Method of determination of appropriate heat treatment of animal meal by immunoassay developed for detection of cooked beef: interlaboratory study.

An interlaboratory trial was conducted for the validation of an enzyme-linked immunosorbent assay (ELISA) method for determination of appropriate heat treatment of animal meal. A commercially available ELISA test kit developed for the identification of beef in cooked food was used in the study. Twelve laboratories from 7 European countries examined 2 different analytical protocols to establish the most appropriate analytical method. Three different samples were used, 2 animal waste materials sterilized at 129 and 134 degrees C (wet conditions), respectively, and a meat and bone meal material processed at dry conditions (maximum temperature, 140 degrees C). Statistical evaluation applying t-statistics showed that the animal meal treated according to European legislation (>133 degrees C) was clearly distinguishable from the 2 other test materials at a 99% confidence level using both analytical protocols. This method can be considered as a complementary test to the immunoassay developed for the detection of pork in cooked food that is already applied in routine analysis for the surveillance of rendering plants.     

Detection of pork in heat-processed meat products by monoclonal antibody-based ELISA

An enzyme-linked immunosorbent assay (ELISA) using a monoclonal antibody to a porcine thermal-stable muscle protein was developed for detection of pork in cooked meat products. The assay specifically detects porcine skeletal muscle, but not cardiac muscle, smooth muscle, blood, and nonmuscle organs. No cross-reactivity was observed with common food proteins. Validity of the assay was evaluated with laboratory formulated and commercial meat samples. The detection limit was determined as 0.5% (w/w) pork in heterologous meat mixtures. Overall, intra- and inter-assay coefficients of variation were 5.8 and 7.9%, respectively. The accuracy in analyzing market samples was 100% as verified by product labeling and confirmed by a commercial polycolonal antibody test kit.     

Detection of residues of tetracycline antibiotics in pork and chicken meat: correlation between results of screening and confirmatory tests.

Residues of the tetracycline group of antibiotics were quantified in pork and chicken muscle tissue that had previously been screened with a microbiological inhibition test and an immunological method. Pieces of frozen pork and chicken meat were screened on a pH 6 culture medium seeded with Bacillus subtilis. An aqueous extract of the inhibitor-positive samples was then screened with a group-specific commercial ELISA kit, able to detect levels of oxytetracycline, chlortetracycline, tetracycline and doxycycline corresponding with the European MRL or lower. The cut-off value of the ELISA was set at a B/B0 value of 75%. Finally, confirmation and quantification were performed using a validated HPLC method with fluorescence detection. The fluorescence was induced by complexation of the tetracyclines with the zirconium cation which is added post-column to the HPLC eluate. This fluorescence makes it possible to quantitate residues below one-half of the MRL. To gain additional qualitative information some samples were also analysed with LC-MS-MS. ELISA analysis demonstrated the presence of residues of tetracyclines in 12 out of 19 inhibitor-positive pork samples and in 19 out of 21 inhibitor-positive chicken samples. Doxycycline was detected with HPLC in 10 of these 12 pork samples and in 18 out of 19 chicken samples. The two other ELISA positive pork samples contained oxytetracycline, while no tetracyclines were found in one ELISA positive chicken meat sample. The correlation between the ELISA B/B0 values and the actual levels determined with the HPLC method was poor, whereas a better correlation was observed between the inhibition zones and the doxycycline levels. Our results indicate that an inhibition test with a medium at pH 6 and B. subtilis as test organism is well suited to screen pork and chicken muscle tissue for residues of tetracycline antibiotics. Since many positive samples contained doxycycline levels below the MRL, a confirmatory method is necessary to quantify the residues.     

Immunoassay kit used to detect the presence of bovine material in processed foods

The Tepnel Bio Kit for the detection of beef in cooked foods was assessed to determine its validity in demonstrating if food being imported into New Zealand contains beef material. The test suffered no interference from the presence of other common nonbovine species meats accepted as food within New Zealand and it detected beef in cooked samples of mixed meats when the proportion of beef in the mixture was >2 or >1%, depending on other meat species present. The documentation supplied with the kit indicates that the specific proteins it measures in cooked beef are stable to 130 degrees C. This was confirmed in the literature when the kit was used to test meat and bone meal cooked to at least 133 degrees C. However, our results showed these proteins to be much less stable when heated to elevated temperatures in moist food under pressure, and samples containing beef ceased to be positive by the immunoassay test after being autoclaved to 121 degrees C. This suggests that the test may not be able to detect even relatively high levels of beef in low-acid canned foods, which are normally retorted under pressure to approximately 121 degrees C.     

Meat species identification by linear discriminant analysis of capillary electrophoresis protein profiles

The objective of this study was to utilize linear discriminant analysis (LDA) in the interpretation of capillary electrophoresis-sodium dodecyl sulfate polymer-filled capillary gel electrophoresis (CE-SDS) meat protein profiles for the identification of meat species. The specific objectives were 1) to collect quantitative data on water-soluble and saline-soluble proteins of different meat species obtained by CE-SDS and 2) to apply LDA on collected CE-SDS protein data for the development of a pattern recognition statistical model useful in the differentiation of meat species. Samples were raw beef top and eye round, boneless fresh pork ham and loin, turkey leg and breast meat, and mechanically deboned turkey meat collected on six different occasions, making a total of 42 samples. Additionally, 14 samples were used as test samples to determine the classification ability of the procedure. Quantitative protein data obtained by CE-SDS was used to generate separate LDA models for either water- or saline-soluble protein extracts. Although a saline solution was a more efficient meat protein-extracting agent, as shown by a higher total protein concentration and a larger number of peaks, water-soluble CE-SDS protein profiles gave more distinctive discrimination among meat species. The correct classification given by LDA on water-soluble protein data was 100% for all meat species, except pork (94%). Conversely, the correct classification on saline-soluble protein data was 88% for beef and mechanically deboned turkey meat, and 94% and 100% for turkey and pork meat, respectively. LDA proved to be a useful pattern recognition procedure in the interpretation of CE-SDS protein profiles for the identification of meat species.     

Chemometric Analysis of Fatty Acid Composition of Raw Chicken,Beef, and Pork Meat with Plant Extract Addition during Refrigerated Storage

During the shelf-life, meat undergoes a number of processes that negatively affect the quality of the product, including fatty acid composition. The application of various plant extracts in meat could affect the changes of fatty acids during storage. Thus, the aim of this study was to investigate the effect of various spice and herb extracts on fatty acid composition in raw pork, beef, and chicken meat when stored at 4 °C for 13 days. Based on multivariate statistical analysis, two datasets were extracted from each type of meat. One dataset included samples with allspice, bay leaf, black seed, cardamom, caraway, clove, and nutmeg with the high share of total MUFA (monounsaturated fatty acids) in chicken and pork meat and high MUFA and PUFA (polyunsaturated fatty acids) contribution in beef meat after storage. The second dataset included basil, garlic, onion, oregano, rosemary, and thyme with high PUFA share in chicken and pork meat and high SFA (saturated fatty acids) contribution in beef meat. From the regression analysis, a significant effect of time on fatty acid composition in meat was reported. Generally, the rates of fatty acid changes were dependent on the plant extract incorporated into the meat. The most visible effect of plant extracts was obtained in chicken meat. In chicken meat with plant extracts, the rates of SFA and PUFA changes with time were slower compared to the control sample. In summary, the fatty acid composition of intramuscular fat varied during storage, and the addition of plant extracts significantly affected the rate of these changes, which was dependent on the meat matrix.     

Determination of selenium in meat products by hydride generation atomic absorption spectrophotometry

Wet digestion using a mixture of nitric, sulfuric, and perchloric acids and an aluminum block digester effectively and rapidly decomposed meat samples for selenium determination by hydride generation atomic absorption spectrophotometry. Digestion did not require constant attention by an operator. Selenium recoveries (range, 94-105%) from National Institute of Standards and Technology standard reference materials and spiked samples were used to validate method accuracy. Coefficients of variation (CVs) of repeatability of in-house reference materials used for precision study were 6.4 and 5.6%, respectively, for seafood mix and mutton liver. Selenium levels in meat products from Brisbane markets varied widely: 0.042-0.142, 0.081-0.42, and 0.050-0.198 microgram/g (wet weight) respectively, for beef, chicken, and pork. Overall, selenium levels in manufactured meat ranged from 0.041 to 0.189 microgram/g. The levels of selenium found in this study were generally lower than those reported in Finland but comparable with those reported in some parts of the United States.     

Establishment of a multiplex-PCR detection method and development of a detection kit for five animal-derived components in edible meat

A multiplex polymerase chain reaction (PCR) detection method for the simultaneous detection of animal-derived components from deer, cow, sheep, pig and horse in edible meat was established, and a multiplex PCR detection kit for the rapid detection of animal-derived components was developed. According to the mitochondrial cytochrome b (Cyt b) gene of bovine species, sheep species, pig species and horse species and the mitochondrial cytochrome c oxidase subunit I (COX 1) gene of sika deer and red deer as the target gene sequences of primers, the specific primers of five different species were designed, the PCR system was optimized, and the multiplex PCR identification method of five animal-derived components was established. The minimum detection amount was determined by sensitivity test. The results showed that five meat specific amplification bands could be found at the same time in the same reaction system, including 173 bp fragment for venison, 148 bp for beef, 261 bp for pork, 100 bp for mutton and 424 bp for horse, indicating that the method is specific and stable. The minimum detection limit by this method was 1 ng/μL, showing a high sensitivity. According to the different sites in different areas of animal mitochondrial genes, a multiplex PCR detection method was established and a detection kit was developed, and the rapid, sensitive, stable and high-throughput detection of five animal-derived components and adulterated animal components in edible meat can be realized by using the kit.     

双羟基复合金属氧化物的晶面生长选择性及晶粒尺寸控制

采用成核／晶化隔离法合成镁铝双羟基复合金属氧化物（LDH）,考察了晶化温度及晶化时间对晶体结构,晶面生长选择性及晶粒尺寸的影响规律。结果发现晶化温度相同时,随晶化时间延长,LDH的晶体结构趋于完整,晶粒尺寸增大;晶化时间相同时,随晶化温度升高,晶体结构趋于完整,晶粒尺寸显著增大,实验条件下得到的LDH,其沿a轴方向的尺寸均大于沿c轴方向的尺寸,即共沿a轴方向的生长速率比沿c轴方向的生长速度快,亦即[110]晶面的生长速率比[003]晶面的生长速率快。根据LDH晶粒尺寸随晶化时间及晶化温度的变化规律,选择合适的晶化条件,制备得到了粒径分布窄的纳米LDH。     

超高效液相色谱-串联质谱法鉴定猪肉的特异性肽生物标志物

以亲缘关系较近的猪、牛和羊3个物种的肌肉组织为研究对象,采用超高效液相色谱-串联质谱(UPLC-MS),筛选并确认了猪物种肉特异性肽生物标志物.3种纯肉样品经蛋白质提取、胰蛋白酶消化和UPLC-TripleTOF-MS分离鉴定,得到的总离子流图谱(TIC)与Uniprot蛋白质数据库对比分析,筛选出3个物种肉的3种高丰度同源蛋白和8种潜在的肽生物标志物;潜在的肽生物标志物经QTRAP-MS质谱的多反应模式(MRM)分析,最终确认了猪物种肉的5种肽生物标志物,其中3种肽生物标志物未见文献报道.     

Determining the temperature dependence of the emulsion particle size of polyacrylate core–shell and polystyrene by photon correlation spectroscopy

The dependence of the particle size of a polyacrylate core-shell emulsion on temperature has been investigated in the temperature range of 10–55°C by photon correlation spectroscopy. To compare, the dependence of the particle size of an aqueous suspension of a polystyrene standard on temperature has been also investigated under the same conditions. This showed that as the temperature increases, the particle size of both samples decreases, but the rate of size decrease of the polystyrene standard is larger than that of the polyacrylate core–shell emulsion. By linear regression analysis, two regression equations of both samples have been set up. Furthermore, the apparent moving activation energy has been worked out from the size–temperature data.     

Screening of salbutamol residues in swine meat and animal feed by an enzyme immunoassay in Taiwan

An ELISA was developed for routine examination for extensive monitoring and screening programs for the residues of salbutamol in swine serum, animal feed, meat, and meat-related products destined for human consumption in Taiwan. Objectives of the study were to investigate the use of a new immunoassay for the detection of salbutamol residues in swine meat and animal feed samples, and to compare with a commercial kit in field test screens. A fast, simple and reliable sample preparation method for the determination of salbutamol was established. Field trials with 222 swine meat and 120 animal feed samples that were taken from local meat markets, auction markets and feed mills. The application and the results of two ELISA kits (a homemade and a commercial kit) for the screening of salbutamol were presented. Adopting 2 μg kg⁻¹ salbutamol as a cut-off value for swine meat, the commercial β-agonist ELISA had a sensitivity of 85.3% and a specificity of 95.2% versus GC-MS at a cut-off of 2 μg kg⁻¹. The homemade salbutamol ELISA had a sensitivity of 100% and a specificity of 90.9% and gave no false-negative rate results. Furthermore, adopting 20 μg kg⁻¹ salbutamol as a cut-off value for animal feed, both the commercial and homemade ELISA showed 100% sensitivity and 100% specificity of the assays. In conclusion, a sensitive, specific salbutamol polyclonal antibody-based ELISA has been developed that could serve as a rapid screening assay, and the detection of positive samples at the place of sampling can result in more effective control of the illegal use of β-agonists.     

Evaluation of two commercial solid-phase microextraction fibres for the analysis of target aroma compounds in cooked beef meat

The aroma profile of cooked beef meat has been investigated by solid-phase microextraction (SPME) combined with gas chromatography-mass spectrometry (GC-MS). Out of more than 200 volatile compounds, 36 key odour-active compounds were selected for analysis. Several extraction times, desorption times, temperature conditions and fibre types were tested to achieve a fast and reproducible extraction, and a representative analysis of the aroma profile of cooked beef. Extraction conditions and fibre type significantly affected the majority of the target compounds. Divinylbenzene-carboxen-polydimethylsiloxane (DVB-CAR-PDMS) fibre presented a better reproducibility at all extraction times and extracted more efficiently the less volatile compounds than the carboxen-polydimethylsiloxane (CAR-PDMS) fibre. The high molecular weight compounds seemed to achieve faster the equilibrium between the headspace and DVB-CAR-PDMS fibre. The use of SPME was shown to be a simple, sensitive, selective, representative, fast, and low-cost method for the evaluation of key odour-active compounds in cooked beef meat, even if further research on quantitative analysis of volatiles using SPME on solid samples has to be done.     

镁铁双羟基复合金属氧化物的可控合成及晶面生长特征研究

采用成核/晶化隔离法合成镁铁双羟基复合金属氧化物MgFe-LDH,考察了Mg ∶ Fe摩尔比对MgFe-LDH晶形的影响,探讨了晶化温度及晶化时间对晶面生长选择性及晶粒尺寸的影响规律.结果表明,随Mg ∶ Fe摩尔比增大,层板阳离子排列更为规整.晶化温度对晶粒尺寸的影响显著大于晶化时间的影响.晶化温度相同,随晶化时间延长, MgFe-LDH的晶体结构趋于完整,晶粒尺寸变化不大;晶化时间相同,随晶化温度升高,晶体结构趋于完整,晶粒尺寸明显增大.所得到的MgFe-LDH沿a轴方向的晶粒尺寸对晶化温度变化的敏感程度远大于对晶化时间变化的敏感程度,但总是沿a轴方向的晶粒尺寸大于沿c轴方向的尺寸,即[110]晶面的生长速率比[002]晶面的生长速率快.     

A Novel and Reusable Electrochemical Genosensor for Detection of Beef Adulteration

This work describes the construction of a genosensor based on a graphite electrode modified with an reduced graphene oxide/poly(3-hidroxybenzoic acid) nanocomposite with an specific DNA oligonucleotide for detection of cattle mitochondrial DNA, in order to certify beef purity. Electrochemical and morphological analyses indicate that the genosensor allows duplex formation with the DNA of pure sample of beef lysate. The genosensor was selective, identifying up to 1 % (w/w) of pork in beef samples, showing good reproducibility and stability within six weeks of storage, and can be reused four times, being a great tool for the evaluation of beef purity, with application in the meat production and marketing chain.     

Effect of gamma radiation on microbial population of natural casings

The high microbial load of fresh and dry natural casings increases the risk of meat product contamination with pathogenic microorganisms, agents of foodborn diseases.

The aim of this work is to evaluate the killing effect of gamma radiation of the resident microbial population of pork and beef casings, to improve their hygiene and safety.

Portions of fresh pork (small intestines and colon) and dry beef casings were irradiated in a Cobalt 60 source with with absorbed doses of 1,2,5 and 10 kGy.

The D₁₀ values of total aerobic microorganisms in the pork casings were 1.65 kGy for colon and 1.54 kGy for small intestine. The D₁₀ value found in beef dry casings (small intestine) was 10.17 kGy. Radurization with 5 kGy was able to reduce, at least, 6 logs the coliform bacteria in pork casings. The killing effect over faecal Streptococci was 4 logs for pork fresh casings and 2 logs for beef dry casings. Gamma radiation with 5 kGy proved to be a convenient method to reduce substantially the microbial population of pork fresh casings. Otherwise, the microbial population of beef dry casings still resisted to 10 kGy.     

Pre-column derivatization of sulfa drugs with fluorescamine and high-performance liquid chromatographic determination at their residual levels in meat and meat products.

A rapid, sensitive and selective high-performance liquid chromatographic method is described for simultaneous determination of eight sulfa drugs in meat and meat products using pre-column derivatization with fluorescamine. The drugs are sulfisomidine, sulfadiazine, sulfamerazine, sulfadimidine, sulfamonomethoxine, sulfamethoxazole, sulfadimethoxine and sulfaquinoxaline. The method includes blender extraction of 3-g samples with chloroform, partition with 3 M hydrochloric acid, derivatization with fluorescamine at pH 3.0 and subsequent high-performance liquid chromatographic analysis on a C18 column with fluorescence detection at an excitation wavelength of 405 nm and an emission wavelength of 495 nm. The drugs were separated with a mobile phase of acetonitrile-2% acetic acid (3:5) at 55 degrees C. The average recovery from samples fortified at 0.1 ng/g was 92.6% with a coefficient of variation of 6.2%. The detection limit was 0.01 ng/g for sulfaquinoxaline and 0.005 ng/g for the other seven drugs. The method was field-tested in a survey of 37 samples including beef (five), pork (seven), chicken (seven), ham (five), sausage (eight), bacon (two) and roast beef (three). Sulfadimidine was detected in one pork sample at the level of 0.295 ng/g and in ham at 0.178 ng/g.     

T形微反应器共沉淀法制备Mg-Al层状双金属氢氧化物及其粒径可控性

采用T形微反应器通过共沉淀法制备了Mg-Al层状双金属氢氧化物(LDHs)纳米颗粒, 考察了流速、混合盐溶液浓度和温度等对产物粒径及其分布的影响. 实验结果表明, 所制备的LDHs样品的形貌和晶体结构与传统共沉淀法结果一致, 但本方法制备的样品粒径小、分布窄. 随着流速增大, 温度升高, 所合成的LDHs样品平均粒径减小, 分布变窄; 而随着混合盐溶液浓度的增大, 所得LDHs样品粒径增大, 分布变宽.     

Identification of low level gamma-irradiation of meats by high sensitivity comet assay

The detection of low levels of irradiation in meats (pork, beef, and chicken) using the new comet assay was investigated in order to assess the capability of the procedure. The new assay includes a process that improves its sensitivity to irradiation and a novel evaluation system for each slide (influence score and comet-type distribution). Samples used were purchased at retailers and were irradiated at 0.5 and 2 kGy at 0°C. The samples were processed to obtain comets. Slides were evaluated by typing comets, calculating the influence score and analyzing the comet-type distribution chart of shown on the slide. Influence scores of beef, pork, and chicken at 0 kGy were 287(SD=8.0), 305 (SD=12.9), and 320 (SD=21.0), respectively. Those at 500 Gy, were 305 (SD=5.3), 347 (SD=10.6), and 364 (12.6), respectively. Irradiation levels in food were successfully determined. Sensitivity to irradiation differed among samples (chicken>pork>beef).     

Quantification of meat proportions by measuring DNA contents in raw and boiled sausages using matrix-adapted calibrators and multiplex real-time PCR

The quantification of meat proportions in raw and boiled sausage according to the recipe was evaluated using three different calibrators. To measure the DNA contents from beef, pork, sheep (mutton), and horse, a tetraplex real-time PCR method was applied. Nineteen laboratories analyzed four meat products each made of different proportions of beef, pork, sheep, and horse meat. Three kinds of calibrators were used: raw and boiled sausages of known proportions ranging from 1 to 55% of meat, and a dilution series of DNA from muscle tissue. In general, results generated using calibration sausages were more accurate than those resulting from the use of DNA from muscle tissue, and exhibited smaller measurement uncertainties. Although differences between uses of raw and boiled calibration sausages were small, the most precise and accurate results were obtained by calibration with fine-textured boiled reference sausages.