Page 16 - Shimadzu Journal vol.3 Issue3

Drug Discovery

Utility of high resolution MALDI imaging in drug discovery:
Histological distribution of gentamicin in proximal renal tubules of rats

*Part of this work was presented in a poster session of the 63th ASMS Conference, May 13 - June 4, 2015, St. Luis.
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Hidefumi Kaji ; Hiroyuki Hashimoto ; Masayoshi Saito ; Takushi Yamamoto ; Noriyuki Ojima 2
1: Mitsubishi Tanabe Pharma Corporation, Saitama, Japan
2: Shimadzu Corporation, Kyoto, Japan
Overview
Observation MALDI imaging
⾣ MS/MS imaging of gentamicin was performed from high-selectivity
analysis of endogenous metabolites in biological tissue.
⾣ High resolution (10 µm) imaging of rat renal cortex showed that Movement
gentamicin is specifically distributed in proximal renal tubules.
⾣ New matrix application method is useful for the sensitive MALDI
imaging.
Imaging Mass Microscope
1. Introduction
Laser: Nd:YAG laser (335 nm)
In pharmacology and toxicology, localization of a drug molecule in the Collision energy: 50
Laser shot: 200 shot/pixcel
target tissue of organs provides very important in vivo biological Laser diameter: 50 or 10 µm
information. Imaging mass spectrometry (IMS) is increasingly used in Matrix: CHCA
drug discovery and development during preclinical studies.
Gentamicin is an antibiotic to treat infections, but it possesses renal Structures of Gentamicin
toxicity. As it is excreted in the urine, the kidney tissues of a person We analyzed gentamicin C1, because it had the highest efficacy.
being treated with gentamicin are almost constantly bathed in
gentamicin. The objective of the study is to define the specific
distribution of the parent drug in proximal renal tubules of rats dosed
gentamicin using MALDI imaging.

2. Methods
At 2 h after single intravenous (I.V.) administration (3, 10, or 30
Sample preparation
mg/kg) of gentamicin to male SD rats, 5-µm sections taken from the
single kidney of each rat were prepared. The tissue sections were
coated with CHCA by sublimation using an automated sample
treatment system (iMLayer™, Shimadzu Corporation, Japan), and
analyzed using an ion trap-time of flight (IT-TOF) tandem mass
spectrometer equipped with MALDI source†. This instrument is a
I.V. administration Extraction of kidneys Snap-freeze in
combination of an optical microscope which allows the observation of liquid nitrogen
high-resolution morphological images, with a mass spectrometer
which identifies and visualizes the distribution of specific molecules.
Pretreatment
The other kidney was used for determining the concentrations of LC-MS/MS
gentamicin by LC-MS/MS.
Homoginized kidney

IMS

Cryosectioned (10 µm)

Matrix application
iMLayer™
Spray: 10 mg/mL CHCA, 0.1% TFA in 60% acetonitrile solution
(Matrix Vapor Deposition System, Shimadzu)
Sublimation: time; 20 min, thickness; ca.2 µm
High reproducibility and minute matrix crystals
Nebulizer: applied methanol mist, 5 min, particle size: <5 µm
Pre-coated matrix: sublimation before thaw-mounted tissue section

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