A strategy for quantitative bioanalysis of non-polar neutral compounds by liquid chromatography/electrospray ionization tandem mass spectrometry: determination of TS-962, a novel acyl-CoA:cholesterol acyltransferase inhibitor, in rabbit aorta and liver tissues.
A strategy for the sensitive and reliable quantitative determination of non-polar neutral compounds in biological matrices by liquid chromatography/electrospray ionization tandem mass spectrometry is described in the context of assay development for TS-962, a novel acyl-CoA:cholesterol acyltransferase (ACAT) inhibitor, in rabbit aorta and liver tissues. The electrospray ionization (ESI) mass spectrum of this compound with a mobile phase of water/acetonitrile did not give abundant [M + H]+ ions, but did give alkali metal cation adducts such as [M + Na]+, [M + CH3CN + Na]+ and [M + K]+ ions. The cationized species are acknowledged as unsuitable precursor ions for selected reaction monitoring (SRM) for various reasons, such as difficulty in obtaining characteristic product ions in low-energy collision-induced dissociation, and irreproducibility of the adduct-ion intensities. To overcome this problem, a solution of 3.4 mM trifluoroacetic acid in 2-propanol was added to the mobile phase as a postcolumn additive, resulting in a decrease of the undesirable adduct formation and significant enhancement of [M + H]+ ion intensity. An attempt was then made to prevent the matrix effect by employing a column-switching system, which allowed direct injection of a large volume of 2-propanolic tissue homogenate (950 microL) followed by sufficient clean-up, separation, and ESI-SRM on-line. This enabled development of a sensitive and reliable assay method for TS-962 in rabbit aorta and liver tissues in the concentration range of 5-500 ng/g wet tissue using a 25-mg aliquot of tissue sample. Application of this method to the determination of aortic TS-962 levels at 24 h after repeated oral administration of this compound (3 mg/kg) once a day for 12 weeks to 1% cholesterol-fed rabbits is also presented. Results showed that TS-962 is well distributed to both the thoracic and abdominal aorta tissues, at levels higher than the 50% inhibitory concentration value of this compound for microsomal ACAT activity from rabbit aorta.
Yamaguchi
J
J
Drug Metabolism Laboratory, Pharmaceutical Research Laboratories, Taisho Pharmaceutical Co., Ltd., 403 Yoshino-cho 1-chome, Ohmiya-shi, Saitama 330-8530, Japan.
Matsuno
Y
Y
Hachiuma
K
K
Ogawa
N
N
Higuchi
S
S
D000081:Acetamides / Q000032:analysis*; D000818:Animals; D001013:Aorta, Thoracic / Q000737:chemistry; D002138:Calibration; D002851:Chromatography, High Pressure Liquid; D004791:Enzyme Inhibitors / Q000032:analysis*; D008099:Liver / Q000737:chemistry*; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000737:chemistry*; D011786:Quality Control; D011817:Rabbits; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization; D002785:Sterol O-Acyltransferase / Q000037:antagonists & inhibitors*
D000081:Acetamides; D004791:Enzyme Inhibitors; C401889:TS 962; D002785:Sterol O-Acyltransferase