Validation of the performance of a GMO multiplex screening assay based on microarray detection

A new screening method for the detection and identification of GMO, based on the use of multiplex PCR followed by microarray, has been developed and is presented. The technology is based on the identification of quite ubiquitous GMO genetic target elements first amplified by PCR, followed by direct hybridisation of the amplicons on a predefined microarray (DualChip^® GMO, Eppendorf, Germany). The validation was performed within the framework of a European project (Co-Extra, contract no 007158) and in collaboration with 12 laboratories specialised in GMO detection. The present study reports the strategy and the results of an ISO complying validation of the method carried out through an inter-laboratory study. Sets of blind samples were provided consisting of DNA reference materials covering all the elements detectable by specific probes present on the array. The GMO concentrations varied from 1% down to 0.045%. In addition, a mixture of two GMO events (0.1% RRS diluted in 100% TOPAS19/2) was incorporated in the study to test the robustness of the assay in extreme conditions. Data were processed according to ISO 5725 standard. The method was evaluated with predefined performance criteria with respect to the EC CRL method acceptance criteria. The overall method performance met the acceptance criteria; in particular, the results showed that the method is suitable for the detection of the different target elements at 0.1% concentration of GMO with a 95% accuracy rate. This collaborative trial showed that the method can be considered as fit for the purpose of screening with respect to its intra- and inter-laboratory accuracy. The results demonstrated the validity of combining multiplex PCR with array detection as provided by the DualChip^® GMO (Eppendorf, Germany) for the screening of GMO. The results showed that the technology is robust, practical and suitable as a screening tool.     

Ultrasound‐based,in‐line monitoring of anaerobe yeast fermentation: model,sensor design and process application

In order to implement process analytical technology in beer manufacturing, an ultrasound‐based in‐line sensor was developed which is capable to determine sound velocity and density via the multiple reflection method. Based on a systematic study of the ternary system water–maltose–ethanol, two models were established to estimate the critical process parameters: sugar and ethanol mass fraction. The sound velocity‐based model showed unreasonable high errors although temperature variations and deviations due to dissolved CO₂ were corrected. In contrast, the sound velocity–density–temperature model provided an average root mean square error of 0.53%g/g sugar and 0.26%g/g ethanol content for the main fermentation. Method, sensor and model showed the capability to capture the process signature which may be related to product and process quality.     

Expression and characterization of three important panallergens from hazelnut

Several hazelnut allergens with different clinical relevance and crossreactive properties have been identified and characterized so far. The aim of this study was to develop protocols for producing relatively large amounts of three recombinant hazelnut allergens Cor a 1.04, Cor a 2, and Cor a 8 in a folded and immunologically active form. The availability of well-characterized, pure recombinant allergens will improve diagnostic in vitro tests for food allergy, by allowing a highly sensitive component resolved diagnosis. Depending on the individual hazelnut allergen, protocols for heterologous production - either as fusion or nonfusion protein - were developed to obtain homogenous protein batches. The resulting proteins were purified by a two-step FPLC method and their IgE antibody reactivity was verified. Identity was verified by N-terminal sequencing and MALDI-TOF-MS analysis. Their secondary and tertiary structure was controlled by circular dichroism (CD)-spectroscopy and NMR analysis. Decisions on the strategies for expression and purification of allergens on a large scale were made on a case by case basis: Preparation of rCor a 1.04 and rCor a 2 as fusion proteins in E. coli from inclusion bodies resulted in approximately 10 mg pure protein per liter whereas rCor a 8 expression in yeast as nonfusion protein yielded 30 mg/L.     

Consumer preferences and willingness to pay for value-added chicken product attributes

A growing demand for convenient and ready-to-eat products has increased poultry processors' interest in developing consumer-oriented value-added chicken products. In this study, a conjoint analysis survey of 276 chicken consumers in Edmonton was conducted during the summer of 2009 to assess the importance of the chicken part, production method, processing method, storage method, the presence of added flavor, and cooking method on consumer preferences for different value-added chicken product attributes. Estimates of consumer willingness to pay (WTP) premium prices for different combinations of value-added chicken attributes were also determined. Participants'"ideal" chicken product was a refrigerated product made with free-range chicken breast, produced with no additives or preservatives and no added flavor, which could be oven heated or pan heated. Half of all participants on average were willing to pay 30% more for a value-added chicken product over the price of a conventional product. Overall, young consumers, individuals who shop at Farmers' Markets and those who prefer free-range or organic products were more likely to pay a premium for value-added chicken products. As expected, consumers' WTP was affected negatively by product price. Combined knowledge of consumer product attribute preferences and consumer WTP for value-added chicken products can help the poultry industry design innovative value-added chicken products. Practical Application: An optimum combination of product attributes desired by consumers for the development of a new value-added chicken product, as well as the WTP for this product, have been identified in this study. This information is relevant to the poultry industry to enhance consumer satisfaction of future value-added chicken products and provide the tools for future profit growth.     

Influence of Whey Protein Addition and Feed Moisture Content on Chosen Physicochemical Properties of Directly Expanded Corn Extrudates

Production of extrudates from cereals is an often-used technological process in today’s world food industry. Extrudates from corn flour produced using the twin-screw extrusion process and enriched with whey protein concentrate represent high-quality source of proteins and fats. Whey protein concentrate (WPC) as a valuable source of proteins and minerals is one of the highest-quality components for possible extrudate enrichment. In this paper, the influence of various WPC addition and some extrusion process parameters such as feed moisture content ( Q_{textHtext2 textO} Q_{{{text{H}}_{text{2}} {text{O}}}} ) on physicochemical properties of directly expanded corn flour extrudates manufactured in twin-screw co-rotating extruder was investigated. Whey protein concentrate was added in the following ratios 7.5%, 15% and 22.5% and water in 10.08, 12.18 and 14.28 L/h. Final composition of products is determined with measuring of protein, fat and water shares, water absorption index (WAI) and water solubility index (WSI). With added WPC and with increase of water volume flow, there was a significant rise in total protein, fat and water content in final products, as well as lowering of WSI and rising of WAI indexes. The statistical analysis of the obtained data shows that the lowest WSI and the highest WAI had samples with the largest share of WPC (22.5%) and water volume flow of 14.28 L/h. Colour is measured for each sample, and results were represented with hue angle (H), chroma (C) and lightness (L) values. Process parameters, WPC and Q_{textHtext2 textO} Q_{{{text{H}}_{text{2}} {text{O}}}} influence the increase of saturation of C and lightness of L colour value, while H value stays unchanged. Mean value of H was 90.14 ± 1.06, which corresponds to dominance of yellow colour of samples.     

Development of an event-specific detection method for genetically modified rice Kefeng?6 by quantitative real-time PCR

The genetically modified (GM) rice Kefeng?6 has gained resistance against several rice pests by inserting the cpti and cry1Ac genes. As this transgenic line is not approved for import, processing and cultivation in the European Union (EU), sensitive and specific detection methods need to be available to monitor any illegal presence of Kefeng?6 in food products within the EU. The aim of this study was to develop and validate an event-specific detection method by means of quantitative real-time PCR (qPCR) for the detection of Kefeng?6 in foodstuff. A primer pair and hydrolysis probe were designed according to the right border junction sequence of the transgene. The qPCR assay was validated according to the ENGL/EURL-GMFF guidelines for GMO testing and is presented according to the MIQE guidelines. The in-house validation process resulted in a limit of detection of 5 DNA copies of the transgene with confidence intervals (95?%) between 0.07 and 0.52, a PCR efficiency of 105?% and a correlation coefficient (R ²) value of 0.9997. The specificity of the assay was tested by end-point PCR, gel electrophoresis and subsequent sequencing of the PCR products. By testing DNA of several GM and non-GM crops, cross reactivity of the assay was not observed. Further, 35 food products were analyzed for the presence of Kefeng?6 by means of the event-specific detection method. For 9 out of 35 samples, PCR products for Kefeng?6 DNA were observed.     

Temporal trends of synthetic musk compounds in mother's milk and associations with personal use of perfumed products

We analyzed two nitro musks (musk xylene and musk ketone) and five polycyclic musks (HHCB, AHTN, ADBI, ATII, and AHDI) in mother's milk from primiparae women (N = 101) living in Uppsala County, Sweden, 1996-2003. Possible temporal trends in musk concentrations and associations with lifestyle/medical factors, such as use of perfumed products during pregnancy were studied. HHCB showed the highest median concentration (63.9 ng/g lipid) followed by AHTN (10.4 ng/g) and musk xylene (MX) (9.5 ng/g). Concentrations of the other substances were, in most cases, below the quantification limit (2.0-3.0 ng/g). Women with a high use of perfume during pregnancy had elevated milk concentrations of HHCB, and elevated concentrations of AHTN were observed among women reporting use of perfumed laundry detergent. This strongly suggests that perfumed products are important sources of musk exposure both among the mothers and the nursed infants. Concentrations of AHTN and MX declined significantly between 1996 and 2003, suggesting a decline in the industrial use of the compounds in consumer products, or alterations in the consumer use pattern of perfumed products. No temporal trend in HHCB concentrations was seen. The lack of toxicity data makes it difficult to generalize about the safety of musk exposure of breast-fed infants.     

Humidity-Regulating Trays: Moisture Absorption Kinetics and Applications for Fresh Produce Packaging

For packaged fresh produce, inappropriate high relative humidity (RH) levels and condensation of water vapour cause premature spoilage. Humidity-regulating trays were developed to solve this issue. They were made from a thermoformed multilayer structure: polyethylene (outside)/foamed hygroscopic ionomer (active layer) with 0 or 12 wt% NaCl/hygroscopic ionomer (sealing layer, inside). Moisture absorption kinetics of the humidity-regulating trays with 0 and 12 wt% NaCl (T-0 and T-12, respectively) was investigated under different RH conditions (76, 86, 96 and 100 %) at 13 °C for 16 days. Additional trays containing 7 g of distilled water were closed with a high barrier lidding film, and the headspace RH was continuously monitored as a function of time. As control, a polypropylene (control-PP) tray was used. Strawberries and tomatoes were used to test capability of the trays to regulate in-package RH. The amount of water absorbed by the T-0 and T-12 trays was 7.6 and 13.2 g, respectively. Active hygroscopic ionomer layer was effective in water vapour absorption, and the integration of NaCl into this active layer increased the water vapour absorption capacity of the tray. The Weibull model adequately described the moisture sorption kinetics of the individual packaging trays as a function of time. The headspace RH of trays covered with a lidding film was found to be 89.8, 99.6 and 100 % in the T-12, T-0 and control-PP trays, respectively. The T-12 trays containing fresh produce best regulated the in-package RH below 97 % and maintained overall quality, but at the expense of slightly higher product weight loss (2–3 wt% for strawberry, 1 wt% for tomatoes) compared to the control-PP trays (0.3–0.6 wt%).     

An Innovative Submerged Entry Nozzle Design for Billet and Bloom Casting

Fluid flow in the mold has a significant impact on the quality of continuously cast steel products.Since it is strongly influenced by the SEN design,attention h...