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ABSTRACT: A rapid multiplex real-time PCR assay was developed to achieve highly specific, simultaneous detection of two kinds of mushrooms, Omphalotus guepiniformis and Lentinula edodes. Primers and TaqMan minor groove binder probes were designed according to the internal transcribed spacers 1-5.8S region of rDNA and evaluated by the specificity for fruiting bodies of 17 O. guepiniformis, 16 L. edodes and samples from 57 other species. DNA extracts of all the target species had positive signals with no cross-reaction, the limit of detection being 0.00025 ng of DNA. Threshold cycle (Ct) values for raw and processed fruiting bodies and for fruiting bodies (1% (w/w)) mixed with foodstuffs or artificial gastric juice contents ranged from 17.16 to 26.60 for both examined species. This new assay proved specific to the target species, highly sensitive, and applicable to processed food samples and gastric juice contents, making it useful for rapidly identifying O. guepiniformis and L. edodes.
Bioscience Biotechnology and Biochemistry 07/2012; 76(7):1343-9. · 1.28 Impact Factor
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ABSTRACT: A rapid and sensitive TaqMan real-time PCR assay for the detection of Pacific cod (Gadus macrocephalus) and capelin (Mallotus villosus) roes in Alaska pollack (Theragra chalcogramma) roe product was developed. The primers and the TaqMan MGB (minor groove binder) probes were designed based on the gene encoding cytochrome b for the specific detection of Alaska pollack, Pacific cod and capelin. This real-time PCR assay had the detection limit of 0.002 ng/microL mitochondrial DNA and showed no cross-reaction with 48 other species. The calculated r(2) values of the standard curves for the three species were 1.000. This assay was applied for the detection of Pacific cod and capelin roes in mixture samples: Pacific cod or capelin roes were added to Alaska pollack roes at 0.1, 1 and 10%. The threshold cycle values were obtained from both of the mixture samples at 0.1%. Practical applicability of this assay was examined with 64 samples of Alaska pollack roe products. In all cases, the species detected from the samples corresponded with species described on the food label.
Journal of the Food Hygienic Society of Japan (Shokuhin Eiseigaku Zasshi) 01/2010; 51(3):110-4. · 0.43 Impact Factor
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ABSTRACT: A method for the determination of nonvolatile amines (putrescine, cadaverine, histamine, tyramine and spermidine) in foods by solid-phase extraction and excimer-forming derivatization was investigated. Nonvolatile amines in a solid sample were extracted with 3% trichloroacetic acid, and the amines in a liquid sample were extracted with water. The extract was applied to polymer-based strong cation exchange mini-column, which was then rinsed with phosphate buffer of pH 6.8 and water. Nonvolatile amines were eluted with 100 mmol/L potassium carbonate solution. The solution was mixed with 6 mmol/L 1-pyrenebutylyl chloride solution and derivatized. Derivatives of nonvolatile amines were analyzed by LC-FLD, and the identity of the amines was confirmed by LC-MS/MS without derivatization. The limit of detection (S/N> or =3) of nonvolatile amines in all samples was 0.04 microg/g, and the limit of quantitation (S/N> or =10) was 0.1 microg/g. Recoveries of nonvolatile amines from fish tissues, miso, shoyu and red wine were in the range of 80.4-111%.
Journal of the Food Hygienic Society of Japan (Shokuhin Eiseigaku Zasshi) 01/2010; 51(3):115-21. · 0.43 Impact Factor
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ABSTRACT: A simple method was developed for the analysis of hyoscyamine and scopolamine in human serum and urine using liquid chromatography with tandem mass spectrometry (LC/MS/MS). Hyoscyamine and scopolamine in serum and urine were cleaned up with an Oasis HLB cartridge and a PSA cartridge. The LC separation was carried out on an ODS column, using linear gradient elution with 5 mmol/L IPCC-MS3-methanol as the mobile phase. The mass spectral acquisition was done in the positive ion mode by applying selected reaction monitoring (SRM). The recoveries of hyoscyamine and scopolamine were 86.0-105% from human serum and urine fortified at 0.2 ng/mL and 10 ng/mL. The detection limits of hyoscyamine and scopolamine were 0.02 ng/mL. Four serum and three urine samples of humans poisoned by eating Datura innoxia Mill. were analyzed by this method. Hyoscyamine and scopolamine were detected at the levels of 0.45-3.5 ng/mL in all serum samples and 170-670 ng/mL in all urine samples.
Journal of the Food Hygienic Society of Japan (Shokuhin Eiseigaku Zasshi) 09/2008; 49(4):266-71. · 0.43 Impact Factor
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ABSTRACT: A simple and rapid method was developed for the analysis of tetrodotoxin in puffer-fish tissues, and in serum and urine of humans poisoned after consuming puffer-fish, by means of high-performance liquid chromatography with tandem mass spectrometry (LC/MS/MS). Tetrodotoxin was extracted with 2% acetic acid. The extracted solution from puffer-fish tissues was diluted with water, and the extracted solution from human serum and urine was cleaned up by LC/MS/MS with a methacrylate-styrenedivinylbenzene cartridge. The LC separation was performed on a C18 column (50 mm x 2.1 mm i.d.) using 10 mmol/L IPCC-MS7-methanol (65 : 35) as the mobile phase at a flow rate of 0.2 mL/min. The mass spectral acquisition was done in the positive ion mode by applying selected reaction monitoring (SRM). The recoveries of tetrodotoxin were 79-90% from puffer-fish tissues fortified at 0.1 microg/g and 1 microg/g, and 93-101% from human serum and urine fortified at 0.5 ng/mL and 5 ng/mL. The detection limits of tetrodotoxin were 0.01 microg/g in puffer-fish tissues and 0.1 ng/mL in human serum and urine. Thirty samples of puffer-fish from wholesale markets, and 7 serum and 5 urine samples of humans poisoned after consuming puffer-fish were analyzed by this method. Tetrodotoxin was detected in all puffer-fish tissues, and all serum and urine samples at the levels of 0.04-140 microg/g, 0.9-1.8 ng/mL and 15-150 ng/mL, respectively.
Journal of the Food Hygienic Society of Japan (Shokuhin Eiseigaku Zasshi) 05/2006; 47(2):46-50. · 0.43 Impact Factor
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Kouichi Akaki
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ABSTRACT: A simple and rapid method was developed for the analysis of mepiquat chloride in grape, wine and juice by high-performance liquid chromatography with electrospray tandem mass spectrometry (LC/MS/MS). Mepiquat chloride was extracted with water-methanol (1:1). Extracted solution was adjusted to pH 10 with ammonia solution. A part of the extracted solution was cleaned up on a styrenedivinylbenzene (SDVB) cartridge for LC/MS/MS. The LC separation was performed on a C18 column (50 mm x 2 mm i.d.) using 0.1% IPCC-MS7-methanol (60:40) as the mobile phase at a flow rate of 0.2 mL/min. The mass spectral acquisition was done in the positive ion mode by applying selected reaction monitoring (SRM). The recoveries of mepiquat chloride from fresh grape, wine and juice fortified at 5 microg/kg and 50 microg/kg were 84.5-96.1%. The lower limit of quantification was 1 microg/kg. Fourteen fresh grape samples, 14 wines (white), 36 wines (red) and 11 juice samples were analyzed by this method. Mepiquat chloride was detected in 5 fresh grape samples, 3 wines (white) and 1 wine (red) at the level of 12.8-199 microg/kg, 5.7-47.7 microg/kg and 24.1 microg/kg, respectively.
Journal of the Food Hygienic Society of Japan (Shokuhin Eiseigaku Zasshi) 09/2004; 45(4):197-200. · 0.43 Impact Factor
