A validated liquid chromatography-tandem mass spectrometry method for the determination of l-hyoscyamine in human plasma: Application in a clinical study.

Atropine is a racemic mixture of d- and l-hyoscyamine, but only l-hyoscyamine is the effective ingredient. In this study, a new, sensitive, stable, and selective LC/MS assay was developed for the determination of l-hyoscyamine and applied to a clinical study. The parent-product (m/z) transition pair of l-hyoscyamine was 290.1 → 124.1. Chromatographic separations were performed using a chiral MZ column (250 mm × 4.6 mm, 5.0 µm) by a stepwise gradient elution mode with n-hexane, isopropanol, and diethylamine as mobile phases. l-Hyoscyamine in human plasma was extracted by liquid-liquid extraction. This assay displayed a good linearity over a concentration range of 20.0-400 pg/mL for l-hyoscyamine. The accuracy of the validation assay for l-hyoscyamine ranged from -2.7% to 4.5%, and the precision was within 6.3% coefficient of variation. l-Hyoscyamine in human plasma remained stable at different storage conditions. The method has been successfully applied to plasma samples obtained from a safety study in humans.

Simultaneous quantification of marine neutral neoagaro-oligosaccharides and agar-oligosaccharides by the UHPLC-MS/MS method: application to the intestinal transport study by using the Caco-2 cell monolayer.

A sensitive and robust ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was established for the first time to simultaneously quantify marine neutral neoagaro-oligosaccharides (NAOS) and agar-oligosaccharides (AOS) with different degrees of polymerization (DP) in Hanks' balanced salt solution (HBSS). The separation was achieved on a BEH amide column using a mobile phase of acetonitrile-10 mmol L-1 ammonium acetate (58 : 42, v/v) with an isocratic elution program. The total analysis time was 3.5 min. The mass spectra were acquired in the multiple reaction monitoring (MRM) pattern by using a heated-electrospray ionization (H-ESI) source operating in the positive ionization mode. The linear range was 40-20 000 nmol L-1. The accuracy and precision ranged from 91.5 to 110.0% and 0.9 to 10.4%, respectively. The extraction recovery was consistent and reproducible. The stability was within 90.3-110.8%. The matrix effect, carryover, and dilution integrity were all satisfactory. Moreover, the validated method was successfully applied to the intestinal transport study by using the Caco-2 cell monolayer in vitro. The results revealed that neoagarobiose, neoagarotetraose, neoagarohexaose, agarotriose, agaropentose, and agaroheptose were transported by a paracellular pathway.