Food Safety and Technology Department PDF

MST:5164 Food Safety and Technology Department Faculty of Veterinary Medicine Beni-Suef University Table Eggs, Edible Fats and Oils Safety and Technology By Staff Member 2024...

MST:5164 Food Safety and Technology Department Faculty of Veterinary Medicine Beni-Suef University Table Eggs, Edible Fats and Oils Safety and Technology By Staff Member 2024 1  MST:5164 Edible fats and oils Oils and fats are important parts of the human diet and more than 90 % of the world production from vegetable, animal and marine sources is used as food or as ingredient in food products. Oils and fats serve as a rich source of dietary energy. They contain certain fatty acid components which are essential nutrients and their of unctional textural characteristics contribute to the flavour and platability of many natural and prepared foods. Fats or lipids are esters of glycerol and fatty acids are mostly long, straight, hydrocarbon chains, with varying degrees of hydrogen saturation of the carbon atoms, having a carboxyl group linked to one of the end carbon atoms. They are usually triglyceride in which one molecule of glycerol combined with three molecules of fatty acids with liberation of three molecules of water e.g. butyrin, palmetine. C3H5 (OH)3 + 3 C16H32O2 Tripalmetine Also, all fatty oils contain smaller amount of free fatty acids and small amount of unsaponifible matter as (hydrocarbons, sterols, colouring matter and dissolved impurities). If the fatty oils are liquid at ordinary temperature, it is termed oil, while if it is solid it is termed fat. Coconut oil (in tropical countries) it called oil and in European countries it called fat. Classification of fatty oils: 2  MST:5164 1. Crude oils or fats Which were obtained from oil seeds or from animals in their natural state without any treatment Products prepared from crud oils by a 2. Refined oils refining process (deodorization, decolorization and bletching) 3. Processed oils or fats As hydrogenated oils or margarine Hydrogenation of oils: This process performed to produce solid fat from oils by treatment of oils with hydrogen in presence of heavy metal as catalyst (Nicle) by which the saturation of fatty acid with hydrogen happened. Catalyst C3H5 (C17H33COO)3 + 3 H2 C3H5 (C17H35COO)3 This process gives the new product (solid fat) stability against oxidative spoilage. Grouping of oils and fats: It is grouped according to resemblance of chemical and physical properties: Olive oil group Include Olive oil, Peanut oil and Almond oil Cotton seed oil group Include Cotton seed oil and Sesame oil Line seed oil group Include Line seed oil and Sunflower oil Tallow group Include Beef fat, Butterfat, Matton fat and Lard fat General properties of fats & oils: 3  MST:5164 (1) Pure fats and oils are tasteless and odorless, the usual taste or odor of some oils are due to presence of impurities. (2) They are insoluble in water but soluble in fat solvents (ether, carbon tetrachloride, carbon bisulphide and hot alcohol dissolve considerable amount of them) (3) If acted upon by oxygen of air in presence of light and moisture free fatty acids are liberated with alteration of taste and odor. It is termed rancidity and give disagreeable odor and acrid taste. Methods of analysis: (Examination of Edible fats and oils) Sampling: All sampling bottles and jars must be dry, clean and glass stoppered. Samples should be protected from light. From oils: Samples must be taken from each oil tank separately and must be homogenized if not. Three samples should be taken at three different levels. From fats: Samples should be taken from different parts of the bulk of fat. Preparation of sample: If the sample is oil it must be thoroughly mixed before examination. 4  MST:5164 If the sample is solid fat must be melted at low temperature as possible and then mixed. The sample must be clear and clean free from visible dirts and suspended matter to be fit for examination. If turbidity is noticed due to suspended matter or moisture, samples are filtered tell clear bright color is obtained. Object of analysis: To determine the purity of a given sample of an oil or fat, the analysis depends upon the determination of certain analytical constants. Analytical constants: They are physical and chemical tests, which give a certain values specific for each oil or fat. The analytical constant varies from one oil or fat to another due to differences in the fatty acids, which are present. Analytical constants Physical Chemical 1. Specific gravity 1. Iodine number 2. Refractive index 2. Saponification number 3. Melting point 3. Reichert-Meissel and Polenske number Physical constants (1) Determination of specific gravity of fats or oils: 5  MST:5164 It must be determined on fresh and pure oil or fat not affected by rancidity or any deterioration and must be determined at a standard temperature 15.5 0C by using Westphal’s balance. If the test is done at room temperature the result should be corrected as the following: Sp. Gravity at 15.5 0C = K x Sp While Sp = Specific gravity at room temperature. K = Factor varying with the temperature. Test: 1. Placing the sample (oil or melted fat) in a cylinder containing dilute alcohol, by continuous addition of alcohol or water. 2. The mixture may be so adjusted that the oil globules neither rise nor fall, but remain in equilibrium in the liquid which should be at 15.5 0C. 3. The specific gravity of liquid is taken and is obviously the same as that of the oil or fat. Specific gravities of some edible oils and fats Edible fat Sp. Gravity Beef-tallow 0.947 Butter fat 0.936 Coconut oil 0.926 Cotton seed oil 0.922 Sesame oil 0.922 Hydrogenated Cotton seed oil 0.88 - 0.87 (2) Determination of Refractive Index: 6  MST:5164 This made by using Abbe-Refractometer at the standard temperature 20 0C for oils and 40 0C for solid fat and this adjusted by passing warm water at the desirable degree of temperature around the prism recording the temperature degree by the thermometer present. Ref. I. At 20 0C (of oils) = Ref.I. obtained + 0.00035 Ref. I. At 40 0C (of fat ) = Ref.I. obtained + 0.00036 Ref. Index of some edible oils Fat Ref. Index Butter fat 1.447 Tallow fat 1.451 Hydrogenated cotton seed oil 1.457 – 1.460 Sesame oil 1.465 Cotton seed oil 1.471 (3) Determination of Melting point: By using capillary tube method 7  MST:5164 In capillary tubes (about 5 cm. Long and 1.4 mm wide) of thin glass wall. A small quantity of the melted fat is introduced in a capillary tube, the amount of fat in the tube is adjusted by placing a piece of filter paper underneath till reaching the desirable highest. The tubes are placed in ice for few hours and attach the tubes thus prepared to a delicate thermometer graduated to tenth of degrees using a small rubber ring. The tubes and thermometer put in water in a beaker of 50 cc capacity, which in turn rests on the neck of a round bottomed flask containing water. The water is heated gradually at a rate not exceeding 2 0C per minute until the fat melts. The temperature at which the fat becomes transparent liquid is taken as the melting point. Chemical constants of fats and oils (1) Determination of Iodine number (Iodine value): It is No. of grams of iodine absorbed by 100 grams of the oil or fat Test: 8  MST:5164 1. In a 250 ml white clean dry glass stoppered bottle weigh 0.5 gm. of the oil or fat. 2. Add 10 ml. of chloroform to dissolve the sample. 3. By using safety bulb pipette 25 ml capacity add 25 ml of standard iodine solution and allow to stand for 30 minutes in dark place with occasional shaking. 4. Add 10 ml of 15 % potassium iodide solution and shake thoroughly and then add 50 ml of freshly boiled and cooled water. 5. Titrate against a standard N/10 sodium thiosulphate solution until faint yellow color appears. 6. Add few mls of soluble starch solution (1-2 cc) and repeat the titration till the blue color completely disappears. Record the number of ml of thiosulphate taken as (b) 7. Make a blank determination, using the same procedure described previously (without any fat or oil) and record the No. of ml of sod. thiosulphate exhausted as (a) (a – b) x 0.01269 Iodine No. = X 100 Weight of fat used Iodine No. of some edible oils and fats Fat Iodine value Butter fat 32 – 40 Beef tallow 34 – 44 Hydrogenated cotton seed oil 60 – 80 Cotton seed oil 101- 115 9  MST:5164 Sesame oil 101- 115 Formation of yellow color 1 ml hot starch 0.5% gives blue color End point disappearance of blue color 10  MST:5164 Determine the iodine number of a given oil sample 1- Technique 2- Result 3- Judgement 4-Significance Date: Supervisor signature:  MST:5164 11 (2) Saponification number: It is the number of milligrams of KOH which are required to saponify completely 1 gm of fat or oil. 12  MST:5164 Content of the flask + 1 ml ph. ph. Saponification No. of some edible fat and oils Fat Saponification No. Sesame oil 188 – 195 13  MST:5164 Hydrogenated cotton seed oil 190 – 199 Beef tallow 193 – 199 Butter fat 224 – 230 Coconut oil 252 - 255 (3) Reichert-Meissel Number and Polenske Number (i) Reichert-Meissel Number {R.M.N.} It is the No. of ml of N/10 NaOH (alkali) required to neutralize the water soluble volatile fatty acids distilled from 5 gm of fat or oil. (ii) Polenske Number {P.N.} It is the No. of ml of N/10 NaOH (alkali) required to neutralize the water insoluble volatile fatty acids distilled from 5 gm of fat or oil. Fatty acids Volatile Non-volatile Water soluble water insoluble - Butyric - Lauric - Caproic - Myrestic - Caprylic - Capric These acids are These acids are In high % in In high % in Butterfat Coconut oil Therefore, with the aid of determination of R.M.N. and P.N., it is easy to differentiate between butter and coconut 14  MST:5164 oil as well as margarine formed from oils, as the R.N. of butter is higher than that of coconut oil. While P.N. of coconut oil is higher than that of butterfat. Test: Reichert-Meissel Number (A) Saponification: Condenser 5 gm of sample (oil or extracted fat + 20 m l glycerol soda solution Heat continuously tell formation of clean mixture (complete saponification) While still hot add 135 ml of free CO2 water drop by drop to prevent foaming. (B) Distillation: 15  MST:5164 0.5 gm pumice stone + 6 ml H SO 20% + 2 4 Soap solution (fat + glycerol soda solution + water) Complete distillation till having 110 ml of distillate (C) Filtration: Take the distillate and filter through dry filter paper after cooling Filter paper contain the water 110 ml of Water distillate insoluble volatle soluble fatty acids volatile F.A Filter (D) Titration: (E) Calculation: N/10 NaOH Polenske R.M.N. = R x 110 /100 Number 100 ml of filtrate Wash the + 1 ml ph.ph. filter paper which contain the End Point = water faint pink color insoluble volatile fatty acids with 16  MST:5164 water (15 cc in each) three times to get rid of all the rest of soluble fatty acids. Pour neutral alcohol (45 ml divided into 3 portion, 15 each) on the filter paper and receive the filtrate in clean dry flask. Titrate the filtrate against NaOH N/10 using ph.ph. tell having faint pink color Filter paper NaOH N/10 45 ml of contain water neutral insoluble alcohol volatule F.A. Dissolved F.A. in alcohol Fatty acids (FA) + 1 ml ph.ph. End point = Faint pink color P.N. = R Butter Animal fat Coconut oil R.M.N. 24 - 30 0.35 – 1.0 6–8 P.N. 1.3 – 1.5 0.25 – 0.70 16 - 18 Acidity of oils and fats May be expressed either as acid value or in percentage of free acids expressed as the free acids predominating oleic, palmetic or lauric. 17  MST:5164 (A) Acid value: It is the number of mg. of KOH required to neutralize the free fatty acids in 1.0 gm. of fat or oil. N/10 KOH solution (R) 1 ml of 1 % alcoholic ph.ph. 50 ml of mixture of equal parts of Ether & alcohol 10 gm of fat or oil End point = faint pink color 5.61 x (R) Acid value = --------------------------------------------- Weight of tested oil or fat (B) Acidity degree: It is the number of cc of N/10 KOH exhausted to neutralize the free fatty acids in 10 grams of oils Specific tests: (1) Kreis test: Used for detection of rancidity of fat or oil 18  MST:5164 Test: Then added 0.1 % of phluroglucinol in ether , then shake 5 ml conc. HCL 5 ml of oil Shake vigorously for 30 seconds Allow standing for 10 minutes Result: If a pink or red color appears make 2 mixture of original fat in petroleum ether or Kerosene in a ratio of 1:9 & 1:19 Test 5 ml of each mixture as above and read the color Sample Result Judgement Original sample (+) pink color No, rancidity Rancidity not detected 1:9 (+) pink color by odor or taste 1:19 (+) pink color Rancidity detected by odor and taste (Definite rancidity) (2) Halphen’s test: It used for detection of Cotton seed oil Test: 2.5 ml of sulpher in carbondisulphide 19  MST:5164 2.5 ml of amylalcohol 2.5 ml of sample Water bath for 30 minutes Result: Positive result (+) gives Crimson color (3) Baudouin’s test: It is used for detection of Sesame oil Test: 1 ml sucrose in conc. HCL 1 % 2 ml of sample Left for 5 minutes Result: Positive result (+) gives Crimson color ❖ Examine the given oil sample for detection of rancidity 20  MST:5164 1- Technique 2- Result 3- Judgement 4-Significance 21  MST:5164 ❖ Identify the given oil sample A- Cotton seed oil 1- Technique 2- Result B- Sesame oil 3- Technique 2- Result 3- Judgement 4-Significance Date: Supervisor signature:  MST:5164 21 Margarine Margarine or “artificial butter” for many years it was made from beef and animal fat. Nowadays a large number of vegetable oils are used, chiefly cottonseed. Sesame, palm kernel and coconut. Margarine is usually prepared by churning melted and clarified vegetable and animal fats, with milk or cream. The fats are having mechanical agitators. The milk to be used is held in vats and has a ripening culture added. The fats and milk are churned in cylindrical churns through which the mixture is forced to the front of the churn from which it passes into vats. After storage and hardening, it is finally packed or packaged. Ingredients: (1) Animal fat: Beef fat, Matton or lard fat. Fish and marine oils as whale oils after deodourization and bleaching. (2) Plant oils: Used after hydrogenation e.g. cottonseed oil, sesame oil, coconut oil or peanut oil (3) Aqueous ingredients: Ripened skim milk or whole milk after being ripened to give aroma and flavour of natural butter (using butter starter; St. cremoris, lactis, diacetylactis Citrovorus and Paracitrovorus.) 23  MST:5164 (4) colouring matter: Annato or carotins to give yellow colour. Manufacture process: {1} Selection and preparation of suitable mixture of animal fat or hydrogenated plant oils or both (deodourization, bleaching, melting and mixing) {2} Preparation of aqueous ingredients by ripening the skim or whole milk. {3} Mixing and emulsifing of melted fat mixture and aqueous ingredients. {4} Churning of the emulsion (continuos agitation) {5} Cooling and working of product by addition of salt and well mixing and pressing to remove the excessive water, then the coloring and flavouring matter was added, molding and wrapping. Legal requirements of margarine: 1. Fat % not less than 80 % 2. Water % not more than 18 % 3. NaCl % not more than 3 % 4. Milk fat (if present) not more than 10 % of fat content (70 %+ 10 %) 5. Sesame oil must be present as identifiable substance (5- 10%) 6. Must be free from deterioration, rancidity and ingerous matter. Concentrated margarine 24  MST:5164 (Prepared from original margarine) 1. Fat % not less than 98 % 2. Water % not more than 1% 3. NaCl % not more than 1% Kinds of margarine: Plant margarine from plant oil Animal margarine from animal fat Plant and animal margarine Mixed Concentrated margarine from original margarine Examination of margarine and concentrated margarine: As in butter and ghee Comparison and differentiation between butter and margarine (I) Comparison Item Butter Margarine Water 16-20 % 18% Fat % 78-80 % 80% 25  MST:5164 Milk fat 78-80 % 0-8% Foreign fat No 72-80% NaCl 3% 3% (II) Differentiation Item Butter Margarine (A) Physical tests (home test) {1} Spreading test: Glistening Spread a piece of sample on Dull surface surface grease proof paper or bread {2} Foaming test: Will foam Will not foam Heat 2-3 gm of sample in a spoon on the flame {3} Curdling test: Curd will settle Curd will settle In a beaker, 50 gm of sample down leaving clear down leaving turbid melted in water bath(50-600C) supernatant fluid supernatant fluid (B) Chemical tests: using Rose Gottlieb. or Soxhlet method for obtain pure fat {1} Halphen’s test - ve + ve {2} Baudouin’s test - ve + ve Lower (except if {3} Sap. No. 224-230 coconut present) {4} R.M.N. 24-30 Lower {5} Polenske No. 2-3 lower ❖ Differentiate between butter and margarine physically 1-Technique 26  MST:5164 2- Result 3- Judgement 4-Significance Date: Supervisor signature: Table Eggs Hygiene The word egg is of ancient Nordic origin. In popular usage an egg is a vehicle for the reproduction of birds and also a store of food for human consumption. 27  MST:5164 Human interest in the commercial production of eggs depends on their availability as a source of excellent nutrients. The egg is in fact used as a nutritional standard by which other foods are evaluated and judged. Eggs in human nutrition: Eggs supply the three main nutritional requirements of vertebrates, namely: Energy, Protein and essential accessory factors (vitamins, minerals and certain fatty acids and amino acids). The yolk lipid provides energy very largely, while the others are present in the albumen (white) as well as the yolk. According to the WHO (1985), the protein in egg has the highest true digestibility of major foods and together with milk and meat proteins is used as a standard. The nutritive value of egg proteins is also high because they contain the essential amino acids in the required proportions. The lipids in egg yolk provide metabolic energy and they are often more useful as a low-caloric source of other nutrients. Egg yolk lipids from hens on normal diets are relatively low in saturated fatty acids (about 1/3 is saturated). Eggs are rich in linoleic acid (unsaturated fatty acid) which is essential in human nutrition and contain the biologically important minerals (iron, phosphorus, trace elements), but they are not a good source of calcium (discarded in shell). Eggs contain most of the vitamins needed in human nutrition, exception vitamin C and vitamin B (niacin). Each of these is present in very small amount in eggs. 28  MST:5164 The flavour and quantity of nutritive value of an egg depend on the producing hen as well as the mode of their nutrition. Uses of eggs: (A) The use of eggs in old technologies: Leather manufacture Painting Cosmetics Bakery products and ice cream (B) The use of eggs in new technologies: Egg yolk as a preservative for spermatozoa (A.I) Eggs as source of antibodies. The egg as a culture medium. (Microbiology) Medical uses in pharmacology. Other uses for parts of egg: i. The egg shell ii. The vitelline membrane iii. Proteins of the albumen. iv. Egg lipids (C) Commercial uses of avian embryos. (Virology) (D) Gene transfer Composition of the egg The composition of the egg is fairly constant but there are some variations between the egg laid, by one bird, 29  MST:5164 by birds of one strain, breed, or species and by birds from different strains of one species. It is clear that a part from water, proteins and lipids are the main constituents of avian eggs (see following table) The average composition of the hen’s egg Whole Constituents egg White Yolk Shell % by weight 100 58.5 31.0 10.5 Water 73.6 88.5 47.5 1.66 Proteins 12.8 10.5 17.4 6.40 Lipids 11.8 0.02 33.0 0.03 Carbohydrate 1.0 0.5 0.2 0.0 Solids 26.4 11.5 0.0 97.4 Organic matter 25.6 0.0 0.0 0.0 Inorganic matter 0.8 0.5 1.1 91.1 Calcium carbonate 0.0 0.0 0.0 97.25 Magnesium carbonate 0.0 0.0 0.0 0.71 Tri-calcium phosphate 0.0 0.0 0.0 0.61 Formation of an egg: The complete reproductive system, consisting of ovary and oviduct, is shown in the following figure. In the hen usually only the left ovary and oviduct are functional, while the others are rudimentary 30  MST:5164 Figure: The reproductive system of the hen. Showing the ovary (upper part) and oviduct (lower part) Each egg starts as a single cell (ovum) in the ovary. As ovum matures, it is filled with yolk material. After a period of 7-10 days, rapid growth of the vascular tissue (follicle) surrounding an individual mature yolk is cleanly ruptured and the ovulated yolk enclosed in the inner thin, transparent vitelline membrane is released into the infundibulum (funnel-like mouth) of the oviduct. 31  MST:5164 (1) Infundibulum: The yolk generally spends 15-30 minutes in the infundibulum and probably acquires the outer layer of the vitelline membrane and the chalazal layer of the albumen (white) from the tubular or chalaziferous region (narrow posterior end) which contains chalaziferous tubular glands. The infundibulum is capable of limited movement and it functions by grasping the completed ovum at ovulation as well as the probable site for fertilization. (2) Magnum (albumen-secreting region) While the yolk is passing through it, most of the albumen is deposited in 2-3 hours. (3) Isthmus: Both of the shell membranes are deposited on the albumen in 60-75 minutes. It has been suggested that formation of the shell particularly the mammillary cores, which are the centers of mineralization, starts in the isthmus or at the junction with the next region (isthmo-uterine junction). (4) Uterus (shell gland): The egg stays in this region for 18-20 hours and the synthetic processes started in the isthmus continue here to complete the formation of the shell. (5) Vagina:  MST:5164 It plays little part in the secretory processes that lead to the formation of the egg. 31 Structure of an egg: The egg consists of three main parts, the shell, the albumen and the yolk. Longitudinal section through a hen’s egg showing the macroscopic parts Figure: (1) Egg Shell: The parts of the shell will be discussed from the outside starting with the cuticle and proceeding to the membranes. (1.1) The shell cuticle (Bloom): The cuticle is a thin layer of variable thickness of mucoid protein found on the shell of the fresh egg and now known that nearly 90 % of it, is proteins, with some carbohydrate and a smaller amount of lipid. 33  MST:5164 The average dry weight of the cuticle of 60 grams egg is about 12 mg. Once the cuticle is dry, it is very resistant to damage. It is not very soluble in water or salt solution, but it can be partly removed from the shell by washing with water at 40 0C. It can also, be removed from the shell by mechanical abrasion or bad handling. It is normally disintegrated after about 14 days. When the cuticle dry it gives the freshly laid egg a glossy appearance. It is possible that the cuticle helps repel water, it plays a part in controlling loss of water vapor under dry conditions, and it may facilitate oviposition because the cuticle is sticky when the egg is laid and it acts as a protective layer in repelling microbes and small predators. (1. 2) Egg shell pigments: These pigments are confined to the cuticle and the outer part of the calcified layer. The three main pigments are protoporphyrin, biliverdin IX and its zinc chelate. Protoporphyrin tends to give brownish shell colours and the biliverdins blue and green colours, while most white egg shell contain small amounts of pigments. The shell pigments are from two sources, in the oviduct, the shell gland contins enzymes of the porphyrin biosynthetic pathway or synthesis of 34  MST:5164 protoporphyrin from glycine in avian erythrocytes at the same time as heme is synthesized. (1.3) Egg shell (shell): It comprises about 8-13 % of the total egg weight. It is known as the spongy or crystalline layer and is largely responsible for mechanical strength of the shell. The calcified layer consists very largely of inorganic material (97 % or more), usually calcium carbonate (97%), magnesium carbonate (0.7%), calcium phosphate (0.6%) and organic matters. The shell of hen’s egg contains 7 x 103 to 17 x 103 pores that range in diameter from 15-65 m on the outside and from 6-23 m on the inside. They may be partially obstructed by the cuticle or by specialized structures. Pores of the domestic hen’s eggshell are capped with organic plugs (keratin depris-protein) known as cuticular plug. Pores are being more per mm. at the broad end than the narrow end. The pore system performs the indispensable function of permitting uptake of oxygen and loss of CO2 and water vapor through the shell during incubation. Also, pores are used as channel for penetration of microorganisms. (1.4) Shell membranes: They are two thin membranes (outer and inner) formed from insoluble protein (thin keratin-like membranes). They form an excellent barrier against invading microorganisms, particularly the denser inner membrane. (1.5) Air sac (air cell or air space): 35  MST:5164 It is formed between the shell membranes due to the contraction of egg contents at cooling period as body temperature of the hen is 41.5 0C. It is formed normally at the broad end of the shell as this part has many pores per cm2 than the narrow end. Its normal size (depth) is 3-7 mm, but increases during storage due to the evaporation of water content. (2) Albumen (White): It comprises about 60% of total egg weight and consists of four main parts: Two thick whites and two thin whites, in addition, the chalazae may be included (see the following figure) Figure: Diagram of a longitudinal section through a fresh hen’s egg, showing the four parts of the albumen (inner and outer thick, inner and outer thin) plus the chalazae. The proportion of thick white decreases as the hen becomes older and dietary magnesium is deficient. The thick white is firm gels. Most of the albumen in a fresh egg 36  MST:5164 consists of the outer thick white (middle dense layer) which is attached to the egg membrane at the ends of the egg. It gives the albumen its structure in the egg and is responsible for the jelly-like appearance of the freshly isolated albumen. It is also, largely responsible for the quality of commercial eggs. The inner thick white (chalazal or chalaziferous layer or membrane) is the smallest of the four main parts of the albumen. It consists of several membranes surrounding the yolk and forming a narrow gelatinous layer that merges with the vitelline membrane. It is apparently continuous with the chalazae. The chalazae are gelatinous rope-like structures that tie the yolk in the center of the egg and consist of twisted layers of material resembling the thick white. At their outer ends they merge with the outer thick white. The chalaza at the narrow end of the egg is larger and apparently consists of two strands. The other is a single strand. The chalazae are slightly elastic and permit limited rotation of the yolk, but not much lateral displacement. They contain lysozyme. On aging, the chalazae do not deteriorate as rapidly as the rest of the albumen. The thin whites consist of a sticky fluid. The outer thin white (outer liquid layer) is between the outer thick white and the egg membrane, except at the ends of the egg where thick white is attached to the membrane. An increase in the activity of antibacterial enzymes in the thick white has been reported. The pH value of the newly laid egg (white) is about 7.5-7.9, but 37  MST:5164 during storage and due to diffusing of CO2, pH increases up to 9.6 and remains constant. N.B. The movement of the yolk is restricted by thick white, which is attached to the shell membrane at both sides as well as the yolk is anchored with the thick white by the chalazae at both sides. (3) Yolk: It is considered the main source of nutrients and comprises about 31% of the total egg weight. The main parts of a typical avian egg yolk are shown in the following figure. Figure: Longitudinal section through a hen’s egg showing the macroscopic parts of the yolk. The light and dark bands represent different shades of yellow Most of the yolk is yellow yolk, that is depending on the bird’s diet and it has the capacity for heavy pigmentation. It is surrounded by vitelline membrane. It has the blastodisc (Germinal disc) which contains the egg’s DNA. Its pH value is generally about 6, but increases 38  MST:5164 gradually to 6.4 and 6.9 if stored at 2 0C for 50 days or kept at 37 0C for 18 days. Changes of the egg after laying: (1) Ageing: As the egg ages, changes in its structure occur. It is a continuous process. The rate of deterioration is closely related to the loss of carbon dioxide (CO2) through the shell and the increase alkalinity of the egg contents. With higher temperature, the rate of CO2 loss is higher and the egg appears stale within a few days. At low temperature of 8-10 0C, the loss is slight and the structure of the egg is maintained at an acceptable level for several weeks. 1. Basically, It is the thick albumen, which slowly breaks down resulting in more and more thin albumen. The changes occurring in the rest of the egg are associated with the change in proportion of thick and thin albumen. 2. As the egg deteriorates, water slowly migrates from the albumen into the shell and yolk giving them a mottling appearance. 3. The yolk becomes enlarged and flattened during that the thick albumen is decreases. 4. With advancing deterioration the chalazae may become detached allowing the yolk to move freely within the egg and eventually rest against the shell. 39  MST:5164 5. Water evaporates from the liquid contents through the shell pores and air replaces the water lost leading to enlargement of air cell. These changes give the egg a stale appearance, which is unacceptable. In addition, the egg becomes extremely suitable for invasion by microorganisms. (2) Rot and Mold growth: Freshly laid eggs are generally sterile. However, in a relatively short period of time after laying, numerous microorganisms may be found on the outside and may enter eggs under the proper conditions where they grow and cause spoilage. (A) Bacterial spoilage (Rotting): The most common form of bacterial spoilage of eggs is a condition known as Rotting. Types of Rotting: (Bacterial spoilage) Type Causes Green rot with ammonia odour Ps. fluorescens Blue rot Ps. aeruginosa Custard rot with offensive odour Proteus vulgaris Black rot Proteus species Pink rot Serratia species Yellow rot Flavobacterium species Hydrogen sulphide odour Bacillus subtilis Mold spoilage: It is generally referred as pin-spots from the appearance of mycelial growth on the inside shell membrane upon candling. 40  MST:5164 Several genera of molds could be isolated from shell eggs as Alterneria, Cladosporium, Mucor, Penicillium and Thamnidium. They found their way to the contents of the egg through cracks or pores of the shell. The different molds cause spots of different colours. Molds generally show growth first in the region of the air cell where oxygen favours the growth. Whiskers is a form of fungal spoilage which covers the surface of the egg shell specially when the eggs are stored in high humidity. Egg handling and processing by producers: Once a good-quality egg has been laid. It must then be properly handled to minimize any loss of that quality. Eggs are a perishable product. The care they receive between the time of laying and delivery to the first buyer is most crucial. Since the physical and chemical changes in the egg responsible for quality decline are accelerated by high temperatures, it is important to cool eggs promptly. This means frequent gathering, especially in the summer months, to minimize exposure of the eggs to laying house temperatures. Eggs are often placed directly in one-piece filler-flats, which may be stocked on open racks for transporting from the lay houses to the refrigerated holding rooms. Although temperatures just above the freezing point are the most effective in maintaining quality, eggs are generally held between 10 and 16 0C where storage times 41  MST:5164 are limited to a few days. Quality decline at this higher level is slightly greater, but the costs of refrigeration are considerably less. Also, the problem of moisture condensation on the cold egg on removal from the cooler, commonly called “sweating”, is reduced at the higher 42  MST:5164 holding temperature. A relatively high moisture level in the air of the holding room is desirable to minimize loss of water from the eggs. Above 80% relative humidity, mustiness and off-odour may develop. In addition to holding eggs under refrigeration to preserve quality, they may be oil-processed to seal the pores of the shell. This treatment not only retards loss of moisture but also reduce loss of CO2 from the egg. As a result, the pH of the albumen rises less rapidly, therefore, eggs are often oiled as they are gathered in the laying house or shortly thereafter. Microbiology of eggs: The interior of the newly laid egg of healthy stock is usually free from microorganisms but contamination of egg contents occasionally occurs either before the egg is laid or shortly after. As a result, the egg may be decomposed and become unfit for consumption or may responsible for transmitting diseases among poultry and man. (I) The nature and sources of bacterial contamination of eggs There are large variations in the species and varieties of microorganism that infect eggs or grow on their surfaces. In the following table some of the bacterial species found on the surface of hens’ eggs are compared with those found inside infected eggs. Although the surface of the egg 41  MST:5164 contains mostly gram-positive organisms, those on the insides of spoiled eggs are mostly gram-negative. This difference is almost certainly related to the antimicrobial defenses of the egg. Table: bacteria detected on the shell surface and on the inside of rotten eggs Bacterium On shell Inside shell Gram-positive Micrococcus +++ + Staphylococcus ++ - Streptococcus + + Sercina + - Bacillus ++ + Gram-negative Aeromonas + ++ Achromobacter ++ + Aerobacter ++ - Alcaligenes ++ +++ Cytophaga ++ + Escherichia ++ +++ Flavobactericum ++ +++ Pseudomonas ++ +++ Serratia + - Proteus + +++ 44  MST:5164 {1} Contamination before laying: Before they are laid, only microorganisms inside the bird obviously can contaminate eggs. Such contamination, which is difficult to detect and probable accounts for less than 5% of infected eggs and has two possible sources: {a} The bird’s blood: From which microorganisms could pass into the yolk or albumen. {b} The oviduct: From which microorganisms could be incorporated into the vitelline membrane, the albumen, the shell membranes, or the inside of the shell. The oviduct wall can be infected by (1) Microorganisms that migrate from the cloaca by their own motility. (2) Microorganisms that are sucked into the oviduct along with larger foreign bodies by antiperistaltic motion-fortunately a rare occurrence. (3) Incorrect artificial insemination of commercial birds. It has proved difficult to establish the relative importance of the sources of ovarian and oviductal infection, largely because of the problem of contamination by native microflora. The transovarian transmission of Staphylococcus aureus and Pasteurella haemolytica has been established, but not Listeria monocytogenes or Pseudomonas aeruginosa. 45  MST:5164 Salmonella can reach ovaries via the blood from the intestine and can contaminate eggs. It is also possible that contaminated water is a more plausible candidate for the transmission of Salmonella than contaminated feed. (2) Contamination after laying: This is the principal reason for the bacterial infection of eggs. After the egg has been laid, the main sources of bacteria on the surface are the cloaca, the atmosphere, and the place where the egg is deposited, that is the nest or cage. For commercial eggs, the washing and handling procedures are important. The nature and number of contaminating bacteria therefore depend largely on the hygienic conditions of environment where the eggs are laid. Depending on such conditions the number of organisms per egg could vary from 100 to 107. Duck eggs contain a rather high percent of contamination as they lay their eggs nearer to damp places (ponds) with high moisture. They picked up flies and other infective materials. The antibacterial activity of the albumen (conalbumen) deteriorates rapidly on storage. Eggshell is thinner than that of hen’s egg. (3) Contamination during commercial handling: For commercial eggs, the extent of contamination is related not only to the immediate environment where the eggs are laid, but also to factors such as the methods of 46  MST:5164 collecting and handling, the temperature, and the washing procedures. Such factors also influence the subsequent microbial growth. Evidence from several parts of the world has established that much of the early trouble with the infection of commercial eggs during storage and transport was because of overzealous or incorrect washing of the shells. Most of the commercial eggs in developed countries get some washing treatment. The type of detergent used, the temperature, and the pH of the washing solution affect the nature of the microflora and the final load. (II) Barriers to the penetration of microorganisms into eggs: Before it can become established in an egg, a microorganism must pass through the external structures: cuticle, shell, shell membranes. (1) The cuticle: This mucilaginous layer is synthesized and deposited on the egg just before it is laid, so the fresh egg has a sticky, moist surface. The cuticle, which almost completely covers the shell, could evidently play an important part in preventing the entry of microorganisms. Furthermore, shell-less egg and egg removed from the uterus before deposition of the cuticle becomes infected much faster. 47  MST:5164 It was found that the protective action of the cuticle lasts only for 96 hours. Afterward, the cuticle dries and cracks and the egg becomes more prone to spoilage. The proposed role of the cuticle in preventing microbial growth and penetration could be related to : {1} The extent and amount of cuticle deposited on the egg. {2} The amount of egg handling, which could result in removal of the cuticle. {3} A storage condition, for example, the cracking of the cuticle is influenced by length of storage, humidity and temperature. {4} The procedure used for washing and how mach cuticle it removes. (2) The shell: The role of the calcified shell in protecting the contents of the egg from microorganisms is also controversial, which based on the extremely low spoilage rates of eggs with intact shells compared with much faster rates for cracked eggs. Several different factors have been implicated in the penetration of bacteria through the shell. The shell thickness, which varies during the clutch cycle and is related to the time spent in the uterus, influences the ability of bacteria to pass through the pores. Other factors that could influence penetration include abrasion and other damage to the shell. The number of 48  MST:5164 microorganisms and the duration and extent of their contact with the shell. (3) The shell membranes: These structures are probably the most important single barrier to penetration of microorganisms into the egg. They consist of a network of fibers that closely resemble bacterial filters. It has observed bacteria sticking to glycoprotein mantle of the membrane fibers, and lysozyme in the membrane has this function. It is possible that some of the albumen proteins help the shell membranes resist microbes. Thus, the surface-active properties of the ovoglobulins could help plug defects in the membranes. The shell membranes are obviously not an infallible barrier to microorganisms, but how they are penetrated is uncertain. Enzymic digestion could be involved. It is possible that the chelating effects of hydroxymate and phenolates produced by the microorganisms aid their passage through the membrane. (4) The vitelline membrane: For physical and chemical reasons it would be expected that this membrane is a barrier to microorganisms attempting to enter the yolk. Like the shell membranes, the vitelline membrane consists largely of close-packed fibers that should impede the passage of bacteria. 49  MST:5164 The continuous membrane between the fibrous layers appears capable of physically excluding microorganisms completely. In addition, the outer layer of the membrane is composed largely of antimicrobial proteins: ovomucin and an insoluble form of lysozyme that retains antibacterial activity. (III) Antimicrobial properties of the albumen: The albumen of hen’s eggs rarely contains bacteria at oviposition, so any contamination must have penetrated the outer egg structures. The next barrier before the yolk is the albumen. The structure of the albumen is an important physical factor because it impedes the movement of microorganisms and influences the availability of nutrients. The addition of small amounts of water or glucose greatly enhances the growth of microorganisms in the albumen. Furthermore sonication, which physically damages the albumen, causes a similar effect. The water activity of the albumen is not low enough to inhibit microbial growth. The main defense mechanisms in albumen are related to its chemical constituents, principally the proteins. An important additional factor is the pH of albumen. This is variable but is normally in the range 7.6 to 7.9 immediately on laying. Most microorganisms will grow at this pH, although it is not ideal for some, but as the egg ages the pH increases and may reach 9.5, which should, in general, have an inhibiting effect on bacterial growth. 50  MST:5164 (1) Lysozyme: Lysozyme splits the wall of certain bacteria-specifically those that are Gram-positive. Probably for this reason Gram-positive bacteria are rarely isolated from rotten eggs. Gram-negative bacteria are less easily available to enzymes. Gram-negative bacteria can be made sensitive to lysozyme by freezing and thawing or by treatment with EDTA or alkali. (2) Ovotransferrin (Conalbumen) This substance belongs to a class of protein (the transferrins) that is notable for their iron-binding abilities. It has been suggested that ovotransferrin chelates iron in the albumen and so denies this essential element to microorganisms. Iron is not an essential element for all microorganisms. Micrococcus species were most sensitive, followed by Bacillus species and Gram-negative bacteria. A number of other factors, including pH and concentration of CO2, are also important in the binding of iron by ovotransferrin in albumen. While reviewing the antimicrobial properties of albumen, have indicated that ovotransferrin may play the most important part in preventing the growth of bacteria. Several types of bacteria can produce hydroxymates and phenolates, which can counter the effect of ovotransferrin. 51  MST:5164 (3) Avidin: The ability of this minor albumen protein to bind the vitamin biotin is well known. This ability has a role in preventing the growth of microorganisms depends on the requirement of microorganisms for this vitamin. (4) The antiproteases of albumen: These include ovomucoid, ovoinhibitor, cystatin and ovomacroglobulin. They can inhibit the action of a wide range of bacterial proteases. It is possible that ovalbumin, the most plentiful protein in albumen, should be included. (5) Other constituents of albumen: Ovomucin helps prevent the spread of microorganisms. It also has antiviral properties. A minor protein of albumen that may be associated with ovomucin is the enzyme, -N- acetylglucosaminidase, which inhibits Gram-negative bacteria Egg quality defects: Egg quality is compounded of those characteristics of an egg that affect its acceptability to the consumer.  MST:5164 (1) Shell quality defects: {a} Consistency: The eggshell may be weak, rough in consistency and / or misshape due to calcium deficiency. {b} Cracked shell: The eggshell is broken while the 2 shell membranes are sound. {c} Leaking shell: In which the shell is badly broken also, the 2 shell membranes from which egg contents may escape outside. {d} Dirty shell: Dirties, mud, blood and / or faecal matter may found on the shell. Some of the abnormalities that affect hen’s egg shells are listed Table: Shell defects of hens’ egg Appearance Possible causes Wrinkled Copper deficiency; infection Encrusted material from abnormal Pimply or lumpy surface albumen or shell membranes Not certain; abnormal shell matrix; Mottled or glassy shell nicarbazide in feed Shell broken early in uterus then Sealed cracks (checks or body checks) repaired Metabolic disorder, abnormal Chalky or powdery surface phosphate metabolism Two concentric shells Interrupted passage down oviduct Metabolic disorder, pesticides in No shell or soft shell dietb Abnormal pigmentation Unknown b Not common with hens’ eggs 51 (2) Albumen (White) quality defects: 53  MST:5164 {a} Discoloured albumen: In which albumen has a Grey, red, green or blue colour. The different colours are due to the different spoilage microorganisms either bacteria or molds. {b} Cloudy albumen: In which albumen appears turbid (muddy) due to either bacterial rot or washing egg with too hot water. {c} Watery albumen: It is due to a defect in ovomucin synthesis (3) Air cell quality defects: {a} Large air cell: Its depth is more than 7 mm. {b} Ringed air cell: Its depth is very large and sharply defined air cell. {c} Running air cell: Air bubbles are found between the 2 shell membranes and usually due to faulty packaging. (4) Yolk quality defects: {a} Sided yolk:The yolk is presented at any extent from its central position, which may be attributed to faulty packaging. {b} Stucky yolk:Yolk stucks to the inner shell membranes thus favours microbial growth. {c} Flattened yolk: The water content of the egg migrates from white to yolk through vitalline membrane, so the yolk becomes enlarged and flattened.  MST:5164 {d} Spready yolk: Vitelline membrane may ruptured leading to spreading of yolk contents in the white. {e} Yolk mottling: It is due to a non-uniform distribution of water in the mottled yolk or from a separation of the vitelline membrane and the chalaziferous layer of the albumen. Also, it is due to presence of harmful chemicals in feed. {f} Patchy yolk: (Heat spot): It is the enlargement of the germinal disc due to storage of fertile eggs in a temperature over 20 0C Table: Yolk defects in hens’ eggs Defect Causes Close release of two ova Delay in movement of first ovum Abnormal Two yolks ovary with compound follicles Three yolks(rare) Similar Two blastodiscs Early fusion of two yolks Odd shape Weak vitelline membrane Constriction in oviduct Pasty yolks Cylopropenes in diet Abnormal pigmentation Absence of pigment in diet Certain bacterial infections 55  MST:5164 General quality defects: {a} Blood spots: Blood spots or clots or streaks are found in the white or adhere to yolk which lowers the grade of eggs. It is due to rupture of blood capillaries in the yolk follicle during ovulation. This defect is not detected by candling in eggs with brown shell or cloudy white. {b} Bloody egg (Bloody albumen): Blood diffuses in white or around yolk. It is caused by a blood clot breaking and spreading through the albumen. {c} Incubated egg: Fertile egg has a halo around the germinal disc. In advanced incubation, blood vessels or even embryo may found on the yolk. {d} Meat spots: They are fatty, fleshy or liver-like materials floating freely in the white or embedded in chalazae or attached to the yolk. Some meat spots degenerate and change in colour to reddish browning. {e} Rot and mold growth: Microbial contamination of the egg contents either by bacteria or molds produce different colours and odours depending on the kind of the contaminants. {f} Tainted egg: Tainted eggs are those with abnormal odours. Defects that render eggs Defects that render eggs unfit for suitable for Human consumption rapid consumption Bloody egg Large air cell 56  MST:5164 Incubated egg Ringed air cell Cloudy albumen Small blood spots Discoloured albumen Sided yolk Meat spots(Foreign bodies) Patchy yolk Rot and mold growth Mottled yolk Stucky yolk Dirty egg Spready yolk Tainted egg Testing eggs for freshness (A) Egg shell: 57  MST:5164 (1) On small scale: {a} Brine test: Eggs, under test, are transferred to a brine solution (10% Sod. chloride). Fresh eggs, with small air cell, will sink to the bottom while old eggs, with a large air cell, will float at various position and depths depending on size and site of their air cells. Old (large air cell) Suspected Fresh (small air cell) {b} Shaking test: On shaking eggs, fresh one gives no sound while old egg (aged) gives a sound as thick white becomes thin and chalazae are loosened allowing yolk to move freely within the egg. (2) On large scale: {a} Candling: Candling is one commercially suitable way of testing the quality of eggs without breaking their shells. There are two types of candling: 58  MST:5164 i – Individual egg candling: In which the egg is held with the broad end uppermost at a slight angle to the aperture of the candle lamp and withstands to and fro around its long axis. ii- Mass egg candling: In which eggs pass in front of the candler either in a single raw or up to 12 parallel raws in a suitable source of light. More than 150 egg can be illuminated in front of the candler at any one time. The candling involves providing a beam of light sufficiently strong to penetrate the shell and outline the contents to detect the size of the air cell as well as the different quality defects of the egg. {b} Examination of eggs under ultraviolet-rays: (Quartz lamp) The fresh eggs (till 10 days old) when subject to U.V. rays, show a red pink shiny appearance. If the eggs are more than 10 days old., the colour will gradually change from violet to bluish. The basis of this test depends on the presence of oporphyrin in the cuticle present on the shell of fresh eggs, and as the egg becomes older and consequently having less amount of the bloom, as well as due to the effect of day light on the shell, the colour under U.V. lamp will be changed from red to bluish. This test is uncertain because the oporphyrin is present not only in cuticle but also in the shell itself. Moreover, the colour of the shell interferes with the judgment (yellowish brown or brown shells). (B) Broken-out appearance: (egg contents) 59  MST:5164 The eggshell is broken and the contents are transferred to a Petri-dish to detect abnormal colour and/or smell as well as any quality defects in egg contents. {a} Fresh egg {b} Old egg: (Stale egg) The yolk stands up well and is No thick albumen is apparent. held centrally by the albumen. The chalazae become The yolk colour is an even unattach-ed and very weak. yellow. The yolk is displaced, flattened Both chalazae are distinct and and severely patchy or mottled. firmly attached to the yolk. On touching, the yolk easily There is no sign of mottling ruptures. and/ or foreign bodies. Blood and meat spots may be present. The albumen is clear with no tint or colour. Differentiation between the outer thick and thin albumen is distinct. There are no blood spots and other foreign bodies. The germinal disc is just visible. Test the given eggs for freshness Technique 60  MST:5164 Result Judegement significance Grading of egg quality: Egg grading involves inspection of the shell for soundness, cleanliness, apparent strength and shape, checking the interior of the egg by candling and sorting into sizes on the basis of weight. (I) Eggs are classified according to size as: Size Weight/dozen Jumbo 30 oz. Extra large 27 oz Large 24 oz Medium 21 oz Small 18 oz Peewee 15 oz 61  MST:5164 (II) Eggs are graded according to the interior quality: The interior qualities as well as condition and appearance of the shell to Class A, Class B and Class C (clean unbroken eggs). Class C includes all eggs, which do not satisfy the requirements of Class A and Class B but are suitable for the manufacture of foodstuffs for human use. All eggs which do not fall into these classes are classed as (Industrial eggs) and may not be used for human consumption either in the manufacture of foodstuffs or otherwise. See the following table. (III) Dirty or broken eggs: They may not be graded 62 MST:5164  MST:5164 61 Relation to public health: Pathogenic bacteria that can enter the egg contents either before laying or after are able to multiply rapidly as yolk is highly nutritive medium. If such eggs are consumed raw or semi-raw may be responsible for sporadic or epidemic diseases. Many members of Salmonellae e.g. Sal. Typhimurium etc… causing food poisoning may be found in egg contents. Duck eggs are responsible for many cases of food poisoning outbreaks than hen eggs. It has been found that soft boiling, coddling or frying on one side did not always render an egg free from salmonellae. Egg contents may contaminated with other food poising organisms as S. aureus, E. coli and other members of Enterobacteriacaea. Tubercle bacilli (avian type) can be reach and infect consumers through eggs produced from diseased poultry. To secure consumers from being infected, eggs must be obtained from clean farms applying the hygienic measures. Producing hens should be tested regularly for T.B. and Pullorum diseases. Boiling eggs for 5 minutes (hen egg) and 8 minutes (duck egg) with slow cooling destroys microorganisms may be found in egg contents. Cleaning of eggs: 64  MST:5164 Clean and dry eggs are essential to the production and maintenance of good quality. Most eggs are cleaned by washing with modern egg washers designed to minimize defects may occur. Eggs are sprayed with water rather than immersing them, using a sanitizer in water along a detergent for cleaning them, using rinse water warmer than the wash water and finally drying the eggs with hot air. Rotating the eggs during washing and using pressure, sprays and oscillating brushes, provides scrubbing action by the washers. The most often used chemical is any of several chlorine compounds at level of over 50 ppm active chlorine. Preservation of eggs: Preservatives may be used on the shells of eggs, in the atmosphere around them, or on warps or containers for eggs. The chief idea of preservation is to prevent entrance of organisms inside the eggs and to prevent multiplication of the microorganisms, which may gain access inside the eggs. A large number of different substances have been applied to the surface of the shells of eggs or used as packaging material about eggs to aid in their preservation. Some of these substances are primarily to keep the shell dry and reduce penetration of oxygen into the egg and passage of CO2 and moisture out: waxing, oiling the shells and otherwise sealing are examples. Other materials inhibit the growth of microorganisms and some are germicidal. Material 65  MST:5164 used for the dry packaging of eggs in the home includes bran, salt, lime, sand, sawdust and ashes. Methods of preservations: (A) On small scale: {1} Water glass: The eggs are immersed in a solution of sodium silicate, the shell becomes dull in appearance and the eggs can store for many months. The solution is inhibitory because of its alkalinity. {2} Lime-water: Composed of: 4 parts slaked lime 1 part sodium chloride 20 parts of water, Allow solution to stand for a week, decant and rinse the eggs in the resulting liquid. The shells become rough in texture and white dull in appearance. Both methods of treatment can be detected by applying a drop of phenolphthalin, pink colour appears. {3} Using oil and sawdust: The eggs are wrapped with oil and place in layers surrounded with sawdust. The preserved eggs should be stored in a cool place and should not subjected to violent agitation. (B) On large scale: {1} Cold storage (chilling): 66  MST:5164 Most shell eggs are preserved by chilling. They are selected for storage on the basis of their general appearance and a result of candling. The eggs should be cooled as promptly as is practicable after production and held at a temperature and relative humidity that will depend upon the anticipated time of storage. Over seas, temperatures of 0 to 1 0 C are more common, with 80 to 85 % relative humidity. Air circulation in the storage room is important, in order that the desired relative humidity will be maintained around the eggs, and a constant storage temperature is essential to avoid the condensation of moisture on the eggshells. Impregnation of the egg shell with a colourless and odourless mineral oil is a commonly employed method that keeps out moisture, shows down desiccation and air penetration, retains CO2, and retards physical and chemical changes within the egg. At present, the egg is immersed for a few seconds in a thinner mineral oil at atmospheric temperature, or preferably at about 40 0C and then drained. Oiling is used primarily for eggs to be stored commercially for long periods, after several months of cold storage atypical flavour may develop in the stored eggs. To prevent such conditions, periodical sterilization should be done with disinfectant gas as ozone (O3). It has been claimed from 0.6 ppm of ozone (in clean eggs) to 1.5 ppm (for dirty eggs) at 0 0C and 90% relative humidity will keep eggs fresh for 8 months. {2} Oil dipping: Eggs to be oiled should be dipped in light paraffin oil (i.e. viscosity of 55 to 60), preferably at 20 0C immediately 67  MST:5164 after they have been properly washed and dried. All excess oil should be allowed to drain from the eggs. Dipping the egg in water containing 4 % sodium propionate before oil dipping can prevent growth of molds. {3} Vacuum process: The air exhausted from the cell and the shell is coated with oil. The oil is carried into the pores of the shell and acts as seals. Treated eggs have a less oily appearance. {4} Storage under carbon dioxide atmosphere: Storage rooms are charged by CO2 in concentrations of 2.5 % at 85 % relative humidity. The effect is most potent at higher temperature of environment when refrigeration is not available. The quality of such stored egg is improved as eggs deteriorate at first due to loss of CO2. Eggs processing: (I) Heat-treating shell eggs: The pasteurization process has been applied to shell eggs to inactivate the inherent enzymes, destroy the bacteria present on the shell, in the shell and shell membranes as well as in the albumen and yolk of shell eggs. Obviously, several factors must be considered such as temperature of eggs before pasteurization, size, age, grade and type of heating medium. The best results have been obtained by adjusting the eggs at room temperature (28-290C) before immersing and rotating them in oil. The temperature of oil is held at 60 0C 68  MST:5164 and eggs are rotated for 10 minutes. This timetemperature factor is adjusted to prevent coagulation of egg proteins. (II) Pasteurization of liquid egg products: Eggs are pasteurized primarily to eliminate Salmonellae but reduction in other microflora is also of considerable value. The temperature at which egg white proteins are denatured is very close to the temperature required to kill Salmonellae. For this reason, accurate temperature regulation is essential to prevent the loss of functional properties in the white. The procedure is as follows: {A} Eggs whites are acidulated with lactic acid to pH 6.87.3, then aluminum sulphate is added to prevent damage to egg whites, by heat. The eggs can be pasteurized at 60 0C for 3.5 minutes. {B} Egg whites are heated to 51.7 0C for 1.5 minutes to inactivate the enzyme catalase. Hydrogen peroxide is added and pasteurization process continues at 51.7 for 2 minutes. After cooling, the enzyme catalase is again added to remove H2O2. {C} Egg whites are heated at 56.7 0C for 3.5 minutes under vacuum: Normally, less than 1% of the bacteria in raw egg product survive pasteurization. The genera may be found in the pasteurized egg products are: Alcaligenes, Bacillus, Proteus, Escherichia, Flavobactericum and Gram positive cocci. Proposed temperature for pasteurization in 3.5 minutes Product ( 0C) calculated Proposed Whole egg 60 60 69  MST:5164 Yolk plasma 61.4 61.4 + 10 % sugar 63.13 63.8 + 10 % salt 65.4 63.8 Whites: - pH 7 59.4 60 - pH 9 56.6 57.12 Egg products: (1) Liquid eggs: Eggs for processing are broken out either by hand or machine, examined for evidence of spoilage by sight or smell, then homogenized as whole egg or separated into white or yolk. Egg products must now be pasteurized prior to freezing or drying in order to destroy Salmonella. After screening to remove chalaza, shell fragments, etc… eggs are pasteurized at 60-62.8 0C for 1-4 min. prior to cooling. Cooling may be carried out in tanks provided with cooling coils and paddles, which agitate the product to facilitate cooling. (2) Frozen eggs: After cooling to 4.4 0C, the pasteurized eggs may be placed in metal cans holding about 30 lb. of product. The filled cans are then placed in a cold room at -17.8 0C to – 20.5 0C until the product is frozen, after which the product is held at –17.8 0C or lower until shipped out to the distributor or to the point of utilization. Frozen whole egg magma and frozen yolks are subjected to deterioration during frozen storage. Ingredients in the yolk tend to form a gummy mass during frozen storage. In order to prevent this, 5-7.5% salt or 70  MST:5164 glycerin or 5-10% sugar may be added and mixed with the product. {3} Dried eggs: Egg white, yolk or whole egg magma may be spraydried by forcing the product through a nozzle (to form droplets) into a chamber of heated air where most of the moisture is removed from the droplets to the heated air, which is vented to the outside, and the dried product falls to the bottom of the drier from where it is collected. Another method is the roller or drum process in which the liquid egg is passed over a heated drum, with or without vacuum. Air-drying is accomplished by means of open pans or by the belt system where the egg liquid is on a belt that passes through a heated tunnel (60 to 71.1 0C). Spray drying or pan drying, combined with tunnel drying is used for egg white. Formerly eggs were dried to a moisture content of about 5%, but it has been shown that the keeping quality of dried white or whole egg is improved as the moisture content is decreased toward 1 %, and then trend in that direction. A mixture of two enzymes glucose oxidase and catalase, may be used to remove sugars from eggs, which must be carried out prior to drying. Microbiological Examination of eggshell, eggs contents and egg products (I) Egg shell: (A) Swab contact method: 1. A sterile swab, in peptone water, is wrapped on the surface of the eggshell. 71  MST:5164 2. The swab stands in a test tube containing sterile peptone water or broth and incubated for 24 hours at 370C. 3. The different microbiological examinations could be done. (B) Rinse solution method: 1. The egg is immersed in a beaker contains 100 ml sterile tryptic soy broth. 2. The beaker is rotated gently for 15 minutes on a mechanical rotating shaker till complete washing the eggshell. 3. The rinse solution of each egg or 5 pooled eggs, from a part, 10-folds serial dilution is prepared for: Total bacterial count Detection and enumeration of Coliforms. Enterococci count Yeast and mold count 4. The other part is centrifuged and sediment is prepared for the different microbiological examinations. (II) The contents of egg: 1. The eggshell is sterilized by either Tr. iodine or by adding few drops of alcohol then ignited. 72  MST:5164 2. By sterile scissors cuts the eggshell is cutted out around air cell. 3. The contents are removed aseptically and homogenized using a blender. 4. Examinations can be carried out on single egg or on the bulked contents of a number of eggs. 5. Serial decimal dilutions are prepared from the homogenate to estimate total viable count, presumptive test for Coliforms, Enterococci count and yeast and mold count. 6. Also, isolation and identification of S. aureus as well as Enterobacteriacaea, especially Salmonellae, are done. (III) Egg products: (A) Frozen whole eggs: 1. The product is sampled while still frozen. 2. The lid and top of the tin are cleaned and swabbed with alcohol and flamed. 3. The lid is removed, with sterile suitable instrument, 2 cores are removed, one from the center of the can and one at the edge extending from the top surface to a deep level as possible with the instrument used. 4. Samples are transferred to sterile container and examined as soon as possible. 5. The frozen samples are held to soften slightly and while still very slightly frozen, sample is blended thoroughly. 6. Serial decimal dilutions to 10 –5 are prepared to detect; general viable count, presumptive test for Coliforms,  MST:5164 71 DMC using Breed’s method with dilation 10 –2 and isolation as well as identification of Salmonellae. {B} Dried egg: The same procedure for sampling is used as in frozen eggs. {C} Frozen, dried and flakes albumen: The same method used for frozen egg is adopted for such products. The Food and Agriculture Organization of the United Nation (FAO/WHO, 1975) has tabulated the various microbiological criteria imposed for dried and frozen eggs by 19 counties. A summary of these criteria is presented in the following table: Microbial numbers / gm. Dried eggs Frozen eggs Standard plate count 15,000 to 600,00 10,000 to 500,000 Coliform group 10 to 100 10 to 300 Coagulase positive NDa 0.1 to 1000 NDa 0.1 to 1000 Staphylococcus NDb 20 to NDb 20 to NDc Salmonella NDc 25 x30 25 x 30 Yeast and moulds 20 to 100 50 a- ND 0.1 = not detectable in 0.1 g. b- ND 20= not detectable in 20 g. c- ND 25 x 30 not detectable in 30 samples of 25 g. each In addition, the FAO has established its over criteria of these two commodities, summarized as follows: 74  MST:5164 n c m M Salmonella (24 g. portions) 10 0 0 0 Salmonella for special diets 30 0 0 0 Standard plate count 5 2 5x104 106 Coliform group 5 2 10 103 In which n samples are analyzed of which c samples may exceed m, but none may exceed M. Detection of inhibitory substances in eggs: Antibiotics and sulfa drugs are now used on a large scale in poultry industry as feed additives to promote growth and prophylactic agents as well as for treatment of some infectious diseases. Preparation of egg contents: 1. Each egg is carefully washed and the contents are mixed. 2. From the mixture, 2 ml are homogenized with 20 ml of solvent (40 gm Potassium hyroxide, 0.3 gm creatine are dissolved in 100 ml distilled water) 3. The homogenate is centrifuged at 3000 rpm for 10 minutes to separate the supernatant, which is used for the detection of residues. Procedure: 1. Microbiological assay by agar diffusion method is used with Bacillus subtilis as test organism and Iso-sensitest agar as substrate medium. 2. Circular wells (well technique) of 10 mm diameter are punctured carefully in the inoculated agar, with test 75  MST:5164 organism, (1 x 10 6 concentration), 0.1 ml of the egg content sample (supernatant) is transferred to the well. 3. The plates are incubated at 55 1 0C for 6 hours. 4. Any zone of inhibition is recorded as positive results. 76  MST:5164 Food Preservation Food preservation involves the action taken to maintain foods with the desired properties or nature for as long as possible. Food spoilage can occur by: a) Internal biological deterioration b) External biological deterioration Microorganisms and their preferred environments for propagation: ⚫ A large part of food preservation depends on the control of microorganisms. ⚫ Bacteria or microbes are unicellular microscopic organisms that reproduce by binary fission. ⚫ Certain bacterial species can form spores that are highly resistant to killing. ⚫ Molds or fungi are multicellular and unicellular plantlike organisms. ⚫ Yeasts are similar organisms that reproduce by budding. The propagation and spread of molds and yeasts is typically slower than for bacteria because of the reproduction method. Steps that precedes food preservation methods: 1- Controlling access of the microorganisms in the food: This was achieved by adopting of proper sanitation starting from production of milk at the farm till its processing into different dairy products this will helps in reduction of the microbial load to desired levels. 2- Physical removal of microorganisms: 77  MST:5164 a. Centrifugation: It used in liquid foods as milk to remove suspended undesirable particles (dust, leukocytes, some microorganisms and spores). Under high force, as much as 90% of population can be removed. b. Filteration: It's used in heat sensitive nutrients. Filtration of air is used in processing of spray drying of milk to reduce the microbial level from air used for drying. Basic methods, which are used alone or in combination, can extend the normal biological shelf life of the food: Reduced temperatures Heat Treatment Water reduction Chemical preservation Modified atmospheres Irradiation Each method can slow the natural biological maturation and spoilage of a food product, reduce biological activity or inhibit the chemical activity that leads to abiotic spoilage. Each method requires its own unique blend of packaging materials and technology I- Reduced Temperature and Freezing: Reducing temperatures below the ambient temperature has many beneficial effects that will lead to a longer shelf life. 1). Slows chemical activity 2). Slows loss of volatiles 3). Reduces or stops biological activity. Refrigeration: It's advisable to refrigerate the milk directly after its withdrawn from the cow during storage on the farm, during 78  MST:5164 transportation to plant and after pasteurization in plant or in retail market and during delivering and in the home or restaurant until consumption. The desired refrigeration temperature is 4 -5oC. Some of refrigerated foods have a storage life of 60 days or more. Freezing: Lowering the temperature of food so that microbes and enzymes are inactivated. Moisture is changed to ice and microbes become inactive without water. Fast freezing (-25ºC) helps maintain nutritive value and texture of food. Quick or fast freezing occurs at – 25ºC or less. Ice crystals are small and do not damage food cells. While slow freezing occurs at -24 ºC or above. Ice crystals are big and damage the food cells causing loss of texture, nutrients, colour & flavour on thawing. Bacteria and molds stop developing at about -8oC, and by - 18oC, chemical and microorganism activity stops for most practical purposes. II- Heat Treatment: The main objective of heating food is to destroy vegetative cells and spores of microorganisms that include molds, yeasts, bacteria and viruses. Heating of foods also helps to destroy undesirable enzymes that would otherwise adversely affect the acceptance quality of food. Some microorganisms can release toxins in food, sufficient heat will destroy the heat sensitive one, so consumption of such a food will not cause health hazard but the heat stable toxins are not completely destroyed. 1- Low heat treatment: heating of the product at temperature less than 100oC. The best example is the pasteurization. 2- Boiling: Milk is heated at 100.5 °C for 10 - 15 minutes with continuous mixing. But this method changes the appearance, palatability, digestibility and nutritive properties of milk. 79  MST:5164 3- High heat treatment: Heating of the food (milk) at temperature above 100oC for a specific period of time. UHT milk processing at which the milk is sterilized at 135-150oc for 1-2 sec. by direct injection of hot steam or plate heating of the milk at 140oC for 3-4 sec. using steam. The shelf life of milk varies from 6 - 12 months. 4- Steam under pressure (Canning): Putting the milk in clean, dry and sterile jars or bottles and close it then put it in pressure canning container at 10lb pound pressure. Close the container firmly then open the heat and once the pressure reaches 10lb for 10 min, then turn off the heat and wait until the pressure falls down to zero and open the pressure canning container, you will have a canned milk that having good colour and flavor that could be preserved for long time (one year or more). III- Reducing Water activity: Drying is an old and well-established method of preserving food. The essential feature of drying is that moisture content is reduced below that required for the support of microorganisms. An added advantage is reduced bulk and reduction of other chemical activity. Methods: by simple heat drying or by the addition of salt or sugar. IV- The control of the pH: The pH is the negative logarithm of the hydrogen ion activity. It is evident that pH is an important factor affecting growth of microorganisms in food because it affects (i) microbial energy metabolism involving the build-up of gradients of hydrogen ions across 80  MST:5164 membranes, and (ii) microbial enzyme activity and stability of cellular macromolecules. Organic acids are most important in food preservation. The stability and safety of many dairy products are completely or partially due to lactic acid formed by lactose-fermenting lactic acid bacteria. The maximum levels of lactic acid and the final pH in fermented milk products depends on the acid tolerance of the starter culture and their proteolytic activity which they require to utilize milk proteins. Of the thermophilic (i.e., with optimal growth at about 40±45oC) homofermentative lactobacilli, strains of the species Lactobacillus delbrueckii (ssp. bulgaricus, helveticus, lactis) may accumulate up to 250mM lactic acid and lower the pH to about 3.7 while most commercial starter cultures belonging to the Lactobacillus acidophilus group stop fermenting lactose at pH values of 4.0±4.3. When growing in axenic culture, Streptococcus thermophilus lowers the pH to about 4.3±4.6. The final pH of milk soured by mesophilic Lactococcus species is about 4.5. V- Reducing Oxidation- reduction potential(O-R)= Modified atmosphere packaging (MAP): The main purpose of MAP technology is to extend shelf life of food products by slowing down their rates of spoilage while maintaining their safety and general quality. The absence of O2 in the atmosphere would, of course, inhibit the growth of aerobic microorganisms. The removal of O2 can be used to effectively control obligated aerobes such as moulds. Various combinations of CO2, O2 and N2 are usually used as gas atmospheres of choice. Carbon dioxide constitutes less than 0.05% by volume in air. It has a strong bacteriostatic effect on aerobic microorganisms, especially 81  MST:5164 gram-negative bacteria, and inhibitory effects on some enzymes. Researchers listed four proposed mechanisms of action attributed to CO2: ▪ lowering the pH of the food ▪ cellular penetration followed by a decrease of the cytoplasmic pH of the cell ▪ specific actions on cytoplasmic enzymes. ▪ specific actions on biological membranes. Some dairy products, e.g., cheeses, with the exception of those relying on mould cultures (e.g., blue cheese and Camembert), are susceptible to mould spoilage. Packaging them in the atmosphere of CO2 and N2 may extend the refrigerated shelf life of these products. Mould-free shelf life of shredded cheese, for example, can be extended with MAP using 30% CO2 and 70% N2. VI- Irradiation: Food irradiation is the process of exposing foodstuffs to ionizing radiation. Ionizing radiation is energy that can be transmitted without direct contact (radiation) capable of freeing electrons from their atomic bonds (ionization) in the targeted food. This treatment is used to preserve food, reduce the risk of food borne illness. Irradiated food does not become radioactive. Radiation is energy categorized by wavelength and includes radio waves, microwaves, infrared radiation, visible light, ultraviolet light and X rays. These types of radiation increase in energy from radio to X rays; the shorter the wavelength, the greater the energy. Short wavelength radiations have enough energy to cause energy to ionization of molecules, mainly water. Ionization can disrupt complex molecules and leads to the death of living organisms 82  MST:5164 The radiation can be emitted by a radioactive substance or generated electrically. Radiation proved to completely or partially destroy molds, yeasts, bacterial cells and spores and viruses but cannot destroying toxins or undesirable enzymes in food. Irradiation of any food commodity up to an overall average dose of 10 kilogray causes no toxicological hazard. UV rays used in irradiation of processing rooms in dairy plants to reduce the numbers of microorganisms in their air in where sweetened condensed milk is being prepared or cut cheese is being packaged and in cheese curing rooms. UV light is also used in storage tanks containing liquid sugar or other ingredients of ice cream to prevent mold growth on the surface. VII- Natural Antimicrobials: Food antimicrobials are chemical compounds added to or present in foods that retard microbial growth or kill microorganisms. The functions of food antimicrobials are to inhibit or inactivate spoilage microorganisms and pathogenic microorganisms. In addition to potential benefits associated with natural antimicrobials in foods, there are a number of potential concerns that need to be examined with respect to food safety. For example, if an antimicrobial is to be used exclusively to inhibit a pathogenic microorganism, it must be uniformly effective, stable to storage, and stable to any processes to which it is exposed. Standardized assays for activity need to be developed to ensure that the antimicrobial compounds retain potency.