Determination of the Peptide AWRK6 in Rat Plasma by Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) and Its Application to Pharmacokinetics

AWRK6 was a synthesized peptide developed based on the natural occurring peptide dybowskin-2CDYa, which was discovered in frog skin in our previous study. Here, a quantitative determination method for AWRK6 analysis in rat plasma by using liquid chromatography-tandem mass spectrometry (LC-MS/MS) was established and validated following U.S. FDA guidelines. A combination of plasma precipitation and liquid–liquid extraction was applied for the extraction. For pharmacokinetics study, the rats were administrated with AWRK6 via intraperitoneal and intravenous injection. The prepared plasma samples were separated on an ODS column and analyzed by tandem MS using precursor-to-product ion pairs of m/z: 533.4→84.2 for AWRK6 and m/z: 401.9→101.1 for internal standard Polymyxin B sulfate in multiple reaction monitoring mode. AWRK6 concentrations in rat plasma peaked at about 1.2 h after intraperitoneal injections at 2.35, 4.7 and 9.4 mg/kg bodyweight. The terminal half-life was around 2.8 h. The absolute bioavailability of AWRK6 was 50% after 3 doses via injection, and the apparent volume of distribution was 4.884 ± 1.736 L. The obtained determination method and pharmacokinetics profiles of AWRK6 provides a basis for further development, and forms a benchmark reference for peptide quantification.

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Pharmacokinetics and bioavailability of hapepunine in mice by ultra-performance liquid chromatography–tandem mass spectrometry

Acta Chromatographica ◽

10.1556/1326.2019.00559 ◽

2020 ◽

Vol 32 (1) ◽

pp. 44-48

Author(s):

Lianguo Chen ◽

Qingwei Zhang ◽

Yijing Lin ◽

Xiaojie Lu ◽

Zuoquan Zhong ◽

...

Keyword(s):

Mass Spectrometry ◽

Liquid Chromatography ◽

Tandem Mass Spectrometry ◽

Multiple Reaction Monitoring ◽

Ultra Performance Liquid Chromatography ◽

Absolute Bioavailability ◽

Tandem Mass ◽

Mouse Blood ◽

Chromatography Tandem Mass Spectrometry ◽

Liquid Chromatography Tandem Mass

An ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) method was established to determine the hapepunine in mouse blood, and the pharmacokinetics of hapepunine after intravenous (1.0 mg/kg) and intragastric (2.5, 5, and 10 mg/kg) administrations was studied. Delavinone was used as an internal standard. The UPLC ethylene bridged hybrid (BEH) C18 column was used for chromatographic separation. The mobile phase consisted of acetonitrile and 0.1% formic acid with a gradient elution flow rate of 0.4 mL/min. Multiple reaction monitoring (MRM) mode was used for quantitative analysis of hapepunine in electrospray ionization (ESI) positive interface. Proteins from mouse blood were removed by acetonitrile precipitation. The verification method was established in accordance with the US Food and Drug Administration (FDA) bioanalytical method validation guidelines. In the concentration range of 1–1000 ng/mL, the hapepunine in the mouse blood was linear (r2 > 0.995), and the lower limit of quantification was 1.0 ng/mL. In the mouse blood, the intra-day precision coefficient of variation (CV) was less than 12%, the inter-day precision CV was less than 14%. The accuracy ranged from 91.7% to 109.3%. The average recovery was higher than 76.7%, and the matrix effect was between 86.0% and 106.4%. The UPLC–MS/MS method was sensitive, rapid, and selective and was successfully applied to the pharmacokinetic study of hapepunine in mice. The absolute bioavailability of hapepunine was 22.0%.

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Pharmacokinetic Effects of l-Tetrahydropalmatine on Ketamine in Rat Plasma by Ultraperformance Liquid Chromatography Tandem Mass Spectrometry

BioMed Research International ◽

10.1155/2020/9259683 ◽

2020 ◽

Vol 2020 ◽

pp. 1-8

Author(s):

Yan Du ◽

Hongliang Su ◽

Jie Cao ◽

Zhiwen Wei ◽

Yujin Wang ◽

...

Keyword(s):

Mass Spectrometry ◽

Liquid Chromatography ◽

Tandem Mass Spectrometry ◽

Multiple Reaction Monitoring ◽

Internal Standard ◽

Rat Plasma ◽

Tandem Mass ◽

Chromatography Tandem Mass Spectrometry ◽

Relative Standard ◽

Liquid Chromatography Tandem Mass

Male Sprague-Dawley rats (n=18) were randomly divided into three groups: a saline group (20 mL/kg by gavage), a ketamine (KET) group (100 mg/kg by gavage), and a KET (the same routes and doses) combined with levo-tetrahydropalmatine (l-THP; 40 mg/kg by gavage) group (n=6). Blood samples were acquired at different time points after drug administration. A simple and sensitive ultraperformance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was established to determine the concentrations of KET and its metabolite, norketamine (NK), in rat plasma. Chromatographic separation was achieved using a BEH C18 column (2.1 mm×50 mm, 1.7 μm) with chlorpheniramine maleate (Chlor-Trimeton) as an internal standard (IS). The initial mobile phase consisted of acetonitrile–water with 0.1% methanoic acid (80 : 20, v/v). The multiple reaction monitoring (MRM) modes of m/z 238.1→m/z 179.1 for KET, m/z 224.1→m/z 207.1 for NK, and m/z 275→m/z 230 for Chlor-Trimeton (IS) were utilized to conduct a quantitative analysis. Calibration curves of KET and NK in rat plasma demonstrated good linearity in the range of 2.5–500 ng/mL (r>0.9994), and the lower limit of quantification (LLOQ) was 2.5 ng/mL for both. Moreover, the intra- and interday precision relative standard deviation (RSD) of KET and NK were less than 4.31% and 6.53%, respectively. The accuracies (relative error) of KET and NK were below -1.41% and -6.07%, respectively. The extraction recoveries of KET and NK were more than 81.23±3.45% and 80.42±4.57%, respectively. This sensitive, rapid, and selective UPLC-MS/MS method was successfully applied to study the pharmacokinetic effects of l-THP on KET after gastric gavage. The results demonstrated that l-THP could increase the bioavailability of KET and promote the metabolism of KET. The results showed that l-THP has pharmacokinetics effects on KET in rat plasma.

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Method Development and Validation of Darifenacin in Rat Plasma by Liquid Chromatography-Tandem Mass Spectrometry: Application to a Pharmacokinetic Study

ISRN Spectroscopy ◽

10.5402/2012/659257 ◽

2012 ◽

Vol 2012 ◽

pp. 1-9

Author(s):

Thejomoorthy Karavadi ◽

B. R. Challa

Keyword(s):

Mass Spectrometry ◽

Liquid Chromatography ◽

Tandem Mass Spectrometry ◽

Pharmacokinetic Study ◽

Method Development ◽

Multiple Reaction Monitoring ◽

Rat Plasma ◽

Tandem Mass ◽

Chromatography Tandem Mass Spectrometry ◽

Liquid Chromatography Tandem Mass

A selective, sensitive, and high-throughput liquid chromatography-tandem mass spectrometry (LC-ESI-MS/MS) method has been developed and validated for the quantitation of darifenacin in rat plasma. Sample clean up involved liquid-liquid extraction (LLE) and used 100 μL of rat plasma. Zorbax, SB C18, 4.6×75 mm, 3.5 μm particle size analytical column using 10 mM ammonium acetate buffer (pH 5) and methanol (10 : 90, v/v) as the mobile phase was used. The parent → product ion transitions for the drug (m/z 427.3 → 147.3) and IS (m/z 431.4 → 151.2) were monitored on a triple quadrupole mass spectrometer, operating in the multiple reaction monitoring (MRM) and positive ion mode. The method was validated over the concentration range of 10.00–20000.00 pg/mL for darifenacin. The method was successfully applied into a pharmacokinetic study in rat plasma under fasting conditions.

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