Effect of Iron Form, Temperature, and Inoculation with Clostridium botulinum Spores on Residual Nitrite in Meat and Model Systems

The effect of iron form (ferrous, ferric, heme), temperature and botulinal spores on nitrite level was determined in meat. In model systems, ferritin iron was also included, and ascorbate was used as a reducing agent. Reduced hemoglobin caused the most rapid nitrite depletion in both systems. Ferrous iron caused faster nitrite depletion in model systems than in meat. Ferrous iron reduced nitrite readily in model systems at 27°C, but not at 5°C. Ferritin iron did not affect nitrite level. In meat at 27°C, nitrite depletion was much faster in inoculated samples. Protein-bound nitrite levels were higher in meat with added ionic iron. In cured meat with added ionic iron, iron-NO-protein complexes may form, lowering the amount of nitric oxide (NO) available to inhibit botulinal spore outgrowth.     

Nitric Oxide Complex of Iron(II) Myoglobin Converted from Metmyoglobin by Staphylococcus xylosus

With Staphylococcus xylosus FAX-1, metmyoglobin in MRS broth (pH 5.8) was found to undergo conversion to hexacoordinate nitric oxide (NO) complex of Fe(II) myoglobin. When the pH of the MRS culture containing myoglobin changed from 5.8 to 4.0, it affected the conversion from hexacoordinate to pentacoordinate NO complex of Fe(II) myoglobin. This conversion process was reversible. Salami without nitrite or nitrate addition was prepared by inoculating S. xylosus FAX-1, and pentacoordinate NO complex of Fe(II) myoglobin (nitrosylmyoglobin formed in cured meat) was formed in the salami.     

Carbon Monoxide as a Colorant in Cooked or Fermented Sausages

ABSTRACT:  The study aimed at substituting nitrite with carbon monoxide (CO) in cooked or fermented meat batter products by investigating color and color stability in myoglobin solutions, model meat systems, and full-scale hotdog and salami sausages of pork and beef. For cooked model meat systems and hotdogs at 75 to 80 °C core temperatures, direct flushing with a 1% CO gas mixture during the last stage of the batter chopping produced an initial red color equal to nitrite or more intense than with nitrite. For fermented model meat systems and salami sausages with a final pH of 4.7, pretreatment and storage of ground raw meat in a 1% CO mixture, and later use of the pretreated meat in batters, also formed an initial red color in the final products. The color stability during air and light display of cooked and fermented meat products with CO was inadequate compared to products with nitrite, although the red color of CO products was largely maintained by anaerobic packaging and storage. Spectra of carboxy- and nitrosomyoglobin at pH 4.7 demonstrated higher absorbance for carboxymyoglobin.     

Irradiation effects on meat color - a review

Color changes in irradiated fresh meat occur because of the susceptibility of the myoglobin molecule, especially the iron, to alterations in the chemical environment and to energy input. The potential for iron electrons to exist in various states makes the environment adjacent to the iron atom particularly vulnerable to the presence of electron-donating compounds and high energy inputs (irradiation). Initial condition of the myoglobin (Fe(++)O(2), Fe(+++)), modification of oxidation-reduction potential of the tissue, and generation of ligand-forming compounds (CO) from endogenous organic compounds and water are enhanced or suppressed depending on the gas atmosphere, temperature, pH and myoglobin concentration of the system. Generation of stable red pigments or brown pigments which become red over time appears to be due to binding of irradiation-generated reactive oxygen species (()O(2)(-)) or gasses (CO) which become ligands bound by iron under altered reducing conditions. Rapid generation of large amounts of metmyoglobin when irradiation is conducted in an oxygen-containing environment appears to be an acceleration of the normal process by which myoglobin undergoes oxidation. Generation of green pigments appears to be due to breakdown of the porphyrin integrity and/or formation of sulfmyoglobin. Maintenance of ideal meat color during irradiation can be enhanced by various combinations of pre-slaughter feeding of antioxidants to livestock, optimizing the condition of the meat prior to irradiation, addition of antioxidants, gas atmosphere (MAP), packaging, and temperature control.     

Effect of water-soluble haem and non-haem iron complexes on lipid oxidation of heated muscle systems

The distribution of soluble iron between three main components—ferritin, haemoglobin plus myoglobin and a low molecular weight fraction—and the pro-oxidant activities of each fraction in heated water-washed muscle systems from beef, pork and mackerel was determined. Over 75% of soluble iron was associated with the haem fraction. Even so, in all meat systems, the low molecular weight fraction was the major catalyst for lipid oxidation. Ferritin contributed significantly to lipid oxidation in all systems. The addition of nitrite significantly inhibited oxidation in all systems.     

Nitrite,nitrite alternatives,and the control of clostridium botulinum in cured meats

Historically, nitrite has been a component of meat‐curing additives for several centuries. In recent years the safety of nitrite as an additive in cured meats has been questioned mainly because of the possible formation of carcinogenic nitrosamines. Nitrite has many important functions in meat curing including its role in color development, flavor, antioxidant properties, and antimicrobial activity. The inhibition of Clostridium botulinum growth and toxin production is an especially important antimicrobial property of nitrite. This review discusses the effects of processing, curing ingredients (especially nitrite), and storage of cured meats in relation to the control of C. botulinum. If nitrite is eliminated from cured meats or the level of usage decreased, then alternatives for the antibotulinal function of nitrite need to be considered. Several potential alternatives including sorbates, parabens, and biological acidulants are discussed.     

Lipid Stability of Beef Model Systems with Heating and Iron Fractions

Catalytic effects of different temperatures (55, 70, 85, and 100°C) on lipid oxidation were studied in aqueous- and chloroform/methanol-extracted beef model lipid systems containing iron forms inherent in beef (water-extractable, diffusate, nondiffusate, ferritin, myoglobin, hemoglobin), hematin, FeCl₂, or FeCl₃. Heating increased thiobarbituric acid and peroxide values in both systems. All forms of iron catalyzed lipid oxidation in aqueous systems, with greatest oxidation by heme and low molecular weight iron fractions. Oxidation in lipid extracts was not increased by ferritin, FeCl₂, or FeCl₃, but heme iron was the major oxidation catalyst. Lipid stability decreased with addition of any iron forms inherent in beef or with increased heating, which helps understanding of rapid oxidation of meat during refrigerated storage or after cooking.     

Nitrite, nitrite alternatives, and the control of Clostridium botulinum in cured meats

Historically, nitrite has been a component of meat-curing additives for several centuries. In recent years the safety of nitrite as an additive in cured meats has been questioned mainly because of the possible formation of carcinogenic nitrosamines. Nitrite has many important functions in meat curing including its role in color development, flavor, antioxidant properties, and antimicrobial activity. The inhibition of Clostridium botulinum growth and toxin production is an especially important antimicrobial property of nitrite. This review discusses the effects of processing, curing ingredients (especially nitrite), and storage of cured meats in relation to the control of C. botulinum. If nitrite is eliminated from cured meats or the level of usage decreased, then alternatives for the antibotulinal function of nitrite need to be considered. Several potential alternatives including sorbates, parabens, and biological acidulants are discussed.     

Citrus Co-Products as Technological Strategy to Reduce Residual Nitrite Content in Meat Products

ABSTRACT:  Sodium or potassium nitrite is widely used as a curing agent in cured meat products because it inhibits outgrowth and neurotoxin formation by  Clostridium botulinum , delays the development of oxidative rancidity, develops the characteristic flavor of cured meats, and reacts with myoglobin and stabilizes the red meat color. As soon as nitrite is added in the meat formulation, it starts to disappear and the nitrite that has not reacted with myoglobin and it is available corresponds to residual nitrite level. Health concerns relating to the use of nitrates and nitrites in cured meats (cooked and dry cured) trend toward decreased usage to alleviate the potential risk to the consumers from formation of carcinogenic compounds. Recently, some new ingredients principally agro-industrial co-products in general and those from the citrus industry in particular (albedo [with different treatments], dietetic fiber obtained from the whole co-product, and washing water used in the process to obtain the dietetic fiber) are seen as good sources of bio-compounds that may help to reduce the residual nitrite level in meat products. From these co-products, citrus fiber shows the highest potential to reduce the residual nitrite level, followed by the albedo and finally the washing water. The aim of this article is to describe the latest advances concerning the use of citrus co-products in meat products as a potential ingredient to reduce the nitrite level.     

The antioxidant activities of nitrite and nitrosylmyoglobin in cooked meats

Low concentrations of nitrite (20 mg/kg) caused significant (p < 0·001) inhibition of lipid oxidation, measured by the TBA test, in a cooked muscle system and 50 mg/kg nitrite resulted in a highly significant (p < 0·001) reduction in TBA values. Similar antioxidant effects of nitrite were observed in heated water-extracted pork muscle systems catalysed by 5 mg/g metmyoglobin (Mb) or 5 mg/kg Fe(2+), Cu(2+) or Co(2+). The cured meat pigment, nitrosylmyoglobin per se exhibited significant (p < 0·05) antioxidant effects in pork muscle systems catalysed by Mb or metal ions. Progressive depletion of nitrite occurred during refrigerated storage of heated and unheated nitrite-treated pork muscle, muscle aqueous extract and in systems containing Mb, Cu(2+) ot Co(2+). Nitrite depletion occurred much more rapidly in Fe(2+)-containing systems and nitrite concentration had decreased to 5% of the original concentration immediately after heating. In addition, nitrite caused a significant (p < 0·05) reduction in the concentration of non-haem iron in heated aqueous-extracts of beef muscle, whereas, in nitrite-free extracts, a highly significant (p < 0·001) increase in the concentration of non-haem iron, probably due to heat denaturation of the haem structure with release of iron, was observed. Based on the results of this study, three co-operative mechanisms for the antioxidative activities in meat are proposed: (a) by the formation of MbNO which has antioxidant properties per se, (b) on heating, MbNO forms a stable complex, nitrosylhaemochrome, which blocks the catalytic activity of haem iron and also prevents release of haem iron as non-haem iron, which is a highly effective catalyst and (c) nitrite appears to 'chelate' non-haem iron-and possibly copper and cobalt-forming a stable complex, thus inhibiting catalytic activity.     

Color, Pigment and Iron Content of Meat Loaves with Blood, Blood Emulsion, or Mechanically Deboned Meat Added

The hemoglobin, myoglobin, and iron content in slaughter-house blood, a blood emulsion, mechanically deboned meat (MDM) and meat minces with these raw materials added, were determined. Blood and blood emulsion were added in the range 0-5%. MDM was added as a substitute for ground beef. The pigment and iron content in the minces were compared with the surface color of heated meat loaves produced. Minces containing blood yielded lighter meat loaves upon heating (i.e. a higher lightness L* was observed) than did minces containing blood emulsion or MDM, even if the amounts of extractable hemoglobin and myoglobin in the minces were equal. At equal concentrations of iron, minces with blood and MDM gave lighter meat loaves than did minces to which blood emulsion had been added.     

Blood Fraction Effects on the Antibotulinal Efficacy of Nitrite in Model Beef Sausages

Toxin production by Clostridium botulinum was studied in a model cured beef sausage containing 0–5% added dried bovine blood fractions. Controls which contained nitrite but without blood fractions, were toxic by bioassay in 3 wk at 27°C. Controls without nitrite or blood fractions were toxic at 1 wk, while model sausages supplemented with hemoglobin, red cells, or whole blood were toxic in 1–3 wk. Plasma yielded no detectable toxin for 4 to > 10 wk depending upon addition level. Sausages showing delayed toxigenesis had pH values lower than those which developed toxin earlier. These results demonstrated that use of blood fractions that increased iron levels in beef above 30 μ.g/g interfered with the antibotulinal efficacy of sodium nitrite.     

Relationship between nitrate/nitrite reductase activities in meat associated staphylococci and nitrosylmyoglobin formation in a cured meat model system

Quantitative determination of catalase, nitrate reductase, nitrite reductase and nitric oxide synthase activities (NOS) was performed on 11 different bacterial strains, mainly staphylococci, isolated from fermented sausages, bacon brine or cured meat products. All except one strain possessed catalase activity in the range from 1.0 to 6.1 μmol min^− 1 ml^− 1. Ten out of 11 bacteria strains showed nitrate reductase activity in the range between 50 and 796 nmol min^− 1 ml^− 1 and nine showed nitrite reductase activity in the range between 6 and 42 nmol min^− 1 ml^− 1. No evidence of NOS activity of the selected strains was detected. In a colour formation assay containing myoglobin all strains affected nitrosylmyoglobin (MbFe^IINO) formation in assays containing nitrite, whereas only strains having nitrate reductase activity generated MbFe^IINO in assays containing nitrate as the sole nitrosylating agent. The quantitative nitrate and nitrite reductase activity did not fully explain or correlate well with the observed rate of formation of MbFe^IINO, which seemed to be more affected by the growth rate of the different strains. The mechanism of the reduction of nitrite into NO of strains not having nitrite reductase activity remains to be fully elucidated, but could be due to a dual-mode action of nitrate reductase capable of acting on nitrate.     

Effect of Nitrite and Sorbate on Total Number of Aerobic Microorganisms in Chicken White and Dark Meat Patties

The effects of nitrite, sorbate and combinations of these two ingredients plus salt on the number of aerobic microorganisms in chicken white and dark meat patties were examined. All patties were stored at 4–5°C for 12 days. Nitrite concentrations of 400 and 2500 ppm were effective in preventing bacterial growth in chicken white meat patties while 2500 ppm was required to prevent growth in dark meat. A reduction in bacterial growth (4–10 days) was demonstrated with 100 and 150 ppm nitrite in chicken white meat patties. Four hundred ppm nitrite reduced bacterial growth in dark meat patties for 6 days. In white meat patties, the use of sorbate (0.26%) in combination with nitrite (40 ppm) was as effective in reducing bacterial growth as higher concentrations of nitrite, salt (2.5%) alone or in combination with sorbate (0.26%) or nitrite (40 ppm) inhibited bacterial growth in both white and dark meat patties during the first 4–6 days of storage.     

Nitric oxide inhibits the formation of zinc protoporphyrin IX and protoporphyrin IX

The aim of this study was to elucidate the mechanism by which curing agents, especially nitrite, inhibit the formation of zinc protoporphyrin IX (ZPP) in dry-cured hams such as Parma ham. The oxidation–reduction potential of model solutions was increased by the addition of nitrite, but it was not clear whether the formation of ZPP is inhibited by the oxidizing property of nitrite. The effect of nitric oxide (NO) produced from nitrite on the formation of ZPP was examined. The amount of ZPP formed was decreased by the addition of NO donors. The amount of protoporphyrin IX (PPIX), which is the precursor of ZPP, was also decreased by the addition of NO donors. It is concluded that NO produced from nitrite inhibited the formation of PPIX and ZPP was therefore not formed in cured meat products with the addition of nitrite or nitrate.     

Prevention of Clostridium outgrowth in heated and hermetically sealed meat products by nitrite – a review

This paper discusses several hypotheses regarding the inhibition of Clostridium in cured meat. It focuses on the binding of iron, as there is some evidence that inhibition of Clostridium botulinum outgrowth in nitrite-cured meat products is mainly due to iron binding in such a way that it is no longer available for outgrowth of Clostridium spores. This strong binding also explains the antioxidative properties of nitrite in these products. The binding of iron, in all probability, requires no more than a moderate amount of nitrite which, after binding as nitric oxide to the haem group, is unavailable for reactions leading to N-nitroso compounds either. Other reactions of nitrite, which also could lead to Clostridium inhibition, are not discussed in detail here.     

The effect of the initial gas volume to meat weight ratio on the storage life of chilled beef packaged under carbon dioxide

Normal pH (5·5-5·7) and high pH (>6·0) beef cuts of 400 g were vacuum packaged in polyvinylidine chloride (PVDC) laminate, in aluminium foil laminate or in foil laminate with a CO(2) scavenger (Ba(OH)(2)); or were packaged under CO(2) in foil laminate with CO(2) added at 200, 400, 700, 1000 or 2000 ml per kg of meat. The process used to prepare meat before packaging resulted in an initial flora containing a high fraction of eneterobacteria but with undetectable numbers of lactobacilli. During storage at +1°C, all vacuum packaged meat developed floras containing substantial fractions of enterobacteria. The enterobacteria fractions were larger on high pH than on normal pH meat, and on normal pH meat in PVDC or in foil laminate plus CO(2) scavenger packs than on normal pH meat in foil laminate without the CO(2) scavenger. All vacuum packaged meat was spoiled by putrid flavours, high pH meat at 7 weeks and normal pH meat at 12 weeks. Increasing amounts of added CO(2) progressively retarded the development of putrid spoilage, as growth of the spoilage floras was slowed while the relative numbers of lactobacilli in the floras were enhanced. With the two largest amounts of added CO(2), spoilage was delayed until 15 weeks and 21 weeks for high pH meat and normal pH meat, respectively, and enterobacteria were detected in the spoilage floras only at later sampling times. With lesser amounts of added CO(2), enterobacteria persisted or re-emerged at earlier times, in the floras.     

Zn-porphyrin formation in cured meat products: Effect of added salt and nitrite

Zn-porphyrin (Zn-pp) was quantified by fluorescence spectroscopy in the cured and dry cured meat products: Parma ham, Iberian ham, dry-cured ham with added nitrite, cooked ham with added nitrite, raw ham meat, raw bacon and Karree-Speck. The highest amount of Zn-pp was found in dry-cured Parma ham and Iberian ham, while the use of nitrite as curing agent was found to inhibit completely the formation of Zn-pp in meat products. A positive correlation between both Zn content and Fe content and the logarithmic transformed Zn-pp content (measured as fluorescence intensity I_fl) was found for the different cured and dry cured meat products, with correlation coefficients of 0.79 (p < 0.001) and 0.71 (p < 0.01), respectively. Log I_fl correlates best with the Zn content, indicating that the formation of Zn-pp is proportional to the Zn content. A model system with vacuum packed pork in brine with different added levels of sodium chloride with or without nitrite and Zn acetate was investigated in order to further elucidate the mechanism of Zn-pp formation. Zn-pp increased with time (up to 42 days investigated) in non-cured meat and for meat cured solely with NaCl lower than 9%. Addition of nitrite or Zn(II) in the curing brine was found to inhibit formation of Zn-pp confirming the observations from the various cured meat products. It is suggested that a chloride anion assisted dissociation of iron from myoglobin could be rate-determining for Zn-pp formation in meat products.     

Lipid oxidation and metmyoglobin formation in sausages containing chickpea flour

Rapid lipid oxidation and metmyoglobin formation in sausages containing up to 30% chickpea flour is due to the presence of lipoxidase in chickpea flour. This enzyme oxidises the unsaturated fats present to peroxides (or related compounds) which then catalyse myoglobin oxidation. Heat treatment of chickpea flour at 80° C for 1 h in water prior to its addition to the batter will prevent both accelerated lipid and myoglobin oxidation in these sausages. An antioxidant containing α-tocopherol and ascorbyl palmitate inhibited the lipid oxidation in these products but had no effect on myoglobin oxidation in sausage batter containing unheated chickpea flour. The relevance of these results to the interdependence of lipid and myoglobin oxidation in meat and meat products is discussed.     

Microbial formation of nitrite-cured pigment,nitrosylmyoglobin, from metmyoglobin in model systems and smoked fermented sausages by<Emphasis Type="Italic"> Lactobacillus fermentum</Emphasis> strains and a commercial starter culture

Two Lactobacillus fermentum strains (JCM1173 and IFO3956) were evaluated for their ability to generate nitrosylated derivatives of myoglobin either in broth media or fermented sausages. For comparison, a commercial starter culture was also included. All bacteria species investigated converted brown metmyoglobin into red myoglobin derivatives when incubated separately in broth, but only the two lactobacilli showed a signal for nitrosylmyoglobin as measured by electron spin resonance spectroscopy. In smoked sausages with added bacteria culture the highest amount of nitrosylmyoglobin was observed in the centre of sausage with added L. fermentum, but colour formation in sausages with 60 ppm of nitrite added was more pronounced. An outer peripheral zone of all fermented sausages contained levels of nitrosylmyoglobin comparable to nitrite-cured sausages. Nitrogenous gasses from smoke may, however, cause this zone to be formed. Depending on a further optimisation of the processing parameters, the bacteria's ability to generate NO could form the basis for production of cured meat products without the use of nitrite/nitrate.