Enzyme immunoassay for determination of gluten in foods: collaborative study

A collaborative study was performed in 15 laboratories to validate a monoclonal antibody-based enzyme immunoassay (EIA) for determination of gluten in foods. The study included 13 samples: maize starch, "gluten-free" baking mixes, wheat flours, cookies, cooked meats, and a soup. Gluten was present in these samples at either zero or 0.02 to 10% by weight, i.e., over almost 3 orders of magnitude. The mean assay values for the foods varied from 88 to 105% of the actual amounts. The assay was quantitative for cereal products and the soup with repeatability (RDS-r, relative standard deviation) and reproducibility (RSD-R) of 16-22% and 24-33%, respectively. The assay was semiquantitative for the processed meat products (RSD-r 14 and 26% and RSD-R 46 and 56%), probably because gluten was unevenly distributed in the small (1 g) samples that were analyzed. The ELISA method produced no false positive results, and false negatives obtained with tannin-containing foods could be avoided by use of a modified sample extractant. None of the collaborators reported problems in following the protocol. The method has been adopted official first action by AOAC for determination of wheat gluten in foods.     

Reliable enzyme-linked immunosorbent assay for the determination of walnut proteins in processed foods

Among food allergens of tree nuts, walnuts are a frequent cause of adverse food reactions in allergic patients. A novel sandwich enzyme-linked immunosorbent assay (ELISA) for the detection and quantification of walnut soluble proteins in processed foods was developed. The sandwich ELISA is highly specific for walnut soluble proteins. The recovery ranged from 83.4 to 123%, whereas the intra- and interassay coefficients of variation were less than 8.8 and 7.2%, respectively. This study showed that the proposed method is a reliable tool for detection in the presence of hidden walnut proteins in processed foods.     

Reliable enzyme-linked immunosorbent assay for the determination of soybean proteins in processed foods

Among allergenic foods, soybean is known as a food causing adverse reactions in allergenic patients. To clarify the validity of labeling, the specific and sensitive detection method for the analysis of the soybean protein would be necessary. The p34 protein, originally characterized to be p34 as an oil-body associated protein in soybean, has been identified as one of the major allergenic proteins and named Gly m Bd 30K. A novel sandwich enzyme-linked immunosorbent assay (ELISA) for the detection and quantification of the soybean protein in processed foods was developed using polyclonal antibodies raised against p34 as a soybean marker protein and the specific extraction buffer for extract. The developed sandwich ELISA method was highly specific for the soybean protein. The limit of detection (LOD) and the limit of quantification (LOQ) of the developed ELISA were 0.47 ng/mL (equivalent to 0.19 microg/g in foods) and 0.94 ng/mL (equivalent to 0.38 microg/g in foods), respectively. The recovery ranged from 87.7 to 98.7%, whereas the intra- and interassay coefficients of variation were less than 4.2 and 7.5%, respectively. This study showed that the developed ELISA method is a specific, precise, and reliable tool for the quantitative analysis of the soybean protein in processed foods.     

Reliable enzyme-linked immunosorbent assay for the determination of coconut milk proteins in processed foods

This study was designed to develop a novel sandwich enzyme-linked immunosorbent assay (ELISA) for the detection and quantification of coconut milk proteins in processed foods. The developed sandwich ELISA was able to detect coconut milk proteins from various coconut milk products and did not show any cross-reactivity with 41 of 42 kinds of popularly used food ingredients, thus reflecting great specificity for coconut milk proteins. In addition, the established ELISA is highly sensitive and allowed the detection of 0.31 μg/g of coconut milk protein in complex food matrices. This proposed assay could serve as a useful tool for the detection of the presence of hidden coconut milk proteins in processed foods.     

Gas chromatography-mass spectrometry determination of phosphine residues in stored products and processed foods

A gas chromatography-mass spectrometry (GC-MS) method was used for the quantitative confirmation of phosphine residues in stored products and processed foods. An established extraction technique was utilized for the preparation of headspace samples, which were analyzed by GC-MS and gas chromatography-nitrogen-phosphorus detection (GC-NPD). Wheat, oats, maize, white rice, brown rice, cornflakes, tortilla cornchips, groundnuts, and raisins were validated, showing excellent agreement between detectors when spiked at levels equivalent to 0.001 and 0.01 mg/kg phosphine and for samples containing incurred residues. The GC-MS method was reproducible and accurate when compared to the GC-NPD method and allowed five samples to be quantified in a working day. Subambient GC-MS oven temperatures were most suitable for phosphine residues ranging from 0.001 to 0.005 mg/kg, and a GC oven temperature of 100 degrees C was appropriate for residues >0.005 mg/kg. The method was sufficiently robust to be evaluated for other similar commodities as the need arises.     

Fast real-time PCR for the detection of crustacean allergen in foods

Crustaceans are one of the most common allergens causing severe food reaction. These food allergens are a health problem, and they have become very important; there are various regulations that establish that labeling must be present regarding these allergens to warn consumers. In the present work a fast real-time PCR, by a LNA probe, was developed. This allows the detection of crustaceans in all kinds of products, including processed products in which very aggressive treatments of temperature and pressure during the manufacturing process are used. This methodology provides greater sensitivity and specificity and reduces the analysis time of real-time PCR to 40 min. This methodology was further validated by means of simulating products likely to contain this allergen. For this, products present on the market were spiked with crustacean cooking water. The assay is a potential tool in issues related to the labeling of products and food security to protect the allergic consumer.     

Fluorescent excitation transfer immunoassay for the determination of spinosyn A in water.

A fluorescent excitation transfer immunoassay for spinosyn A, a fermentation derived insect control agent, has been developed and applied to the analysis of tap water and wastewater effluent from manufacturing plants. Fluorescein (F) and tetramethylrhodamine (TMR) were chosen as donor and quencher, respectively, for the excitation transfer. Fluorescence quenching was observed from the binding of F-labeled antigen to TMR-labeled antibody. By employing nonlabeled antigen in a competitive immunoassay format, we reversed fluorescence quenching. The assay provides a limit of detection of 0. 01 ppb and a working range of 0.05-1 ppb and allows for the rapid determination of spinosyn A in water with recovery values ranging from 96% to 120%. With the exploitation of the small size of optical fibers, fluorescence from an assay volume of 24 microL could be measured without special vessels.     

Enzyme immunoassay for detection of Salmonella in foods: collaborative study

A collaborative study was performed in 25 laboratories to validate an enzyme immunoassay (EIA) procedure utilizing 2 specific monoclonal antibodies for rapid detection of Salmonella in foods. The EIA was compared with the standard culture procedure for detection of Salmonella in 6 food types: ground black pepper, soy isolate, dried whole eggs, milk chocolate, nonfat dry milk, and raw deboned turkey. Uninoculated and inoculated samples were included in each food group analyzed, with the exception of poultry which was naturally contaminated. There was no significant difference in the productivity of the EIA and culture procedures at the 5% level for any of the 6 foods. The enzyme immunoassay screening method has been adopted official first action.     

Visual immunoassay for detection of Salmonella in foods: collaborative study

A collaborative study was performed in 13 laboratories to validate a visual enzyme immunoassay (EIA) procedure, TECRA, for rapid detection of Salmonella in foods. The EIA method was compared with the standard culture procedure for detection of Salmonella in 6 food types: ground black pepper, soy flour, dried whole eggs, milk chocolate, nonfat dry milk, and raw deboned turkey. Uninoculated and inoculated samples were included in each food group analyzed. There was no significant difference in the productivity of the EIA and culture procedures at the 5% level for any of the 6 foods. The enzyme immunoassay screening method has been approved interim official first action.     

A review of bromine determination in foods.

Organic compounds containing bromine, including methyl bromide, ethylene dibromide, and 1,2-dibromo-3-chloropropane, have been used extensively for the fumigation of foods, or soils in which foods grow, making it necessary to determine residues of bromine and bromine-containing organic compounds. A large number of methods for the determination of bromine in foods, as organic, inorganic, and combined total bromide, have been developed. In methods for organic bromide, the bromine is converted to the inorganic form for measurement by titration, photometry, or other means. In recent years, instrumental methods have been developed in which the total bromine in the sample is determined, regardless of the state in which it exists. X-ray fluorescence and neutron activation analysis are the 2 instrumental methods used most widely. Residue data are presented for some typical bromine-containing samples.     

A multiresidue method for the determination of insecticides and triazine herbicides in fresh and processed olives

A simple, rapid, and low-cost gas chromatographic multiresidue method has been developed for the analysis of pesticide residues in raw and processed olives. This has been validated for 19 insecticides and triazine herbicides, covering a wide range of polarities. The method uses low-temperature precipitation to remove lipids and gives good cleanup for gas chromatography analysis with nitrogen phosphorus and electron capture detection. Recoveries are between 71 and 99%, with relative standard deviation values of 5-15%.     

Enzyme immunoassay for detection of Drosophila melanogaster antigens in the juice of various foods

An enzyme immunoassay (EIA) was developed to identify and quantitate soluble antigens originating from Drosophila melanogaster eggs. Polystyrene microtiter plates were coated with anti-egg antibody. Egg antigen standards or samples were reacted with the sensitized wells. The immobilized antigen was reacted with biotinylated antibody, and the complex was quantitated by an avidin-horseradish peroxidase conjugate. The assay can detect egg antigens equivalent to 0.03 eggs/mL. The antibody was directed against a number of antigens and cross-reacted with extracts of adult Drosophila melanogaster flies. Homogenized eggs diluted in various juice samples were detectable by EIA. Heating the antigen diluted in buffer or juice at a temperature used in processing of juice reduced the immunological response approximately 40-45%. Brine separated from fresh-packed sauerkraut showed EIA responses equivalent to 0.3-3 eggs/mL. Electrophoretic, chromatographic, and immunological analyses suggested that the EIA responses for sauerkraut brine were related to the Drosophila melanogaster egg antigens.     

Enzyme immunoassay for detection of Salmonella in low-moisture foods: collaborative study

A collaborative study was performed in 15 laboratories to evaluate a modification of the enzyme immunoassay (EIA) method for detection of Salmonella in foods (46.B21-46.B29). The modified EIA requires 18-24 h pre-enrichment, 6-8 h selective enrichment, and 14-18 h M-broth post-enrichment prior to performing the assay, which requires 1-2 h. Total assay time is 40-52 h. The modified method was compared with the standard culture method for detection of Salmonella in 5 low-moisture foods: nonfat dry milk, milk chocolate, meat and bone meal, dry whole egg, and ground pepper. The modified method has been adopted official first action for use with low-moisture foods.     

A quantitative stable-isotope LC-MS method for the determination of folic acid in fortified foods

A stable-isotope liquid chromatography-mass spectrometry (LC-MS) assay was developed for the quantitative determination of folic acid in fortified foods. Folic acid was extracted from food samples into a phosphate buffer, purified on a C-18 Sep-Pak cartridge, and analyzed by LC-MS in the negative ion mode using electrospray ionization. The analyte was quantified using (13)C(5)-folic acid as an internal standard. The coefficient of variation for the precision of the method was 5.6% based on the analysis of four sample replicates. The accuracy of the method was assessed using a standard method of addition of folic acid to a shredded whole-wheat cereal. The quantitative determination of folic acid in this matrix was linear over 1 order of magnitude having a concentration range of 2.4 to 24 microg/g of food (or 0.05 to 0.5 microg of analyte injected into the LC-MS). The overall quantitative efficiency of the method was evaluated using a standard reference material (infant formula SRM 1846). The method was applied to the determination of folic acid in several test samples (fortified breakfast cereals), and the values were in accord with the manufacturer's claim. This method advances a LC-MS technique for the determination of folic acid in fortified foods based on stable-isotope dilution methodology. The specificity of the technique and quantitative accuracy of the method in various food substrates suggests that the method may be adapted for routine analysis in other fortified foods.     

A resuscitation/selection system for rapid determination of Salmonella in foods

A resuscitation medium was developed consisting of a trypticase soy broth base supplemented with 0.5% yeast extract, 0.25% sodium pyruvate, 0.01% sodium thioglycollate, and 0.1% chicken fat. After a resuscitation period of 4 h, the medium was made selective by addition of either sodium thiosulfate, bile salts and iodine, or sodium selenite and L-cystine. The now selective medium was incubated for 16 h. The presence or absence of Salmonella was determined by the Salmonella-Tek antibody-based detection kit. The present system was compared with a method of the Bacteriological Analytical Manual (BAM) for naturally contaminated foods. Nineteen egg products were screened; 3/19 were positive using the BAM method, 9/19 were positive using the present system. Seventeen chicken samples were assayed; 10/17 were positive using the BAM method; 13/17 were positive using the present system. Of 8 pepper samples, 4/8 were positive using the BAM method; 6/8 were positive using the present system. Of 8 spice samples, 6/8 were positive using the BAM method, 7/8 were positive using the present system. Of 6 onion products sampled, 5/6 were positive using the BAM method; 6/6 were positive using the present system.     

Improved method for determination and identification of serotonin in foods

A previously described method to identify and quantitate serotonin in foods has been improved. The extraction and separation of serotonin from interfering substances has been improved, and the scope of material to which the method may be applied has been widened. The relative standard deviation (RSD) for repeated determinations of serotonin in canned fried tomato purée and the average recovery of serotonin added to the same sample was 6.25 and 89.9%, respectively. The method showed the presence of serotonin in apricots, cherries, and peaches.     

Sandwich immunoassays for the determination of peanut and hazelnut traces in foods

People suffering from food allergies are dependent on accurate food labeling, as an avoidance diet is the only effective countermeasure. Even a small amount of allergenic protein can trigger severe reactions in highly sensitized patients. Therefore, sensitive and reliable tests are needed to detect potential cross-contamination. In this paper two fast sandwich immunoassays are described for the determination of peanut (Arachis hypogaea) and hazelnut (Corylus avellana) traces in complex food matrices. Mouse monoclonal antibodies were used as capture antibodies, and labeled rabbit polyclonal antibodies were used as detection antibodies in both assays. The assay time was 30 min in total, and cross-reactivities against a variety of fruits and seeds were found to be in the low 10(-4)% (ppm) level or in some cases not detectable. The recoveries in all tested food matrices ranged from 86 to 127%, and the limits of detection were in the range of 0.2-1.2 mg/kg (ppm) in food for both peanut and hazelnut, respectively.     

Fluorescent enzyme immunoassay for rapid screening of Salmonella in foods: collaborative study

A collaborative study was performed in 13 laboratories to validate an enzyme immunoassay (EIA) procedure for rapid detection of Salmonella in foods. The EIA was compared with the standard culture procedure for detection of Salmonella in 6 food types: ground black pepper, soy flour, dried whole eggs, milk chocolate, nonfat dry milk, and raw deboned turkey. Uninoculated and inoculated samples were included in each food group analyzed. There was no significant difference in the proportion of samples positive by the EIA and culture procedures at the 5% level for any of the 6 foods. The enzyme immunoassay screening method has been adopted official first action as a rapid screening method for detection of Salmonella.     

A mass spectrometric validated high-performance liquid chromatography procedure for the determination of folates in foods

A series of five food reference materials (RM) that had certified values of folate concentrations and five frozen food samples were analyzed for 5-methyltetrahydrofolic acid (5-MTHFA) and folic acid (FA) using a high-performance liquid chromatography (HPLC) method with fluorescence detection that was validated using an HPLC mass spectrometry (MS) method with electrospray ionization. Identical sample specimens were extracted and analyzed in triplicate using both instrumental methods, and a comparison was made of the mean values of 5-MTHFA and FA resulting from these determinations. The analytes were isolated on either a high capacity strong anion exchange solid phase extraction column (HPLC method) or a phenyl Bond Elut column (MS method) prior to analyses. For quantification of the analytes by MS, (13)C-labeled 5-MTHFA and FA were added to samples as internal standards prior to enzymatic digestion and conversion of the polyglutamate forms of 5-MTHFA to the monoglutamic acid. Quantification of FA and 5-MTHFA using the HPLC analysis was carried out using external standards. With the exception of one RM (pig liver), the values established for 5-MTHFA using these methods were highly comparable. In determining the variance associated with these two procedures, it was observed that the mean relative standard error for 5-MTHFA was 12 (range, 2-27%) and 11% (range, 5-25%) for the HPLC and MS methods, respectively. FA was detected in only three of the samples, and the values obtained for it by either method were similar. This is the first paper that describes a mass spectrometric method used in the validation of an HPLC determination of food folates across a wide range of sample matrixes. The comparable values for 5-MTHFA and FA suggest that HPLC analysis with fluorescent detection may be used to accurately quantify folates present in a variety of food matrixes.     

Catechin contents of foods commonly consumed in The Netherlands. 1. Fruits, vegetables, staple foods, and processed foods

Catechins, compounds that belong to the flavonoid class, are potentially beneficial to human health. To enable epidemiological evaluation of these compounds, data on their contents in foods are required. HPLC with UV and fluorescence detection was used to determine the levels of (+)-catechin, (-)-epicatechin, (+)-gallocatechin (GC), (-)-epigallocatechin (EGC), (-)-epicatechin gallate (ECg), and (-)-epigallocatechin gallate (EGCg) in 24 types of fruits, 27 types of vegetables and legumes, some staple foods, and processed foods commonly consumed in The Netherlands. Most fruits, chocolate, and some legumes contained catechins. Levels varied to a large extent: from 4.5 mg/kg in kiwi fruit to 610 mg/kg in black chocolate. (+)-Catechin and (-)-epicatechin were the predominant catechins; GC, EGC, and ECg were detected in some foods, but none of the foods contained EGCg. The data reported here provide a base for the epidemiological evaluation of the effect of catechins on the risk for chronic diseases.