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OFF-FLAVOUR FORMATION IN HEAT STERILIZED MEAT IN TRAYS
OFF-FLAVOUR F O R M A T I O N IN H E A T STERILIZED M E A T IN TRAYS
S. Langourieux and F.E. Escher Department of Food Science, Laboratory of Food Chemistry and Food Technology, Swiss Federal Institute of Technology, Zurich (ETH), CH-8092 Zurich, Switzerland
1 INTRODUCTION Heat sterilization is one of the most important preservation techniques for producing microbiologically safe foods storable at room temperature. The increasing consumer demand for easy-to-use and smaller portions, and with a favourable geometry to reduce heat damage, has led to heat processed convenience foods packed in trays rather than in cans. Nevertheless, sterilization and storage still lead to a considerable loss of organoleptic and nutritional quality, due to oxidative and heat-induced changes, ' in particular if these products contain meat preparations. Off-flavours are developed via the autoxidation of polyunsaturated fatty acids and oxidative effects on proteins, peptides and amino acids. ^ Oxidative stability of various heat-processed menu components has been studied in a comprehensive project at the ETH, Zurich, in collaboration with the food canning industry. The influence of the atmosphere above the product and the influence of a natural antioxidant were particularly investigated. For meat products in aluminium trays, removal of air by flushing with nitrogen during filling associated with a storage temperature of 20 °C significantly increased the storage stability. But in this case, it also leads to the formation of a strong sulfurous off-flavour after the sterilization process. In turn, addition of a rosemary extract as an antioxidant resulted in a very effective stabilization of the product and no sulfurous off-flavour was detected sensorially when the trays were opened. Our objective in this work was first to investigate this off-flavour and then to determine the influence of the process parameters on its formation. 1
2 3
4
1
7
2 MATERIAL AND METHODS 2.1 Sample Preparation The meat-fat mixture was prepared according to Giintensperger and Escher. To this ground meat-fat mixture (90 g) was added 88 mg per kg fat of citric acid or 44 mg per kg fat of desaromatized rosemary extract (RE-S, FIS S.A., Switzerland) or samples were further processed without antioxidant. Deionized water (180 g) was added and the samples 1
*Present address: Laboratoire de Genie Biologique et Sciences des Aliments, Unite de Microbiologie et Biochimie industrielles, Universite de Montpellier II, F-34095 Montpellier cedex 5, France
Thermally Generated Flavour
195
were put into aluminium trays (Type 73300, 138 x 138 x 28 mm, W. Wagner GmbH, Germany). The trays were sealed with a peelable aluminium lid foil (Flexalpeel 45/1000, Alusingen, Germany) with or without prior removal of air by evacuating to 200 mbar and flushing back with nitrogen to atmospheric pressure (CPM H175VG, Packaging Machinery, England). This operation was repeated three times before sealing. The trays were then sterilized in hot water in a batch retort (Pilot Rotor 400, Stock GmbH, Germany) without agitation at 121 °C or 131 °C to an F value of 8-11 min. The trays were then stored in the dark at 20 °C for 14 hours or one month before analysis. To investigate the effects of the different experimental parameters on the response variable (headspace concentration of volatile compounds), an ANOVA analysis was performed (SAS v.6.10, SAS Institute Inc., U.S.A.). 0
2.2 Analysis of the Volatile Compounds of the Headspace 2.2.1 Purge and Trap of the Headspace. The headspace from each tray was collected (without opening the tray) using two needles. The first one purged the tray with purified nitrogen (20 ml min ) and the second one drove the headspace gas into a PTFE column (14 cm length, 0.5 cm internal diameter) packed with Tenax GR, (Alltech). After an hour of purging, the Tenax trap was removed and replaced by another and this purge lasted a further three hours. Diethyl ether (20 ml) was used for the extraction of the traps with 1 -butanethiol (Aldrich) and n-dodecane (Sigma) as internal standards. The total ether solution was further concentrated with a Kuderna-Danish concentrator to a volume of approximately 0.1 ml. No internal standard was added to the extracts for olfactometry. 2.2.2 Gas Chromatography (GC) and Gas Chromatography-Olfactometry (GC-O). A Hewlett-Packard model 5890 Series II gas chromatograph equipped with an HP-5 column (30 m length, 0.32 mm internal diameter, 0.25 mm film thickness) was used for the analysis of the extracts (1 ul). Nitrogen was used as the carrier gas (1.1 ml min ). The temperature programme started at 30 °C and increased at a rate of 1 °C min to 40 °C, then increased at a rate of 10 °C min" to 250 °C and held for 10 min. Injector, FID and FPD (flame photometric detector) temperatures were set at 220, 250, and 150 °C respectively. The gas chromatograph was also equipped with an olfactometric detector (SGE, Australia). The data were analysed according to the CHARM analysis with the help of software derived from Origin 3.5 (DMP Ltd., Switzerland). 2.2.3 Gas Chromatography-Mass Spectrometry Analyses (GC-MS). A Fisons GC 8000 series gas chromatograph equipped with the same column and operated at the same conditions as for the GC analysis and coupled to a Finnigan MAT SSQ710 mass spectrometer was used with helium as the carrier gas. The source, analyser and transfer line temperatures were 150, 70 and 220 °C respectively. The ionization voltage applied was 70 eV, the emission current was 0.2 mA and the electron multiplier voltage 1000 kV. Mass spectra obtained from a scan range of 33-250 m/z were compared with those of known compounds in the NIST library. 2.2.5 Analysis of Hydrogen Sulfide. Based on method of Jacobs et al. 2.2.6 Ethane Determination. As described by Giintensperger and Escher -1
-1
-1
1
11
9,10
1
3 RESULTS AND DISCUSSION 3.1 Determination of the Volatile Compounds Responsible of the Off-flavour A 2 ml sample of the headspace of a non-flushed and a nitrogen-flushed tray were directly injected and a G C - 0 analysis carried out. The results are given in Figure 1. Short
Flavour Science: Recent Developments
196
chain alkanes from CI to C6 and hydrogen sulfide were identified. No odour was detected for the 'C-tray' (no flushing) but a strong spoiled egg like odour was detected for the 'Ftrays' (flushing with nitrogen) at a retention of 1.8 min corresponding to hydrogen sulfide. The same odour was perceived when opening the 'F-trays'. C-' and 'F-trays' were also extracted using the purge and trap method and GC, G C MS and G C - 0 analysis were performed. CHARM profiles of C and F headspace samples are shown in Figure 2. CHARM profiles revealed differences between the two samples. Dimethyl trisulfide, 2-pentyl furan and octanal are important contributors to the odour FID
FPD
1E5 5000
40
— 10
Time (min)
0
Figure 1 FID and FPD chromatograms of 2 ml directly injected tray headspace (C: no removal of air before sealing; F: flushed with nitrogen)
••
1
Figure 2
20-
CHARM Response
*.
•• • *• • • • •• *
0
I
i
4
I
CHARM Response 8 12 i
l
i
I
i
I
i
I
i
l
i
16 I
n.i., butter 3-Methylbutanal, herbaceous 2-Ethylfuran, coffee, roast Dimethyl disulfide, onion Hexanal, herbaceous, pungent Heptanal, fresh, pungent n.i., biead, cooked potato Dimethyl trisulfide, fresh cut onion 2-Pentylfuran, fruity Octanal, fruity (orange-like), pungent 2-Octenal, honey n.i., fresh, light Heptanoic acid, mushroom Nonanal, fatty 2-Buryloctanol, fruity 2-Nonenal, waxy 2,4-Nonadienal, hay n.i., wood 2,4-Decadienal, hay, pungent 2-Undecenal, hay, earth n.i., fried onion
CHARM profiles of headspace purge and trap extracts of two different trays (C: no removal of air before sealing; F: flushed with nitrogen)
Thermally Generated Flavour
197
perceived in the 'C-trays' while only octanal persists in 'F-trays'. It can be concluded that hydrogen sulfide is mainly responsible for the off-flavour in the 'F-trays' but flushing with nitrogen also greatly modifies other odour active compounds. No typical roasted or meaty notes were found in the purge and trap extracts. This is due to the fact that the samples were sterilized without prior pre-cooking and in the presence of a large proportion of water. All the identified odour active compounds were already described in the literature as meat aroma compounds. They are formed by the autoxidation of lipids and by the degradation of sulfur-containing amino acids. Hydrogen sulfide and dimethyl trisulfide were chosen as criteria for the off-flavour. To follow lipid oxidation, headspace concentration of ethane was also measured. Headspace concentration of hydrogen sulfide, ethane and dimethyl trisulfide, analysed 14 hours and one month after the sterilization are summarized in Table 1. The most significant parameter in the ANOVA of the data was the atmosphere above the product. Flushing the trays with nitrogen significantly reduced the concentration of ethane and dimethyl trisulfide, whereas it increased hydrogen sulfide concentration. 11-15
Table 1
Calculated Means of Headspace Concentrations of Hydrogen Sulfide, Ethane and Dimethyl Trisulfide from ANOVA Temperature (°C) 121
Hydrogen sulfide (mg 1 ) TO 12.05 T4 7.79
Atmosphere
131
C
F
9.16 5.37
4.57 0.09
16.64 13.06
9.24 87.59
18.47 175.02
5.04 13.30
3.48 n.d.
8.04 n.d.
2.35 n.d.
Antioxidant AO
0
AO;
A0
2
l
Ethane (mg f ) TO T4
9.29 6.39*
12.21 6.30*
10.31 7.04*
1
13.12 93.97
Dimethyl trisulfide (mg l" ) TO 6.96 T4 n.d.
13.36 84.22
9.48 63.38
11.21 123.52
1
4.86* n.d.
3.05* n.d.
6.27* n.d.
TO, T4: 14 h and one month storage respectively; * Not significant at 0.05 level; n.d. Not determined. 3. 2 Influence of the Process Parameters on Lipid Oxidation In previous work, ' ' flushing with nitrogen and use of a rosemary extract were very effective in preventing lipid oxidation. Similar effects of vacuum and/or modified atmosphere have already been reported for beef patties, sliced cooked meat and whole milk 1 7 10
i
6,16-18
powder. ' 3. 3 Influence of the Process Parameters on Off-flavour 3.3.1 Influence of the Sterilization. For the two sterilization temperatures, the total sterilization effect was the same but the thermal treatments were very different with a cooking value of 80 min and 120 min for 131 °C and 121 °C respectively. A 10 °C increase of the sterilization temperature led to a decrease of about 25% in hydrogen sulfide and 30% in dimethyl trisulfide concentrations after 14 hours of storage. Hydrogen sulfide is formed by thermal denaturation of proteins and/or thermal degradation of sulfur-containing amino
Flavour Science: Recent Developments
198
acids. ~ This can easily be explained since hydrogen sulfide is formed from the - S H groups of the proteins or the free amino acids. Dimethyl trisulfide is formed indirectly in the Strecker degradation of methionine, under the effect of a thermal process. This produces methional, which is further degraded to methanethiol. Methanethiol can be transformed into dimethyl disulfide in the presence of oxygen. Dimethyl trisulfide can be generated from dimethyl disulfide easily. The role of the temperature in these reactions explains the differences in the concentration of dimethyl trisulfide for the two sterilization temperatures. 3.3.2 Influence of Flushing with Nitrogen. Flushing with nitrogen has a very significant role on the hydrogen sulfide concentration in two ways. First, it increases the concentration initially formed (16.64 ug l" in 'F-trays' against 4.57 ug l" in 'C-trays'). The redox potential is strongly reduced in the absence of oxygen and a low redox potential favours the breakdown of the disulfide groups of the proteins and the formation of cysteine from free cystine. As a consequence, formation of hydrogen sulfide is also favoured. Secondly, the decrease in hydrogen sulfide concentration after one month's storage was higher for 'C-trays' (about 97%) than for 'F-trays' (about 22%). One can assume that either the reactivity of hydrogen sulfide is strongly reduced without oxygen or that the compounds with which it can react (e.g., the secondary products of the lipid oxidation) were not present in 'F-trays'. It is known that protein oxidation can occur through radical reactions promoted by lipid hydroperoxides. A radical can be formed at the oc-carbon of the amino acids, but the side chains are also susceptible to damage and cysteine/cystine are among the most labile amino acids. In the presence of oxygen, the radical initiation on sulfur substituents through the formation of thiyl radicals takes place very easily. One can assume that hydrogen sulfide is also able to form thyil radicals. This has already been described for high temperatures. Thus the reaction between lipid hydroperoxides and thiyl radicals is highly probable. In the absence of oxygen, such as in the case of 'F-trays', hydrogen sulfide formed in excess due to the redox potential decrease, cannot react further because of the absence of the secondary products of the lipid oxidation. It is assumed that the decrease of hydrogen sulfide concentration after one month's storage was due only to its dissolution in water. 23
1
1
24
25-27
19
For the formation of dimethyl trisulfide, the role played by the oxygen explains why its concentration was smaller in 'F-trays' than in 'C-trays'. 3.3.3 Influence of Antioxidant. Rosemary extract is a primary antioxidant which can react with radicals to convert them into more stable products. Citric acid is a secondary antioxidant and has the ability to chelate metal ions which catalyse the lipid oxidation and so inhibits the formation of lipid peroxides. The significant role played by the antioxidant on the initial concentration of hydrogen sulfide supports the hypothesis that thyil radicals are involved. With the more efficient antioxidant (rosemary extract), the concentration of hydrogen sulfide was higher. Consequently, the concentration of hydrogen sulfide was the smallest without antioxidant. The antioxidants have no effect on the concentration of dimethyl trisulfide. 4 CONCLUSION This study investigated the influence of different antioxidative treatments applied to sterilized meat in trays on the formation of off-flavour. The removal of oxygen by flushing with nitrogen prior to sterilization was very effective in inhibiting lipid oxidation but gave rise to off-flavour due to an accumulation of hydrogen sulfide and to a diminution of other odour active compounds such as dimethyl trisulfide. Both hydrogen sulfide and dimethyl
Thermally Generated Flavour
199
trisulfide are characteristic compounds of meat aroma, but when the balance between the concentrations is modified, the flavour turns into off-flavour. A residual oxygen quantity must remain in the tray in order to permit formation of thiyl radicals and to prevent accumulation of hydrogen sulfide. In this perspective, use of rosemary extract as an antioxidant could be the optimal solution, both preventing lipid oxidation and formation of off-flavour. REFERENCES 1. B. Giintensperger and F.E. Escher, J. Food Science, 1994, 59, 689. 2. J. Kanner, in 'Lipid Oxidation in Food', ed. A.J. St. Angelo, American Chemical Society, NY, 1992, p. 55. 3. J R . Vercellotti, A.J. St. Angelo and A.M. Spanier, in 'Lipid Oxidation in Food', ed. A.J. St. Angelo, American Chemical Society, NY, 1992, p. 1. 4. J.I. Gray and A.M. Pearson, 'Advances in Food Research', eds. A.M. Pearson and T R . Dutson, Van Nostrand Reinhold, NY, 1987, Vol. 3, p. 221. 5. D. Ladikos and V. Lougovois, Food Chem., 1990, 35, 295. 6. A.M. Spanier, A.J. St. Angelo and G.P. Shaffer, J. Agric. Fd. Chem., 1992, 40, 1656. 7. B. Giintensperger and F.E. Escher, 86th American Oil Chem. Soc. Annual Meeting, San Antonio, Texas, 1995. 8. T.E. Acree, J. Barnard and D.G. Cunningham, Food Chem., 1984, 14, 273. 9. M B . Jacobs, M.M. Braverman and S. Hochheiser, Anal. Chem., 1957, 29, 1349. 10. S. Langourieux and F.E. Escher, J. Food Science, submitted for publication. 11. A.M. Gait and G. MacLeod, J. Agric. Fd. Chem., 1984, 32, 59. 12. G. MacLeod and J.M. Ames, Flavour and Fragrance J., 1986, 1, 91. 13. D.W. Baloga, G.A. Reineccius and J.W. Miller, J. Agric. Fd. Chem., 1990, 38, 2021. 14. T.D. Drumm and A.M. Spanier, J. Agric. Fd. Chem., 1991, 39, 336. 15. N. Ramarathnam, L.J. Rubin and L.L. Diosady, J. Agric. Fd. Chem., 1993, 41, 933. 16. H. Stapelfeldt, H. Bjorn, L.H. Skibsted and G. Bertelsen, Z Lebensm. Unters. Forsch., 1993, 196, 131. 17. G. Hall and H. Lingnert, J. Food Quality, 1984, 7, 131. 18. G. Hall, J. Andersson, H. Lingnert and B. Olofsson, J. Food Quality, 1985, 7, 153. 19. M. Fujimaki, S. Kato and T. Kurata, Agr. Biol. Chem., 1969, 33, 1144. 20. M. Boelens, L.M. Van der Linde, P.J. De Valois, H.M. Van Dort and H.J. Takken, J. Agric. Fd. Chem., 1974, 22, 1071. 21. C.-K. Shu, M L . Hagedorn, B.D. Mookherjee and C.T. Ho, J. Agric. Fd. Chem., 1985, 33, 438. 22. C.-K. Shu, M L . Hagedorn, B.D. Mookherjee and C.T. Ho, J. Agric. Fd. Chem., 1985, 33, 442. 23. L. Schutte, CRC Critical Reviews in Food Technology, 1974, 4, 457. 24. K. Hofmann, Die Fleischwirtschaft, 1977, 10, 1818. 25. H.W. Gardner, J. Agric. Fd. Chem., 1979, 27, 220. 26. H.W. Gardner, in 'Xenobiotics in Foods and Feeds', eds. J.W. Finley and D.E. Schwass, American Chemical Society, Washington, D.C., 1983. 27. D.A. Lillard, in 'Warmed-Over Flavor of Meat', eds. A.J. St. Angelo and M.E. Bailey, Academic Press, Inc., 1987, p. 63.
S. Langourieux and F.E. Escher Department of Food Science, Laboratory of Food Chemistry and Food Technology, Swiss Federal Institute of Technology, Zurich (ETH), CH-8092 Zurich, Switzerland
1 INTRODUCTION Heat sterilization is one of the most important preservation techniques for producing microbiologically safe foods storable at room temperature. The increasing consumer demand for easy-to-use and smaller portions, and with a favourable geometry to reduce heat damage, has led to heat processed convenience foods packed in trays rather than in cans. Nevertheless, sterilization and storage still lead to a considerable loss of organoleptic and nutritional quality, due to oxidative and heat-induced changes, ' in particular if these products contain meat preparations. Off-flavours are developed via the autoxidation of polyunsaturated fatty acids and oxidative effects on proteins, peptides and amino acids. ^ Oxidative stability of various heat-processed menu components has been studied in a comprehensive project at the ETH, Zurich, in collaboration with the food canning industry. The influence of the atmosphere above the product and the influence of a natural antioxidant were particularly investigated. For meat products in aluminium trays, removal of air by flushing with nitrogen during filling associated with a storage temperature of 20 °C significantly increased the storage stability. But in this case, it also leads to the formation of a strong sulfurous off-flavour after the sterilization process. In turn, addition of a rosemary extract as an antioxidant resulted in a very effective stabilization of the product and no sulfurous off-flavour was detected sensorially when the trays were opened. Our objective in this work was first to investigate this off-flavour and then to determine the influence of the process parameters on its formation. 1
2 3
4
1
7
2 MATERIAL AND METHODS 2.1 Sample Preparation The meat-fat mixture was prepared according to Giintensperger and Escher. To this ground meat-fat mixture (90 g) was added 88 mg per kg fat of citric acid or 44 mg per kg fat of desaromatized rosemary extract (RE-S, FIS S.A., Switzerland) or samples were further processed without antioxidant. Deionized water (180 g) was added and the samples 1
*Present address: Laboratoire de Genie Biologique et Sciences des Aliments, Unite de Microbiologie et Biochimie industrielles, Universite de Montpellier II, F-34095 Montpellier cedex 5, France
Thermally Generated Flavour
195
were put into aluminium trays (Type 73300, 138 x 138 x 28 mm, W. Wagner GmbH, Germany). The trays were sealed with a peelable aluminium lid foil (Flexalpeel 45/1000, Alusingen, Germany) with or without prior removal of air by evacuating to 200 mbar and flushing back with nitrogen to atmospheric pressure (CPM H175VG, Packaging Machinery, England). This operation was repeated three times before sealing. The trays were then sterilized in hot water in a batch retort (Pilot Rotor 400, Stock GmbH, Germany) without agitation at 121 °C or 131 °C to an F value of 8-11 min. The trays were then stored in the dark at 20 °C for 14 hours or one month before analysis. To investigate the effects of the different experimental parameters on the response variable (headspace concentration of volatile compounds), an ANOVA analysis was performed (SAS v.6.10, SAS Institute Inc., U.S.A.). 0
2.2 Analysis of the Volatile Compounds of the Headspace 2.2.1 Purge and Trap of the Headspace. The headspace from each tray was collected (without opening the tray) using two needles. The first one purged the tray with purified nitrogen (20 ml min ) and the second one drove the headspace gas into a PTFE column (14 cm length, 0.5 cm internal diameter) packed with Tenax GR, (Alltech). After an hour of purging, the Tenax trap was removed and replaced by another and this purge lasted a further three hours. Diethyl ether (20 ml) was used for the extraction of the traps with 1 -butanethiol (Aldrich) and n-dodecane (Sigma) as internal standards. The total ether solution was further concentrated with a Kuderna-Danish concentrator to a volume of approximately 0.1 ml. No internal standard was added to the extracts for olfactometry. 2.2.2 Gas Chromatography (GC) and Gas Chromatography-Olfactometry (GC-O). A Hewlett-Packard model 5890 Series II gas chromatograph equipped with an HP-5 column (30 m length, 0.32 mm internal diameter, 0.25 mm film thickness) was used for the analysis of the extracts (1 ul). Nitrogen was used as the carrier gas (1.1 ml min ). The temperature programme started at 30 °C and increased at a rate of 1 °C min to 40 °C, then increased at a rate of 10 °C min" to 250 °C and held for 10 min. Injector, FID and FPD (flame photometric detector) temperatures were set at 220, 250, and 150 °C respectively. The gas chromatograph was also equipped with an olfactometric detector (SGE, Australia). The data were analysed according to the CHARM analysis with the help of software derived from Origin 3.5 (DMP Ltd., Switzerland). 2.2.3 Gas Chromatography-Mass Spectrometry Analyses (GC-MS). A Fisons GC 8000 series gas chromatograph equipped with the same column and operated at the same conditions as for the GC analysis and coupled to a Finnigan MAT SSQ710 mass spectrometer was used with helium as the carrier gas. The source, analyser and transfer line temperatures were 150, 70 and 220 °C respectively. The ionization voltage applied was 70 eV, the emission current was 0.2 mA and the electron multiplier voltage 1000 kV. Mass spectra obtained from a scan range of 33-250 m/z were compared with those of known compounds in the NIST library. 2.2.5 Analysis of Hydrogen Sulfide. Based on method of Jacobs et al. 2.2.6 Ethane Determination. As described by Giintensperger and Escher -1
-1
-1
1
11
9,10
1
3 RESULTS AND DISCUSSION 3.1 Determination of the Volatile Compounds Responsible of the Off-flavour A 2 ml sample of the headspace of a non-flushed and a nitrogen-flushed tray were directly injected and a G C - 0 analysis carried out. The results are given in Figure 1. Short
Flavour Science: Recent Developments
196
chain alkanes from CI to C6 and hydrogen sulfide were identified. No odour was detected for the 'C-tray' (no flushing) but a strong spoiled egg like odour was detected for the 'Ftrays' (flushing with nitrogen) at a retention of 1.8 min corresponding to hydrogen sulfide. The same odour was perceived when opening the 'F-trays'. C-' and 'F-trays' were also extracted using the purge and trap method and GC, G C MS and G C - 0 analysis were performed. CHARM profiles of C and F headspace samples are shown in Figure 2. CHARM profiles revealed differences between the two samples. Dimethyl trisulfide, 2-pentyl furan and octanal are important contributors to the odour FID
FPD
1E5 5000
40
— 10
Time (min)
0
Figure 1 FID and FPD chromatograms of 2 ml directly injected tray headspace (C: no removal of air before sealing; F: flushed with nitrogen)
••
1
Figure 2
20-
CHARM Response
*.
•• • *• • • • •• *
0
I
i
4
I
CHARM Response 8 12 i
l
i
I
i
I
i
I
i
l
i
16 I
n.i., butter 3-Methylbutanal, herbaceous 2-Ethylfuran, coffee, roast Dimethyl disulfide, onion Hexanal, herbaceous, pungent Heptanal, fresh, pungent n.i., biead, cooked potato Dimethyl trisulfide, fresh cut onion 2-Pentylfuran, fruity Octanal, fruity (orange-like), pungent 2-Octenal, honey n.i., fresh, light Heptanoic acid, mushroom Nonanal, fatty 2-Buryloctanol, fruity 2-Nonenal, waxy 2,4-Nonadienal, hay n.i., wood 2,4-Decadienal, hay, pungent 2-Undecenal, hay, earth n.i., fried onion
CHARM profiles of headspace purge and trap extracts of two different trays (C: no removal of air before sealing; F: flushed with nitrogen)
Thermally Generated Flavour
197
perceived in the 'C-trays' while only octanal persists in 'F-trays'. It can be concluded that hydrogen sulfide is mainly responsible for the off-flavour in the 'F-trays' but flushing with nitrogen also greatly modifies other odour active compounds. No typical roasted or meaty notes were found in the purge and trap extracts. This is due to the fact that the samples were sterilized without prior pre-cooking and in the presence of a large proportion of water. All the identified odour active compounds were already described in the literature as meat aroma compounds. They are formed by the autoxidation of lipids and by the degradation of sulfur-containing amino acids. Hydrogen sulfide and dimethyl trisulfide were chosen as criteria for the off-flavour. To follow lipid oxidation, headspace concentration of ethane was also measured. Headspace concentration of hydrogen sulfide, ethane and dimethyl trisulfide, analysed 14 hours and one month after the sterilization are summarized in Table 1. The most significant parameter in the ANOVA of the data was the atmosphere above the product. Flushing the trays with nitrogen significantly reduced the concentration of ethane and dimethyl trisulfide, whereas it increased hydrogen sulfide concentration. 11-15
Table 1
Calculated Means of Headspace Concentrations of Hydrogen Sulfide, Ethane and Dimethyl Trisulfide from ANOVA Temperature (°C) 121
Hydrogen sulfide (mg 1 ) TO 12.05 T4 7.79
Atmosphere
131
C
F
9.16 5.37
4.57 0.09
16.64 13.06
9.24 87.59
18.47 175.02
5.04 13.30
3.48 n.d.
8.04 n.d.
2.35 n.d.
Antioxidant AO
0
AO;
A0
2
l
Ethane (mg f ) TO T4
9.29 6.39*
12.21 6.30*
10.31 7.04*
1
13.12 93.97
Dimethyl trisulfide (mg l" ) TO 6.96 T4 n.d.
13.36 84.22
9.48 63.38
11.21 123.52
1
4.86* n.d.
3.05* n.d.
6.27* n.d.
TO, T4: 14 h and one month storage respectively; * Not significant at 0.05 level; n.d. Not determined. 3. 2 Influence of the Process Parameters on Lipid Oxidation In previous work, ' ' flushing with nitrogen and use of a rosemary extract were very effective in preventing lipid oxidation. Similar effects of vacuum and/or modified atmosphere have already been reported for beef patties, sliced cooked meat and whole milk 1 7 10
i
6,16-18
powder. ' 3. 3 Influence of the Process Parameters on Off-flavour 3.3.1 Influence of the Sterilization. For the two sterilization temperatures, the total sterilization effect was the same but the thermal treatments were very different with a cooking value of 80 min and 120 min for 131 °C and 121 °C respectively. A 10 °C increase of the sterilization temperature led to a decrease of about 25% in hydrogen sulfide and 30% in dimethyl trisulfide concentrations after 14 hours of storage. Hydrogen sulfide is formed by thermal denaturation of proteins and/or thermal degradation of sulfur-containing amino
Flavour Science: Recent Developments
198
acids. ~ This can easily be explained since hydrogen sulfide is formed from the - S H groups of the proteins or the free amino acids. Dimethyl trisulfide is formed indirectly in the Strecker degradation of methionine, under the effect of a thermal process. This produces methional, which is further degraded to methanethiol. Methanethiol can be transformed into dimethyl disulfide in the presence of oxygen. Dimethyl trisulfide can be generated from dimethyl disulfide easily. The role of the temperature in these reactions explains the differences in the concentration of dimethyl trisulfide for the two sterilization temperatures. 3.3.2 Influence of Flushing with Nitrogen. Flushing with nitrogen has a very significant role on the hydrogen sulfide concentration in two ways. First, it increases the concentration initially formed (16.64 ug l" in 'F-trays' against 4.57 ug l" in 'C-trays'). The redox potential is strongly reduced in the absence of oxygen and a low redox potential favours the breakdown of the disulfide groups of the proteins and the formation of cysteine from free cystine. As a consequence, formation of hydrogen sulfide is also favoured. Secondly, the decrease in hydrogen sulfide concentration after one month's storage was higher for 'C-trays' (about 97%) than for 'F-trays' (about 22%). One can assume that either the reactivity of hydrogen sulfide is strongly reduced without oxygen or that the compounds with which it can react (e.g., the secondary products of the lipid oxidation) were not present in 'F-trays'. It is known that protein oxidation can occur through radical reactions promoted by lipid hydroperoxides. A radical can be formed at the oc-carbon of the amino acids, but the side chains are also susceptible to damage and cysteine/cystine are among the most labile amino acids. In the presence of oxygen, the radical initiation on sulfur substituents through the formation of thiyl radicals takes place very easily. One can assume that hydrogen sulfide is also able to form thyil radicals. This has already been described for high temperatures. Thus the reaction between lipid hydroperoxides and thiyl radicals is highly probable. In the absence of oxygen, such as in the case of 'F-trays', hydrogen sulfide formed in excess due to the redox potential decrease, cannot react further because of the absence of the secondary products of the lipid oxidation. It is assumed that the decrease of hydrogen sulfide concentration after one month's storage was due only to its dissolution in water. 23
1
1
24
25-27
19
For the formation of dimethyl trisulfide, the role played by the oxygen explains why its concentration was smaller in 'F-trays' than in 'C-trays'. 3.3.3 Influence of Antioxidant. Rosemary extract is a primary antioxidant which can react with radicals to convert them into more stable products. Citric acid is a secondary antioxidant and has the ability to chelate metal ions which catalyse the lipid oxidation and so inhibits the formation of lipid peroxides. The significant role played by the antioxidant on the initial concentration of hydrogen sulfide supports the hypothesis that thyil radicals are involved. With the more efficient antioxidant (rosemary extract), the concentration of hydrogen sulfide was higher. Consequently, the concentration of hydrogen sulfide was the smallest without antioxidant. The antioxidants have no effect on the concentration of dimethyl trisulfide. 4 CONCLUSION This study investigated the influence of different antioxidative treatments applied to sterilized meat in trays on the formation of off-flavour. The removal of oxygen by flushing with nitrogen prior to sterilization was very effective in inhibiting lipid oxidation but gave rise to off-flavour due to an accumulation of hydrogen sulfide and to a diminution of other odour active compounds such as dimethyl trisulfide. Both hydrogen sulfide and dimethyl
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trisulfide are characteristic compounds of meat aroma, but when the balance between the concentrations is modified, the flavour turns into off-flavour. A residual oxygen quantity must remain in the tray in order to permit formation of thiyl radicals and to prevent accumulation of hydrogen sulfide. In this perspective, use of rosemary extract as an antioxidant could be the optimal solution, both preventing lipid oxidation and formation of off-flavour. REFERENCES 1. B. Giintensperger and F.E. Escher, J. Food Science, 1994, 59, 689. 2. J. Kanner, in 'Lipid Oxidation in Food', ed. A.J. St. Angelo, American Chemical Society, NY, 1992, p. 55. 3. J R . Vercellotti, A.J. St. Angelo and A.M. Spanier, in 'Lipid Oxidation in Food', ed. A.J. St. Angelo, American Chemical Society, NY, 1992, p. 1. 4. J.I. Gray and A.M. Pearson, 'Advances in Food Research', eds. A.M. Pearson and T R . Dutson, Van Nostrand Reinhold, NY, 1987, Vol. 3, p. 221. 5. D. Ladikos and V. Lougovois, Food Chem., 1990, 35, 295. 6. A.M. Spanier, A.J. St. Angelo and G.P. Shaffer, J. Agric. Fd. Chem., 1992, 40, 1656. 7. B. Giintensperger and F.E. Escher, 86th American Oil Chem. Soc. Annual Meeting, San Antonio, Texas, 1995. 8. T.E. Acree, J. Barnard and D.G. Cunningham, Food Chem., 1984, 14, 273. 9. M B . Jacobs, M.M. Braverman and S. Hochheiser, Anal. Chem., 1957, 29, 1349. 10. S. Langourieux and F.E. Escher, J. Food Science, submitted for publication. 11. A.M. Gait and G. MacLeod, J. Agric. Fd. Chem., 1984, 32, 59. 12. G. MacLeod and J.M. Ames, Flavour and Fragrance J., 1986, 1, 91. 13. D.W. Baloga, G.A. Reineccius and J.W. Miller, J. Agric. Fd. Chem., 1990, 38, 2021. 14. T.D. Drumm and A.M. Spanier, J. Agric. Fd. Chem., 1991, 39, 336. 15. N. Ramarathnam, L.J. Rubin and L.L. Diosady, J. Agric. Fd. Chem., 1993, 41, 933. 16. H. Stapelfeldt, H. Bjorn, L.H. Skibsted and G. Bertelsen, Z Lebensm. Unters. Forsch., 1993, 196, 131. 17. G. Hall and H. Lingnert, J. Food Quality, 1984, 7, 131. 18. G. Hall, J. Andersson, H. Lingnert and B. Olofsson, J. Food Quality, 1985, 7, 153. 19. M. Fujimaki, S. Kato and T. Kurata, Agr. Biol. Chem., 1969, 33, 1144. 20. M. Boelens, L.M. Van der Linde, P.J. De Valois, H.M. Van Dort and H.J. Takken, J. Agric. Fd. Chem., 1974, 22, 1071. 21. C.-K. Shu, M L . Hagedorn, B.D. Mookherjee and C.T. Ho, J. Agric. Fd. Chem., 1985, 33, 438. 22. C.-K. Shu, M L . Hagedorn, B.D. Mookherjee and C.T. Ho, J. Agric. Fd. Chem., 1985, 33, 442. 23. L. Schutte, CRC Critical Reviews in Food Technology, 1974, 4, 457. 24. K. Hofmann, Die Fleischwirtschaft, 1977, 10, 1818. 25. H.W. Gardner, J. Agric. Fd. Chem., 1979, 27, 220. 26. H.W. Gardner, in 'Xenobiotics in Foods and Feeds', eds. J.W. Finley and D.E. Schwass, American Chemical Society, Washington, D.C., 1983. 27. D.A. Lillard, in 'Warmed-Over Flavor of Meat', eds. A.J. St. Angelo and M.E. Bailey, Academic Press, Inc., 1987, p. 63.