Irradiation to Ensure the Safety and Quality of Prepared Meals

D6.20.07 RC 864.2

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Proceedings of the 2nd Research Coordination Meeting FAO/IAEA Coordinated Research Project held in Pretoria South Africa, 26-30 April 2004

Reproduced by the IAEA Vienna, Austria, 2004

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JOINT FAO/IAEA DIVISION OF NUCLEAR TECHNIQUES IN FOOD AND AGRICULTURE

INTERNATIONAL ATOMIC ENERGY AGENCY FOOD AND AGRICULTURE ORGANIZATION OF THE UNITED NATIONS

Irradiation to Ensure the Safety and Quality of Prepared Meals

Report of the 2nd Research Coordination Meeting of FAO/IAEA Coordinated Research Project held in Pretoria, South Africa, 26-30 April 2004

Working Material Produced by the IAEA Vienna, Austria 2004

EDITORIAL NOTE

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TABLE OF CONTENTS

Introduction ...... 1

Achievements ...... 2

Conclusions and Recommendations...... 13

Country Reports ...... 16

A. ARGENTINA...... 16 B. CHINA ...... 20 C. GHANA...... 24 D. GREECE...... 29 E. HUNGARY...... 32 F. INDIA ...... 38 G. INDONESIA ...... 45 H. ISRAEL...... 51 I. KOREA...... 62 J. SOUTH AFRICA ...... 68 K. SYRIA...... 73 L. THAILAND...... 78 M. UNITED KINGDOM ...... 82 N. UNITED STATES ...... 93

Annex I, List of Participants ...... 98

Annex II, Programme...... 108

Annex III, Publications to date...... 104

1. INTRODUCTION

The prepared convenience foods sector has become a significant part of the economy of many developed countries with a similar trend evolving in developing countries, where many types of ethnic foods are now also prepared as convenience foods. For example, the prepared convenience foods sector in Ireland is a significant part of the Irish economy. In 2001, just under half of the sector's total output was exported for a value of 841 million Euro, representing a 12% annual increase. The sector's strong growth both in exports and in total sales has made it one of the fastest growing sectors of the food industry in many countries.

Consumer studies carried out on convenience foods have shown that perceived time pressures contribute positively to the purchase of both prepared meals and take-away meals. Other reasons found to contribute positively to the purchase of prepared meals include not enjoying cooking for oneself, a value-for-money perception of convenience foods and different eating times of family members.

As a consequence of the increased market for convenience foods particularly, the food industry is interested in developing ways for the production of prepared meals, which are safe to eat, have an acceptable shelf-life and are of good sensorial and nutritional quality. One technology with the potential to achieve these objectives is food irradiation, which is one of the most thoroughly researched food processing technologies ever developed.

It is thought that research into the application of ionizing radiation to products such as prepared meals could be of unique benefit to the food industry, particularly in developing countries where the microbiological safety of many ethnic dishes is questionable and their shelf-life limited due to the conditions under which they are produced and stored. Food irradiation used on its own or in combination with other processes such as chilling could significantly enhance the microbial safety of such products as well as extending shelf-life. This is of special importance for the most vulnerable individuals in society such as the immunocompromised.

Although extensive research has been carried out on the microbiological and sensorial effects of irradiating individual uncooked food items, little work has been reported on the irradiation of mixed food systems such as prepared meals. In this CRP, the potential of using the irradiation technology for convenience foods is being investigated with regard to safety, shelf- life and overall quality, particularly in terms of sensory and nutritional quality.

The products being investigated include a wide range of ethnic meals such as waakye from Ghana, biltong from South Africa, galbi from Korea, spicy chicken basil rice from Thailand, and kubba and borak from Syria. Other aspects of the CRP include the development of HACCP systems for prepared meals and research into consumer acceptance of irradiated food.

The overall objective of this CRP is to evaluate the effectiveness of irradiation as a method to ensure the microbiological safety and extend the shelf-life of prepared meals, stored under ambient, chilled or frozen conditions and to evaluate the sensory quality of the treated products.

The specific objective of the CRP is to use validated procedures for irradiation treatment and process control, and to use validated methods for assessing microbiological safety and quality as well as sensory evaluation of prepared meals mainly of ethnic origin.

2. MEETING

The meeting was held at the University of Pretoria, South Africa from 26 - 30 April 2004 and was attended by Research Contract/Agreement holders from Argentina, China, Ghana, Greece, India, Indonesia, Israel, Korea, Syria, South Africa, Thailand, United Kingdom and United States of America, as well as five observers from South Africa. The list of participants is attached as Annex 1. Greece is a new participant within the CRP. Hungary is also a Research Contract holder but the investigator could not attend the meeting although a report was submitted and is included as part of Annex 3. The main objective of the meeting was to evaluate the achievements of the CRP during the last 18 months.

The meeting was opened by Professor Johann Kirsten, Chairperson of the School of Agricultural & Food Sciences, University of Pretoria, who welcomed the participants of the RCM to South Africa and the University. Dr Tatiana Rubio-Cabello of the Food & Environmental Protection Section of the Joint FAO/IAEA Joint Division for Nuclear Techniques in Food & Agriculture, Vienna, thanked Professor Kirstein for his kind words of welcome and the University of Pretoria for hosting the RCM. In particular, she thanked Professor Amanda Minnaar and her colleagues for organising the meeting and for her excellent cooperation prior to the RCM.

Dr Rubio emphasized the objective of the meeting as well as the new requirements of the Agency to publish the research results in peer-reviewed scientific journals. A list of peer- reviewed journals in Food Science and Technology as well as Microbiology and Food Safety was distributed.

Professor Minnaar was elected Chairperson of the Meeting with Dr Rudy Nayga and Dr Eileen Stewart agreeing to act as rapporteurs.

All Research Contract/Agreement holders presented a report on the work they had undertaken since the previous RCM in Vienna, 2002. The meeting agenda included an introduction of each participant followed by 45 minute presentations and a 15 minute discussion on each presentation. The program of the meeting is attached in Annex 2.

As a result of the discussion, participants agreed to the following definition for prepared meals:

“Have had sufficient preparation, before being bought, for consumers to eat either as is or after a short heat treatment.”

Most products in this category are perishable, have a short shelf-life and require refrigeration to maintain their freshness.

3. ACHIEVEMENTS

3.1. General achievements

3.1.1. Meals for the immunocompromised

The investigator from Argentina presented findings on the production of safer meals prepared specially for immunocompromised hospital patients. It is estimated that this group comprises 20% of the total world population, whether they are hospitalized or not. It is important to note that the participant from Argentina worked together with nutritionists and the Dietary &

Alimentary Service of the Clinical School Hospital “José de San Martín”, Buenos Aires. This made it possible to have whole irradiated lunches tasted by hospitalized immunocompromised patients. The Ethics Committee of the hospital did not have any objections to the work being carried out. The microbial decontamination levels attained by irradiation of this lunch composed of salad, main dish and dessert, made it possible to afford these longed-for, more varied, nutritious and palatable unusual meals to immunocompromised patients, without foodborne disease risks. The patients participating in these studies showed a keen interest in the meals provided. The research was taken up by the press and published in a widely distributed newspaper in Argentina (La Nación, April 5, 2004). Some television interviews were also conducted.

3.1.2. Hazard Analysis of Critical Control Points

The objectives of the work carried out by the investigator from Israel is to introduce a modified HACCP (Hazard Analysis of Critical Control Points) analysis route for irradiated prepared meals that addresses health hazards as well as sensorial failures, and economic risks, while pin-pointing failure modes specific to the radiation pasteurization aspect. The suggested modified analysis should serve attempts to transfer processes and products from the laboratory-research type into an industrial one. While in the laboratory “failure” samples or lots, or parts of a lot that are of inferior quality can be discarded, an industrial process must be profitable and, hence, stable and producing reproducible high quality products. The analysis route exemplified covers all the steps involved in the food production, from farm to fork. The analysis comprises foreseen failure modes related to the physical, chemical and bacterial aspects of all raw materials, including packaging, all food processing steps and tools involved therein, packaging process and the subsequent irradiation, storage and distribution steps and, last but not least, the consumer’s expected handling. The serious consequences of an inadequate HACCP system in an increasingly global food market, where control points may be in different continents, were exemplified by a retrospective hazard analysis of a widely exported kosher snack. Finally, a practical 10-step approach to implement the suggested combined HACCP, from comprehensive analysis to validated protocol was provided. In conclusion, HACCP protocol has to be carefully structured to specifically answer the needs of each product. The forthcoming stage of this work will be to collaboratively produce combined specific HACCP protocols for radiation-pasteurized prepared meals of other groups in the CRP. [See report from Israel under Annex 3].

3.1.3. Consumer studies

The objective of the consumer project carried out by the USA investigator is to assess and evaluate consumers’ perceptions, acceptance and willingness to pay for irradiated ready-to- cook and fully cooked prepared beef products. Determining consumers’ willingness to pay a premium for irradiated food products is important because this is a major factor that would determine the potential marketability and success of the product. In addition, food irradiation adds costs to the production of the product and these costs must be able to be covered by the price premium before any food manufacturer or retailer will consider selling the irradiated product. Studies carried out to date, using both survey and experimental economics methodologies, generally suggest that information about the nature of food irradiation technology increases consumer acceptance of irradiated prepared and processed ground beef and fully cooked beef brisket. The research findings also indicated that consumers are willing to pay a premium for irradiated ground beef.

3.2. Specific achievements

The participants carried out research into almost 20 different prepared meals. Table 1 summarizes the list of the dishes investigated as well their composition, intrinsic qualities, and the analyses carried out in order to determine their overall safety and quality.

Table 1. List of prepared meals studied Country of Origin Animal Based Products Vegetable Based Products/Miscellaneous Argentina Cannelloni Vegetables & hard boiled egg Empanadas salad; Fruit based dessert China Chinese dumpling Wuxi chop Ghana Poached chicken meals Hungary Cordon Bleu (chicken & ham) Sous-vide mixed vegetables Filled pasta products (Tortellini) Kubba India Prawn masala Poha Chicken biryani Upma Prawn pulao Mixed vegetables Khichadi Rice Vegetable pulao Indonesia Chicken vegetable soup Black soup Chicken sweet-corn soup Ox-tail soup

Korea Bulgogi Galbi South Africa Beef Biltong Syria Kubba Borak Thailand Thai spicy chicken basil rice UK Chicken Masala

3.2.1. Animal based prepared meals

The efficacy of radiation processing for microbiological safety and quality of more than ten prepared meals with beef, chicken, pork or mutton as a major component was investigated. The meals included cannelloni, empanadas, biltong, soups, Thai spicy chicken, poached chicken, chicken chile, Chinese dumpling, wuxi chop, chicken masala, bulgogi, galbi, kubba and borak. The optimum gamma radiation doses were found to be in the range of 2 to 3 kGy for majority of the meals to achieve microbiological safety and desired sensory quality. A dose dependent increase in the spoilage microflora was observed during storage trials. Challenge studies with pathogens such as Escherichia. coli, Listeria monocytogenes, Staphylococcus aureus, and Salmonella spp., revealed that doses employed eliminated the test organisms thus unequivocally establishing the safety of these products. Shelf-life of the meals

was extended from one week to more than three weeks at chilled temperatures depending upon the characteristics of the meals. No significant changes were observed with regard to physical and chemical properties such as pH, water activity, lipid peroxidation of the meals. There was no significant difference in the overall acceptability of the meals at the optimal doses of gamma radiation.

3.2.2. Vegetable based prepared meals

Radiation processing of two of the most popular vegetarian meals consumed in India namely vegetable pulav and mixed vegetables has been standardised. The initial bacterial load in the non-irradiated control samples was found to be 2.6 log10 cfu/g and increased to 6.4 log10 cfu/g during storage at 0-3°C. These samples were also found to be contaminated by potentially pathogenic bacteria such as S. aureus and spoiled within two weeks. Contrary to this, no viable bacterial growth was observed in samples treated with gamma radiation (2 kGy) up to 30 days of storage period. Malonaldehyde formation, a measure of lipid peroxidation increased marginally on irradiation and on further storage. No significant difference in sensory acceptability was observed between the untreated samples and the irradiated ones. Shelf-life of the meals was extended by more than two weeks by gamma radiation treatment. Thus it can be concluded that microbiologically safe, convenient vegetable pulav and mixed vegetables with a shelf-life of a month could be prepared by radiation processing and be of advantage to the processor, retailer and consumer.

3.2.3. Miscellaneous meals

The sensory quality of cooked rice irradiated at more than 2 kGy was found to be unacceptable in terms of texture and colour.

Gamma irradiation with doses of 5-7 kGy of four frozen soups made of different basic materials, having moisture contents between 69 and 86%, could reduce microbial load by 2-3 log cycles and extend the shelf-life to three months at 4oC, without impairing sensory quality. Soups were vacuum packaged with laminated pouch of polyester/aluminum foil/LLDPE.

A dessert composed of fresh apples and pears cubes mixed with strawberry flavoured gelatine jelly and soft white cheese, packaged in polypropylene recipes and refrigerated at 5oC, was successfully decontaminated by 1.5 kGy of gamma radiation, attaining a 3 log cycles reduction in total viable counts with acceptable sensory quality throughout a week of storage, which doubled its shelf-life. A Salmonella enteritidis challenge test showed that this dose was sufficient to reduce its counts by 6 log cycles, which assures a good security level.

Gamma radiation of a carrot, hard-boiled egg and tomato salad at a dose of 2 kGy, packaged in polypropylene, covered with PVC film and stored at 5°C was sufficient to attain a 6 log cycle reduction in Salmonella enteritidis counts. Total viable counts were reduced by 3-4 log cycles with few detrimental effects on sensory quality.

3.3. Publications

Some of the participants have already published the results obtained in relevant journals. A list of these publications is attached as Annex 3.

4. OTHER ACTIVITIES

On the afternoon of 29 April, the participants visited the irradiation facility of Isotron, South Africa (SA) in Isando. Participants were met by the Managing Director, Mr Gúnther von Ketelhodt. He presented an outline on the status of food irradiation in South Africa in general and Isotron, SA specifically. Discussions were held on the technological, quality and safety aspects, and future prospects of food irradiation as a pasteurization method for prepared meals.

5. SUMMARY OF THE RESULTS

A summary of the products studied, their composition, intrinsic qualities and analyses undertaken is given in Table 2.

Table 2. Summary of Safety and Quality Parameters of Ethnic Prepared Meals Country Prepared Composition of prepared meals Intrinsic Safety and Quality Parameters Meal(s) Parameters Microbiological Sensorial Chemical studied 1 Argentina Cannelloni in Wheat dough wrapping (raw) PH, aw L. innocua, Overall acceptability using a 9- tomato sauce Salmonella spp, point hedonic scale Filling; cooked spinach, veal meat, cheese S. aureus Total Viable Count, Mould & Yeast Count, Total Coliforms, Sporulated anaerobes Salad Grated carrot Whole tomatoes (“cherry”), hardboiled egg S. enteritidis, Salmonella spp, S. aureus Total Viable Count, Mould & Yeast Count, total Coliforms, Sporulated anaerobes

Empanadas Wheat dough wrapping (raw) Filling: boiled chicken breast, vegetables

Desert Fresh apples, pears, commercial strawberry gelatin, soft cheese 1 1 2 2 China Chinese Wheat flour, pork, onion, spices Moisture , aw pH Total Viable Scores based on colour, texture, POV , TVBN , dumpling Count, Salmonella taste, flavour, overall acidity1 spp.,S. aureus acceptability using a 9-point protein1 hedonic scale fat1 Wuxi chop Soya sauce, sugar, spices E. coli + all the above 1 1,2 Ghana Poached Chicken, rice, carrots pH, aw , FFA Total Viable Triangle test, Overall chicken meat Count acceptability using a 9-point hedonic scale

Country Prepared Composition of prepared meals(s) Intrinsic Safety and Quality Parameters Meal(s) parameters studied Microbiological Sensorial Chemical

1 2 India Poha, Upma, Vegetables, chicken, rice, prawns, garlic, ginger aw Total Viable Count Overall acceptability using a TBARS Mixed paste S. aureus, 10-point hedonic scale vegetables, Faecal coliforms, Chicken chilli, Mold and Aerobic Prawn Masala, spore Count Vegetable pulav, Chicken biryani, Prawn rice Indonesia Black soup Beef meat, Pangium edule, shallot, garlic, roasted pH, fat1, Total Viable Count Overall acceptability using a 5- coriander, red chilli, ginger, lemon leaf, roasted fish carbohydrate1 Mold & Yeast Count, point hedonic scale paste, turmeric, ginger root, lemon grass, Coliforms, E. coli, Kaempferia galanga, salt, palm sugar, bay leaf, Salmonella spp., palm oil, water S. aureus and Cl. perfringens Oxtail soup Oxtail, shallot, garlic, salt, palm oil, water, onion, ground nutmeg, cloves, white pepper, onion leaf, celery, margarine Chicken and Chicken, shallot, garlic, salt, water, nutmeg, white vegetable soup pepper, onion leaf, celery, margarine, sugar, carrot, green beans, broccoli, sugar peas

Chicken sweet Chicken, salt, water, nutmeg, sweet corn, chicken corn soup sausage, carrot, egg, corn starch Israel A variety of Mixed meat, vegetables HACCP parameters HACCP parameters HACCP parameters HACCP meals parameters 1 Korea Bulgogi Beef, soy sauce, grain wine, sugar, kiwi, green pH, aw Thermophilic bacteria, Overall acceptability using 9- Galbi onion, garlic, onion, sesame oil, black pepper Coliforms, point hedonic scale S. aureus, E. coli, S. typhimurium, B. cereus, Total Viable Count

Country Prepared Composition of prepared meals(s) Intrinsic Safety and Quality Parameters Meal(s) parameters studied Microbiological Sensorial Chemical

South Biltong Salted, dried, intermediate moisture meat product Moisture1, NaCl1 S. aureus Multiple difference testing TBARS2 1 Africa aw , Overall acceptability using a fat1, 9-point hedonic scale pH Syria Kubba Ground wheat, beef, spices, lamb, onion, fat, Moisture1, fat1, ash1, Total Viable Count Overall acceptability using a Lipid oxidation, 1 1 1 pistachio protein , aw , pH 5-point hedonic scale TVBN , total acidity2 Borak Dough, eggs, lamb, onion, spices As above. Total Viable Count, As above. As above. Coliforms, Yeast Count, Salmonella spp., E. coli

Thailand Spicy chicken Cooked rice, chicken, vegetable oil, chilli, fish Moisture1, pH l. monocytogenes, basil rice sauce, water, Basil leaves e. coli, S. typhimurium U.K. Chicken Chicken, onion, tomato, water, yoghurt, coconut, Total Viable Count, TBARS2, Masala red pepper, tomato puree, rapeseed, oil modified Pseudomonas spp., Vitamin B1 and starch, ground coriander leaf, salt, ginger, cayenne Psychrotrophs, Lactic E, pepper, malt extract, turmeric acid bacteria, Cyclobutanone Coliforms (EN1786) USA Fully cooked Beef brisket, spices Consumer Consumer beef brisket demand and demand and acceptance acceptance parameters parameters

Ground beef Ground beef patties As above. As above

Abbreviations: 1(%), 2 TBARS: Thiobarbituric Acid Reactive Substances, POV: Peroxide Value, TVBN: Total Volatile Basic Nitrogen, DPPH: 1,1-Diphenyl-2-Picrylhydrazyl, FFA: Free Fatty Acids

FUTURE WORK PLANS

For future work, participants will use the following protocol to describe the experimental work that will be carried out. The completed protocols will be forwarded to Dr Rubio- Cabello by 31 May 2004. The future work plan contractual periods are shown in Table 3.

Table 3. Contractual periods Country Period Argentina 15-02-04 to 14-02-05 China 15-05-04 to 14-05-05 Ghana 15-04-04 to 14-04-05 Greece 01-03-04 to 28-02-05 Hungary 01-04-04 to 31-03-05 India 30-09-04 to 29-09-05 Indonesia 01-11-03 to 31-10-04 Korea 01-04-04 to 31-03-05 South Africa 01-12-03 to 30-11-04 Syria 15-05-04 to 14-04-05 Thailand 30-04-04 to 31-03-05

Experimental Protocol

Name of participant Country Type of meal Intrinsic properties: For example: (please indicate error pH margins) Aw Moisture content Fat content Others: Experimental objective

Treatments: Number of samples required: Irradiation Dose: Temperature: Dosimetry (please indicate error 0 kGy used: margins) x kGy Min dose: Max dose:

Type of packaging Oxygen Gas mixture (if Laminate used Permeability relevant) Unpackaged PVC over-wrap Vacuum packed MAP Other? Other treatments (e.g use of preservatives)

Storage period / shelf- Temperature: Time: Intervals for life period testing (please indicate safety samples: margins)

Treatments Number of samples Inoculation study Pathogen 1: Inoculation level: 0 (please indicate error 10x cfu/g margins) Pathogen 2: Inoculation level: 0 10x cfu/g Pathogen 3: Inoculation level: 0 10x cfu/g Replication of experiment (minimum 3 separate occasions) Analyses: Microbiological Chemical tests: Sensory tests: Tests: Consumer acceptability/preferences (at least 50 consumers ): Yes / No

Analytical discrimination tests: (n = 30): Yes / No:

Descriptive tests (trained panel of 10- 15 panellists(recommended for shelf-life studies) Yes / No Duplicate analyses per samples for Analyses per Analyses per descriptive tests: Y/N sample tested: sample tested: Duplicate Y/N Duplicate Y/N Triplicate Y/N Triplicate Y/N Application of HACCP Yes No Statistical analyses to be used: Number of envisaged publication

7. CONCLUSIONS AND RECOMMENDATIONS

7.1. Conclusions

From the results obtained it is evident that the use of ionizing radiation has the potential to improve the safety and quality as well as extending the shelf-life of the prepared meals studied.

This technology could potentially be advantageous to food manufacturers and consumers.

In view of the increasing trend in consumer demand for safe prepared foods, the importance in the use of radiation pasteurization is likely to increase in the future. This was confirmed by the consumer studies carried out in the USA, the results of which were reported during the course of this RCM.

7.2. Recommendations

The current trend of increasingly approaching the industry and greater industrial collaboration of the RCM members on prepared meals should be intensified. In particular, awareness of the trend in the food industry of increasing production of prepared meals should allow further research on a wider variety of products. Nevertheless, a higher level of “industrial thinking” and market and economic-testing, theoretical as well as experimental, should be incorporated in the individual projects.

Receiving national approval for new radiation-pasteurized prepared meal products is crucial, legally and economically alike, since it is crucial to allow their market-tests and consequent marketing as soon as their development is complete. Hence, it is most recommended to take care of all legal aspects of the petitions for new products, as soon as completion of their development seems viable.

The currently existing rather limited variety of prepared meals manifest the state of the art that is available before the application of radiation-Pasteurization. New prepared meals, which are facilitated only with the aid of radiation Pasteurization, are a great challenge. Hence, their research and development will greatly exemplify the unique capability of radiation Pasteurization in serving the food industry and its future markets.

The selection of bacteria ensemble for inoculation-pasteurization studies should be harmonized in the RCM. This is highly important to ensure that the bacterial studies, that are crucial for industrial development and the concurrent liability, have been properly addressed. Further, inoculation-Pasteurization studies of ethnic foods should preferably be carried out in the presence of the expected natural flora typical of each food, to ensure relevancy of the results, conclusions and practical recommendations.

A common finding in most of the prepared meals was radiation-induced lipid peroxidation. The concurrent sensorial degradation was often poorly detected by the sensory panel members in the research, probably due to the fact that mostly untrained consumer panels were used instead of trained panelists using, for example, descriptive sensory tests. It is recommended that this failure mode should be considered in advance, in all the projects, and experiments to address it will be carried out accordingly. Counter measures such as increased quantity of antioxidants, oxygen-free packaging (i.e. MAP or Vacuum) or fat-level reduction should preferably be incorporated in future studies from rather early stages. These measures are

expected to increase the sensorial quality and public satisfaction and willingness to buy, consequently.

A common finding in many of the prepared meals was radiation-induced change of acidity and sensorial quality thereafter. In some cases, the sensorial quality was degraded, yet in other cases it was improved. It is recommended firstly to measure the preferred pH sensorial range in a specific population. Consequently, it is recommended to manipulate the post-irradiation acidity, via the use of food acidifiers such as vinegar, to maximize consumer satisfaction and food marketability.

The oversimplified attitude to employ ionizing radiation as a technology to pasteurize an existing industrial meal-product is often inadequate to meet the goal of extended shelf-life. The meal could be modified first, to extend its sensorial shelf-life (e.g. texture), regardless of bacterial growth, and then pasteurized by the irradiation.

The attempt to employ radiation to pasteurize a highly contaminated existing industrial meal- product often results in needing a relatively high dose, which often results in poor sensorial quality. In principle, the preparation should meet Good Manufacturing Practices (GMP), thus lowering the bacterial burden as much as possible. In particular, this is applicable to hand- made high-moisture foods (e.g. Borak, Wuxi Chop), which are currently over contaminated.

A distinction should be made between bulk and surface contamination of specific foods. Bulk contamination is typical of ground or minced foods, which benefit highly from radiation pasteurization. Foods with a small surface to volume ratio carry almost only external microbial contamination. That bacterial load should be preferably lowered first by surface- specific pasteurization methods. Post-packaging irradiation of the product can thus be minimized to ensure optimum sensorial quality, as well as minimizing unnecessary irradiation costs.

Prepared ethnic meals, which are currently hand-made in some countries, will sooner or later be mechanized to reduce production costs as labor costs permanently rise. Hand hygiene close to that achievable in mechanized production should be practiced, to lower the bacterial burden, the irradiation dose needed to pasteurize the product, and the consequent loss of sensorial quality.

It is generally recommended to always bear in mind the perception that radiation pasteurization is a means and not an objective. It is one among many techniques to ensure the safety and quality of food. Its use should always be considered with open mind, taking into account its advantages and disadvantages alike, technologically, sensorially, and economically. Radiation pasteurization is particularly advantageous as the final stage, applied on the packed and sealed prepared meal. Nonetheless, the bio-burden of the ingredients at early stages of preparing the meal can most often be reduced by alternative methods, which may be advantageous to specific foods. This attitude includes a pre-treatment of radiation- disinfection of specific ingredients of high bio-burden. Thus, the radiation dose at the final stage, which affects all the ingredients, can be substantially lowered, resulting in improved sensorial quality and as important - economic profitability.

Sufficient numbers of available consumer panelists for sensorial evaluation is a common obstacle in this type of research. It is recommended that participants use descriptive sensory panels (10-15) for shelf-life testing instead of consumer panels.

It is highly recommended to carry out primary characterization of the products with regard to chemical and microbiological parameters.

If changes are made to the agreed Work Plans, immediate notification and request for approval of the new plan should be made to the IAEA.

It is expected, and highly recommended, that research results should be published in peer- reviewed journals, at least one publication per project.

ARGENTINA

“Safer Prepared Meals for Immunocompromised Patients and the General Consumer, by Gamma Irradiation”

Chief Scientific Investigator: Patricia Narvaiz

National Atomic Energy Commission- Argentina Food Irradiation Section- Technological and Agricultural Applications

Research Contract No.: 11903

INTRODUCTION

Caneloni in tomato sauce, “empanadas”, vegetables and hard-boiled egg salad, and fruit salad in gelatine with soft cheese, were the meals considered in this period. Caneloni was studied by Mr. Gastón Liendo , an advanced student of the Food Technology career of the Belgrano University, Buenos Aires, who chose food irradiation as subject of his graduation thesis. Microbiological experimental work was carried out by Mr. Liendo at Lyca Food Control Laboratories, a private institution where he was working, and experimental design, irradiation and sensory analyses, at this institute. Contacts with the technical staff of the company which manufactures and commercializes these caneloni were made.

The other three meals composed a whole lunch for immunocompromised patients, as designed by Miss Paola Veronesi Bohl and Mrs. Carolina Kratzer, advanced students of the Nutrition career of the Entre Ríos National University who chose food irradiation as subject of their graduation thesis, under the supervision of Lic. Elena Cossani, nutritionist and professor. Dr. Liliana Lound, from the Food Science and Technology career of the same University, performed the microbiological analyses. Experimental design, irradiation and sensory analyses (consumer panel) were carried out at this Ezeiza Atomic Center. Sensory analysis was also carried out by some immunocompromised patients of the Clinical School Hospital “José de San Martín”, Buenos Aires, after the approval of the Ethics Committee of that Hospital.

The aim was to offer immunocompromised patients some ready-to-eat meals which they usually would wish to eat, highly nutritious besides, but forbidden due to high risk of food borne pathogens transmission. They were selected after visiting hospitalized persons who declared their longing for colourful, fresh fruits and vegetables, and “empanadas”. These are very popular among the Argentine population; the wrapping, made of wheat pasta, is raw, and the filling is half cooked, containing usually some kind of meat. Empanadas are hand-closed by means of a twisting on the pasta borders, and it is not possible to perform this operation with gloves.

2. MATERIALS AND METHODS

For every meal, a first experience consisted of evaluating survival of a selected microbial pathogen after increasing radiation doses, and a preliminary out of panel sensory evaluation of samples irradiated at the estimated minimum and maximum doses, to detect coarse differences attributable to irradiation.

Samples were stored at 5 ± 2 C and 60 ± 8 % RH along storage time. Irradiation was carried out in the semi-industrial Cobalt-60 facility of the Ezeiza Atomic Center, 1.1 x 1010 MBq of activity , at a dose rate of 0.15 kGy/min. Doses were measured with silver dichromate.

A second experience consisted of irradiating at the minimum dose found in the first experience as that required to get a 6 log cycle reduction in pathogen counts, and at twice that dose (Dmax). Microbiological analysis required by the Argentine Alimentary Code (AAC) were performed on control and irradiated samples along storage: total aerobic bacteria counts (TABC), moulds and yeasts, sporulated anaerobes, Salmonella spp, Staphylococcus aureus, coliforms, faecal coliforms, according to ICMSF methods. These samples were sensorily evaluated by a ~ 50 member healthy consumer panel twice along their estimated shelf-life. The evaluated attributes were: aroma, external appearance, colour, tenderness, juiciness, flavour and general acceptability, with a nine point hedonic scale. Results were statistically analyzed by Dunnet test, p< 0.05. Four immunocompromised patients also tasted a whole irradiated lunch, consisting of salad, empanadas and dessert.

Water activity and pH of samples were measured, the first one by sample equilibration with standard salts solutions, AOAC (1995); the second one, by means of a pH meter.

Meat and vegetables cannelloni composition: 34.65% water, 31.7% wheat flour, 12.7% grated cheese, 10.7% spinach, 8.1% cooked veal meat, 1.1% pasteurized egg, 0.8% sodium chloride, 0.14% sodium glutamate, 0.1% nutmeg), purchased from a commercial pasta factory of wide distribution in Argentina, were placed into polystyrene commercial recipes with lid and covered with tomato sauce (90% industrially canned tomato puree, 8% chopped fresh onions, 2% dehydrated marjoram). Inoculated with Listeria innocua. Cooking was performed in a microwave oven.

Grated carrot, whole tomato (“Cherry” variety) and hard-boiled egg salad: Packaging was polypropylene trays, covered with PVC (polyvinylchloride) film.

“Empanadas” filling: chicken breast, onion, red pepper, carrot, tomato, hard boiled egg, salt, fresh parsley, dried marjoram. This mixture was fast boiled with a little water, cooled, wrapped with pasta discs, and placed into aluminum recipes with lid. Cooking was performed in a gas oven.

Fruit (fresh peeled apples and pears pieces), strawberry flavoured gelatine, and soft cheese dessert was placed in two layers into polystyrene recipes with lid.

The last three meals were inoculated with Salmonella enteritidis.

Sensory evaluation by the consumer panel was performed on Days 2 and 15 for cannelloni, and for the rest of the meals, on Days 2 and 8 after irradiation. Control samples were only evaluated on the first analysis date. Immunocompromised patients evaluated on days 2 and 8.

RESULTS

MEAL WATER ACTIVITY (25C) pH Cannelloni in tomato sauce 0.990 5.5 Salad 0.995 Tomato 4.20 Carrot 6.40 Hard-boiled egg 7.65 Empanadas 0.990 5.93 Dessert 0.995 3.90

Results from the inoculation experience showed that radiation doses of 1.5, 2, and 3 kGy were needed to reduce 6 log cycles of Salmonella enteritidis in dessert, salad, and empanadas, respectively, and 3 kGy for reducing Listeria innocua in the same amount in cannelloni.

Preliminary sensory evaluation showed the following differences between control and irradiated samples: the salad had slight egg yolk and carrot discolouration, vegetables softening and egg and tomato off-odour and flavour, especially at 4 kGy; irradiated empanadas had more attractive (golden) colour after cooking; dessert showed fruit softening and gelatin discolouration with irradiation, especially at the 4 kGy dose, fruit browning was observed in every sample.

The preliminary sensory evaluations indicated a reasonably good acceptability of irradiated samples, provided some changes were performed:

• Salad: Radiation doses, or at least maximum radiation dose, should be diminished. Tomato should be placed whole, and not cut in halves.

• Empanadas: The filling should be less cooked, so as to be more juicy. A little bit more marjoram should be incorporated, to make filling more tasteful.

• Dessert: The addition of a red colourant, “Punzo Red”, authorized by the AAC, which had proved to be fairly radiation resistant in a previous experience, should be incorporated. Fruit pieces would be soaked in orange juice previous to their incorporation to the gelatin, with the aim of evaluating the scavenging action of ascorbic acid in reducing browning. If possible, radiation doses will also be reduced, to prevent softening to some extent.

Microbiological and sensory results (consumer panel):

• CANNELLONI: Microbiological results along storage showed a three log cycles reduction in total aerobic bacterial (TABC) and moulds and yeasts counts, and at least one log cycle reduction in total coliforms after irradiation at 3 kGy, being pathogens absent in every sample. Sensory evaluation showed no significant differences between control and irradiated samples, even at 6 kGy (Dmax), being every attribute evaluated with qualifications above the acceptability threshold, between 6.5 to 7.5.

• SALAD: Irradiation at 2 kGy renderred a 3 to 4 log cycles reduction in TABC and moulds and yeasts, and a 2 log cycles in total coliforms, being pathogens absent in every sample. Sensory evaluation showed qualifications between 6.5 and 7.5 for every sample and

attribute. Irradiation at 2 and 3 kGy (Dmax) significantly , but slightly, lowered texture, flavour and acceptability.

• EMPANADAS: A five log cycle reduction in TABC, three log cycles in moulds and yeasts, and two log cycles in total coliforms was obtained with 3 kGy, being pathogens absent in every sample. Sensory evaluation showed no significant differences between control and irradiated samples, even at the 6 kGy dose (Dmax). Attributes got qualifications around 7, with the exception of juiciness, between 6 and 7.

• DESSERT : Samples irradiated at 1.5 kGy showed a three log cycles reduction in TABC , fulfilling the AAC along storage time . Regarding sensory evaluation, the only significant differences were found between control and 2.5 kGy (Dmax) samples in texture and acceptability, which were slightly lowered by irradiation. Every attribute in every sample got qualifications higher than 7. Four immunocompromised patients evaluated the three meals irradiated at Dmin in their lunch with higher qualifications than the consumer panel, ranging from 7 to 9, being acceptability between 8.5 and 9.

CONCLUSIONS

Gamma irradiation of these meals can offer microbiologically safe and sensorily acceptable products at refrigeration temperatures, at least triplicating their shelf-life.

This level of decontamination would afford offering these longed-for, more varied, nutritious, and palatable unusual meals to immunocompromised patients, without risk of food borne diseases, as well as to the general public.

On-going and planned work for 2004:

• Widen the number of immunocompromised patients trying the above depicted irradiated lunch, in order to attain some statistical results.

• Studying commercial refrigerated cooked ham and cheese sandwiches, and pies of varied fillings, under a similar protocol as described for the other meals in this report.

• Looking for ways of lowering the radiation damage on colour and texture of those fruits and vegetables employed in this work.

CHINA

Use of Irradiation to Ensure the Safety and Quality of Chinese Dumpling

Chief Scientifi Investigator: Sun Baozhong

Co-investigators: Jia Chunfeng , Guo Yaping,,Li Haipeng, Ma Aijin Institute of Animal Science Chinese Academy of Agricultural Science

Gao Meixu, Li Shurong Institute of Atomic Application in Agricultural Science Chinese Academy of Agricultural Science

Research Contact No.: 11995

Abstract

Chinese Dumplings are popular ethnic prepared meal in China. The shelf-life of fresh cooked dumpling state is needed to extend. In this study, we mainly make the research on the aspect of the effects of irradiation on the survival of Salmonellae or Staphylococcus aureu s which are pathogenic bacteria normally in the prepared meal and its effects on the sensory, nutrition and physic and chemical characteristics of dumpling with vacuum package. The results shows that the safety of and composition of the experimental dumpling is a quality according to China industrial standard and Chinese enjoy the it for which has high likeness. The D10 values and D value of Salmonella in cooked dumpling were 0.9928, 0.31kGy and 1.63kGy, and the R value, the D10 values and D value of staphylococcus aureus were 0.9727, 0.44 kGy and 2.55 kGy. The vacuum packaging and irradiation treatment with2and 3kGy dosage may extend the shelf-life of dumpling remarkable and have some influences on and sensory quality.

1. INTRODUCTION

Chinese Dumplings are popular ethnic dish in China which contains abundant nutrient and many kind of food, such as every kind of edible meat, fish, sea food, vegetable, egg, wheat and rice flour and so on, all the housewife, cooker, and producer may adjust its composition according to their performance .The shelf-life of cooked Chinese Dumplings prepared in traditional manner is about 1~2d under ambient conditions and 2~5d under chilling condition, so Chinese Dumplings are mainly prepared and marketed at raw and frozen state. The flavor and convenience of commercial product of Chinese Dumplings need to be improved by irradiation or other technique.

In the first stage, we have found Enterococcus gallinarum and Bacillus macerans are the main type of bacteria exist in Chinese Dumpling and found that irradiation may extend the shelf- life of Chinese Dumpling remarkable. The aim of this stage is to determine the microbial type contaminated in Chinese Dumpling under commercial condition, to determine the D10 values of pathogenic bacteria potential in cooked dumpling, and study the effect of a selected dose of irradiation on the sensorial and nutritional quality of cooked dumpling.

2. MATERIALS AND METHODS

2.1. Dumpling samples and its treatment

Dumpling samples were acquired from Beijing Duolingduo Dumpling Company, which is processed with pork, lard, Chinese onion, spices and wheat flour. Salmonella (50040) and Staphylococcus aureus (26073) were obtained from National Institute For The Control of Pharmaceutical And Biological Products.

For the test of the D10 values of potential pathogenic bacteria in cooked dumpling: firstly the sample of dumpling were cooked in the boiling water for seven minutes, and then minced and sterilized at 121°C for 20 min. After the minced and sterilized dumplings were chilled to 20~25°C and inoculated the 105cfu/ml solution of Salmonellae or Staphylococcus aureus into the minced and sterilized dumpling and then packaged them in vacuum bags with a thickness 3 2 -1 of 0.075mm, a O2 transmission rate of 5 ml/cm 100 µm(m .24h) , which is a compound of Biaxial Oriented Polyamide (BOPA) and Low-Lensity Polyethylene (LDPE). The vacuum degree is -0.1MPa, the sealed temperature and time is 135°C for 3s incubated for 12h at 36°C. The packages were randomly divided into seven groups and irradiated with 0.5, 0.75, 1.0, 1.25, 1.5, 2.0 kGy. The control and the irradiated sample stored in the refrigerator at 0-4°C and the count of Salmonellae or Staphylococcus aureus were detected at 0, 3, 7, 14 and 21d during the storage.

Dumpling samples for the test of the sensory and the chemical and nutritional character: After cooked, the dumplings were packaged One group was used as the control (0kGy), while the other two groups were designated for the irradiation treatment at the target dosages of 2.0 and 3.0kGy. After irradiated, all dumpling samples were stored in the refrigerator at 0-4°C. The sensory, chemical and nutritional analysis were made at 0,7,14 and 21d during storage.

2.2. Microbiological analysis

The total count of bacteria was carried using 25 g of each bag of sample and blending in 225 ml peptone water. Additional decimal dilutions were prepared for plating in Plate Count agar and the plates were incubated at 36±1°C for 48h before counting the colony forming units. The results were expressed as the logarithm of the colony forming units per gram.

2.3. Sensory analysis

According oneself likeness, seven semi-trained panellists aged between 28-40 years using a 9 point hedonic scale. A score of 4= like extremely, 3= like very much, 2= like moderately, 1= like slightly, 0= neither like nor dislike, -1= dislike slightly, -2= dislike moderately, -3= dislike very much, and -4= dislike extremely.

2.4. Chemical analysis

Sample pH was measured using a pH/mV/Thermometer IQ150 according to GB/T9695.5- 1988. The analysis methods of POV, TVBN, Aw were following The Handbook of Food Chemistry edited by Yashan Han.

2.5. Nutritional analysis

The content of protein, fat of the Chinese Dumpling was analyzed by the Feed and Food Analysis Center of Institute of Animal Science, Chinese Academy of Agricultural Science according to GB/T9695.11-1988, GB/T9595.7-1988.

2.6. Statistical analysis

Data were processed by the General Linear Model (GLM) of Statistical Analysis System (SAS, 2000). Mean values and standard deviations were reported, and the differences in the mean values were compared by the SNK (Student-Newman-Keuls) multiple range test.

3. RESULTS

3.1. Characteristics of experimental dumpling products

According to the quality requirement of SB/T10289-1997 “The acceptable quality of the frozen flour and rice food. The result of the determination of protein, fat and water content, water activity and pH showed that the experimental dumpling products is acceptable product in which the microbial flora may easily increase and cause it decayed and unsafe.

3.2. Determination of D10 and D value of irradiation on Salmonella and Staphylococcus aureus

The R value, the D10 values and D value of Salmonella in cooked and vacuum packaged dumpling were 0.9928, 0.31kGy and 1.63kGy, and the R value, the D10 values and D value of Staphylococcus aureus were 0.9727, 0.44 kGy and 2.55 kGy. If the cooked dumpling contaminated by Salmonellae and Staphylococcus aureus, the irradiation dose should not less than their D value.

3.3. Effects of selected irradiation on the sensory, nutrition and physicochemical characteristics of the cooked and vacuum packaged dumpling

All the sensory characters except shape, freshness, tenderness, juiciness of control samples, 2kGy samples and 3kGy samples had significant difference with the storage time extended. Especially, the characters of wrapper of cooked dumpling have very significant difference. At seventh day, the most of characters reach their peak value, then, the most of characters of the wrapper of cooked dumpling continued to decrease, however, the changes of most of characters of the filling and whole of cooked dumpling are not significant. At the fourteenth day, the control sample produce unpleasant flavour, and scores of biting force, stickiness, elasticity were very lower, so we detected the control samples were decayed. At the twenty eighth day, biting force, stickiness, elasticity of irradiated samples were significantly lower than the other days.

The effect of irradiation dose and storage time on moisture content of the cooked dumplings is insignificant. With the storage time extended, the moisture content values of samples decrease very slowly. The effect of irradiation dose and storage time on Aw of the cooked dumplings was insignificant. The Aw values of samples were situated during 0.94-0.99. With the increase of irradiation, the pH value decreased. The effects of irradiation dose and storage time on POV(%) and TVBN(mg/100g) of the cooked dumplings are significant. With the increase of irradiation doses and the storage time extension, the value of POV and TVBN increase. But, at 28d, the POV values does not exceed standard. With irradiation doses increased, the TVBN

values increase very slowly. The shelf-life of cooked dumpling could be extended by irradiation.

4. CONCLUSIONS

Considering D10 value and D value of Salmonella of irradiation and the survival of Salmonella during storage, the irradiation dose to prevent the food-borne disease of Salmonella for cooked dumpling should not be less than and 1.63kGy, 2kGy dose would be the useful irradiation dose for Salmonella safety and more one weeks shelf-life. The irradiation dose to prevent the food-borne disease of staphylococcus aureus for cooked dumpling should not be less than and 2.55kGy, 3kGy dose would be the useful irradiation dose for Salmonella safety and more one weeks shelf-life .The effects of 2 and 3kGy irradiation on the survival of Salmonella and Staphylococcus aureus, and the vacuum packaging and irradiation treatment with 2 and 3kGy may extend the shelf-life of dumpling remarkable, but the decrease of sensory quality of should be confirmed by consumer test or improved with other methods.

Work plan for next year:

1. Confirm the effects of selected irradiation dose on the base of new challenge test and the exceeded research on the sensory quality of cooked and vacuum packaged dumpling further by consumer test.

2. Complete the research of the effects of irradiation on the nutrient, such as the vitamin, necessary unsaturated fatty acid, on the phys-chemical traits, such as TBA and other character missed in the research compared with other projects.

3. Find an acceptable method to improve defects such as the sticky wrapper or the off- odour filling with irradiation and storage.

GHANA

Irradiation Of Prepared Meals For Microbiological Safety and Shelf-Life Extension

Chief Scientific Investigator: Josephine NKETSIA-TABIRI

Co-investigators: A. Adu-Gyamfi, Alex Owusu-Biney, F. Apea Bah, F. Akomeah-Adjei Biotechnology & Nuclear Agriculture Research Inst. Ghana Atomic Energy Commission LEGON/GHANA

Research Contract No: 11904

1. INTRODUCTION

Current trends suggest increasing world-wide demand for convenience foods such as prepared meals and those that require minimum preparation before consumption [1]. In Ghana, the trend is the same with ready-to-eat or prepared meals mostly sold in the informal sector, playing a major role in meeting the nutritional needs of the people these meals are prepared on daily basis as in most developing countries [2]. With increasing urbanization and rapid adoption of refrigeration for food preservation in developing countries, the demand for such foods in supermarkets should be expected.

Over 90% of ready-to-eat meals prepared in Ghana are marketed under ambient conditions and have shelf-life of less than 12 hours. However, there are a few caterers who prepare cook- chill meals using hygienic and preventative approach (eg HACCP) in order to enhance the safety of their meals. Preliminary investigations have demonstrated that these cook-chill meals are potential sources of pathogenic bacteria and have been implicated in food-borne disease outbreaks [3,4]. There is therefore the need to ensure the safety of ready-to-eat meals through effective processing such as irradiation.

2. OBJECTIVES a) To characterize poached chicken meal; b) To determine the effect of irradiation on the microbiological quality of chilled poached chicken meal; c) To determine the effect of irradiation on the sensorial quality of chilled poached chicken meal; and d) To determine the shelf-life of chilled non-irradiated and irradiated poached chicken meal.

3. METHODOLOGY

3.1. Preparation, packaging and irradiation of poached chicken meals

Two batches of the cook-chill poached chicken meals consisting of rice, carrots, chicken and gravy were supplied by the caterer at different times. The chicken and gravy were together packaged in aluminum tray with covers while the fried rice and boiled carrots were also together packaged in aluminum trays with covers. The chilled meals were transported on ice to the gamma irradiation facility. The chicken and gravy packs were treated with 2 kGy and

the rice and carrot packs with 1 kGy in a gamma radiation facility at a dose rate of 2.6 kGy/h. After irradiation both the controls and irradiated meals were transported to the laboratory and stored in a refrigerator (4°C±1°C). Samples of the meals were subjected to physico-chemical, microbiological and sensory analyses during storage period of 22 days.

3.2. Microbiological, physico-chemical and sensory tests

Microbiological examination of the samples was carried out using standard methods [5]. Total viable count was determined on plate count agar; incubated at 36°C/48 h. The pH and free fatty acids of the meals were determined using standard methods [6]. Water activity was determined using water activity meter. Triangle, hedonic and acceptability tests were used by twenty-four panelists for the detection of differences between and preference for the sensory quality of irradiated and non-irradiated meals. All the meals were heated in a microwave oven (600 watts) for four minutes for rice/carrots and six minutes for chicken/gravy before the sensory tests.

4. RESULTS OBTAINED

4.1. Characteristics of chilled poached chicken meals

The water activity of the various components of the non-irradiated and irradiated meals during storage were similar, ranging between 0.930 and 0.948 (Table 1). The high water activity of the meals make them suitable for microbial proliferation hence the need to reduce initial microbial population and control temperature variations during storage. With respect to free fatty acids, the component of most interest is the chicken and gravy because of their relatively higher fat content. The free fatty acid content of the rice and carrots was low (<0.01%) and stable likewise the free fatty acid content of the chicken and gravy (0.03%). The results suggested that the irradiation treatment and chilled storage did not enhance fat hydrolysis. These observations may be attributed to the low radiation doses and chilling temperature applied. The pH of non-irradiated carrots experienced greater variations (4.7-6.3) during storage than their irradiated equivalents (6.3-6.4) than rice, chicken and gravy components. The sharp drop in pH of the non-irradiated carrots might in part be attributed to temperature abuse experienced by the meals due to power outage within the first week of refrigerated storage. It is note-worthy that the treatment of carrots with 1 kGy radiation stabilized pH for over 14 days refrigeration irrespective of the unexpected abuse temperature experienced.

4.2. Microbiological quality of non-irradiated and irradiated cook-chill meal during chilled storage

From the results, in Table 2 the initial TVC for the non-irradiated meals was low. Each of the meal components had counts below the detection limit of 102 CFU/g. The observed low counts can be attributed to the supplier’s strict adherence to the HACCP plan in the preparation and post-production handling of the meals. After irradiation, TVC was also below detection limit of 102 CFU/g. Although counts increased during storage, TVC for the irradiated meals were generally lower than the control. For example, while TVC for irradiated rice was below 104 CFU/g on day 21, TVC for non-irradiated rice exceeded 106 CFU/g. The TVC for non-irradiated carrots on day 14 was 103CFU/g but below detection limit for the treated ones. The TVC for non-irradiated chicken and gravy exceeded 106 CFU/g on day 14 but was below detection limit for irradiated chicken and gravy. After 21 days storage TVC of irradiated chicken was below 105CFU/g. Thus irradiation (2kGy) extended microbiological shelf-life of chicken and gravy to 14 days compared to seven days for the control.

4.3. Sensory Evaluation

No significant difference (p>0.05) was observed between the irradiated and non-irradiated meal components (i.e. rice, carrots, chicken/gravy) on day 0 and 3 during refrigerated storage. On the seventh day, significant difference (p>0.05) was observed between irradiated and the non-irradiated carrots, but panelists found both samples acceptable in terms of all three attributes (taste, colour, texture). After 11 days storage, highly significant differences (p<0.01) between the irradiated and non-irradiated carrots were observed; panelists however found the irradiated carrots acceptable in all three attributes but the non-irradiated carrots were unacceptable in terms of taste. In the case of rice, panelists detected significant differences (p<0.05) between the irradiated and non-irradiated samples after 14 days storage. Panelists did not detect significant differences (p>0.05) between the irradiated and control chicken/gravy after seven days storage. After 14 days storage, panelists detected significant differences between the irradiated and non-irradiated chicken/gravy; however, the irradiated chicken/gravy was acceptable to the panelists. The results of the preference tests using nine- point hedonic scale indicated that after 15 days of storage, non-irradiated gravy was scored (4.1) below the limit of acceptability and was rejected after 22 days. Irradiation however controlled adverse changes in sensory quality and extended the shelf-life of gravy to 15 days. The results further suggested that irradiation extended sensory shelf-life of rice to 22 days since the non-irradiated rice was rejected by the 22nd day. No significant difference was observed between scores for irradiated and control chicken after nine days refrigeration. After 15 days, however, non-irradiated chicken was poorly scored (3.9 for taste) and rejected on day 22. Irradiated chicken however had scores significantly higher scores for taste (7.2), on day 15 and slightly lower score for taste (6.5) on 22 day.

5. CONCLUSIONS

The sensory quality of the non-irradiated poached chicken meal, was limited to seven days under refrigeration. Treatment of rice and carrots with 1 kGy and chicken and gravy with 2 kGy improved microbiological quality and extended sensory quality of the meal to 14 days.

6. FUTURE STUDIES

1. Determination of fecal coliform and staphylococci counts on the meals during storage.

2. Determination of radiation sensitivity of Staphylococcus aureus and E. coli in poached chicken meal.

3. Determination of shelf-life of the meals under potential abuse temperature during refrigerated storage.

4. Characterization of all of rice with chicken and vegetables (pepper and carrots)

5. Determination of the microbiological profile of the meal.

6. Determination of radiation sensitivity of spoilage and pathogenic bacteria isolated from the meal.

7. Determination of sensory quality and shelf-life of the irradiated meal stored at ambient and under refrigeration.

Acknowledgements

The authors gratefully acknowledge the financial support by the International Atomic Energy Agency and the technical assistance of Messrs E. Akolmolga, T. Mahami, D. Datohe and C. Owulah.

REFERENCES

[1] FSIS (1999) Refrigeration and food safety, USDA, http://www.fsis.usda.gov. [2] GLSS: Ghana Living Standards Survey, Ghana Statistical Department (1988/89; 1998/99. [3] Nketsia-Tabiri, J., A. Adu-Gyamfi, and A. Owusu-Biney, Irradiation of prepared meals for microbiological safety and shelf-life extension, in Microbiological quality of waakye and cook-chill meals. Technical Report (2003). [4] Bryan (1990) Application of HACCP to ready-to-eat chilled foods. Food Technology (7), 70-77. [5] Speck, M.K., Compendium of methods for the microbiological examination of foods, American |Public Health Association], Washington, D.C. (ed) 1976. [6] Pearson, D., Chemical analysis of foods (1976).

Table 1. Range of selected characteristics of Poached Chicken Meal during 22 days chilled storage Component Treatment pH %FFA Aw Rice 0 kGy 5.8 - 6.2 <0.01 0.937 - 0.942 Rice 1 kGy 5.9 - 6.1 <0.01 0.940 - 0.946 Carrots 0 kGy 4.7 - 6.3 <0.01 0.938 - 0.953 Carrots 1 kGy 6.3 - 6.4 <0.01 0.944 - 0.946 Chicken/Gravy 0 kGy 5.3 - 5.7 0.03 0.934 - 0.943 Chicken/Gravy 2 kGy 5.2 - 5.7 0.03 0.936 - 0.944

Table 2. Changes in microbiological quality (Total Viable Count, CFU/g) of components of poached chicken meal during 21 days chilled storage. Meal Component Dose (kGy) Storage Time (days) 0 7 14 21 Rice 0 < 102 < 102 9.7 x 103 4.9 x 106 Rice 1 < 102 2.0 x 103 < 102 3.1 x 102 Carrots 0 < 102 < 102 2.5 x 103 NA Carrots 1 < 102 < 102 < 102 NA Chicken/gravy 0 < 102 < 102 > 106 NA Chicken/gravy 2 < 102 < 102 < 102 4.7 x 104 < 102 = below detection limit NA = not analysed

Table 3. Sensory scores for irradiated and non-irradiated poached chicken and rice stored at 4 ± 1ºC for 22 days Component/Dose Storage Time (days) 9 Days 15 Days 22 Days Taste Colour Texture Taste Colour Texture Taste Colour Texture Rice 0 kGy 7.2 7.4 6.5 7.4 7.4 7.4 rejected rejected rejected Rice 1 kGy 7.6 7.3 6.6 7.3 7.0 7.3 7.3 7.2 7.1 Chicken 0kGy 7.1 7.5 6.8 3.9 5.3 5.8 rejected rejected rejected Chicken 2 kGy 7.4 7.6 6.9 7.2 6.4 6.6 6.5 7.3 7.5

GREECE

Improving the Microbiological Quality and Safety of Prepared Meals Consumed in Greece by Low Dose Irradiation

Chief Investigator: I.N. Savvaidis

Co-investigators: K. Riganakos, M.G. Kontominas Laboratory of Food Chemistry and Microbiology Department of Chemistry University of Ioannina Ioannina Greece

Research Contract No.: 12271

1. SUMMARY OF THE PROPOSED RESEARCH

The proposed research of this CRP will address the use of low-dose-gamma irradiation (< 3 kGy) to extend the shelf-life and ensure the microbiological safety of prepared meals, marketed as refrigerated products. Such products are recently of increasing demand in Greece. Prepared meals will include salad meals that are very popular in Greece i.e. (eggplant salad, yoghurt/garlic salad), traditional sausage products, smoked trout etc. The first part of the proposed project will focus on the effect of low dose irradiation on shelf-life extension of above products. Assessment will include microbiological, chemical, and sensory parameters. Chemical parameters will include: pH, TBA (degree of lipid oxidation), TVB-N, TMA-N, vitamin and biogenic amines content. Microbiological parameters will include: Total viable counts, Enterobacteriaceae, Pseudomonas spp., Yeasts/Molds, Clostridium spp., Listeria spp. and Salmonella spp. Sensory assessment of non- and irradiated meals will include: evaluation of appearance (color, texture), taste, and odor. In the second part of the proposed project, a series of challenge experiments will be conducted regarding the microbiological safety of prepared meals. Food products will be inoculated with pathogens i.e. Listeria monocytogenes and their survival will be monitored as a function of irradiation dose and storage time. Experiments will be carried out at two different temperatures i.e. 4 and 10oC.

2. SCIENTIFIC SCOPE OF THE PROJECT

The overall objective of this CRP is to benefit the food industry and consumers in countries that use irradiation in terms of microbiological safety, increased shelf-life, quality, storage and distribution, and convenience of such food products. It must be mentioned that the use of gamma irradiation for microbial decontamination and disinfection is currently limited and applied only to certain types of foods in Europe (Directive 1999/2/EC, 1999/3/EC).

It is hoped that the use of irradiation will be approved in the immediate future for categories of foods such as prepared meals and ready-to eat-foods, since there is an increasing demand worldwide for such products.

The specific objectives of this CRP are:

1. Evaluation of the effect of gamma irradiation as to its ability to extend the shelf-life of Greek prepared meals, stored under refrigeration;

2. Determination of sensory changes (texture, appearance, taste etc.) using subjective and instrumental methods;

3. Evaluation of the effectiveness of low-dose irradiation process in controlling growth of Listeria monocytogenes in Greek prepared meals;

4. Determination of D10 values of Listeria monocytogenes in prepared meals consumed in Greece;

5. Determination of the effect of temperature (4, 10oC) on survival of Listeria monocytogenes population following gamma irradiation.

3. MATERIALS AND METHODS

3.1. Samples preparation and storage conditions

Samples (selected salads, sausage products, smoked trout, etc.) commercially produced and packaged will be transported to the irradiation plant in ice and irradiated immediately after sample collection. Irradiation of samples will be carried out at 4oC. A set of control samples (non-irradiated) will be stored under refrigeration at 4 and 10oC (abuse temperature). After irradiation, samples will be transported back to the laboratory in ice, and stored at 4 and 10oC until analyses.

3.2. Detailed Work Plan for First Year-Proposed Methods

During the first year, the work plan of the proposed study will focus on the determination of shelf-life of prepared meals. The prepared meals will include eggplant salad, tzatziki (yoghurt/garlic salad) as well as sausage products (salami aeros) and smoked trout.

3.3. Irradiation

Irradiation processing will involve the use of gamma irradiation at low doses in the range 0-3 kGy. Gamma irradiation will be carried out in the ELVIONY (Attica, Greece) plant, being the only commercial facility available in Greece using a 60Co source. It should be mentioned that the research team proposing this study is the only team in Greece working on Food Irradiation.

3.4. Analyses

Analyses at predetermined time intervals will include evaluation of microbiological, chemical, and sensory parameters of control (non-irradiated) and irradiated samples, stored under refrigeration at 4 and 10oC.

3.4.1. Microbiological analysis

Microbiological analyses will include the determination of the following bacterial species: Mesophilic aerobic flora, Pseudomonas spp., Enterobacteriaceae, Lactic acid bacteria, Brochothrix thermosphacta (meat and meat products), H2S-producing bacteria (including Shewanella putrefaciens) (fish products), Salmonella spp., Staphylococcus spp., Listeria spp. and Yeasts/Molds.

3.4.2. Chemical analysis

Chemical analyses will include determination of the following parameters: pH, lipid oxidation (TBA), volatile bases (TVB-N and TMA-N), biogenic amines (BA’s), and vitamin content and organic acids.

3.4.3. Sensory analyses

Sensory analyses will be conducted on prepared meals and will include evaluation of meals’ color, taste, odor and texture. Color and flavor will also be determined objectively using a Hunter lab colorimeter and GC/MS combination respectively.

4. EXPECTED OUTPUTS

Results of this CRP should lead to the formation of a database with specific data on the effectiveness of low-dose irradiation to extend the shelf-life of prepared meals. Data so far are lacking with regard to microbiological, nutritional and sensorial quality of prepared meals consumed in Greece.

1. Useful data will also be generated on the improvement of microbiological safety of Greek prepared meals, stored under refrigeration.

2. Through this CRP an opportunity will be given to postgraduate students of the University of Ioannina to be trained in the important fields of Food Preservation (Food Irradiation), Food Microbiology and Food Safety.

HUNGARY

Improvement of the Microbiological Safety and Shelf-Life of Selected Chilled Meals By Gamma Irradiation*

Chief Investigator: József Farkas BKÁE Department of Refrigeration and Livestock Products’ Technology Budapest, Hungary

Contract No. 11905, time period covered: 1 January 2002 - 31 December 2003

* A summary prepared for the information of participants of the 2nd Research Coordination Meeting of the Coordinated Research Project on Irradiation to Ensure the Safety and Quality of Prepared Meals to be held in Pretoria, 26-30 April 2004.

1. INTRODUCTION

There is a growing interest to market semi-prepared and prepared meals, eventually packaged in modified atmosphere packaging (MAP) and distributed under chilled conditions instead of frozen state. Such products are less energy-demanding and more attractive for the consumers than frozen meals. However, they are non-sterile and potential survival of some pathogenic microorganisms and/or post-processing contamination before packaging create microbiological risks and a considerable limitation of shelf-life, especially under abusive temperature conditions, frequently occurring during retail display period and home storage.

The objective of our research contract work was the demonstration of the technological feasibility of preventing the above risk by gamma irradiation of commodities such as a reconstituted breaded turkey breast meat with cheese and ham filling (called „Cordon Bleu”), filled pasta products (an European type: ”Tortellini”, and a Syrian type „Kouba”), and „sous vide” mixed vegetables.

2. EXPERIMENTS AND RESULTS OBTAINED

(Materials and methods have been described in detail in our first and second progress reports submitted to the IAEA in due course).

2.1 Cordon Bleu

2.1.1. Product parameters

o The aw of the product revealed to be 0.96 at 24 C, the pH was in the range of 6.24 - 6.30. The individual „steaks” were vacuum-packaged into „Multibarrier-4” laminated foil puches, and refilled to 750 bar pressure with a gas mixture of 40% CO2 and 60% N2.

2.1.2. Preliminary tests

Microbiological testing of unirradiated batches of Cordon Bleu showed that even some vegetative microorganisms can either survive inside the product the pre-frying process or recontaminate it before packaging. A radiation treatment by 3 kGy, which did not diminish significantly the sensorial properties, resulted more than three log cycles reduction of the total viable cell count and reduced the bacterial spore count by approx. 1 log cycle.

Testing of the unirradiated product during refrigerated storage showed that:

• in aerobic packaging the microbial shelf-life even at 4o C would be less than 14 days because of mould growth on the surface;

• under modified atmosphere packaging the microbiological shelf-life was longer, but variable, two to three weeks at 8o C, and three to longer than four weeks at 4o C;

• degradation of visual quality (the appearance) occurred within three to four weeks of chilled storage;

• in the majority of unirradiated experimental samples the psychrotrophic total viable counts increased steadily and latcobacilli seemed to become the dominant spoilage bacteria in the MAP product;

• no sulphite-reducing clostridia could be found in any samples during the entire storage;

• the counts of the aerobic/facultative anaerobic bacterial spores detected remained practically unchanged during the whole duration of refrigerated storage.

2.1.3. Inoculated pack studies

Due to re-organization of the producing companies, only the first challenge test of those planned for 2003 could be performed: namely using a psychrotrophic strain of Bacillus cereus, the same organism, which has been utilized formerly by us studying the improvement of microbiological safety of sous-vide meals by gamma irradiation (Farkas et al., 2002, 2003). The test organism and the preparation of the spore population as an inoculum have been detailed also in those papers. The aqueous spore suspension has been „heat-activated” at 60oC for 30 min heat treatment prior inoculation. On the basis of an average mass of 125 g of the individual Cordon Bleu steaks, 0.15 ml spore suspension was distributed in 10 µl aliquots on the surface of the lightly pre-fried steaks, in order to obtain a spore contamination of appr. 104 spores per g of the test material. Untreated and 4 kGy irradiated samples formed the experimental batches. Both batches were divided into sub-batches for 5 and 9o C storage temperatures. Storage was continued until four weeks and microbiological testings were performed at the day following irradiation and every week during storage.

Radiation treatment did not change the appearance of the samples and soon after irradiation no off-odour was noted at the opening of the pouches. However, after two weeks of storage, the bread-crumb surface layer started to develop a less freshly fried appearance, independent from the radiation treatment and the cheese slices inside the unirradiated samples started to liquify, more intensely at 9o C. During the next two weeks, the samples were gradually loosing their fresh appearance and produced some oily exudation. The untreated samples developed a „stale” odour, while in the irradiated samples a rancid odour was noted when they were prepared for microbiological testings.

Microbiological investigations revealed that even vegetative bacteria and yeasts survived the mild pre-frying, which makes it possible that eventually pathogenic microorganisms could not be inactivated by the producing technology. Detectable mould propagules in the surface layer showed that environmental contaminants might re-contaminate the pre-fried product before packaging. In the untreated samples, the initial total aerobic count was determined by the appr. 2.3x104 CFU/g level of the Bacillus cereus spores inoculated into the experimental batches. However, these spores remained dormant at both storage temperatures. After seven days of storage the lactobacilli, which have been in less than 103 CFU/g level at the beginning

of storage, became dominant in the viable count of the unirradiated samples, reaching unacceptable level (higher than 107 CFU/g) between the sevent and fourteenth days at 9o C, and between 21st and 28th days at 5o C. Enterobacteriaceae and sulphite reducing clostridia could not be found. Investigations of irradiated samples showed that the 4 kGy gamma radiation dose reduced only less than 1 log cycle the count of Bacillus cereus spores. This confirms the considerable radiation resistance of these spores as demonstrated already by our earlier experiments (Farkas et al., 2003). However, the spores of the test organism did not germinate or grow in the present product even at 9o C during the four weeks storage, perhaps because of the MAP and the 0.96 aw level. They were readily germinating and developing at 5 - 7o C in our earlier experiments in BHI broth (Farkas et al., 12003). The radiation treatment greatly eliminated the lactic acid bacteria, thus, the irradiated experimental batch remained microbiologically stabile for at least 4 weeks at 5 oC, and only one sample has shown growth of lactic acid bacteria and only on the 28th day sampling at 9o C, and only up to a 106 CFU level, i.e. two log-cycles less than the average count of lactobacilli on the 14th day sampling of the unirradiated batch.

Survival and growth of lactic acid bacteria in the unirradiated experimental batches make it probable that Listeria monocytogenes, an ubiquiter environmental contaminant might eventually be present, too. Its elimination by combining the pre-frying with irradiation will be the aim of our further challenge testing planned.

2.2. Tortellini

The water activity levels of the pasta products investigated in our experiments would not exclude the opportunity of growth and enterotoxin formation of Staphylococcus aureus if the pathogen would contaminate this type of product in the early stage of dehydration before marketing and retail storage. Because of the high heat resistance of Staphylococcus enterotoxins, the final cooking of these filled pastas cannot inactivate pre-formed toxins. Considering also the frequently abusive temperature of retail display, and the home storage, a challenge testing using Staphylococcus aureus was performed to investigate the feasibility of its elimination by a radiation dose of 3 kGy.

2.2.1. Product parameters

The composition of this pasta product made from grits of durum wheat flour, eggs and water, filling made from bread crumbs, smoked ham, cheese, salt, pepper and Na-glutamate. The product prepared by a commercial company had a pH of 5.5 - 6.1 and water activity level of o aw 0.96 at 25 C.

2.2.2. Inoculated pack studies

Tortellini pieces were inoculated individually to appr. 104 CFU/g level with 10 µl aliquouts of the suspension of Staphyloccoccus aureus and ten pieces each were packaged in the commercial packaging foil of the product under a gas atmosphere of appr. 20 % CO2 and 80% of N2. (The residual O2 content was 0.23 v%). Numerous replicate packages of the inoculated pasta were irradiated at 3 kGy dose, a sensorially acceptable dose level. Untreated and irradiated experimental batches were stored at 15o C representing a strong but not infrequent abusive temperature and total aerobic viable counts and Staphylococcus aureus counts were estimated periodically during storage.

The initial total aerobic cell counts of the inoculated unirradiated samples was dominated by the Staphylococcus aureus inoculum, because the total aerobic cell counts of the

uninoculated samples were two log cycles less (appr. log CFU = 2.3), composing mainly from aerobic bacterial spores. As an effect of irradiation, the Staphylococcus aureus count has been reduced below the detection limit, i.e. less than log CFU=0.26. Thus, the radiation treatment resulted more than three log-cycles reduction of the test organism in the pasta product. The aerobic spores of the „native” microbiota were reduced by somewhat less than one log cycle by the 3 kGy dose. The spore formers were unable to grow in the samples under the experimental conditions during eight days of storage, while the Staphylococcus aureus test organism grew up to 108 CFU/g level in the unirradiated samples. No Staphylococcus aureus cells recovered in the irradiated samples, even after 28 days storage at 15o C. The aerobic total viable cell counts in the irradiated samples started to increase only after eight days storage and their slow increase was due to some yeasts and micrococci forming distinctly different white colonies than the test organism. The thiamin content of the untreated samples was found to be 34 µg/100 g, with an accuracy of +/- 10%, and it was not changed by the radiation dose applied.

2.3. Kouba

A small batch of this Syrian pasta product were carried to our laboratory by Ms. YAKOUB, an IAEA fellowship holder from Damascus arrived for a research training in our laboratory. With Kouba, storage studies were performed at 10o C after re-packaging the samples aerobically into plastic pouches and threating the small experimental batches by 0, 2 and 4 kGy doses, respectively. Microbiological tests of this product were carried out estimating total and psychrotrophic aerobic bacterial counts, Enterobacteriaceae, moulds and yeasts and suspected Staphylococcus aureus counts, the latters by Baird-Parker agar and additional confirmation tests. The pH of the pasta product was 5.8 and the aw value 0.96. The total bacterial counts were reduced by four log-cycles at 2 kGy dose and by five log-cycles at 4 kGy dose level, respectively. The initial Enterobacteriaceae count (3.9x104 CFU/g) has been reduced under the detection limit (5 CFU/g) even by 2 kGy. A slow re-growth of Enterobacteriaceae was noted to the initial level in the irradiated samples during two-week storage but in the unirradiated samples their value was around 108 CFU/g already after one week. Considerable numbers of suspected Staphylococcus colonies were also isolated from the untreated samples, however, their number showed a 5 log-cycle reduction in the irradiated samples. The unirradiated samples showed unacceptable numbers of suspected Staphylococcus already after seven days. In this sense the radiation treatment extended the safe storage time by one week. Yeasts were more radiation resistant compared to the other investigated components of the microbiota of Kouba.

2.4. Sous-vide mixed vegetables

2.4.1. Test material

Mixed vegetables composed from pre-cut beans, carrot cubes, shelled maize corn and cauliflower pieces has been pre-cooked to doneness before the inoculated pack studies were performed with vacuum-packaging and sous vide heating, either without or with radiation treatment. The pH of the product was almost 7.0 and the water activity was higher than 0.98.

2.4.2. Inoculated pack studies

Listeria monocytogenes 4ab strain. No. 10 was used as test organism and for inoculation the vegetable mixture was dipped into its aqueous suspension for 10 min. After inoculation vacuum-packaged portions in laminated foil (Farkas et al., 2003) have been heat treated in a

controlled temperature water bath following the internal temperature of certain samples by a heat-penetrometer until a heat-treatment equivalent of 70 oC, 2 min was recorded. After cooling several heat-treated packages have been irradiated at 2 kGy, a sensorically acceptable dose and stored together with the unirradiated batch of packages at 10o C as a slightly abusive temperature for two weeks.

In the uninoculated samples the mesophilic aerobic total count amounted to log CFU/g=4.88, with non-detectable level of bacterial spores (log CFU/g < 0.3). This „native” microbiota has increased to approx 107 CFU/g level during the two-week storage at 10o C. The dense inoculum of Listeria monocytogenes caused a Listeria contamination of 108 CFU/g level in the non-irradiated samples before the sous-vide heating. The very mild sous-vide cooking resulted in only 1.5-2 log units reduction of the Listeria count, whereas irradiation resulted 4.5-5 log units reduction. During one week of storage at 10o C, the Listeria grew up to a maximum level, close to 109 CFU/g in the unirradiated sous-wide cooked samples. The irradiated sous-vide samples showed also a considerable re-growth, although their Listeria count was only1 to 10 % of those of the non-irradiated respective samples.

3. WORK PLAN FOR 2004

A. Inoculated pack studies (challenge test) with the pre-fried meal „Cordon Bleu” packaged under modified atmosphere (CO2 + N2). Test organism: Listeria monocytogenes 4ab No.10, radiation dose: 2 kGy. Storage of experimental batches both at 5o C and 9o C. Besides microbiological testings, lipid oxidation in the product as affected by irradiation and storage time and temperature will be followed. Detailed experimental plan has been described in the completed request for renewal of research contract, submitted to the IAEA in December 2003.

B. Utilizing an EU-sponsored international microbiological data base called „eComBase” (Baranyi and Tamplin, 200) to estimate growth potential of selected spoilage and pathogenic bacteria on the basis of our experiences and observations gathered during our present project with various chilled semi-prepared or prepared meals as a function of the relevant microbiological ecological parameters of the specific food products. This extensive data base will be made available to us.

C. The results of these computer studies will be compared to those of our previous studies, and they will assist to validate our former observations. They can serve similar evaluations of other projects involved in the present coordinated research programme. The use of this data base might minimize the need for time consuming and expensive traditional challenge tests. Our expected experience with this predictive microbiological project will be presented to other CRP partners at future research coordination meetings.

REFERENCES

[1] BARANYI, J., and TAMPLIN, M. L., ComBase: a combined database on microbial responses to food environments. 1st International Conference on Microbial Risk Assessment: Foodborne Hazards, College Park, Maryland. p. 23 (2002). [2] FARKAS, J., POLYÁK-FEHÉR, K., ANDRÁSSY, É., MÉSZÁROS, L., Improvement of microbiological safety of sous-vide meals by gamma irradiation. Rad. Phys. Chem., 63: 345- 348 (2002). [3] FARKAS, J., POLYÁK-FEHÉR, K., MOHÁCSI-FARKAS, C., MÉSZÁROS, L., ANDRÁSSY, É., SÁRAY, T., Studies on irradiation of pre-packaged prepared vegetables and improvement of microbiological safety of some sous-vide meals by gamma radiation. In: Radiation Processing for Safe, Shelf-stable and Ready-to-Eat Food. IAEA-TECDOC-1337. IAEA, Vienna, pp. 27-46 (2003).

INDIA

Radiation processing to ensure the safety and quality of ethnic prepared meals

Chief Scientific Investigator: R. Chander

Co-investigators: S. R. Kanatt, S. P. Chawla, V. Dhokane, A. S. Bawa* and Arun Sharma Food Technology Division, Bhabha Atomic Research Centre Mumbai, India

*Defence Food Research Laboratory, Mysore-570 011, India

Research Contract No.: 11996

1. INTRODUCTION

With rapid urbanization and change in socio-economic status there is an increase in demand for convenience ready-to-cook /ready-to-eat foods. Ready cooked meals, which are stored under frozen condition at -18° C or below are available in developed countries but none in Indian markets. In India, meals are consumed after cooking within a short period. No ready- to-eat (RTE) meals are marketed. However, a number of retort processed RTE meal components such as lentils (dals) and vegetables have started appearing in Indian markets. The retort process like freezing is high energy consuming and costly. The RTE meal market is becoming increasingly popular among consumers looking for high quality and safe products. Freezing facilities are expensive and inadequate in our country. Also freezing affects the texture of these products. A developing country such as India can ill afford high energy consuming food processes like freezing or retorting and are not economically viable. Consumers prefer chilled foods over frozen, these are perceived to be fresh-like and more convenient. The RTE chilled meals are of particular interest to India and other developing countries. However, cook-chill prepared meals have short shelf-life of maximum seven days at 4° C thereby limiting the geographical area in which they can be marketed. There are significant concerns about microbiological safety. Cooking eliminates vegetative pathogens but sometimes undercooking may lead to some of them surviving. In addition, there is always a risk of recontamination after cooking. L. monocytogenes is of particular concern being psychrotropic, it can grow at chilled temperatures. Technologies that allow for several fold extension of the shelf-life are therefore required. Radiation processing (1-2.5 kGy) of cook chill meal will not only result in a much safer food by eliminating non-sporulating pathogens but also in extension of shelf-life by decreasing the number of spoilage bacteria without jeopardizing their taste, and nutritive value. Therefore, investigations were carried out to examine the effectiveness of irradiation for extending shelf-life and improving microbiological safety of some of the ethnic preparations that included items from breakfast menu, meal components and complete meals.

2. MATERIALS & METHODS

2.1. Preparation of ready-to-eat-meals

A number of RTE meals were prepared with the ingredients as mentioned in Table 1. 100 g of each product was packed in sterile, low-density polyethylene bags (700 gauge; WVTR 0.4 g/m2/day); OTR 1800 ml/ m2 /day). For sensory evaluation 200 g of the sample were packed separately. Three sets of experiments were carried out for each product.

2.2. Irradiation

Irradiation of prepackaged products was carried out at melting ice temperature (1-3°C) in a Package Irradiator (Nordion Intl. Inc. Canada) with a 60Co source at a dose rate of 3kGy h-1. Non-irradiated lot served as control. All samples were then stored at 0-3°C.

2.3. Microbiological analysis

Samples (25 g) in duplicate from the irradiated and their corresponding control batches, were aseptically homogenized for one min with sterile saline in a Stomacher. Appropriate serial dilutions of the homogenate were carried out. Total plate count was determined using Plate Count Agar (PCA) incubated at 30° C for 48 h. Selective and differential media used were Baird Parker’s Agar (37° C for 24 h) for enumeration of Staphylococcus spp, Violet Red Bile Agar (44° C for 24 h) for fecal coliforms and Potato Dextrose Agar for molds incubated at 30° C for five days. Total aerobic spore count was also determined. 5 ml of the 10% homogenate was heated at 80° C for 10 min, cooled and then serial dilutions of it plated on PCA (30°C for 48 h).

2.4. Measurement of lipid peroxidation

Thiobarbituric acid-reactive substances (TBARS) produced from lipid peroxidation were determined using the method of Alasnier et al., (2000).

2.5. Sensory evaluation

The sensory attributes evaluated were appearance, flavor, texture and overall acceptability of the products using a 10-point hedonic scale. The panel consisted of 15-20 experienced members of the staff who were familiar with meals characteristics. Sensory analyses were carried out after steaming the samples for one to two minutes.

3. RESULTS AND DISCUSSION

3.1. Microbiological quality

The total viable counts of poha, upma, meals and meal components are depicted in Tables 2 and 3. The initial total aerobic count in non-irradiated varied with products and increased to more than 6 log cfu/g in many products in less than 14 days of storage and the samples spoiled. In irradiated samples, there was a dose dependent reduction and at 2 kGy total bacterial counts were significantly lower or non-detectable in some samples even after 28 days of storage. The enhanced safety of radiation-processed products was evident from the complete elimination of potentially pathogenic Staphyloccocus spp. Fungal growth was detected in some of the non-irradiated samples during chilled storage. Thayer and Boyd (1992) have reported that 90% of S. aureus in mechanically deboned chicken meat were killed by a dose of 0.36 kGy.

Fecal coliforms were detected in only one batch of non-irradiated control chicken chilly samples. In all the irradiated samples they were not detected. Aerobic spore count in non- irradiated chicken chilly also varied with samples and could not be detected in most of the samples on irradiation except in few samples. Microbial load, in particular aerobic spore counts in poha were fairly high and not completely eliminated by 2 kGy of gamma dose. Effect of low dose irradiation in reducing the bacterial load of meat and meat products has

been reported earlier from our laboratory (Naik et al., 1994; Kanatt, et al., 1997). Under aerobic conditions, the dominant spoilage organisms are the strictly aerobic Pseudomonas (Gill, 1986), which are very sensitive to irradiation (Maxy, 1982).

3.2. Lipid peroxidation

It was measured in terms of thiobarbituric acid reactive substances (TBARS). Non-irradiated control samples showed lower TBARS values than irradiated samples. The increase in TBARS values was dose dependent and also on storage (Fig.1).

3.3. Sensory evaluation

The overall sensory scores of irradiated and non-irradiated samples were not significantly different (Fig. 2). Appearance, flavor and texture of irradiated samples were not different from its non-irradiated control and all the samples were acceptable (data not shown).

4. CONCLUSIONS

Radiation processing of ethnic RTE meals results in a dose dependent decrease in the total viable counts and elimination of potentially pathogenic organisms such as Staphyloccocus spp. Irradiation at 2 kGy extends the storage life of these meals by more than two weeks compared to non-irradiated samples. Lipid peroxidation increases with radiation and should be controlled by packaging/addition of natural antioxidants Thus this study showed that irradiation in conjunction with chilled storage could be of advantage to the processor, retailer and consumer.

5. FUTURE PLAN OF WORK

The studies carried so far indicated the need to minimize lipid peroxidation of radiation processed meals to improve its acceptability. Plan of work for the next year is as follows:

• Vacuum and modified atmosphere packaging will be evaluated for decreasing the lipid peroxidation of the samples.

• Suitability of natural antioxidants from potato peel, mango peel, shrimp waste and curry leave extract will be examined for enhancing acceptability of radiation- processed meals.

• Migration of constituents from the packaging material during irradiation would be evaluated.

Table 1. Description of Ready to Eat (RTE) Meals ITEMS INGREDIENTS REMARKS BREAKFAST POHA Dried pounded rice, onion, spices, Warm and serve, few vegetable oil drops of fresh lime juice enhance its taste UPMA Semolina, onion, spices, vegetable oil Warm and serve with coconut chutney MEAL COMPONENTS MIX VEGETABLES Peas, beans, carrots, onion, ginger-garlic To be consumed with paste, spices, vegetable oil rice/roti (Indian bread) after heating CHICKEN CHILLY Boneless chicken, ginger-garlic paste, Goes well with rice/roti spices, vegetable oil when served hot PRAWN MASALA Deveined prawns, ginger-garlic paste, Warm and serve with spices, vegetable oil rice RICE Rice Serve with mix vegetables/ chicken or prawn curry COMPLETE MEALS VEGETABLE Rice, peas, French beans, carrots, onion, Serve hot and is a perfect PULAO ginger-garlic paste, spices, vegetable oil combination with curd CHICKEN BIRYANI Rice, chicken drumsticks, ginger-garlic Serve hot with curd paste, spices, vegetable oil PRAWN PULAO Rice, deveined prawns, ginger-garlic Serve hot and garnish it paste, spices, vegetable oil with coconut flakes KHICHDI Rice, lentils, spices, vegetable oil A good meal for patients and those with stomach upset

Table 2. Microbiological Quality of Breakfast and Meal Items Meal Dose Control 1 kGy 2 kGy Storage period 0 15 30 0 15 30 0 15 30 (days) log CFU/g Poha* TVC 3.4 5.5 6.7 2.7 4.05 4.4 2.3 2.1 4.0 SC 3.2 3.7 3.6 2.6 2.24 2.7 2.3 2.1 2.2 Upma* TVC 1.9 3.54 5.6 Nil Nil Nil Nil Nil Nil SC 1.3 1.3 1.7 Nil Nil Nil Nil Nil Nil Mix TVC 1.0 Nil 3.6 Nil Nil Nil Nil Nil Nil vegetable* SC Nil Nil Nil Nil Nil Nil Nil Nil Nil Chicken TVC 3.7 6.7 $ 2.0 3.7 5.9 1.4 2.0 5.0 chilly SC 1.2 1.4 $ Nil Nil Nil Nil Nil Nil S.aureus 2.3 3.6 $ Nil Nil Nil Nil Nil Nil Prawn TVC 2.2 2.3 2.3 1.5 1.5 1.7 Nil Nil Nil masala* SC 1.9 2.2 2.2 1.0 Nil Nil Nil Nil Nil Rice* TVC 1.0 2.6 5.6 Nil Nil Nil Nil Nil Nil SC Nil Nil Nil Nil Nil Nil Nil Nil Nil

Table 3. Microbiological Quality of Complete meals Meal Dose Control 1 kGy 2 kGy Storage period 0 15 30 0 15 30 0 15 30 (days) log CFU/g Vegetable TVC 2.6 6.5 6.4 Nil Nil Nil Nil Nil Nil pulao* SC 2.0 1.3 2.8 Nil Nil Nil Nil Nil Nil Chicken TVC 2.9 3.0 4.3 2.4 2.3 1.6 2.3 2.2 2.0 biryani* SC 1.5 2.7 2.4 2.0 1.3 1.7 2.1 1.8 Nil Prawn TVC 2.7 3.7 5.6 1.0 1.5 Nil Nil 1.3 Nil pulao* SC 1.7 Nil 1.7 Nil Nil Nil Nil Nil Nil Khichadi* TVC 1.69 1.84 # Nil Nil Nil Nil Nil Nil SC Nil Nil # Nil Nil Nil Nil Nil Nil TVC= Total Viable Count SC= Spore Count # = visible fungus seen and so sample not analysed $= sample not analyzed as it had spoiled *= Staphylococcus spp., fungus and fecal coliforms were not detected

Control 1 kGy 1.5 2.5 5 Poha Upma Chicken chilly 2 kGy

4 2.0

1.0 3 1.5

2 1.0

0.5 1

0.5

0 015300153001530

2.0 2.0 4 Prawn m asala Mix vegetable Chicken biryani

1.5 1.5 3

1.0 1.0 2

0.5 0.5 1 TBA value (mg MDA/kg sample) MDA/kg (mg value TBA 0.0 0.0 0 015300153001530

2.5 1.5 1.5 Vegetable pulao Prawns pulao Khichadi

2.0

1.0 1.0 1.5

1.0 0.5 0.5

0.5

0.0 0.0 0.0 015300153001530

Storage period (days) Fig. 1 Lipid peroxidation measured as TBARS of radiation processed meals

Control 10 1 kGy 2 kGy

0 Chicken biryani Vegetable pulao Prawns pulao Khichadi 10

5 Sensory score (Hedonic scale) (Hedonic score Sensory

0 Chicken chilly M ix vegetable Praw ns curry Fig. 2 Sensory evaluation of radiation processed m eals

REFERENCES

[1] ALASNIER, C., MEYNIER, A., VIAU, M., & GANDMER, G., Journal of Food Science, 65, 9 (2000). [2] GILL, C. O., In: Advances in meat research-meat and poultry microbiology, Pearson, A. M. & Dutson, T. R. Eds., AVI Publishing Co., Westport, CT (1986). [3] KANATT, S. R., PAUL, P., D’SOUZA, S.F., AND THOMAS, P., Journal of Food Safety, 17, 283 (1997). [4] MAXCY, R. B., Journal of Food Protection, 45, 363 (1982). [5] NAIK, G. N., PAUL, P., CHAWLA, S. P., SHERIKAR, A.T., & NAIR, P.M. Meat Science, 38, 307 (1994). [6] THAYER, D. W., & BOYD, G., Journal of Food Science, 57, 848 (1992).

INDONESIA

Irradiation To Ensure the Safety and Quality of Home Style Frozen Foods: 1. Liquid based materials

Chief Scientific Investigator: Z. Irawati*

Co-investigators: L. Natalia**, C.M. Nurcahya* and F. Anas*

*Centre for Research and Development of Isotopes and Radiation Technology National Nuclear Energy Agency JKSKL, Jakarta 12070 Indonesia

**Research Institute for Veterinary Science Bogor West Java, Indonesia

Research Contract No.: 11906

1. INTRODUCTION

The ready to eat meals which offer convenient and less time for preparation are marketed in most big cities in the world either under chilled with limited shelf-life or frozen for long term sale in supermarkets. Application of irradiation technology at high doses in combination with cryogenic condition along the process of some traditional shelf-stable foods could accelerate their safety and extend the shelf-life of various types of Indonesia ethnic dishes has been conducted. It is recognized that irradiation at pasteurization doses has also a potential role to improve the microbiological safety and shelf-life of a number of chilled prepared meals. These types of foods currently marketed under frozen condition could possibly be replaced through irradiation and in combination with chilled storage to ensure not only microbiologically safe but also sufficient shelf-life to meet market requirement resulting in more practical, quick preparation, saving energy and cost.

Objective of this study is to evaluate the effectiveness of irradiation as a method to ensure microbiological safety and extend the shelf-life of prepared meals such as home style frozen foods stored at chilled temperatures. The chemical characteristics and sensory quality of the treated products has also been conducted.

2. MATERIALS AND METHODS

2.1. Materials

2.1.1. Commercially prepared of home style frozen soups: black soup, ox-tail soup, chicken vegetable soup, chicken sweet corn soup were purchased from supermarket.

The product was put in a High Density PolyEthylene (HDPE) pouch followed by waxy-thin carton as secondary packaging material, and stored at –18o C prior to use for the research work. The netto quantity of each box was 400cc.

2.1.2. Self-making home style frozen soups

Formulation and basic materials used in making four different type of soups in order to develop Standard Operating Procedure (SOP) as follows:

a) Black soup: beef, pangium edule, shallot, garlic, roasted coriander, seedless red chili, ginger, small aromatic lemon leaf, roasted fish paste, turmeric, ginger root, lemon grass, kaemferia galanga, salt, palm sugar, bay leaf, palm oil, and water. b) Oxtail soup: oxtail, shallot, garlic, salt, palm oil, water, onion, ground nutmeg, cloves, ground white pepper, onion leaf, celery, and margarine. c) Chicken vegetable soup: Chicken, shallot, garlic, salt, water, ground nutmeg, ground white pepper, onion leaf, celery, margarine, sugar, carrot, green beans, brocolli, and frozen sugar peas. d) Chicken sweet corn soup: Chicken, salt, water, ground nutmeg, young sweet corn, chicken sausage, carrot, eggs, and corn starch.

2.2. Sample preparation and treatments

2.2.1. “Rawon”/Black soup

Beef -> boiled -> removed ->cut into cubical shape, 2x2x2 cm3. Shallot, garlic, candlenut, coriander, red chili and Pangium edule -> ground into form of mix-ingredients. Palm oil was heated and then mixed with the ingredients followed by adding roasted fish paste, turmeric, lemon leaf, ginger root and lemon grass ->scenting fragrance appeared. The cubical beef -> added into the mixture and fried for 10 min. 50% of broth ->poured into the spicy cooked beef -> added with some salt and sugar. The soup was ready after final cooking for 30 min.

2.2.2. Ox-tail soup, Chicken vegetable soup, and Chicken sweet corn soup had almost similar preparation but different in materials and ingredients used along the process.

2.2.3. Packaging material

Laminate pouch of PET 12µm/LDPE as adh.2 µm/Al-foil 7 µm/LDPE as adh./LLDPE 50 µm; and styrofoam boxes ( 33.75 x 36.25 x 51.25 cm3) filled with dry ice to maintain the internal temperature of the samples during radiation process.

2.2.4. Irradiation treatment

IRPASENA irradiator at the CRDIRT, NNAE, Jakarta. Cobalt -60 (capacity of ca. 20 kCi) at a dose rate of 3 kGy/h. Perspex calibration dosimeter to determine absorbed dose. Samples were irradiated with minimum doses of 1, 3, 5, and 7 kGy respectively.

2.2.5 Storage condition and Methods of analysis

Both treated and untreated samples were stored at 4o C in order to determine the effect of gamma irradiation on the microbiological assessments according to different international standards i.e., Total Plate Count (TPC), Total mould and yeast count (TMYC), Coliform (MPN), E. coli (MPN), Salmonella spp., Staphylococcus aureus, and Clostridium perfringens. It was considered to conduct CCP at each step of the process to determine the microbiological load of meat, ox-tail, tap water, ground seasonings and the processed soups. Quality evaluation for all type of soups was carried out after storage periods of 0 and three months. Moisture content was measured using gravimetry method while pH were measured using a Microcomputer pH meter LEC 60. Organoleptic attributes (colour, appearance, odour, taste,

and texture) were also measured. It was done by 10 to 20 selected panellists using a five-point hedonic scale. A score of 5 = excellent, 4 = good, 3 = fair, 2 = poor, and 1 = extremely poor.

3. RESULTS

Table 1. Microbiological assessments of commercially prepared four different type of soups stored for three weeks at 4oc before and after irradiation Parameters Dose Commercially prepared home style frozen soups (kGy Black soups Ox-tail soup Chicken Chicken sweet ) vegetable soup corn soup TPC 0 3.7 x 103 4.6 x 103 7.6 x 103 6.1 x 103 (cfu/g) 1 8.5 x 102 2.4 x 102 2.9 x 103 1.5 x 103 3 1.4 x 102 1.7 x 102 1.4 x 102 1.6 x 102 5 0 0.6 x 102 0.7 x 102 0.4 x 102 7 0 0 1.8 x 10 4.0 x 102

0 1.9 x 103 3.0 x 103 4.1 x 103 3.4 x 103 TMYC 1 1.0 x 102 1.5 x 102 2.7 x 103 1.4 x 103 (cfu/g) 3 0 1.3 x 102 1.5 x 102 1.2 x 103 5 0 0.5 x 102 0.6 x 102 0.5 x 102 7 0 0 0.2 x 10 0 Coliform, Escherichia coli, Salmonella spp., Staphylococcus aureus, and Clostridium perfringens did not grow nor negative either in untreated or treated samples by irradiation.

Table2. Microbiological load of water, seasonings, beef meat and beef ox-tail, and the soups at each step of processing of black and ox-tail soups prior to irradiation. Samples Total Plate Total Mould and Count/TPC (cfu/g) Yeast Count/TMYC (cfu/g) Tap water (not potable) 1.6 x 103 1.3 x 10 Cooked water (potable) 2.6 x 102 0 Raw beef meat 8.5 x 107 0 Raw ox-tail 5.3 x 107 0 Raw seasonings for black soup 1.5 x 107 3.9 x 103 Raw seasonings for ox-tail soup 1.1 x 106 0 Cooked seasonings for black soup 8.0 x 104 0 Cooked seasonings for ox-tail soup 6.0 x 103 0 Black soup after cooking 3.6 x 103 0 Ox-tail soup after cooking 5.6 x 103 3.6 x 103 *Average of three replications

Table 2 shows that neither TPC nor TMYC were not found in the broth of the soups.

Table 3. Microbiological load of water, seasonings, chicken meat and vegetables, and the soups at each step of processing of chicken vegetable and chicken sweet corn soups prior to irradiation Samples Total Plate Total Mould and Count/TPC (cfu/g) Yeast Count/TMYC (cfu/g)

Tap water (not potable) 1.5 x 103 0.3 x 103

Cooked water (potable) 1.6 x 102 0

Raw chicken meat 2.1 x 103 0

Raw vegetables for chicken vegetable soup 1.0 x 103 3.0 x 103

Raw vegetables for sweet corn soup 1.9 x 103 1.2 x 103

Raw seasonings for chicken vegetable soup 5.1 x 104 3.6 x 104

Raw seasonings for sweet corn soup 8.5 x 103 1.0 x 103

Chicken vegetable soup after cooking 7.7 x 103 1.2 x 103

Chicken sweet corn soup after cooking 1.4 x 103 1.1 x 103

* Average of three replications

Table 4. Microbiological assessments of irradiated black and ox-tail soups packed in vacuum sealed laminate pouch of stored at pet12µm/ldpe as adh.2 µm/al-foil 7 µm/ldpe as adh./lldpe 50 µm, before and after storage at 4oc for 3 months Parameters Irradiation Type of soups and storage time (s) dose Black soup Ox-tail soup (kGy) 0 month 3 months 0 month 3 months TPC (cfu/g) 0 0 1.3 x 103 103 > 106 1 0 0 0 2.5 x 105 TMYC (cfu/g) 0 0 3.8 x 103 0 0 Coliform 0 Negative 9 negative negative 1 Negative 13 negative negative Staph. aureus 0 Negative negative negative negative 1 Negative negative negative negative 3 Negative 40** 20** negative

Table 5. Microbiological assessments of irradiated chicken vegetable and chicken sweet corn soups packed in vacuum sealed laminate pouch of stored at pet 12µm/ldpe as adh.2 µm/al-foil 7 µm/ldpe as adh./lldpe 50 µm, before and after storage at 4oc for 3 months. Parameters Irradiatio Type of soups and storage time (s) n dose Chicken vegetable soup Chicken sweet corn soup (kGy) 0 month 3 months 0 month 3 months TPC (cfu/g) 0 32.9 x 103 1.7 x 105 1.0 x 106 2.5 x 105 1 12.3 x 103 1.3 x 102 0.5 x 106 2.0 x 102 3 5.4 x 102 0 10.6 x 103 0 TMYC (cfu/g) 0 0 3.3 x 103 0 2.4 x 103 Coliform 0 Negative 1.1 x 102 negative negative 1 Negative 0.8 x 102 negative negative E.coli 0 Negative negative 1.1 x 102 negative 1 Negative negative 0.8 x 102 negative *Average of three replications

The results from Table 5 showed that gamma irradiation at doses of 5-7 kGy could reduce significantly the TPC and starting from 1 kGy the mold and yeast were not found the soups observed. Gamma irradiation at doses 3-7 kGy could eliminate Coliform and E.coli in the samples.

Gamma irradiation and storage condition did not affect the moisture content nor pH of the soups observed. The data showed that moisture content (m.c) and pH as follows: m.c.black soup was 69% and pH was 6.7; m.c. ox-tail soup was 83% and pH was 7.1; m.c. chicken vegetable soup was 86% and pH was 6.9; m.c. chicken sweet corn soup was 81% and pH was 6.8. Meanwhile, the average fat content among soups were in the range of 21-30% and carbohydrate were in the range of 0.65-1.09%.

Data on organoleptic score of the irradiated 4 different types of soups at two different doses showed that after three months of storage the soups were still acepted and favourable by panellists and the average value was 4.5. while the control was evaluated as 2.8 but it was not orally tasted during the test.

4. CONCLUSIONS

Gamma irradiation at pasteurization doses 5-7 kGy in combination with cryogenic condition, -79oC, and storage temperature at 4oC could maintain the safety and quality of, black soup, ox-tail soup, chicken vegetable soup and chicken sweet corn soup packed in an individual laminate pouch of Polyester/Aluminum foil/LLDPE under vacuum during 3 months of storage respectively. Obviously, the soups could be accepted by the panellists until 3 months of storage.

5. FUTURE STUDIES

IRRADIATION TO ENSURE THE SAFETY AND QUALITY OF HOME STYLE FROZEN FOODS : 2. Solid based materials

5.1. Programme of Work

5.1.1. Materials used are consisting of 3 different model systems :

a) Spring rolls: wheat flour dough, chicken meat, marine products, typical vegetables, seasonings and spices.

b) Croquette: mashed potato, ground beef, vegetables, seasonings and spices.

c) Rissole: wheat flour dough, eggs, milk, vegetables, seasonings and spices.

5.1.2. Source of materials

a. Solid state frozen prepared meals are commercially made by food industries are normally sold in the supermarket.

b. Home industry solid state frozen prepared meals as self-making products in order to develop Standard Operating Procedure (SOP) and indicate CCP along the process.

5.1.3. Methods and treatments

The methods and treatments will be conducted as almost similar to the previous study as described in the irradiated 4 different types of soups but some chemical characteristics and nutritional values of the new products might also be conducted as additional information.

5.2. Objective

1. To determine the microbial profiles of selected frozen prepared meals as solid state materials: spring rolls, croquette and rissole as well as their shelf-life under chilled condition.

2. To determine the effectiveness of irradiation to extend the shelf-life and eliminate pathogenic bacteria in the prepared meals.

3. To determine the sensory quality of the irradiated prepared meals.

ISRAEL

Hazard, Sensorial and Economic Analysis Critical Control Point (HACCP) Check List for Radiation Sterilized Ready-to-Eat Meals

Chief Scientific Investigator: Yaara F. Haruvy Soreq Nuclear Research Centre Operations Division, QA Department Yavne Israel

Research Contract No.: 11997

OBJECTIVES

The primary objective of this work is to introduce a modified HACCP analysis route, for irradiated prepared meals that addresses not only health hazards but also sensorial failures, economic risks. Above all, this method aims at pinpointing failure modes specific to the radiation pasteurization aspect of the production and the product. The suggested modified analysis should serve all researchers in the CRP in their attempts to transfer their process and product from the laboratory-research type into an industrial one. While in the laboratory we can discard “failure” samples or lots, or parts of a lot that are of inferior quality, an industrial process must be profitable and, hence, stable and reproducibly producing high quality products.

The basic aspects of the HACCP route are listed below.

HACCP* describes a system of control for assuring food safety and provides a more structured and critical approach to the control of identified hazards than that achievable by traditional inspection and quality control procedures. It has the potential to identify areas of concern where failure has not yet been experienced, making it particularly useful for new operations.

The analysis route is exemplified in the Table below, that covers all the steps involved in the food production, from farm to fork.

CONTROL FAILURE MODE / HAZARD PREVENTIVE CONTROL DETECTABILITY Haz. Sens. Econ Rad. POINT MEASURES . Specifi c Raw Materials: Vegetables Bacterial Bacterial contamination due to irrigation Irrigation inspection Microbiological smear with sewage contaminated water testing " " Contamination in the field by birds, Bird, rodent and insect control field Inspection; Microbiological rodents or insects program smear testing " " Post-harvest contamination due to injuries Discard of injured or broken vegetables Inspection for injured or broken vegetables " " Storage: Mold or bacterial growth due to Dry storage / humidity control and Microbiological smear damp storage conditions moisture control in storage warehouse testing " " Storage: Mold or bacterial growth due to Temperature control in storage Microbiological smear excessive temperature storage conditions warehouse testing " " Storage: Contamination by birds, rodents Bird, rodent and insect control storage Inspection; Microbiological or insects program smear testing " " Supplier: High microbial loading of Selection of suppliers of hygiene Microbiological smear unknown source practices testing

Bio-toxins Accumulation of bio-toxins due to Post detection: Discard lot Biotoxins smear testing sustained exposure to bacteria and/or molds Pesticides Contamination by pesticides Post detection: Discard lot Pesticide smear testing

Physical Faulty texture, rigidity, stem-cutting, or Harvesting quality protocols, Inspection damaged skin monitoring, and sorting " " Contamination by foreign matter: stones, Inspection, sieving and washing. Set Inspection, sieving and metal objects, shells threshold for lot discarding. washing

" " High concentration of hard fibers or Inspection, sorting. Set threshold for lot Inspection, mechanical grainy material. discarding. testing

" " Storage: Spoilage by exposure to extreme Temperature control in storage Microbiological smear 52

temperatures warehouse testing Temperature Excessively high temperature upon Inspection, sorting. Set threshold for lot Inspection, mechanical harvesting, resulting in damaged skin discarding. testing

" " Excessively low temp. upon intermediateInspection, sorting. Set threshold for lot Inspection, mechanical storage, resulting in damaged skin discarding. testing

CONTROL FAILURE MODE / HAZARD PREVENTIVE CONTROL DETECTABILITY Haz. Sens. Econ Rad. POINT MEASURES . Specifi c Raw Materials: Spices - all the above as well as: Active material Variation in concentration of active Tune recipe to maintain spice-level Analysis of spices concentration material - e.g. capsicum, ginger - within set limits bacterial flourishing Acidity Variation in concentration of acid: e.g. Tune recipe to maintain spice-level Analysis of spices concentration lemon, vinegar etc - bacterial flourishing within set limits Fermentation Variation in concentration of active Tune recipe to maintain spice-level Analysis of spices Products material and bacterial flourishing within set limits thereafter Salt mixtures Variation in concentration salt and Tune recipe to maintain spice-level Analysis of spices bacterial flourishing thereafter within set limits Raw Materials: Animal products Bacterial Bacterial contamination from broken Discard parts contaminated beyond Inspection and intestines permitted level Microbiological testing " " Bacterial contamination from cuts, Discard parts contaminated beyond Inspection and bruises and abcesses permitted level Microbiological testing " " Bacterial contamination from insect bites Discard parts contaminated beyond Inspection and and larvae therein permitted level Microbiological testing Physical Bones, teeth, scale, and their fragments Discard parts physically contaminated Inspection beyond permitted level Fat Content Fat degradation during irradiation and Discard parts containing fat beyond a Inspection production of off odors pre-determined level Raw Materials: Packaging Bacterial - pre- Contamination by birds, rodents or Bird, rodent and insect control program Packaging lots Inspection packaging insects " " Contamination by dirt from machinery Clean storage environment Packaging lots Inspection and workers Bacterial - post- Contamination by penetration of birds, Bird, rodent and insect control program Packaged lots inspection packaging rodents or insects " " Contamination (external) by dirt Clean storage environment Packaged lots inspection 54

Packaging injuries Contamination by penetration of insects Discard damaged packaging materials Inspection of packaging and cuts and bacteria prior and after packaging

CONTROL FAILURE MODE / HAZARD PREVENTIVE CONTROL DETECTABILITY Haz. Sens. Econ Rad. POINT MEASURES . Specifi c Processing: Cutting (or pressing) Bacterial Inclusion of bacteria from contaminated Pre-cleaning and temperature control of Microbio. inspection of cutting equipment and/or boards cutting equipment; Discard contaminated cutting equipment and lots processed lots Bacterial Inclusion of bacteria from chopped Inspection of pre-processed lots for Microbio. / foreign- insects or small rodents or their litter insects or rodents or litter BAMBA particles inspection of CANDY STORY processed lots Bacterial Bacterial growth due to processing Temperature control of cutting Microbiological inspection temperature equipment; Discard contaminated lots of processed lots Foreign objects Inclusion of foreign objects from cutting Pre and post processing inspection of Pre/post processing equipment and/or boards cutting equipment; Discard suspicious inspection of cutting lots equipment Processing: Mixing Bacterial Same as all the above and: Bacterial Inclusion of bacteria from a specific Discard any contaminated ingredient Microbiological inspection ingredient of mixed lots Foreign objects Inclusion of foreign objects from cutting Pre and post processing inspection of Pre/post processing equipment and/or boards mixing equipment; Discard suspicious inspection of mixing lots equipment Processing: Cooking Bacterial Inclusion of bacteria from cookware Discard lots contaminated beyond Inspection and or transfer tools permitted level Microbiologic. testing of cookware and food Foreign objects Inclusion of foreign objects from Pre and post cooking inspection of Pre/post processing cookware or transfer tools cookware; Discard suspicious lots inspection of cookware Over-Cooking Spoiled food from over-cooking or Discard spoiled lots Monitor stirring, defective stirring; Radiation enhancement temperature, time. of off-taste

Texture spoilage Spoilage of food texture by tools used for Discard spoiled lots Monitor stirring tools and transfer or stirring process Processing: Frying Bacterial Inclusion of bacteria from fry-ware Discard lots contaminated beyond Inspection and or transfer tools permitted level Microbiologic. testing of fry-ware and food Foreign objects Inclusion of foreign objects from fry- Pre and post frying inspection of Pre/post processing ware or transfer tools fry-ware; Discard suspicious lots inspection of fry-ware Over-Frying Inclusion of spoiled food from over- Discard spoiled lots Monitor stirring, frying or defective stirring temperature, time. Texture spoilage Spoilage of food texture by tools used for Discard spoiled lots Monitor stirring tools and transfer or stirring process

CONTROL FAILURE MODE / HAZARD PREVENTIVE CONTROL DETECTABILITY Haz. Sens. Econ Rad. POINT MEASURES . Specifi c Processing: Frying (cont.) Rancidization Off taste due to use of spoiled oil Discard spoiled lots Frying Oil packaging QA; Radiation enhancement of off-taste Lab and/or sensory inspection Rancidization Off taste due to oil spoilage upon Discard spoiled lots Monitor temperature, time, overheating stirring Radiation enhancement of off-taste Oil inclusion Excessive inclusion of oil in the food Discard spoiled lots Monitor temperature upon upon under-heating introduction of food Radiation enhancement of off-taste Sticking Spoilage of food texture by sticking onto Discard spoiled lots Monitor stirring tools and fry-wear process Processing: Structuring and Stuffing Bacterial Inclusion of bacteria from processing Discard lots contaminated beyond Inspection and tools permitted level Microbiological testing of tools and food Foreign objects Inclusion of foreign objects from Pre and post processing inspection of Pre/post processing processing tools tools; Discard suspicious lots inspection of tools Internal texture Spoilage of inner food texture by Discard spoiled lots Monitor shear forces on spoilage processing tools food External texture Spoilage of envelope food texture by Discard spoiled lots Monitor food structuring spoilage processing tools operations Processing: Packing Bacterial Inclusion of bacteria from packaging Discard contaminated lots Pre/post packaging equipment inspection of food Microbiological level Foreign objects Inclusion of foreign objects from Discard spoiled lots Pre/post packaging packaging equipment inspection of tools and equipment External texture Spoilage of envelope food texture by Discard spoiled lots Pre/post packaging

spoilage packaging equipment inspection of food texture Processing: Irradiation Dose mal control Spoilage due to insufficient dose control Discard spoiled lots Double dose control on all product lots Temperature mal Spoilage due to insufficient temperature Discard spoiled lots Prompt temperature control control control on all product lots Mechanical mal Spoilage due to mechanical damage Discard spoiled lots Post irradiation inspection control

CONTROL FAILURE MODE / HAZARD PREVENTIVE CONTROL DETECTABILITY Haz. Sens. Econ Rad. POINT MEASURES . Specifi c Storage: Bacterial Microbial growth in moist products Storage of moist products @ 5 + 2 oC, Monitor temperature and Limited shelf-life. food spoilage Distribution: Bacterial Microbial growth in moist products Distribution chain of moist products Chill distribution chain 5 C @ 5 + 2 oC, Limited shelf-life. + - 2 C

Bacterial Distribution of out of date product Date label and stock rotation control Date label and stock rotation control

Consumer: Bacterial Storage abuse of product leading to Clear instructions to the consumer on Clear instructions to the microbial growth storage, shelf-life and product consumer on storage, shelf- preparation life and product preparation

All Steps: Bacterial - Contamination due to unsanitary Personnel hygiene enforcement. Discard Personnel hygiene Personnel handling practices contaminated lots, inspection. " " Cross-contamination from raw material, Enforcement of controlled movement of Controlled movement of germination and growing areas staff and equipment, Discard staff and equipment. contaminated lots. " " Metal fragments in product Use of metal detector Use of metal detector

To demonstrate the consequences of an inadequate HACCP in an increasingly global food market, where control points may be in different countries or even different continents, an example is given:

Retrospective Hazard Analysis: The "Bamba" Story 1. Children epidemic Salmonella was reported in London, 2 mortalities. 2. Investigation reveals all sick kids are Orthodox Jewish. 3. All kids eat Special Kosher food (BADATZ), snacks imported from Israel 4. Favorite snack: "Bamba" – a puffed corn-peanut snack 5. All imported snacks are recalled and tested. 6. "Bamba" found highly contaminated with Salmonella. 7. A special investigator [I. Klinger] inspects all "Bamba" ingredients and equipment. 8. Contamination source identified: Peanut butter imported from South Africa 9. Contamination source tracked, in SA: Rodents litter found on raw pre-pressed peanuts. 10. Contamination mechanism: Peanut storage is open to rodents who eat while leaving their litter.

A practical approach to implement the suggested combined HACCP is given below:

PRACTICAL STEPS OF COMBINED HACCP 1. Carry out a comprehensive HACCP analysis of the actual food to be prepared. 2. Assess the most risky (less controlled) points in the process. 3. Set the necessary QA system to minimize the need for control points. 4. Set the necessary control-measures, inspection-points and inspection-frequency. 5. Run a few experimental preparations, employing markers for bacteria, foreign bodies, texture damage etc., introduced in reasonable points. 6. Analyze the effectiveness of the HACCP system. 7. Improve the HACCP set: add missing control points and remove redundancy. 7. Run Validation preparations as before. 8. Write the protocol and manual of performing the HACCP. 9. Train the HACCP personnel, verify their knowledge and certify accordingly. 10. Start production.

CONCLUSIONS

HACCP protocol has to be carefully structured to specifically answer the needs of each product. Hence, the forthcoming stage of this work is of course to collaboratively produce specific combined specific HACCP protocols for radiation-pasteurized ready-to-eat meals of other groups in the CRP.

KOREA

Irradiation application on ready-to-cook Galbi, Korean traditional meat product for safety and extending shelf-life

Chief Scientific Investigator: Cheorun JO

Radiation Food Science & Biotechnology Korea Atomic Energy Research Institute Yuseong, Daejeon, Korea

Research Contract No. 11908

1. INTRODUCTION

Bulgogi and Galbi are the most popular traditional dishes in Korea with 1,500 years of history and it has become popular gradually worldwide. Ready-to-cook Bulgogi and Galbi are similar marinated meat products using thin slices of beef marinating in sauce composed of soy sauce, sesame oil, garlic, onion and other seasonings; and cooking over a hot charcoal grill. While a commercialization of these foods by small meat benders is increased fast, safety of the products during distribution and storage is not well understood because of possible high contamination of pathogens or spoilage bacteria from fresh vegetables, soy sauce, and raw beef.

2. MATERIALS AND METHODS

2.1. Sample preparation, strains, and culture condition

Freshly prepared marinated beef rib (deboned, uncooked Galbi) was purchased from a local manufacturer and a 10 g sample was packed in oxygen-impermeable nylon bags (2 mL 2 O2/m /24 h at 0°C, 0.09 mm thickness; Sunkyung Co. Ltd., Seoul, Korea). The samples were exposed to an irradiation dose of 30 kGy (point source, AECL, IR-79, NDS, Nordion, Ontario, Canada) to achieve the complete inactivation of the indigenous microflora.

2.2. Strains and Culture Condition

Four food-borne microorganisms of Staphylococcus aureus (KCTC 1916), Bacillus cereus (KCTC 1012), Salmonella Typhimurium (KCTC 1925), and Escherichia coli (KCTC 1682) were used in this research and obtained from a Korean collection for type culture (KCTC, Daejeon, Korea). All the strains were grown in a tryptic soy broth (Difco, Laboratories, Sparks, MD, USA) and incubated at 37°C. The bacterial cultures were prepared for 24 h in a sterilized broth medium from 1 colony from an agar slant, from which 0.1 mL was transferred to a new broth medium and grown for 18 h. The cultures were centrifuged (5,000 rpm for 10 min at 4°C) in a refrigerated centrifuge (Vs-5500, Vision Scientific, Co., Seoul, Korea). Cultures were washed twice with sterile peptone water. The pellet was finally suspended in sterile peptone water to have a cell density above the 107~108 CFU/g levels.

2.3. Inoculation of barbecued ribs with test organism

Sterile marinated beef rib was inoculated with a cell suspension of the four test organisms, respectively. The test culture suspension (200 µL) was uniformly and aseptically spread on

the marinated beef ribs. It was kept in a sterile workstation for 10 minutes to allow it to be absorbed. Samples were then packed in to sterile stomacher bags and sealed.

2.4. Irradiation

Packed marinated beef ribs inoculated with the test organisms were irradiated in a cobart-60 irradiator (point source AECL, IR-79, MDS Nordion International Co. Ltd., Ottawa, ON, Canada) at the Korea Atomic Energy Research Institute, Daejeon, Korea. The ice bag with a thickness of 1 cm was covered to avoid a temperature increase during irradiation. The source strength was approximately 100 kCi with a dose rate of 10 kGy h-1 at 12±0.5°C. Dosimetry was performed using 5 mm diameter alanine dosimeters (Bruker Instruments, Rheinstetten, Germany), and the free-radical signal was measured using a Bruker EMS 104 EPR Analyzer. The dosimeters were calibrated against an International standard set by the International Atomic Energy Agency (Vienna, Austria). The applied doses in this study were 0, 1, 2, 3, 4 and 5 kGy. After irradiation, the samples were transferred to an incubator (4°C and 20°C) and microbiological analysis was performed during storage at a commercial storage condition (4°C) and an abused temperature storage condition (20°C).

2.5. Microbiological Analysis

A sample (10 g) was aseptically homogenized for 2 min in a sterile stomacher bag containing 90 mL of sterile peptone water using a stomacher lab blender (Model 400, Tekmar Co, Cincinnati, OH. USA). Media for enumeration of the S. arueus, B. cereus, Salmonella Typhimurium, and E. coli were prepared by baird parker agar (Difco Laboratories, Detroit, MI, USA), cereus selective agar (Oxoid, Basingstoke, Hampshire, England), xylose lylsine deoxycholate agar (Difco Laboratories, Detroit, MI, USA), and eosin methylene blue agar (Difco Laboratories, Detroit, MI, USA), respectively. Plates were incubated at 37°C for 48 h, colony forming units (CFU) per gram were counted at a dilution of 30 to 300 CFU per plate. D10 values (the dose required to inactivate 90% of a population) for each of the organisms were determined by calculating the reciprocal of the slope. Experiments with each bacteria culture were conducted independently twice.

2.6. Statistical Analysis

3. RESULTS

Commercial marinated beef rib distributed in Korea has 5.68 log CFU/g of viable cells (Table 1). In addition, B. cereus and S. aureus was found at a 3.81 and 2.57 log CFU/g level, respectively. No viable cell of Salmonella Typhimurium and E. coli was observed. These results indicated that, even if the marinated beef rib is cooked before consumption, the undercooked meat might be potentially hazard. Radiation-sterilized marinated beef rib was found to be devoid of any viable vegetative microorganisms as determined by the analyses on different media.

D10-vlaues of Bacillus cereus, Staphyloccoccus aureus, Salmonella Typhimurium, Escherichia coli were calculated to be 0.66±0.01, 0.59±0.05, 0.64±0.02, and 0.54±0.01 kGy, respectively after gamma irradiation (Table 2).

Viable cells in non-irradiated marinated beef rib were initially 6 log CFU/g but the irradiation at 4 kGy showed no viable cell growth of S. aureus, B. cereus, Salmonella Typimurium, E. coli at day 0. Viable cells of all the four pathogens increased slightly during 9 days of storage during an abusive storage condition at 20°C (Tables 3-6). During 9 days of storage there were approximately a log reduction of viable cells by irradiation at 0, 1, 2, 3, 4, and 5 kGy, respectively. E. coli showed the most radiation-sensitive, as it decreased more rapidly with irradiation and during storage after irradiation than other pathogens (Table 4).

With a commercial storage condition (4°C), irradiation at 3 kGy had initially about a 1 log CFU/g of viable cells detected (Tables 7 and 8). However, at 2-weeks and 4-weeks, there was no viable cell counted. At refrigeration storage, 4 and 5 kGy of irradiation eliminated the viable cells of 4 kinds of pathogens with no growth during storage (Tables 7 and 8).

In conclusion, irradiation of raw marinated beef rib before cooking at the level of 4 kGy of irradiation may ensure the safety of the product when stored at a refrigerated temperature (4°C). However, when the storage temperature is abused (20°C), irradiation at 5 kGy may not be enough to control the remaining viable cells from multiplying in the raw marinated beef rib.

3.1. Effect of irradiation on microbiological, physicochemical, and sensory characteristics of prepared “Galbi” during storage at 4°C.

3.1.1. Sample preparation

Prepared seasoned beef rib, Galbi, was purchased from W-mart in Daejeon, Korea. The half of Galbi (approximately 50 g each) was packaged with oxygen impermeable 2 nylon/polyethylene bags (2 mL O2/m /24 h at 0°C, 0.09 mm thickness; Sunkyung Co. Ltd., Seoul, Korea) and packaged with vacuum packaging machine. The other half of the packs were flushed air into the bag without sealing to develop oxygen contact environment. The samples were exposed to an irradiation dose of 0, 2.5, 5.0, and 7.5 kGy (point source, AECL, IR-79, NDS, Nordion, Ontario, Canada) at the Korea Atomic Energy Research Institute, Daejeon, Korea.

3.1.2. Microbiological analysis

A sample (10 g) was aseptically homogenized for 2 min in a sterile stomacher bag containing 90 mL of sterile peptone water using a stomacher lab blender (Model 400, Tekmar Co, Cincinnati, OH. USA). Media for enumeration of the total aerobic bacteria, S. arueus, B. cereus, Salmonella Typhimurium, and E. coli were prepared by total plate count agar (Difco Laboratories, Detroit, MI, USA), baird parker agar (Difco), cereus selective agar (Oxoid, Basingstoke, Hampshire, England), xylose lylsine deoxycholate agar (Difco), and eosin methylene blue agar (Difco), respectively. Plates were incubated at 37°C for 48 h and colony forming units (CFU) per gram were counted at a dilution of 30 to 300 CFU per plate. Experiments with each bacteria culture were conducted independently twice.

3.1.3. Water activity (Aw) and pH

Water activity (aw) was determined with a thermoconstanter (Novasina RA/KA, Switzerland). The pH was determined using a pH meter (Orion 520A, Orion Research Inc., Boston, MA, USA) using sample (1 g) mixed with deionized distilled water (DDW, 9 mL).

3.1.4. DPPH radical scavenging effect

The sample (2 g) was transferred to a 20 mL-test tube and homogenized (DIAX 900, Heidolph Co., Ltd., Germany) for 15 s at speed setting 6 (approximately 23,000 rpm) with 8 mL of deionized distilled water (DDW). The mixture was filtered by a filter paper (No. 4., Whatman International Ltd., Springfield Mill, Maidstone, Kent, England) and chloroform (2 mL) was added into the filterate. The tube with filterate was vortexed vigorously to remove any fat in the sample and centrifuged (VS 5500, Vision Scientific Co., Ltd. Bucheon, Korea) at 2,400 rpm for 30 min. The upper layer of the mixture was diluted 5 times with DDW, and the free radical scavenging effect was estimated according to the method of Blois (1958) with some modification. A sample (1 ml) was added into the 0.2 mM DPPH radical (1 ml). The mixture was shaken and left to stand for 30 min at room temperature and measured at 517 nm with a spectrophotometer.

3.1.5. 2-thiobarbituric acid reactive substances (TBARS) value

Lipid oxidation was determined as a 2-thiobarbituric acid reactive substances (TBARS) value by using a spectrophotometer (UV 1600 PC, Shimadzu, Tokyo, Japan) as described by Ahn et al. (1999). The lipid oxidation development was reported as mg malondialdehyde/kg meat sample.

3.1.6. Sensory analysis

Preference test was performed to evaluate sensory properties of the raw and cooked Galbi. For raw meat status, odor and color were tested using 200 g of sample in a dish served individually to the panelists. For cooked meat status, Galbi was placed on a preheated pan at about 170°C for minute with 2 – 3 turnover. Average pan temperature during cooking was about 160°C. After cooling for 2 min at room temperature, the sample (approximately 20 g) were served to the panelists individually. The sensory parameters for the cooked Galbi were taste, tenderness, and overall acceptance. The sensory test was performed at each of the 3 different occasions. A 9-point hedonic scale was provided to the panelists and it was anchored as like extremely (9) to unlike extremely (1) for all sensory parameters.

3.1.7. Statistical analysis

Each data represents the mean of two different experiments with three measurements in each experiment. Mean values and standard deviation (SD) were calculated using a Statistical Analysis System (SAS Institute, 1996) and are reported for microbiological analysis. The mean values and standard error (SE) were reported for other measurements. Ducan’s multiple range test was adapted with significance defined at P<0.05.

3.2. Results

The number of total aerobic bacteria of the seasoned Galbi being distributed in the commercial market was 106 viable cells/g sample (Table 9). Irradiation of seasoned Galbi induced 1, 3, and 4 decimal reduction at irradiation doses of 2.5, 5.0, 7.5 kGy, respectively. The number of viable cells were 6.17±0.40, 4.95±0.03, 3.26±0.43, and 2.80±0.51 CFU/g in the samples with irradiated at 0, 2.5, 5, and 7.5 kGy, respectively.

Growth of pathogenic bacteria including B. cereus, Salmonella Typhimurium, E. coli, and S. aureus in irradiated seasoned Galbi during storage was shown in Table 10-13. Irradiation of 7.5 kGy almost eliminated the pathogenic bacteria except for B. cereus. The number of B.

cereus of non-irradiated control reached 4.5 log CFU/g but irradiation of 2.5, 5, and 7.5 kGy reduced the number to 3.71, 2.01, and 1.60 log CFU/g, respectively. In spite of irradiation treatment up to 7.5 kGy, the Bacillus cereus was not eliminated. In microbiological study, vacuum packaging showed slightly effect in controlling the total aerobic bacteria but not very significant.

The pH reduction of irradiated seasoned Galbi during storage was less than that of non- irradiated control (Fig. 1). It can be explained that microbiological control of irradiation may reduce the pH changes. The water activity tested was 0.99±0.02 with no significance in irradiation treatment.

DPPH radical scavenging effect was measured because the raw materials in the seasoning contain vegetable and sometimes fruits, which have beneficial polyphenolic compounds. The data of radical scavenging effect of samples were not consistent, however, during storage for seven days, the effect was decreased in irradiated sample (Fig. 2). Especially aerobic packaging the decreasing rate was faster than that of vacuum packaging, which may came from the oxidation-likely environment.

Development of lipid oxidation was measured using TBARS method. Irradiated samples had higher TBARS value than non-irradiated control at day 7 in vacuum packaging but 0 and 3 days of storage did not except for 7.5 kGy-irradiated sample at day 0 (Fig. 3). The TBARS value of seasoned Galbi packaged aerobically increased significantly by irradiation treatment and storage period. The results indicated that irradiation of oxygen-available environment is not desirable in terms of development of lipid oxidation.

Sensory analysis with raw seasoned Galbi indicated that irradiation decreased odor preference in the aerobically packaged sample at day 3 but increased color preference significantly (Table 14). Vacuum packaged sample had relatively higher sensory scores of the irradiated Galbi at raw state. After cooking the sensory scores of taste in the sample vacuum-packaged and irradiated at 7.5 kGy decreased (Table 15). Overall acceptance also decreased by irradiation at 5.0 and 7.5 kGy in vacuum state. However, other treatment combination did not show any difference.

In microbiological point of view, the seasoned Galbi commercially available in Korea need at least 7.5 kGy of irradiation to eliminate the pathogenic bacteria and secure the safety of the food, even though proper cooking method can control the pathogenic bacteria. The combination of aerobic packaging and irradiation may increase lipid oxidation dramatically during storage when irradiated at 7.5 kGy and may decrease the beneficial radical scavenging effect. These adverse changes may minimize by packaging as vacuum state. Although the vacuum packaging was provided, however, there is one more problem that should be solved in sensory, especially in taste of the cooked meat product. This would be one important point before commercializing the seasoned Galbi and also other prepared meals.

3.3. Work Plan for the Next Stage

In the last 2 years, irradiation application of Bulgogi and Galbi, two most popular prepared meat products in Korea, had been studied. Irradiating these products resulted the safe products without compromising their characteristic sensory quality. Commercial use of this technology collaborating with a local catering service company may be postponed since the consumer acceptance of our country does not reach the level that a private company takes an action. The approval of governmental authority is also prerequisite. However, the data obtained from the

IAEA CRP project are very helpful to apply for the approval of the irradiated ready-to-cook food products for our government in near future.

Now, Korea has a popular prepared meal, Kimbab. Kimbab is a rolled by a dried laver with cooked rice and vegetables, seasoned meat products, and seafood products in it. The Kimbab is now a most rapidly growing item in food industry, especially by franchised special small benders. However, the Kimbab has only 12 hrs or less of shelf-life and many foodborne diseases were reported from Kimbab. One of the important reasons is that the Kimbab cannot refrigerate because of its sensory change, which majorly come from the retrogradation of starch of the cooked rice. Therefore, it is very important to achieve the public safety from this special prepared food. Also, the extension of shelf-life to minimize the loss of food materials will be helpful for the industry. Irradiation of Kimbab is one of the most beneficial techniques to reduce the food-borne disease outbreak and extension of its shelf-life. The outcome from this proposed project can be used for application of the prepared meals for irradiation to governmental authorities as a basic research data.

3.4. The studies using Kimbab for following year are:

• Determination of irradiation sensitivity of pathogens in food materials for preparation of Kimbab. Pathogens to be used are Vibrio, Salmonella, Staphylcoccus, Listeria, and E. coli.

• Determination of the growth of total aerobic bacteria, molds, and yeast in food materials for preparation of Kimbab during storage at 10, 20, and 30°C for shelf-life extension and safe distribution.

• The food materials used for this experiment will be dried laver, boiled fish paste, crab- flavored surimi products as seafoods, seasoned beef, ham, fried egg as animal- originated products, and seasoned and fried spinach, washed cucumber, seasoned burdock as vegetables. These are the most common food materials for Kimbab preparation.

SOUTH AFRICA

Use of Irradiation to Improve the Safety and Quality of Ethnic South African Foods

Chief Scientific Investigator: Amanda Minnaar

Co-investigators: Karlien Nortjé, Elna M Buys and Department of Food Science, Faculty of Natural and Agricultural Sciences, University of Pretoria, South Africa

Research Agreement No. 11909

1. INTRODUCTION

Beef biltong is a popular, traditional South African salted dried raw meat product (Van den Heever, 1970; Van der Riet, 1976). Nowadays many consumers prefer ‘moist’ beef biltong with relatively high moisture contents (greater than 40%) and water activity (aw) levels ranging between 0.85 and 0.93 (Van den Heever, 1970; Osterhoff & Leistner, 1984). Since Staphylococcus aureus can grow under these conditions, moist biltong may potentially present a health risk (Van den Heever, 1970; Osterhoff & Leistner, 1984). Furthermore, it has been found that raw meat from South African abattoirs typically used for biltong production may be significantly contaminated with S. aureus (103 to 105 cfu/g) (Nel, Lues, Buys & Venter, 2004). The fact that biltong is predominantly sold unpackaged together with raw meat in butcheries is compounding the problem. Spoilage of biltong by yeast and mould growth (especially when moist) is also problematic because fungal contamination of biltong is often rather high (105 to 106 cfu/g) (Van der Riet, 1976).

The use of irradiation, alone or in combination with an edible coating may potentially solve these problems. Irradiation is very effective in inactivating bacterial pathogens as well as some spoilage yeasts and moulds. The rational for using an edible coating is that it is believed that it may potentially inhibit yeast and mould growth by acting as an oxygen barrier. It has also been shown that the combined use of a casein-whey protein based coatings and irradiation can synergistically inhibit microbial growth (Ouattara, Sabato & Lacroix, 2001; 2002).

2. OBJECTIVES

1. To determine the maximum irradiation dose that can be used to irradiate moist beef biltong without adversely affecting the sensory acceptability (Phase 1).

2. To determine the effect of irradiation, alone and in combination with a casein-whey protein based edible coating on inoculated S. aureus and the indigenous microbial flora of whole moist beef biltong (Phase 2).

3. EXPERIMENTAL DESIGN

Phase 1 Vacuum packaged moist beef biltong (47% moisture; 3.7% NaCl; 1.5% crude fat; aw 0.92)

Irradiate (0, 2, 4, 6 and 8 kGy) at ambient temperature (20°C – 25°C)

Analyses

Lipid oxidation products

(Thiobarbituric acid reactive substances – Ouattara, Smoragiewicz, Saucier & Lacroix, 2002) Sensory multiple difference testing (n=96)

(Same-different/R-index method – O’Mahony, 1986)

Sensory acceptability/hedonic testing (n=50)

(To be conducted if significant differences are found between control and irradiated samples)

Figure 2 Experimental design for determination of the effect of irradiation on the sensory perception of moist beef biltong Phase 2 Analysis/tests Aerobic plate count Whole moist beef biltong (n=3) (Petrifilm™ - AOAC Method 990.12) No inoculation (control) S. aureus inoculation Yeast & mould count (≈ 106 cfu/g) (Petrifilm™) S. aureus count

No coating Coating No coating Coating % Moisture content (control) (control) (AOAC Method 24.002) % NaCl content Irradiate (0 and 4 kGy) vacuum packed (AOAC Method 24.010) (ambient temperature: 20°C –25°C) aw (Novasina Thermoconstanter) Store unpackaged in incubator for 7 days (25°C) pH (Van den Heever, 1970) Figure 2 Experimental design 1,2 for determination of the effect of irradiation and a casein- whey protein edible coating on the microbiological safety and quality of moist beef biltong

1 Experiment repeated three times (samples tested in triplicate during individual experiments) 2 Results statistically analysed by ANOVA and Fisher’s LSD-test

4. RESULTS AND DISCUSSION

4.1. Phase 1: Effect of irradiation on the sensory perception of moist beef biltong

A dose dependant increase in TBARS values was observed. However, only biltong irradiated at 8 kGy had significantly higher levels of oxidation products than untreated biltong, probably due to higher levels of free radicals produced. This was not totally unexpected given the fact that irradiation processing was done under vacuum conditions and the fat content of the moist beef biltong was relatively low (i.e. 1.53 %).

In contrast, it was found that all irradiated moist beef biltong samples (2, 4, 6 and 8 kGy) could be distinguished from untreated biltong when sensory difference ratings of an analytical panel were analysed using the R-index method and the Wilcoxon-Mann-Whitney rank sums test. The degrees of differences between the irradiated samples were perceived to be rather small, since only ratings of 2 and 8 kGy samples differed significantly (p<0.05).

A consumer acceptability (hedonic) test performed to determine the relation of perceived differences between untreated and irradiated moist beef biltong to the acceptability of the different samples revealed that none of the irradiated biltong samples (2, 4, 6 and 8 kGy) were significantly less acceptable than the control (0 kGy) (p>0.05). However, biltong irradiated at the lower doses (2 and 4 kGy) was liked significantly more than untreated biltong or biltong irradiated at higher doses (i.e. 6 and 8 kGy). The preference for low dose irradiated biltong above untreated biltong was unexpected. It may be because the slight irradiation flavour that developed in these samples contributed to a fuller/more meaty flavour, since moist beef biltong is usually more bland in flavour than drier biltong types due to the higher residual moisture content.

It was therefore concluded that for Phase 2 irradiation at 4 kGy would be the most suitable irradiation treatment, since this was the highest irradiation dose used for which the sensory quality of the biltong were deemed most acceptable and preferable.

4.2. Phase 2: Effect of irradiation and a casein-whey protein based edible coating on the microbiological safety and quality of moist beef biltong

Irradiation at 4 kGy reduced S. aureus counts by approximately 6 log cycles, irrespective of application of the edible coating. Since viable S. aureus counts found on non-inoculated biltong samples never exceeded 103 cfu/g it can be concluded that irradiation at this dose would under normal conditions be very effective to render moist beef biltong safe from this pathogen. Even in the event of unexpectedly high contamination with S. aureus, the counts are unlikely to exceed 106 cfu/g directly after production when irradiation would typically be employed, and a dose of 4 kGy will thus probably also be effective for elimination in such a scenario. In contrast, the edible coating had no effect on the level of S. aureus when compared to uncoated samples (p>0.05).

Similar results were found for the total aerobic plate counts. This reduction is highly significant in terms of potential spoilage bacteria like lactic acid bacteria, which at high counts may cause spoilage of vacuum packaged moist biltong. In addition, the edible coating did not affect (p>0.05) the total aerobic counts significantly.

As for bacteria, the edible coating did not affect (p>0.05) yeast and mould counts. No inhibitory effect on microbial growth was therefore observed for the casein-whey protein coating, and it can thus be concluded that it did not have any significant oxygen barrier

properties. A possible reason for this is that the biltong used during the experiments had higher than expected moisture levels (ca. 54% moisture and aw 0.98) under which conditions barrier properties of protein coatings are diminished. Since the effect of the edible coating was determined after 24 h (48 h in conjunction with irradiation) it may be that the time period was not long enough for a significant effect to be observed. For this reason, it was planned to test samples after 7 days of storage. Due to gross fungal spoilage in less than 48 h under the specific storage conditions used, these samples were however deemed unfit for human consumption and consequently not tested. No visual difference between coated and non- coated samples could however be seen, but it is not known whether it would have had any effect if lower numbers of microbes survived irradiation and/or if less favourable conditions were employed.

As anticipated, irradiation caused a significant reduction (p<0.05) in yeast and mould counts. However, because fungi are more resistant to irradiation than bacteria (Monk, Beuchat & Doyle, 1995), a relatively smaller reduction in counts (1 to 2 log cycles) compared to that of bacteria (i.e. 6 log cycles) was observed. When good manufacturing practices are used during manufacture of biltong, levels of yeasts and moulds are not expected to exceed 100 cfu/g to 1000 cfu/g (as found for the biltong samples during the third repetition of the experiment - Fig. 10). Therefore, reductions in fungal counts by 1 log to 2 log cycles due to irradiation at4 kGy could extend the shelf-life of moist beef biltong significantly under these circumstances for the different biltong batches tested during the three repeats of the experiment

However, in the biltong samples tested during the first two repetitions, yeast and mould counts up to 104 cfu/g to 105 cfu/g were found. Irradiation at 4 kGy would clearly not be adequate in this case to prevent microbial spoilage due to surviving yeasts and moulds. (i.e. 103 cfu/g to 104 cfu/g). This will especially be true if storage conditions are highly favourable for fungal growth (i.e. 75% RH, 25°C, aerobic atmosphere and no preservatives), as was the case when the biltong samples were stored for 7 days. Irradiated samples spoiled visually within 48 h, only 24 h longer than it took for non-irradiated samples to spoil. These highly favourable conditions are however unlikely to occur in practice, and therefore some extension in shelf-life will probably be obtained if moist biltong with high fungal counts is irradiated at 4 kGy.

5. CONCLUSIONS & RECOMMENDATIONS

Considering the findings, irradiation at 4 kGy will be adequate to ensure safety of moist beef biltong in terms of S. aureus. This dose will however not be effective to control fungal growth in moist beef biltong if initial contamination levels are high (≥104 cfu/g) and conditions are favourable for fungal growth. It is therefore clear that strict hygienic practices and microbiological standards must be adhered to during and after the manufacture of moist biltong, since irradiation at 4 kGy will not be effective to eliminate high fungal loads.

Since it was found that irradiation at doses up to 8 kGy did not affect the sensory quality of moist beef biltong detrimentally, irradiation at higher doses may potentially be used to control yeast and mould growth on moist biltong. Otherwise, the use of a combination of hurdles (eg. vacuum packaging and use of protein coatings with incorporated preservatives) together with irradiation at 4 kGy may be employed to attain extended microbiological stability.

6. FUTURE WORK – RTE BOVINE TRIPE

1. To determine the microbial ecology of fresh tripe (concentrating specifically on enteric pathogens such as E. coli and sporeformers such as B. cereus or C. perfringens).

2. To determine the effect of selected doses of irradiation on the sensory acceptability of ready-to-eat bovine tripe.

3. To determine the effect of irradiation combined with suitable pre-treatments (e.g. acid treatment and cooking) on inoculated spores and indigenous microflora of ready-to-eat tripe stored for at least 2 weeks at normal (5 °C) and abuse (15 °C) temperatures.

REFERENCES

[1] ASSOCIATION OF OFFICIAL ANALYTICAL CHEMISTS,. Meat and meat products. In: Horwitz, W. (Ed.) Official Methods of Analysis of the Association of Official Analytical Chemists. 13 th Edition. Washington: Association of Official Analytical Chemists. pp. 376-377 (1980). [2] BI, J & O’MAHONY, M., Table for testing the significance of the R-index. Journal of Sensory Studies 10, 341-347 (1995). [3] MONK, J.D., CLAVERO, R.S., BEUCHAT, L.R., DOYLE, M.P. & BRACKETT, R.E., Irradiation inactivation of Listeria monocytogenes and Staphylococcus aureus in low- and high-fat, frozen and refrigerated ground beef. Journal of Food Protection 57, 969-974 (1994). [4] NEL, S., LUES, J.F.R., BUYS, E.M. & VENTER, P.,. Bacterial populations associated with meat from the deboning room of a high throughput red meat abattoir. Meat Science 66, 667-674 (2004). [5] O’MAHONY, M., Sensory evaluation of foods: Statistical Methods and Procedures. New York: Marcel Dekker, Inc. pp. 153-182, 389-397 (1986). [6] OSTERHOFF, D.R. & LEISTNER, L., South African beef biltong – another close look. Journal of the South African Veterinary Association 55 (4), 201-202 (1984). [7] OUATTARA, B., GIROUX, M., SMORAGIEWICZ, W., SAUCIER, L. & LACROIX, M., Combined effect of gamma irradiation, ascorbic acid and edible coating on the improvement of microbial and biochemical characteristics of ground beef. Journal of Food Protection 65, 981-987 (2002). [8] OUATTARA, B., SABATO, S.F. & LACROIX, M., Combined effect of antimicrobial edible coating and gamma irradiation on shelf-life extension of precooked shrimp (Penaeus spp.). International Journal of Food Microbiology 68, 1-9 (2001). [9] OUATTARA, B., SABATO, S.F. & LACROIX, M., Use of gamma-irradiation technology in combination with edible coating to produce shelf-stable foods. Radiation Physics and Chemistry 63, 305-310 (2002). [10] VAN DEN HEEVER, L.W., Sekere gesondheidsoorwegings van biltong. M.Med.Vet (Hyg) thesis. University of Pretoria, South Africa, 138p (1970). [11] VAN DER RIET, W.B., The mycoflora of biltong and microbiological aspects of the mouldy biltong problem. M.Sc thesis. University of Stellenbosch, 144p. (1976).

SYRIA

Effect of Gamma Irradiation on the Microbial Load, Chemical and Sensory Characteristics of Locally Prepared Meals

Chief Scientific Investigator: Al-Bachir, M. Radiation Technology Dept. Syrian Atomic Energy Commission Damascus, Syria

Research Contract No.: 11910

Abstract

Locally prepared meal Kubba and Borak were treated with 0, 2, 4 and 6 kGy doses of gamma irradiation. Treated and untreated meals were kept in a refrigerator (1-4oC). Microbiological and chemical analyses were performed on each treatment sample immediately after processing, and weekly throughout storage period, which lasted for 3 and 6 weeks for Kubba and Borak respectively. Sensory evaluation and proximate analysis was down within one week of irradiation. The results indicated that 4 and 6 kGy doses of gamma irradiation decreased the total counts of mesophilic aerobic bacteria and increased the shelf-life of Kubba and Borak. Irradiation decreased the major constituents of Kubba (moisture, protein and fats) and did not had a significant effect of Borak. The three chemical parameters, total acidity, lipid peroxide and volatile basic nitrogen, which were chosen as the indices of freshness, were all well within the acceptable limit for up to one week for Kubba treated with 0 and 2 kGy, and for up to 3 weeks for samples treated with 4 and 6 kGy and for up to 1, 3 and 6 weeks at 1 - 4 0C for Borak treated with 0 or 2, 4 and 6 kGy respectively . Sensory evaluation showed no significant differences between irradiated and non-irradiated samples.

1. INTRODUCTION

The Syrian consumers have recently started using prepared meals, which are prepared and marketed by local supermarkets. The Syrian food industry is traditionally dominated by Kubba. Indeed a large amount of Kubba is consumed not only in Syria, but also in neighbor countries. The objectives of this study were to investigate the use of gamma irradiation in order to improve the microbiological quality of Kubba and Borak, as precooked prepared meals, by extending their shelf-life at refrigeration temperature (1-4 oC), while preserving the nutritional and organoleptic qualities.

2. MATERIALS AND METHODS

2.1. Preparation and formulation of Kubba and Borak

The Kubba and Borak were prepared by a local caterer. Kubba have two parts, in which the outer layer consists of ground pre-boiled wheat (borgel) mixed with minced beef and spices. Outer layer was stuffed with precooked lamb, onion, fat, pistachio and spices. Borak have two parts, the outer part is kind of dough. The dough was stuffed with precooked lamb, onion, fat, pistachio and spices. Eight pieces of precooked Kubba or Borak were placed on polystyrene trays covered with lids made of polyethylene film.

Samples from packed precooked Kubba and Borak were exposed to gamma radiation at doses of 0, 2, 4 and 6 kGy. Irradiated and non-irradiated Kubba or Borak were stored at (1 – 4°C)

temperature. Microbiological, chemical, and sensory analyses were performed on each treatment sample immediately after processing, and weekly throughout storage period.

2.2. Microbiological, chemical and sensory evaluation and statistical analysis

Microbial load were evaluated according to the standard methods (AOAC, 1986). A cut- of value of 107 CFU/ g for MACs (Ayres, 1960), were used for the samples that were not acceptable. Moisture, ash, fat and protein were determined using standard methods (AOAC, 1990). The total acidity was obtained by a direct titration and calculated as ml of (0.1 N) NaOH = 0.0090 g lactic acid (Egan, et al., 1987). Total volatile basic nitrogen (VBN) was determined in term of mg VBN/kg Kubba or Borak (ppm) (Pearson, 1976). Lipid peroxidation in terms of g iodine / 100g fat of Kubba or Borak was determined by the modified method of Buege and Aust, (1978).

Taste, odor, color and texture of the irradiated and non-irradiated Kubba or Borak were evaluated within one week of irradiation, by 20 people using 5- point scale (1= very bad, 2 = bad, 3 = excepted, 4 = good, 5 = very good), according to Lavrova and Krilova (1975).

The data were subjected to the analysis of variance test (ANOVA).

3. RESULTS AND DISCUSSION

3.1. Kubba and Borak characteristics

Mean Kubba characteristics were: fat content (12.16 ± 0.10 %), protein content (9.88 ± 0.75 %), ash content (1.93±0.06 %), moisture content 52.73 ±0.43 %) and Carbohydrates (25.59 ± 2.47). Mean Borak characteristics were: fat content (5.25 ± 0.60 %), protein content (8.06 ± 0.53 %), ash content (2.88±0.27 %) and moisture content (39.03 ±2.70 %). The aw value for Borak was 0. 95 at 24°C and the pH value was (6.03 ± 0.04).

3.2. Microbiological quality

Data in Table 1 indicate that 2, 4 and 6 kGy doses of gamma irradiation significantly (p> 0.05) decreased the aerobic microorganism counts of Kubba and Borak compared with control. A reduction in the CFUs values as a result of irradiation at time 0 found in samples in which the control was about 103 CUF/ g. The reduction in Kubba was a log cycle of more than 1 or 2 for 4 and 6 kGy, respectively. The reduction of total microbial load of Borak was a log cycle about 2, 4 or 6 for 2, 4 and 6 kGy, respectively. However, control Kuba and Borak samples have reached the generally accepted spoilage number of microorganism counts 107/g (Ayres, 1960) after one week of storage. Whereas, treated Kubba with 4 or 6 kGy had not reached the same number after 3 weeks. Overall, there were significant differences in results from Borak treated with 2, 4 and 6 kGy, although 6 kGy was better than those given 2 or 4 kGy in controlling microbial numbers. Using a dose of 6 kGy resulted in counts of total microbial load that were close to those of sterility. Therefore, it concluded 6 kGy should be used in future studies. Borak Samples irradiated at 2, 4 or 6 kGy remained acceptable, in order of microbial numbers, throughout the 2, 5 or 6 weeks of storage at (1 – 4°C) temperature respectively.

3.3. Chemical quality of irradiated Kubba

Total acidity: Table 2 shows that, immediately after treatment, all used doses (2, 4 and 6 kGy) had no effect on total acidity of Kubba and Borak. After one week of storage, 4 and 6

kGy doses of gamma irradiation significantly (p <0.05) decreased the total acidity. Throughout storage periods, the total acidity of both irradiated and non-irradiated samples increased.

Volatile basic nitrogen VBN: Immediately after treatment, the value of VBN of irradiated Kubba and Borak were significantly (p<0.05) higher than those of the control. After one week of storage, there were no significant (p<0.05) differences between Borak treatments concerning the VBN. Whereas, after one week of storage, the values of VBN of irradiated Kubba were significantly lower than those of control.

Lipid oxidation: Effects of gamma irradiation on lipid oxidation of Kubba and Borak were compared (Table 4). Immediately after treatment, lipid oxidation values for irradiated Kubba and Borak were not different than those of non-irradiated controls. After one week of storage, 4 and 6 kGy of gamma irradiation significantly (p<0.05) increased lipid oxidation. During storage, lipid oxidation values of irradiated Kubba samples tended to increase. Whereas, lipid oxidation values of irradiated Borak samples tended to decrease.

3.4. Sensory quality of irradiated Kubba and Borak

Sensory data presented in Table 6 indicates that taste, odor, color and texture characteristics initially of Kubba and Borak were unaffected by gamma irradiation treatment. Sensory change was not observed in the irradiated Kubba and Borak, and this was judged to be acceptable by a trained sensory panel. A correlation between sensory evaluation and chemical parameters was observed in relation to irradiated Kubba and Borak.

4. CONCLUSIONS

From the results of this research it can concluded that irradiation at doses of 4 and 6 kGy combined with refrigeration (1- 4 0C) does not affect the chemical quality (total acidity, lipid peroxide and volatile basic nitrogen), alter the organoleptic quality of the Kubba and Borak, and can extend the shelf-life of locally prepared meals (Kubba), from one week for the control to more than three and six weeks weeks Kubba and Borak respectively.

REFERENCES

[1] Association of Official Analytical Chemists (AOAC).1986. In Official Methods of Analysis (14th ed.), Washington D. C. [2] Association of Official Analytical Chemists (AOAC). 1990. In Official Methods of Analysis (15th ed.), Washington, D. C. [3] Ayres, J. C. 1960. The relationship of organisms of the genus pseudomonas to the spoilage of meat, poultry and eggs., J. Appl. Bacteriol. 23, 471. [4] Buege, J. A. and Aust, S. D. 1978. Microsomal lipid peroxidation. In Methods in Enzymology, 52, 302. AP, NewYork. [5] Egan, H., Kirk, R. S. and Sawyer, R. 1987. Pearson’s Chemical Analysis of Foods, 8th ed. Longman scientific & Technical, pp. 185. [6] Lavrova, L. P. and Krilova, V. X. 1975. Luncheon meat technology. Food’s industry, pp.325-326, Moscow, (in Russian).

Table 1. Effect of gamma irradiation on microbial load of Kubba & Borak stored at 1-4°C (CFU/g) Treatment 0kGy 2kGy 4kGy 6kGy LSD5% Kubba 0 2.96±0.04 2.20±0.181.71±0.30 0.93±0.81 0.83 1 R* R 1.52±0.45 0.87±8.81 1.49 3 R R 3.89±0.01 2.15±0.21 0.65

Bourak 0 6.17±0.10 4.08±0.082.05±1.38 0 1.07 1 R 6.72±0.296.18±0.42 0 0.39 3 R R 5.88±0.27 1.93±1.42 0.35 6 R R R 6.6±0.40 *R

Table 2. Effect of gamma irradiation and on Total acidity (%Lactic acid) of Kubba & Borak Treatment 0kGy 2kGy 4kGy 6kGy LSD5% Kubba 0 0.01±0.18 0.01±0.16 0.18±0.00 0.17±0.02 0.03 1 0.317±0.030 0.244±0.0240.202±0.031 0.224±0.009 0.0473 3 R* R 0.236±0.0520.211±0.005 0.084 Bourak 0 0.28±0.056 0.34±0.06 0.25±0.006 0.26±0.05 0.091 1 57.88±17.09 46.22±10.03 25.64±2.60 26.53±0.81 18.83 3 R R 45.04±4.24 41.86±9.52 16.7 6 R R R 27.09±8.87 *R

Table 3. Effect of gamma irradiation on volatile basic nitrogen (VBN) of Kubba & Borak (p.p.m.) Treatment 0kGy 2kGy 4kGy 6kGy LSD5% Kubba 0 76.0±7.0 106.0±5.0 138.0±7.0 75.7±2.5 10.7 1 119.0±1.0 62.3±1.5 77.0±8.0 72.3±10.5 12.6 3 R* R 123.0±11.0 105.0±5.0 19.4 Bourak 0 251.47±56.82 209.14±43.14137.52±3.44 106.77±9.16 67.79 1 126.84±43.47 142.23±16.06173.15±7.06 164.63±16.43 46.76 3 R R 234.46±13.47238.52±11.21 28.03 6 R R R 150.66±8.09 *R

Table 4. Effect of gamma irradiation on lipid peroxide (m. mol O2/ Kg) of Kubba & Borak Treatment 0kGy 2kGy 4kGy 6kGy LSD5% Kubba 0 0.035±0.001 0.038±0.0080.045±0.02 0.038±0.005 0.023 1 0.042±0.006 0.043±0.0070.055±0.005 0.060±0.0002 0.010 3 R* R 0.065±0.0110.078±0.023 0.041 Bourak 0 0.15±0.02 0.16±0.03 0.13±0.03 0.16±0.02 0.05 1 0.07±0.01 0.08±0.06 0.13±0.08 0.11±0.04 0.1 3 R R 0.17±0.07 0.18±0.05 0.07 6 R R R 0.0076±0.00365 *R

Table 5. Effect of gamma irradiation on the taste, texture, color and flavor of Kubba & Borak Treatment 0kGy 2kGy 4kGy 6kGy LSD5% Kubba Taste 4.00±0.95 4.00±0.85 4.08±0.90 4.08±0.79 0.72 flavor 4.26±0.97 4.26±0.75 4.08±0.79 3.83±1.19 0.78 color 4.27±1.19 4.46±0.82 4.64±0.51 4.27±0.91 0.77 Texture 3.83±1.27 3.83±1.03 4.33±0.99 4.25±0.75 0.84 Bourak Taste 3.82±1.02 3.35±1.16 3.58±1.07 3.85±0.9 0.57 flavor 3.58±1.36 3.50±1.48 3.77±1.28 3.58±1.39 0.76 color 3.77±1.11 3.27±1.15 3.27±1.43 3.73±1.0 0.65 Texture 3.50±1.27 3.23±1.24 3.35±1.20 3.46±1.42 0.73 * Data represent a 5 point scale ranging from 1 (very bad) to 5 (very good).

THAILAND

Use Of Irradiation To Improve The Safety And Quality of Thai Prepared Meal

Chief Scientific Investigator: A. Noomhorm Asian Institute of Technlogy Food Engineering and Bioprocess Technology Klongluang, Pathumthani THAILAND

Co-investigators: T. Koomsanit, S. Biramontri, P. Sirisoontaralak, W. Srisawas, P. Vongsawasdi

Research Contract No.: 11911

1. INTRODUCTION

Due to the change of life style of people in developed and developing countries, there is an increased economic growth of prepared meals. In Thailand, there is a dynamic growth of market for chilled prepared meals because of the growth of food services in supermarkets and convenient stores. However, the shelf-life for ready-to-eat chilled food including chilled prepared meals is quite short and sometimes insufficient to meet market requirements (FSIS, 1999) and it is implicated in a number of serious foodborne disease outbreaks (Bryan, 1990). Irradiation could provide a potential to improve the microbiological safety and extend the shelf-life of chilled prepared meals. Therefore, the study about the possibility to use irradiation for extending shelf-life and ensure microbiological safety of chilled prepared meals should be conducted. Also, information relevant to the application of food safety control system like Hazard Analysis and Critical Control Point (HACCP) should be gathered to ensure more safety of the irradiated prepared meals. Many traditional Thai dishes are popular in Thai restaurant over the world. Thai dishes are normally composed of herb and spicy with different types of meat. All dishes are eaten along with rice. Aromatic rice is the most preference for Thai consumers and foreigners. Thai spicy basil rice dish with selected meat such as chicken is selected as a prepared meal in this study due to its popularity and feasibility of commercial production.

2. METHODOLOGY

The optimum of irradiation dose for Thai spicy chicken basil rice was determined. Firstly effect of different irradiation doses (1, 2, 3 and 4 kGy) for cooked rice and spicy chicken basil was evaluated based on microbial, physical and sensory quality. Secondly shelf-life of irradiated Thai spicy chicken basil under chill condition (2±2oC) was evaluated. In this test, cooked rice was prepared with three different cooking methods and spicy chicken basil was packed with three different methods and irradiated at 1 and 2 kGy respectively. To modify for practical irradiation process, the same dose of 2 kGy was applied for both cooked rice and spicy chicken and storage study was conducted for four weeks under chilled contamination. Thirdly safety of chilled prepared Thai spicy chicken basil rice from food pathogens was confirmed in contamination study. L.monocytogenes and Escherichia coli were separately inoculated into sterilized cooked rice samples and L. monocytogenes and Samonella tiphimurium into sterilized spicy chicken samples under aseptic condition at initial pathogen concentration around 106 cell per gram of sample. The effect of selected irradiation dose of 2 kGy on decontamination of the three microorganisms was evaluated and also the growth of

survivals during storing under chilled condition was determined. Finally, HACCP plan for irradiated chilled prepared meal was proposed.

3. SUMMARY OF THE RESULTS

Irradiation (1 to 4 kGy) causes some changes in qualities of cooked rice and spicy chicken basil. It could significantly reduce microbial population. However the applied dose was limited by acceptance of the consumers. From the results, the dose of 1 and 2 kGy was selected as optimum dose for cooked rice and spicy chicken basil.

Shelf-life of chilled irradiated cooked rice (1 kGy) and spicy chicken basil (2 kGy) was evaluated based on total viable count and sensory quality. For irradiated cooked rice, it could be kept longer than 1 month with very less total viable count. With separate packing of spicy chicken from basil leaf, better microbial quality was obtained. It could be kept for more than 4 weeks compared to control samples, which had shelf-life only 3 weeks. With proper cooking method, irradiated cooked rice stored > 14 days was still accepted by panelists. However, after irradiation the acceptability of spicy chicken with basil leaf decreased during storage mainly due to the development of off-flavor. When basil leaf was removed and packed separately, panelists satisfied irradiated spicy chicken stored >4 weeks and gave higher scores compared to control sample, which was rejected after 2 weeks.

Modified to practical irradiation process, cooked rice was irradiated at the same dose of spicy chicken basil at 2 kGy. After irradiation, irradiated cooked rice was still accepted after 4 weeks compared to control sample, which rejected at 3 weeks during storage under chill condition.

Previous optimization of 2-kGy dose was based only on microbiological evaluation of total plate count. In actual case, the contamination from food pathogens could be occurred. From the contamination study, it was found that irradiation dose of 2 kGy applied for cooked rice and spicy chicken was effectively sufficient to reduce E. Coli in cooked rice and Salmonella in spicy chicken (contamination rate 108 and 104 respectively) to safe level. The samples were free from these two pathogens until the end of storage (4 weeks) at chilled condition (2+2°C). Irradiation at 2 kGy did not enough to decontaminate 2*107 L. monocytogenes (cfu/g) in spicy chicken. However, inoculation rate was quite high (2*107 cfu/g) compared to microbiological quality of ready-to-eat foods at point of sales (PHLS, 1992). Efficiency of irradiation to decontaminate pathogens was depended also on type of irradiated food. In this case, irradiation of 2 kGy could kill all L. monocytogenes in cooked rice sample contaminated at the same rate of 2*107 cfu/g and sample was safe during storage under chilled condition (Table 1).

When HACCP principles were applied, critical control points of the model ready-to-eat meal were the receiving point of raw materials, cooking, irradiation and metal detection for spicy chicken basil, while for cooked rice were only steaming, irradiation and metal detection. Controlling cooking temperature, checking irradiation dose, using metal detector and analysis of antibiotic and pesticide residue in chicken were proposed as monitoring procedures and each corrective actions were given for each critical control point.

Table 1. The amount of survival microorganism in the samples after irradiated with 2 kGy gamma ray and storage under chilled condition Initial Concentration after irradiated and storage under 2+2°C Samples Concentration (cfu/g) (cfu/g) 0 day 1 week 2 week 3 week 4 week Control Cooked rice <10 <10 <10 <10 <10 <10 <10 <10 <10 <10 <10 <10 Spicy chicken <10 <10 <10 <10 <10 <10 <10 <10 <10 <10 <10 <10 L.monocytogenes Cooked rice 2.12 x 107 ND ND ND ND ND 2.25 x 107 ND ND ND ND ND 2.23 x 107 4.40 x 1.35 x 1.25 x 2.76 x Spicy chicken 160 103 102 102 103 2.05 x 107 4.85 x 1.40 x 1.30 x 2.58 x 130 103 102 102 103 Samonella Spicy chicken 1.35 x 104 ND ND ND ND ND 1.48 x 104 ND ND ND ND ND E. coli Cooked rice 2.64 x 108 ND ND ND ND ND 2.42 x 108 ND ND ND ND ND

4. DETAILED WORK PLAN FOR NEXT COMING YEAR

From the progress work of this project, three constituents (cooked rice, spicy chicken, basil leaf) of Thai spicy chicken basil rice were suggested to packed separately. Cooked rice and spicy chicken basil was able to irradiate at 2 kGy but not for basil leaf, which sensitive to flavor and odor loss. However, from contamination study, dose of 2 kGy did not enough to decontaminate Listeria monocytogenes in spicy chicken with innoculation at 2x107 cfu/g.

1. Therefore, the higher dose irradiation (3 kGy) will be used for contamination study of L. monocytogenes in spicy chicken.

2. For basil leaf, fresh leaf will be preserved using combination of low dose (0.1-0.3 kGy) of irradiation and modified atmosphere packaging. Beside of this, drying techniques will be introduced with three different methods as follows: (1) hot air drying; (2) vacuum drying; (3) freeze drying. Operation factors in term of temperature

and time will be varied to determine the optimum drying conditions. Dried basil leaf could be packed with irradiated Thai spicy chicken basil rice.

3. Consumer test will be done by irradiation of cooked rice at 2 kGy and spicy chicken basil at selected dose (regarding to the result in experiment 1). A best method concerning quality and storability (regarding to the result in experiment 2) will be selected to preserve basil leaf. Consumer acceptance to this food will be evaluated during storing under chilled condition (2±2oC).

UNITED KINDGDOM

Effect of Gamma-Radiation On the Quality of Prepared Meals and Their Components

Chief Scientific Investigator: E.M. Stewart Agriculture, Food & Environmental Science Division Department of Agriculture and Rural Development (DARD), and Department of Food Science Queen’s University Belfast Belfast, UK

Research Agreement No.: 11912

1. INTRODUCTION

The studies reported in this paper, and to be carried out in the future, focus on ready-to-eat meals with an exotic origin such as Chicken Masala, Beef Madras etc. and on ingredients such as minced beef which could be used as components of such meals. The research looks at the effects of ionizing radiation on the quality attributes of the meals of importance to the consumer such as microbiological quality and nutritional quality.

2. MATERIALS AND METHODS

2.1. Effect of gamma radiation on the quality of a Chicken Masala ready meal

Chilled Chicken Masala prepared meals were obtained from a local manufacturer in Northern Ireland, the ingredients of which (in descending order) were as follows: [Chicken, onion, tomato, water, yoghurt, coconut, red pepper, tomato puree, rapeseed oil, modified starch, chicken bouillon, Garam Masala, garlic extract, green chillies, chilli powder, ground coriander leaf, salt, ginger, cayenne pepper, malt extract and turmeric]. Irradiation was carried out using a cobalt 60 source (Gammabeam 650, Nordion International Inc., Canada) at a dose rate of 0.42 kGy h-1 and an environmental temperature of 2-3oC. In order to confirm the dose received by the meals, gammachrome YR dosimeters (AEA Technology, Harwell, UK) were placed on a number of meals receiving dose of 1 and 2 kGy while three amber dosimeters were placed on those receiving 3 kGy. The samples were analysed for vitamins B1 (thiamine) and E (α-tocopherol), TBA (thiobarbituric acid) values, 2-alkylcyclobutanones (in order to determine if irradiation treatment of the samples could be confirmed) and for microbiological quality. The vitamin, TBAs and 2-alkylcyclobutanone analyses were carried out on the same samples. A total of 72 meals were given doses of 1, 2 or 3 kGy or left unirradiated to serve as controls thereby giving 18 meals per treatment. Within each batch of 18, six samples were analysed immediately while six were stored for 7 days and six for 14 days at 3 ± 1°C. Upon analysis, 3 meals at each dose level were analysed in the fresh state while the other three meals were reheated in a convection oven at 180°C for 20 min to give an internal temperature of 80°C. The meals were allowed to cool at ambient temperature prior to being homogenised by a food processor and analysed. For the microbiological analysis, a total of 64 samples from the same batch of meals were given the same irradiation doses as those described previously, with 16 meals being treated per dose. Within each batch of 16, four meals were analysed at each storage time of 0, 7, 11 or 14 days, two being stored at 3 ± 1°C and two at 10 ± 1°C. These meals were not reheated prior to analysis and on each sampling day were assessed for total viable count (TVC) (30°C for 48 h) on typtone soya agar

containing 0.6% yeast extract (TSAYE); coliform count (37°C for 18-24 h) on McConkey No 3 agar; and psychrotrophic count of TSAYE (7°C for 10 days).

2.2. Effect of gamma radiation on the oxidative rancidity of vacuum packed beef burgers irradiated at 5°C and 20°C and stored for 8 weeks at 3°C

A batch of freshly minced beef, containing approximately 10% lipid, was obtained from a local butcher in the Belfast area. A total of 90 burgers (100 g) were prepared and vacuum packaged. The samples were irradiated (at a dose rate of 0.48 kGy h-1) as described in Experiment 1 being given doses of 2.5 or 5.0 kGy at an environmental temperature of 5°C or 20°C or left unirradiated to serve as controls, thereby giving 15 burgers for each dose at each temperature. The actual doses received by the samples were measured using amber perspex dosimeters. Following irradiation, 18 samples at each dose were randomly allocated to storage treatments of 0, 2, 4, 6 or 8 weeks at temperatures of 3 ± 1°C. The samples were analysed for oxidative rancidity by the TBA method as described previously.

2.3. Effect of gamma radiation on the oxidative rancidity and microbiological quality of vacuum packed and over-wrapped beef burgers stored for 21 days at 3°C

On each of three separate occasions, a batch of freshly minced beef, containing approximately 15% lipid, was obtained from a local butcher in the Belfast area. Under aseptic conditions a total of 65 burgers (50 g) were produced and vacuum packaged. The samples were irradiated (at a dose rate of 5.33 kGy h-1) as described in Experiment 1 being given doses of 2.5, 5.0, 7.5 or 10.0 kGy or left unirradiated to serve as controls, thereby giving 13 samples at each dose. Following irradiation, 35 samples were retained within the vacuum packages while the other 30 were removed aseptically from the packaging in a laminar flow cabinet, placed in sterile cartons and covered in cling film in order to give 30 over-wrapped samples. Five samples were either analysed (day 0) immediately and five at each treatment after 1, 2, 5, 7, 14 or 21 days at 3 ± 1°C for microbiological quality or after 1, 2, 5, 7, 14 or 21 days at 3 ± 1°C for oxidative rancidity, thereby giving two samples per treatment. A total of 195 samples were analysed over the three occasions. The same burgers were used for both the oxidative rancidity and microbiological assessment, the samples for microbiological analysis being taken under aseptic conditions prior to the burgers being analysed for oxidative rancidity. On each sampling day, the microbial quality of the vacuum packed samples was assessed using total viable, anaerobic, coliform; psychrotrophic and lactic acid counts. For the over-wrapped samples, total viable, psychrotrophic, pseudomonad and coliform counts were carried out.

2.3. Statistical Analysis

The results for all the experiments were subjected to analysis of variance.

3. RESULTS

3.1. Effect of gamma radiation on the quality of a Chicken Masala chilled ready meal

The results are presented in Tables 1-4 and in Figures 1-2. It can be seen that the TBA values (Table 1) decreased with increasing irradiation dose over the range 0 to 3 kGy. The results obtained for oxidative rancidity were surprising as it was expected that the TBA values would increase upon irradiation, storage and cooking. One possibility for this result is the presence of antioxidants in the meals which were quenching the free radicals produced by the ionising radiation. The meals proved to be a good source of vitamin E (α-tocopherol) (Table 2), a well

known natural antioxidant, and contained a range of ingredients such as onion, tomato, red pepper and spices, which are all known for their antioxidant activity. Another explanation could be that there is a preferential attack mechanism by the free radicals on the α-tocopherol and thiamine (Table 3) present in the meals, thereby reducing the concentration of these vitamins. This consequently resulted in decrease in the TBA values with an increase being observed in the amount of α-tocopherol and thiamine destroyed. The effects of irradiation on the vitamin B1 and E content of the meals are given in Tables 2 and 3, respectively. It was found that irradiation dose, storage and re-heating all had a very highly significant effect on the concentration of both of these vitamins. Both 2-DCB and 2-DCB were detected in the meals although both ions m/z 98 and 112 were not detected in all of the samples analysed. Irradiation had a very highly significant effect on the concentration of both 2-DCB and 2- TCB in the chilled meals, with higher levels of 2-DCB being detected compared to 2-TCB. Cooking or storage did not have a significant effect on the levels of the cyclobutanones. With regard to microbiological quality (Figs. 1 – 2), in all cases, irradiation significantly reduced the levels of bacteria in the prepared meals.

3.2. Effect of gamma radiation on the oxidative rancidity of vacuum packed beef burgers irradiated at 5°C and 20°C and stored for 8 weeks at 3°C

Results for this experiment are given in Table 5. There was a significant in the TBA values of the beef burgers made from minced beef upon irradiation. Overall, a reduction of 12% was measured between 0 and 2.5 kGy with a further decrease of only 1% between 2.5 and 5.0 kGy. The TBA values were lower for the meals irradiated at the higher temperature of 20°C compared to those treated at 5°C, an overall difference of 10% being observed. t is worthy of note that as the TBA values were <1 over the 0 to 5 kGy dose range and 8 week storage period, the minced beef was considered to be of acceptable quality from an organoleptic point of view as no off-flavours due to oxidative rancidity would occur at such as low values. The low TBA values obtained for the samples were not unexpected as the samples were vacuum packaged thereby would have had a restricted supply of oxygen.

3.3. Effect of gamma radiation on the oxidative rancidity and microbiological quality of vacuum packed and over-wrapped beef burgers stored for 21 days at 3°C

The results for this experiment are presented in Table 6 and Figures 3 to 7. Table 7 shows a comparison of the TBA values obtained for beef burgers irradiated in vacuum samples followed by subsequent storage for 21 days in the vacuum packages or after being transferred to over-wrapped packages. Unlike the vacuum packed samples, it can be seen that the TBA values of the over-wrapped samples increased with increasing storage time, although it was noted that little change occurred over the first two days of storage. Both irradiation and storage significantly affected the microbiological quality of both the over-wrapped and vacuum packaged beef burgers (Figs. 3 – 7). Irradiation significantly reduced the numbers of organisms present in both the vacuum packed and over-wrapped burgers.

4. CONCLUSIONS

The work carried out to date on both the prepared meals and minced beef, the latter being a potential component of many types of prepared meals, showed that:

• irradiation doses of 2 to 3 kGy can extend the shelf-life for at least 14 days at refrigeration temperatures;

• irradiation under vacuum packaging will significantly enhance shelf-life as numbers of bacteria are significantly reduced for up to 21 days with oxidative rancidity being well below the acceptable levels;

• storing irradiated meals or their components in over-wrapped packages will significantly increase the microbial numbers present and result in an increase of oxidative rancidity thereby resulting in a decrease in overall quality and acceptability. It may therefore be beneficial to add natural antioxidants to the meals in order to reduce the rate of oxidation. The addition of natural antimicrobials may also lead to an improvement in microbial quality;

• irradiation will reduce the levels of vitamins in the meals, although the diminution would not prove to have a significant impact on nutritional value; and

• The 2-alkylcyclobutanone (EN1785) standard method can be used to detect irradiation treatment of the meals analysed and as demonstrated from previous work will detect irradiated minced beef contained within meals.

Table 1. Effect of gamma radiation on the TBA values (mg malonaldehyde kg-1) of a Chicken Masala ready-meal stored for up to 14 days at 3 ± 1°C Irradiation Storage Uncooked Cooked Mean Mean dose time (Not reheated) (Reheated) (kGy) (days) TBARS (mg malonaldehyde kg1) 0 0 0.388 0.288 0.313 0.356 7 0.322 0.361 0.342 14 0.457 0.367 0.412 1 0 0.283 0.276 0.280 0.355 7 0.452 0.359 0.406 14 0.372 0.390 0.381 2 0 0.284 0.247 0.266 0.343 7 0.408 0.374 0.391 14 0.377 0.369 0.373 3 0 0.263 0.237 0.250 0.309 7 0.341 0.317 0.329 14 0.390 0.309 0.350 Mean 0.357 0.325

Table 2. Effect of gamma radiation on the thaimine content (mg g-1) of a Chicken Masala ready-meal stored for up to 14 days at 3 ± 1°C

Irradiation Storage time Uncooked Cooked Mean Mean dose (kGy) (days) (Not reheated) (Reheated) Thiamine concentration (µg g-1) 0 0 6.65 6.52 6.58 5.98 7 6.04 5.40 5.72 14 5.89 5.39 5.64 1 0 5.36 5.11 5.23 5.03 7 5.49 4.54 5.01 14 5.17 4.51 4.84 2 0 5.13 4.77 4.95 4.70 7 4.44 4.49 4.46 14 4.86 4.49 4.67 3 0 4.40 4.38 4.39 4.15 7 4.03 3.94 3.98 14 4.06 4.11 4.08 Mean 5.13 4.80

Table 3. Effect of gamma radiation on the Vitamin E content (mg g-1) of a Chicken Masala ready-meal stored for up to 14 days at 3 ± 1°C Irradiation Storage time Uncooked Cooked Mean Mean dose (kGy) (days) (Not reheated) (Reheated) Vitamin E concentration (µg g-1) 0 0 23.05 21.99 22.52 19.42 7 19.55 16.23 17.89 14 19.42 16.25 17.84 1 0 19.72 19.91 19.81 18.44 7 19.14 16.69 17.92 14 19.22 15.95 17.59 2 0 18.91 15.12 17.02 16.71 7 17.49 16.42 16.95 14 17.30 15.02 16.16 3 0 17.32 16.52 16.92 16.12 7 16.63 14.82 15.73 14 16.60 14.83 15.71 Mean 18.70 16.65

Table 4. Effect of gamma radiation on the 2-alkylcyclobutanone content (mg g-1 lipid) of a Chicken Masala ready-meal stored for up to 14 days at 3 ± 1°C Irradiation Storage time Uncooked Cooked dose (kGy) (days) (Not reheated) (Reheated) Concentration (µg g-1 lipid) DCB TCB DCB TCB 1 0 0.064 0.009 0.071 0.015 7 0.072 0.022 0.063 0.013 14 0.079 0.010 0.063 0.015 Mean 0.071 0.014 0.066 0.015 2 0 0.115 0.031 0.113 0.025 7 0.129 0.039 0.114 0.029 14 0.106 0.031 0.116 0.028 Mean 0.117 0.034 0.117 0.028 3 0 0.165 0.048 0.193 0.055 7 0.199 0.065 0.143 0.036 14 0.152 0.045 0.190 0.057 Mean 0.172 0.053 0.175 0.049

(a) (b)

10 10 0 kGy 1 kGy 2 kGy 3 kGy L.O.D 0 kGy 1 kGy 2 kGy 3 kGy L.O.D

8 8

6 6 cfu/g 10

Log 4 4

2 2

0 0 0246810121402468101214 Storage (days) Storage (days) Figure 1: Effect of irradiation treatment and storage on the total viable counts (TVCs) of Chicken Masala ready-meals stored at 3 ± 1°C (a) or 10 ± 1°C (b) [L.O.D. = limit of detection (1.70 -1 Log10 cfu g )]

(a)

10 0 kGy 1 kGy L.O.D (b)

10 8 0 kGy 1 kGy 2 kGy 3 kGy L.O.D

8 6

4 cfu/g 10

Log 4

2 2

0 0 02468101214 02468101214 Storage (days) Storage (days)

Table 5. Effect of gamma radiation on the TBA values (mg malonaldehyde kg-1) of vacuum packaged minced beef treated at different temperatures and stored for 8 weeks at 3 ± 1°C

Irradiation Storage time Irradiated at Irradiated at 20°C Mean Mean dose (kGy) (weeks) 5°C TBARS (mg malonaldehyde kg-1) 0 0 0.666 0.606 0.636 0.589 2 0.568 0.509 0.539 4 0.663 0.549 0.606 6 0.653 0.517 0.585 8 0.655 0.504 0.580 Mean 0.641 0.537 2.5 0 0.681 0.606 0.644 0.516 2 0.429 0.505 0.467 4 0.557 0.481 0.519 6 0.458 0.406 0.433 8 0.515 0.520 0.517 Mean 0.528 0.503 5.0 0 0.634 0.549 0.592 0.513 2 0.499 0.437 0.468 4 0.494 0.541 0.517 6 0.484 0.502 0.493 8 0.554 0.434 0.494 Mean 0.533 0.493

Table 6. Effect of gamma radiation on the TBA values (mg malonaldehyde kg-1) of over- wrapped and vacuum packaged minced beef stored for up to 21 days at 3 ± 1°C Irradiation Storage time Overwrapped Vacuum packed Mean Mean dose (kGy) (days) TBARS (mg malondialdehyde kg-1) 0 1 0.346 0.281 0.313 0.458 2 0.351 0.283 0.317 5 0.577 0.273 0.425 7 0.640 0.289 0.464 14 0.887 0.458 0.672 21 0.640 0.479 0.559 Mean 0.573 0.344 2.5 1 0.387 0.309 0.348 0.671 2 0.515 0.335 0.425 5 0.835 0.387 0.611 7 1.027 0.346 0.686 14 1.231 0.538 0.885 21 1.633 0.504 1.069 Mean 0.938 0.403 5.0 1 0.559 0.348 0.454 0.844 2 0.598 0.307 0.452 5 0.866 0.328 0.597 7 1.113 0.317 0.715 14 2.106 0.374 1.240 21 2.834 0.382 1.608 Mean 1.346 0.343 7.5 1 0.551 0.354 0.453 1.135 2 0.757 0.325 0.541 5 1.173 0.351 0.762 7 1.693 0.380 1.036 14 2.982 0.361 1.672 21 4.321 0.377 2.349 Mean 1.913 0.358 10.0 1 0.541 0.405 0.473 1.160 2 0.762 0.385 0.573 5 1.232 0.408 0.820 7 1.615 0.380 0.997 14 3.356 0.364 1.860 21 4.059 0.416 2.237 Mean 1.927 0.393

(a)

12 0 kGy 2.5 kGy L.O.D (b)

10 12 0 kGy L.O.D

10 8

8 6

6 4 4

2 2

0 0 0 3 6 9 12 15 18 21 0 3 6 9 12 15 18 21 Storage (days) Storage (days) Figure 3: Effect of irradiation treatment and storage on the total viable counts (TVCs) of vacuum packaged (a) or over-wrapped (b) beef burgers stored at 3 ± 1°C [L.O.D. = limit of -1 detection (1.7 Log10 cfu g )]

(a) (b)

12 12 0 kGy 2.5 kGy 5.0 kGy L.O.D 0 kGy 2.5 kGy L.O.D

10 10

8 8

6 6

4 4

2 2

0 0 0 3 6 9 12 15 18 21 0 3 6 9 12 15 18 21 Storage (days) Storage (days) Figure 4: Effect of irradiation treatment and storage on the psychrotrophic counts of vacuum packaged (a) or over-wrapped (b) beef burgers stored at 3 ± 1°C [L.O.D. = limit of -1 detection (1.7 Log10 cfu g )]

(b) (a)

12 12 0 kGy L.O.D 0 kGy L.O.D

10 10

8 8

6 6

4 4

2 2

0 0 0 3 6 9 12 15 18 21 0 3 6 9 12 15 18 21 Storage (days) Storage (days) Figure 5: Effect of irradiation treatment and storage on the pseudomonad counts of vacuum packaged (a) or over-wrapped (b) beef burgers stored at 3 ± 1°C [L.O.D. = limit of detection (1.7 -1 Log10 cfu g )]

8 0 kGy 2.5 kGy L.O.D

0 0 3 6 9 12 15 18 21 Storage (days) Figure 6: Effect of irradiation treatment and storage on the anaerobic bacteria in vacuum packaged (a)

or over-wrapped (b) beef burgers stored at 3 ± 1°C [L.O.D. = limit of detection (0.70 Log10 cfu g-1)]

8 0 kGy 2.5 kGy 5.0 kGy L.O.D

0 0 3 6 9 12 15 18 21 Storage (days) Figure 7: Effect of irradiation treatment and storage on the lactic acid bacteria in vacuum packaged (a) or over-wrapped (b) beef burgers stored at 3 ± 1°C [L.O.D. = limit of detection (0.70 -1 Log10 cfu g )]

UNITED STATES

Consumer Acceptance of Irradiated Prepared and Processed Food

Chief Scientific Investigator: Rodolfo M. Nayga, Jr. Professor Department of Agricultural Economics Texas A&M University USA

1. OUR STUDY

Although the FDA has approved the irradiation of food, at specified levels, to prevent foodborne diseases, it is unclear if consumers are willing to accept and pay for irradiated food products. In spring 2002, we conducted a study to assess (1) consumers’ knowledge and acceptance of food irradiation, (2) the effects of information regarding the food irradiation technology on consumer acceptance, and (3) the willingness to pay (WTP) for irradiated ground beef. We interviewed 474 consumers at various stores of a regional supermarket chain in Austin, San Antonio, Waco, and Houston.

Potential buyers’ of irradiated foods can be grouped into four segments, namely, strong buyers, interested, doubters and rejecters. During the interview process, we provided each respondent with Information I (nature and benefits of food irradiation) and Information II (difference between use of electron beam and gamma rays to irradiate food products) (Figure 1). The respondents were asked, before and after the presentation of information, in which of the four segments they think they belong in regards to irradiated foods. We also asked the respondents who were willing to buy irradiated ground beef about their willingness to pay (WTP) a premium for a pound of irradiated ground beef.

Figure 1. Information I and II

______

Information I

General statement about the benefit of food irradiation excerpted from United States General Accounting Office (GAO), Washington, D.C.

Food irradiation is the process of exposing food to controlled levels of ionizing radiation. Ionizing radiation is a type of energy similar to radio and television waves, microwaves, and infrared radiation. The high energy produced allows it to penetrate deeply into food, killing microorganisms without significantly raising the food’s temperature.

Irradiation, within approved dosages, has been shown to destroy at least 99.9 percent of common foodborne pathogens, such as Salmonella (various species), E.coli O157:H7, and Listeria monocytogenes, which are associated with meat and poultry. Irradiation can also prolong shelf-life, decontaminate spices and control insect infestation.

Information II

The FDA mandates that all foods treated with ionizing energy be labeled as having been irradiated. Note that ionizing radiation has the same effect on food, but its source of energy is different. In the United States, Gamma rays and Electron beams are used for food irradiation.

1. Gamma rays

The source of gamma rays is radioactive cobalt-60. The cobalt is held in thin stainless steel rods that are placed on a rack in an irradiation facility. The rack holding the cobalt containing rods is stored in a deep pool of water in the irradiation facility. When products are to be irradiated, the rack is raised from the pool and products are conveyed past the rack to absorb the gamma rays being emitted from the cobalt rods. Gamma rays are also produced from radioactive cesium-137, but those from cesium are not used for food irradiation.

2. Electron Beam

The source of electron beam is a machine, much like a television set, which utilizes electricity to generate electrons and accelerates the electrons to nearly the speed of light. When products are to be irradiated, the electron beam accelerator is switched on and products are conveyed through the stream of electrons. The Electron beam process is also called Electronic pasteurization. No radioactive materials are ever used, produced or deposited with this process.

2. WTP EXPERIMENT

We gave each respondent a pound of non-irradiated ground beef and some money (first bid value that is equivalent to one of the bid values calculated from DWEABS model we used to determine optimal bid values) as a gift for the survey participation. The respondent was then asked his/her willingness to exchange the pound of non-irradiated ground beef and first bid money “for” a pound of irradiated ground beef. If the respondent accepted the bid, the WTP value was recorded as the first bid value. However, if the respondent rejected the bid, he/she was asked again his/her willingness to exchange “a pound of non-irradiated ground beef and a half value (second bid) of the money” for a pound of irradiated ground beef. If the answer was “Yes” the second bid value is recorded as his/her WTP, otherwise, the WTP is assumed to be lower than the second bid value. 2.1. Information Effects

Information plays a very important role on consumer buying decisions. Before Information I and II were presented, about 45% of our sample had zero knowledge of food irradiation, 51 % would not buy irradiated ground beef, and only 8.5% considered themselves "strong buyers". After the presentation of Information I and II, 94% of the sample indicated that they would buy irradiated ground. In addition, the percentage of respondents indicating that they would be a "strong buyer" increased from 8.5% to 42.2% while the percentage of respondents indicating that they would be "doubters" or "rejecters" decreased significantly from 14.3% to 3.1% for "doubters" and from 3.9% to 0.6% for "rejecters" (Figure 2).

Figure 2

80% 73.24% 66.46% 70% 53.99% 60% Before After Info1 50% 42.23% After Info2 40% 28.13% 30% 14.32% 20% 8.51% 3.54% 3.94% 10% 3.15% 1.88% 0.63% 0% Strong Buyer Interested Doubter Rejecter

Figure 3 shows segment movement after presentation of the two types of information regarding food irradiation. About 68% of the strong buyers prior to the presentation of the information remained in the "strong buyer" segment after the information was presented while 43% of the interested buyers prior to the presentation of information switched to become "strong buyers" after the presentation of information. About 24% of "doubters" and 41% of " rejecters" prior to the presentation of information switched to the "strong buyer" segment after the presentation of information.

Figure 3 80% 68% 70% 65% Strong buyer Interested 60% 56% Doubters Rejecters 50% 43% 40% 41% 35% 30% 25% 24% 18% 20% 9% 5% 6% 10% 3% 1% 2% 0% Strong buyer Interested Doubters Rejecters

2.2. Willingness to purchase and pay more for irradiated prepared ground beef

Before the presentation of Information I and II, only about half of the respondents indicated willingness to buy irradiated ground beef. After the presentation of Information I, 88.5 % of the respondents indicated a willingness to buy irradiated ground beef. Even more (94.12 %) indicated a willingness to buy irradiated ground beef after the presentation of Information II. The percentage of respondents in this study who are willing to buy and pay a premium for irradiated ground beef appear higher than estimates from previous study conducted by FoodNet Population Survey (1998-1999). According to the CDC as well, typically at least half will buy the irradiated food, if given a choice between irradiated product and the same product non-irradiated. If consumers are first educated about what irradiation is and why it is done, approximately 80% will buy irradiated products.

2.3. Results from WTP experiment

The willingness to pay experiment on the first bid values show that among the respondents who were asked their willingness to pay 10 cents more per pound of irradiated ground beef,

97.3% said, “Yes” to the offer. As the bid values increased, the proportion of respondents saying “Yes” also went down (Table 1). In the second bid offer, among those who said, “No” to the first bid offer (once we cut the offer into half), 100% said “Yes” to the 5 cents per pound offer and 67% said “Yes” to the 20 cents per pound offer.

Table 1. First bid offer 10 40 60 80 120 (cents)

Accept 97.3% 72.22% 69.8% 56.86% 46.15%

Reject 2.7 % 27.78% 30.2% 43.14% 53.85%

Second bid 5 20 30 40 60 offer (cents)

Accept 100% 66.7% 75% 59.9% 42.86%

Reject 33.3% 25% 40.1% 57.14%

2.4. Some Perspective

Despite the high level of safety in the US food supply, microbiological hazards exist. Illnesses and death due to foodborne pathogens costs society billions of dollars due to lost productivity and medical expenses. Processing by irradiation enhances food safety through the reduction of potential pathogens in raw meat and poultry products, thus reducing the likelihood of illness from cross contamination or inadequate cooking Our study investigated consumers' willingness to buy and pay for irradiated ground beef. In general, we found that information about the nature and benefits of food irradiation is a major factor affecting consumers' perception and attitudes toward irradiated foods. Many consumers are quite willing to buy irradiated foods. This is particularly true if the purpose of the irradiation is clearly indicated. Consumers showed interest in a process that eliminates harmful microbes from the food and reduce the risk of foodborne disease. This finding reflects the importance of educating the public to the hazards of foodborne pathogens and to the potential benefits of consuming irradiated foods. Food irradiation, however, is not a short cut that means food hygiene efforts can be relaxed. Irradiation is a major step forward, but it does not replace other important efforts, including efforts to improve sanitation on the farm and in the food processing plan and educating the consumers about proper food handling and cooking techniques.

2.5. Future Work

Plan for the next year includes writing a journal article focused on the details of willingness to pay experimental results. Also, I would not mind working with other members of the CRP who would like to include a marketing and economic component similar to what we have done in their respective projects.

2.6. Published Work So Far

[1] Nayga Jr., R.M., W. Aiew, and J. Nichols, “Information Effects on Consumers’ Willingness to Purchase Irradiated Food Products”, Review of Agricultural Economics, 26(2004): forthcoming. [2] Aiew, W., R. Nayga, Jr., and R. Woodward, “The Treatment of Income Variable in Willingness to Pay Studies”, Applied Economics Letters, 11(2004): forthcoming. [3] Nayga Jr., R.M., A. Poghosyan, and J. Nichols, “Will Consumers Accept Irradiated Food Products?” International Journal of Consumer Studies, 28,2 (March 2004): 178- 185. [4] Nayga Jr., R.M., “Food Safety Through Food Irradiation: Should it be Adopted More by the EU?”, EuroChoices, 2,3(2003): 36-39. [5] Aiew, W., R.M. Nayga, Jr., and J.P. Nichols, “The Promise of Food Irradiation: Will Consumers Accept It”, Choices, Third Quarter 2003, pp. 31-34. [6] Aiew, M., R.M. Nayga, Jr., and J. Nichols, “Experimental Study on Willingness to Purchase Safer Foods: The Case of Irradiated Foods”, Quality Assurance, Risk Management and Environmental Control in Agriculture and Food Supply Networks, Proceedings of the 82nd Seminar of the European Association of Agricultural Economists, G. Schiefer and U. Rickert, eds., University of Bonn-ILB Press, Bonn, Germany, 2004, pp. 405-412. [7] Nayga Jr., R.M., A. Poghosyan, and J. Nichols, “Consumer Willingness to Pay for Irradiated Beef: Initial Phase”, Paradoxes in Food Chains and Networks, J.H. Trienekens and S.W.F Omta (editors), Wageningen Academic Publishers, The Netherlands, 2002, pp. 250-259.

ANNEX 1

List of Participants

Dr. Patricia Narvaiz CNEA, Food Preservation Section Technological and Agricultural Applications Ezeiza Atomic Center Pbtro. Juan Gonzalea y Aragon NO. 15 B 1802AYA – Pcia. de Buenos Aires ARGENTINA Tel.: +54 011 6779 8556 Fax: +54 011 6779 8322 Email: [email protected]

Dr. Josephine Nketsia –Tabiri Ghana Atomic Energy Commision Biotechnology and Nuclear Agriculture Research Institute Department of Food Science and Radiation Technlolgy P.O.Box 80, Legon- Accra GHANA Tel.: +233 21 402286/ 401454 Fax: +233 21 400807 Email: [email protected] OR [email protected]

Dr. Zubaidah Irawati Koenari Centre for Research and Development of Isotopes and Radiation Technology P.O. Box 7002, JKSKL Jakarta 12070 INDONESIA Tel.: +62 21 769 0709 Fax: +62 21 769 1607; +62 21 751 3270 Email: [email protected]

Dr. Cheorun Jo KAERI, Radiation Food Science and Biotechnology Team P.O. Box 105 Yuseong Daejeon 305-600 Republic of KOREA Tel: +82 42 868 8065 Fax: +82 42 868 8043 Email: [email protected]

Dr. Amanda Minnaar Department of Food Science University of Pretoria Lynnwood Road Pretoria 0002 SOUTH AFRICA Tel.: +27 12 420 3239 Fax: +27 12 420 2839 Email: [email protected]

Dr. Mahfouz Al-Bachir Syrian Atomic Energy Commission Radiation Technology Department Food Irradiation Division P.O. Box 6091 Damascus, SYRIA Tel.: +61 11 926 17 Fax: +61 12 22 89 Email: [email protected]

Dr. Athapol Noomhorm Asian Institute of Technlogy Food Engineering and Bioprocess Technology Km 41, Phaholyothin Hwy. P.O. Box 4 Klongluang Pathumthani, 12120 THAILAND Tel.: +66 2 524 5476 Fax: +66 2 524 6200 Email: [email protected]

Dr. Eileen Stewart Queen’s University Belfast (QUB) Department of Agriculture and Rural Development (DARD) Agriculture and Food Science Centre Newforge Lane Belfast BT9 5PX UK Tel.: +44 2890 255 345 Fax: +44 2890 255 006 Email: [email protected]

Dr. Baozhong Sun Institute of Animal Scince Chinese Academy of Agricultural Science Beijing, P.R. CHINA 100094 Tel.: +86 10 628 16010 Fax: +86 10 628 95351 Email: [email protected] OR [email protected]

Dr. Ramesh Chander (replacing Dr. Sharma) Food Technology Division Bhabha Atomic Research Centre Trombay, Mumbai 400 085 INDIA Tel.: +91 22 5595374 Fax: +91 22 5505 151 OR + 91 22 2551 9613 Email: [email protected] [email protected]

Dr. Yaara Haruvy Israel Atomic Energy Commission Soreq Nuclear Research Centre Operations Division, QA Department Yavne 81800 ISRAEL Tel.: +972 8943 4414 Fax: +972 89 43 4403 Email: [email protected]

Dr. Rudy Nayga Department of Ag. Economics Texas A&M University College Station, TX 77843-2124 USA Tel.: 1 979 845 8376 Fax: 1 979 862 3019 Email: [email protected]

Dr. Ioannis Savvaidis University of Ioannina; School of Natural Sciences Dept. of Chemistry; Laboratory of Food Chemistry & Technology P.O. Box 1186 GR-45110, Ioannina GREECE Tel.: +30 265 10 983 43 Fax: +30 265 10 987 95 Email: [email protected]

IAEA

Dr Tatiana Rubio-Cabello (Scientific Secretary) International Atomic Energy Agency Joint FAO/IAEA Division of Nuclear Techniques in Food and Agriculture Wagramer Strasse 5, P.O. Box 100 A- 1400 Austria Tel: +43 1 2600 21639 Fax: +43 1 26007 E-mail: [email protected]

Observers:

Mr. Joseph Abu Department of Food Science University of Pretoria Lynnwood Pretoria 0002 SOUTH AFRICA Tel.: +27 12 420 3413 Fax.: +27 12 420 2839 Email: [email protected]

Dr. Elna Buys Department of Food Science University of Pretoria Lynnwood Pretoria 0002 SOUTH AFRICA Tel.: +27 12 420 3209 Fax.: +27 12 420 2839 Email: [email protected]

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Ms. Edith Fombang Department of Food Science University of Pretoria Lynnwood Pretoria 0002 SOUTH AFRICA Tel.: +27 12 420 3413 Fax.: +27 12 420 2839 Email: [email protected]

Mr. Obert Gurira Department of Food Science University of Pretoria Lynnwood Pretoria 0002 SOUTH AFRICA Tel.: +27 12 420 3413 Fax.: +27 12 420 2839 Email: [email protected]

Ms. Karlien Nortjé Department of Food Science University of Pretoria Lynnwood Pretoria 0002 SOUTH AFRICA Tel.: +27 12 420 3413 Fax.: +27 12 420 2839 Email: [email protected]

Ms. Angela Parry-Hanson Department of Food Science University of Pretoria Lynnwood Pretoria 0002 SOUTH AFRICA Tel.: +27 12 420 3413 Fax.: +27 12 420 2839 Email: [email protected]

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ANNEX 2

Program

Monday 26 April 09:00 - 09:30 Inaugural Session: Prof J. Kirsten (Chair Person: School of Agricultural and Food Sciences, University of Pretoria ) Dr T. Rubio-Cabello (FAO/IAEA) Prof A. Minnaar (SAF) 09:30 - 10:00 Objectives of the RCM. Dr. Tatiana Rubio-Cabello 10:30 - 11:00 Break 11:00 - 12:00 Publication of results. IAEA-TECDOC Administrative matters 12:00- 13:00 Lunch 13:00 - 14:00 Safer prepared meals for immunocompromised patients and the general consumer, by gamma irradiation. P. Narvaiz (Argentina) 14:00 - 15:00 Improving the microbiological quality and safety of prepared meals consumed in Greece by low dose irradiation. I. Savvaidis (Greece) 15:00 - 15:30 Break 15:30 - 16:30 Irradiation of prepared meals for microbiological safety and shelf-life extension. J. Nketsa-Tabiri (Ghana) 16:30 - 17:30 Radiation processing to ensure the safety and quality of ethnic prepared meals R. Chander (India) Tuesday 27 April 09:00 - 10:00 Irradiation to ensure the safety and quality of home style frozen foods: 1. Liquid based materials. Z.I. Koenari (Indonesia) 10:00 - 11:00 HACCP protocols for ready to eat meals pasteurised with ionising radiation. Y. Haruvy (Israel) 11:00 - 11:30 Break 11:30 - 12:30 Irradiation application on ready-to-cook Galbi, Korean traditional meat product for safety and extending shelf-life. C. Jo (Korea) 12:30 - 13:30 Lunch

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13:30 - 14:30 Use of irradiation to improve the safety and quality of ethnic South African foods. K. Nortjé, E.M. Buys, A. Minnaar (South Africa) 14:30 - 15:30 Use of irradiation to improve the safety and quality of Thai prepared meal. A. Noomhorm (Thailand) 15:30 - 16:00 Break 16:00 - 17:00 Effect of gamma-radiation on the quality of prepared meals and their components. E.M. Stewart (UK) Wednesday 28 April 09:00 - 10:00 Consumer acceptance of irradiated prepared and processed food R. Nayga (USA) 10:00 - 11:00 Effect of gamma irradiation on the microbial load, chemical and sensory characteristics of locally prepared meals. M. Al-Bachir (Syria) 11:00 - 11:30 Break 11:30 - 12:30 Use of irradiation to ensure the safety and quality of Chinese dumpling. B. Sun (China) 12:30 - 13:30 Lunch 13:30 - 15:30 Working groups 15:30 - 16:00 Break 16:00 - 17:30 Working report Thursday 29 April 09:00 - 10:30 Conclusions and recommendations 10:30 - 11:00 Break 11:00 - 12:30 Final draft and revision of the written report 12:30 - 14:00 Lunch 14:00 - 17:30 Visit to Irradiation facility Friday 30 April 09:00 - 10:30 Approval of the report 10:30 - 11:00 Break 11:00 - 11:30 Closing ceremony

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ANNEX 3

Publications to Date

Aiew, W., R.M. Nayga, Jr., and J.P. Nichols, “The Promise of Food Irradiation: Will Consumers Accept It”, Choices (published by the American Agricultural Economics Association), Third Quarter 2003, pp. 31-34.

Aiew, M., R.M. Nayga, Jr., and J. Nichols, “Experimental Study on Willingness to Purchase Safer Foods: The Case of Irradiated Foods”, Quality Assurance, Risk Management and Environmental Control in Agriculture and Food Supply Networks, Proceedings of the 82nd Seminar of the European Association of Agricultural Economists, G. Schiefer and U. Rickert, eds., University of Bonn-ILB Press, Bonn, Germany (2004) pp. 405-412.

Aiew, W., R. Nayga, Jr., and R. Woodward, “The Treatment of Income Variable in Willingness to Pay Studies”, Applied Economics Letters, 11 (2004): forthcoming.

Farkas, J., “Irradiation on minimaly processed foods”,. Chapter 10, In: R.A. Molins (ed.) Food Irradition: Principles and Applications, John Wiley & Sons, Inc., NewYork, (2001) pp.273-290.

Farkas, J., Radiation decontamination of spices, herbs,condiments, and other dried food ingredients. Chapter 11. In: R.A. Molins (ed.) Food Irradiation: Principles and Applications. John Wiley & Sons, Inc., New York, ( 2001) pp. 291-312.

Farkas, J. Food irradiation. In: A. Mozumder and Y. Hatano (eds.) Charged Paticle and Photon Interactions with Matter. Marcel Dekker, Inc., New York, Basel (2004) pp. 785-812.

Farkas, J., The potential of new physical preservation methods in improving microbiological safety of minimally processed foods. In: L. Gasparlin and B. Zlender (eds.) Proceedings of 'Food Safety', 22nd Food Technology Days 2004, organized by the University of Ljubljana, held in Radenci, 18-19 March 2004, pp. 15-28

Farkas, J., Elimination of food-borne pathogens by ionising radiation. In: F.J.M. Smulders, J.D. Collins (eds.) Food Safety Assurance and Veterinary Public Health, Vol.2. Safety Assurance during Food Processing. Wageningen Academic Publishers, The Netherlands, 2004. pp. 157-176.

Farkas, J. 'World-wide status of food irradiation' keynote lecture presented at the 2nd Central European Congress on Food, Budapest, 26-28 April 2004, submitted for publication in the journal Acta Alimentaria, will be available also on the CD of the Congress Proceedings.

Jo, C., D. Kim, M. Shin, M. Kang, and M. Byun, “Irradiation Effect of Bulgogi Sauce for Manufacturing Korean Traditional Meat Products, Bulgogi”, Radiat. Phy. Chem. 68,5 (2003): 851-856.

Jo, C., C. Han, K. Chung, and M. Byun, “Gamma Irradiation of Ready to Cook Bulgogi Improves Safety and Extends Shelf-life without Compromising Organoleptic Quality”, Nutraceutical Foods, 8,2 (2003):1 91-195.

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Jo, C., N. Lee, H. Kang, D. Kim, and M. Byun, “Inactivation of Foodborne Pathogens in Marinated Beef Rib by Ionizing Radiation”, Food Microbiology, 21,5 (2004):543-548.

Kang, H., C. Jo, N. Lee, and M. Byun, “Effect of Gamma Irradiation on Microbial Growth, Electron Donating Ability, and Lipid Oxidation of Marinated Beef Rib (Galbi) with Different Packaging Methods”, Korean Journal of Food Science Technology, 36,1(2004):168-173.

Lee, N., C. Jo, H. Kang, D. Kim, and M. Byun, “Radio-sensitivity of Pathogens in Gamma Irradiated Marinated Beef Rib (Galbi) and its sensory property”, Korean Journal of Food Science Technology, 36,1 (2004):168-173.

Nayga Jr., R.M., W. Aiew, and J. Nichols, “Information Effects on Consumers’ Willingness to Purchase Irradiated Food Products”, Review of Agricultural Economics, 26(2004): forthcoming.

Nayga Jr., R.M., A. Poghosyan, and J. Nichols, “Will Consumers Accept Irradiated Food Products?” International Journal of Consumer Studies, 28,2 (March 2004): 178-185.

Nayga Jr., R.M., “Food Safety Through Food Irradiation: Should it be Adopted More by the EU?”, EuroChoices (published by the European Association of Agricultural Economists), 2,3(2003): 36-39.

Nayga Jr., R.M., A. Poghosyan, and J. Nichols, “Consumer Willingness to Pay for Irradiated Beef: Initial Phase”, Paradoxes in Food Chains and Networks, J.H. Trienekens and S.W.F Omta (editors), Wageningen Academic Publishers, The Netherlands (2002) pp. 250-259.

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