Detection of 6-(methylsulfinyl)hexyl isothiocyanate (6-MITC) and its conjugate with N-acetyl-L-cysteine (NAC) by high performance liquid chromatograpy-atmospheric pressure chemical ionization mass spectrometry (HPLC-MS/APCI).
A method using high-performance liquid chromatography (HPLC) and atmospheric pressure chemical ionization (APCI) mass spectrometry (MS) was established for the detection of 6-(methylsulfinyl)hexyl isothiocyanate (6-MITC) and its conjugate with N-acetyl-L-cysteine (NAC). The optimal chromatographic conditions were obtained on an ODS column (150 x 4.6 mm, 3 microm) with the column temperature at 37 degrees C. The mobile phase consisted of a methanol-0.1% trifluoroacetic acid (TFA) mixture (50 : 50, v/v), and the flow rate was 0.3 ml/min. The detection wavelength was set at 220 nm. The identities of the peaks were accomplished by comparing retention times (tR), UV and mass data. All calibration curves showed good linear regression (correlation coefficients for 6-MITC and NAC>0.999) within test ranges. The developed method provided satisfactory precision calculated as percent coefficient of variation with overall intra-day and inter-day variations of less than 5% (4.1 and 4.9% for 6-MITC; 4.2 and 4.9% for NAC). Both 6-MITC and NAC had good responses in the positive APCI and formed strong [M+H]+ ions in the full scan spectra at an m/z of 206 and 164, respectively. The presence of the [M+H]+ ion for the 6-MITC/NAC conjugate was also observed at an m/z of 369. To our best knowledge, this is the first report that describes the formation of the 6-MITC/NAC conjugate and its detection method by HPLC-MS.
Yamaguchi
Hideaki
H
Department of Clinical Pharmacy, Faculty of Pharmaceutical Sciences, Aomori University, Aomori, Japan. yamaguch@aomori-u.ac.jp
Noshita
Toshiro
T
Kidachi
Yumi
Y
Umetsu
Hironori
H
Fuke
Yoko
Y
Ryoyama
Kazuo
K
D000111:Acetylcysteine / Q000032:analysis*; D002138:Calibration; D002851:Chromatography, High Pressure Liquid; D007202:Indicators and Reagents; D017879:Isothiocyanates / Q000032:analysis*; D013058:Mass Spectrometry; D015203:Reproducibility of Results; D013056:Spectrophotometry, Ultraviolet
C505386:6-(Methylsulfinyl)hexyl isothiocyanate; D007202:Indicators and Reagents; D017879:Isothiocyanates; D000111:Acetylcysteine