Our innovative LC-MS/MS method enables precise quantification of 15-Hydroxy Lubiprostone in human plasma, addressing challenges like low detection levels and complex matrix interferences. This validated approach ensures accurate and reliable results, crucial for bioequivalence studies and clinical applications.
Lubiprostone is a medication used in the management of idiopathic chronic constipation. As a derivative of prostaglandin E1, Lubiprostone is a chloride channel agonist, which is proposed to act by selectively activating voltage-dependent type 2 chloride channels in the small intestine. This action stimulates intestinal chloride secretion and triggers the diffusion of sodium and fluid into the intestine.
Primary Metabolic Pathway of Lubiprostone In Vivo:
Lubiprostone has low systemic exposure after oral administration, resulting in very low plasma concentrations. Consequently, pharmacokinetic parameters such as Cmax, AUC, and t1/2 cannot be reliably calculated. After oral administration, Lubiprostone is rapidly metabolized to 15-Hydroxy Lubiprostone. Therefore, FDA guidance for evaluating the bioequivalence of Lubiprostone capsules recommends 15-Hydroxy Lubiprostone as the evaluation indicator. Hence, 15-Hydroxy Lubiprostone is measured in plasma following a dose of Lubiprostone.
At Lambda Therapeutic Research, we have developed and validated a highly sensitive bioanalytical method for the quantification of 15-Hydroxy Lubiprostone.
Challenges:
- Developing a method for derivatives of prostaglandins is challenging due to the presence of other prostaglandins in the plasma matrix.
- The low dosage of Lubiprostone results in detection levels in the low picogram range for 15-Hydroxy Lubiprostone.
- As a derivative of prostaglandin, other prostaglandins with similar structures and molecular weights are also present in plasma, leading to multiple impurity peaks at or near the retention time of the analyte.
- Due to the structural similarity with other prostaglandins, derivatization approaches are ineffective.
- The low detection level and the use of a highly sensitive mass spectrometer resulted in a high baseline, hindering low-level detection in the matrix.
Given these challenges, mass parameters were optimized, and the selection of the daughter ion was crucial. While there are many published methods for the quantification of 15-Hydroxy Lubiprostone, at Lambda, we have developed a robust method by selecting a unique daughter ion. Detection and sufficient recovery were achieved using a liquid-liquid extraction technique. Reproducible chromatography was achieved through an extensive gradient program with a 10-minute run time using a core-shell technology analytical column.
Chromatography of LLOQ Sample:
Chromatography of Blank Sample:
Results:
Below are the global statistics for three precision and accuracy runs in human plasma.
| Quality control samples Id | DQC (1/5) | HQC | MQC | LMQC | LQC | LOQQC |
|---|---|---|---|---|---|---|
| Mean (pg/mL) | 837.2842 | 238.1477 | 139.1158 | 40.3883 | 2.8057 | 1.0428 |
| SD ± | 45.7294 | 13.49126 | 8.80338 | 2.56019 | 0.29704 | 0.17826 |
| Precision (%CV) | 5.5 | 5.7 | 6.3 | 6.3 | 10.6 | 17.1 |
| Accuracy (%) | 93.2 | 100.7 | 98 | 101.6 | 93.6 | 101.1 |
| n | 18 | 18 | 18 | 18 | 18 | 18 |
We generated matrix stability data, including benchtop stability for 16.0 hours and freeze-thaw stability for five cycles as part of the validation process. Interim long-term stability in the matrix for 13 days has also been performed.
Application:
The validated method has been successfully applied to analyze study samples in a pivotal study. The incurred sample reproducibility experiment was conducted and found to be well within the acceptable criteria, thus demonstrating the method’s reproducibility for incurred samples.
Conclusion:
A selective, precise, accurate, and highly sensitive LC-MS/MS method is now available for the quantification of 15-Hydroxy Lubiprostone in human plasma.
Extensive Experience:
Lambda Therapeutic Research offers a wide range of bioanalytical capabilities specializing in cell-based assays, biomarkers, immunogenicity, and pharmacokinetics (PK). With state-of-the-art facilities and a team of experienced scientists, Lambda provides comprehensive solutions for the analysis of small molecules, biomarkers, biologics, and therapeutic trace elements.
