The sensory quality of dry fermented sausages as affected by fermentation stage and curing agents

The effects on the quality of dry fermented sausages of applying either a slow or rapid fermentation stage, and the addition of either nitrate or nitrite was studied. In the slow fermentation process, TBARS were higher in the batch made with nitrite than in the batch containing nitrate. For rapid fermentation, both batches, with added nitrate and nitrite, presented intermediate TBARS levels. Volatile compounds coming from lipid oxidation and microbial lipid β-oxidation were more abundant (p < 0.05) in the samples submitted to a rapid fermentation stage, whilst compounds coming from amino acid catabolism and carbohydrate fermentation were higher (p < 0.05) in samples subjected to a slow fermentation stage. A significant preference (p < 0.05) was shown during sensory analysis for the batch made with nitrate and submitted to the slow fermentation process in aroma, taste and overall quality, whilst no differences in sensory analysis were detected among batches in the rapid fermentation process. Therefore, the use of nitrate is only justified in slow fermented processes due to its positive effect on the sensory quality of fermented sausages.     

Dried sausages fermented with Staphylococcus xylosus at different temperatures and with different ingredient levels - Part I. Chemical and bacteriological data

Sausages with added Staphylococcus xylosus were fermented at different temperatures and with different added levels of salt, glucose, nitrite, nitrate and Pediococcus pentosaceus in accordance with a six factor fractional design. The numbers of surviving Staphylococcus xylosus, lactic acid bacteria, pH, free fatty acids and residual amounts of nitrite and nitrate were measured. The effects of temperature and different ingredients on the chemical and bacterial data were tested using multiple linear regression and analysis of variance. The study showed that numbers of surviving Staphylococcus xylosus were reduced by high fermentation temperature, especially when salt concentration was low and glucose concentration high. High levels of nitrite and addition of Pediococcus pentosaceus had adverse effects as well. pH was lowered by high fermentation temperature, low salt concentration and addition of glucose and Pediococcus pentosaceus. On the other hand p H was increased by addition of nitrate. The pH-lowering effect of glucose was small when temperature was low. The residual levels of nitrite and nitrate were increased by addition of nitrate, but then increased and decreased, respectively, by increasing temperature. Addition of glucose and Pedio coccus pentosaceus lowered the residual amount of nitrite. The level of free fatty acids was reduced by salt and increased by higher temperature. Of the individual acids, the amount of linoleic acid was also increased by addition of Pediococcus pentosaceus and decreased by nitrate.     

Effect of nitrate and nitrite curing salts on lipolysis in dry sausages produced using a rapid fermentation process

 The effect of nitrite and nitrate curing salts on lipolysis in a typical non-smoked Spanish sausage (salchichón), manufactured using a rapid dry fermentation process, was studied. From the results obtained, one could deduce that nitrite and nitrate per se do not affect the endogenous hydrolysis of the lipids during the rapid fermentation process used to produce dry sausage. Polyunsaturated fatty acids (PUFA) are liberated in greater quantities than monounsaturated (MUFA) and saturated (SFA) ones, which suggests that they are derived mainly from phospholipids. The samples that contained nitrate curing salts contained more free fatty acids (FFA) (548 mg/100 g) than those that contained nitrite curing salts (478 mg/100 g) at the end of the fermentation step, as a consequence of having undergone a preripening stage, during which significant amounts of FFA are formed. However, at the end of the ripening process, although there is a significant increase of FFA levels in all the sausages, no significant differences between samples with nitrite and those with nitrate could be found. This result could have occurred if the oxidative effects of the samples with nitrate balanced the antioxidative properties of nitrite, such that, through different processes, similar concentrations of FFA are obtained at the end of ripening. Phospholipids (PL) and free fatty acids (FFA) are the lipid fractions that undergo the most important changes during ripening. PL are moderately degraded during the drying process, probably due to the combined effects of the drying temperature (10°C) and the presence of starters. However, this degradation does not differ significantly between the samples that contain nitrite and and those that contain nitrate in each control. Received: 23 July 1997 / Revised version: 8 October 1997     

Effect of nitrate and nitrite curing salts on lipolysis in dry sausages produced using a rapid fermentation process

 The effect of nitrite and nitrate curing salts on lipolysis in a typical non-smoked Spanish sausage (salchichón), manufactured using a rapid dry fermentation process, was studied. From the results obtained, one could deduce that nitrite and nitrate per se do not affect the endogenous hydrolysis of the lipids during the rapid fermentation process used to produce dry sausage. Polyunsaturated fatty acids (PUFA) are liberated in greater quantities than monounsaturated (MUFA) and saturated (SFA) ones, which suggests that they are derived mainly from phospholipids. The samples that contained nitrate curing salts contained more free fatty acids (FFA) (548 mg/100 g) than those that contained nitrite curing salts (478 mg/100 g) at the end of the fermentation step, as a consequence of having undergone a preripening stage, during which significant amounts of FFA are formed. However, at the end of the ripening process, although there is a significant increase of FFA levels in all the sausages, no significant differences between samples with nitrite and those with nitrate could be found. This result could have occurred if the oxidative effects of the samples with nitrate balanced the antioxidative properties of nitrite, such that, through different processes, similar concentrations of FFA are obtained at the end of ripening. Phospholipids (PL) and free fatty acids (FFA) are the lipid fractions that undergo the most important changes during ripening. PL are moderately degraded during the drying process, probably due to the combined effects of the drying temperature (10°C) and the presence of starters. However, this degradation does not differ significantly between the samples that contain nitrite and and those that contain nitrate in each control. Received: 23 July 1997 / Revised version: 8 October 1997     

氮源对低温发酵脐橙果酒挥发性成分的影响

为解决低温发酵周期长的问题,并进一步分析其挥发性成分。通过考察不同氮源质量浓度对低温发酵脐橙果酒酒精度和发酵时间的影响,确定氮源添加量,并采用顶空-固相微萃取-气质联用法(HS-SPME-GC-MS)分析其挥发性成分。结果表明,与未添加氮源的空白组相比,添加DAP 0.4 g/L、硫酸铵1.0 g/L、碳酸氢铵1.5 g/L的脐橙果酒酒精度可提高4.6%~5.3%,缩短发酵时间22%~56%。四种低温发酵脐橙果酒共检测出挥发性成分32种,其中高级醇5种、萜烯类化合物10种、中链脂肪酸2种、高级醇乙酸酯3种、中链脂肪酸乙酯7种、醛酮类5种。添加三种氮源的果酒中,萜烯类和乙酸酯类挥发性成分总含量均大于空白组,中链脂肪酸和乙基酯类挥发性成分总含量均小于空白组。其中,添加磷酸氢二铵的果酒中乙酸酯类挥发性成分总含量最高(1.170 mg/L),添加硫酸铵的果酒中萜烯类挥发性成分总含量最高(11.378 mg/L);与另外两种氮源相比,添加磷酸氢二铵的果酒中乙酸-2-苯乙酯、庚酸乙酯等挥发性成分含量相对更高,添加硫酸铵的果酒中D-柠檬烯、乙酸乙酯、癸酸乙酯等挥发性成分含量相对更高,添加碳酸氢铵的果酒中β-水芹烯、4-萜烯醇、辛酸乙酯等挥发性成分含量相对更高。实验证明了氮源对促进脐橙果酒低温发酵具有明显效果,并对挥发性香气成分含量具有一定的促进作用,为脐橙果酒低温发酵工艺的应用提供了理论依据。     

The influence of additives on the oxidation of pork back fat and its effect on water and fat binding in finely comminuted batters

We have found in previous experiments that slaughter-fresh processed pork back fat enhances batter stability in frankfurter-type sausages. In an attempt to clarify whether the decrease of batter stability with storage time of the fatty tissue is due to the fat oxidation a series of experiments was conducted using sodium nitrite and nitrate, 6- O -palmitoyl- l -ascorbic acid and dl-ga-tocopherol as additives to the fatty tissue. It was found that sodium nitrite and nitrate, although the first is regarded as an antioxidant in meat, had a pro-oxidant effect when added to the fatty tissue. The palmitoyl-l-ascorbic acid showed the best antioxidative activity, and the dl-α-tocopherol was less effective as an antioxidant. Contrary to its effect on the increase in fat oxidation, sodium nitrite addition gave a remarkable improvement in batter stability, expressed as jelly and fat separation, which was more pronounced on the second day of storage of the fatty tissue. The pro-oxidant effect of nitrite increased up to a concentration of 375 ppm added to the fatty tissue. Concentrations of nitrite in fatty tissue up to 125 ppm had no influence on jelly release but at higher concentrations the jelly separation decreased. All nitrite concentrations, from 62·5 to 2250 ppm, showed significantly lower fat separation than for the control treatment, especially on the eighth and 15th day of storage of the fatty tissue. The results show that fat oxidation has little effect on batter stability. Further experiments are needed to clarify the mode of action of nitrite in improving batter stability.     

亚硝酸钠对自然发酵哈尔滨风干肠微生物生长、脂肪氧化及挥发性化合物的影响

研究不同添加水平的亚硝酸钠（NaNO2）对自然发酵哈尔滨风干肠微生物生长、脂肪氧化和挥发性化合物的影响。结果表明,添加NaNO2对风干肠发酵过程中的pH值、水分活度、总菌落数和乳酸菌菌落数无显著影响（P＞0.05）,但添加0.10?g/kg以上的NaNO2可以显著降低风干肠中大肠杆菌菌落数（P＜0.05）。色差分析表明当NaNO2添加量达到0.10?g/kg时可以起到良好的发色作用,过氧化物值和硫代巴比妥酸反应产物显示NaNO2具有良好的抗氧化作用,NaNO2残留量随发酵时间的延长而逐渐降低,发酵结束后,各实验组间差异不显著（P＞0.05）。与此同时,挥发性化合物的分析结果显示,NaNO2添加量不影响风干肠中由碳水化合物发酵和氨基酸降解所产生的挥发性化合物的相对含量（P＞0.05）,但显著影响由脂肪氧化所产生的醛类和酮类化合物的相对含量（P＜0.05）,NaNO2添加量越大,对脂肪氧化产物的抑制作用越显著。     

Colour formation in fermented sausages by meat-associated staphylococci with different nitrite- and nitrate-reductase activities

Three Staphylococcus strains, S. carnosus, S. simulans and S. saprophyticus, selected due to their varying nitrite and/or nitrate-reductase activities, were used to initiate colour formation during sausage fermentation. During fermentation of sausages with either nitrite or nitrate added, colour was followed by L^*a^*b measurements and the content of nitrosylmyoglobin (MbFe^IINO) quantified by electron spin resonance (ESR). MbFe^IINO was rapidly formed in sausages with added nitrite independent of the presence of nitrite reducing bacteria, whereas the rate of MbFe^IINO formation in sausages with added nitrate depended on the specific Staphylococcus strain. Strains with high nitrate-reductase activity showed a significantly faster rate of pigment formation, but other factors were of influence as well. Product stability for the sliced, packaged sausage was evaluated as surface colour and oxidation by autofluorescence and hexanal content, respectively. No significant direct effect of the Staphylococcus addition was observed, however, there was a clear correspondence between high initial amount of MbFe^IINO in the different sausages and the colour stability during storage. Autofluorescence data correlated well with hexanal content, and may be used as predictive tools. Overall, nitrite- and nitrate-reductase activities of Staphylococcus strains in nitrite-cured sausages were of limited importance regarding colour development, while in nitrate-cured sausages strains with higher nitrate reductase activity were crucial for ensuring optimal colour formation during initial fermentation stages.     

Nitrate reduction during fermentation by Gram-negative bacterial activity in carrots

Carrots were subjected to lactic acid fermentation through the action of a starter culture, Lactobacillus plantarum, and changes in the amount of both the naturally present and added nitrate were recorded. The nitrate content in the carrots decreased to about 10% of the original amount during the initial stage of the fermentation process. By using irradiation-sterilized carrots it was shown that the decrease in the nitrate content is a result of the activity of the Gram-negative bacteria, which dominate the flora during the initial stage of the fermentation process, and that the lactic acid bacteria present were unable to affect the nitrate content. The nitrite concentration was also determined and was found not to exceed 0.2 mg/kg on any occasion. The conclusion is that if the nitrate content of carrots is to be lowered in a fermentation process, this process must be controlled in such a way as to allow the original Gram-negative flora to reduce the nitrate amount before the starter organism takes over.     

甘兰乳酸发酵过程中亚硝峰消长机制及抑制途径的研究

本文研究了甘兰乳酸发酵过程中微生物区系及硝酸盐、亚硝酸盐和硝酸还原酶的变化规律。发现甘兰乳酸发酵过程中硝酸盐降低,亚硝酸盐累积并产生亚硝峰,之后又迅速消失。实验证明,亚硝峰的产生是由于发酵初期的微生物将甘兰体内的硝酸盐还原为亚硝酸盐所致。同时,在发酵的酸性环境下,亚硝酸盐又被化学降解,另外,乳酸菌本身对亚硝酸盐的降解也有作用。采用纯接种技术可有效地降低亚硝峰,并且以接种肠膜明串珠菌(Leumesenteroides)效果最佳。添加适量蒜汁,无论对自然发酵,还是纯接种发酵都可以极明显地抑制亚硝峰的出现,且整个发酵过程中,亚硝酸盐含量都维护在一个极低的水平。     

Dried sausages fermented with staphylococcus xylosus at different temperatures and with different ingredient levels. Part IV. Amino acid profile

Sausages with added Staphylococcus xylosus were fermented at different temperatures and with different added levels of salt, glucose, nitrite, nitrate and Pediococcus pentosaceus in accordance with a six-factor fractional design. The amounts of individual amino acids were measured and the effects of temperature and different ingredients on the amino acid profile were tested using multiple linear regression and analysis of variance. Also, the amino acid profile was correlated to the level of volatile compounds by partial least squares analysis. The study showed that the level of free amino acids was significantly affected by the different factors. High fermentation temperature and nitrite content increased the amount, while high contents of salt, nitrate, glucose and P. pentosaceus lowered the amount of free amino acids. In general, temperature and nitrate had the greatest influence. It was shown that the amounts of the volatile compounds, 2-methyl propanal, 2- and 3-methyl butanal, were inversely correlated with the amounts of valine, isoleucine and leucine, respectively, indicating that those volatiles were degradation products of the latter.     

亚硒酸钠对薇菜发酵中亚硝酸盐含量的影响

该试验研究了亚硒酸钠对薇菜发酵过程中亚硝酸盐含量的影响。分别添加亚硒酸钠于自然发酵和接种发酵的薇菜,发现亚硒酸钠可以降低“亚硝峰”;接种发酵薇菜,“亚硝峰”比自然发酵的低,且提前1 d出现,发酵8 d时,亚硝酸盐和硝酸盐含量皆比自然发酵的低。以接种发酵结束后的亚硝酸盐含量为评价指标,确定了发酵薇菜中亚硒酸钠的适宜添加量为0.3 g/L。通过添加Na2CO3维持发酵薇菜的pH值为5.0,发酵薇菜的“亚硝峰”提前1 d出现,且发酵结束时亚硝酸盐含量由自然pH时的2.16 mg/kg降为0。亚硒酸钠分3次添加,每次0.10 g/L时,“亚硝峰”提前至发酵3 d时出现,且添加次数增多时,发酵结束时亚硝酸盐含量也由一次性添加时的1.26 mg/kg降低到0.34 mg/kg,而硝酸盐也由0.13 g/kg降低到0.032 g/kg。     

Monitoring nitrite and nitrate residues in frankfurters during processing and storage

Frankfurter-type sausages were prepared in a pilot plant with different concentrations of NaNO(2) (75, 125 or 250 ppm) combined or not with 200 ppm KNO(3). A meat system, free of curing agents, was also used as control. Nitrite and nitrate levels were tested in various processing steps and over 120 days storage at 3 °C of the vacuum-packaged frankfurters. Little influence of the originally added nitrite level on the amount of nitrate formed was observed. Important losses of nitrite and nitrate were due to cooking. Thereafter about 50% of the nitrite added initially remained in this form in all samples (39, 59 and 146 ppm, respectively) and between 10 and 15% as nitrate. When only nitrate was initially added, formation of nitrite after cooking was observed (maximum level 43 ppm NaNO(2)). Formulations prepared with both nitrate and nitrite showed no significant differences (p < 0.01) respect to their nitrite or nitrate counterparts. A good correlation among nitrite and nitrate levels and storage time was showed by multiple linear regression analysis. It is concluded that the use of nitrate in combination with nitrite in cooked meat products seems to have little technological significance and adds to the total body burden of nitrite.     

大白菜发酵过程中亚硝酸盐消长规律的研究

对大白菜在发酵过程中亚硝酸盐的形成规律进行了研究。结果发现 ,亚硝酸盐的出现期主要集中在发酵前期 ,随发酵液酸度的增加亚硝酸盐逐渐消失。亚硝峰的形成主要是由杂菌将硝酸盐还原为亚硝酸盐的作用 ,与大白菜体内硝酸还原酶无直接关系。亚硝峰消失的原因一是在酸性条件下的化学降解 ,二是乳酸菌对亚硝酸盐的还原作用。纯接种发酵可降低亚硝峰 ,无论是接入单一菌株还是接入混合菌株都能明显地降低亚硝峰 ,尤以混合接种效果最佳     

Survival of Listeria innocua in dry fermented sausages and changes in the typical microbiota and volatile profile as affected by the concentration of nitrate and nitrite

The involvement of nitrate and nitrite in the formation of N-nitrosamines in foods is a matter of great concern. This situation has led to revise the real amount of nitrate and nitrite needed in meat products to exert proper technological and safety activities, and also to extensive research to find alternatives to their use. The present study addresses the possibility of reducing the ingoing amounts of these additives below the legal limits established by the current European regulations. Different concentrations of nitrate and nitrite were tested on Spanish salchichón-type dry fermented sausages concerning their role in the microbiota and volatile profile. Sausages were manufactured with the maximum ingoing amounts established by the EU regulations (150 ppm NaNO₃ and 150 ppm NaNO₂), a 25% reduction and a 50% reduction; control sausages with no nitrate/nitrite addition were also prepared. The mixtures were inoculated with 5 log cfu/g of Listeria innocua as a surrogate for Listeria monocytogenes. L. innocua numbers in the final product were approximately 1.5 log cfu/g lower in the batch with the maximum nitrate/nitrite concentration when compared to 25 and 50% reduced batches, and about 2 log cfu/g in comparison to the control sausages. The final numbers of catalase-positive cocci were 1 log cfu/g higher in the 50% nitrate/nitrite reduced batch and 2 log cfu/g higher in the control sausages, compared to products manufactured with the maximum nitrate/nitrite concentration. This increase was related to a higher amount of volatile compounds derived from carbohydrate fermentation and amino acid degradation. Sausages with no addition of nitrate/nitrite showed higher amount of volatiles from lipid oxidation. Enterobacteriaceae counts reached detectable values (1-2 log cfu/g) in both nitrate/nitrite reduced sausages and in the control batch, while these organisms were not detected in the batch with the maximum ingoing amount. Nitrate and nitrite exerted a significant effect on the typical microbiota of dry fermented sausages and effectively contributed to control Listeria. These considerations should be taken into account in view of a future restriction in the use of these curing additives.     

Headspace concentration of selected dry-cured aroma compounds in model systems as affected by curing agents

The effect of curing agents (sodium chloride, nitrate, nitrite, glucose, and ascorbic acid) on the headspace concentration of six volatile compounds (2-methyl-butanal, 3-methyl-butanal, 2-pentanone, hexanal, methional, and octanal) has been studied. These volatile compounds were selected based on their presence and contribution to the flavour of typical Spanish dry-cured meat products. The release of volatile compounds to the headspace has been studied by using solid-phase microextraction (SPME) and gas chromatography analysis. The main effect was produced by sodium chloride, because it produced a salting-out effect on all the volatile compounds studied. Furthermore, the presence of nitrite, nitrate, and ascorbic acid in the solution also produced an increase in the headspace concentration of the volatile compounds while glucose decreased the headspace concentration of octanal and hexanal. The changes in the relative proportions of volatile compounds due to the effect of curing agents should be taken into account as they may change the flavour perception of dry-cured meat products.     

副干酪乳杆菌HD1.7发酵酸菜与商品酸菜代谢物比较与品质评价

以分离自酸菜发酵液的副干酪乳杆菌(Lactobacillus paracasei)HD1.7为发酵剂,构建酸菜发酵微生物生态系统研究模型,利用高效液相色谱和气相色谱-质谱联用技术,分析L.paracasei HD1.7发酵酸菜、自然发酵酸菜及8种商品酸菜中有机酸、酸度、亚硝酸盐、VC、甘油、乙醇、甘露醇、2,3-丁二醇及挥发性风味成分的差别。结果表明:不同酸菜样品中有机酸含量与酸度正相关,黑大酸菜、L.paracasei HD1.7发酵酸菜和袁滋袁味酸菜有机酸含量较高,分别为(13.21±0.13)、(13.20±0.17)、(13.13±0.13)g/L。此外,黑大酸菜、L.paracasei HD1.7发酵酸菜、边府记酸菜中不仅亚硝酸盐含量显著低于其他供试酸菜,VC含量显著高于其他供试酸菜(P0.05),安全性、营养价值更高,而且风味物质含量丰富、感官品质更优。研究结果有助于酸菜生产工艺监控和质量标准建立,对酸菜质量稳定性的提高和产业发展水平的提升具有实践指导意义;同时展现了L.paracasei HD1.7在酸菜发酵的工业生产中的应用前景。     

Metabolism of nitrate in fermented meats: the characteristic feature of a specific group of fermented foods

Within the universe of food fermentation processes the multi-purpose use of nitrate and/or nitrite is a unique characteristic of meat fermentations. These curing agents play a decisive role in obtaining the specific sensory properties, stability and hygienic safety of products such as fermented sausages, ham and, more recently, emulsion type of sausages. The use of nitrate is the traditional method in curing processes and requires its reduction to reactive nitrite. Thus, nitrate reduction is the key event that is exclusively performed by microorganisms. Under controlled fermentation conditions starter cultures are used that contain staphylococci and/or Kocuria varians, which in addition to strongly affecting sensory properties exhibit efficient nitrate reductase activity. To obtain clean label products some plant sources of nitrate have been in use. When producing thermally treated sausages (e.g. of emulsion type), starter cultures are used that form nitrite before cooking takes place. Staphylococci reduce nitrite to ammonia after nitrate has been consumed. K. varians is devoid of nitrite reductase activity. Nitrate and nitrite reductases are also present in certain strains of lactobacilli. It was shown that their application as starter cultures warrants efficient activity in sausages made with either nitrate or nitrite. NO is formed from nitrite in numerous chemical reactions among which disproportionation and reaction with reductants either added or endogenous in meat are of practical importance. Numerous nitrosation and nitrosylation reactions take place in the meat matrix among which the formation of nitrosomyoglobin is of major sensory importance.Safety considerations in meat fermentation relate to the safe nature of the starter organisms and to the use of nitrate/nitrite. Staphylococci (“micrococci”) in fermented meat have a long tradition in food use but have not received the QPS status from the EFSA. They require, therefore, thorough assessment with regard to toxigenicity and pathogenicity determinants as well as presence of transferable antibiotic resistance. Nitrate and nitrite are still considered basically undesired in food. The main objections are based on their potential to form nitrosamines with carcinogenic potential. In view of new results from intensive research of NO, potential risks are opposed by positive effects on human health.     

Malondialdehyde-Nitrite Interactions in Meat and Model Systems

Nitrite retarded formation of carbonyl compounds in meat when added before cooking, but had no effect after cooking and storage even though the 2-thiobarbituric acid (TBA) number was very low due to reaction of malondialdehyde with nitrous acid. Although tripolyphosphate/ascorbate reacted similary preventing formation of carbonyl compounds, they had no effect on the TBA number when added after cooking. In a model system, nitrite reacted with malondialdehyde at pH 1.3 at room temperature. The percentage of reacting malondialdehyde decreased with higher pH and increased with nitrite concentration. The reaction of malondialdehyde with nitrite yielded high-molecular-weight products. Sulfanilamide prevented the reaction of malondialdehyde with nitrite, but only when added before nitrite.