Effects of substrate, water activity, and temperature on growth and verrucosidin production by Penicillium polonicum isolated from dry-cured ham

Penicillium polonicum, a common mold on dry-cured meat products, is able to produce verrucosidin, a potent neurotoxin. The ability of P. polonicum isolated from dry-cured ham to grow and produce verrucosidin from 4 to 40 degrees C at water activities (a(w)) of 0.99, 0.97, and 0.95 on malt extract agar (MEA) and a medium made up with meat extract, peptone, and agar (MPA) was evaluated. Verrucosidin was quantified by high-pressure liquid chromatography and mass spectrometry. P. polonicum was able to grow on MEA and MPA at all the a(w) values tested from 4 to 37 degrees C but not at 40 degrees C. The optimal environmental conditions for growth were 20 degrees C, 0.99 a(w) on MEA and 20 to 25 degrees C, 0.97 a(w) on MPA, but the highest amount of verrucosidin was obtained at 25 degrees C, 0.99 a(w) in both media. No direct correlation between extension of mold growth and verrucosidin production was found. Temperature appears to be the most important factor ruling mycelial growth, whereas verrucosidin accumulation is mostly influenced by a(w). However, analysis of variance of the data showed that there was a complex interaction among all the environmental factors (medium, temperature, and a(w)) that significantly (P < 0.0001) affected growth and verrucosidin production. The reduction of a(w) to intermediates values of 0.95 has a stronger effect on growth on MEA than on MPA. Given that the meat-based medium proved to be an appropriate substrate for the biosynthesis of verrucosidin by P. polonicum, the ability of this mold to produce the toxin on meat products should be established.     

Proteolytic activity of Penicillium chrysogenum and Debaryomyces hansenii during controlled ripening of pork loins

The role of micro-organisms on the ripening process of dry-cured ham, particularly with respect to proteolysis, is not clear. This is partially due to the lack of an adequate system to study changes on a sterile control meat product for long ripening times. Using a meat system based on sterile pork loins ripened under aseptic conditions for 106 days, the contribution to the proteolysis of two micro-organisms isolated from dry-cured ham has been established. Changes were studied by SDS-PAGE of sarcoplasmic and myofibrillar proteins, capillary zone electrophoresis (CZE) of low ionic strength-soluble nitrogen compounds, and HPLC of free amino acids. Debaryomyces hansenii Dh345 did not show any significant proteolytic activity. However, Penicillium chrysogenum Pg222 showed high proteolytic activity on myofibrillar proteins resulting in an increase in soluble nitrogen compounds. For this, P. chrysogenum Pg222 should be considered to be used as starter culture in meat products made using long ripening times.     

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.     

Penicillium populations in dry-cured ham manufacturing plants

Seven ham manufacturing plants were sampled for 1 year to assess the mycoflora present in the air and on hams, with special attention given to potential mycotoxin producers. Temperature and relative humidity were recorded in the ripening rooms. Maturing rooms held hams from 2 to 3 through 6 to 7 ripening months, and aging rooms held hams for the following 6 to 7 months, until the 14-month ripening point, when they were ready for the market. Mean temperatures and relative humidities registered during the study were 14.9 degrees C and 62.4%, respectively, in maturing rooms and 16.3 degrees C and 57.6% in aging rooms. Aspergilli and penicillia, potential mycotoxin producers, were isolated in all the plants from the air and the ham. Aspergilli represented 5% of the isolates, while penicillia were largely dominant, with Penicillium nalgiovense being the most represented species (around 60% of the penicillia), followed by Penicillium nordicum, with 10 and 26% of the penicillia isolated, respectively, from the air or the ham. Ochratoxin A production ability, checked in vitro at 250C, was observed in 50% of the P. nordicum isolates obtained both from the air and the ham. Air and ham surface contamination by penicillia was greater in the ripening rooms, where higher temperatures were registered. A certain correlation was also observed between air and ham surface contamination. On the basis of this study, P. nordicum, the ochratoxin A producer that is notable on proteinaceous substrates, is normally present in ham manufacturing plants in Italy, even though not a dominant species. Further studies are necessary to clarify and ensure if dry-curing conditions minimize the potential risk of ochratoxin A formation in the product.     

Proteolytic activity of lactic acid bacteria strains and fungal biota for potential use as starter cultures in dry-cured ham

During the processing of dry-cured meat products, sarcoplasmic and myofibrillar proteins undergo proteolysis, which has a marked effect on product flavor. Microbial proteolytic activity is due to the action of mostly lactic acid bacteria (LAB) and to a lesser extent micrococci. The proteolytic capacity of molds in various meat products is of interest to meat processors in the Mediterranean area. Eleven LAB and mold strains from different commercial origins were tested for proteolytic activity against pork myosin, with a view to possible use of these strains as starter cultures for Iberian dry-cured ham. Proteolytic activity was tested by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The LAB strains with the highest proteolytic activity were Lactobacillus plantarum (L115), Pediococcus pentosaceus (Saga P TM), and Lactobacillus acidophilus (FARGO 606 TM). The best fungal candidate was Penicillium nalgiovense LEM 50I followed by Penicillium digitatum, Debaryomyces hansenii, and Penicillium chrysogenum.     

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.     

Effect of vaccination against gonadotrophin-releasing factor on growth performance, carcass, meat and fat quality of male Duroc pigs for dry-cured ham production

The aim of this study was to compare production, carcass and meat quality parameters, boar taint compounds and fat composition of green and dry-cured hams, between immunocastrated (IM), surgically castrated (CM) and female (FE) Duroc purebred pigs (n=75, 138.7±8.27kg). Liveweight and fat and muscle thicknesses were measured and average daily gain was calculated during growth. Carcass, meat and fat quality parameters were measured. Immunocastrated grew faster than CM or FE after the second dose of vaccine. IM had the lowest dressing percentage but similar % of ham and carcass lean to FE and CM. The effect of the immunocastration on carcass fatness depended on the location, did not affect fat and meat quality and reduced skatole and androstenone levels. Both in green and dry-cured ham, immunocastration slightly altered FA composition. Thus, Duroc pigs vaccinated with Improvac are suitable for the production of high quality dry-cured ham.     

Influence of aw value and storage temperature on the multiplication and enterotoxin formation of staphylococci in dry-cured raw hams.

Growth and enterotoxin A production in dry-cured raw ham, with different aw and pH values, was investigated for Staphylococcus aureus strain FRJ-100 over a temperature range of 20-35 degrees C when stored for up to 7 days. Enterotoxin production took place at all temperatures tested in the meat of ham that was cured, but not dried (aw 0.95). In dry-cured raw ham with an aw-value of about 0.92, and stored at 20 degrees C, no enterotoxin production was detectable within the 7-day storage period, whereas enterotoxin was produced at the higher temperatures. In ham with an aw of about 0.89, enterotoxin was produced only when stored for 7 days at 35 degrees C. Critical points for S. aureus multiplication and enterotoxin-formation during production and storage of dry-cured raw ham are discussed.     

The mycobiota of three dry-cured meat products from Slovenia

The surface mycobiota of three types of Slovenian dry-cured meat products were isolated from a total of 75 items of product that were sampled periodically during the drying/ripening stage of processing. The predominant filamentous fungal genus isolated was Penicillium. Eurotium spp., Aspergillus versicolor and Cladosporium spp. were isolated from only two of the products. Eight Penicillium species were identified. Penicillium nordicum was recovered frequently. Penicillium nalgiovense was recovered less frequently, from one product only (a salami), while a yet-to-be described species Penicillium “milanense” was isolated from 21 items. The other penicillia were rarely isolated. Of the isolated and identified species, those that can produce mycotoxins are: A. versicolor, Penicillium brevicompactum, Penicillium chrysogenum, P. nordicum, and Penicillium polonicum. Their growth on dry-cured meat products is undesirable.     

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.     

Mycological examination of domestic unpolished rice and mycotoxin production by isolated Penicillium islandicum

Fungi growing on domestic rice were examined from April to June, 2003. One hundred samples of rice, which had been harvested in the autumn of 2002, were collected from the local market, and 15 samples of stored rice, which had been harvested in 2001 and stored in warehouses under government control, were used as samples. From each sample, 50 grains (100 grains in total) were plated on potato-dextrose agar (PDA) and malt yeast 40% sucrose agar (M40YA) containing chloramphenicol after being washed with sterile distilled water to remove any microorganisms on the surface, and incubated at 25 degrees C for a week. For most of the rice samples harvested in the preceding year, the proportion of grains infected with fungi was less than 20% of the total grains tested. In about half the samples of rice stored for one and half years, more than 80% of the grains were infected with fungi that grew on M40YA. The major genera of fungi isolated from the rice harvested in the preceding year were Penicillium and Alternaria, and those from the rice stored for one and a half years were Aspergillus, Penicillium and Eurotium. P. islandicum, A. versicolor, A. ochraceus and others were isolated as possible mycotoxin-producers in the mycoflora of domestic rice. P. islandicum was isolated from 3 samples, and 82% of the grains were infected with this fungus in one sample. All three isolates from these samples appeared to produce luteoskyrin on Czapek yeast extract agar, based on TLC and HPLC analysis.     

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.     

Influence of loading method and stocking density during transport on meat and dry-cured ham quality in pigs with different halothane genotypes

The effect of loading method and stocking density in transit on meat and dry-cured ham quality was investigated in pigs with different halothane genotypes. A total of 507 Italian heavy pigs, supplied by two farms, were loaded by ramp or lift and transported unmixed for 35–55 min to the abattoir at a stocking density of either <0.4 or >0.6m² per 100 kg pigs. After overnight lairage in separate pens with free access to water, the pigs were slaughtered. Halothane genotype was assessed post mortem. Four hundred and thirty-nine pigs had a homozygous dominant (NN) genotype and 68 pigs were heterozygous (Nn). Carcass skin damage, meat quality traits and ham curing parameters were evaluated. Loading method and stocking density showed a negligible effect on meat and dry-cured ham quality while the predominant factor affecting these was the halothane genotype. Nn pigs produced meat with a faster rate of pH fall and lower water holding capacity as well as ham with higher weight losses in salting and greater incidence of defects in the dry-cured product. There were insignificant interactions between halothane genotype and loading method or stocking density. Overall, irrespective of pre-slaughter treatment, the Nn pigs were less suitable for the production of high quality products such as dry-cured ham.     

Assay of Cathepsin D activity in fresh pork muscle and dry-cured ham

Different assays of cathepsin D activity in both porcine muscle and dry-cured ham extracts have been tested in order to find the best conditions for a reliable detection of cathepsin D in dry-cured meat products. The enzyme was effectively extracted with 0·2% (v/v) of Triton X-100.

Nucleases if present were not observed to interfere with the assays. The best conditions for a reliable detection of cathepsin D in dry-cured ham were found to be the incubation of the enzyme extract for 1 h at 45°C in a reaction mixture containing 0·60% (w/v) of haemoglobin in 0·2 sodium citrate buffer, pH = 3·7.     

Effect of pressure and holding time on colour,protein and lipid oxidation of sliced dry-cured Iberian ham and loin during refrigerated storage

The effect of high pressure (HP) treatments (200 MPa 15 min, 200 MPa 30 min, 300 MPa 15 min, 300 MPa 30 min) on colour, lipid and protein oxidation in sliced vacuum-packed dry-cured Iberian ham and loin during refrigerated storage (90 days, + 4 °C) was evaluated. Pressure level and holding time increased the extent of lipid oxidation in both products. Dry-cured ham showed a higher susceptibility to lipid oxidation than dry-cured loin since HP treatment increased TBA-RS values in dry-cured ham samples while HP treatment decreased TBA-RS values in dry-cured loin samples. However, HP treatment did not affect protein oxidation in both meat products. On the other hand, HP treatment affected instrumental colour since non-pressurized dry-cured meat products showed higher redness than pressurized ones. Regarding changes under storage, after 90 days of refrigerated storage lipid and protein oxidation increased while redness decreased in both HP treated and non-treated dry-cured meat products. Changes induced by HP were only noticeable after HP treatment, as storage reduced the initial differences between HP treated and non-treated samples. Therefore, the lack of differences in long stored dry-cured ham and loin HP treated and non-treated indicates that the application of HP (200–300 MPa/15–30 min) could not affect the quality of dry-cured meat products.Industrial relevanceDry-cured meat products are the meat-based products with the highest sensory quality in Spain and have a high projection in exterior markets. High pressure processing is effective in controlling pathogen and spoilage microorganisms in meat and meat products although it can promote color and oxidation changes that modify sensory characteristics. The study aimed the evaluation of pressure and holding time on color changes and protein and lipid oxidation at vacuum packed slices of Iberian dry-cured ham and loin during subsequent extended chilled storage. High pressure treatment of dry-cured Iberian ham and loin induce changes after treatment although initial differences are not maintained along refrigerated storage.     

Bind, Sensory and Chemical Properties of Restructured Dry-cured Hams

Restructured, boneless, dry-cured ham product was processed from intact, bone-in, dry-cured hams (DC). Processing variables included tumbling and addition of ground, dry-cured ham meat. Restructured treatments were more tender (P<0.05), less salty (P<0.01), and had less off-flavors (P<0.05) with similar country ham flavor compared to the DC control. Addition of ground meat decreased tenderness (P<0.05) and increased Instron binding values. Tumbling decreased off flavors (P<0.05). Sensory bind scores of restructured hams were greater (P<0.05) than fresh, restructured controls (FR); however, Instron binding values were higher (P<0.001) for FR. An acceptable restructured product could be made from traditionally processed dry-cured hams.     

Lipolytic and proteolytic properties of dry-cured boneless hams ripened in modified atmospheres

We studied proteolytic and lipolytic properties of dry-cured boneless ham (porcine quadriceps femoris) made with chilled (10°C, 48 h) or frozen/thawed meat (frozen at -20°C frozen for 90 days and followed by thawing at 10°C for 48 h) were determined. Dry-cured meats were stored in modified atmosphere packages (100% N(2) and a mixture of 75% N(2)+25% CO(2)) at 15°C with the intention of reducing ripening space. Results showed that dry-cured hams made with frozen/thawed raw meat had more salt, volatile fatty acids and free fatty acid content after salting and smoking. Whereas, samples prepared with chilled meats contained more nitrogenous compounds (water-soluble nitrogen, non-protein nitrogen, and free amino acids). Volatile and free fatty acid contents in all samples significantly increased with storage. Acetic acid was the predominant volatile fatty acid. To confirm lipolytic activity in dry-cured ham stored in modified atmospheres, we calculated the lipolytic coefficient. The lipolytic coefficients of all samples were positive values and significantly (P<0.05) increased with storage indicating lipolysis in samples were still active. Furthermore, nitrogenous compounds in dry-cured ham significantly (P<0.05) increased with storage indicating proteolysis in samples were not affected by modified atmosphere storage. Aerobic, anaerobic and lactic acid bacteria counts in dry-cured meats were stable to modified atmospheres storage for 20 weeks at 15°C. Flavor, texture and color score in sensory evaluation for dry-cured ham made with chilled meat were significantly higher than that made with frozen/thawed meat. All samples had high overall acceptance scores in sensory evaluation. Results in this study suggested that dry-cured boneless ham stored in modified atmospheres for 20 weeks at 15°C was another feasibility to ripen the meat without affecting lipolysis, proteolysis, microbiology and sensory quality.     

Advances in the ultrasound characterization of dry-cured meat products

In this work, the feasibility of using non-contact ultrasonic techniques (air-coupled and scanning acoustic microscopy, SAM) for characterizing different dry-cured meat products was assessed. Air-coupled ultrasonic measurements were performed on vacuum packaged sliced dry-cured ham, and compared with contact measurements. The average ultrasonic velocity in dry-cured ham was 1846 ± 49 m/s and 1842 ± 42 m/s for air-coupled and contact measurements, respectively. The deviation (1% relative error) between both techniques was related to the influence of the heterogeneous structure and composition of dry-cured ham and the transducer focusing. The SAM was used to characterize dry-cured ham and chorizo samples. B-scan images for dry-cured ham and chorizo showed two dominant reflections from the sample, linked to reflections in the lean and fatty tissues. The results indicate that contact ultrasonic measurements could be replaced by the air-coupled technique, reducing the measuring time and the material handling. On the other hand, SAM technique allows the microscopic characterization of dry-cured meat products.     

云南干腌火腿加工工艺及其品质影响因素研究进展

云南干腌火腿是一种重要的传统发酵肉制品,以猪后腿为原料,经腌制、风干、发酵等加工而成。云南干腌火腿主要有宣威火腿、撒坝火腿、三川火腿、老窝火腿、诺邓火腿及鹤庆火腿等,本文对云南干腌火腿加工工艺及干腌火腿品质影响因素进行综述,从原料肉、腌制条件、色泽及风味等几方面做了详细讨论,旨在为干腌火腿品质改进提供一定的理论参考。     

A comparative study of boar taint in cooked and dry-cured meat

A comparative study of boar taint in cooked and dry-cured pig meat products was performed. Forty-eight cooked loins and 48 dry-cured hams from entire males and castrates were studied. The samples were classified according to the androstenone (AN) and skatole (SK) fat content determined using HPLC. A trained sensory panel evaluated taste, aroma, boar odour, boar flavour, juiciness, tenderness and fatness in cooked and dry-cured samples. Threshold levels of AN and SK to separate (P<0.05) entire and castrate samples according to their boar odour and flavour were determined. The effects of castration and processing on the eating quality were studied. Finally, the relationships among boar taint and other sensory attributes in cooked and dry-cured meat were compared. Results from these studies show that the eating quality of processed meat was affected negatively by boar taint. Threshold values were higher in dry-cured ham (2 μg g(-1) AN and 0.12 SK) than cooked loin (0.5 AN and 0.1 SK). AN and SK had a synergistic effect on boar odour and flavour in both products. AN had a greater influence than SK on the aroma and taste, especially in cooked meat. Boar odour was perceived more intensely than boar flavour in both of the products studied. Castration favoured fatness and improved the aroma and taste of cooked and dry-cured meat. The loss of aroma and taste due to boar taint was more noticeable in cooking than drying and curing. In dry-cured meat boar taint was associated with less aroma, taste, juiciness and tenderness. However, in cooked meat, boar taint affected the aroma and taste more strongly, but was not related with juiciness and tenderness, probably because these attributes are influenced by cooking.