The purpose of this study was to clarify the effects of enteral formulas and their food protein and dietary fiber components on the pharmacokinetics of warfarin in rats. Blood samples were collected from 0 to 30 h after the oral administration of 0.5 mg/kg warfarin with or without an enteral formula (Mei Balance R^®, Mei Flow^®, F2 Light^®, or PG Soft EJ^®), food protein (milk protein or casein), or dietary fiber (indigestible dextrin or carrageenan). Plasma warfarin concentration was determined by liquid chromatography-tandem mass spectrometry. Plasma warfarin concentration was lower at the initial phase after co-administration of F2 Light^®, milk protein, or casein compared with the control group, which might have been caused by the suppression of the gastrointestinal absorption of warfarin in accordance with the binding to food protein. Plasma warfarin concentration was significantly higher after co-administration of Mei Flow^®, indigestible dextrin, or carrageenan compared with the control group. This is the first study to find that the plasma concentration of warfarin increases when warfarin is co-administered with enteral formula Mei Flow^® or their dietary fiber components such as indigestible dextrin or carrageenan. Therefore, close attention should be paid to decreases or increases in the plasma warfarin concentration when warfarin is used with enteral formula.

Two steps of enzymatic oxidations from hypoxanthine to uric acid with xanthine oxidase (XOD) were kinetically analyzed by capillary electrophoresis/dynamic frontal analysis. When a substrate solution of hypoxanthine was introduced into a capillary with a separation buffer containing XOD, the enzymatic reaction continuously proceeded during the electrophoresis and a product of xanthine was continuously resolved from the substrate zone. A plateau signal of the product xanthine was detected based on the constant reaction rate with XOD. The plateau height was directly related with the reaction rate, and a MichaelisMenten constant K_M,HXA was successfully determined as 770±40 μmol L⁻¹. When xanthine was used as a substrate, a slope response of uric acid was obtained because of the low concentrations of the substrate and its significant decrease. However, the Michaelis-Menten constant was successfully determined by using the initial reaction rate, and a Michaelis-Menten constant of K_M,XA was determined as 85±6 μmol L⁻¹.

Three-dimensional (3D) HPLC methods are useful tools for the selective analysis of biomolecules in complicated real-world matrices, however, the columns applicable for the second dimension are still limited. In the present study, two types of mixed-mode columns (Singularity MX-102 and 103) were newly designed/developed focusing on the separation of N-protected chiral amino acids. The retention profiles of the two mixed-mode columns were evaluated using 10 proteinogenic amino acids with various chemical properties, namely, alanine, asparagine (Asn), glutamine, glycine, lysine (Lys), phenylalanine, proline, serine (Ser), threonine and valine (Val), and compared to that of a mixed-mode column (Singularity MX-001) previously adopted for the 3D-HPLC analysis of chiral amino acids. The amino acids were derivatized with 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F) and detected by their fluorescence (ex. 470 nm, em. 530 nm). For the mobile phases, mixed solutions of acetonitrile/methanol containing formic acid were used. As a result, NBD-Val eluted the fastest for all the tested mixed-mode columns. However, the NBD-amino acids most strongly retained on the columns were different (Asn for MX-001, Ser for MX-102 and Lys for MX-103). Also, the elution orders of the 10 NBD-amino acids on the three mixed-mode columns were not the same as that on the reversed-phase C18 column, and characteristic elution orders and retention properties were observed for the individual mixed-mode columns. These results indicated that mixed-mode columns reported in the present study have different retention mechanisms, and the development of highly selective 3D-HPLC systems using these columns is expected.

Osimertinib is a third-generation epidermal growth factor receptor tyrosine kinase inhibitor commonly used in Japan. This study aims to develop a simultaneous rapid assay for gefitinib, erlotinib, afatinib, and osimertinib using liquid chromatography-tandem mass spectrometry (LC-MS/MS). These drugs in plasma samples were purified by liquid-liquid extraction with tert-butyl methyl ether, a simple and inexpensive method. The purified drugs were rapidly separated in less than 5 min by isocratic elution using an XBridge Shield RP18 column as the separation column and were sensitively quantified by MS/MS in the electrospray ionization positive mode. The quantification range in plasma for the four drugs was 5–200 nM with a linearity of >0.994 and a lower limit of quantification of 5 nM. The developed method was applied to therapeutic drug monitoring of patients with non–small cell lung cancer treated with oral osimertinib preparation and demonstrated its usefulness.