Determination of soybean proteins in commercial heat-processed meat products prepared with chicken,beef or complex mixtures of meats from different species

The addition of foreign proteins (mainly soybean proteins and milk proteins) to heat-processed meat products is a common practice. This work approaches the determination of additions of soybean proteins in heat-processed meat products prepared with chicken meat, beef meat, and complex mixtures of meats from different species (chicken, pork, beef, and turkey) by perfusion reversed-phase high-performance liquid chromatography. The applied method was previously developed for the determination of soybean proteins in pork and turkey meat products but it has never been tested for the determination of soybean proteins in other heat-processed meat products containing other kinds of meats. This paper demonstrates the validity of this method for the detection of soybean proteins in heat-processed meat products containing different varieties of meats and even in the presence of other foreign proteins such as milk proteins. The specificity and existence of matrix interferences have been checked for these samples and accuracy has been evaluated by the comparison of the soybean protein contents determined by the proposed method and the official ELISA method.     

Nutritive value of meat-soy mixture

The protein value of meat soy blend was evaluated by utilizing the net protein ratio (NPR), the relative protein value (RPV) and the serum urea content methods. It was compared with that of meat. Casein was used as a reference protein. NPR values indicated that utilization of meat and meat soy proteins are comparable. The study of amino acid pattern shows that sulphur containing amino acids are limiting to almost the same degree in meat and meat soy blend. However, the RPV of meat soy bean blend is slightly higher than that of meat. The lowest serum urea content was that of rats fed meat soy blend. Therefore, the mixing of meat with soy bean did not reduce the nutritive value of meat, on the contrary there is tendency towards improvement.     

Specific degradation of myosin in meat by bromelain

The specificity of bromelain and papain for degradation of actin and myosin in meat was compared. Meat was treated with 0·1% enzyme for 0–60 min at 24°C and proteins, extracted with 6 urea containing 2% sodium dodecylsulfate (SDS) solution, were separated by SDS-polyacrylamide gel electrophoresis (PAGE). The profile of the extracted proteins clearly indicated that papain degrades myosin and actin at similar rates, whereas bromelain degrades myosin preferentially. This distinction could be useful in the development of freeze-dried meat products in which toughening of the meat due to the formation of high molecular weight aggregates from myosin cross-linking is minimized.     

Comparison of immunohistochemical,histochemical and immunochemical methods for the detection of wheat protein allergens in meat samples and cooked,dry, raw and fermented sausage samples

Nowadays is it a common practice to add vegetable protein in the production of meat products. Because of the possible substitution of high-quality raw meat with vegetable protein without the labelling the product package and because of the allergenic potential of many vegetable proteins, it is important to develop accurate methods for its detection. The objective of the study was to compare histochemical, immunochemical (ELISA, ALERT gliadin screening test) and immunohistochemical methods for the detection of wheat protein in meat samples and sausages. Histochemical methods were useful for the detection of flour in meat samples, but the immunohistochemical method was better for the detection of wheat protein. ALERT gliadin screening test detected gliadin from 10?mg?kg^?1, while an immunohistochemical method detected wheat protein concentrations from 1?g?kg^?1 and an ELISA method detected wheat protein concentrations from 4?g?kg^?1. ALERT gliadin screening test showed results within 1 day, whilst an ELISA detection method took 2 days, and an immunohistochemical procedure took 5 days at the soonest, all including sample preparation. This study also focused on optimisation of an immunohistochemical method for samples of cooked sausage. In addition, three samples were sufficient for wheat protein detection at a concentration of 1?g?kg^?1 (and greater) with a confidence level greater than 95%.     

Effect of water washing of shark (Scoliodon laticaudus) meat on the properties of proteins with special reference to gelation

The effect of water washing of shark meat on the properties of proteins has been investigated. The contents of low-molecular-weight proteins and urea were reduced significantly with three washing cycles. The gel forming ability showed marked improvement with the number of washing cycles. The dynamic viscoelastic behavior of washed and unwashed meat revealed a structure build-up reaction that was more pronounced in the washed meat. The concentration of myosin heavy chain of washed meat increased as revealed by Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Addition of urea at biological concentration (approximately 250 mM) to the washed meat reduced the gel forming ability significantly as compared to unwashed meat. The emulsion capacity showed an increase with the number of washing cycles.     

Recombinant DNA in meat additives: Specific detection of Roundup Ready™ soybean by nested PCR

Soybean proteins are widely used by the meat industry as technological coadjutor when producing processed products such as emulsified and ground meat products. Since regulations for the use and labeling of GMOs and derived ingredients are in force in Brazil, a PCR‐based method capable of detecting Roundup Ready^? (RR) soybean was employed for meat additives. Thirty‐two samples of meat additives containing soy proteins were tested for the presence of soybean amplifiable DNA and RR soybean DNA. Twenty‐five samples gave a positive signal for the lectin gene, confirming the presence of soybean amplifiable DNA and 15 samples returned a positive signal for specific RR detection confirming the presence of genetically modified soy. These results demonstrate for the first time the presence of RR soybean in meat additives. This method may be useful for meat industries interested in controlling the presence of RR soybean in additives used for meat products manufacture. Copyright © 2007 Society of Chemical Industry     

Detection of casein, soy proteins and coagulated egg-white in meat products by electrophoresis on cellulose acetate membranes

Summary A rapid method has been developed for the detection of casein, soy protein and coagulated egg-white in meat products. Fat and serum proteins are removed by hot water extraction. Casein and soy proteins are extracted with 8 M urea. Coagulated egg-white is subsequently extracted with a fresh mixture of 8 M urea/0.15 M thioglycol (9:1). The proteins are precipitated with an acetate buffer of pH 4.6 which is added in 4-fold excess to the urea extract. After isolation and washing of the precipitate the proteins are dissolved in urea or urea/thioglycol and subjected to electrophoresis on cellulose acetate membranes in a suitable buffer system, viz. barbitone/urea pH 8.5 for casein and formic acid/urea pH 4.0 for soy proteins e.q. egg-white. About 0.1% of casein, soy protein and 0.5% of egg-white added to meat products can be detected.

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Zusammenfassung Es wurde eine schnelle Methode zum Nachweis von Casein, Soja-Protein und coaguliertem Eiklar in Fleischerzeugnissen (Bratwurst usw.) entwickelt. Fett und Serumproteine werden mit heißem Wasser extrahiert; Casein und Soja-proteine mit einer 8m-Harnstoff-Lösung. Coaguliertes Eiklar wird dann mit einer frisch hergestellten Lösung von 8m-Harnstoff und 0,15 m-Thioglycol (9:1) extrahiert. Die Proteine werden mit einer Acetat-Puffer-Lösung von pH 4,6 in 4fachem Überschuß präzipitiert. Nach Isolierung und Waschung des Präcipitats werden die Proteine wieder in Harnstoff oder Harnstoff/Thioglycol gelöst und der Elektrophorese auf Celluloseacetat-Membranfiltern in einem geeigneten Puffersystem unterworfen, d. h. Barbitone/Harnstoff pH 8,5 für Casein und Ameisensäure/Harnstoff pH 4,0 für Soja-proteine und Eiklar. Die untere Nachweißgrenze liegt bei 0,1% für Casein und Soja-Proteine und bei 0,5% für Eiklar.

     

Performance of dairy goats fed soybean meal or meat and bone meal with or without urea during early lactation

Use of meat and bone meal for early lactation rations was studied. In Experiment 1, 18 Alpine goats were used in a 15-wk lactation trial. Isonitrogenous and isoenergetic diets were fed (15% CP and 2.3 Mcal/kg metabolizable energy) containing soybean meal, meat and bone meal with urea, or meat and bone meal without urea. Dry matter intake was 2.21, 2.30, and 2.34 kg/d for does fed soybean meal, meat and bone meal with urea, and meat and bone meal without urea, respectively. Milk production was 2.50, 2.67, and 2.66 kg/d in the same sequence. Rumen ammonia N (mg/dl) and total VFA (mM) were 12.9 and 81.5, 21.4 and 76.3, and 12.2 and 81.6 for does fed soybean meal, meat and bone meal with urea, and meat and bone meal, respectively. Serum urea N was higher in does fed meat and bone meal with urea, and no differences were observed in serum total protein and plasma glucose concentrations. In Experiment 2, 4 mature, castrated male goats were used to estimate digestibilities and retention of nutrients in three diets. Digestibilities of NDF and P and retention of P were higher in goats fed the meat and bone meal diet. Allowance of absorbed protein for milk production was calculated to be 81 to 83 g/kg 4% FCM. Meat and bone meal may be utilized efficiently by lactating does as a protein less degradable in the rumen, Ca, and P source; and may be beneficial for higher milk production during early lactation.     

Easy determination of the addition of soybean proteins to heat-processed meat products prepared with turkey meat or pork-turkey meat blends that could also contain milk proteins

The addition of non-meat proteins to processed meat products is limited by regulations. Therefore, this work has investigated the determination of added soybean proteins in commercial heat-processed meat products prepared with turkey meat or pork-turkey meat blends that could also contain milk proteins. The method consisted of extracting proteins from the meat products in a Tris-HCl buffer (pH 8) and analysing the extract by high-performance liquid chromatography with a linear gradient water-acetonitrile containing 0.05% (v/v) TFA. This method enabled the detection and quantitation of up to 0.08 and 0.28% (w/w), respectively, of soybean proteins (related to 6 g initial product) in these products. Satisfactory precision and recovery data were established. Accuracy was evaluated by a comparison of soybean protein contents determined by the proposed method and the existing AOAC official method based on an enzyme-linked immunosorbent assay (ELISA) from which no statistically significant differences were observed.     

Evaluation of a rapid PCR-based method for the detection of animal material

A rapid PCR-based analytical method for detection of animal-derived materials in complete feed was developed. Using a commercially available DNA forensic kit for the extraction of DNA from animal feed, a sensitive method was developed that was capable of detecting as little as 0.03% bovine meat and bone meal in complete feed in under 8 h of total assay time. The reduction in assay time was accomplished by reducing the DNA extraction time to 2 h and using the simpler cleanup procedure of the kit. Assay sensitivity can be increased to 0.006% by increasing the DNA extraction time to an overnight incubation of approximately 16 h. Examination of dairy feed samples containing either bovine meat and bone meal, porcine meat and bone meal, or lamb meal at a level of 0.1% (wt/wt basis) suggested that this method may be suitable for regulatory uses. The adoption of this commercially available kit for use with animal feeds yields an assay that is quicker and simpler to perform than a previously validated assay for the detection of animal proteins in animal feed.     

Incorporation of Blood Proteins into Sausage

The possibility of introducing beef plasma and decolorized globin proteins as ingredients in meat emulsions of cooked sausages was evaluated. The emulsifying capacity (EC) of plasma proteins was similar to that of meat, but globin proteins showed significantly lower EC values. The latter could be enhanced when plasma proteins were added in proportions ranging from 10 to 65%. Combinations of meat, plasma and globin proteins, containing up to 20% blood proteins, yielded acceptable EC values. These values were affected by the fat content of the meat in the emulsions and especially by the total content of proteins. Up to 12% of plasma protein and different combinations of plasma: globin proteins, containing up to 5% of the latter (12% of total protein replaced), was used in the preparation of cooked sausages. This yielded an acceptable product as ranked by a sensory panel.     

A SYBR Green real-time PCR assay to detect and quantify pork meat in processed poultry meat products

Species identification in meat products has grown in interest in recent years since these foodstuffs are susceptible targets for fraudulent labelling. In this work, a real-time PCR approach based on SYBR Green dye was proposed for the quantitative detection of pork meat in processed meat products. For the development of the method, binary meat mixtures containing known amounts of pork meat in poultry meat were used to obtain a normalised calibration model from 0.1 to 25% with high linear correlation and PCR efficiency. The method revealed high specificity by melting curve analysis, being successfully validated through its application to blind meat mixtures, which confirmed its adequacy for pork meat determination. The fully applicability of the method was further demonstrated in commercial meat products, allowing verification of labelling compliance and identification of meat species in processed foods.     

Investigation of methods to detect mechanically recovered meat in meat products - IV: Immunology

The possibility of using immunological techniques as a method for the detection of mechanically recovered chicken meat in meat products has been investigated in this preliminary study. Antibodies were raised against a low molecular weight fraction (≤ 30 kDa) of chicken bone marrow proteins and an enzyme-linked immunosorbent assay (ELISA) developed. The system was used to test for the presence of mechanically recovered meat (MRM) in a range of product types, from raw chicken meat through to heat processed samples. The results show that it is possible to raise antibodies to chicken bone marrow proteins which show a strong reactivity with chicken and turkey MRM but show little reaction with extracts of MRM and hand deboned meat of other common meat species. However, blood, skin and soya all affected the accuracy of the ELISA.

This study has demonstrated the potential for the use of an immunological procedure as a rapid test for MRM. The selectivity of the antiserum would, however, have to be increased before this procedure could be considered as a suitable technique for the detection of MRM in meat products.     

Production of a horse-specific monoclonal antibody and detection of horse meat in raw meat mixtures by an indirect ELISA

A stable hybridoma cell line (DD3) has been produced secreting a monoclonal antibody specific for horse muscle proteins. The DD3 monoclonal antibody (mAb) did not show significant cross-reactivity when tested against beef, chicken, pig and soya proteins or bovine caseins, gelatine and bovine serum albumin. The DD3 mAb was used in an indirect ELISA format for the detection of defined amounts of horse meat (10–500 g kg-¹) in beef meat mixtures. Immunorecognition of monoclonal antibodies adsorbed to horse meat adsorbed onto the ELISA plate was made with rabbit anti-mouse immunoglobulins conjugated to the enzyme horseradish peroxidase. Subsequent enzymic conversion of the substrate gave clear optical density differences when assaying mixtures of minced beef containing different amounts of horse meat.     

Detection, isolation and enumeration of Yersinia enterocolitica from raw pork

The methods available for the isolation of Yersinia enterocolitica from foods are generally considered to be less than optimal, and methods for estimation of numbers are lacking. Such methods are needed to understand better the significance of foodborne yersiniosis and to provide data for exposure assessment. We describe a method for the detection and enumeration of Y. enterocolitica containing the pYV virulence plasmid (YeP+) in samples from pork surfaces. The method uses a multiplex PCR targeting the ail and virF genes to detect Y. enterocolitica after incubation of surface swabs in Yersinia enrichment broth according to Ossmer. Enumeration was achieved by adapting the enrichment to a most probable number (MPN) method format. A presumptive result was available within 24 h of sample receipt, and YeP+ isolates were confirmed within four days. The presence/absence and MPN methods were evaluated in a pilot survey of 34 packs of raw pork meat purchased from retail outlets in Christchurch, New Zealand. YeP+ was detected by PCR on meat from 32% of the packs, and YeP+ isolates were obtained from 18% of the samples. YeP+ were present at numbers ranging from 0.30 to 5.42 MPN/cm(2). This improved method for the detection and enumeration of YeP+ from meat samples can be used for microbiological surveys to obtain data for assessments of consumer exposure to virulent Y. enterocolitica, and in outbreak investigations.     

Semiquantitative detection of male pork tissue in meat and meat products by PCR

Consumer awareness has increased concerning castration of piglets without analgesia or anaesthesia. On the other hand the occurrence of boar taint is not tolerated by consumers. Currently no reliable methods exist for the on-line detection of boar taint in the slaughterhouse or for genetic sexing of pigs. Therefore, as an alternative the detection of male pork meat was sought. Based on detection of a length polymorphism of the sex chromosomal amelogenin gene a reliable, specific and highly sensitive PCR method for qualitative and semi-quantitative determination of male pork tissue in meat and meat products was determined. A set of 25 male and 25 female meat samples could be correctly identified and mixtures with as little as 0.1% male meat content could be detected. Therefore the method can be used for production and control of specific meat products containing low amounts of male pork meat and thus avoiding boar taint.     

Use of phytate determination for estimation of soya proteins in meat products

Phytates present in vegetable protein preparations were used to measure the degree of substitution of meat proteins by vegetable proteins. In two series of beef blends with soybean protein preparations 6–30% of meat proteins was substituted by soybean proteins. To some blends pickling solutions containing i.a. Na₄P₂O₇ were added. Phytic phosphorus was determined colorimetrically, after extraction with HCl, in raw, pasteurized, and sterilized blends both with and without addition of inorganic phosphate. In blends with no pickling solution the contents of phytic P increased linearly with increasing contents of soybean proteins and the linearity was not influenced by the heat treatment. In blends containing the pickling solution the amount of phytic P increased with increasing heat treatment conditions and with increasing amount of inorganic phosphate added. These relationships reduce the applicability of the method for practical estimation of substitution degree of meat by vegetable proteins.     

Quantitative Estimation of Dark Muscle Content in the Mackerel Meat Paste and its Products Using Antisera Against Myosin Light Chains

ABSTRACT: Based on the high level of extractability of myosin subunits (light chains), even after prolonged heat treatment of muscle, a new method to evaluate the dark muscle content in the fish meat and products of mackerel is proposed. Tissue-specific rabbit antisera with myosin light chains (A1 from ordinary muscle and D1 from dark muscle) from mackerel Scomber japonicus were obtained. Mackerel meat paste (surimi) was dissolved in 8 M urea containing 1% SDS, and diffused on agar plates containing antiserum against A1 or D1 by single radial immunodiffusion (SRID). The results obtained showed that the area of halos formed in the plates was quite proportional to the content of dark muscle.     

Sandwich ELISA for detection of horse meat in raw meat mixtures using antisera to muscle soluble proteins

A double-antibody sandwich ELISA (enzyme-linked immunosorbent assay) has been successfully developed for the detection of defined amounts of horse meat (1-50%) in unheated meat mixtures. The assay uses horse-specific antibodies obtained by immunoadsorption of the crude horse antisera onto immobilised sarcoplasmic extracts from chicken, beef and pig to remove cross-reacting antibodies. The purified antibodies bound to a solid support sequester horse muscle soluble proteins from meat mixtures. Further immunorecognition was made with the same antibodies conjugated to the enzyme horseradish peroxidase. Subsequent enzymic conversion of substrate gave clear optical density differences when assaying mixtures of minced beef and pig containing variable amounts of horse meat.     

Analysis of soyabean proteins in meat products: a review

The use of soyabean proteins as meat extenders has spread significantly due to the interesting nutritional and functional properties that are present in soyabean proteins. Together with these, health and economical reasons are the major causes for the addition of soyabean proteins to meat products. Nevertheless, despite the good properties associated to soyabean proteins, there are many countries in which the addition of these proteins is forbidden or in which the addition of soyabean proteins is allowed up to a certain extent. Thus, the need of analytical methods enabling the detection of added soyabean proteins in meat products is obvious. Microscopic, electrophoretic, immunologic, and chromatographic methods are the most widely used for this purpose. However, the detection of soyabean proteins in meat products presents difficulties related to the composition (meat species, meat quality, soyabean protein source, presence of other non-meat proteins, etc.) and the processing of the meat products, and, although these analytical methods have tried to overcome all these difficulties, there is still not a method enabling quantitative assessment of soyabean proteins in all kinds of meat products.