Hisao Iwasaki

Determination of nitrate in meat products and cheese by gas-liquid chromatography with electron-capture detection

Abstract A simple, sensitive and practical method for the determination of nitrate in various meat products and cheeses is described. The method is based on the nitration of 2-sec.-butylphenol in ca. 57% sulphuric acid t form 4-nitro-2-sec.-butylphenol. After nitration, the toluene extract is re-extracted with an alkaline solution and subsequently analysed by gas-liquid chromatography with an electron-capture detector (GLC-ECD) using a column of OV-17 on Chromosorb W HP after pentafluorobenzoylation and extraction with n-hexane. The nitrate concentration is calculated from the peak height. Amounts of 0.05-1.0 μg of nitrate can be determined. The detection limit for nitrate is 0.006 μg/ml. The procedure for determining nitrate in meat products and cheeses involves direct analysis by GLC-ECD without clean-up; the detection limit is 0.07 ppm and the recovery from meat products and cheeses ranged from 96.8 to 99.0% at the 5 ppm level and from 94.7 to 98.6% at the 10 ppm level. The nitrated compound of 2-sec.-butylphenol was identified as 4-nitro-2-sec.-butylphenol by thin-layer chromatography and nuclear magnetic resonance spectroscopy and the final pentafluorobenzoylated product was confirmed by combined gas chromatography-mass spectrometry.

Spectrophotometric determination of nitrate in vegetable products using 2-sec-butylphenol

A procedure is described for the determination of nitrate in vegetable products, based on the quantitative reaction of nitrate and 2-sec-butylphenol in sulphuric acid (5 + 7), and the subsequent extraction and measurement of the yellow complex formed in alkaline medium. The colour reaction is sensitive and stable and absorbances measured at 418 nm obey Beers law for concentrations of nitrate-nitrogen between 0.13 and 2.50 µg ml–1. Various possible interferents in vegetable products do not interfere with the nitration of 2-sec-butylphenol. Recoveries of nitrate from vegetable products were satisfactory and the standard deviation for the whole procedure was 1.41% for the 42 determinations; the detection limit of the method is 1.3 p.p.m. for nitratenitrogen.

Determination of the stable isotope of nitrite flux in the blood of mice by gas chromatography—mass spectrometry with selected ion monitoring

A simple and reliable method for the determination of the stable isotope of nitrite (15NO2) flux in the blood of mice is described. It is based on the reaction of 15NO2 with 1-hydrazinophthalazine in acidic solution to form [15N]tetrazolophthalazine, a stable compound which can be extracted with an organic solvent and then determined by gas chromatography-mass spectrometry with selected ion monitoring using the pentafluorobenzoyl ester of 2,4-dinitro-6-sec.-butylphenol as an internal standard. Amounts of 0.2-10 micrograms of 15NO2 can be determined. The detection limit of 15NO2 was 0.1 microgram/ml. This is a specific method for 15NO2. The procedure for determining 15NO2 in the blood of mice involves extraction with solvent, followed by further clean-up by alumina column chromatography; the detection limit is 0.05 microgram. With the new technique we were able to perform a metabolic fate study of nitrite in the blood of mice following a single oral dose of 15NO2 or the stable isotope of nitrate (15NO3).