Lipid Fractions of Dry Fermented Sausages Change when Starter Culture and/or Aspergillus Lipase are Added

Six levels of Aspergillus sp. lipase were evaluated and 0.400u/g was selected. A sausage with a starter culture (Lactobacillus plantarum (10%)+Staphylococcus carnosus (90%)), one with lipase and one with both starter culture and lipase were produced in a pilot plant. A lower amount of Micrococcaceae was found in sausages without starter culture. Total FFA quantity at the 15th day was higher in sausages with lipase. Higher amounts of acetic, propionic and butyric acids were found in sausages with both lipase and starter culture. Odor intensity had slightly higher scores in sausages with added lipase.     

Addition of lipase from Candida cylindracea to a traditional formulation of a dry fermented sausage

The objective of this work was to study the manufacture of sausage containing a traditional starter culture (Lactobacillus plantarum and Staphylococcus carnosus) together with an enzyme lipase from Candida cylindracea as compared with that of a sausage with only starter. The acidity value showed more intense lipolysis in the sausage with lipase with this increase being especially important in the second week of drying. In spite of this, there was no significant (p > 0.05) increase in the oxidative rancidity processes in this sausage. The analysis of short chain fatty acids suggested the enzyme and starter together produced a greater amount of such acids than did the enzyme or the starter separately. Almost all free fatty acids showed significantly higher values in the sausage with lipase with the exception of linolenic acid. The addition of enzyme lipase produced a higher proportion of free saturated acids and a lower proportion of polyunsaturated acids during the drying of the sausage.     

浓缩型冻干发酵剂在鸭肉发酵香肠中的应用

将浓缩型冻干发酵剂应用于鸭肉发酵香肠中,对比液体发酵剂和自然发酵对香肠发酵过程中的影响。结果表明：接种冻干发酵剂的香肠48 h内pH值降到5.3以下,能够快速发酵产酸达到发酵香肠安全控制范围,且水分含量呈下降趋势,发酵末期降到30%左右;对各处理组鸭肉香肠的质构进行分析比较,接种冻干发酵剂处理组（S2处理组）以及液体发酵剂（L2处理组）的香肠硬度、弹性和咀嚼性均显著（P＜0.05）高于自然发酵（对照组）;此外,香肠发酵过程中酸价呈上升趋势,过氧化值先上升后下降,各处理组差异不显著（P＞0.05）,推断内源性脂酶比微生物发酵剂产生的脂酶发挥更大作用;各类型脂肪酸占总游离脂肪酸的比例大小顺序为：饱和脂肪酸＞单不饱和脂肪酸＞多不饱和脂肪酸,接种发酵剂的处理组游离脂肪酸总量显著高于对照组（P＜0.05）,表明发酵剂的添加对游离脂肪酸的释放起到一定促进作用,在一定程度上丰富了香肠风味的前体物质。     

Proteolytic and lipolytic starter cultures and their effect on traditional fermented sausages ripening and sensory traits

In this study, three starter formulations including Lactobacillus curvatus and Staphylococcus xylosus strains selected in vitro on the basis of their lipolytic and proteolytic activities were employed for the manufacture of traditional fermented sausages of southern Italy. Microbial population, proteolysis, lipolysis, changes in free amino acids (FAA) and free fatty acids (FFA) and development of characteristic taste and flavor of the final product were investigated. Proteolysis and lipolysis were observed in sausages inoculated with proteolytic and lipolytic S. xylosus coupled with L. curvatus, while the sausage started with only S. xylosus without lactobacilli was identical to the non-inoculated control, indicating that the proteolysis could be due to both microbial activity and endogenous proteases activated by the decrease in pH. The statistical analysis applied to the instrumental and sensory data showed that there was an effect of the starter used on the characteristics of the sausage obtained. In particular, the control samples showed very close features different from the sausages obtained by adding starter cultures. Finally, analyzing the sensory parameters the sausages ripened without starter addition and those started without the L. curvatus AVL3 showed similar features indicating an influence of the presence of the lactobacilli on the final organoleptic quality of the sausages. An appropriate choice of a combination of strains in a starter formulation is fundamental to obtain products of the expected quality.     

Changes in selected biochemical and sensory parameters as affected by the superficial inoculation of Penicillium camemberti on dry fermented sausages

An atoxigenic strain of Penicillium camemberti was superficially inoculated on fermented sausages in an attempt to improve their sensory properties. The growth of this mould on the surface of the sausages resulted in an intense proteolysis and lipolysis, which caused an increase in the concentration of free amino acids, free fatty acids (FFA) and volatile compounds. Many of these were derived from amino acid catabolism and were responsible for the "ripened flavour", i.e. branched aldehydes and the corresponding alcohols, acids and esters. The development of the fungal mycelia on the surface of the sausages also protected lipids from oxidation, resulting in both lower 2-thiobarbituric acid (TBARS) values and lipid oxidation-derived compounds, such as aliphatic aldehydes and alcohols. The sensory analysis of superficially inoculated sausages showed clear improvements in odour and flavour and, as a consequence, in the overall quality of the sausages. Therefore, this strain is proposed as a potential starter culture for dry fermented sausage production.     

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.     

CHANGES IN FREE FATTY ACID AND DIACYLGLYCEROL COMPOUNDS IN SHORT-RIPENING DRY-CURED SAUSAGE

A study on lipolysis of Southern Italy short-ripened dry-cured sausages manufactured without and with starters was carried out. Samples were submitted to microbiological and chemical analysis during drying and ripening, whereas sensory analysis was performed at the end of the ripening. The starters, addition caused the inhibition of the Enterobacteriaceae. An increase of free fatty acids (FFAs) and diacylglycerols (DAGs) during the ripening was observed. The addition of starter did not affect FFA and DAG release. The DAG profile including 1,2- and 1,3-isomers was analyzed for the first time in dry-cured sausages. Oleic, linoleic and palmitic acids, and 1,2-, 1,3-palmitinolein, 1,2- and 1,3-diolein were the most abundant compounds in FFA and DAG, respectively. A decrease in the concentration of some 1,2-DAG and an increase of 1,3-OO and 1,3-OP species during the process was observed. Finally, the addition of starter cultures slightly influenced the sensory properties of sausages.

PRACTICAL APPLICATIONS

In this work the development and application of an analytical method for the determination of free fatty acids (FFAs) and diacylglycerols (DAGs) in sausages was achieved. DAGs are intermediate products of hydrolysis and also constitute a substrate for hydrolysis reactions leading to FFA release. These latter compounds undergo a series of reactions leading to the formation of low-molecular-weight products responsible for sensory properties of dry-cured sausages. Therefore, the assessment of these lipolysis products is a useful tool for evaluating the ripening process of dry-cured sausages.     

Technological characterization of a bacteriocin-producing Lactobacillus sakei and its use in fermented sausages production

The aim of this paper was the technological characterization of a Lactobacillus sakei strain, able to produce the bacteriocin sakacin P, that was originally isolated from naturally fermented sausages. Experiments were conducted in situ, using MRS-based medium, and in situ, when the strain was inoculated as starter culture in real sausage fermentation. The results obtained underlined that the strain was able to grow in conditions that are commonly used in the production line, and only lactose and high concentrations of NaCl (5% w/v) reduced the capability for bacteriocin production. When inoculated in sausages, the strain showed a good performance, being able to colonize rapidly the ecosystem. A high number of isolates, capable of producing sakacin P, were already isolated after the third day of fermentation, and persisted throughout the course of the fermentation. The inoculated strain also affected other microbial colonization trends; in fact the total bacterial count and fecal enterococci showed a rapid decrease at the end of the fermentation. Moreover, during sensory evaluation, the final sausage product received high scores for the parameters of tenderness and juiciness, with medium acidity and low rancidity. Lastly, the panelists preferred the sausages produced with the L. sakei characterized in this study when compared to a fermented sausage produced with a commercial starter.     

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.     

The effects of sodium lactate and starter cultures on pH, lactic acid bacteria, Listeria monocytogenes and Salmonella spp. levels in pure chicken dry fermented sausage

Two starter cultures (A and B) and seven sodium lactate concentrations were evaluated for chicken raw dry-fermented sausage processing. Starter culture B contained more lactic acid bacteria and less staphylococci than starter A. Their effects on acidification and inhibition of pathogens (Listeria monocytogenes and Salmonella spp.) were monitored. Starter culture B grew faster and was less inhibited by sodium lactate, thus inducting a faster and more important pH drop into the sausages. With lower pH, sausages processed with B starter were less contaminated by Listeria monocytogenes. The type of starter was found to influence the end-product pH, lactic acid bacteria content and extent of Listeria monocytogenes contamination. A 30-member panel did not note differences between sausages processed with the different starter cultures when lactate was added. Adding sodium lactate to the sausage mix reduced the pH drop in the dry sausage product. This acidification inhibiting effect of sodium lactate was greater for A. Sodium lactate significantly inhibited lactic acid bacteria development but did not reduce Listeria monocytogenes contamination frequency of the batches, unlike in many literature data. Sodium lactate may however control the acidification of the sausage processed with starter B, in order to obtain moderately acidified fermented sausages. A simple kinetic model was applied to our data. The sodium lactate content and especially the type of starter culture often had a significant effect on the four parameters of this empirical model (lag time, acidification time, initial and final pH).     

Lipolysis in dry cured sausages as affected by processing conditions

Changes of phospholipids and free fatty acids during fermentation and ripening steps of the Spanish dry cured sausage ('salchichón') process have been analyzed by studying the effects of manufacturing conditions; namely starter addition, pre-ripening and drying temperature. Phospholipid concentration of all types of samples considered (with or without starter, preripened or not) showed no important changes during fermentation, but different behaviour was observed during ripening in relation to sample type and drying temperature. At 8 °C concentrations did not change in sausages without starter and suffered a moderate decrease in samples with starter. At 16 °C the decrease was pronounced in samples without starter and much more in those with starter. Pre-ripening had no effect on phospholipid concentration. Free fatty acids increase slightly with pre-ripening, but during fermentation pre-ripened samples showed no more variation, whereas in non-preripened samples an important increase was observed. These results were similar in samples with and without starter. During the ripening step, samples with starter dried at 8 °C showed a higher increase of free fatty acids than those without starter, with no influence of the pre-ripening. At 16 °C all samples showed a higher increase than any sample dried at 8 °C. The maximum was found with non-preripened samples without starter and the minimum with pre-ripened ones, also without starter. Sausages with starter showed an intermediate increase with no effect of pre-ripening.     

Biochemical and sensory characteristics of traditional fermented sausages of Vallo di Diano (Southern Italy) as affected by the use of starter cultures

In this study, two strains of Staphylococcus xylosus isolated from traditional fermented sausages of Vallo di Diano (Southern Italy) were used in combination with an acidifying strain of Lactobacillus curvatus as starter culture for the production of fermented sausages. Two starter formulation were developed combining the proteolytic but not lipolytic (prt(+), lip(-)) S. xylosus CVS11 with the L. curvatus AVL3 (starter S1) and the S. xylosus FVS21 (prt(-), lip(+)) with the same strain of L. curvatus (starter S2). Proteolysis and lipolysis were observed during ripening by the increase in total free amino acids (FAA) and free fatty acids (FFA), respectively. Such activities were observed in both started and non started sausages (control). Moreover, the proteolytic and lipolytic activities were detected in products started by both formulations irrespective of the presence of such activities in the strains used. Therefore, it was not possible to conclude whether the effect of proteolysis and lipolysis during ripening of the started fermented sausages was due to the activity of the starter cultures or to the action of meat endogenous enzymes.     

Use of lipase from Rhizomucor miehei in dry fermented sausages elaboration: Microbial, chemical and sensory analysis

Three different amounts of lipase (0.075, 0.100 and 0.150 LU/g) from Rhizomucor miehei (Palatase M 200L Novo Nordisk) were used to determine the correct amount to use in dry fermented sausages. Determination of acidity values through fourteen days of ripening showed that 0.100 LU/g was the most appropriate. Two types of fermented sausages were manufactured, addition of the enzyme being the only difference between them. Addition of Palatase did not affect product stability (pH and A(w)), and the growth of micro-organisms. In spite of the increase in acidity value, no rancidity developed as determined by both chemical and sensory analysis. Increases in the liberation of palmitic, palmitoleic, stearic, oleic and linoleic acids were found when lipase was used. Juiciness and taste were slightly better in the sausages with Palatase than in those without, but these differences were not reflected in the overall acceptability.     

Aroma components from dried sausages fermented with Staphylococcus xylosus

Sausages with and without Staphylococcus xylosus were manufactured with four replicates. Antibiotics and a fungicide to inhibit growth of naturally occuring microorganisms were added to the control sausages. The volatile compounds from the sausages were collected and identified by gas chromatography/mass spectrometry, the effluent evaluated by sniffing and the sausages assessed by a trained panel. The study showed that sausages fermented with Staphylococcus xylosus contained several fragrant esters that were not found in control sausages without microbial growth. Control sausages had an unpleasant, rancid odour compared to sausages with Staphylococcus xylosus, indicating that the esterase activity of Staphylococcus xylosus or other microorganisms is very important in order to obtain the proper fermented sausage aroma. Although sausages with Staphylococcus xylosus contained the highest amounts of free fatty acids, it seemed to be of no importance to the aroma development. It is therefore questionable whether lipolytic activity of starter cultures has any influence on sausage flavour.     

Survival of Listeria monocytogenes strains in a dry sausage model

The survival of five inoculated Listeria monocytogenes strains (DCS 31, DCS 184, AT3E, HT4E, and HR5E) was studied in dry fermented sausages prepared using two different starter cultures (starter A and B) with or without a protective Lactobacillus plantarum DDEN 2205 strain. L. monocytogenes was detected throughout ripening in every sausage sample in which the L. plantarum DDEN 2205 strain had not been used. The use of either starter A, with a high concentration of protective culture, or starter B, with a low concentration of protective culture, resulted in L. monocytogenes-negative sausages after 17 days of ripening. Differential survival was noted among the L. monocytogenes strains during fermentation. Strains AT3E and DCS 31 survived in sausages with protective cultures more often than did the other strains, whereas HT4E and HR5E were inhibited during ripening by all starter and protective cultures used. Protective cultures such as L. plantarum may be used as part of a hurdle strategy in dry sausage processing, but variations in susceptibility of different L. monocytogenes strains can create problems if other hurdles are not included.     

发酵剂对发酵香肠食用品质的影响及其在不同直径香肠中的应用

以商业发酵剂（木糖葡萄球菌＋戊糖片球菌）为对照组,不同发酵剂组合（木糖葡萄球菌＋副干酪乳杆菌、木糖葡萄球菌＋戊糖片球菌＋副干酪乳杆菌）为实验组,通过对发酵香肠水分含量、pH值、水分活度、色泽、质地、风味和感官品质等指标进行测定,确定最佳发酵剂及适宜发酵的香肠直径。结果表明：相较于商业发酵剂,木糖葡萄球菌与副干酪乳杆菌组合发酵的香肠总体可接受性相对较高,且庚醛、1-辛烯-3-醇、乳酸乙酯、戊酸乙酯、癸酸乙酯、2-甲基丙酸乙酯等愉悦风味物质为该组独有,其特征主要表现为更加浓郁的清新味、甜香味、果香味和花香味;适宜的直径（21 mm）、水分含量（（25.40±0.00）%）和硬度（（2 812.46±767.93）g）以及相对较高的pH值（pH 5.57±0.02）是该组发酵香肠口感显著高于其他2 组的重要原因。因此,木糖葡萄球菌与副干酪乳杆菌组合发酵的小直径香肠食用品质最佳。     

微生物酶与肉组织酶对干发酵香肠中游离脂肪酸的影响

以戊糖乳杆菌31-1和木糖葡萄球菌为发酵剂生产干发酵香肠,就发酵剂对干发酵香肠成熟过程中游离脂肪酸的影响进行了研究。研究结果表明,在灌肠后的1周内,脂肪的水解主要是由组织酶引起的,微生物对脂肪微弱的水解能力主要表现在干发酵香肠成熟的后期,并且发酵剂不能改变发酵香肠中游离脂肪酸的变化模式。     

The influence of Debaryomyces hansenii and Candida utilis on the aroma formation in garlic spiced fermented sausages and model minces

The influence of the yeast starter cultures Debaryomyces hansenii and Candida utilis on fermented meat aroma was studied in model minces and in commercial-type fermented sausages. Volatile compounds from model minces and sausages were collected using diffusive and dynamic headspace sampling respectively and were identified by gas chromatography/mass spectrometry (GC/MS). A triangle test was carried out on the sausages to detect whether the yeast influenced the sausage odour. C. utilis demonstrated high metabolic activity in the model minces, producing several volatile compounds, in particularly esters. C. utilis also seemed to ferment the amino acids valine, isoleucine and leucine into compounds important for the aroma of sausages. D. hansenii on the contrary, had very little effect on the production of volatile compounds in the model minces. In the sausage experiment both yeast cultures died out before the ripening process ended and the sensory analysis showed only a slight difference between the sausages. A fungistatic test of the garlic powder added to the sausages indicated that garlic inhibits the growth of the yeast starter cultures.     

直投式发酵剂制备的哈尔滨风干肠在成熟过程中的理化及微生物特性变化

采用三种由自然发酵分离鉴定出的微生物,按一定比例混合制备复合发酵剂,将其用于哈尔滨风干肠中,研究其对风干肠的理化性质、微生物变化和游离氨基酸含量的影响。实验分为三组,分别为对照组,添加普通发酵剂组和添加直投式发酵剂组。结果表明:添加发酵剂的风干肠pH明显低于对照组,且添加直投式发酵剂的pH最低(P<0.05);添加发酵剂会使产品的游离氨基酸含量显著提高(P<0.05)、感官品质也较好(P<0.05),而各组水分活度变化差异不明显,这说明直投式发酵剂具有很好的活力和发酵性能,能够显著提高风干肠的质量,缩短发酵时间。     

A fermented meat model system for studies of microbial aroma formation

A fermented meat model system was developed, by which microbial formation of volatiles could be examined. The model was evaluated against dry, fermented sausages with respect to microbial growth, pH and volatile profiles. Fast and slowly acidified sausages and models were produced using the starter cultures Pediococcus pentosaceus and Staphylococcus xylosus. Volatiles were collected and analysed by dynamic headspace sampling and GC-MS. The analysis was primarily focused on volatiles arising from amino acid degradation and a total of 24 compounds, of which 19 were quantified, were used for multivariate data analysis. Growth of lactic acid bacteria was comparable for model and sausages, whereas survival of S. xylosus was better in the model. Multivariate analysis of volatiles showed that differences between fast and slowly acidified samples were identical for model and sausage. For both sausage and model, fast-acidified samples had a high content of ketones, sulphides and methyl-branched acids, whereas slowly acidified samples had the highest content of methyl-branched alcohols, aldehydes, their ethyl esters, phenylacetaldehyde and methional. Furthermore, model repeatability with respect to pH, microbial growth and volatile profiles was similar to sausage production. Based on these findings, the model system was considered valid for studies of aroma formation of meat cultures for fermented sausage.