Ripening Of Sausages Research Articles

Effectiveness of a Lactobacillus sakei Starter Culture in the Reduction of Biogenic Amine Accumulation as a Function of the Raw Material Quality

The effectiveness of an amine-negative starter culture (Lactobacillus sakei CTC494) in the reduction of biogenic amine production during the ripening of fermented sausages was examined. Four batches were manufactured in parallel:spontaneously fermented and starter-mediated sausages were manufactured from two lots of raw materials of different hygienic quality. Besides the biogenic amine contents, changes in the microbial counts, nitrogenous fractions, pH, and water content were measured at several sampling points during the ripening process. In sausages manufactured from good quality meat, the starter strain of L. sakei reduced and even inhibited biogenic amine accumulation during sausage fermentation, the end products showing extremely low biogenic amine contents (tyramine levels less than 15 mg/kg of dry matter and putrescine and cadaverine levels less than 5 mg/kg of dry matter). Nevertheless, starter-mediated sausages made from poorer-quality raw materials showed much higher amine contents (308, 223, and 36 mg/kg of dry matter of cadaverine, tyramine, and putrescine, respectively), which were only slightly lower than those of the spontaneously fermented sausages made from the same raw materials. The relatively high bacterial numbers of raw materials of poorer-hygienic quality diminished the beneficial effect of the starter strain. Therefore, the effectiveness of the starter was strongly dependent on the hygienic quality of the raw materials used.

Influence of Hygienic Quality of Raw Materials on Biogenic Amine Production during Ripening and Storage of Dry Fermented Sausages

The effect of the hygienic status of raw materials on biogenic amine production during ripening and storage (at 4 and 15°C) of fermented sausages was studied. Two portions of fresh lean and back fat pork were stored for 5 days at −20°C (treatment A) and at 4°C (treatment B), respectively. Raw materials of treatment A maintained their hygienic quality high and low amine content. Raw materials of treatment B showed from 1 to 3 log (CFU/g) higher microbial counts and a biogenic amine index near 50 mg/kg, indicating poorer hygienic quality. The quality of raw materials influenced the composition and the concentration of biogenic amines produced during the ripening sausages. Sausages of treatment A (A-sausages) showed a large accumulation of tyramine (up to 100 mg/kg dm) followed by putrescine and cadaverine (<15 mg/kg). In contrast, B-sausages resulted in earlier and much greater amine production, and cadaverine, tyramine, and putrescine levels were 50-, 2.6-, and 6.5-fold higher than those of A-sausages. Other biogenic amines, such as octopamine, tryptamine, phenylethylamine, and histamine were also produced in B-sausages. The higher proteolysis and the lower pH of B-sausages might have favored the decarboxylase activity of microorganisms. Biogenic amine contents of sausages during storage depended on the raw materials used and storage temperature. No significant modification on the amine contents was observed during the storage of A-sausages at either temperature. Greater changes occurred in B-sausages stored at 15°C than in those stored at 4°C. Higher temperatures favored proteolytic and decarboxylase reactions, resulting in increased amine concentrations after storage.

Microbiological events during the elaboration of "fuet", a Spanish ripened sausage

The development of different microbiological groups, most of them reported previously as histamine- or tyramine-forming in foods, as well as pH and water activity (aw) were studied during the elaboration of "fuet", one of the most popular and consumed Spanish ripened sausages. Lactic acid bacteria became the predominant microorganisms after the first 4 days of ripening. Enterococci counts showed a significant statistical relationship with the counts of tyrosine-decarboxylase-containing bacteria during the first days of ripening, but they could not explain the tyramine-forming activity detected during the later stages of the process. The number of enterobacteria and pseudomonads decreased steadily during the production process, disappearing after the ripening period. Both showed a statistically significant relationship with histidine-decarboxylase-containing bacteria (HDB) counts. No histamine was formed during the elaboration process. However, HDB were still detectable on the last day of our study. Although no statistically significant relationships were found between pH development and the decrease in Gram-negative and HDB, a highly significant statistical relationship was found between their development and the decrease in water activity (aw).

Changes in the Components of Dry-Fermented Sausages during Ripening

Several chemical changes occur during the ripening of dry-fermented sausages that determine the flavor and odor of the end product. The phenomena that take place during fermentation, that is, both acidification of the sugars by lactic acid bacteria and reduction of nitrates and nitrites to nitric oxide by micrococci have been known for several years. However, the chemical changes involved in this process, and, particularly, the agents responsible have not yet been established, although they have been attributed to changes in the majority components (proteins and lipids) and to the ingredients added (spices and condiments) in the preparation of the original mixture.The typical flavor and odor of dry-fermented sausages cannot be attributed to volatile substances alone, but to a large number of volatile and nonvolatile compounds present in the product in suitable proportions. Microbial growth in the sausage together with activity of the meat endogenous enzymes are undoubtedly partially responsible for the development of a number of aromatic and sapid compounds. However, lipid autooxidation reactions are also an important source of these substances, and it is not yet known which of these processes is more important in sausage ripening. Much research has focused on the break up of triglycerides into free fatty acids, diglycerides, and monoglycerides during ripening and the progressive increase in the amounts of different carbonyl oxidation products. Carbonyl compounds probably play a significant role in determining the flavor because, in general, these have very low perception thresholds, in the ppm and ppb range. Similarly, the protein breakdown to yield peptides and amino acids has been studied extensively, the latter being substrates of several microbial and chemical reactions that generate many flavor compounds.

Influence of Meat Processing and Meat Starter Microorganisms on the Degradation of Organochlorine Contaminants

The influence of processing of different dry-cured sausages on the degradation of organochlorine pesticide residues such as hexachlorobenzene (HCB), α-, β-, γ-hexachlorocyclohexane (HCH), and p,p'-DDE was investigated. The residual contamination was analyzed at different stages of the sausage's ripening by capillary gas-liquid chromatography with electron capture detector. Furthermore, the degradative capability of the microorganisms used as starters in the manufacture of such meat products (Pediococcus acidilactici, Pediococcus pentosaceus, and Micrococcus varians) was studied in vitro. The M. varians strain was able to degrade some of the compounds tested. Significant reductions (p 0.05).

Tyramine degradation by micrococci during ripening of fermented sausage

The ability of tyramine oxidase exhibiting strains of Micrococcus varians to degrade tyramine in vivo during sausage fermentation was investigated. Fermented sausage was produced with a tyramine forming strain of Lactobacillus curvatus which acquired a final tyramine concentration of 190ppm. The addition of either one of two strains of M. varians exhibiting a potential to oxidise tyramine decreased the amount of tyramine formed to 160 and 150ppm, respectively. No effect on growth of L. curvatus and pH development in the fermented sausage was observed when micrococci were present during a three weeks ripening period. Sausage fermentation was further carried out with a non-amine forming strain of L. sake and M. varians after addition of 100ppm tyramine to the raw material. The amount of tyramine in the end product was 60ppm, and during the first 10 days of ripening the decrease was faster on the outside of the sausage.

Application of liquid chromatography‐atmospheric pressure chemical ionization mass spectrometry and tandem mass spectrometry to the determination of volatile nitrosamines in dry sausages

A high‐performance liquid chromatographic‐atmospheric pressure chemical ionization mass spectrometric method was developed for the determination of volatile nitrosamines in dry sausages. Tandem mass spectrometry was applied for the detection of N‐nitrosopyrrolidine N‐nitrosodiethylamine and N‐nitrosopiperidine. N‐nitrosomethylamine was detected by using the selected ion monitoring mode. The occurrence of the four different nitrosamines was monitored in 27 dry sausage samples and a correlation was observed between N‐nitrosopyrrolidine and biogenic amines. Nitroso compounds are thus not only formed in heated conditions but formation can also occur during ripening of dry sausages by reaction between residual nitrite and amines formed during the fermentation process.

Dry fermented sausages

Two major aspects of fermented sausages are discussed in the article. After a brief history of enterohemorrhagic Escherichia coli with special regard to fermented meat products the basic experimental designs and results are tackled on the basis of which safety issue in this respect is thoroughly discussed. The results show a rather limited opportunity for meeting food safety requirements with short or medium time ripened raw fermented sausages while traditionally, long ripened sausages are in a much better position. Fermented meat products as probiotics are also discussed. After evolving the EHEC outbreaks caused by fermented sausages the possibilities for finding reliable methods and products have been narrowed and more research is needed to find optimum solution.

Evaluation of three decarboxylating agar media to detect histamine and tyramine-producing bacteria in ripened sausages.

Histidine- and tyrosine-decarboxylase activity of 175 strains of bacteria isolated from eight retail samples of Spanish ripened sausages was tested in three decarboxylating agars (Niven medium, Joosten and Northolt medium and modified decarboxylating agar of Maijala) and confirmed by an enzymic method (histamine) and thin-layer chromatography (tyramine). Enterobacteria and pseudomonads showed the highest percentage of positive responses to histamine and tyramine in the three decarboxylating agars, but only enterobacteria were subsequently confirmed as histamine-producing. Confirmed tyramine-producing strains were all identified as enterococci or lactic acid bacteria. The medium described by Joosten and Northolt was more sensitive and faster at detecting tyramine-producing microorganisms. However, all three media failed to detect one histamine-positive strain of lactic acid bacteria used as a control.

The importance of meat enzymes in ripening and flavour generation in dry fermented sausages. First results of a European project

The importance of bacterial and meat enzymes in lipolysis and proteolysis was evaluated during dry sausage ripening. The data suggest that lipolysis is to a great extent brought about by muscle and fat tissue. Polyunsaturated fatty acids are liberated from the polar lipid fraction and their specific liberation is higher than for monounsaturated and saturated fatty acids. Lipolysis seemed to be more pronounced in sausages with pork than with beef. Initial protein degradation seemed to be originating from cathepsin D like muscle enzymes. In a later stage, bacterial enzymes become more important in further degradation of the protein fragments formed.

Effect of nitrate and nitrite curing salts on microbial changes and sensory quality of rapid ripened sausages

The effect of the use of either nitrite or nitrate curing salts on microbial changes and sensory quality in rapid ripened sausages inoculated with a mixed starter culture ( Lactobacillus sake and Staphylococcus carnosus) was investigated. Lactic acid bacteria and Micrococcaceae were not greatly affected by the added curing salt. Conversely, the inhibition exerted by nitrite on the undesirable flora ( Enterobacteriaceae and psychrotrophs) was evident from the early stages of the processing keeping highly significant differences (P<0.01) with respect to nitrate made sausages till the end of the ripening stage. The use of nitrite in sausage processing was found to reduce hygienic risks.

OCCURRENCE OF TYRAMINE PRODUCING MICROORGANISMS IN “SALCHICHON” AND TYRAMINE PRODUCTION IN SAUSAGES INOCULATED WITH A TYRAMINE PRODUCING STRAIN OF LACTOBACILLUS BREVIS

Tyrosine decarboxylase activity of different isolates obtained from 20 samples of raw (before the ripening process) and 10 of ripened “salchichón”, a cured sausage produced in Spain, was evaluated. Although 33% of the isolates obtained from raw sausages showed a positive response in the decarboxylation medium described by Joosten and Northolt (JNM), only 8.4% were subsequently confirmed as tyramine formers by thin layer chromatography (TLC). Pseudomonads and enterobacteria showed the highest percentage of positive isolates in JNM, but none was confirmed by TLC. In contrast, 100% of positive enterococci were confirmed. Most of confirmed isolates obtained from raw sausages were identified as Enterococcus faecalis while Enterococcus faecium and Lactobacillus curvatus showed the highest prevalence in ripened sausages. Inoculation with a tyramine forming strain of Lactobacillus brevis, isolated from raw sausage, did not increase the amounts of tyramine formed during the production process of the sausage.

Histidine Decarboxylase Activity of Enterobacter cloacae S15/19 during the Production of Ripened Sausages and Its Influence on the Formation of Cadaverine

The histidine decarboxylase activity of Enterobacter cloacae S15/19 was studied during the production process of salchichón, a Spanish ripened sausage. Counts of fecal coliform and histidine decarboxylase bacteria decreased during the production process, showing a good correlation in both inoculated and control samples. In the samples inoculated with Enterobacter cloacae S15/19, fecal coliforms were undetectable the last day of the survey, while the population of histidine decarboxylase bacteria was over 2 log MPN/g. Despite the fact that inoculation with Enterobacter cloacae S15/19 increased histidine decarboxylase bacteria counts, no differences were observed in the histamine concentration reached, which was undetectable in most of the control and inoculated samples. In contrast, cadaverine concentration increased significantly (P < 0.01) in the inoculated samples, suggesting that cadaverine could be used as a hygienic-quality indicator of the raw materials employed in sausage processing.

Bacterial Proteinase Reduces Maturation Time of Dry Fermented Sausages

ABSTRACTThe effect of proteinases, from Lactobacillus paracasei subsp. paracasei NCDO151 and from Bacillus licheniformis (Alcalase, NOVO Nordisk), on the ripening of dry sausages was compared. Increased levels of starter bacteria, d‐lactic acid, a sharper pH‐drop and changes in chromato‐graphic profiles (by gas chromatography/mass spectrometry) indicated that the enzyme from Lactobacillus paracasei accelerated the dry sausage ripening. This effect was not detected in sausages with the Alcalase enzyme. Higher sensory scores on maturity flavor, color, hardness and others, confirmed that sausages containing proteinase from L. paracasei showed the same maturity after 14 days as those without enzymes or with the Alcalase enzyme after 28 days.

Lipolysis in a Belgian sausage: Relative importance of endogenous and bacterial enzymes

The importance of bacterial and meat enzymes in lipolysis and carbonyl formation was evaluated during dry sausage ripening. Sausages were prepared with and without addition of an antibiotic-antimicotic mixture. In some experiments, an extra inoculum of Micrococcaceae was added and in two experiments, glucose was omitted. Total viable bacterial counts after 21 days were lowered by at least 2 log units in the presence of antibiotics. Free fatty acid productions after 3 and 21 days, in the presence of antibiotics were not significantly lower than observed in the control sausages. Total carbonyl compounds (benzidine reaction compounds) were significantly lowered by the presence of antibiotics compared to the control sausages except when glucose was omitted from the recipe. The data suggest that lipolysis is almost exclusively brought about by muscle and fat tissue. Polyunsaturated fatty acids are liberated from the polar lipid fraction and their specific liberation is higher than for monounsaturated and saturated fatty acids. Carbonyl production from lipids seems to be independent of bacterial activity.

Histidine Decarboxylase Activity of Bacteria Isolated from Raw and Ripened Salchichón, a Spanish Cured Sausage

Twenty samples of raw sausages (before the ripening process) and 10 of ripened sausages of two different sizes of salchichón, one of the most-consumed ripened meat products in Spain, were microbiologically analyzed. Histidine decarboxylase activity of different isolates obtained from different culture media was evaluated by a decarboxylase agar medium and confirmed afterwards by an enzymatic method. Of the isolates obtained from raw sausages, 15.8% showed histamine-forming activity, most of them (78.1%) belonging to the Enterobacteriaceae family. Klebsiella oxytoca, Enterobacter aerogenes (one isolate), and Enterobacter cloacae (six isolates) showed a powerful histamine-forming activity, producing more than 3,000 μg/ml. In spite of histamine-forming activity being detected in eight of ten analyzed samples of ripened sausages, only one histamine-forming bacterium, identified as Lactobacillus curvatus, was isolated; it showed an ability to form 1,994 μg/ml of histamine in experimental conditions.

Formation of Biogenic Amines during Ripening of Dry Sausages as Affected by Starter Culture and Thawing Time of Raw Materials

ABSTRACTRaw materials affect formation of biogenic amines in dry sausages. Effects of thawing time of raw materials and amine‐negative starter culture on amine formation were studied on a pilot scale. The levels of biogenic amines, precursor amino acids, pH, water activity, and microbial counts were measured. Use of starter culture significantly decreased levels of histamine, tyramine and cadaverine formed. The effect of thawing time on formation of biogenic amines was dependent on the use of starter culture.

Effect of the addition of pancreatic lipase on the ripening of dry-fermented sausages — Part 1. Microbial, physico-chemical and lipolytic changes

The effect of the addition of nine different amounts (3–500 units) of pancreatic lipase on the microbial and physico-chemical parameters and lipid fractions during the ripening of dry fermented sausages has been studied. No differences between conventional and lipase-added sausages were found for pH, dry matter and water activity. The addition of lipase caused a greater accumulation of products resulting from the triglyceride breakdown, mainly diglycerides and free fatty acids (FFA). The maximum rate of lipolysis was observed during the first week of the ripening process, specially in the fermentation phase. The greater the pancreatic lipase added, the higher lipolysis observed. At the end of the ripening, the levels of total FFA were clearly higher (1·5 to 5-fold) in all lipase-added batches than in the controls. This fact gave rise to the accumulation of a great amount of FFA, which can contribute either by themselves to the flavour of the sausage or can be available as substrates for further transformations which may generate other flavour compounds.

Effect of the addition of pancreatic lipase on the ripening of dry-fermented sausages — Part 2. Free fatty acids, short-chain fatty acids, carbonyls and sensory quality

The effect of the addition of nine different amounts ( 3–500 units) of pancreatic lipase on the composition of free fatty acids (FFA), short-chain fatty acids and carbonyls, as well as the sensory quality, has been studied on dry fermented sausages. The lipase produced a greater release of all fatty acids in relation to the control sausages. The greater the pancreatic lipase content, the higher the release of these fatty acids, the most important ones being myristic, palmitoleic and oleic acids. A lower release of linoleic acid was observed, probably because of its oxidative degradation. At the end of the ripening in all lipase-added batches, a clear increase of the carbonyl content was noticed in relation to conventional sausages. No consistent changes were observed in the short-chain fatty acid fraction. In the sensory evaluation, the highest significant differences ( P < 0·01) between control sausages and lipase-added batches were observed when 60 and 90 lipase units were used. According to the chemical and sensory analyses, it can be concluded that the addition of 60 and 90 units to sausages seems to be useful to enhanced the flavour of these products.

Partial replacement of sodium chloride with potassium chloride in dry fermented sausages: Influence on carbohydrate fermentation and the nitrosation process

Two types of dry fermented sausages were manufactured: type A with traditional formulation and type B with reduced sodium content. pH and a w values and the counts of aerobic mesophiles, Enterobacteriaceae and lactobacilli micro-organisms during the ripening of both fermented sausages were similar. Final lactic acid contents were also similar, but carbohydrate content was significantly lower in type B sausage. Content of acetic, propionic and butyric acids were significantly higher in type B, suggesting a higher heterofermentative activity. Also type B sausage showed lower nitrate level and higher chemical conversion percentage, suggesting a higher nitrosation intensity.