application to pharmacokinetic study.

The purpose of this study was to develop a method for simultaneous analysis of schizandrin, ephedrine, paeoniflorin, and cinnamic acid as constituents of Socheongryong-tang tablet in human plasma using UPLC-MS/MS. These four components were separated using water containing 0.01% formic acid and methanol as a mobile phase by gradient elution at a flow rate of 0.3 mL/min with a HALO-C18 column (2.1 mm × 100 mm, 2.7 µm particle size). Quantitation was performed on a triple quadrupole mass spectrometer employing electrospray ionization technique operated in multiple reaction monitoring mode. Mass transitions were m/z 432.9 → 384.1 for schizandrin, 165.8 → 148.1 for ephedrine, 525.0 → 449.2 for paeoniflorin, 146.8 → 102.9 for cinnamic acid, and 340.0 → 324.0 for papaverine as internal standard. Liquid-liquid extraction and protein precipitation with ethyl acetate-methanol (1:2, v/v) were used to obtain these four components. Chromatograms showed high resolution, sensitivity, and selectivity without interference by plasma constituents. Calibration curves of schizandrin, ephedrine, paeoniflorin, and cinnamic acid in human plasma ranged from 0.02 to 8 ng/mL, 0.5 to 200 ng/mL, 0.2 to 80 ng/mL, and 1 to 400 ng/mL, respectively. Calibration curves of each analyte displayed excellent linearity, with correlation coefficients > 0.99. For all four components, both intra- and inter-day precisions (CV%) were <5.99%. The accuracy was 99.35-103.30% for schizandrin, 98.48-104.38% for ephedrine, 97.06-103.34% for paeoniflorin, and 99.97-104.36% for cinnamic acid. This analytical method developed in this study satisfied the criteria of international guidance. It could be successfully applied to pharmacokinetic studies of schizandrin, ephedrine, paeoniflorin, and cinnamic acid after oral administration of Socheongryong-tang tablet to humans.

Identification and Quantification of Thujone in a Case of Poisoning Due to Repeated Ingestion of an Infusion of Artemisia Vulgaris L.

Plants of the Artemisia genus are used worldwide as ingredients of botanical preparations. This paper describes the case of a 49-year-old man admitted to the emergency room at a Zurich hospital in a manic state after the ingestion of 1 L of an infusion of Artemisia vulgaris. Two monoterpenic ketones, α- and ß-thujone, are present in various concentrations in Artemisia spp., but adverse effects have previously been associated only with essential oil from Artemisia absinthium and attributed to the inhibition of gamma-aminobutyric acid receptors, with consequent excitation and convulsions. The aim of this work was to examine and quantify the possible presence of thujone in the patient's serum and urine. A High Performance Liquid Chromatography (HPLC) method with isocratic separation and fluorescence detection (FLD) was set up and validated. Serum thujone concentrations were found to be 27.7 ± 3.48 µg/mL at day 0 and 24.1 ± 0.15 µg/mL on day 1. Results were confirmed by a gas chromatography with flame ionization detection (FID). Poisoning due to thujone was thus confirmed, suggesting four possible scenarios: (1) an unusually high concentration of thujone in the A. vulgaris ingested; (2) chronic exposure as the cause of the poisoning; (3) low metabolic efficiency of the patient; (4) contamination or adulteration of the plant material with other Artemisia spp., for example, A. absinthium. PRACTICAL APPLICATION: These results could aid research in the field of adverse effects of botanicals, lead to better understanding and management of similar cases of poisoning, and promote more informed use of natural products.

Comparison of the pharmacokinetics of plant-based treatments in milk and plasma of USDA organic dairy cattle with and without mastitis.

Organic dairy products are the second largest sector of the organic food market. Organic dairy products come from United States Department of Agriculture (USDA) certified organic dairy cattle that meet USDA organic standards. Organic dairy cattle in the US cannot be treated with antibiotics for mastitis, one of the costliest diseases of dairy cattle, and thus effective alternatives are needed. When any compound (medication or other non-food product) is used in a food producing animal, a withhold time for that compound that meets US Food and Drug Administration (FDA) standards for food safety must be applied to the animal and its products (like milk). However, there are no US FDA products approved for mastitis that maintain USDA certified organic dairy cattle's organic status. Thus, we studied the pharmacokinetics of 3 compounds (garlic, thymol and carvacrol) used on organic both healthy and mastitic organic dairy cattle. We also used this information to estimate a milk withhold time using methods consistent with US FDA requirements. For thymol intra-mammary and carvacrol intra-mammary or topical administration, all compounds were partially absorbed into the body from the milk or skin. Thymol and carvacrol are measurable in plasma (at 0.0183 and 0.0202 µg/mL, respectively) after intramammary administration with similar elimination half lives of 1.7 h. Milk concentrations of thymol and carvacrol are much higher at 2.958 and 4.487 µg/mL in healthy cattle, respectively. Concentrations are not significantly different in cows with mastitis as compared to those in healthy cows. Despite these compounds being natural products, they should have a withhold time for milk of at least 24 h after administration. For garlic, levels remained below the limit of detection in milk and plasma and thus no withdrawal time appears to be needed for milk.

Identify super quality markers from prototype-based pharmacokinetic markers of Tangzhiqing tablet (TZQ) based on in vitro dissolution/ permeation and in vivo absorption correlations.

BACKGROUND: A quality marker (Q-marker) is defined as an inherent chemical compound that is used for the quality control of a drug. Its biological activities are closely related to safety and therapeutic effects. Generally, a multiple-component herbal medicine may have many Q-markers. We therefore proposed a concept of "super Q-marker" satisfying both the criterion of Q-markers and PK-markers to be used in more effective quality control of herbal medicine. PURPOSE: The first aim was to find suitable prototype-based PK-markers from Tangzhiqing tablets (TZQ), a Chinese patent medicine. Then super Q-markers were expected to be identified from the prototype-based PK-markers based on an in vitro-in vivo correlation study. METHODS: Potentially eligible prototype-based PK-markers were identified in a single- and multiple-dose pharmacokinetic study on TZQ in 30 healthy volunteers. The in vitro dissolution and permeation profiles of the prototype-based PK-markers of TZQ were evaluated by the physiologically-based drug dissolution/absorption simulating system (DDASS). An in vitro-in vivo correlation analysis was conducted between the dissolution/permeation behaviors in DDASS and the actual absorption profiles in human to test the transferability and traceability of the promising super Q-markers for TZQ. RESULTS: In human, plasma paeoniflorin and nuciferine as prototype-based PK-markers exhibited the appropriate pharmacokinetic properties, including dose-dependent systemic exposure (AUC, Cmax) and a proper elimination half-life (1∼3h). In DDASS, it was predicted that paeoniflorin and nuciferine are highly permeable but the absorption rates are primarily limited by the dissolution rates. Moreover, the established in vitro-in vivo correlations of paeoniflorin and nuciferine were in support of the super Q-markers features. CONCLUSION: Paeoniflorin and nuciferine are identified as the super Q-markers from the prototype-based PK-markers of TZQ based on findings from a combination of in vitro, in vivo, and in vitro-in vivo correlation studies. This method is practical for optimal identification of qualified Q-markers, thus helping improve the quality control of herbal medicines.

Application of TQSM polypharmacokinetics and its similarity approach to ascertain Q-marker by analyses of transitivity in vivo of five candidates in Buyanghuanwu injection.

BACKGROUND: It is well-known that the public still have been facing on a severe issue about the inconsistency of quality and therapeutic efficacy of traditional medicines. Recently, Professor Chang-Xiao Liu has created a new promising concept for identifying relevant quality-markers (Q-marker) from herbs, their formulas and manufacturing products. Therefore, building up a new approach is necessary for us to bridge over quality to efficacy of pharmaceutical products. STUDY DESIGN: In this paper, five candidate Q-markers, astragaloside IV, paeonflorin, amygdalin, tetramethylpyrazine, ferulic acid in Buyanghuanwu injection (BYHWI) had been designed to carry out in rat by using single and polypharmacokinetic models for total quanta to ascertain adequate Q-marker. METHODS: The Q-marker transitivity in vivo was studied with polypharmacokinetic model and its similarity approach, which were modeled with TQSM principle. The Q-marker was ascertained with transitive similarity and bioavailability in polypharmacokinetics. Their concentrations in plasma sample of white rat were determined by RP-HPLC. Data analyses were used by the DAS software for singles and myself-written-program with EXCEL for multiples. RESULTS: In BYHWI, five candidate Q-marker pharmacokinetic profiles were singly fixed to two compartmental models in rat using classical compartmental analysis, but there were tremendous differences among which the candidate parameters were fluctuated from nearly 3552 folds to equivalency. The theoretical value of TQSM polypharmacokinetic parameters such as AUCT, MRTT, VRTT, CLT, VT over the mixure of five drugs were 110.8 ±â€¯51.91 mg min ml-1, 176.0 ±â€¯36.5 min, 39,921 ±â€¯4311 min2, 0.3116 ±â€¯0.02347 ml min-1 kg-1, 54.83 ±â€¯7.683 ml kg-1 respectively. The TQSM polypharmacokinetic parameters in astragaloside Ⅳ ordered by AUCT, MRTT, VRTT, CLT, VT were 110.8 ±â€¯51.91 mg min ml-1, 176.0 ±â€¯36.5 min, 39,921 ±â€¯4311 min2, 0.3116 ±â€¯0.02347 ml min-1 kg-1, 54.83 ±â€¯7.683 ml kg-1, respectively, which were closed to the theoretical values. TQSM similarity versus astragaloside Ⅳ was 0.9661. CONCLUSION: The results represented that the optimum Q-marker in BYHWI is astragaloside Ⅳ, whose transitivity in vivo similarity was close to the behavior of polypharmacokinetics with maximum bioavailability to the total quanta. It is feasible for Q-marker in CMMs to screen on the comparison of single pharmacokinetic behavior and bioavailability to the total quanta.