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[Detecting penehyclidine hydrochloride in rats plasma and tissues with HPLC-MS/MS]
Qin Yu1, Mao-Zhi Liang, Feng Nan
1Laboratory of Clinical Pharmacology, GCP Center, West China Hospital, Sichuan University, Chengdu, China.
Objective:
To develop a method for detecting penehyclidine hydrochloride (PH) in mouse plasma and tissues using HPLC-MS/MS.
Methods:
The plasma and tissue samples were extracted with petroleum ether, ethyl ether (70:30). The isolation of PH was achieved on an Allure C18, (50 x 2.1 mm, 5 microm) column, with methanol/5 mmol/L ammonium acetate/triethylamine = 90/10/0.05 (adjusted pH 5.8 by formic acid as mobile phase. Benzhydramine was used as the internal standard.
Results:
The calibration curves were linear over the range of 0.39-200 ng/mL for plasma and 0.391-100 ng/mL for tissues. The extraction recovered 67.32-70.80% of PH in plasma, 84.99%-89.27% of PH in lung tissues, and 76.86%-81.98% of PH in brain tissues. The HPLC detected 97.66%-99.97% of PH in extracted samples of plasma, 96.55%-101.65% extracted samples of lung and 95.18%-100.21% of extracted samples of brain. The within-day RSD were 1.94%-2.93%, 1.73%-3.70% and 2.35%-2.79% for plasma, lung, and brain, respectively. The between-day RSD were 3.00%-3.85% 2.86%-3.94% and 2.77%-5.00% for plasma, lung, and brain, respectively. Both the long-term and freeze-thaw stabilities were acceptable. The long-term RSD were 1.52%-5.26%, 1.88%-2.93%, and 2.22%-3.76% for plasma, lung, and brain, respectively. The freeze-thaw RSD were 0.38%-3.55%, 2.79%-9.60%, and 1.35%-2.29% for plasma, lung, and brain, respectively.
Conclusion:
The assay is simple and accurate, with good reproducibility, and can satisfy to the needs of pharmacokinetics and relative bioavailability studies on penehyclidine hydrochloride.
Insights
This study presents a validated HPLC-MS/MS method for detecting penehyclidine hydrochloride (PH) in mouse plasma and tissues. The assay demonstrates accuracy and reproducibility, suitable for pharmacokinetic studies.
Area of Science:
- Pharmacology
- Analytical Chemistry
- Biomedical Science
Background:
- Penehyclidine hydrochloride (PH) is a drug requiring accurate quantification.
- Existing methods may lack sensitivity or specificity for biological matrices.
- Pharmacokinetic and bioavailability studies necessitate reliable analytical techniques.
Purpose of the Study:
- To develop and validate a sensitive and accurate HPLC-MS/MS method.
- To quantify penehyclidine hydrochloride (PH) in mouse plasma and various tissue homogenates.
- To establish a robust assay for supporting preclinical drug development studies.
Main Methods:
- Sample preparation involved liquid-liquid extraction using petroleum ether and ethyl ether.
- High-Performance Liquid Chromatography (HPLC) separation was performed on an Allure C18 column.
- Detection and quantification were achieved using tandem mass spectrometry (MS/MS) with benzhydramine as the internal standard.
Main Results:
- Linear calibration curves were established for plasma (0.39-200 ng/mL) and tissues (0.391-100 ng/mL).
- Extraction recoveries ranged from 67.32% to 89.27% across different matrices.
- The method exhibited high accuracy (95.18%-101.65%) and excellent reproducibility (RSD within-day: 1.73%-2.93%; between-day: 2.77%-5.00%).
- Stability studies confirmed the integrity of PH in samples under various storage conditions.
Conclusions:
- The developed HPLC-MS/MS assay is simple, accurate, and highly reproducible.
- The method is suitable for quantifying penehyclidine hydrochloride (PH) in mouse plasma and tissues.
- This validated assay meets the requirements for pharmacokinetic and relative bioavailability studies.
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