Enzyme-assisted extraction for the HPLC determination of ochratoxin A in pork and dry-cured ham

The extraction of ochratoxin A from meat products is generally carried out using chlorinated organic solvents, such as chloroform or methyl chloride, acidified with hydrochloric or o-phosphoric acid. In this study, an innovative method was developed to extract ochratoxin A from pork and dry-cured ham samples. The method was based on an enzyme-assisted extraction with pancreatin in phosphate buffer pH 7.5. Pancreatin hydrolyses the proteins, so that ochratoxin A, kept in the ionised form, is easily extracted by the aqueous solution. After purification through an immunoaffinity column, ochratoxin A is determined by HPLC with fluorescence detection. The average recovery values were higher than 90.0% and the relative standard deviations were below 5.5%. The limits of detection and of quantification were 0.06 and 0.12?µg?kg^?1, respectively. A comparison between the new enzyme-assisted extraction and an established chloroform method was carried out on six naturally contaminated samples of pork and on 40 samples of dry-cured ham. Significantly higher (p?<?0.001) values of ochratoxin A were obtained on dry-cured ham samples by the enzyme-assisted method.     

Determination of sodium chloride in pork meat by computed tomography at different voltages

Ground pork samples simulating the widely different chemical composition of hams during dry-cured ham production were produced and scanned by x-ray computed tomography (CT). Chemical composition accounted for most of the variation in CT values (97%). Tube voltage (80, 110, and 130 kV) affected CT value and the effect varied between different types of tissue. Sodium chloride (NaCl) was predicted in the ground samples with average prediction errors (RMSEP) as low as 0.2% to 1.0% NaCl. NaCl was also predicted in small samples of raw to dry-cured ham. When dry and fat ham samples were left out of the models, NaCl was predicted with a high precision (RMSEP 0.2% to 0.4% NaCl, R(2) > 0.99). CT can be used as a valuable, nondestructive tool to analyze distribution of and quantify NaCl in ham during dry-curing.     

原料肉特性对湖南腊肉品质的影响

为比较瘦肉型的杂交商品猪肉和本地猪肉对湖南腊肉品质的影响,实验选取三元杂交商品猪的背最长肌(A)和腹部五花肉(B)、本地宁乡猪的背最长肌(C)和腹部五花肉(D)为原料加工湖南腊肉。测定了加工过程中腊肉理化指标变化和腊肉的颜色、挥发性性成分等。实验结果表明:在腊肉成品中,相同部位原料肉加工的腊肉,杂交商品猪肉的水分含量显著高于宁乡猪加工的腊肉样品(AC,BD),但食盐含量和亚硝酸盐含量呈现相反的变化趋势,即商品杂交猪肉腊肉具有较低的食盐含量和亚硝残留量(AC,BD);4组实验腊肉样品中,L*值均呈下降趋势,两个品种的猪肉中背最长肌的a*值呈现先下降后上升趋势,腹部五花肉的变化规律不明显;样品A、B、C、D分别鉴定出挥发性成分40种、24种、54种、23种。说明了不同原料肉品种和不同部位对湖南腊肉的品质有一定影响。     

不同微生物菌株对干腌肉块挥发性风味化合物的影响

以干腌猪肉块为原料,分别接种表皮葡萄球菌(Staphylococcus epidermidis)、木糖葡萄球菌(Staphylococcus xylosus)和耳氏葡萄球菌(Staphylococcus auricularis)后按火腿加工工艺进行发酵生产干腌肉块。采用固相微萃取(SPME)和气相色谱- 质谱联用(GC-MS)进行挥发性风味化合物的测定,研究不同干腌发酵肉块中挥发性风味化合物的种类和相对含量,以探讨不同微生物发酵剂对干腌发酵肉块中挥发性风味化合物的影响。本实验共鉴定出149 种挥发性风味化合物,包括17 种醛、32 种醇、44 种烷烃、28 种酯、10 种酮、10 种酸、5 种杂环化合物、1 种酚和1种醚。不同微生物菌株对干腌发酵肉块挥发性风味化合物的种类、数量和相对含量都产生了非常明显的影响。对照处理组的发酵肉块中烃类物质的相对含量较高,木糖葡萄球菌处理的发酵肉块中杂环类化合物相对含量较高,木糖葡萄球菌处理的发酵肉块中醛类物质相对含量较高、耳氏葡萄球菌处理的发酵肉块中酯类物质的相对含量较高。     

Comparison of different methods for total lipid quantification in meat and meat products

This study was aimed to evaluate the efficiency of six extraction methods for the quantification of total lipid content in meat and meat products: standard Soxhlet method (with and without previous acid hydrolysis), continuous Soxhlet method (with and without previous acid hydrolysis), and those methods based in the use of a mixture of chloroform and methanol, and described by Folch, Less, and Sloane (1957) and Bligh and Dyer (1959). Lipid content was determined in nine different meat products with different fat contents and physico-chemical features: cooked turkey breast, fresh pork loin, cooked ham, dry-cured ham, mortadella, beef burger, fresh sausage, dry-cured sausage and salami. The most effective methods for determining fat content in the studied meat products were the method described by Folch et al. (1957) and the Soxhlet with previous acid hydrolysis method. The Soxhlet method without previous acid hydrolysis adequately extracted lipids only in those meat products with very high fat content. The use of the method described by Bligh and Dyer (1959) gave rise to the lowest lipid contents in all the studied meat products.     

Occurrence of ochratoxin A in raw ham muscle,salami and dry-cured ham from pigs fed with contaminated diet

Pork meat-derived products can contribute to the overall ochratoxin A intake, either by carry-over effect, or by environmental mould population cross-contamination. In order to assess the role of these different contamination routes, a study was carried out with pigs challenged orally with OTA contaminated feed at subchronical level. After slaughtering, thighs and minced meat from control and treated groups were transformed into dry-cured hams and salami, respectively, which were analysed for OTA determination after ripening. From collected data, the carry-over in muscle was generally low, whereas a significant contribution to the OTA contamination in dry-cured hams was due to toxinogenic mould population growing on their surface during ripening. Finally, a survey of different types of dry-cured ham (n = 110), from the Italian market, was performed, showing the occurrence of OTA on the surface portion in 84 out of 110 samples with a median value of 0.53 μg/kg and in the inner core in 32 out of 110 samples with a median value lower than 0.1 μg/kg.     

The effects of castration on the eating quality of dry-cured ham

The influence of the castration of entire male pigs on the eating quality of dry-cured ham was evaluated. Forty-eight dry-cured hams (435-day aged) were studied from entire and castrated males of two different crossbreeds. The proximate composition of the meat and the androstenone, indole and skatole fat content were determined. The entire hams were classified according to the androstenone and skatole content. Sensory analysis was carried out by a trained panel, evaluating marbling, juiciness, saltiness, graininess, toughness, overall flavour, boar odour and boar flavour. Also carried out was a preference and acceptability paired test by consumers. Castration increased meat fattening and reduced the androstenone and skatole levels of the fat. Significant differences (P<0.05) were found between entires and castrates for the average values given in all the sensory attributes studied. The dry-cured ham from castrates was scored as more flavoured, more marbled and softer. It was also perceived as less grainy, less salty and having less boar odour and flavour. The sensory perception of boar odour was more intense than that of flavour in dry-cured ham and appears to be related to the level of androstenone and skatole in fat. Dry-cured ham from castrated males was also more accepted and more preferred by consumers, especially women and habitual consumers. Castration of male pigs contributes to improve the quality of dry-cured ham. The rejection caused by boar odour and flavour is reduced, improving the overall flavour, texture and juiciness. In addition, the saltiness is less pronounced in ham from castrates. For this reason, the production of high quality dry-cured ham will have to shoulder the extra costs associated with processing castrated pigs.     

High pressure treatment of dry-cured Iberian ham. Effect on radical formation,lipid oxidation and colour

The effect of high-pressure treatment (200, 400, 600 and 800 MPa) on radical formation in solid samples of dry-cured Iberian ham and in ham slurries was evaluated using the spin-trapping technique and electron spin resonance (ESR) spectroscopy. In addition, hexanal was, as a secondary lipid oxidation product, assessed by gas chromatography. The effect of pressure treatments on ESR signals determined in the spin-trapping assay following pressure treatment in solid samples of dry-cured Iberian ham was not statistically significant. However, the lowest level of pressure applied initiated radical formation and promoted lipid peroxidation, whereas intermediate to high levels of pressure seemed to promote further reaction and disappearance of free radicals. Pressure significantly affected hexanal content in the same way as seen for formation of free radicals. Regarding the slurries prepared from dry-cured ham, pressure significantly increased the tendency of radical formation as seen from the ESR signals and significantly increased the hexanal content. The different pattern between solid ham and ham slurries points toward an initiation mechanism associated with the membrane phospholipids for oxidation. In addition, surface colour (L*, a* and b* values and percent reflectance values) of non-pressurized and pressurized (200–800 MPa) dry-cured Iberian ham samples was assessed. Non-pressurized samples showed a higher lightness than pressurized samples, and redness significantly decreased with pressure treatment.     

Effect of irradiation on colour and lipid oxidation of dry-cured hams from free-range reared and intensively reared pigs

The effect of irradiation dose (0, 5 and 10 kGy) of vacuum-packaged Iberian dry-cured ham slices from pigs fed on concentrate (CON) or free-range reared (FRG) was studied in relation to TBA-RS, hexanal content and instrumental colour changes. TBA-RS values increased after irradiation in the two sets of hams and the increase was dose-dependent. FRG samples showed higher TBA-RS values than CON samples in all treatments, although differences in the rate of formation of thiobarbituric acid-reactive substances were different in the two types of hams, being higher in CON samples than in FRG ones. Irradiation of dry-cured ham slices significantly increased hexanal contents in both sets of hams. Irradiation produced statistically significant increases in vacuum-packed dry-cured ham slices lightness (L^⁎-value), yellowness (b^⁎-value) and chroma (C^⁎-value). Irradiation resulted in significantly lower hue angle (h°) values and higher a^⁎-values in both sets of hams indicating a redder colour of irradiated samples than non-irradiated, and these changes were greater in FRG samples than in CON samples. Differences in composition characteristics of raw material could play an important role in the irradiation-induced changes on colour and lipid oxidation of vacuum-packaged dry-cured ham slices.Industrial relevanceIberian ham is dry-cured meat product with a high sensory quality due to pig fattening feature and the characteristics of its processing. Both factors lead to a product with characteristic cured red colour of the lean, an intense, characteristic and pleasant flavour, unique in dry-cured hams, derived from intense lipid oxidative phenomena. New commercialization formats of dry-cured Iberian hams include vacuum-packed sliced ham. Although the low Aw, NaCl and nitrite/nitrate contents make dry-cured ham not adequate for the growth of pathogens and spoilage microorganisms (cocci gram+ and yeast), those may be present on the surface of whole hams and reach the sliced product during the final boning, slicing and packaging operations even with the observation of strict hygienic procedures despite the implementation of modern technologies and HACCP systems, constituting a permanent risk of contamination. Dry-cured Iberian ham, as a result of its chemical characteristics, is a product with a long shelf-life at refrigeration temperature, however the risk of spoilage or pathogen growth could increase in the case of temperature abuse for a long stored period. Finally, due to health concerns, salting and curing process of dry-cured Iberian hams tend to a reduction in levels of NaCl and nitrite used. That could increase the risk associated to microorganism presence in dry-cured sliced hams. The use of irradiation could minimizes this risk, however irradiation could have important effects on overall quality of dry-cured Iberian products in which colour and extent of lipid oxidation and lipid oxidation-derived volatile compounds play an important role in consumer acceptation of the product.     

Consumer satisfaction with dry-cured ham in five European countries

The objective is to investigate consumer satisfaction with dry-cured ham in five European countries. A logistic regression model has been fitted using data collected through a cross-sectional web-based survey carried out in Belgium, Germany, Denmark, Poland and Greece during January 2008 (n=2437 of which 2156 were dry-cured ham consumers). Satisfaction was evaluated as overall satisfaction, as well as specific satisfaction with healthfulness, price, convenience and taste. The findings show that the main determinant of overall satisfaction is taste satisfaction, hence, producers are recommended to focus on matching sensory acceptability of dry-cured ham. No significant between-country differences were found, reflecting the wide availability of this product in all countries. Consumer characteristics influenced their level of satisfaction. Men, older (age > 52 years) and frequent consumers of dry-cured ham consumption were more likely to be satisfied with dry-cured ham. Consumers trust the butcher's advice and they preferred purchasing dry-cured ham at a butcher shop rather than in a supermarket.     

Development of a rapid, specific and efficient procedure for the determination of proteolytic activity in dry-cured ham: definition of a new proteolysis index

A method was adapted to determine proteolytic activity in dry-cured ham using fluorescamine-specific labelling of N-terminal α-amino groups of peptides and amino acids. Fluorescence of the complex was measured using a microplate procedure and optimum excitation and emission wavelengths of 375nm and 475nm, respectively. A new proteolysis index (PI) was defined as the percentage ratio of the N-terminal α-amino group content to the total protein content of the ham extract. The robustness of the method was evaluated by measuring PI in pork meat samples subjected to standardized processing conditions and in samples extracted from industrial hams taken at different stages of processing. For the industrial samples, a comparison with the classic nitrogen procedure of PI determination was performed and a formula relating the two PIs was established. The rapidity, sensitivity and specificity of the procedure make it a good candidate for a screening test to evaluate ham quality in industry.     

Optimization of rapid detection of Escherichia coli O157:H7 and Listeria monocytogenes by PCR and application to field test

For rapid detection of Escherichia coli O157:H7 and Listeria monocytogenes, simple methods for sample preparation and PCR were established and applied to a field test. To improve specificity, primer sets LP43-LP44 and C(+)-D(-) were selected for E. coli O157:H7 and L. monocytogenes, respectively. Through centrifugation and partial heat treatment after enrichment, E. coli O157:H7 and L. monocytogenes were detected at 1 initial CFU without genomic DNA extraction in the culture and with artificially inoculated food samples including milk, chicken, ham, and pork. Based on the optimized PCR method, a feasibility test was carried out using randomly collected field samples. To remove false positives and false negatives, a PCR method using several primer sets, including the optimized primer set, and a standard culture method were used. With the PCR detection and standard culture methods, two pork samples were positive for L. monocytogenes after enrichment, indications that the PCR assay could be effectively used for rapid, sensitive, and species-specific detection of foodborne pathogens.     

Data Mining on MRI-Computational Texture Features to Predict Sensory Characteristics in Ham

In this study, data mining technique was applied on computational texture features obtained from the analysis of magnetic resonance imaging (MRI) of hams, with the main objective of determining sensory attributes of dry-cured ham non-destructively. For that, fresh and dry-cured hams were scanned and then the MRI images were analyzed by three methods of computational texture features. Data mining was applied on the computational texture features from fresh and dry-cured hams for obtaining prediction equations of the sensory attributes of dry-cured hams. The correlation coefficient (R) was used to analyze the results. Accurate prediction was found for 13 sensory attributes as a function of computational texture features of fresh ham, and three from dry-cured ham. In addition, a sensory analysis of dry-cured hams was also carried out to validate the predicted results. Similar values were found between the predicted attributes and those determined by sensory analysis. Thus, it is possible to predict sensory attributes of dry-cured hams by applying data mining on computational texture features of MRI from fresh and dry-cured hams. This supposes the chance of determining non-destructively sensory attributes of dry-cured hams, even before the curing process starts.     

不同来源和年份诺邓火腿的理化和呈味性质

为研究市售常见诺邓火腿的差异,对具有地域特色的诺邓火腿进行综合评价。以洋三元白猪、诺邓黑猪为原材料,按照诺邓火腿工艺加工1、3 年火腿,测定其理化指标、质构特性和游离氨基酸含量,并用电子舌分析其滋味特性差异。结果表明：4 组火腿质地存在明显差异;4 组火腿中均分离出18 种游离氨基酸,精氨酸含量最丰富,亮氨酸、赖氨酸和谷氨酸次之;洋三元白猪火腿（W组）的游离氨基酸含量高于诺邓黑猪火腿（B组）,B组的鲜味氨基酸相对含量高于W组,氨基酸的释放量可明显将1 年和3 年的火腿区分开;电子舌结果表明,B1组的滋味丰富性最大,B3组的咸味最大但鲜味最小,W1组的鲜味较大但丰富性较小,W3组的咸鲜味和丰富性适中。综上,鲜味、丰富性和咸味是诺邓火腿重要的味觉指标,诺邓黑猪火腿滋味较为丰富,B3组硬度、咸味最大,加工过程中要控温控湿。     

Ochratoxin A determination in ham by immunoaffinity clean-up and a quick fluorometric method.

A simple and rapid method for the determination of ochratoxin A (OA) in ham was developed using a basic methanolic extraction, immunoaffinity column clean-up and a fluorometric determination of the toxin contamination levels. A mean recovery of OA from ham samples spiked at levels from 0.7 to 9.7 microg kg(-1) was 83 +/- 6% using the fluorometric method, with a detection limit of 0.7 microg kg(-1). Recovery data were compared statistically with those obtained using reversed-phase high-performance liquid chromatography with acetonitrile-water-acetic acid (99:99:2) as mobile phase and fluorescence detection, commonly used for OA determination in food. A good correlation between the two analytical techniques was obtained. Both methods were successfully applied to 42 ham samples, 21 in the middle of the ripening period (after 6 months from the process beginning) and the other 21 at the end of the maturation, after 12 months. Twenty-seven samples (64%) showed an OA contamination level <1.0 microg kg(-1), the Italian Ministry of Health guideline. The maximum contamination level found was 2.3 microg kg(-1). A good agreement (R(2) = 0.980) between HPLC and fluorometer analysis on naturally contaminated samples was obtained.     

Effect of Penicillium chrysogenum and Debaryomyces hansenii on the volatile compounds during controlled ripening of pork loins

During ripening of meat products such as dry-cured ham, the moulds and yeasts that proliferate on the surface may contribute to flavour development. However, their contribution to volatile components of dry-cured meat products is not known. One strain each of Penicillium chrysogenum and Debaryomyces hansenii, selected from dry-cured ham by their proteolytic activity, were tested to determine their effect on the volatile compounds during ripening. Sterile pork loins were inoculated and ripened for 106 days. Volatile compounds collected with a Solid Phase Micro-Extraction (SPME) fibre were analysed by GC/MS. Inoculation of pork loins with P. chrysogenum lead to a decrease in compounds attributed to lipid oxidation and to an increase of compounds derived from free amino acids. Inoculation with D. hansenii seemed to favour the formation of complex alcohols.     

Contribution of a selected fungal population to the volatile compounds on dry-cured ham

Dry-cured ham is obtained after several months of ripening. Different fungi strive on the surface, including toxigenic molds. Proteolysis and lipolysis by the endogenous and microbial enzymes seem to play a decisive role in the generation of flavor precursors in dry-cured meat products. In addition, fungi show a positive impact on the volatile compounds of ripened pork loins. However, the contribution of the fungal population to flavor formation in dry-cured ham remains unclear. One selected strain each of Penicillium chrysogenum and Debaryomyces hansenii was inoculated as starter cultures on dry-cured ham. Volatile compounds extracted by solid phase micro-extraction technique were analyzed by gas chromatography/mass spectrometry. A trained panel evaluated flavor and texture of fully ripened hams. The wild fungal population on non-inoculated control hams correlates with higher levels of short chain aliphatic carboxylic acids and their esters, branched carbonyls, branched alcohols, and some sulfur compounds, particularly at the outer muscle. Conversely, P. chrysogenum and D. hansenii seem to be responsible for higher levels of long chain aliphatic and branched hydrocarbons, furanones, long chain carboxylic acids and their esters. The very limited impact of P. chrysogenum on pyrazines in inoculated hams can be due to the activity of the yeast. Lower levels for some of the more volatile linear carbonyls at the ham surface suggest an anti-oxidant effect by micro-organisms. The differences in volatile compounds did not show a neat impact on flavor in the sensorial analysis. Nonetheless, inoculated hams got a better overall acceptability, which has to be attributed to their improved texture. The lower toughness of inoculated hams is a direct consequence of an early settling of a highly proteolytic mold. Thus, the use of selected fungi as starter cultures may be useful to obtain high-quality and safe dry-cured ham.     

Phenotypic correlations among quality traits of fresh and dry-cured hams

In this study, fresh and processing quality traits were collected on a total of 312 Country Hams. Phenotypic correlations between traits were estimated and numerous values were significantly different (P<0.05) from zero. Yield was significantly correlated with several fresh pork quality traits measured on the fresh hams, including subjective color (0.34) and lipid percentage (0.32). Some meat color traits were significantly correlated, including associations between the color scores taken on the fresh and cured hams. Correlations between fresh pork quality traits were also determined, with results showing significant correlations between ultimate pH and other pork quality traits. The results indicate that yield can be maximized if hams with good fresh pork quality parameters are used for curing. The correlations estimates are helpful for understanding of the biological mechanisms behind the production of dry-cured hams and can potentially be used to improve the efficiency of the dry-cured ham production chain.