Qili Zhang

Simultaneous determination of cinnamaldehyde, cinnamic acid, and 2-methoxy cinnamic acid in rat whole blood after oral administration of volatile oil of Cinnamoni Ramulus by UHPLC-MS/MS: An application for a pharmacokinetic study

A simple and rapid ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed for the simultaneous determination of cinnamaldehyde, cinnamic acid, and 2-methoxy cinnamic acid in rat whole blood. It was the first time to study the pharmacokinetics of 2-methoxy cinnamic acid in rat whole blood. Samples were processed by a one-step protein precipitation with acetonitrile-37% formaldehyde (90:10, v:v). Chromatographic separation was performed on a Thermo Scientific C18 column (2.1mm×50mm, 1.9μm) at room temperature. The total run time was 4min. The detection was accomplished by using positive and negative ion electrospray ionization in multiple reaction monitoring mode. The method was linear for all of the analytes over 1000 times concentration range with correlation coefficients greater than 0.99. The lower limits of quantification (LLOQ) were 0.1ng/mL for cinnamaldehyde, 5.8ng/mL for cinnamic acid, and 10ng/mL for 2-methoxy cinnamic acid, respectively. To our knowledge, this was the first time that the LLOQ for cinnamaldehyde in validated methods for biological samples was as low as 0.1ng/mL. Intra- and inter-day precision and accuracy were within ±9% for all of the analytes during the assay validation. Assay recoveries were higher than 80% and the matrix effects were minimal. The half-life were 8.7±0.7h for cinnamaldehyde, 1.0±0.5h for cinnamic acid, and 1.4±0.4h for 2-methoxy cinnamic acid, respectively. The validated assay was firstly applied to the simultaneous quantification of cinnamaldehyde, cinnamic acid, and 2-methoxy cinnamic acid, especially for 2-methoxy cinnamic acid in rat whole blood after oral administration of 15mg/kg essential oil of Cinnamoni Ramulus. It was observed that the Cmax and AUC of 2-methoxy cinnamic acid (0.01% in essential oil of Cinnamoni Ramulus) were greater than those of cinnamaldehyde (83.49% in essential oil of Cinnamoni Ramulus), which implied that 2-methoxy cinnamic acid might be the major bioactive constitutes in essential oil of Cinnamoni Ramulus.

Metabolic profile of 2-(2-hydroxypropanamido) benzoic acid in rats by ultra high performance liquid chromatography combined with Fourier transform ion cyclotron resonance mass spectrometry

An ultra high performance liquid chromatography combined with Fourier transform ion cyclotron resonance mass spectrometry (UHPLC-FT-ICR-MS) method was developed to investigate the in vivo metabolism of 2-(2-hydroxypropanamido) benzoic acid (HPABA), a marine-derived anti-inflammatory drug, for the first time. Plasma, urine, feces and bile samples were collected from male and female rats after a single intragastric administration of HPABA at a dose of 100mg/kg. Besides the parent drug, a total of 13 metabolites (3 phase I and 10 phase II metabolites) were detected and tentatively identified through comparing their mass spectrometry profiles with those of HPABA. Results demonstrated that metabolic pathways of HPABA in rats included decarboxylation, hydroxylation, dehydrogenation, glucuronidation, glycine conjugation and N-acetyl conjugation. In summary, this work provided valuable information regarding the metabolism of HPABA in rats, which would contribute to better understanding of its safety and mechanism of action.

The determination of 2-(2-hydroxypropanamido) benzoic acid enantiomers and their corresponding prodrugs in rat plasma by UHPLC–MS/MS and application to comparative pharmacokinetic study after a single oral dose

A simple and sensitive UHPLC-MS/MS method was developed and validated to determine the pharmacokinetic profile of 2-(2-hydroxypropanamido) benzoic acid (HPABA) enantiomers and their prodrugs in rat plasma. Separation was performed on a Thermo Syncronis C18 column (50mm×2.1mm, 1.7μm; Thermo, USA), which was protected by a high pressure column prefilter (2μm) at a flow rate of 0.4ml/min. Liquid-liquid extraction with ethyl acetate was used to process plasma samples. The separation of two enantiomers, prodrugs of (R)-/(S)-HPABA and internal standard was obtained within a cycle time of 4.5min. The lower limit of quantification of (R)-/(S)-HPABA and prodrugs of (R)-/(S)-HPABA in plasma were 0.01μg/ml and 0.2μg/ml, respectively. (S)-HPABA showed significantly higher AUC, Cmax and a longer t1/2 than (R)-HPABA, indicating higher bioavailability of the (S)-HPABA. Additionally, inversion between HPABA enantiomers was not observed in rats. (R)-/(S)-HPABA showed higher Cmax and AUC than those of their prodrugs. However, the values of t1/2 of prodrugs were higher than those of (R)-/(S)-HPABA. Furthermore, the higher Vz values of prodrugs might improve the targeting of (R)-/(S)-HPABA in rat tissues.

Anesthetic constituents of Zanthoxylum bungeanum Maxim.: A pharmacokinetic study

A sensitive and selective ultra high performance liquid chromatography with tandem mass spectrometry method was established and validated for the simultaneous determination of hydroxy-α-sanshool, hydroxy-β-sanshool, and hydroxy-γ-sanshool in rat plasma after the subcutaneous and intravenous administration of an extract of the pericarp of Zanthoxylum bungeanum Maxim. Piperine was used as the internal standard. The analytes were extracted from rat plasma by liquid-liquid extraction with ethyl acetate and separated on a Thermo Hypersil GOLD C18 column (2.1 mm × 50 mm, 1.9 μm) with a gradient elution system at a flow rate of 0.4 mL/min. The mobile phase consisted of acetonitrile/0.05% formic acid in water and the total analysis time was 4 min. Positive electrospray ionization was performed using multiple reaction monitoring mode for the analytes. The calibration curves of the three analytes were linear over the tested concentration range. The intra- and interday precision was no more than 13.6%. Extraction recovery, matrix effect, and stability were satisfactory in rat plasma. The developed and validated method was suitable for the quantification of hydroxy-α-sanshool, hydroxy-β-sanshool, and hydroxy-γ-sanshool and successfully applied to a pharmacokinetic study of these analytes after subcutaneous and intravenous administration to rats.

Simultaneous determination and pharmacokinetic study of Atractylenolide I, II and III in rat plasma after intragastric administration of Baizhufuling extract and Atractylodis extract by UPLC-MS/MS.

A simple and rapid ultra high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed for the simultaneous determination of Atractylenolide I, II and III in rat plasma. Plasma samples were processed by liquid-liquid extraction with ethyl acetate, using schisandrin as internal standard (IS). Chromatographic separation was accomplished on a Thermo Hypersil GOLD C18 column (2.1mm×50mm, 1.9μm) with mobile phase consisting of acetonitrile and 0.1% formic acid-water (50:50, v/v). The detection was carried out by ESI-MS (positive ionization mode) and low-energy collision dissociation tandem mass spectrometric analyses using the multiple-reaction monitoring (MRM) scan mode. The quantification was performed using the transitions of the protonated molecule→product ion at m/z 231.0→185.1 for Atractylenolide I, at m/z 233.1→187.1 for Atractylenolide II and at m/z 249.1→231.1 for Atractylenolide III, respectively. Method validation revealed excellent linearity over investigated range together with satisfactory intra- and inter-day precision, accuracy, matrix effects and extraction recoveries. This method was successfully applied to the comparative pharmacokinetic study of Atractylenolide I, II and III in rat plasma after intragastric administration of Baizhufuling extract and Atractylodis extract.

Application of an UHPLC–MS/MS method to tissue distribution and excretion study of 2-(2-hydroxypropanamido) benzoic acid in rats

2-(2-Hydroxypropanamido) benzoic acid (HPABA) is a potential anti-inflammatory agent isolated from the marine fungus Penicillium chrysogenum. In vivo tissue distribution and excretion of HPABA were unknown. Therefore, the purpose of this paper was to establish an UHPLC-MS/MS method for the determination of HPABA and proceed to apply for the study of tissue distribution and excretion in rats. The results obtained indicated that HPABA in tissues was rapidly distributed and cleared. The highest level was found in intestine and stomach, followed by kidney, liver, lung and heart. These observations showed that HPABA was well distributed into abundant blood-supply tissues, suggesting that the blood flow or perfusion rate of organs plays a major role. A small amount of HPABA was detected in brain indicated that the ability of it in crossing the blood brain barrier (BBB) was very weak in rats. Excretion studies demonstrated that HPABA was mainly excreted by kidney. The total cumulative excretion of HPABA were 54.3% and 44.1% in male and female rats, respectively. In addition, gender-related differences in the excretion and distribution profile of HPABA were observed in rats.

Plasma Pharmacokinetics, Bioavailability, and Tissue Distribution of Four C-Glycosyl Flavones from Mung Bean (Vigna radiata L.) Seed Extracts in Rat by Ultrahigh-Performance Liquid Chromatography–Tandem Mass Spectrometry

The main polyphenols in mung bean (Vigna radiata L.) seed (MBS), an edible legume with various biological activities, are C-glycosyl flavones (vitexin, isovitexin, isovitexin-6″-O-α-l-glucoside, and dulcinoside). In our study, a validated ultrahigh-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed to quantitate the concentrations of four C-glycosyl flavones from MBS extracts in the plasma and various tissues of rats and successfully applied to study their pharmacological profile and tissue distribution in vivo. Four C-glycosyl flavones were rapidly absorbed after oral administration, achieving a Cmax at around 1.5 h, and they could be distributed widely and rapidly in tested tissues. The concentrations of four C-glycosyl flavones in all of the tested tissues decreased obviously in 4 h, which indicated that there was not a trend of long-term accumulation of them. This is the first time to report on pharmacokinetic and tissue distribution studies of four C-glycosyl flavones in rat. The results provided a significative basis for the application of MBS.

Study on degradation kinetics of 2-(2-hydroxypropanamido) benzoic acid in aqueous solutions and identification of its major degradation product by UHPLC/TOF-MS/MS.

A RP-HPLC method was developed and validated for the degradation kinetic study of 2-(2-hydroxypropanamido) benzoic acid (HPABA), a promising anti-inflammatory drug, which would provide a basis for further studies on HPABA. The effects of pH, temperature, buffer concentration and ionic strength on the degradation kinetics of HPABA were discussed. Experimental parameters such as degradation rate constants (k), activation energy (Ea), acid and alkali catalytic constants (k(ac), k(al)), shelf life (t1/2) and temperature coefficient (Q10) were calculated. The results indicated that degradation kinetics of HPABA followed zero-order reaction kinetics; degradation rate constants (k) of HPABA at different pH values demonstrated that HPABA was more stable in neutral and near-neutral conditions; the function of temperature on k obeyed the Arrhenius equation (r = 0.9933) and HPABA was more stable at lower temperature; with the increase of ionic strength and buffer concentration, the stability of HPABA was decreased. The major unknown degradation product of HPABA was identified by UHPLC/TOF-MS/MS with positive electrospray ionization. Results demonstrated that the hydrolysis product was the primary degradation product of HPABA and it was deduced as anthranilic acid.

Simultaneous determination of five bioactive components of Gancao in rat plasma by UHPLC-MS/MS and its application to comparative pharmacokinetic study of incompatible herb pair Gansui-Gancao and Gansuibanxia Decoction

HighlightsA LC–MS/MS method was developed for determining components of Gancao in rat plasma.Comparative pharmacokinetics of incompatible herb pair and GSBXD was investigated.Gansui decreased the bioavailability of liquorice.Shaoyao increased the bioavailability of Gancao and relived the inhibition of Gansui on liquorice.The mechanism of incompatible herb pair Gansui‐Gancao and GSBXD was firstly elucidated in pharmacokinetic viewpoint. ABSTRACT Incompatible herb pair Gansui‐Gancao is recorded in “eighteen incompatible” medicaments in many monographs of TCM (Traditional Chinese Medicine) which means the two herbs can not be co‐used in most cases. However, Gansuibanxia decoction composed of Gansui(Kansui), Banxia(Pinellia), Shaoyao(Peony) and Gancao (Liquorice) is a traditional Chinese formula which has been clinically employed for the treatment of cancerous ascites, pleural effusion, peritoneal effusion, etc. The purpose of the study was to investigate the pharmacokinetics of main bioactive components in Gancao to explore the reasons why Gansui‐Gancao can be used in Gansuibanxia decoction. A simple, rapid and sensitive UHPLC‐MS/MS method for simultaneous determination of liquiritigenin, isoliquiritigenin, liquiritin, glycyrrhetinic acid and glycyrrhizic acid of liquorice in rat plasma was developed and validated. After extraction from plasma, the analytes and internal standard were separated on a C18 column with the mobile phase consisting of 0.1% acetic acid containing 0.2 mM ammonium acetate in water and acetonitrile via gradient elution. The electrospary ionization source was adopted under the multiple reaction monitoring mode. The method was succesfully applied to a comparative pharmacokinetic study of main bioactive components of Gancao in rat plasma after oral administration of the extracts of Gancao (GC), Gansui‐Gancao (GS‐GC), Shaoyao‐Gancao (SY‐GC), Gansui‐Shaoyao‐Gancao (GS‐SY‐GC) and Gansuibanxia decoction (GSBXD), respectively. The pharmacokinetic parameters had significant differences (P < 0.05) in different groups which showed that Gansui decreased the bioavailability of Gancao, while Shaoyao increased the bioavailability of Gancao. Hence, these may be the pharmacokinetic mechanism of incompatible herb pair Gansui‐Gancao and the reasons why the herb pair can be used in Gansuibanxia decoction.

Studies on New Activities of Enantiomers of 2-(2-Hydroxypropanamido) Benzoic Acid: Antiplatelet Aggregation and Antithrombosis

R-/S-2-(2-Hydroxypropanamido) benzoic acid (R-/S-HPABA), a marine-derived anti-inflammatory drug, however, the antiplatelet and antithrombotic effects have not been investigated. In this paper, the in vitro antiplatelet activities and in vivo antithrombotic effects of R-/S-HPABA were investigated, for the first time. The effects of R-/S-HPABA on platelet aggregation induced by adenosine diphosphate (ADP), collagen (COLL) and arachidonic acid (AA) were evaluated. In addition, the in vivo bleeding time, clotting time, collagen-epinephrine induced pulmonary thrombosis and common carotid artery thrombosis were also investigated in rats. R-/S-HPABA significantly inhibited ADP, COLL and AA induced platelet aggregation in rabbit platelet rich plasma in vitro compared with control group, to a degree similar to that of aspirin. Besides, R-/S-HPABA prolonged bleeding time and clotting time as well as increased the recovery rate obviously in pulmonary thrombosis. Moreover, the level of thromboxane B2 (TXB2) was decreased while the production of 6-keto-prostaglandin F1α (6-keto-PGF1α) was increased markedly by R-/S-HPABA. Furthermore, R-/S-HPABA reduced carotid artery thrombosis weight. These results illustrated that R-/S-HPABA could be a potent antiplatelet aggregation and antithrombotic agent.