Background: Many phenolic C-glycosides possess nutritional benefits and pharmacological efficacies. However, the MS/MS fragmentation pattern of phenolic C-glycosides analysis is understudied. This paper aims to determine the MS/MS fragmentation patterns of phenolic C-glycosides. Method: Ten compounds with different sugar moieties, aglycones, and substitutes were analyzed to determine the impact of these structural features on MS/MS fragmentation using UPLC-QTOF-MS analysis. Results: The results showed that water loss followed by RDA reaction and alpha cleavage in the C-C bonded sugar moieties are the major fragmentation pathways. Additionally, the sugar cleavage was not affected by the skeleton and the substitute of the aglycones. These results suggested that the fragmentation patterns of phenolic C-glycosides differ from those in the O-glycosides, where the O-C glycosidic bond is the most cleavage-liable bond in MS/MS analysis. Conclusions: These MS/MS fragmentation patterns can be used for the identification of C-glycosides from dietary components and herbal medicine as well as developing robust methods using MRM methods to quantify C-glycosides.

Oxaliplatin (OXP), a third-generation platinum-based chemotherapy drug, was often indirectly analyzed via total platinum by an ICP-MS because it was difficult to directly quantify using an LC-MS/MS method, due to its instability, bad column separability and severe MS signal inhibition. Here, we developed and validated a specific, sensitive and reproducible LC-MS/MS method for the quantification of OXP itself in rat plasma and tongue tissue on a SCIEX 4000 QTRAP^® MS/MS system equipped with a Phenomenex Lux 5u Cellulose-1 column (250 × 4.6 mm, 5 μm). This method was validated at the lower limit of detection (LOD) and the lower limit of quantitation (LLOQ) of 5 ng/mL and 10 ng/mL, with linearity of 10–5000 ng/mL (r² > 0.99) and 10–2500 ng/mL (r² > 0.99), in rat plasma and tongue homogenates, respectively. The intra- and inter-day precision (CV%) and accuracy (RE%) were within 15% for LLOQ, low-, medium- and high-quality control samples. The mean extraction recoveries were around 50% and 80% for plasma and tongue homogenates, respectively. This assay was successfully applied to pharmacokinetics study following intravenous administration of OXP, as well as tongue tissue distribution after 1 h and 4 h of a novel oral mucosal patch application.

Objectives: A thorough understanding of pharmacokinetics and metabolism is critical during early drug development. This study investigates the absorption, distribution, metabolism, and excretion (ADME) profile of R14, a novel compound, using a combination of in vitro and in vivo approaches. Methods: In vitro studies included Caco-2 permeability assays, metabolic stability evaluations in liver microsomes and hepatocytes, and identification of CYP isoforms responsible for R14 metabolism. In vivo pharmacokinetic and metabolic profiling was conducted in rats following oral administration. R14 was quantified using UHPLC-MS/MS. Metabolites were identified using high-resolution UHPLC- QTOF MS/MS, and relative exposure was estimated using peak area-derived AUCs. Results: R14 exhibited low oral bioavailability (13.4%) and high systemic clearance (2.63 L/h/kg), indicating high hepatic extraction. A total of 21 plasma and 38 urine metabolites were identified. Major metabolic pathways included initial hydroxylation and hydrogenation, followed by sequential methylation and Phase II conjugations (glucuronidation and sulfation). Key metabolites (M3, M4, M22, M38) accounted for the majority of systemic exposure. Less than 1% of the unchanged drug was excreted in urine, confirming extensive metabolism. Notably, discrepancies between in vitro and in vivo metabolite profiles suggested rapid further transformation of initial metabolites in vivo, which were not fully captured in vitro. Conclusions: This study demonstrates an efficient and integrated strategy for early-phase ADME characterization. The combined use of in vitro assays and in vivo studies, guided by advanced analytical techniques, provides a robust framework for understanding drug metabolism. These findings can inform drug optimization and help minimize risks in later stages of development.

MPI8, a peptidyl aldehyde, is a potent antiviral agent against coronavirus. Due to unique tri-peptide bonds and the formyl functional group, the bioassay of MPI8 in plasma was challenged by a strong interference from water MPI8. Using QTOF LC-MS/MS, we identified MPI8•H₂O as the major interference form that co-existed with MPI8 in aqueous and biological media. To avoid the resolution of MPI8 and MPI8•H₂O observed on reverse phase columns, we found that a Kinetex hydrophilic interaction liquid chromatography (HILIC) column provided co-elution of both MPI8 and MPI8•H₂O with a good single chromatographic peak and column retention of MPI8 which is suitable for quantification. Thus, a sensitive, specific, and reproducible LC-MS/MS method for the quantification of MPI8 in rat plasma was developed and validated using a triple QUAD LC-MS/MS. The chromatographic separation was achieved on a Kinetex HILIC column with a flow rate of 0.4 mL/min under gradient elution. The calibration curves were linear (r² > 0.99) over MPI8 concentrations from 0.5–500 ng/mL. The accuracy and precision are within acceptable guidance levels. The mean matrix effect and recovery were 139% and 73%, respectively. No significant degradation of MPI8 occurred under the experimental conditions. The method was successfully applied to a pharmacokinetic study of MPI8 after administration of MPI8 sulfonate in rats.

Background: Diacerein, a prodrug of Rhein, is commonly prescribed for the management of joint disorders, specifically osteoarthritis. This study aimed to develop and validate an LC-MS/MS method to quantify Rhein and its major metabolites, Rhein-G1 and Rhein-G2, in plasma samples. Method: An ACE C18 column was used for chromatographic separation with a mobile phase comprising ammonium acetate at a concentration of 1.0 mM and acetonitrile. Detection was achieved using a Sciex 4000 Q-Trap LC-MS/MS, operated in negative ion mode with multiple reaction monitoring (MRM). Results: The analytical results indicated that the lower limit of quantification (LLOQ) for Rhein and its glucuronides was 7.81 nM. Precision was consistently below 9.14%, while accuracy remained within the acceptable range of 80.1–104.2%. We also verified the method’s matrix effect recovery and stability variance, which were less than 12.60% and 10.37%, respectively. The pharmacokinetic study demonstrated that diacerein is swiftly metabolized into Rhein, and then Rhein subsequently undergoes glucuronidation, forming detectable concentrations of Rhein-G1 and Rhein-G2 in plasma. Conclusions: This new LC-MS/MS method proved to be both sensitive and selective, allowing for pharmacokinetic studies in rats.