Flow cytometric evaluation of nanoparticles using side-scattered light and reactive oxygen species-mediated fluorescence-correlation with genotoxicity

We recently clarified that the side-scatter(ed) light (SSC) of flow cytometry (FCM) could be used as a guide to measure the uptake potential of nanoparticles [ Suzuki et al. Environ. Sci. Technol. 2007 , 41 , 3018 - 3024 ]. In this paper, the method was improved so as to be able to determine simultaneously the uptake potential of nanoparticles and the production of reactive oxygen species (ROS), and correlations with genotoxicity were evaluated. In the FCM analysis, SSC and fluorescence of 6-carboxy-2,7'-diclorodihydrofluorescein diacetate, di(acetoxy ester) based on ROS production were concurrently detected after treatments with ZnO, CuO, Fe(3)O(4), TiO(2), and Ag nanoparticles. The ZnO and CuO nanoparticles caused high ROS production, which was more significant in the cells with higher SSC intensity. The increase of SSC intensity was more significant for TiO(2) than ZnO and CuO, whereas ROS production was higher for ZnO and CuO than TiO(2), suggesting that the extent of ROS production based on the uptake of nanoparticles differed with each kind of nanoparticle. ROS production was correlated with generation of the phosphorylated histone H2AX (γ-H2AX), a marker of DNA damage, and an antioxidant, n-acetylcysteine, could partially suppress the γ-H2AX. This method makes it possible to predict not only uptake potential but also genotoxicity.     

鉴别红葡萄酒中添加人工色素或天然色素的简易方法

目的:鉴别红葡萄酒中是否添加合成人工色素以及天然色素(以葡萄皮红色素为例)。方法:对葡萄酒样品、人工色素模拟酒和天然色素模拟酒样品在碱性测试试纸上显示的颜色进行表征并对比。结果:当测试试纸显示的颜色为红色时,该红葡萄酒样品中含有人工色素;当测试试纸显示的颜色为蓝色,并在室温下暴露在空气中5min后不变色,则该红葡萄酒样品中添加天然色素,在5min内若由蓝色变为黄色,则说明该红葡萄酒样品中未添加天然色素,为全葡萄汁酿制而成。结论:该法简易快速,适合初步定性鉴别红葡萄酒中是否掺杂色素使用。     

Polarised light stress analysis and laser scatter imaging for non-contact inspection of heat seals in food trays

This paper introduces novel non-contact methods for detecting faults in heat seals of food packages. Two alternative imaging technologies are investigated; laser scatter imaging and polarised light stress images. After segmenting the seal area from the rest of the respective image, a classifier is trained to detect faults in different regions of the seal area using features extracted from the pixels in the respective region. A very large set of candidate features, based on statistical information relating to the colour and texture of each region, is first extracted. Then an adaptive boosting algorithm (AdaBoost) is used to automatically select the best features for discriminating faults from non-faults. With this approach, different features can be selected and optimised for the different imaging methods. In experiments we compare the performance of classifiers trained using features extracted from laser scatter images only, polarised light stress images only, and a combination of both image types. The results show that the polarised light and laser scatter classifiers achieved accuracies of 96% and 90%, respectively, while the combination of both sensors achieved an accuracy of 95%. These figures suggest that both systems have potential for commercial development.     

Simple and selective analysis of different antibiotics in milk using molecularly imprinted polymers: a review

ABSTRACT

The frequent, sometimes illegal, use of antibiotics for therapeutic and prophylactic purposes in dairy cattle management may cause residues in milk. Because of problems concerning bacterial resistance or allergies in consumers, monitoring of residues is required. In spite of the huge development of analytical instrumentation during recent years, sample preparation is still a bottleneck of the analytical process. In this regard, efforts have been directed towards improving selectivity during extraction and clean-up of samples. Compared to traditional sorbents, molecularly imprinted polymers (MIPs) are excellent materials to provide selectivity for sample pretreatment. This review presents the application of MIPs as effective sorbents for separation, clean-up, pre-concentration and analysis of different antibiotics in milk.     

Non-destructive analysis of anthocyanins in cherries by means of Lambert-Beer and multivariate regression based on spectroscopy and scatter correction using time-resolved analysis

In high-value sweet cherry (Prunus avium), the red coloration - determined by the anthocyanins content - is correlated with the fruit ripeness stage and market value. Non-destructive spectroscopy has been introduced in practice and may be utilized as a tool to assess the fruit pigments in the supply chain processes. From the fruit spectrum in the visible (Vis) wavelength range, the pigment contents are analyzed separately at their specific absorbance wavelengths.A drawback of the method is the need for re-calibration due to varying optical properties of the fruit tissue. In order to correct for the scattering differences, most often the spectral intensity in the visible spectrum is normalized by wavelengths in the near infrared (NIR) range, or pre-processing methods are applied in multivariate calibrations.In the present study, the influence of the fruit scattering properties on the Vis/NIR fruit spectrum were corrected by the effective pathlength in the fruit tissue obtained from time-resolved readings of the distribution of time-of-flight (DTOF). Pigment analysis was carried out according to Lambert-Beer law, considering fruit spectral intensities, effective pathlength, and refractive index. Results were compared to commonly applied linear color and multivariate partial least squares (PLS) regression analysis. The approaches were validated on fruits at different ripeness stages, providing variation in the scattering coefficient and refractive index exceeding the calibration sample set.In the validation, the measuring uncertainty of non-destructively analyzing fruits with Vis/NIR spectra by means of PLS or Lambert-Beer in comparison with combined application of Vis/NIR spectroscopy and DTOF measurements showed a dramatic bias reduction as well as enhanced coefficients of determination when using both, the spectral intensities and apparent information on the scattering influence by means of DTOF readings. Corrections for the refractive index did not render improved results.     

PCR-RFLP using a SNP on the mitochondrial Lsu-rDNA as an easy method to differentiate Tuber melanosporum (Perigord truffle) and other truffle species in cans

Canned truffle products labeled Tuber melanosporum, the famous Perigord truffle, may contain other less tasty and cheaper truffle species. To protect consumers from fraud, a PCR DNA-based method was used to unequivocally identify the nature of the product. Several rapid and simple cell lysis procedures, used in conjunction with a commercially available DNA purification kit, were evaluated for their effectiveness in recovering DNA from canned truffle. In parallel, a marker for T. melanosporum was tested on the mitochondrial rDNA. These two techniques were then combined to differentiate T. melanosporum from other truffle species like T. aestivum, T. brumale or T. indicum up to the legal threshold in canned products. These findings not only allow a comparison of the effectiveness of the different DNA extraction methods but also provide a preliminary indication of the specificity and sensitivity of the detection with the mitochondrial marker that might be attainable for truffle species in a quantitative PCR-based analysis method.     

Development of easy and efficient methods for quantitative analysis of ethyl carbamate using GC-MS in various fermented foods

To develop more efficient quantitative analysis of ethylcarbamate in fermented food, the major disturbance was identified as diethyl succinate on GC-MS through matching with mass fragmentation pattern on mass library and comparison of retention time. And the clean-up method was developed based on the difference of partition coefficient between ethyl carbamate and diethyl succinate. Finally, a washing step was added with 1% dichloromethane in n-hexane on diatomaceous earth cartridge, and successfully applied to remove diethyl succinate from the matrix. The developed method showed acceptable recovery rate (69.5–88.4%) and limit of quantitation (1.0–1.2 ng/mL) on several fermented foods.     

Determination of heavy metals in honey by potentiometric stripping analysis and using a continuous flow methodology

A methodology for the determination of Zn(II), Cd(II) and Pb(II) directly in dissolved honey samples by potentiometric stripping analysis with a flow cell is proposed. Heavy metals in honey are of interest not only for quality control, but can be used also as an environmental indicator. In this work honey samples were collected in different places of Burgos (Spain). Lead (II) and cadmium (II) can be directly determined. It is necessary to avoid the Zn–Cu interference for zinc (II) determination by adding Ga(III) in the adequate concentration, which has been calculated by using an experimental design methodology. The results were compared with inductively coupled mass plasma spectrometry as reference method.     

LC-MS method development to evaluate major triterpenes in skins and cuticular waxes of grape berries

Predominant triterpenes (free oleanolic acid, β-sitosterol and β-sitosterol-3-O-β- d -glucoside) in skin and cuticular wax of grape berries have been analysed by liquid chromatography–mass spectrometry (LC-APCI-MS). The validity of the developed method was established by determining linearity, recovery, precision, accuracy, limit of detection and quantification. Detection limits were in the range of 0.12–0.25 mg L⁻¹, and linearity values ranged up to 105 mg L⁻¹. The repeatability of the method was good. High variability was found among the measured grape varieties based on triterpene content, quantitites of oleanolic acid of cuticular wax ranged from 31.53 mg kg⁻¹ to 162.01 mg kg⁻¹ of the twelve analysed samples. The highest sitosterol content was measured in the sample 'Othello' (73.12 mg kg⁻¹), while maximum sitosterol glucoside content was also found in 'Othello' variety (13.68 mg kg⁻¹). The results showed that the study on triterpenes could be an informative tool to characterise grape varieties.     

Improvement of the ammonia measurement using a clarifying reagent and application to evaluate heat damage in commercial milk samples

The enzymatic Boehringer procedure for ammonia measurement in milk was improved to be rapid and simple using a clarifying reagent without any pretreatment of the sample. A good correlation (r=0.99, n=36) existed between the proposed enzymatic method (EM) and the specific electrode method (SEM) when applied on raw and commercial milk samples. The repeatability of the EM applied to UHT milk was satisfactory with a relative standard deviation of 5.4% (n=26). The EM was used to evaluate heat damage in commercial half-skimmed pasteurised, direct or indirect or unlabelled UHT and in-bottle sterilised milk. The EM was well correlated with absorbance at 340 nm (A₃₄₀) of transparent modified milk (r=0.97, n=42), with lactulose determined by HPLC and by capillary electrophoresis (r=0.95, n=17 and r=0.95, n=26, respectively). All parameters differentiated significantly (P<0.001) between pasteurised, UHT- and in-bottle sterilised milk, and also direct-UHT and indirect-UHT samples. More intensive heat treatment produced higher contents of ammonia, lactulose and A₃₄₀ values.     

Evaporative light scattering: a novel detection method for the quantitative analysis of humic-like substances in aerosols

The chemical composition of organic atmospheric aerosols is only poorly understood. Although a significant fraction of organic aerosols consists of humic-like substances (HULIS), only little is known about this class of compound, and accurate quantification remains difficult, partly due to the lack of appropriate standards. Here, evaporative light-scattering detection (ELSD) was applied for the first time to quantify water-soluble HULIS in aerosol particles smaller than 1 microm. This detection method was shown to be suitable for the quantification of compounds with unknown structures and lacking appropriate quantification standards. As compared to organic carbon determination of isolated HULIS, no organic carbon/organic mass (OC/OM) conversion factor needs to be applied with ELSD and therefore eliminates this significant uncertainty factor of the OC/OM method, which is frequently used to quantify HULIS. Solid-phase extraction and size-exclusion chromatography were applied to separate inorganic ions and low molecular weight compounds from HULIS before ELSD quantification. The ELSD itself provides an additional separation step where low volatility HULIS are separated from high volatility, small compounds. Electrospray ionization mass spectrometry was used to identify the molecular weight range of the compounds quantified with ELSD. The most intensive peaks were in the range of m/z 200-500, with some masses upto m/z800. We showed that UV detection using fulvic acid as surrogate quantification standard underestimates the HULIS concentration by a factor of 1.1 to 2.5, which is in agreement with earlier studies. During a 6 week winter 2005-2006 campaign at a suburban site near Zurich, Switzerland, an average of 1.1 microg/m(3) HULIS was found, which is about4-6% of the total particle mass smaller than 1 microm (PM1) and 10-35% of the organic matter in PM1.     

Determination of sugarcane deterioration at the factory: Development of a rapid,easy and inexpensive enzymatic method to measure mannitol

Mannitol, formed mainly by Leuconostoc mesenteroides bacteria, is a very sensitive indicator of sugarcane deterioration that directly affects processing and can predict problems from dextran and levan polysaccharides. An enzymatic method has been developed to measure mannitol in juice pressed from consignments of sugarcane delivered to the factory. This screening tool will allow factory staff to rapidly know which consignments of cane will affect processing negatively or reject consignments that will cause unacceptable processing problems. Mannitol is directly measured on a spectrophotometer using mannitol dehydrogenase as the enzyme catalyst. The stability of the reagents, limited cane juice preparation and linearity are described. This method is accurate, comparing favorably with an ion chromatography method, and can be easily performed using existing equipment in sugarcane factories. The coefficient of variation (CV) for cane juices ranged from 1.73% to 5.13% with the highest CVs occurring for low mannitol concentrations in undeteriorated cane. Mannitol can be measured after ∼7 min at room temperature and within 4 min if a 40 °C waterbath is used. The method is highly specific for mannitol and was not affected by the presence of sucrose, glucose, fructose, or dextran. The current cost is only ∼60 US cents per analysis. Further studies on the viability of the method at the factory, and as a screening tool for breeding programs for cane freeze tolerance, are being undertaken.     

Monitoring of beef aging using a two-line flow injection analysis biosensor consisting of putrescine and xanthine electrodes

A two-line flow injection analysis (FIA) biosensor was developed which can simultaneously detect bacterial spoilage and the progress of aging. This FIA biosensor was composed of a putrescine oxidase immobilized electrode and a xanthine oxidase immobilized electrode as detectors. The putrescine electrode measures putrescine and cadaverine which are produced by bacteria, and the xanthine electrode measures hypoxanthine and xanthine which accumulate in meat with aging.

The analytical conditions for this system were set as follows; flow rate, 1 ml/ min; water bath temperature, 30 °C; flow buffer, 0.1 M phosphate buffer (pH 7.0); injection volume for putrescine electrode, 200 μl; injection volume for xanthine electrode, 40 μl; and measurement cycle, 2 min.

The linear relationship for standard solution was between 20 and 800 nmol/ml in the putrescine electrode and between 0.1 and 3.0 μmol/ml in the xanthine electrode. The coefficients of variation in standard solutions were 2.14% with the putrescine electrode and 2.83% with the xanthine electrode. The coefficients of variation values in the specimen solution were 3.22% and 3.76%, respectively.

This two-line FIA biosensor was applied to the vacuum-packed beef stored at 0, 5 and 10 °C. The progress of aging could be monitored at all temperatures, and the bacterial spoilage could be detected before the appearance of putrid odor at 5 and 10 °C. However, at 0 °C the putrid odor did not appear during storage, and neither putrescine nor cadaverine was detected. Thus, this FIA biosensor was confirmed to be useful for the quality control of beef aging at 5 and 10 °C, but not at 0 °C.     

Development of an objective and uniform scoring method to evaluate the quality of rearing in Dutch dairy herds

Young stock rearing is an essential part of dairy management, and it is important that the quality of rearing can be monitored and altered if necessary. In this study, a young stock rearing quality system (KalfOK) was developed with the aim to provide an objective and standardized means to evaluate and monitor the quality of young stock rearing in Dutch dairy herds. In the project, 201 dairy farmers participated. Twelve key indicators were defined that were related to calving and successful rearing, antimicrobial use, and herd health. For each of the key indicators, the value was calculated per herd and quarter of the year between January 2014 and April 2017. Benchmark values were determined to compare herd-specific results and for selection of threshold values. Each of the key indicators was graded when the value scored above the threshold. Combining the grades resulted in the herd-specific KalfOK score, which could vary between 0 and 100. Subsequently, 100 of the participating dairy herds were visited and the quality of young stock rearing was scored by a trained veterinarian. Using principal component analysis, the results of the herd health checks were combined into a factor score that represented the observed quality of young stock rearing during the visit. The amount of variance in observed quality of rearing during the herd health check that was explained by the key indicators in KalfOK was evaluated. Additionally, the validity of KalfOK to distinguish herds with an excellent or insufficient quality of young stock rearing was assessed by comparing the top and bottom 10% herds in the herd health check with the proportion of herds with a KalfOK score above or below a prespecified cutoff value. The results of the linear regression model showed that the key indicators included in KalfOK accounted for 56% of the variation in the score of the herd visits by a veterinarian. The moving average of the annual KalfOK score, which was the sum of the grades of all key indicators, was 77 points (25th percentile = 71, 75th percentile = 85 points). The combination of the sensitivity (88%, 95% confidence interval = 47–100%) and specificity (67%, 95% confidence interval = 54–78%) of KalfOK to correctly classify herds with an excellent quality of young stock rearing was highest when a cutoff value of 80 points was applied. Detection of dairy herds with an insufficient quality of young stock rearing was best at a cutoff value of 70 points (sensitivity 86%, 95% confidence interval = 42–100%; specificity 77%, 95% confidence interval = 66–86%). The KalfOK score that was based on routinely collected herd data provided an indication of the quality of young stock rearing in individual Dutch dairy farms. The KalfOK score illustrates how such data can be transferred into herd-specific information in support of animal health and welfare. Given the increasing availability of automatically assembled data, the development of similar monitoring tools seems a feasible option to enhance herd-specific management.     

Roughness analysis of the skin as a secondary evaluation criterion in addition to visual scoring is sufficient to evaluate ethnic differences in wrinkles

Visual scoring has been used to evaluate ethnic differences in skin wrinkling, but it is not sufficient to fully evaluate those differences in wrinkles. We examined whether both the roughness analysis of the skin and visual scoring are sufficient to characterize ethnic differences in wrinkles in Japanese, Chinese and German women. One hundred and five Japanese, 96 Chinese and 90 German age-matched women participated in this study. The severity of their wrinkles in the skin at two sites at the periphery of the eye was evaluated by visual scoring using a photoscale and by roughness values obtained from three-dimensional analysis of skin replicas. Slight but significant differences were scarcely observed between Japanese and Chinese women as well as between Japanese and German women at the same age group using the visual scoring method. However, significant and clearer differences among those ethnic groups were observed using the roughness analysis of skin replicas. Below the eye, significant differences among those ethnic groups were observed using both visual wrinkle scoring and roughness analysis. However, the extent of increased roughness values with age was relatively small compared with the increased wrinkle scores. These results show that roughness analysis is more sensitive than the visual scoring method when comparing ethnic differences in wrinkles. We conclude that roughness analysis of the skin is an important secondary evaluation criterion to visual scoring necessary to evaluate ethnic differences of wrinkles.     

流动注射化学发光法测定人体血清和金银花中的绿原酸

基于绿原酸对鲁米诺-溶解氧体系化学发光反应的抑制作用,结合流动注射技术,建立了绿原酸化学发光分析法,用于人体血清、金银花水提液中绿原酸测定,实现了样品中超痕量的分析．当绿原酸的质量浓度为1．0ng·mL^-1～100．0ng·mL^-1时,化学发光强度的减小值与其浓度呈线性关系．测定的检出限为0．33ng·mL^-1,线性相关系数r^2=0．9978,加标回收率为95．8％～105．5％．在流速为2．0mL·min^-1时,该方法完成一次绿原酸的测定只需0．5min,相对标准偏差为5．0％,并且选择性和灵敏度较高．     

Bioelectrical impedance analysis: opportunities and prospects of using this method in practical dietology

In clinical practice spectral bioelectrical impedance analysis is a sensitive enough method in an estimation of changes of structure of a body composition and distribution of a liquid on sectors at various modes of a feed in norm and pathology. The analysis of parameters allows to direct the expert on creation of the individual circuits of a feed, is especial at the patients who are taking place on long treatment.