Introduction

The current philosophy for food quality assurance is steadily decreasing the focus on end product testing and verification, traditionally the cornerstones of quality and regulatory control. The efforts of producers and legislation is concentrating on the development and application of structured quality assurance systems based on prevention through monitoring, controlling and recording of critical parameters through the entire product’s life cycle extending from production to final use. Effective application of this approach requires systematic study and modelling of the temperature dependence of shelf life. Ideally this would mean establishing a time correlation between measured microbiological activity and sensory value for the conditions of interest. The objective of the present study was to study the relation between microbial growth and organoleptic characteristics of ground meat and develop and validate a microbial spoilage model for accurate predictions of fresh meat shelf life.

Materials and Methods

Preparation of samples. Ground pork divided into portions of 50g and placed on plastic trains and simply enclosed into permeable polyethylene plastic film transported from open market to the laboratory within 24h at 5oC after their production. The samples stored under controlled isothermal conditions (0, 5, 10, 15 and 20 oC) in high-precision (0.2oC) low-temperature incubators (model MIR 153; Sanyo Electric Co., Ora-Gun, Gunma, Japan). Samples were taken at appropriate time intervals to allow for efficient kinetic analysis of microbial growth. The models developed from the studies at isothermal conditions were validated at dynamic temperature conditions. Four different temperature scenarios were tested: T1: 24 h at 0oC-24 h at 10oC, T2: 12 h at 0oC-6 h at 10oC-6 h at 15oC, T2: 6 h at 0oC-6 h at 10oC-6 h at 20oC, T2: 10 h at 5oC-6 h at 20oC. Τhe temperature dependence of the kinetic parameters was modelled using the Arrhenius equation

where T is the absolute temperature, μref is the growth rate at a reference temperature Tref, EA is the activation energy and R the universal gas constant. A limited number of samples were freeze-stored to serve as controls during sensory evaluation of colour and odour. Microbiological analysis. Samples (25g) of minced meat were aseptically weighed, added to ¼ strength Ringer's solution (225ml), and homogenized in a stomacher (Lab Blender 400, Seward Medical, London) for 60s at room temperature. Decimal dilutions in quarter strength Ringer's solution were prepared and duplicate 1 ml or 0.1 ml samples of appropriate dilutions were poured or spread on the following media: Plate Count Agar (PCA; Merck, 1.05463, Darmstadt, Germany) for total viable count (TVC), incubated at 25o C for 72h; Brochothrix thermosphacta on STAA medium supplemented with streptomycin sulfate, thallous acetate and cycloheximide (actidione); this medium was made from basic ingredients in the laboratory, and incubated at 25o C for 72h; Lactic acid bacteria on MRS (Merck, 1.10660, Darmstadt, Germany) overlaid with the same medium and incubated at 25oC for 96h under anaerobic conditions; Pseudomonas spp. on cetrimide- fucidin- cephaloridine (CFC) agar (Oxoid, CM559 supplemented with selective supplement SR 103E, Basingstoke, UK) incubated at 25o C for 48h; Enterobacteriaceae on Violet Red Bile Dextrose Agar (Merck, 1.10275, Darmstadt, Germany) overlaid with the same medium and incubated at 37o C for 24h. Sensory analysis. Sensory evaluation of minced pork samples was performed during storage by a five member sensory panel composed of staff from the laboratory. The same trained persons were used in each evaluation, and all were blinded to which product was being tested. The sensory evaluation was carried out in artificial light and the temperature of packed product approximated the ambient temperature. Special attention was given to the colour. Organoleptic evaluation: product cooked in aluminum foil at 180oC for 20min. Each attribute was scored on a 3-point hedonic scale where: 1=acceptable; 2=rejection point; and 3=unacceptable.

Results and Discussion

Representative experimental data for growth of the different measured constituents of ground pork natural microflora at 5 and 15 oC are shown in Figures 1 and 2. Pseudomonads were the dominant organisms at all temperature testing following by Br. thermosphacta, lactic acid bacteria and Enterobacteriaceae. At all temperatures, growth of Pseudomonads which is well established spoilage indexes for air stored chilled meat, followed closely the decrease of sensory quality and end of shelf life coincided with a pseudomonads level of 9.0 log10 cfu/g. The growth data of spoilage bacteria from four individual replicated experiments with ground pork stored at different isothermal conditions (0, 5, 10, 15 and 20 oC) were modelled as a function of time using the Baranyi model and the kinetic parameters (max, Lag phase and Nmax) were estimated. The results showed that the storage temperature did not affect the maximum concentration (Nmax) which was found to be constant (for pseudomonas an average value of 9.6 log10 cfu/g. Further, the temperature dependence of the kinetic parameters was modelled using the Arrhenius equation (Figure 3). The estimated values and statistics of the Arrhenius model parameters for the different spoilage bacteria are shown in Table 1. The activation energies were 69.1, 67.6, 98.2 and 96.0 kj/mol for pseudomonads, Br. thermosphacta, lactic acid bacteria and Enterobacteriaceae respectively. The model for pseudomonads growth was validated at non-isothermal conditions using different fluctuating temperature scenarios (figures 4-6). Furthermore, the predicted shelf life (e.g time required for pseudomonads to growth from their initial level N0 to the spoilage level Ns=109 cfu/g) was compared to the observed shelf life estimated by the sensory analysis (Table 2).

Development and validation of a microbial spoilage model for aerobic stored ground meat A. Stamatioua,A. Stamatioua,a,ca,c , K. Koutsoumanis , K. Koutsoumanisbb, M. R. Adams, M. R. Adamscc and G.J. Nychas and G.J. Nychasa, a,

aaLab. Microbiology & Biotechnology of Foods, Dept. Food Science &Technology, Agricultural University of Athens, Iera odos 75, Athens, 11855, Tel.-Fax.: 210-5294693, e-mail: Lab. Microbiology & Biotechnology of Foods, Dept. Food Science &Technology, Agricultural University of Athens, Iera odos 75, Athens, 11855, Tel.-Fax.: 210-5294693, e-mail: gjn@aua.grbbAristotle University of Thessaloniki, Faculty of Agriculture, Dept. Of Food Science and Technology, Lab of Food Microbiology and Hygiene, Thessaloniki,54124, tel.: 2310-471467, Fax: 2310-471257,Aristotle University of Thessaloniki, Faculty of Agriculture, Dept. Of Food Science and Technology, Lab of Food Microbiology and Hygiene, Thessaloniki,54124, tel.: 2310-471467, Fax: 2310-471257,

e-mail: e-mail: kkoutsou@agro.auth.grccSchool of Biological Sciences, University of Surrey, Guildford, Surrey Gu2 5XH, UKSchool of Biological Sciences, University of Surrey, Guildford, Surrey Gu2 5XH, UK tel.: +44-1483-300-800, Fax: +44-1483-300-374, e-mail: tel.: +44-1483-300-800, Fax: +44-1483-300-374, e-mail: m.adams@surrey.ac.uk

AbstractThe growth of the different measured constituents of ground pork natural microflora was monitored at different isothermal temperatures conditions from 0 to 20 oC. At all temperatures, growth of

Pseudomonads which is well established spoilage indexes for air stored chilled meat, followed closely the decrease of sensory quality. The kinetic parameters of pseudomonads (maximum specific growth rate) was modelled as a function of temperature using the Arrhenius model. The model was further validated at non-isothermal conditions using different fluctuating temperature scenarios. The results showed that the kinetic model developed in the present study can accurately predict growth of pseudomonads and shelf life of ground meat at both isothermal and dynamic temperature conditions. Such models can be useful tools for quality optimisation in quality management systems of chilled meat.

References

1.Koutsoumanis, Κ., M. Giannakourou, P. S. Taoukis, and G.J.E. Nychas (2000) Application of SLDS (Shelf life Decision system) to marine cultured fish quality. Int. Journal of Food Microbiol. 73, 375-3822.Koutsoumanis, K. P., Taoukis, P., Drosinos, E. H, and Nychas G-J.E. (2000) Applicability of an Arrhenius model for the combined effect of temperature and CO2 packaging on the spoilage microflora of fish. Applied Environemental Microbiology, 66, 3528-35343.Koutsoumanis, K. and Nychas G-J. (2000) Application of a systematic experimental procedure to develop a microbial model for rapid fish shelf-life prediction. International Journal of Food Microbiology, 60, 171-184

refTT

11

R

E-exp = A

refmax

0

2

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12

0 30 60 90

time (h)

cell

con

c. (l

og

cel

ls/m

l)

-5

0

5

10

15

20

25

30te

mp

era

ture

(C)

=ref x exp(-EA/R*(1/T ref-1/T)) Pseudomonads

EA (kj/mol) 69.1 ref 0.056 r2 0.990

Br. thermosphacta EA (kj/mol) 67.6

ref 0.044 r2 0.981

L. acid bacteria EA (kj/mol) 98.2

ref 0.021 r2 0.981

Enterobacteriaceae EA (kj/mol) 96.0

ref 0.025 r2 0.954

Table 1 =ref x exp(-EA/R*(1/T ref-1/T))

Pseudomonads EA (kj/mol) 69.1

ref 0.056 r2 0.990

Br. thermosphacta EA (kj/mol) 67.6

ref 0.044 r2 0.981

L. acid bacteria EA (kj/mol) 98.2

ref 0.021 r2 0.981

Enterobacteriaceae EA (kj/mol) 96.0

ref 0.025 r2 0.954

Table 2

Temperature profile

SL observed

SL pred. Temp. model

Difference %

T1 85.3 76.5 10.3

T2 98.0 62.8 35.9

T3 68.8 50.5 26.6

T4 71.5 67.7 5.3

Table 2

Temperature profile

SL observed

SL pred. Temp. model

Difference %

T1 85.3 76.5 10.3

T2 98.0 62.8 35.9

T3 68.8 50.5 26.6

T4 71.5 67.7 5.3

Figure 1

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Figure 3

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Figure 4

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Figure 5

Figure 6