Development of High Performance Liquid Chromatography Tandem Mass Spectrometry Method for Quantitative Analysis of Bacopaside-I in Rat Urine and Feces Samples

Significance of the Topic

Bacopaside I is a primary triterpenoid saponin from Bacopa monnieri recognized for its nootropic and neuroprotective properties. Accurate quantification in biological matrices underpins pharmacokinetic and toxicological evaluations.

Objectives and Study Overview

This study aimed to develop and validate a high performance liquid chromatography tandem mass spectrometry method for quantifying Bacopaside I in rat urine and feces. It represents the first LC/MS/MS protocol specifically tailored to this analyte in such complex samples.

Methodology

Sample preparation utilized liquid-liquid extraction from rat urine and fecal homogenates. Chromatographic separation was achieved with a fast gradient on a C18 column under acidic mobile phase conditions. Mass spectrometric detection employed positive electrospray ionization with multiple reaction monitoring transitions of m/z 979.4>483.4 for the analyte and m/z 609.3>195.0 for the internal standard.

Used Instrumentation

• Shimadzu LCMS-8030 triple quadrupole mass spectrometer
• Kinetex C18 column 50 mm × 2.1 mm, 1.7 µm particle size
• Electrospray ion source in positive mode
• Reserpine as internal standard monitored at m/z 609.3>195.0

Main Results and Discussion

The method showed linear response from 8.6 to 900 ng/mL with an r2 of 0.9999. Limits of detection and quantification were approximately 5 and 15 ng/mL, respectively. Urine recovery ranged from 104 to 129 with RSDs below 18, while fecal recovery was lower (29 to 52), highlighting matrix effects that warrant further optimization.

Benefits and Practical Applications of the Method

• Enables sensitive measurement of Bacopaside I in preclinical pharmacokinetic studies
• Supports metabolic profiling and dose optimization in neuropharmacology research
• Applicable for safety assessment and bioavailability evaluation of plant derived saponins

Future Trends and Application Possibilities

• Refinement of extraction procedures to improve fecal analyte recovery
• Extension to related triterpenoid saponins and their metabolites
• Automation and high throughput screening for large scale studies
• Adaptation to human biological fluids for clinical translational research

Conclusion

A robust LC/MS/MS method has been established and validated for Bacopaside I quantification in rat urine and feces, delivering high sensitivity and precision. Further work is required to enhance recovery from fecal matrices and expand analytical scope.

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