Factors controlling the growth of Clostridium botulinum types A and B in pasteurized, cured meats

The effect of combinations of sodium chloride (2.5, 3.5, 4.5% w/v on water), sodium nitrite (100, 200, 300 μg/g), sodium nitrate (0, 500 μg/g), sodium isoascorbate (0,1000 μg/g, or equimolar with nitrite level) and polyphosphate (Curaphos 700; 0, 0.3% w/v), on the growth of Clostridium botulinum types A and B was studied in an experimental pork slurry system, without heating and after two heat treatments (80°C for 7 min and 80°C for 7 min plus 70°C for 1 hr) followed by storage at: 15, 17.5, 20 or 35°C for up to 6 months.
Statistical analyses showed that increasing salt or nitrite levels, adding isoascorbate or nitrate, using the highest heat treatment or decreasing the storage temperature all significantly reduced toxin production by Cl. botulinum . The addition of 0.3% polyphosphate (Curaphos 700) significantly increased toxin production. There were many significant two-factor interactions; the effect of increasing nitrite was relatively less if isoascorbate was present, at 4.5% salt, or at low storage temperature. The presence of isoascorbate also counteracted the increase in toxin production attributed to the presence of polyphosphate.     

Prediction of toxin production by Clostridium botulinum in pasteurized pork slurry

A model pork slurry system was used to study factors controlling the growth of Clostridium botulinum types A & B (Roberts, Gibson & Robinson, 1981a,b). The following factors were studied in combination: sodium chloride (2.5, 3.5, 4.5% w/v on water); sodium nitrite (100, 200, 300 μg/g); sodium nitrate (0, 500 μg/g); sodium isoascorbate (0, 1000 μg/g); polyphosphate (Curaphos 700, 0, 0.3% w/v); heat treatment (none, 8°C/7 min, 80°C/7 min + 70°C/1 hr); at two pH levels and stored at: 15, 17.5, 20 or 35°C.
Analyses of results yielded a statistical model providing two formulae (for 'low' and 'high' pH slurries) which estimate the probability of toxin production in the pork slurry system within the limits defined above.     

Factors controlling the growth of Clostridium botulinum types A and B in pasteurized, cured meats III. The effect of potassium sorbate

The growth of Clostridium botulinum types A and B spores, at 10¹ or 10³ per container, was studied in a pork slurry system containing nitrite (40 μg/g), sodium chloride (2.5, 3.5, 4.5% w/v) sodium isoascorbate (550 μg/g) at varying pH levels, with or without potassium sorbate (0.26% w/v), without heating and after two heat treatments (80°C for 7 min, and 80°C for 7 min + 70°C for 1 hr) followed by storage at 15, 17.5, 20 or 35°C for up to 6 months. At a given spore inoculum, potassium sorbate significantly decreased toxin production, as did increasing NaCl, decreasing pH or decreasing storage temperature. Heat treatment did not significantly affect spoilage or toxin production overall, but interacted significantly with some factors. The effect of sorbate was greater at 3.5% NaCl than at 2.5%, at pH values below 6.0, and at low storage temperature.     

Influence of turkey meat on residual nitrite in cured meat products

A response surface experimental design was employed to estimate residual nitrite level at various initial nitrite concentrations, percent turkey meat in the formula, and heat quantity (F) values using a typical wiener as the test system. Pork and mechanically separated turkey were used as the meat ingredients. Residual nitrite and pH were measured at day 1, 7 days, 14 days, and 49 days after processing. Protein, fat, salt, moisture, and CIE (L*a*b*) color values were also determined. Results showed that the effect of turkey meat on residual nitrite level was significant (P < 0.01). An increased amount of turkey meat in the formula resulted in lower residual nitrite levels at a fixed pH. The residual nitrite level was initially proportional to initial nitrite concentration, but it became a nonsignificant factor during longer storage time. Differences in heat quantity had a significant effect (P < 0.05) on residual nitrite level initially. Greater heat quantity decreased residual nitrite level in finished cured meat products at a fixed pH. However, this effect became nonsignificant during longer storage. Reduction of residual nitrite in wieners because of turkey meat addition at a fixed pH was due to characteristics of the turkey tissue, but the mechanism of action remains unknown. It was also established that commercial wieners had a higher pH if poultry meat was included in the formulation.     

Time-dependent depletion of nitrite in pork/beef and chicken meat products and its effect on nitrite intake estimation

The food additive nitrite (E249, E250) is commonly used in meat curing as a food preservation method. Because of potential negative health effects of nitrite, its use is strictly regulated. In an earlier study we have shown that the calculated intake of nitrite in children can exceed the acceptable daily intake (ADI) when conversion from dietary nitrate to nitrite is included. This study examined time-dependent changes in nitrite levels in four Swedish meat products frequently eaten by children: pork/beef sausage, liver paté and two types of chicken sausage, and how the production process, storage and also boiling (e.g., simmering in salted water) and frying affect the initial added nitrite level. The results showed a steep decrease in nitrite level between the point of addition to the product and the first sampling of the product 24 h later. After this time, residual nitrite levels continued to decrease, but much more slowly, until the recommended use-by date. Interestingly, this continuing decrease in nitrite was much smaller in the chicken products than in the pork/beef products. In a pilot study on pork/beef sausage, we found no effects of boiling on residual nitrite levels, but frying decreased nitrite levels by 50%. In scenarios of time-dependent depletion of nitrite using the data obtained for sausages to represent all cured meat products and including conversion from dietary nitrate, calculated nitrite intake in 4-year-old children generally exceeded the ADI. Moreover, the actual intake of nitrite from cured meat is dependent on the type of meat source, with a higher residual nitrite levels in chicken products compared with pork/beef products. This may result in increased nitrite exposure among consumers shifting their consumption pattern of processed meats from red to white meat products.     

The influence of nitrite and nitrate on microbial, chemical and sensory parameters of slow dry fermented sausage

Nitrate and/or nitrite are used in the manufacture of dry-fermented sausages. However, research has mainly been focusing on nitrite and its effect on flavour development whereas little attention has been paid to nitrate. The aim of the present work was to study the effect of nitrate and nitrite as curing salts on the quality of a slow fermentation process. Two different batches containing nitrate or nitrite were manufactured. Microbial and chemical parameters were monitored during ripening and after vacuum packed storage, as well as their fatty acid composition and their profile of volatile compounds. The oxidation, measured as TBARS (Thiobarbituric acid reactive substances), was greater in the samples with added nitrite than in the samples with added nitrate. FFA (free fatty acids) release was higher in the samples containing nitrite throughout the process. Volatile compounds arising from amino acid degradation and carbohydrate fermentation were generated at higher levels in the samples with added nitrate, probably due to the higher population of microorganisms in these samples and the effect of nitrate on their metabolism.     

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.     

The relationship of added nitrate to pigment formation in native pork-sausage (longaniza)

The nitrosomyoglobin, residual nitrate and nitrite concentrations of the native pork-sausage (longaniza) were analysed within a ten-day storage period. Several levels of nitrate (50, 100, 300, 500 and 2000, 3500 and 5000 ppm) were employed. These were examined to determine the minimum amount of nitrate that could be added to the curing mixture for the making of longaniza whilst still obtaining acceptable organoleptic properties, especially colour.Pigment formation in longaniza due to the added nitrate occurs from the onset and reaches its maximum on the third or fourth day. The desirable pink to red colour in the cured meat was obtained even with amounts of nitrate as low as 50 ppm. All treated longaniza showed high and similar colour scores compared with the control. However, higher colour stability was obtained with the use of greater than 500 ppm nitrate. The amount of residual nitrate and nitrite is directly proportional to the initial quantity of nitrate added. The use of 50 to 500 ppm nitrate poses no danger to public health while the use of 2000 to 5000 ppm nitrate enhances the possibility of carcinogenic nitrosamine formation and incidence of methemoglobinemia.     

2种复合香辛料精油对西式培根安全品质的影响

为验证前期研究的2 种对N-亚硝胺具有阻断作用的复合香辛料精油（compound spice essential oil,CSEO）在西式培根中的实际应用效果,在西式培根加工腌制环节设计3 组实验：1）空白对照组：注射20 mL/100 g基础腌制液（含亚硝酸钠、异抗坏血酸钠、复合磷酸盐等）;2）CSEO-ND组：在基础腌制液中添加由胡椒精油、姜精油、花椒精油、八角精油和丁香精油组成的对N-二甲基亚硝胺（N-nitrosodimethylamine,NDMA）具有较强抑制作用的CSEO-ND;3）CSEO-NP组：在基础腌制液中添加由花椒精油、八角精油、麻椒精油、丁香精油和迷迭香精油组成的对N-亚硝基吡咯烷（N-nitrosopyrrolidine,NPYR）有较强抑制作用的CSEO-NP。对西式培根产品烧烤（200 ℃,5 min）后的感官评分、烧烤前后pH值、亚硝酸盐、生物胺以及N-亚硝胺含量进行测定,并将西式培根产品置于（36±1） ℃条件下进行破坏性实验。结果表明：CSEO-ND可赋予西式培根特有的风味,提高产品的整体可接受性,降低产品中亚硝酸盐残留量（23.52 mg/kg）,提高抗氧化性,抑制生物胺总量的增加（抑制率为43.95%）,有效控制NDMA（抑制率为60.18%）和NPYR（抑制率为64.88%）的形成;CSEO-NP具有较强的抑菌性,能很好地控制产品中组胺、酪胺、腐胺等毒性较强的生物胺形成,对NDMA（65.99%）和NPYR的抑制率（66.13%）均高于CSEO-ND,但产品风味受到一定影响,亚硝酸盐残留量（40.00 mg/kg）超过国家限量标准（30 mg/kg）。     

Effect of light on colour and reaction of nitrite in sliced pork bologna under different chilled storage temperatures

The effect of different lighting conditions (darkness and exposure to 1900 ± 100 lux) on colour stability, conversion of added nitrite (residual nitrite, nitrite converted to nitrate, nitroso heme pigments, and protein-bound nitrite) and oxidative rancidity (2-thiobarbituric acid index) in sliced, vacuum-packaged pork bologna as a function of storage temperature (0 ± 1°C and 7 ± 1°C) was studied. Colour (redness) losses over the storage period were more dependent upon photochemical processes than on thermal processes, and the action of temperature on colour was attributable to its effect on microbial growth, which in turn also affects oxygen availability. Conversion of nitrite into the different fractions studied was chiefly temperature-dependent, but exposure to light lowered the residual nitrite content. Nevertheless, the effect of illumination on the constant rate of residual nitrite depletion was dependent on the storage temperature. Recovery of the added nitrite, i.e. the total of the nitrite in all the fractions combined, was highly dependent upon fluctuations in the residual nitrite levels and varied over the storage period. Under the conditions of the experiment, photo-oxidation did not appear to be a determining factor in lipid oxidation.     

烟叶硝酸还原酶活性在烘烤过程中的变化

研究了不同品种、不同部位烟叶在不同烘烤方式、不同烘烤条件下,烟叶硝酸还原酶（ＮＲ）活性在烘烤过程中的变化。结果表明,在烘烤过程中ＮＲ活性随烟叶的失水干燥而下降,失水干燥快,ＮＲ活性下降也快。在不同品种间,红花大金元的ＮＲ活性下降比Ｋ３２６的下降稍快。不同烘烤方式、不同变黄温湿度对ＮＲ活性变化也有影响,在密集烘烤和较高的变黄湿度条件下,ＮＲ活性下降最慢,酶活性保持的时间较长。     

Effects of Particle Size, Casing Diameter, and Addition of Emulsions and Residual Nitrite on Quality Attributes of Cooked Salami

Cooked salami was manufactured from raw materials of two particle sizes (6.5 and 13.0 mm) with and without the addition of 25% meat batter, and stuffed into two sizes of casings (64 and 90 mm). A fourth variable, three levels of added nitrite (110, 220, and 440 ppm) was also included in the experimental design. Increasing the particle size and casing diameter resulted in less shrinkage and a firmer product. The use of 25% added meat batter reduced shrinkage but made a softer product upon heat processing than without meat batter added. Neither sensory nor textural properties were significantly influenced (p < 0.05) by the level of nitrite added. Levels of residual nitrite decreased upon storage of the product up to 14 days.     

萨拉米模拟发酵过程中猪肉肌红蛋白存在形式影响规律研究

为实现萨拉米色泽的精确控制技术,用480～640nm的扫描图谱分析了萨拉米模拟发酵过程中温度、pH值、亚硝酸钠添加量、异抗坏血酸钠添加量对猪肉肌红蛋白存在形式的影响规律.结果表明,在4h以内,随着发酵温度的上升,氧合肌红蛋白含量显著减少;随着pH值的降低,高铁肌红蛋白显著减少,氧合肌红蛋白含量显著增多,脱氧肌红蛋白显著减少;在同一时间段,随着亚硝酸钠添加量的增加,高铁肌红蛋白显著增加,氧合肌红蛋白显著减少;随着异抗坏血酸钠添加量的增加,肌红蛋白3种形式的比例均无显著变化.     

Influence of an extract of liver on colour and shelf stability of sliced bologna

The influence of the substances present in an extract of liver on the characteristics of bologna sausage was studied in sliced, vacuum-packaged bologna stored under refrigeration for 70 days in darkness or exposed to light (180 ± 10ft-c). The extract replaced 0, 50 or 100% of the water added during formulation. The characteristics studied were colour intensity and stability, residual nitrite content, and oxidative rancidity. Colour fading was nearly non-existent in the batches stored in darkness but was more pronounced in those exposed to light, in which the rate of fading was inversely dependent upon the proportion of extract added. Nitrite loss over the storage period followed a first-order kinetic equation and was dependent upon both liver extract concentration (higher at higher extract levels) and the storage conditions (more pronounced in the batches exposed to light). The changes in the residual nitrite concentration could be attributed to its colour stabilizing action. While the 2-thiobarbituric acid (TBA) index tended to be higher at higher concentrations of liver extract, the increases were not substantial and were not dependent upon the storage period or illumination. Rancidity was not a limiting factor in the refrigerated storage of the product.     

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.     

Evaluation of Iron Binding Compounds as Inhibitors of Gas and Toxin Production by Clostridium botulinum in Ground Pork

Iron binding systems (heme iron binding gases alone or with iron binding salts) were evaluated for antibotulinal activity in ground pork. Compared with meat systems containing nitrite or nitrite plus supplemental iron compounds, carbon monoxide (CO) was not antibotulinal. Nitric oxide (NO) treated meats did swell slower, but nitrite was also found in these systems. Thus, neither CO nor NO would be suitable substitutes for sodium nitrite in meat curing. Addition of ferric chloride or myoglobin decreased the antibotulinal effectiveness of nitrite, but samples containing nitrite plus ethylene-diaminetetraacetate (EDTA) or denatured nitrosylated myoglobin (NOMb) swelled slower. Supplemental iron compounds probably decreased residual nitrite levels in the product, thus permitting botulinal growth, rather than directly stimulating growth by providing iron as an external nutrient.     

Nitrate reductase activity of Staphylococcus carnosus affecting the color formation in cured raw ham

The influence of the nitrate reductase activity of two Staphylococcus carnosus strains used as starter cultures on the formation of nitrate, nitrite and color pigments in cured raw ham was investigated. In this context, microbiological, chemical and multivariate image analyses were carried out on cured raw hams, which were injected with different brines containing either nitrite or nitrate, with or without the S. carnosus starter cultures. During processing and storage, the viable counts of staphylococci remained constant at 6.5 log cfu/g in the hams inoculated with starter cultures, while the background microbiota of the hams processed without the starter cultures developed after 14 days. Those cured hams inoculated with S. carnosus LTH 7036 (high nitrate reductase activity) showed the highest decrease in nitrate and high nitrite concentrations in the end product, but were still in the range of the legal European level. The hams cured with nitrate and without starter culture or with the other strain, S. carnosus LTH 3838 (low nitrate reductase activity) showed higher residual nitrate levels and a lower nitrite content in the end product. The multivariate image analysis identified spatial and temporal differences in the meat pigment profiles of the differently cured hams. The cured hams inoculated with S. carnosus LTH 3838 showed an uncured core due to a delay in pigment formation. Therefore, the selection of starter cultures based on their nitrate reductase activity is a key point in the formation of curing compounds and color pigments in cured raw ham manufacture.     

Microbiological aspects and technological need: technological needs for nitrates and nitrites.

Nitrate and nitrite are used in meat and fish curing, and in the manufacture of certain cheeses. Nitrate itself has little antimicrobial effect and in most applications could be replaced by lower concentrations of nitrite. Further, improved hygiene diminishes the need for nitrite. The antimicrobial activity and technological needs for nitrate and nitrite are reviewed. It is concluded that the technological needs for nitrite in meat products stored at < 10 degrees C could be met by added nitrite concentrations of 50 mg/kg. The overall effect of nitrate in salted fish appears to be marginal. In such products, Vibrio parahaemolyticus does not grow at salt concentrations of 10% and food poisoning by this organism is not related to the absence of nitrate or nitrite; growth of Clostridium botulinum Type E (the predominant cause of botulism from fish products) is arrested by salt concentrations of 3-4%. Listeria monocytogenes in seafood cannot be controlled by nitrite. The use of nitrate in cheese production could be avoided, or at least reduced to a low level by avoiding silage with a high count of gas-producing Clostridia and hygienic milk collection.     

Incubation of curing brines for the production of ready-to-eat, uncured, no-nitrite-or-nitrate-added, ground, cooked and sliced ham

Salt concentration, vegetable juice powder (VJP) concentration and temperature were investigated to determine necessary conditions for incubation of curing brines including VJP and a starter culture containing Staphylococcus carnosus prior to production of naturally cured, no-nitrate/nitrite-added meat products. Subsequently, incubated brines were utilized to produce no-nitrate/nitrite-added sliced ham in which quality characteristics and residual nitrite concentrations were measured to determine feasibility of brine incubation for nitrate conversion prior to injection. Two ham treatments (one with VJP and starter culture; one with pre-converted VJP) and a nitrite-added control were used. No differences (P>0.05) were found for color in the VJP treatments. Control sliced ham was redder after 42 days of storage, retaining significantly (P<0.05) greater a* (redness) than either of the VJP treatments. Residual nitrite concentration was greater (P<0.05) in the control hams during the first week of storage. While the nitrite-added control retained greater red color and initially had more residual nitrite than the VJP treatments, the two VJP treatments did not differ from each other.     

Stability of α-tocotrienol and α-tocopherol in salami-type sausages and curing brine depending on nitrite and pH

We studied the stability of the valuable vitamer nutrients α-tocotrienol and α-tocopherol and options for their protection in salami-type sausages (blended with α-tocotrienol-rich barley oil) and curing brine. Four different sausage formulations were produced containing nitrite curing salt; nitrite curing salt and ascorbic acid (300 mg/kg); nitrite curing salt and carnosic acid (45 mg/kg); or sodium chloride. Initial vitamer contents (100 mg/kg) did not decrease significantly during ripening and decreased only slightly during storage. Ascorbic acid and carnosic acid were found to be effective in preserving the vitamers in fresh sausages. Freeze-drying of sausages resulted in a significant loss of vitamers (97%), particularly after 14-day storage at room temperature, even in the presence of shielding gases. The vitamer content in the curing brine decreased with decreasing pH in the presence of nitrite. A nitrite concentration of 136 mg/L at pH 4 resulted in significant loss (90%) of the vitamers. Sufficient stability of the vitamers in salami-type sausage and curing brine can be achieved by processing, formulation, and storage conditions.