STANDARD PROCEDURE FOR MICROBIOLOGICAL EXAMINATION OF Arthrobacter Sp IN FOOD PRODUCTS

GELLING AGENTS USED IN FOOD INDUSTRY

INTRODUCTION

Within the food industry, stabilisers, thickeners and gelling agents are often more simply referred to as food hydrocolloids. The hydrocolloids traditionally used in food thickening and gelling include, but are not limited to, the following: agar, alginates, arabic, carrageenan, cassia tora, carboxymethyl cellulose, gelatin, gellan, guar, karaya, konjac, locust bean gum, methyl cellulose, hydroxypropylmethyl cellulose, microcrystalline cellulose, pectin, starches, tara, tragacanth and xanthan.

Many of these ingredients are carbohydrates but at least one important hydrocolloid, gelatin, is a protein. Most are agricultural derivatives but some are biotechnology derived, and gelatin, of course, is an animal product.

Many, if not all, of the thickening and gelling agents for food are available in a wide range of differentiated grades. Starch is a good example. There are literally hundreds of different food starches based on different raw materials and production process conditions. A double derivatised, waxy maize starch is totally different from, for example, a pregelatinised potato starch. Cellulose derivatives, such as carboxymethyl cellulose, microcrystalline cellulose, methyl cellulose and hydroxypropylmethyl cellulose, come in a virtually limitless range of differentiated grades depending on the degree of substitution and other processing factors.

A large number of ‘new’ and ‘differentiated’ properties have been and continue to be developed for hydrocolloids that fall under an ‘umbrella category’; for example methyl cellulose, hydroxypropylmethyl cellulose and microcrystalline cellulose can be produced in a multitudeof grades to suit a wide range of specific functional needs. Xanthan is offered in different mesh sizes, rapidly hydrating, brine tolerant and/or as a clarified grade. New versions are constantly being developed and assure the specialty future of at least part of this market.

The following is a brief overview of the key functional properties for which these ingredients are used. Nutritional properties are relatively new and nutraceutical or health-enhancing properties are even more recent. Further work is sure to advance the use of hydrocolloids beyond modification of the rheology of foods.

Viscosity

Viscosity is probably one of the most widely used properties. In this respect, hydrocolloids are often used in systems where the oil or fat content has been reduced or eliminated through substitution with water. The hydrocolloid thickens water, which, in turn, replaces the fat or oil to give a product with similar properties to the full-fat food. A typical application for this function is reduced-fat salad dressings. In other cases, the thickened water simply adds body, texture and mouthfeel to a food such as table syrups, particularly low-calorie syrups.

Stability

If oil or fat is partially removed from a formulation and is replaced with thickened water, an emulsion is usually formed. Often the function of the hydrocolloid is to stabilise the emulsion, to prevent separation and, in the case of frozen foods, to control ice crystal formation. New technology and new ingredients have been developed specifically to address the problem of ice crystals in frozen foods, but hydrocolloids will continue to play a role. Virtually every ice cream product sold in retail outlets is stabilised with carrageenan, locust bean gum and/or guar gum. Low-fat salad dressings, discussed above, also benefit from emulsion-stabilising properties.

Suspension

If insoluble particles are included in the thickened product then separation and settling should be eliminated or at least minimised. Some hydrocolloids create solutions with a ‘yield point’ that will keep particles immobilised in suspension. Salad dressing is a good example of this and xanthan gum is the typical hydrocolloid to supply this functionality.

Gelation

One of the key texturising aspects of hydrocolloids is the ability to gel and solidify fluid products. For example, in gelled milk desserts, even low levels of carrageenan will form a solid milk gel. Other classic gelling agents are pectin, gelatin and agar. Many others, however, will form a gel under specific conditions. Certain grades of alginates form gels with calcium ions. Xanthan and locust bean gum do not gel individually but together they display synergy and form a strong cohesive gel. Methyl cellulose and hydroxypropylmethyl cellulose are unusual in forming solutions that reversibly thicken or gel when heated. The food industry has a myriad of gelling applications ranging from soft, elastic gels to hard and brittle gels.

Nutritional and nutraceutical

There is already a wide use of some hydrocolloids, arabic and guar gum, for example, as sources of soluble dietary fibre. Much research has been conducted in the nutraceutical benefits of hydrocolloids. Potential benefits range from cholesterol reduction to cancer risk prevention. Their use in weight loss programmes is already widespread and likely to expand further.

Acacia Gum (Gum Arabic)

Acacia gum, also known as gum arabic, is a natural gum exudate obtained from acacia trees in the ‘African sub-Sahelian zone’. The gum has a highly branched compact arabinogalactan structure which gives a low-viscosity solution together with a central protein fraction that provides good emulsification properties. The powder hydrates readily in water and concentrations up to 40–50% can be handled easily. Key food applications include a range of confectionery products, flavoured oil emulsions and capsules and health foods as a source of soluble fibre with prebiotic properties.

Acacia gum is a highly branched arabinogalactan polysaccharide with a high molecular weight developing a low viscosity in water. All molecules of acacia gum contain the same sugars: galactose, arabinose, rhamnose and glucuronic acids (Jurasek et al., 1993) partially neutralised with calcium, potassium, sodium and magnesium salts.

Acacia gum is not considered a thickening hydrocolloid when dissolved in water at low concentrations up to 30%. However, when used in sucrose or sugar-free systems at high levels of dry solids, acacia gum provides a unique texture to confectionery products. Acacia gum is used in a wide range of finished products including moulded candies, jujubes, pastilles with sucrose or polyols, for coated and non-coated chewy products, for sugar-free hard candies and in different tableting processes where binding properties are needed.

Moulded candies

For moulded candies made with sucrose, acacia gum is used at different levels depending upon the texture required. For a hard texture, acacia gum from A. senegal is used alone at high concentrations in the finished confectionery. A typical formula contains 35% acacia gum, 30% sucrose, 25% glucose and about 10% water plus flavouring and colouring. Because of the occasional shortage of acacia gum, modified starch has partially substituted the natural gum. And yet, compared to modified starch, it is recognised that hard candy made with acacia gum lasts longer in the mouth, does not stick to the teeth and provides a unique flavour release.

To produce moulded candies with a softer texture, acacia gum is used with other gelling agents, such as gelatine. The combination of the hard texture from acacia gum with the flexible, soft gel from gelatine gives a wine gum-type texture. A typical formula is based on 15% acacia gum from A. seyal or A. senegal with 5% gelatine, 24% sucrose, 44% glucose, colour and flavour, and the remaining part being 12% moisture.

Chewy confectionery

A chewy product is a slightly whipped, soft confection containing 74% sucrose and glucose, 5% hydrogenated vegetable fat, flavour and acid. Specific textures are obtained by including 1% gelatine for aeration and 1% gum from A. seyal for a long-lasting, cohesive chew. These chewy products can also be used as centres and then sugar coated.

Sugar-free hard candies

Because of the water-binding properties of acacia gum, the addition of a low level of 2–5% of gum in the formulation of a sugar-free hard candy, based on sorbitol, maltitol or mannitol, slightly increases the amount of residual water by 1–3% after cooking and, therefore, decreases the cooking temperature between 5 and 15◦C. Hygroscopicity of the candy is reduced, recrystallisation of polyols is avoided and wrapped sweets are not sticky.

Tableting

Tableting includes different techniques: direct compression, wet granulation and making lozenges which are a type of tablet. Agglomerated acacia gum from A. seyal and A. senegal is used as a binder in these different processes to make food and pharmaceutical products. For direct compression, purified and agglomerated acacia gum is mixed with the other powders having the same mesh size before filling the die. In wet granulation, a solution of acacia gum is added to the powders to make a slurry which is dried and sieved to produce a free-flowing material which is then compressed.

For lozenges, two binders are combined: acacia gum and gelatine are dissolved in water and used to bind the flavoured icing sugar. The paste is then sheeted, cut to shape and dried in an oven. The lozenges are usually flavoured with mint and have an old-fashioned traditional appearance with a rough surface.

Coating and panning

The most popular sugar-free product is polyol-coated chewing gum in which the gum base is used as a carrier for an active principle suitable for tooth health. Sorbitol, maltitol and xylitol can be used for hard coating. Each of these polyols has its own specific behavior in terms of stability, hygroscopicity, cooling effect, crunchiness and sweetness. Xylitol and maltitol are the main polyols used for panning sugar-free chewing gum.

Acacia gum is used in a hard-coating syrup containing about 65% polyol, 3% acacia gum, 1% titanium dioxide and 31% water. This is sprayed onto the surface of the chewing gum centre and then dried by air at 30◦C and 40% humidity. One role of acacia gum is to decrease the crystallization temperature of the polyol in order to apply the syrup at a lower temperature of 65–70◦C so as not to damage the shape of the chewing gum centres. Other benefits of including acacia gum in the coating syrup are improving resistance of the polyol layers and increasing shelf life by decreasing hygroscopicity.

Emulsions

Acacia gum is used as an emulsifier and stabiliser in preparing oil-in-water emulsions. Acacia gum is not considered an actual emulsifier which contains a lipophilic and hydrophilic part in its molecule. Acacia gum is a water-soluble polysaccharide. However, it is possible to give it a hydrophilic–lipophilic balance value. The protein contained in the AGP fraction of the molecule gives surface-active behaviour to the molecule and allows formation of a colloidal film around the oil droplets.

Encapsulation

Encapsulation is a general term that includes many different techniques using different carriers including acacia gum (Risch, 1995). The main reasons for encapsulating an active principle are:

• protection against oxidation, water migration or internal reactions;
• production of a water-dispersible, free-flowing powder;
• production of a controlled release powder;
• reduction of hygroscopicity or reduction of dust pollution.

The encapsulation process is generally classified into two groups:

• Matrix encapsulation. The dispersion of oil droplets inside the matrix in which acacia gum may be used. Such a system allows 10–30% oil content in the carrier. The matrix system is normally produced by spray drying.
• Membrane encapsulation. The formation of membrane around a ‘big’ oil droplet from 20 to 500 μm in size. Such a system allows encapsulation of up to 90–95% oil.

Processes used to form the membrane system are mainly:

• complex coacervation where acacia gum could be involved, or
• extrusion where the membrane could be gelatine, agar or alginate.
• 
  

Agar

Agar is a seaweed hydrocolloid, or phycocolloid, with a long history of use as a gelling, thickening and stabilising food additive. Agar is considered to have been discovered in the mid-seventeenth century in Japan, 200 years before it was introduced to the West.

Agar is widely used for solid culture media in microbiology. Although this application is credited to Robert Koch, it is now accepted that his wife, Angelina, first suggested the use of this material. She had learnt of the use of agar-agar in jams and jellies from a Dutch immigrant returning from Java. This gelling agent melted at higher temperatures than gelatine and enabled the jellies to tolerate the hot and humid climate conditions. Subsequently, Koch and his assistant Hesse perfected the extraction of agar for use in microbiological plates leading to Koch’s discovery of the tuberculosis bacillus and the promotion of agar for microbiological media in general.

Agar has been recognised as a polysaccharide since its introduction into Europe in the middle of the nineteenth century. Initially, it was assigned a linear galactan configuration, but later small quantities of sulphate were identified within the macromolecule. The sulphate content of agar is below 4.5% which is very low compared to that of carrageenans, another group of polysaccharides which are obtained from red seaweeds, Rhodophyceae. Typically, the sulphate content of agar is 1.5–2.5%, in contrast to the ester sulphate content of kappa, iota and lambda carrageenans of 22, 32 and 37%, respectively.

Gelation

The ability to form reversible gels by simply cooling hot, aqueous solutions is the most important property of agar. This gel-forming ability has led to the large number of practical applications where agar is used as a food additive or in other applications in microbiology, biochemistry or molecular biology, as well as in industrial applications.

Agar is insoluble in coldwater but hydrates to form random coils in boilingwater. Gelation depends exclusively on the formation of hydrogen bonds, where the random coils associate to form single helices and double helices. These left-handed threefold helices are stabilised by the presence of water molecules bound inside the double helical cavity and exterior hydroxyl groups allow aggregation of up to 10 000 of these helices to form microdomains of spherical microgels. Following the phase separation process, the microgels aggregate to form gels on cooling below 30–40◦C.

This transition occurs at higher temperatures with increasing agar concentration and decreasing cooling rate. For example, the gelling temperature for Gelidium agar increases from 32 to 38◦C and melting temperature rises from 86 to 89◦C as the concentration increases from 0.5 to 2.0%. The gelation process of agar: random coils associate on cooling to form helices followed by further aggregation of the helices to give the aggregated structures of agar gels.

Food applications

The most important advantages for agar in different food applications derive from the characteristic firm texture and heat tolerance of the gels, stability in acidic conditions and limited reactivity to other food components. The key properties are listed below:

• The large gelling capacity enables agar to be used at very lowconcentrations. The threshold concentration for gelation is 0.2% and levels in food products are typically between 0.5% and 2.0%.
• The large gel hysteresis, the difference between setting and melting temperatures, is much greater than that of any other reversible gelling agent, so that liquid solutions may be held at 40◦C before setting and, once gelled, products remain stable up to 80◦C.
• Agar forms gels over a wide pH range. The neutral polymer chain confers good resistance to acid hydrolysis at the normal pH values found in foods, such as in fruit products. However, agar can be hydrolysed by acid at high temperatures, so for pH values below 5, it is recommended that the pH of the food is lowered just before cooling to form the gel.
• No counter ions are needed for gelation and, hence, there is no characteristic metallic taste in the final products as found with alginates or carrageenan, which makes agar a very good product for delicately flavoured foods. Also, variations in the levels of cations in the food product do not produce a change in the textural properties of the gel.
• In cultured foods, such as yogurt, agar does not inhibit the growth of inoculated bacteria.
• It has good compatibility with other polysaccharides and proteins at normal use levels. This enables agar to give consistent gelled dairy desserts and avoid textural variations which can result from differences in milk quality.
• It does not require a minimum sugar solids level for gelation and so it can be used for low-sugar jams and jellies where reduced sugar levels and/or high intensity sweeteners may be employed. In some cases, high sugar concentrations assist the association of the agar network and gel strength increases.
• Agar now has 350 years of safe use in food. It has a high soluble fibre content that is not metabolised in humans and so it does not add calories to foods.

This combination of benefits results in agar maintaining its position as the hydrocolloid of choice in a number of specific food applications, particularly where firm, short-textured gels with good heat stability and good moisture stabilisation are required:

• Water gels, such as water dessert jellies, vegetable, meat and fish aspics and artificial caviar.
• Confectionery including sweets and candies, fruit jellies, nougat, candy fillings, piping gels, jellies and jams.
• Bakery products, such as icings and glazes for pastries, cakes and doughnuts.

Carrageenan

For centuries, red seaweeds (Rhodophyceae) have been harvested and used as foods in the Far East and Europe. There are many different species of red seaweeds but they all contain naturally occurring polysaccharides that fill the voids within the cellulose structure of the plant. This family of natural polysaccharides includes carrageenan, furcellaran and agar.

Carrageenan extracted from seaweed is not assimilated by the human body, providing only fibre with no nutritional value, but it does provide unique functional characteristics that can be used to gel, thicken and stabilise food products and food systems. Multiple commercial red seaweed species provide a sub-family of carrageenan extracts, with differences in composition and molecular conformation, which lead to a wide spectrum of rheological profiles, gel properties and textures, molecular charge densities and interactions with other gums and proteins.

Carrageenans have a backbone of galactose but differ in the proportion and location of ester sulphate groups and the proportion of 3,6-anhydrogalactose. There are three primary types. Kappa carrageenan and iota carrageenan form thermally reversible gels, which range in texture from firm and brittle to soft and elastic. Lambda carrageenan is non-gelling. Kappa carrageenan interacts synergistically with other gums to modify further the gel texture, for example with polymannans such as locust bean gum and konjac. A specific interaction between kappa carrageenan and kappa casein is widely used to stabilise dairy products.

Water gelling applications

Water dessert gels and cake glazes are some of the more traditional uses of carrageenan. These products are based on the firm, brittle gel properties of kappa carrageenan with the texture modified as necessary for elasticity, cohesiveness and syneresis control using iota carrageenan or other gums, such as locust bean gum or konjac. Recent improvements in the combinations used for these applications have resulted in vegetarian products which have a similar appearance and texture to traditional gelatin products while giving additional 

benefits of fast setting and stability at ambient temperatures.

Gelatine

Through the years, mankind discovered that gelatine is a multifunctional protein molecule, used as an ingredient for the food, pharmaceutical and photographic industries in a wide range of applications.

The photographic industry initiated research on gelatine. The manufacturing processes were also found to have a large impact on gelatine characteristics. Research by Ames (1952) clearly made the distinction between two major manufacturing processes: the acid or A-type process, where the raw materials are pre-treated in acid, and the alkaline or B-type (basic) process, where the raw materials are pre-treated in an alkaline solution.

The basic characteristics of gelatine, such as gel strength, viscosity and isoelectric point (IEP), are mainly determined by which process is used. Gel strength is primarily determined by the proportion of proline and hydroxyproline present in the total amino acids. A high content of these two amino acids indicates a high gelling power. Viscosity is primarily determined by the molecular weight distribution of the gelatine. A high molecular weight is linked to a high viscosity. The alkaline process results in IEP values between 4.5 and 5.5, and the IEP from an acid process can vary from 6 to 9.5. These three characteristics are important in determining the appropriate type of gelatine required for a particular application.

Gelatine is obtained by hydrolysis of the collagen that is contained in animal connective tissues. Commercially, skins or bones of different animal species, such as beef, pork, fish and poultry, form the main raw material for gelatine production. These raw materials are collected from animals approved for human consumption by ante- and post-mortem veterinary inspection. Skins can be frozen if there is a long delay between sourcing and eventual use in the gelatine manufacturing process. Some raw materials, mainly bones, are pre-treated by grinding, degreasing and drying before they are used in gelatine production.

Sugar confectionery

In the early days of sugar confectionery, products were made by skilled craftsman working empirically and the science of sugar confectionery came later. The link with more scientific technology for the production of cough sweets and similar products in the pharmaceutical industry was a big step in understanding the basic aspects of sugar confectionery.

The sugar confectionery market offers a wide variety of products around the world. Cultural differences, habits and eating customs can be completely different, so apparently identical sugar confectionery products can have a completely different taste and texture. A good example is the famous gummy bear.

A German gummy bear, with a hard texture, resembles the appearance of a British jelly baby, which has a very soft texture. These are different from the soft Asian gummy bear which has unusual exotic flavours. This means that there is no standard reference for sugar confectionery, but similar basic principles are applied throughout the industry.

Gelatine is one of the most versatile texturising ingredients for sugar confectionery and is perfectly adaptable to most industrial sugar confectionery processes as a result of a number of features:

• the thermo-reversible gel allows confectionery products to be re-melted for easy recyclingand rework of finished products,
• solutions are low viscosity and easy to deposit without tailing,
• it is a protein with good whipping and emulsifying properties,
• it is fully compatible with many other texturising agents,
• strong water binding prevents the finished product from drying out,
• different mesh sizes from 8 mesh (2.35 mm) up to 60 mesh (0.25 mm) allow combination with other ingredients of the same particle size and processing is easy.

Gelatine sugar confectionery products can be divided into two classes: jelly confectionery and aerated confectionery. Jelly confectionery is characterised by the formation of a gel after depositing into starch and cooling down to ambient temperature. Aerated confectionery is made by the introduction of air into the liquid phase and the amount of air is characteristic of the aerated product.

Jelly confectionery

These products are usually called jellies. Occasionally, they are also referred to as gums, although the name ‘gum’ is generally reserved for products made with modified starch. Sometimes modified starch is combined with gelatine, for example in a so-called wine gum.

Variations in texture will be obtained when using gelatine of different gel strengths at the same concentration. Higher gel strength gelatine gives harder textures. Sugar and glucose syrup are the bulking agents in the recipe and are responsible for the sweet taste and the shelf life of the product. Citric acid is used to lower the pH to about 3.2, which will enhance the typical acid fruit flavour.

The gelatine solution is made in a separate tank. Solutions of gelatine up to maximum 40% can be made; at higher concentrations, part of the gelatine will not be dissolved. Gelatine dissolves rapidly in hot water (80–90◦C) and can be kept for 4–6 h maximum at 50–60◦C without significant degradation.

Aerated confectionery

In aerated confectionery, gelatine gives the air cell wall the required mechanical resistance to avoid deformation. The basic principles and the recipes are very similar to jelly confectionery. The big difference is that air is introduced in the gel mass, which results in a mass with densities that can vary from 1.0 to 0.25. The aeration can be done with a planetary beater or a continuous beater under pressure. It is the final density which determines the texture and type of finished product. Four main types can be defined:

• Chewy candy: density 1.0–0.9
• Aerated jelly candy: density 0.9–0.8
• Deposited marshmallow: 0.55–0.5
• Extruded marshmallow: 0.30–0.25

Gellan Gum

Gellan gum is a fermentation polysaccharide produced by the microorganism Sphingomonas elodea (previously identified as Pseudomonas elodea, but later reclassified). CP Kelco found this organism when it undertook a worldwide screening program to discover interesting and useful new gums that could be made by fermentation. At the time of its discovery, gellan gum was thought to be a ‘universal’ gelling agent. The fact that gellan gum gel textures can range from soft and elastic to hard and brittle is unusual. Adding to this appeal, gellan gum can form gels using both monovalent and divalent cations. Gellan gum can be used at very low use levels, and its gels show excellent thermal stability. With its unique combination of properties, it is easy to understand why gellan gum has high appeal to the food industry.

Dessert gels

In dessert gels, low-acyl gellan gum creates a firm, brittle texture with excellent clarity and heat stability. Because it is highly efficient at forming gels, the gellan gum use level is very low, with typical use levels for dessert gels ranging from 0.15 to 0.35%. Gellan gum also provides particularly good flavor release compared to other gums. However, the brittle texture of low-acyl gels is not typical of common dessert gels. Therefore, low-acyl gellan gum is often blended with other gelling agents to provide softer, more elastic textures. A blend of low-acyl gellan gum with xanthan gum and locust bean gum, targeted for a dessert gel 

texture, is commercially available.

Drinking jellies

Drinking jellies are formulated in a similar way to dessert gels, but lower levels of gelling agents are used to create a very soft gel. There are different types and textures of drinking jellies, depending on how they will be consumed. Some drinking jellies are soft enough to be sucked through a straw. Others will be crushed and squeezed out of a pouch. In many Asian countries, drinking jellies are consumed simply for their interesting textures.

Increasingly, however, gellan gum drinking gels are being formulated to provide enhanced nutrition or nutraceutical benefits. Sports gels are an example of such a product. They are typically packaged in pouches with tear-off tops for consumption while exercising. The gelled contents provide water and nutrients without spilling and the juicy gel bits slide down the throat without leaving a sticky coating.

Pectin

Pectin is a natural constituent of all land plants where, together with cellulose, it plays a key role in the cell wall structure. It comprises a group of polysaccharides rich in galacturonic acid units and, to a lesser extent, various neutral sugars. Pectin extracted from plants has been used as a gelling agent in food for many years. In fact, the invention of using pectin as a gelling agent dates from the 1820s when the Frenchman Henri Braconnot prepared a synthetic jelly with alkali-extracted pectin. However, the first recorded commercial production of pectin extract was in Germany in 1908, after which the process spread to the US, where Robert Douglas obtained a patent in 1913.

The centre of production is currently located in Europe and citrus-producing countries such as Mexico and Brazil. Historically, apple pomace was the major pectin source, but in recent years, an increasing use of citrus peel has taken place. An additional but much less important source of pectin is sugar beet pulp. In recent years, new application opportunities have emerged and pectin is no longer just a gelling agent but also used as a stabiliser and thickener.

Pectin raw materials presently used in industrial processes include apple pomace, citrus peel – comprising lime, lemon and orange – and, to a lesser extent, sugar beet pulp. Citrus peel and apple pomace are by-products of the fruit juice industry and sugar beet pulp of the refined sugar industry. Both wet and dry citrus peels are used for pectin production, whereas mainly dry apple pomace is used. Apple pomace must be dried immediately in order to prevent disintegration into a thick mash from which it is uneconomic to extract pectin.

Gelation

The prime physicochemical property of pectin is its ability to gel and gel strength generally increases with increased molecular weight. HM pectin gels in acidic conditions and in the presence of sugar, whereas LM pectin may gel over a broader pH range and lower concentration of sugar but requires the presence of cations which, when referring to food, is generally calcium. Accordingly, one refers to sugar and calcium gelling, respectively.

Pectin is used as a gelling, thickening and stabilising agent in foods and, to a lesser extent, in pharmaceuticals. Basically, pectin is used to control water in products and help to create the desired texture. The traditional and major application is as a gelling agent in jams and jellies which utilises the ability of HM pectin to form gels at low pH and high sugar levels and the ability of LM pectin to form gels at low sugar levels in the presence of calcium. One of the attractive features is that the pH at which pectin has optimal stability matches the natural pH of fruit preserves.

Confectionery

Pectin fruit jellies are delicate confectionery products with a low water content of around 20%. Recommended pectins are shown in Table 13.8. Normally, the jellies are prepared from combinations of sugar and glucose syrup, flavoured with synthetic flavours and acidified with citric acid. However, the jellies may also contain fruit pulp or juice. Pectin is used in the concentration range 1–3%.

Pectin provides a superior texture and flavour release compared to other gelling agents but may be troublesome to handle during processing due to gel formation at high temperatures (∼70◦C) and because of its sensitivity to pH and soluble solids content. In order to gelHMpectin, acid must be added to reduce the pH from 4.1 to 3.5. The acid must be added as a solution (maximum 50% w/v), otherwise pre-gelation will occur around each acid crystal and make the jelly grainy.

When producing non-fruit flavoured pectin jellies, for flavours such as toffee and peppermint, the pH working range must be 4.0–4.5 as the neutral flavours are more compatible with this range. For this type of application, certain amidated LM pectin types can be used. For aerated confectionery products such as fruitflavoured marshmallows, which have a soluble solids content of approximately 80%, HM pectin confers texture and stabilises the foam in the product. Amidated LM pectin may also be used.

Starch

Chefs or food scientists often make a classic choice when they are developing their latest innovative creation: they know exactly how they are going to delight the consumer and where to source all the ingredients in order to do this. Alternatively, they go to their store shelves and see what they have in stock and work out how to create their latest innovation from the materials at hand. The former is exciting and potentially unlimited where their creativity can produce totally new solutions by combining the latest and best materials. 

The latter has the same potential but it is more about being innovative and creative with existing ingredients and obtaining value by pushing the limits of the functionality of these already-available raw materials. Both of these two routes are the purpose and expected delivery of this chapter on starch, a truly flexible and functional thickening and gelling agent.

Gelling starches

At the other extreme, starches can act as gelling agents. Here, the starch granules have broken down and the amylose and amylopectin chains are free in solution and can associate to form a gel. The type of interactions, chain length and interaction with other ingredients has an effect on the gel properties. There are no discrete starch granules and the free starch polymers are dissolved within the continuous phase.

In hot solutions these polymers have a certain degree of freedom to move around and that allows them to start to interact with each other and to create associative networks. It is, in fact, during the cooling phase that these interactions increase and a stronger gelling network is formed. Care should be taken when these are created. Generally with fairly rapid cooling a gelled network is formed but if the cooling is done more slowly, or if there is a higher concentration of starch or low levels of other ingredients that can interact, a process called retrogradation can occur where the starch polymers align to give small, discrete, insoluble particles. This precipitate is generally undesirable for gelling and thickening but important for other food properties such as the creation of insoluble fibrous starch material.

Starches with a higher amylose content are more likely to gel or retrograde but these processes can be avoided by using the 100% amylopectin starches. This shows the two extremes that can be achieved and takes us to the next important step: how to achieve the required thickening or gelling characteristics.

Confectionery

The confectionery market is at the furthest end of the spectrum in the journey from thickening through to gelling and it is in the area of sugar confectionery that there are the most applications for the gelling properties of starches.Within this category there are a number of subdivisions that need to be considered.

Moulded confections

In moulded confections, the end-product tends to be a relatively clear, highly elastic confection that has been formed by pouring a molten solution into a mould which is then cooled and often dried to reduce the moisture content to the desired level. The gelling properties for this product can be achieved by the use of starches. Success has been achieved with a whole range of starches but it is those with a higher amylose content that have proven most effective. In many cases, the goal has been to replace other hydrocolloids, such as gelatin, to allow vegetarian products to be produced in a cost-effective way.

Initially all the ingredients are blended, heated and hydrated. Here, it is important that the starch dissolves and mixes easily and retains fluidity at the high temperatures used in moulding. This fluid mix is poured into moulds and then it must gel rapidly and give an elastic texture. There is always a balance between fluidity at high temperatures and forming good gels at cooler temperatures. In many cases, a further drying step is required to achieve the moisture content before the product is ready to pack. One of the major issues for these products is to retain stable characteristics throughout the shelf life. By careful selection of starches these gelled products have been developed.

Chewy confections

The same approach is taken for chewable products that need to be gelled and also stretch. Again the higher amylose starches are suitable. After moulding the product is stretched in preparation for the final forming and packing so the product must remain elastic for part of this period. In the final form they should not be too sticky to touch but should easily soften when chewed in the mouth.

Xanthan Gum

Xanthan gum is a high-molecular-weight extracellular polysaccharide secreted by the microorganism Xanthomonas campestris and is produced commercially in a fermentation process. It is soluble in cold water and has a very wide range of applications. It was first discovered in the 1960s and commercialised in the 1970s. With an annual sales volume of approximately 46 000 metric tonnes, xanthan gum applications are split approximately 50/50 between food and non-food. Non-food includes oil field, personal care, pharmaceutical and home care. Typical food applications include sauces and dressings, baked goods, beverages, desserts and ice creams. The total textural ingredients market for food is currently estimated to be worth 2.8 billion US dollars and xanthan gum represents approximately 11% of this.

PRESERVATION OF BIOCHEMICAL METHODS WITH SPECIAL EMPHASIS ON ENZYMES - Food preservation technology

1.0          Food preservation and requirement of bio-chemical control methods

In the production of food, it is crucial to take proper measures for ensuring its safety and stability during the shelf-life. Food preservation is carried out to maintain the quality of raw material and physicochemical properties as well as functional quality of the product whilst providing safe and stable products.

Despite improved manufacturing facilities and implementation of effective process control procedures such as Hazard Analysis and Critical Control Points (HACCP) in the food industries, the number of food borne illnesses has increased. Nowadays consumers favor food with few chemical preservatives. As a result there is increased interest in the preservation through biochemical methods because of their safe association with human foods. Several metabolic products produced by these enzymes have antimicrobial effects, including organic acids, fatty acids, hydrogen peroxide and diacetyl.

2.1          Glucanases

Glucanases are the extra cellular enzymes that break down a glucan, a polysaccharide made of several glucose sub-units. As they perform hydrolysis of the glycosidic bond, they are hydrolases which is secrete by Lysobacter enzymogenes and it is characterized for its propensity to lyse fungi and other micro organisms. Glucanases are capable of degrading the major cell wall components of fungi and oomycetes. L. enzymogenes also produces other factors, such as antibiotics that are antagonistic to the growth of microbes.

Functions 

This enzyme is commonly used as a preservative in wine industry where the marine-derived Williopsis saturnus was found to produce very high killer toxin activity against the pathogenic yeast Metschnikowia bicuspidate which is used in wine industry and isolated from the diseased crab. But the purified β-1,3-glucanase from W. saturnus had no killer toxin activity but could inhibit activity of the toxin produced by the same yeast. In contrast, the toxin produced had no β-1,3-glucanase activity.

Mechanisms of the inhibition may be that the β-1,3-glucanase competed for binding to β-1,3-glucan on the sensitive yeast cell wall with the toxin, causing decrease in the amount of the toxin bound to β-1,3-glucan on the sensitive yeast cell wall and the activity of the toxin against the sensitive yeast cells.

Mode of action

Enzyme systems for yeast cell lysis are usually a mixture of several different enzymes, including one or more beta (1-3) glucanase (lytic and nonlytic), protease, beta (1-6)  glucanase, mannanase and chitinase, which act synergistically for lysing the cell wall.

Enzymatic cell lysis of yeast begins with binding of the lytic protease to the outer mannoprotein layer of the wall. The protease opens up the protein structure, releasing wall proteins and mannans and exposing the glucan surface below.

The glucanase then attacks the inner wall and solubilizes the glucan. In vitro, this enzyme cannot lyse yeast in absence of reducing agents, such as dithiothreitol or b-mercaptoethanol, because the breakage of disulphide bridges between mannose residues and wall proteins is necessary for appropriate exposition of the inner glucan layer. When the combined action of the protease and glucanase has opened a sufficiently large hole in the cell wall, the plasma membrane and its content are extruded as a protoplast.

In osmotic support buffers containing 0.55–1.2 M sucrose or mannitol, the protoplast remains intact, but in dilute buffers it lyses immediately, releasing cytoplasmic proteins and organelles, which may themselves lyse. Meanwhile, proteins released from the wall and the cytoplasm could be subject to attack by product-degrading protease contaminants in the lytic system or in the yeast cells themselves.

 2.2         Phenoloxidases

Phenoloxidases (POs) are a group of copper proteins including tyrosinase, catecholase and laccase. In several insects and crustaceans, antibacterial substances are produced through the PO cascade, participating in the direct killing of invading microorganisms. However, although POs are widely recognised as an integral part of the invertebrate immune defense system.

Functions

Among immune defence mechanisms, phenoloxidases are a group of copper proteins including tyrosinase, catecholase and laccase, which are the rate limiting enzymes in melanisation, and play an important role in immune defence mechanisms in invertebrates. In these organisms, phenoloxidases exist as an inactive form, prophenoloxidase.

Mode of action

Pathogen associated molecular patterns (PAMPs), such as peptidoglycans or lipopolysaccharides from bacteria, or β-1,3-glucans from fungi, are recognized by pattern-recognition receptors (PRRs). This will trigger the activation of a cascade of serine proteases that activates polyphenol-activating enzymes (PPAs), and therefore, the conversion of the pro-enzyme prophenoloxidase into phenoloxidase.

The three types of phenoloxidases can oxidise o-diphenols, such as L-3,4-dihydroxyphenylalanine (L-DOPA; catecholase activity). However, among these three enzymes, only tyrosinases can hydroxylate monophenols, such as L-tyrosine (monophenoloxidase activity), and only laccases can oxidise m- and p-diphenols, or aromatic compounds containing amine groups, such as p-phenylenediamine (PPD; laccase activity).

Different roles have been attributed to phenoloxidases, especially in haemolymphatic immune defence mechanisms, and phenoloxidase-generated reactive compounds are known to contribute to the destruction of microbial cells.

2.3          Lactoferrin

Lactoferrin is a non-haem iron binding protein that is part of the transferring protein family, along with serum transferrin, ovotransferrin, melanotransferrin and the inhibitor of carbonic anhydrase, whose function is to transport irons to blood serum.

Some of the numerous properties of lactoferrin, related to its protective functions, can be attributed to its iron binding activity, whereas other properties of lactoferrin are independent.

There are three forms of lactoferrin depending on its iron saturation; apolacto ferrin (iron free), monoferric form (one ferric ion) and hololactoferrin (binding two Fe³⁺ ions). The tertiary structure of hololactoferrin and apolactoferrin is different.

Four amino acid residues are very important for iron binding (histidine, twice tyrosine and aspartic acid), while an arginine chain is responsible for binding the carbonate ion.

Antibacterial activity and mechanisms of action of lactoferrin

The effect of lactoferrin was demonstrated against many bacteria, such as B.subtilis , clostridium spp, micrococcus sp, etc. which can attach themselves to the host cell. Lactoferrin has been also shown to exert anti-microbial activity against some yeasts and fungi such as C.albicans, C.krusei, etc.

Bacteriostatic activity

Lactoferrin ability to bind free iron can inhibit growth of many species of bacteria (and fungi). A lack of iron inhibits the growth of iron-dependent bacteria such as E.coli. In contrast lactoferrin may serve as iron donor, and in this manner, support the growth of some bacteria with lower iron demands such as Lactobacillus spp or Bifido bacterium spp. generally considered as beneficial.

                Bacteriocidal activity

Independent from iron binding and involving the basic N-terminal region of lactoferrin. Lactoferrin can disrupt or possibly even penetrate bacterial cell membranes, and that the isolated N-terminal basic peptides, named lacto ferricins, were more potent than the intact protein.

                Additional antibacterial activities

Biofilm formation, which represents a colonial organization of bacterial cells, is a well studied phenomenon where bacteria also become highly resistant to host cell defense mechanisms and antibiotic treatment. Lactoferrin play an important role in the innate immunity by blocking the biofilm development by Ps. Auruginosa. At concentrations lower than those killing or preventing the growth and with iron chelating activity, lactoferrin stimulates twitching, a specialized form of surface motility, causing the bacteria to wander across the surface instead of forming clusters of biofilm.

                Antifungal activity

Lactoferrin shows a significant antifungal activity by its ability to bind and sequester environmental ion. And the lactoferrin can kill some fungus by altering the permeability of the cell surface, as it does with bacteria.

Applications of lactoferrin in industry

Lactoferrin is already used in a wide range of products including infant formulae, sport and functional foods.

• Milk based infant formulae – improved resistance against pathogens and oro-gastro-intestinal microflora, anti oxidant
• Yoghurt  – improved resistance against pathogens, anti infection and oro-gastro-intestinal microflora, anti oxidant
• Health supplement – aid in iron absorption. Eg :- for pregnant women, immune aid

2.4          glutathione peroxidase

All milks contain a certain amount of somatic cells represented by polymorphonuclear cells (PMN), lymphocytes and macrophages. In bacterial infection and other inflammation processes affecting the mammary tissue, the number of somatic cells in milk increases, especially the PMN level. During mastitis, PMN cells migrate from the peripheral blood into milk, through the mammary epithelium. In many countries, somatic cell count (SCC) is used as an indicator for the hygienic milk quality. An increased SCC in a bulk tank milk indicates that a significant proportion of milk originates from mastitis cows.

More than 140 different microorganisms are recognized to cause mastitis. They are classified into four different groups: contagious, environmental, opportunistic and others. Most mammary gland infections are caused by only a few types of bacteria, including streptococci (Streptococcus agalactiae), staphylococci (Staphylococcus aureus) and coliforms (Corynebacterium bovis).

Functions

Glutathione-peroxidase (GPx) is widespread in the cytoplasm of animal cells. The function of this enzyme is to protect cells against the damaging effects of peroxides, as part of an antioxidant enzymatic system. Milk contains low levels of GPx, more than 90% being represented by extra cellular form. The function of this enzyme in milk is not yet fully known, it is the only known enzyme that fixes 30% of total selenium (Se), an important element of diet. It is also known that milk GPx varies according to species and diet.

Mode of action

When bacteria invade and colonize the mammary gland, macrophages respond by initiating the inflammatory response, attracting polymorphonuclear (PMN) cells in milk to kill bacteria. More than 90% of somatic cells found in infected glands are neutrophils (PMN). Antibacterial activity of neutrophils is mediated via reactive oxygen species (ROS).

Glutathione peroxidase is an antioxidant enzymes in milk, it catalyses the reduction of different peroxides aided by glutathione or other reducing substrates. The average value for GPx activity in normal milk was 23 U.ml^-1. Adding glutathione peroxidase will increase the activity of GPx and preserve the milk in a more effective way by great activity.

2.5          lacto peroxidase

Refrigeration is the most commonly used method to stop or retard the deterioration of milk on its way from the farm to the dairy industry. The lactoperoxidase system (LPS) has been introduced as an alternating way of preserving milk.

The lactoperoxidase system

It consists of lactoperoxidase (LS) and two substrates; thio cyanate ions (SCN^-) and hydrogen peroxide (H₂O₂).

                Lactoperoxidase

Lactoperoxidase is a glycoprotein consisting of a single peptide chain containing 612 amino acid residues. This enzyme is an oxidoreductase and catalyses the oxidation of thiocyanate at the expense of hydrogen peroxide to generate intermediate products with antimicrobial properties against bacteria, fungi and viruses.

                Thiocyanate ion

Thiocyanate ions are present in mammary, salivary and thyroid glands and their secretions, in organs such as the stomach, kidney and in fluids such as synovial, lerebral, cervical, and spinal fluids.

                Hydrogen peroxide

Hydrogen peroxide is not normally detected in raw milk, but it may be generated endogenously, for example, by polymorphoneuclear leucocytes in the process of phagocytosis, in addition many lactobacilli, lactococci, and streptococci produce sufficient hydrogen peroxide under aerobic conditions, to activate LPS.

Antibacterial activity of lactoperoxidase

The oxidation of the thio groups (-SH) of enzymes and proteins is of crucial Importance in the bacteriostatic and/or bacteriocidal effect of the LPS; the structure damage of cytoplasmic membranes by the oxidation of –SH groups results in a leakage of potassium ions, amino acid and peptide into the medium, thus the uptake of glucose, amino acids, purines, pyrinidines in the cell and synthesis of proteins. DNA and RNS are also inhibited.

On the other hand, the anti microbial activity of LPS can be inhibitedby reducing agents containing –SH groups, such as cysteine, glutathione, mercapto-ethanol, dithiothreitol, and sodium hydro sulphite either by direct binding to the haem group or by scavenging OSCN^-.

3.0          References

• Application of Alternative Food-Preservation Technologies to Enhance Food Safety and Stability by Antonio Bevilacqua et al.
• Enzymatic lysis of microbial cells by Oriana Salazar and  Juan A. Asenjo
• Enzyme-Assisted Processing Increases Antimicrobial and Antioxidant Activity of Bilberry by RIITTA PUUPPONEN-PIMIA et al.
• Food preservation by Nicholas J.Russel and Grahame V.Gould
• http://prof.dr.semih.otles.tripod.com/enzymesused/theroleof/theroleof.htm
• http://stalischem.wordpress.com/2010/04/25/major-enzyme-applications-in-food-industry/
• http://www.deepdyve.com/lp/elsevier/xylan-chitosan-conjugate-a-potential-food-preservative-Aqig5Q0HK2
• http://www.hindawi.com/journals/er/2010/862537/tab2/
• Involvement of -Glucans in the Wide-Spectrum Antimicrobial Activity of Williopsis saturnus var. mrakii MUCL 41968 Killer Toxin by Cyril Guyard et al.
• Mutagenesis of β-1,3-Glucanase Genes in Lysobacter enzymogenes Strain C3 Results in Reduced Biological Control Activity Toward Bipolaris Leaf Spot of Tall Fescue and Pythium Damping-Off of Sugar Beet by Jeffrey D. Palumbo et al.
• New Methods of Food Preservation By Grahame W. Gould
• Preservation and fermentation: past, present and future by R. Paul Ross et al.

PRESERVATION OF FOOD BY IRRADIATION

1.      Introduction

Irradiation of food is the use of ionizing radiations from radioactive isotopes of cobalt or cesium or from accelerators that produce controlled amounts of beta rays or x-rays on food. The food does not become radioactive. Irradiation can be used: to destroy insects and parasites in grains, dried beans, dried fruits and vegetables, and meat and seafood; to inhibit sprouting in crops such as potatoes and onions; to delay ripening of fresh fruits and vegetables; and to decrease the numbers of microorganisms in foods. Hence, the incidence of food borne illness and disease can be decreased and the shelf life of food can be extended.

2.      History

The discovery of x-rays by W.K. Roentgen in 1895 and the discovery of radioactive substances by H. Becquerel in 1896 led to intense research of the biological effects of these "radiations". Initially, most of the irradiations made use of x-rays, which are produced when electrons from an electron accelerator are stopped in materials.

These early investigations laid the foundation for food irradiation. Ionizing radiation was found to be lethal to living organisms soon after its discovery. The use of this lethality to control spoilage and other organisms that contaminate foods was demonstrated in the early decades of the 20th century. However, no commercial development of this use occurred then, due to the inability to obtain ionizing radiation in quantities needed and at costs that could be afforded.

In the mid 1940s, the interest in food irradiation was renewed when it was suggested that electron accelerators could be used to preserve food. However, the accelerators in those days were rather costly and too unreliable for industrial application. Early research in the late 1940s and early 1950s investigated the potential of 5 different types of radiation (ultraviolet light, x-rays, electrons, neutrons, and alpha particles) for food preservation.

Researchers concluded at that time that only cathode ray radiation (electrons) had the necessary characteristics of efficiency, safety, and practicality. They considered x-rays to be impractical because of the very low conversion efficiency from electron to x-ray that was possible at that time.

Ultraviolet light and alpha particles were considered to be impractical because of their limited ability to penetrate matter. Neutrons exhibited great penetration and were very effective in the destruction or inactivation of bacteria, but were considered inappropriate for use because of the potential for inducing radioactivity in food.

In the 1940s, sources of proper kinds of ionizing radiation became available. The first sources were machines that produced high energy electron beams of up to 24 million electron volts. This energy was sufficient to penetrate and sterilize a 6-inch No. 10 can of food when electron beams were "fired" from both sides of the can. Also in this same decade, man-made radio nuclides such as Cobalt-60 and Cesium-137 (which in their radioactive decay emit gamma rays) became available through the development of atomic energy. The availability of these sources stimulated research in food irradiation aimed at the development of a commercial process.

3.      Ionizing Radiation Used For Food Irradiation

Energy exists in the form of waves and is defined by its wavelength. As the wavelength gets shorter, the energy of the wave increases. Electric power, radio and television, microwaves (radar) and light have longer wavelengths and lower energies. They cause molecules to move but cannot structurally change the atoms in those molecules.

Ionizing radiation (gamma rays, x-rays) has a very short wavelength and higher energy, high enough to change atoms by knocking off an electron from them to form an ion, but not high enough to split atoms and cause exposed sources to become radioactive. Therefore, the sources of radiation allowed for food processing (Cobalt-60, Cesium-137, accelerated electrons and x-rays) cannot make food radioactive.

3.1   Electron beam irradiation.

High-voltage electron beams (accelerated electrons) generated from linear accelerators are an alternative to radioisotope generators. They lack the penetration depth of gamma irradiation (about 0.5 cm.) per 1,000,000 electron volts (MeV) of energy, however, they require much shorter exposure times (seconds vs. hours for gamma irradiation) to be effective.

Electron beam irradiation is currently being used to disinfest grain at 1.4 MeV at a grain loading facility, and to pasteurize frozen mechanically separated meat products with 10 MeV at a processing facility.

3.2   X-rays

X-rays generated when electrons from an electron beam bombard a heavy metal target such as tungsten, have a greater penetration depth but are less desirable because of the low energy conversion efficiency of electrons to x-rays.

3.3   Gamma rays

Gamma rays used for irradiation processing of food are radioactive fission products of Cobalt-60 and Cesium-137. Gamma rays have good penetration, as do x-rays. With "cross-firing", they can easily deliver a uniform (less than 25% over dose) of energy to a 6-inch No. 10 can of food.

Cobalt-60 is not a waste product from the nuclear industry. It is specifically manufactured for use in radiotherapy, sterilization of medical products, and the irradiation of food. Cesium-137 is one of the fission products contained in used fuel rods. It must be extracted in reprocessing plants before it can be used as a radiation source. Currently, almost all radiation facilities in the world use Cobalt-60 rather than Cesium-137.

3.4   Comparison of gamma rays, x-rays and cathode rays.

Gamma rays, x-rays, and electron beams are equally effective in sterilization for equal quantities of energy absorbed. The greatest drawback at present to the use of x-rays in food preservation is the low efficiency and consequent high cost of their production. For this reason, most research has concentrated on the use of gamma photons and electron beams.

Gamma rays have a maximum of 10 to 25% utilization efficiency, while the maximal efficiency of electrons from electron beam generators ranges between 40 and 80% (depending on the shape of the irradiated material. Radioactive sources of gamma rays (Cobalt-60 or Cesium-137) decay steadily and hence weaken with time, which is another cost. They must be constantly replenished.

The use of electrons from electron beam generators presents fewer health problems than the use of gamma rays, since electron beams are directional and less penetrating, can be turned off for repair or maintenance work, and present no hazard of radioactive materials after a fire, explosion, or other catastrophe. Gamma rays are emitted in all directions, are penetrating, are continuously emitted, and come from radioactive sources. Gamma rays require more shielding to protect workers.

The one overriding requirement for an energy source to be employed in food irradiation is that the energy levels must be below those that could possibly cause the food to become radioactive. After that requirement is met, sources are considered on the basis of their practical and economic feasibility. Machine sources must produce radiation with relatively simple technology. Isotopes must be sufficiently long lived and emit penetrating radiation.

Figure 1.1 Irradiation unit

4.        Effects on Microorganisms

Lethality due to ionizing radiation, as proposed by the target theory, occurs when the irradiated microorganisms are destroyed by the passage of an ionizing particle or quantum of energy through, or in close proximity to, a sensitive portion of the cell. This direct "hit" on the target causes ionization in this sensitive region of the organism or cell and subsequent death.

It is also assumed that much of the germicidal effect results from the ionization of the surroundings, especially water, to yield free radicals, some of which may be oxidizing or reducing and therefore helpful in the destruction of the organisms. This effect is reduced if food is irradiated in the frozen state. Irradiation may also cause mutations in the organisms present.

Bacterial spores are more resistant to ionizing radiation than are vegetative cells. Gram-positive bacteria are more resistant than gram-negative bacteria. The resistance of yeasts and molds varies considerably, but some are more resistant than most bacteria.

The bactericidal efficacy of a given dose of irradiation depends on the following:

• The kind and species of the organism. [Approximate Killing Doses of Ionizing Radiations in Kilorays (kGy) ]
• The numbers of organisms (or spores) originally present. The more organisms there are the less effective a given dose will be.
• The composition of the food. Some constituents [e.g., proteins, catalase, and reducing substances (nitrites, sulfites, and sulfhydryl compounds)] may be protective. Compounds that combine with the -SH groups would be sensitizing.
• The presence or absence of oxygen. The effect of free oxygen varies with the organism, ranging from no effect to sensitization of the organism. Undesirable "side reactions" are likely to be intensified in the presence of oxygen and to be less frequent in a vacuum.
• The physical state of the food during irradiation. Both moisture content and temperature affect different organisms in different ways.
• The condition of the organisms. Age, temperature of growth and sporulation, and state (vegetative or spore) may affect the sensitivity of the organisms.
• 
  

1.4.1      Approximate killing doses of ionizing radiations in Kilograys (kGy)

Organism	Approximate lethal dose (kGy)
Insects	0.22 to 0.93
Viruses	10 teo 40
Yeasts (fermentative)	4 to 9
Yeasts (film)	3.7 to 18
Molds (with spores	1.3 to 11
Bacteria (cells of pathogens):
Mycobacterium tuberculosis        Staphylococcus aureus         Cornybacterium diphtheriae         Salmonella spp.	1.4 1.4 to 7.0 4.2 3.7 to 4.8
Gram-negative:
Escherichia coli        Pseudomonas aeruginosa        Pseudomonas fluorescens         Enterobacter aerogenes	1.0 to 2.3 1.6 to 2.3 1.2 to 2.3 1.4 to 1.8
Gram-positive:
Lactobacillus spp.          Streptococcus faecalis          Leuconostoc dextranicum          Sarcina lutea	0.23 to 0.38 1.7 to 8.8 0.9 3.7
Bacterial spores:
Bacillus subtillus         Bacillus coagulans         Clostridium botulinum (A)         Clostridium botulinum (E)         Clostridium perfringens         Putrefactive anaerobe 3679         Bacillus stearothermophilus	12 to 18 10 19 to 37 15 to 18 3.1 23 to 50 10 to 17

Table 1.1              Approximate killing doses of ionizing radiations in Kilograys (kGy)

5.        uses of food irradiation

Ionizing radiation can be used to process food. Its effect on the food is dependent on the dose level (amount) of irradiation to which the food has been subjected. High-dose levels of irradiation (20 to more than 70 kGy) can be used to sterilize foods by eliminating all vegetative microorganisms and spores in the food. Very low doses of irradiation (less than 0.1 kGy) can be used to inhibit sprouting in potatoes, onions and garlic.

Low doses have also been shown to be as effective as pesticide fumigants for deinfesting grain products prior to shipment and storage, and for reducing microbial and insect contamination on fresh fruits and vegetables. For example, grapefruit grown in Mexico, Central America, and South America frequently are infested with larvae of the Mexican fruit fly, Anastrepha ludens. To prevent entry of this insect into the United States, grapefruits must be quarantined and treated with ethylene dibromide.

A study reported that 20 grays for 0.25, 0.5, 1.0, or 100 minutes reduced adult emergence of Mexican fruit flies from larvae by more than 99%. Therefore, once a quarantine security treatment for the Mexican fruit fly is established, a low irradiation dose rate can be used to reduce adult emergence and should impart little damage to grapefruit peel tissue.

Not all fresh produce is suitable for irradiation. The shelf life of mushrooms, potatoes, tomatoes, onions, mangoes, papayas, bananas, apricots, strawberries, and figs can be extended with low-dose irradiation with no loss in quality. However, the quality of some foods (some citrus fruits, avocados, pears, cantaloupes, and plums) is actually degraded by irradiation.

Pasteurizing doses of irradiation can kill or reduce the populations of both food spoilage and pathogenic microorganisms in food. For example, Salmonella spp. and Campylobacter jejuni can be eliminated from poultry, and trichinae from pork.

6.        Applications of food irradiation

Type of Food	Radiation Dose in kGy	Effect of Treatment
Meat, poultry, fish, shellfish, some vegetables, baked goods, prepared foods	20 to 71	Sterilization. Treated products can be stored at room temperature without spoilage. Treated products are safe for hospital patients who require microbiologically sterile diets.
Spices and other seasonings	Up to a maximum of 30	Reduces number of microorganisms and insects. Replaces chemicals used for this purpose.
Meat, poultry, fish	0.1 to 10	Delays spoilage by reducing the number of microorganisms in the fresh, refrigerated product. Kills some types of food poisoning bacteria and renders harmless disease-causing parasites (e.g., trichinae).
Strawberries and some other fruits	1 to 5	Extends shelf life by delaying mold growth.
Grain, fruit, vegetables, and other foods subject to insect infestation	0.1 to 2	Kills insects or prevents them from reproducing. Could partially replace post-harvest fumigants used for this purpose.
Bananas, avocados, mangoes, papayas, guavas, and certain other non-citrus fruits	1.0 maximum	Delays ripening.
Potatoes, onions, garlic, ginger	0.05 to 0.15	Inhibits sprouting.
Grain, dehydrated vegetables, other foods	Various doses	Desirable changes (e.g., reduced rehydration times).

Table 1.2              Applications of food irradiation

7.         Effect of Ionizing Radiation on Nutrients in Food

When foods are exposed to ionizing radiation under conditions envisioned for commercial application, no significant impairment in the nutritional quality of protein, lipid and carbohydrate constituents was observed. Irradiation is no more destructive to vitamins than other food preservation methods.

It was noted that there were small losses of vitamin E and thiamin. Thiamin in pork is not significantly affected by the FDA-approved maximum radiation dose to control Trichinella, but at larger doses, it is significantly affected. Protection of nutrients is improved by holding the food at low temperature during irradiation and by reducing or excluding free oxygen from the radiation environment. This is accomplished by irradiating vacuum-packaged foods at temperatures below 0°C (32°F).

The effect of irradiation on retention of vitamin E (alpha tocopherol) in chicken breasts was determined when the chicken breasts were irradiated in air with a Cesium-137 source at 0, 1, 3, 5.6, and 10 kGy at 0° to 2°C (32.0° to 35.6°F). The fresh muscle tissue was saponified and the total tocopherols were isolated and quantitated using normal phase high performance liquid chromatography with a fluorescence detector.

Gamma irradiation of the chicken resulted in a decrease in alpha tocopherol with increasing dose. At 3 kGy and 2°C, the radiation level approved by the FDA to process poultry, there was a 6% reduction in alpha tocopherol level. No significant changes were observed for gamma tocopherol.

Free radical scavengers were tested for their ability to reduce the loss of thiamin and riboflavin in buffered solutions and in pork during gamma irradiation. In aqueous solution, the tested compounds were twice as effective for the protection of riboflavin as for the protection of thiamin. The presence of nitrous oxide doubled the rates of loss for thiamin and riboflavin in solution, indicating a predominance of reactions with hydroxyl radicals.

In buffered solutions, niacin was not affected by gamma radiation unless either thiamin or riboflavin was present, in which case, the niacin was destroyed rather than the other vitamin. Ascorbate, cysteine, and quinoid reductants were demonstrated to be naturally present in sufficient quantities to account for the lower rates of loss of thiamin and riboflavin observed during irradiation of pork meat, as compared to irradiation in buffered solution.

A study was made of thiamin content of the skeletal muscles and livers of pork, chicken, and beef after gamma irradiation. Gamma irradiation from a Cesium-137 source was used to irradiate the samples with doses of 0, 1.5, 3, 6, and 10 kGy at 2°C (35.6°F). Samples were also titrated with dichlorophenoindophenol to determine the reducing capacity of the tissue. The rate of loss of thiamin upon irradiation was found to be about 3 times as fast in skeletal muscle as in liver and to be a function of the reducing capacity of the tissues, the loss decreasing with increasing reductant titer. For the same amount of thiamin loss, liver could be irradiated to 3 times the dose as could muscle.

8.        Advantages and disadvantages of food irradiation

Benefits of food irradiation are,

• Disease causing germs are reduced or eliminated.
• The nutritional value of food is preserved.
• Decreased incident of food borne illness.
• Reduced spoilage in global food supply.
• Increased level of quality assurance in international trade of food products.

Disadvantages of food irradiation are,

• It reduces the content of several key nutrients such as Vitamin E (~15-30 %), Thiamin (~10-25%), Vitamin C (5-15%), Riboflavin (~7-10%), Pyridoxine (~10-20%), Vitamin B12 (~15-20%). Other nutrients are also affected however the results are less consistent.
• It creates radiolytic products with unknown short term or long term safety effects.
• Some of the organoleptic properties are affected especially for herbs, spices, essential oils.
• Formation of cholesterol oxides and fatty acid epoxidation and other oxidation products (aldehydes, esters, ketones etc.) posing safety concerns.
• Aggregation of certain proteins has been found for high protein commodities.
• The method is 90-95%% effective in killing microorganisms. The remaining 5-10% remain unaffected and may proliferate thus negating the irradiation steps. The methods can result in 95-100% effectiveness but will substantially affect the quality of the food item (taste, nutrient contents, radiolytic products,denaturing proteins, fatty acids etc.).
• Is ineffective against viruses.

9.        references

• http://www.hi-tm.com
• http://www.physics.isu.edu/radinf/food.htm
• http://uw-food-irradiation.engr.wisc.edu/materials/food_irradiation/sld021.htm
• Food preservation by irradiation by J.van Kooij