Opinion of the scientific committee on food on the use of carbon monoxide as component of packaging gases in modified atmos.

EUROPEAN COMMISSIONHEALTH & CONSUMER PROTECTION DIRECTORATE-GENERAL Directorate C - Scientific Opinions
C2 - Management of scientific committees II; scientific co-operation and networks
Scientific Committee on Food
of the Scientific Committee on Food
on the use of carbon monoxide as component of packaging gases in modified
atmosphere packaging for fresh meat
Telephone: direct line (+32-2) 295.4861, switchboard 299.11.11. Fax: (+32-2) 299.4891 Telex: COMEU B 21877. Telegraphic address: COMEUR Brussels http://europa.eu.int/comm/food/fs/sc/scf/index_en.html Opinion of the Scientific Committee on Food on the use of carbon monoxide as
component of packaging gases in modified atmosphere packaging for fresh meat
Terms of Reference
The Committee was asked to evaluate the safety of carbon monoxide as a packaging gasfor meat in a mixture with carbon dioxide and nitrogen.
Background
The Norwegian Meat Co-operative and the Norwegian Independent Meat Association,representing the Norwegian meat industry, have applied for the use of a gas mixturecontaining 60%-70% carbon dioxide (CO2), 30%-40% nitrogen (N2) and <0.5% carbonmonoxide (CO) as components of the packaging gas in modified atmosphere packaging(MAP) for fresh red meat (mainly beef, pork, lamb but also horse, goat, reindeer, gameetc.). The high level of CO2 inhibits growth of many pathogenic and non-pathogenic micro-organisms thus giving an extended microbiological shelf life. The presence of COprovides a stable cherry red colour of the meat. This MAP is used in Norway to prolong shelf life for displaying and selling fresh retailmeat in all parts of the country (Norwegian Food Control Authority, 2001).
In interpreting the Terms of Reference, the Committee considered both microbial andtoxicological aspects of safety. Technological aspects
The CO used in the MAP has a degree of purity higher than 99.3%, the major impuritiesbeing oxygen and argon, hydrogen and nitrogen, carbon dioxide, water and totalhydrocarbons. The gas is supplied as pre-mixtures in gas containers to prevent potentialhazards from occupational exposure to CO. The proposed gas mixture for fresh meatpackaging is as follows: 60%-70% CO2, 30%-40% N2, and 0.3%-0.5% CO (highCO2/low CO).
Two pigments, myoglobin and haemoglobin, contribute principally to the colour of freshmeat. Myoglobin represents about 80%-90% of the pigment in meat, while haemoglobin,catalase and cytochromes contribute the remainder. The ultimate colour depends on theoxidation state of the Fe-atom in the non-protein haem ring and the compound bound tothe iron at the free binding site. The relatively stable oxygenated oxymyoglobin with the structure {globin-tetrapyrrole ring-Fe+2(O2)} is bright red, myoglobin, with the structure{globin-tetrapyrrole ring-Fe+2(H2O)} inside the reduced atmosphere of the muscle cell, ispurple, the oxidised metmyoglobin with the structure {globin-tetrapyrrole ring-Fe+3(OH)}, is brown. CO ligates to the free binding site on the Fe-atom of the haem to form the cherry redcarboxymyoglobin with the structure {globin-tetrapyrrole ring-Fe+2(CO)}. High partial pressure of oxygen favours the formation of oxymyoglobin for use in themetabolism of the muscle cell while low partial pressure of oxygen favours the formationof myoglobin and metmyoglobin. Carboxymyoglobin is more resistant to oxidation thanoxymyoglobin because of the stronger binding of CO to the Fe-binding site on themyogobin molecule.
Shelf life aspects
Either vacuum packing or MAP improves shelf life of fresh meat. End-productcharacteristics affecting the shelf life depend among others on type of product, initialcontamination, atmosphere, storage temperature, packaging material and design (Churchand Parsons, 1995). The major spoilage flora of fresh meat consists of aerobicPseudomonas spp. This flora is inhibited by the anaerobic conditions in vacuum or non-oxygen MAP systems or due to CO2 concentrations exceeding 10-20% in oxygen MAP(Gill and Molin, 1991; White and Roberts, 1992). The shift in ecology from a gram-negative aerobic spoilage flora to a facultative gram-positive flora of Brochotrixthermosphacta and especially Lactobacillus will exhibit a certain competition towardspathogens that might be associated with fresh meat. A disadvantage with vacuum packing or anaerobic MAP for fresh meat is the change inmeat colour. A mixture of CO2 and oxygen is usually used in MAP to avoid this change.
CO2 is used for its inhibitory effect on the gram-negative spoilage flora and oxygen as astabiliser of the red colour. The shelf life during this storage is not as predominantcompared to anaerobic storage in CO2 (Dainty and Mackey, 1992). Studies were carried out to evaluate the effect of high carbon dioxide/low CO MAP onthe shelf life of fresh meat and meat products under MAP conditions (Sørheim et al.,1997; Sørheim et al., 1999; Nissen et al., 1999).
In comparative studies the shelf lives of fresh meat products packaged in high CO2/lowCO gas mixtures were evaluated against products packaged in alternative MAP gasmixtures having following composition: 70% O2/30% CO2 and 60% CO2/40% N2 with anO2 absorber. The products were stored in the dark at 40C or 80C for up to 21 days.
Meat in the high CO2/low CO mixtures had a stable bright red colour. The shelf lives at40C in this gas mixture were, 11 days for ground beef, 14 days for beef loin steaks and 21days for pork chops. After these storage times off-odours developed. The samples stored under high O2 showed initially a bright red colour of the meat, but the colour wasunstable and off-odours developed rapidly after shorter storage periods of 8, 10 and 14days respectively. Increasing the storage temperature up to 80C reduced the shelf life,under both MAP conditions, to nearly half that at 40C.
The observed off-odours were probably caused by the growth of Brochothrixthermosphacta. Indeed, at chill temperatures above 10C, B. thermosphacta often causesspoilage of meat stored in high O2 atmospheres (Dainty and Mackey, 1992). Highconcentrations of CO2, removal of O2 and low temperatures inhibit the growth of B.
thermosphacta (Gill, 1996; Nissen et al., 1996). Meat in high O2 is often spoiled byPseudomonas spp., but the growth of pseudomonads is retarded under anaerobicconditions (Dainty and Mackey, 1992; Gill, 1996). A shift in the metabolism of lacticacid bacteria under anaerobic conditions can also produce off-odours (Nissen et al.,1996). In the studies that were considered here, the number of coliforms or Escherichiacoli did not exceed 103 colony forming units (cfu)/g in any samples and thus wereprobably not involved in off-odour production.
Microbiological aspects
The inclusion of CO in MAP is controversial because the stable cherry-colour can lastbeyond the microbial shelf life of the meat and thus mask spoilage (Kropf, 1980). Theextended shelf life obtained by MAP may, therefore, under certain conditions implyincreased risk of growth of pathogens (Silliker and Wolfe, 1980; Hintlian and Hotchkiss,1986; Farber, 1991; Lamberts et al., 1991).
Meat packed in high CO2/low CO acquires a stable colour and the shelf life at 40C, basedon odour, is significantly longer than in the other gas mixtures. At this temperatureYersinia enterocolitica and Listeria monocytogenes are considered to be the most seriouspathogens in meat. At abuse temperatures (>80C) E. coli 0157:H7 and Salmonella spp.
also may grow and increase the health risk of the consumer. This issue was discussed bythe Scientific Committee Food (SCF Reports, 1997).
In studies submitted by the petitioner (Sørheim et al., 1997; Sørheim et al., 1999; Nissenet al., 1999; Nissen et al., 2000) growth of the pathogens Y. enterocolitica, L.
monocytogenes, E. coli 0157:H7 and strains of Salmonella was compared in ground beefpacked in high CO2/low CO, in a high O2 mixture (70% O2/30% CO2) and in chub packs.
Ground beef was chosen because it is considered as a risk product since pathogens maybe mixed into the product and not be properly heated before being eaten.
Ground beef was inoculated with rifampicin- or nalidixic acid/streptomycin-resistantstrains (to aid their recovery) at a final concentration of 102 – 103 bacteria/g. Packs werestored at 40C or 100C for up to 14 days.
At 40C shelf life measured on the basis of colour stability and a low background flora,was prolonged for the high CO2/low CO MAP compared with the other packs. At 100Cthe shelf life was below 8 days for all packs.
The growth of Y. enterocolitica was totally inhibited at both 40C and 100C in the highCO2/low CO mixture, while the bacterial numbers in the samples packed in the high O2mixture increased from about 5.102 bacteria/g at day 0, to about 104 on day 5 at 40C andto 105 by day 5 at 100C.
L. monocytogenes showed very little growth at 40C in all treatments. At 100C it grewfrom about 5.103 bacteria/g to about 104 by day 5 in the high CO2/low CO mixture, whilethe numbers in the high O2 and the chub packs were about 10 times higher.
E. coli 0157:H7 does not grow at 40C but even at 100C in ground beef it was almosttotally inhibited in both high CO2/low CO mixture and in the high O2 mixture. In thechub packs growth was much higher reaching 105 bacteria/g on day 5.
Salmonella spp. also do not grow at 40C but at 100C the pathogens S. typhimurium, S.
dublin, S. enteritidis and S. enterica 61:k:1,5,(7) grew better by days 5 and 7 in the highCO2/low CO MAP than the high O2 MAP (Nissen et al., 1999; Sørheim et al., 1999).
The data presented in the studies show that the prolonged shelf life at 40C did notincrease growth of Y. enterocolitica and L. monocytogenes in ground beef stored in thehigh CO2/low CO mixture. However, the observed growth of strains of Salmonella at theabuse temperature of 10ºC in this mixture and in the chub packs does stress theimportance of temperature control during storage.
History of use
The MAP packaging of fresh meat using high CO2/low CO mixtures has been in use inNorway since the mid eighties. Currently 50-60% of retail meat and up to 85% of groundbeef is packaged under such conditions (Norwegian Food Control Authority, 2001).
The Norwegian Food Control Authority has not registered outbreaks or a higherfrequency of sporadic cases of food-borne diseases linked to such products during thistime (Norwegian Food Control Authority, 2001). Toxicological aspects
CO is present in the normal atmosphere, mainly through the incomplete combustion ofcarbon-containing materials, the oxidation of methane in the troposphere and from thedecay of chlorophyll. The natural background levels of CO are 0.01–0.9 mg/m3. In urban areas, 8-h mean concentrations of Cre generally <20 mg/m3. However, maximum 8-hconcentrations of up to 60 mg/m3 have been reported (WHO, 1979). CO is eliminated from the atmosphere through oxidation to CO2 by hydroxyl radicals inthe upper layers of the atmosphere and through oxidation by soil bacteria. CO is formed in the human body through oxidation of the carbon of the methylene bridgein the tetrapyrrole ring in haemoglobin or myoglobin. (Marquardt & Schäfer, 1994). Byfar the most common cause of elevated CO concentrations in the blood is tobaccosmoking (WHO, 1987).
The toxic action of CO is due to the blockage of the oxygen-carrying function ofhaemoglobin through the formation of carboxyhaemoglobin (HbCO) instead ofoxyhaemoglobin (HbO2). The binding of CO to haemoglobin is reversible, with a half-life of ~4.5 h in individuals at rest.
A small amount of CO is formed naturally in the human body from the breakdown ofhaemoproteins. Such production leads to a HbCO concentration of ~0.5% of totalhaemoglobin. The average HbCO concentration in non-smokers is 1.2-1.5%. In smokersthe concentration is in the range from ~3 - 4% (Aunan, 1992). Concentrations above 2% have been observed to have adverse effects ranging fromreduced attention to anoxia and death at concentrations of 30%-50% or more of the totalhaemoglobin (Marquardt & Schäfer, 1994). Concentrations of HbCO below 2% of thetotal haemoglobin do not have any measurable adverse effects in humans. It is thereforeaccepted that, to protect the most vulnerable section of the population, the level of HbCOshould not exceed 1.5% (Coburn et al., 1965).
The mean normal air inhalation of an adult in 24 hr is ~15 m3 (or ~0.625 m3/hr). In orderto prevent a maximum HbCO concentration level in the blood of 1.5% being exceeded,the CO concentration in air for a 1 hr period of moderate physical activity should notexceed 24 mgCO/m3 (Aunan, 1992) Exposure aspects
Very little information exists in the literature on the exposure to CO followingconsumption of meat that has been treated with CO gas. The exposure of beef to anatmosphere containing 1% CO for 3 days resulted in ~30% saturation of the meatmyoglobin (Watts et al., 1978). a mean CO concentrations measured for each possible 8 hr interval during a 24 hr period, then averaged In the absence of carbon monoxide, CO is lost, from previously CO-treated meat duringstorage, with a half-life of ~3 days. When further cooked at 195S only 0.1 mg CO/kgmeat remains. This amounts to a loss on cooking of ~85% (Watts et al., 1978). Fresh packaged meat packaged stored in high CO2/lowCO MAP could contain, next toendogenous CO, an additional 0.7 mg CO/kg meat. This is a worst case estimate as itdoes not take into account losses on cooking which may be up to 85% of the CO boundto carboxymyoglobin and carboxyhaemoglobin present in the packaged meat. An assumed consumption of 250 g fresh meat/24 hrs, could therefore release 0.18 mg CO(equivalent to 0.018 % HbCO) on digestion in the gut. Assuming 100% transfer of COfrom the gut to the blood and complete transformation to HbCO, only a negligibleamount of HbCO would be added to the 0.5% HbCO, resulting from endogenous COproduction, and the 0.7%-1.0% HbCO formed from inhalation of urban air by a non-smoker. These figures are deduced from the observation that an intake of 15.1 mg CO/hr throughinspired air leads to the formation of 1.5% HbCO (Sørheim et al., 1997).
Exposure through inhalation of headspace gas on opening a package of meat with a MAPcontaining 0.3%-0.5% CO would equally contribute insignificantly to the HbCO in theblood when compared to the other sources of inhalation of CO.
Conclusion
Meat packaged in MAP containing a high concentration of CO2 and 0.3%-0.5% COremains microbiologically stable for 11-21 days when stored at a maximum temperatureof 4°C. High CO2/low CO gas mixtures inhibit the growth of L. monocytogenes, Y. enterocoliticaand E. coli O157:H7 during storage at 4°C, however some strains of Salmonella willgrow at 10°C. Thus, close control of temperature throughout packaging, distribution,retailing and storage by the consumer of MAP products is essential.
The use of meats packaged in MAP containing 0.3%-0.5% CO contributes in negligibleamounts to the overall exposure to CO and the HbCO concentration in humans.
The Committee therefore concluded that there is no health concern associated with theuse of 0.3%-0.5% CO in a gas mixture with CO2 and N2 as a modified atmospherepackaging gas for fresh meat provided the temperature during storage and transport doesnot exceed 4°C. However the Committee wishes to point out that, should products bestored under inappropriate conditions, the presence of CO may mask visual evidence ofspoilage. References
Aunan, K. (1992). Effects of ambient air pollution on the health and environment – Air Report No. 92:16, pp. 154-170. State Pollution Control Authority (SFT), Oslo, Norway Church, I.J. and Parsons, A. L. 1995. Modified Atmosphere Packaging Technology: A Review. J. Sci. Food Agric. 67, 143-152. Coburn, R.F., Forster, R.E., Kane, P.B. (1965). Considerations of the physiological variables that determine the blood carboxyhaemoglobin in man. J. Clin. Invest.,44,1899-1910.
Dainty, R.H. and Mackey, B.M. (1992). The relationship between the phenotypic properties of bacteria from chill-stored meat and spoilage processes. J. Appl.
Bacteriology Symp. Suppl., 73, 103S-114°C. European Commission, (1997). The microbiological safety of Modified-Atmosphere Packaged (MAP) and Controlled-Atmosphere Packaged(CAP) Foods. SCF Reports,40th Series, pg 51-62.
Farber, J.M. (1991). Microbial aspects of modified-atmosphere packaging technology – a Gill, C. D. and Molin, G. (1991). Modified atmosphere and vacuum packaging. In: Food Preservatives. Ed. Russel N. J. & Gould, G. W. Blackie, London. UK, pp 172-199. Gill, C.O. (1996). Extending the storage life of meats. Meat Science, 43(suppl.), p99-109.
Hintlian, C.B. and Hotchkiss, J.H. (1986). The safety of modified atmosphere packaging: Kropf, D. H. (1980). Effects of retail display conditions on meat colour. Proc. Reciprocal Lamberts, A.D., Smith, J.P. and Dodds, K.L. (1991). Shelf life extension and microbial safety of fresh meat – a review. Fd. Microbiology, 8, 267-297.
Norwegian Food Control Authority, (2001). Application for assessment of carbon monoxide as component in packaging gases (Modified Atmospheres) for meatproducts. Report submitted to the EU Commission, 09.06.2000.
Marquardt, H. and Schäfer, S.G. (1994). Lehrbuch der Toxikologie, B.I.Wiss.-Verl., Nissen, H, Sørheim, O. and Dainty, R. (1996). Effects of vacuum, modified atmospheres and storage temperature on the microbial flora of packaged beef. Fd. Microbiology,13, 183-191.
Nissen, H., Alvseike, O., Bredholt, S., Holck,A., Nesbakken,T. (1999). Packaging of ground beef in an atmosphere with low carbon monoxide and high carbon dioxiderestrains growth of E. coli O157:H7, L. monocytogenes, Y. enterocolitica, Salmonelladiarizonae. In Proc. 17th Intern. Conf. of ICMFH, 13-17.9.1999, Veldhoven, pg. 285-286.
Nissen, H., Alvseike, O., Bredholt, S., Holck,A., Nesbakken,T. (2000). Comparison between the growth of Yersinia enterocolitica, Listeria monocytogenes, Escherichiacoli O157:H7 and Salmonella spp. in ground beef packed by three commercially usedpackaging techniques. Int. J. Food Microbiology 59, 211-220. Silliker, J.H. and Wolfe, S.K. (1980). Microbiological safety considerations in controlled-atmosphere storage of meats. Fd. Technology, 34, 59-63.
Sørheim, O., Nissen, H., Nesbakken, T. (1999). The storage life of beef and pork packaged in an atmosphere with low carbon monoxide and high carbon dioxide. MeatSc., 52, 157-164 Sørheim, O., Aune, T., Nesbakken, T. (1997). Technological, hygienic and toxicological aspects of carbon monoxide used in modified-atmosphere packaging of meat. TrendsFd. Sc. Techn., 8, 107-112. Watts, D.A., Wolfe, S.K., Brown, W.D. (1978). Fate of 14C-carbon monoxide in cooked or stored ground beef samples. J. Agric. Food Chem., 26, 210-214.
White, R. and Roberts, R. (1992). Developments in Modified Modified Athmosphere and Chilled Foods Packaging. Pira International, Leatherhead, UK. WHO, (1979) Environmental Health Criteria. 13. Carbon Monoxide. World Health WHO, (1987) Air Quality Guide for Europe. World Health Organisation Regional Publications, European Series, No. 23. World Health Organisation, Geneva,Switzerland.

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