A simultaneous rapid test method for pesticide residues in livestock products by LC-MS/MS was developed and validation tests were conducted. For the extraction method, the QuEChERS method was applied, and samples were extracted with acetonitrile, followed by salting out and dehydration. For purification, C18 and PSA columns were used instead of GPC. Based on the validity guidelines of the Ministry of Health, Labor and Welfare, 48 pesticide residues were evaluated at 0.01 μg/g and 0.1 μg/g for cattle muscle, swine muscle and milk. The trueness of the method for 43 pesticides in all three types of food samples was 70-120%, with satisfactory repeatability and within-laboratory reproducibility. Thus, this method demonstrated the usefulness of simultaneous rapid testing for pesticide residues in livestock products.
A simultaneous analytical method was developed for the determination of deoxynivalenol (DON) and ochratoxin A (OTA) in wheat. Analytes were purified from wheat extracts by a multifunctional column and quantified by LC-MS/MS. An interlaboratory study was conducted to validate the analytical method using three spiked and two contaminated wheat samples. The ranges of mean recoveries of DON and OTA were 88-89% and 91-96%, respectively. The RSDs for repeatability and reproducibility were all ≤15%. Furthermore, the levels of DON and OTA in artificially contaminated wheat samples were analyzed by the simultaneous analytical method. The results showed that the developed analytical method yielded analytical values equivalent to those obtained when analyzing DON and OTA individually by existing analytical methods. Based on these results, the simultaneous analytical method for DON and OTA in wheat developed in this study is considered suitable as an alternative to the methods for analyzing each mycotoxin individually.
Molecular typing of enteroviruses (EVs) is essential for surveillance of hand, foot, and mouth disease (HFMD). Conventional reverse-transcription polymerase chain reaction (RT-PCR) targeting the VP4–VP2 region can be insufficiently sensitive, reducing the detectability of Enterovirus A (EV-A). We developed a single-round RT-PCR assay using a modified reverse primer design (C3R) for rapid EV detection and genotyping. Sensitivity was evaluated using EV-A71 and poliovirus type 1 reference strains, across 60 EV-positive clinical specimens. The C3R-based assay showed ~1000-fold higher sensitivity for EV-A71 than for conventional assays (limit of detection: 6.6 copies/reaction). The assay detected 98.3% (59/60) of clinical specimens in a single-round format, whereas the conventional assay detected only 45.0% (27/60) and showed a marked decline in detection at higher Ct values. The C3R-based assay maintained complete detection for clinical specimens with Ct values below 40. The majority of the amplified products yielded high-quality sequences suitable for genotyping. This C3R-based RT-PCR overcomes sensitivity limitations of existing protocols and provides reliable genotyping from low-viral-load specimens, supporting its use in routine diagnostics and large-scale HFMD surveillance.
Ethylene oxide (ETO) is a highly reactive sterilizing agent classified as a carcinogen, and its use in foods is prohibited in many countries. Regulatory monitoring currently relies primarily on the detection of ETO and/or 2-chloroethanol (2CE), a chloride-derived reaction product. However, ETO is volatile and difficult to detect directly in commercial foods, and 2CE may occur naturally or arise from non-food-related sources, raising scientific concerns regarding its specificity as a fumigation marker. In this study, we applied a food-processing metabolomics approach to comprehensively investigate the chemical alterations induced by ETO fumigation of cumin (Cuminum cyminum L.) seeds using gas chromatography-mass spectrometry. Fumigated and non-fumigated samples were clearly discriminated by principal component analysis of the data containing fumigation-specific metabolites. Here, we report that high-resolution gas chromatography quadrupole time-of-flight mass spectrometric analysis identified a characteristic compound with a molecular formula consistent with a trimethylsilylated derivative of C6H10O6 containing a single ETO incorporation. Fragmentation patterns and accurate mass measurements suggested that this compound was an ETO adduct of malic acid. In vitro fumigation experiments using authentic malic acid reproduced the same chromatographic retention time and mass spectrum pattern, confirming its formation. A targeted gas chromatography-tandem mass spectrometric analysis further demonstrated the presence of the malic acid-ETO adduct (m/z 233 → 73) in fumigated sesame and soybean, indicating its potential broader presence across ETO fumigated plant-derived matrices. Collectively, these findings provide the first direct evidence that ETO fumigation induces the formation of a nutrient-derived adduct in foods. The identified ETO-modified nutrient represents a promising chemical marker for verifying ETO fumigation beyond conventional 2CE-based approaches, thereby contributing to improved analytical specificity and regulatory reliability in food safety control.
To accelerate the development of antiviral drugs, we generated human iPS cell-derived myeloid cell lines, which were susceptible to SARS-CoV-2 infection. Viral isolation using clinical specimens showed that some novel cell lines performed better than VeroE6/TMPRSS2. In addition, viral genome mutations that occurred during viral passaging did not happen in VeroE6/TMPRSS2 but accumulated in these cell lines, and those mutations were reported in clinical specimens. Furthermore, the antiviral efficacy of remdesivir differed among the cell types. These findings suggest that the novel myeloid cell lines may serve as a useful platform for virus isolation and the evaluation of antiviral responses.
