BUNSEKI KAGAKU Abstracts

Some agricultural chemicals having high polarity, low vapor pressures or high thermal instability are not directly amenable to GC and GC/MS. We established new analytical methods using LC/MS and LC/MS/MS for such agricultural chemicals as pyrethroids, neonicotinoids and guanidines. The analytical method of a pyrethroid insecticide, tralomethrin, using gas chromatography (GC) with electron-capture and mass spectrometry (MS) detectors is to measure the deltamethrin transformed from tralomethrin in the injector port of the GC system. By using LC/MS, however, both pesticides could be separated and determined simultaneously. Although gas chromatography (GC) and high-pressure liquid chromatography with fluorescence (HPLC-FL) have been used for guanidino pesticides determination, a derivatization step is usually required. Moreover, guanidino pesticides that easily adsorb sample matrixes could also be analyzed by using LC/MS without a derivatization step. In the case of nitenpyram, a neonicotinoid pesticide, its metabolites have been determined by GC after derivatization. However, ten compounds including the metabolites could be simultaneously and selectively analyzed by using LC/MS/MS. In addition, the effect of additives on the extraction efficiency, an important factor for the analytical method, could be outlined.

Angelica keiskei KOIDZUMI (Ashitaba in Japanese) is a perennial plant belonging to the Umbelliferae family. Its original home is Hachijo Island and the leaves have been used as a vegetable and a herbal medicine. This plant contains vast quantities of yellow substances throughout the roots, stems and leaves, and was classified into two types on the basis of morphological characters and HPLC profiles of the coumarins components. Further, these yellow components were studies for a structure elucidation, and were characterized as new chalcones. Further, numerous benefits were found, including the suppression of gastric acid secretion, an action as an antitumor agent, an antithrombotic effect, a reduction in blood pressure, suppression of histamine release etc. For the purpose of promoting the development of the local agricultural in Idzu Islands, line breeding of Angelica keiskei with high chalcone content was carried out.

Trace elements in wheat flour were determined by an X-ray fluorescence (XRF) analysis in order to identify their production area. By optimizing the measurement conditions of an energy-dispersive XRF spectrometer equipped with 3-dimentional polarization optics, the analysis of heavy elements at sub-ppm levels was possible, and linear calibration curves for 17 elements (Na, Mg, Al, P, S, Cl, K, Ca, Mn, Fe, Cu, Zn, Br, Rb, Sr, Mo, Cd) in wheat flour were obtained. We determined the trace elements for 63 wheat flour samples that were produced in Japan or imported from overseas. The analytical results were subjected to multivariate analysis to discriminate the domestic and imported wheat flours. Principal-components analysis successfully distinguished the two origins. The present study has demonstrated that the XRF technique can provide a rapid and easy way for the provenance analysis of wheat flours, which will be further applied to the quality control of commercial food products.

Carbon, nitrogen, and oxygen stable isotope ratios of beef from Japan, the USA, Australia, and New Zealand were examined as potential tool for verifying the geographical origin of beef. Each element of the isotope ratios of beef reflects the environmental fattening condition (region and feed) rather than the breed. Beef from USA is characterized by a higher carbon isotope ratio than that from other countries. Also, beef from Australia is characterized by higher oxygen isotope ratio than that from other countries. Moreover, we determined the carbon, nitrogen and oxygen stable isotope ratios of beef from Okinawa, Mie, Yamagata, and Hokkaido. Carbon and nitrogen isotope ratios did not show much difference among the four regions. On the other hand, the oxygen isotope ratios were the highest in beef from Okinawa, which was the most southerly area among the four regions. Such ratios became lower as the latitude became higher. These results suggest that a comparison of carbon, nitrogen, and oxygen stable isotope ratios is applicable as a potential tool for the discrimination of beef between not only different countries but also different regions within Japan.

A rapid and simple detection for paraquat (PQ) and diquat (DQ) intentionally mixed in beverages was developed. A pipette tip-type micro volume solid-phase extraction device, OMIX^® pipette tip (solid-phase type ; C18, C4 and SCX) was used for the extraction of PQ and DQ. The extracted PQ and DQ were successfully detected by a color test and/or liquid chromatography/mass spectrometry (LC/MS). An 100 μL volume of a sample (coke or coffee spiked with appropriate amounts of PQ and DQ) was alkalinized by adding 100 μL of 0.1 M sodium carbonate buffer (pH 10). Then, PQ and DQ in the mixture were retained onto the C18 or C4 phase of an OMIX^® pipette tip by 10-times repeating the aspirate/dispense cycles using a manual micropipettor. After being washed with water and methanol, PQ and DQ were eluted with 100 μL of 5 mM ammonium formate (pH 2.5) by 5-times repeating the aspirate/dispense cycles. It took only 3 minutes for one sample preparation. The color test was carried out by adding an 1 M sodium hydroxide solution and sodium hydrosulfite to half of the extract and the remaining part was subjected to LC/MS analysis. The extracted PQ and DQ were successfully separated and each of the [M＋H]^＋ ions were observed under the optimized LC/MS condition. The extraction procedure using an SCX-type pipette tip was more simple than the other type of solid phase. PQ and DQ were retained to the SCX phase without any adjustment of the pH and eluted with a saturated aqueous sodium chloride solution. This procedure was suitable for a color test, but not for LC/MS by because of the large amount of sodium chloride in the extract. This method was very useful for suspicious cases of PQ and DQ poisonings in beverages.

A sample preparation method was developed for the determination of qunolones in royal jerry (RJ) using immunoaffinity chromatography. For this purpose, an immunoaffinity column was prepared by packing a quinolone-specific monoclonal antibody immobilized to agarose resin. An RJ sample diluted with phosphate buffer was centrifuged. Then the supernatant was applied to the column. After the column was washed, quinolones were eluted with glycine-HCl. Quinolones in the eluate were determined by HPLC with fluorescence detection. No interfering peak was found on the chromatogram of the RJ sample. The recoveries of quinolones from the samples were over 75％ at fortification levels of 25 ng/g for norfloxacin, 10 ng/g for danofloxacin and 50 ng/g for enrofloxacin and ciprofloxacin. This sample-preprocessing method using immunoaffinity chromatography was found to be effective and suitable for determining residual quinolones in RJ.

A portable total reflection X-ray fluorescence (TXRF) spectrometer has been applied to a trace elemental analysis of wines. Sulfur, K, Mn, Fe, and Rb were detected. These five elements were quantified by using 1 ppm of Co as an internal standard. The quantified concentrations ranged from sub-ppm to several hundred ppm. Because of organic substances in wines, the scattering of the incident X-rays from the dry residues of wines becomes strong. Therefore, a high spectral background appears in TXRF spectra of wines. Because of this background, relative standard deviations of the quantified concentrations were from 4 to 28％. Although the high spectral background appeared in the TXRF spectra of the wines, a detection limit down to several tens of ppb was achieved. The present portable spectrometer can be applied to screening for trace elements in wines before an accurate and precise analysis using a large elemental analyzer.

Laserpitin (LAS), isolaserpitin (ILA) and (8S,9R)-8-angeloxy-8,9-dihyrooroselol (DHO) were identified as bitter constituents from Angelica (A.) keiskei KOIDZUMI, which is becoming popular as vegetables and dietary supplements in Japan. A yellow juice from the aerial parts of 140 samples of A. keiskei could be grouped into 4 types (Type A〜D) according to the HPLC profiles with the peaks of bitter constituents. Type A was from the Hachijyo island strain, which detected LAS and ISO, and Type B was from the Oshima island strain, which only detected DHO. The LAS, ILA and DHO contents were 0.066〜0.092％, 0.040〜0.068％ and 0.064〜0.279％, respectively.

We determined the differences in carbon, nitrogen and oxygen isotopic compositions (δ¹³C, δ¹⁵N, and δ¹⁸O) between short grain rice samples from 9 locations (Hokkaido, Yamagata, Niigata, Nagano, Ibaraki, Tokyo, Mie, Hiroshima and Okinawa) in Japan to discriminate their geographical origin. Oxygen isotopic composition of brown rice is well correlation with that of ambient water (R²＝0.69). The δ¹⁸O values of rice samples are correlated with an increase in the latitude (R²＝0.82). The δ¹³C values of upland rice are higher than those of paddy rice, depending on the water stress. These results suggest that δ¹⁸O and δ¹³C values of Japanese rice samples are reflected the difference in growth condition. The δ¹⁵N values of rice depend on the soil nutrition, where the rice is cultivated. Organic rice is characterized by higher nitrogen isotopic compositions (＋3.0 to ＋8.9‰) than conventional rice (−0.4 to ＋3.2‰). Thus, δ¹⁵N would become a strong tool to discriminate between organic and conventional rice samples. Thus, the δ¹⁸O and δ¹³C values will be potentially useful for the discrimination of organic rice, and the geographical origin of rice within Japan.

Carbon and oxygen stable isotope ratios of plants reflect their growing environment. In this study, we examined the carbon and oxygen stable isotope ratios of apple juice from Japan, China, Australia, and South Africa, in order to confirm them as a potential index for verifying the geographical origin of apple juice. Apple juice from Japan is characterized by lower carbon and oxygen isotope ratios {δ¹³C : −26.7±1.0‰ (mean±sd), δ¹⁸O : 20.3±1.1‰} than those from other countries. Especially, the apple juice from China has a higher δ¹³C value (−24.6±0.5‰), and those from Australia and South Africa have higher δ¹⁸O values (27.8±1.6‰ and 29.4±2.3‰, respectively). These results suggest that carbon and oxygen stable isotope ratios can be used as a potential index for discrimination between domestic and imported apple juice.

Important taste compounds in vegetables are organic acids (sour taste), free sugars (sweetness) and some amino acids (umami-taste). The author tried to analyze these important taste compounds (oxalic acid, citric acid, malic acid, aspartic acid, glutamic acid, fructose, glucose, sucrose) in various vegetables simultaneously using capillary electrophoresis. The electrolyte used was 20 mM 2,6-pyridine dicarboxylic acid and 0.5 mM cetyltrimethylammonium bromide at pH 12.1. The capillary tube was rinsed between every analysis for 1.5 min with water, 3 min with methanol, 4 min with sodium hydroxide (1 M) and 5 min with a running electrolyte, respectively, for the purpose of improving the stability and peak shape of the pherogram. The signal was indirectly detected at 270 nm. The taste compounds of various vegetables purchased in Japanese market were successfully analyzed using this method. These comparative data, based on the same analytical methods are very rare in the literature, and our data clearly indicate the characteristics of each vegetable.

We determined the stable carbon, nitrogen and oxygen isotopic compositions (δ¹³C, δ¹⁵N and δ¹⁸O) of processed eels (Shirayaki and Kabayaki) in order to demonstrate the effect of the processing process on their stable isotopic compositions. There were no significant change in the δ¹³C, δ¹⁵N and δ¹⁸O values between eels and processed eels. We examined processed eels from Japan (n＝84), China (n＝30), and Taiwan (n＝10) to trace their geographical origin. The Japanese eels had lower δ¹⁸O values (＋8.9±1.4‰) than the Chinese (＋12.9±1.8‰) and Taiwanese eels (＋13.1±2.2‰). A principal component analysis using carbon, nitrogen and oxygen isotopic compositions of Japanese, Taiwanese and Chinese processed eels showed a difference tendency. Thus, stable carbon, nitrogen and oxygen isotope analyses would probably be a useful tool for tracing the geographical origin of processed eels.