eChapters

F0 value: It is used to express F-value when z =10oC / 18oF and the temperature is 121.1oC. It is the number of minutes at a specific temperature (250oC) required to destroy or kill a stated number of microorganisms and has specific z-value. It is also called reference value.

Facing or lining board: The layer of the paper on which the flute is embedded is called facing or lining board (Plate 258).

Facultative anaerobic: Organisms that grow either with or without oxygen.

Facultative parasites: Facultative parasites can adapt to live on dead organic matter and often have a wide host range and can easily be cultured on artificial media. These organisms may be aerobic or anaerobic but under changed conditions, they changes to anaerobic or aerobic.

Falling rate period: It is that part of the drying process during which the drying rate varies with time and the instantaneous drying rate per unit surface or weight of dry material continuously decreases. Falling rate takes place when most of the moisture has been evaporated during constant rate period. The falling rate period during dehydration take place in the final stage of dehydration when the food commodities have large amount of free water available for evaporation. Here, rate of removal of the moisture from the food to the surrounding is continuously decreasing / falling per unit time.

E numbers: Each food additive used in the food industry has been given a specific number code for their identification. These numbers are called as E numbers. These E numbers are used within the European Union. The word prefix “E” stands for Europe. They are commonly used on the food label through out the European Union. These are not used in Australia and New Zealand. The different food additives are given different E numbers depending upon the type of food additive like E100-E199 (food colours), E200-E299 (preservative), E300-E399 (antioxidants), E400-E499 (thickners, stabilizers, emulsifier), E500- E599 (pH regulator, anticaking agent), E600- E699 (flavour enhancers), E700-E799 (antibiotics), E900-E999 (miscellaneous like waxes, synthetic glazes, packaging gases, sweetners, foaming agents), and E1000-E1099 (additional chemicals).

Earling: The young lamb of one to two years’ age is called earling or yearling.

Ebullioscopic constant: The boiling point elevation is expressed by Dtb = Km, where m is molality of the solution and K is ebullioscopic constant. It is also called as molal boiling point elevation constant. The unit of an ebullioscopic constant is K Kg mol-1.

Dahi: It is the product obtained from pasteurized or boiled milk by souring, natural or by harmless lactic acid or other bacterial culture. It is also called Curd (Plate 209).

Dairy: It is a room, building, or establishment where milk is kept and butter / cheese is made or it is the department of farming / farm that is concerned with the production of milk, butter, and cheese or an establishment for the sale / and distribution of milk and milk products.

Dairy husbandry: It deals with care, breeding, feeding, milking of the diary cattle and the production and sale of the milk. It is the economical management of a dairy, including looking after the dairy cattle breeding and proper disposal or sale of milk and milk products.

Dairy products: These are the milk preparation, including butter, cream, cheese, ghee, yoghurt, pasteurized milk, tonned milk, butter milk etc processed and prepared in dairy. Ice cream and kulfi or sweets are not included in this.

Damage: Damage is a physical spoilage, often a partial deterioration, are subjectively judge and very difficult to measure and usually reported as per cent damage.

Daminozide: It is a growth regulator which is used for red colour development in apple.

C - enamel can: The sulphur resistant lacquer is golden coloured and the can coated with it are called C-enamel can or sulphure resistant can, used for packing non acid products (Plate 143).

CA storage: Storage of commodities under gaseous atmosphere like gas storage, where the composition of atmosphere is controlled accurately. MA and CA differ only in degree of control and CA is more exact. These storage methods in combination with refrigeration markedly enhance storage life of fruit and vegetables (Plate 144).

CAC: It stands for Codex Alimentarious commission (1962). It is a joint commission of FAO and WHO, comprising of 146 member countries. It ensures consumer food safety, establish fair practices in food trade and promote the development of international food standards. CAC make standards for food additives, veterinary drugs, pesticide residue and other miscellaneous substances that affect consumer safety. These are published in Codex Alimentarius.

B.T.U.: It stands for British thermal unit. The heat required to raise the temperature of one pond of the water through 1oF is called BTU (Plate 82).

Baby back ribs: Meaty pork ribs cut from the lower back rib section.

Bacitracin: A polypeptide antibiotic isolated from a bacillus (Bacillus subtilis or B. licheniformis) and usually used against gram-positive bacteria.

Back Bacon / Canadian bacon: Bacon cut from the loin that has little fat and is cut into round or oblong slices.

Backward feed evaporator: In this evaporator, fresh feed enters the last and the coldest effect and continues on until the concentrated product leaves the first effect (Plate 83).

Abnormal milk: It is milk that is not smooth and homogenous but is flaky, ropy, stringy, slimy, watery, discoloured or bad flavoured (Plate 1 & 2).

Abrassive peeler: It is equipment comprising a rotating drum lined with carborandum (an abrasive material) in which certain vegetables like potato and ginger are tumbled and exposed to the abrasive action of the carborandum lining, thereby facilitating separation of peel from the vegetables (Plate 3).

Absinthe: It is green liqueur which is flavored with wormwood, anise, and other aromatic herbs (Plate 4). Its commercial production is banned in ma ny countries for health concerns.

Absolute alcohol: It is water free alcohol. It contains atleast 99 per cent pure alcohol by weight and obtained by digesting rectified spirit over quicklime and redistilling the same.

Absolute density: It is the ratio of weight of certain volume of a substance at 1oC to that of weight of same volume of water at 4oC.

Absolute humidity: Absolute humidity is defined as kg of water vapour per kg of dry air at a given temperature and pressure. Absolute humidity is the mass of water vapour divided by the mass of dry air in a certain volume of air at a specific temperature. The warmer air absorbs more water than cooler one. Absolute humidity is the measure of water vapour or moisture in the air, regardless of temperature.

Absolute pressure: It is equal to atmospheric pressure minus vacuum pressure (Plate 5).

Absolute zero: It is a theoretical temperature characterized by complete absence of heat and motion and equivalent to exactly -273.15°C or -459.67°F (Plate 6).

Vitamins are naturally occurring, vital nutrients that are needed in small amounts. They are important for bone and tissue health, wound healing, growth and development, immune system function, and other biological processes. These vital organic substances serve a variety of biochemical purposes. Vitamins, like minerals, are not biosynthesized by the body. Consequently, in order to maintain our health, we must obtain them from the food we eat or, in rare circumstances, supplements.

Vitamins in Milk

Together with water- soluble vitamins like vitamin C and B complex and provitamins, milk also contains a variety of fat- soluble vitamins like A, D, E, and K. Some of these vitamins do, however, have very low concentrations.

Vitamin A (Retinol)

Plant foods contain carotenoids, which are the precursors of vitamin A, whereas animal foods provide preformed vitamin A in the form of retinyl esters.

The primary constituents of milk are water, fat, protein, sugar (lactose), minerals, vitamins, and enzymes. Milk is a thin emulsion made up of two phases: A continuous aqueous colloidal phase and a dispersed phase of oil or fat. It is an opaque white liquid containing fat in an emulsion, protein, and a few minerals in a colloidal suspension, and lactose along with soluble proteins and a few minerals in a true solution. The physical characteristics of milk are comparable to those of water, but they are altered by the degree of dispersion of the colloidal and emulsified components as well as the presence of different solutes, such as salts, lactose, and protein, in the continuous phase.

Physical Appearance, Color and Optical Properties

Fat globules and casein micelles scatter light, giving the appearance of turbid and opaque milk. The way that molecules scatter light has an impact on optical characteristics. When the wavelength of light and the particle’s magnitude coincide, light scattering happens. Consequently, light with shorter wavelengths is scattered by smaller particles and vice versa. Because casein micelles scatter blue light more than red light at shorter wavelengths, skim milk appears slightly blue. Cow’s milk has a creamy color because of beta- carotene, a carotenoid precursor to vitamin A. The greenish tinge in whey is due to the presence of riboflavin. Turbid and opaque due to scattering of light mainly by protein micelles and fat globules.

Electrophoresis

The mobility of ions in an electric field is the basis for the separation method known as electrophoresis. Ions that are negatively charged move toward a positive electrode, while those that are positively charged move toward a negative electrode. Usually, one electrode is biased either positively or negatively, and the other is at the ground for safety reasons. Ions can be distinguished from one another based on their size, shape, and overall charge, which determine their migration rates.

Poly Acrylamide Gel Electrophoresis (PAGE)

Introduction

Electrophoresis is an electrochemical process in which substances with a net electric charge migrate under the influence of an electric current. This migration occurs in an agar gel or a liquid- filled buffered matrix such as cellulose acetate. Positively charged substances travel toward the cathode (negative electrode), while negatively charged substances go toward the anode (positive electrode). Different substances move at different rates depending on their charge. This movement is called electrophoretic mobility.

Proteins are charged molecules. They have a positive as well as a negative charge. Among proteins, particularly caseins, they have different contents. The charge density also varies depending on the molecular weight. Proteins have a net negative charge at a pH above 4.6, and hence they move towards the anode (positive electrode) when subjected to an electric field under alkaline conditions. Due to differences in the charge of individual protein fractions, their migration rates also differ and are hence separated into distinct bands. These bands are stained with the dye amido- black, destained with 7% acetic acid, and observed for the electrophoretic pattern.

Food adulteration is a global issue, and because there are insufficient regulations and oversight, developing nations are more vulnerable to it. But in many nations, food adulteration has gone unnoticed. Contrary to popular assumption, milk adulterants can, regrettably, present major health risks that result in fatal diseases. Modern milk adulteration involves increasingly complex techniques, necessitating state- of- the- art research to identify the adulterants. Because milk contains so many nutrients that are needed by both adults and infants, it is referred to as the ideal food. In terms of protein, fat, carbohydrates, vitamins, and minerals, it is among the best sources. Regretfully, milk adulteration is a very common problem worldwide. Potential causes could be the gap between supply and demand, the perishable nature of milk, the low purchasing power of the consumer, and the absence of appropriate detection tests. Because there is insufficient oversight and inadequate law enforcement, the situation is substantially worse in developing and underdeveloped nations. While quantitative detections of adulterants in milk are varied and complex, qualitative detections can be carried out simply using chemical reactions. The type of quantitative detection methods used depends on the milk’s adulterants.

Typical Adulterants in Milk

The addition of vegetable protein, milk from different species, whey addition, and watering are the main adulterants in milk that are known to be economically motivated. There is no serious health risk associated with these adulterations. Some adulterants, though, are too dangerous to ignore. Urea, formalin, detergents, ammonium sulfate, boric acid, caustic soda, benzoic acid, salicylic acid, hydrogen peroxide, sugars, and melamine are a few of the main adulterants in milk that have been shown to have serious negative health effects.

The terms “cultured dairy foods,” “cultured dairy products,” and “cultured milk products” are other terms for fermented dairy products. Lactobacillus, Lactococcus, Leuconostoc, and other lactic acid bacteria, along with yeasts in certain products, are used to ferment milk to make these dairy products. The product’s shelf life is extended through fermentation, which also improves the product’s taste and milk’s digestibility. There is a wide variety of fermented milk products available, each with unique flavors and attributes. The following categories apply to fermented milk products, based on the type of fermentation.

Changes in Nutrients Resulting from Fermentation

Fermentation lowers the lactose content in fermented milk products; however, if the starting milk is concentrated or fortified, the finished product’s lactose content may be higher than that of liquid milk.

The composition and content of protein and total amino acids are comparable to those of raw milk. Casein degradation is small compared to cheese, accounting for not more than 1% of the total protein. While some products of casein degradation are used by the starter microorganisms, others build up as free amino acids in the fermented kinds of milk. The amounts of free amino acids that build up differ depending on the species. Generally, proline is increased in large quantities, followed by serine, alanine, valine, leucine, and histidine. As a result of continuous starter culture activity, the level of free amino acids increases during storage. A limited degree of lipolysis occurs during the manufacture of fermented milk. The amount varies depending on the type of species, but the pattern and concentration of free fatty acids are both impacted. During fermentation and the breakdown of amino acids, trace amounts of volatile fatty acids are also produced.

Being an essential component of the rich Indian past, traditional dairy products and a sophisticated classification of confections have played a significant role in the Indian economy. Depending on how it is processed, it has significant social, cultural, and economic significance. These products have been developed over a long period using a combination of experience, traditional wisdom, and culinary skills. India is emerging as the highest milk- producing country in the world with an annual production of 221.1 million tons in the year 2021–22 (Source: Basic Animal Husbandry Statistics, MoFAHD, DAHD, GoI). An estimated 50% of the milk produced in India is used to make a range of traditional dairy products, including paneer, dahi, ghee, khoa, and shrikhand. The product is not only well- established in the Indian market, but it also has significant export potential due to the large Indian diaspora living all over the world.

All native milk products that have developed over time using locally accessible technologies are referred to as traditional Indian dairy products or Indian indigenous milk products. The Indian masses’ economic, social, religious, and nutritional well- being are greatly influenced by traditional dairy products.

Classification of Traditional Indian Dairy Products

Traditional dairy products in India are typically made, packaged, and/or sold using time- honored methods without the use of machinery or meticulous documentation of the procedure. These products do, however, have several technological advantages, including their widespread appeal, reduced cost of production, straightforward manufacturing process, adoption at the rural level or collection center, large profit margin, and abundance of employment opportunities. These products are categorized according to the manufacturing principle.

One of the most consumed and well- known milk products in the Indian subcontinent is ghee. It is a by- product of tradition, prepared both at organized dairy plants and the household level for a well- established market. Ghee is traditionally made from the milk of cows or buffalo. The tetrapyrrole pigments biliverdin and bilirubin are responsible for the distinctive color of buffalo ghee. On the other hand, ghee is golden yellow due to the presence of carotenoids in cow’s milk. Ghee’s composition changes according to the kind and feeding habits of the animal and can differ from place to place and season to season.

In addition to, being a substantial source of fat- soluble vitamins, essential fatty acids, and other growth- promoting components, ghee is a rich source of energy. Its value as food for humans is enormous. Additional attributes related to medicine are present in ghee. Ghee can improve one’s physical and mental attributes and treat eye conditions like ulcers. Ghee is used in religious and ceremonial rites in addition to being consumed.

Composition

Ghee is a complex lipid composed of mixed glyceride which makes up 99–99.5% of its total content. In addition, it has trace amounts of calcium, magnesium, phosphorus, iron, copper, and fat- soluble vitamins, tocopherols, carbonyls, hydrocarbons, carotenoids, free fatty acids, phospholipid, sterols, and sterol esters

The process of making dried milk involves eliminating water from milk, primarily through spray or roller drying. It is regarded as a significant sector of the dairy industry and is made from whole cow, buffalo, or standardized milk. Skim milk powder (SMP) is made from milk after the fat has been removed whereas whole milk powder (WMP) is made from whole cow or buffalo milk

FSSA (2006) Standard Of Milk Powder According to FSSA (2006), Milk powder is the product obtained by partial removal of water from the milk of cows and/or buffalo. The fat and/or protein content of the milk may be adjusted by the addition and/or withdrawal of milk constituents in such a way as not to alter the whey protein to casein ratio of the milk. It shall be of uniform color and shall have pleasant taste and flavor, free from off-flavor and rancidity. It shall also be free from vegetable oil/fat, mineral oil, thickening agents, added flavor, and sweetening agents. It may contain food additives. It shall conform to the microbiological requirements prescribed by FSSA (2006).

Water activity, high storage temperatures, longer than usual storage times, high product moisture content, high processing temperatures, and container headspace are some of the variables that affect the Maillard reaction in dried milk.

Casein and whey proteins are the major proteins present in milk. Unlike casein, whey proteins are heat-labile. Whey protein fractions present in milk are α-lactalbumin, β-lactoglobulin, bovine serum albumin, immunoglobulin, euglobulin, and lactoferrin. β-lactoglobulin and κ-casein interact during heat treatment. When milk is refrigerated for longer than 24 h, it forms γ-casein and protease-peptone fractions.

Commercially, one of the most significant milk products is ice cream. It is a frozen product which is usually made from milk or cream that has been f lavored with fruits like peaches or strawberries, and spices like vanilla or cocoa, or with sugar or a substitute. Stabilizers are sometimes combined with food coloring. Ice cream mix typically contains 10.0% milk fat, 11.0% solids not-fat, 14.0% added sugar, and 0.3% additives like an emulsifier, stabilizer, f lavor, and color substances.

Standard of Ice Cream

According to FSSA (2006), ice cream, kulfi, chocolate ice cream or softy ice cream means the product obtained by freezing pasteurized milk prepared from milk and/or other products derived from milk with the addition of sweetening agents and fruit products, egg products, coffee, cocoa, ginger, and nuts. It may also contain chocolate, and bakery products such as cake or cookies as a separate layer and/or coating. It may be frozen hard or frozen to a soft consistency. It shall be free from artificial sweeteners. It shall have a pleasant taste and smell free from flavor and rancidity. It may contain food additives. It shall conform to the microbiological requirements as given in FSSA (2006).

Concentrated milk products are obtained through partial removal of water which increases the dry matter content of milk. The increase in dry matter content in concentrated milk is proportional to the degree of concentration. The milk concentrate may be prepared from either full-cream whole milk or fat-free skim milk.

Depending on the nature of raw material and other non-milk additives concentrated milks are classified into four broad categories.

1. Sweetened condensed milk

2. Evaporated milk or unsweetened condensed milk

3. Sterile milk concentrate

4. Reconstituted concentrated milk

Sweetened Condensed Milk

One of the earliest dairy products to be produced industrially is sweetened condensed milk. According to FSSA (2006), sweetened condensed milk is the product obtained by partial removal of water from the milk of cow/buffalo with the addition of sugar or a combination of sucrose with other sugars or by any other process that leads to a product of the same composition and characteristics. The fat and/or protein content of the milk may be adjusted by the addition and/or withdrawal of milk constituents in such a way as not to alter the whey protein to casein ratio of the milk being adjusted. It shall have a pleasant taste and flavor and be free from off-flavor and rancidity. It shall be free from any substance foreign to milk. It may contain food additives. It shall conform to the microbiological requirement as laid down by FSSA. The said product shall also conform to the following requirements

Cheese is made from milk, with the addition of cream, buttermilk, and/or whey. This basic raw material is also called cheese milk. Cheese consists mainly of casein (the major protein of milk), other milk proteins, fat, other milk components, and a certain percentage of water, which is bound. Proteins are precipitated by enzymes and/or acidification as well as heat and are separated by suitable processes from the aqueous phase. (whey, serum). Cheese is most often molded, pressed, and salted and has fungal and bacterial culture additions. colorants, herbs, spices, and other non-dairy ingredients are also added to the cheese. It is consumed either fresh or at different stages of ripening.

Worldwide, there are more than 2000 types of cheese, sometimes made by very different manufacturing processes. A classification can be based on several aspects and is done in different countries according to different criteria. A general classification can be made for three major groups:

a. Rennet or natural cheese: manufactured straight from milk by using proteolytic enzymes (rennet) and acid, with a more or less pronounced ripening process.

b. Fresh cheese or non-ripened Cheese: Quarg, fresh cheese, and white cheese are examples. Its manufacturing process is similar to that of rennet cheese manufacturing, but it has a high degree of acidity and is not subjected to a proteolytic ripening process.

c. Long-life cheese (processed cheese): it is more often made from rennet cheese and is textured by thermal treatment and made shelf-stable.

Chemistry of Creaming

After being drawn from animals, normal milk will form a cream layer on standing. Because of the difference in density between fat globules (920 kg m−3 at room temperature) and milk plasma (density 1030 kg m−3), the granules tend to rise. This causes creaming, which we often want to prevent and it enables milk to be separated into cream and skim milk. Creaming is much enhanced when the globules have been aggregated into flocculates, clusters, or granules.

Cream Ripening

This unit operation serves as an intensive preparation of the cream for the churning process. It is divided into two categories.

1. Physical ripening;

2. biochemical ripening The following criteria are influenced by the cream ripening:

• Consistency, firmness, and spreadability of butter.

• Basic water content of butter during the manufacturing process (bound water in butter, which cannot be removed by any mechanical treatment).

• Buttermilk fat content, which has to be minimized.

The melting and solidification behavior of fat depends on the attached fatty acid, which indicate the hardness of the butterfat.

Natural hardness is caused mainly by seasonal feeding of the milk animals. Winter feed with a great deal of dry feed or roughage yields a hard butterfat with a high percentage of saturated and short-chain fatty acids and few double bonds in the fat molecules.

Summer feeding (green grass) results in fat with longer chains and a higher percentage of unsaturated fatty acids with many double bonds. We therefore distinguish between summer fat and winter fat or summer cream and winter cream.

Types of Enazymes in milk

1. Endogenous and 2. Exogenous.

Three sources of enzyme in milk:

1. The blood via defective mammary cells.

2. Secretory cell cytoplasm. Some of which are occasionally entrapped with fat globules by the encircling MFGM.

3. The outer layer of MFGM is probably the principal source of endogenous enzymes in milk. The outer layer of MFGM is derived from the apical membrane of the secretory cell which in turn originates from the Golgi membrane and acts as the principal source of enzyme in milk.

General Characteristics of Milk Enzymes

1. Deteriorative character e.g. lipase (commercially most important enzyme in milk), proteinase, acid phosphatase, xanthine oxidase.

2. Preservative enzymes. e.g. superoxide dismutase, sulfhydryl oxidase.

3. As an indicator of the thermal treatment history of milk. Related enzymes are alkaline phosphatase, ϒ glutamyl transpeptidase, and lactoperoxidase.

4. Mastitis infection indicator. Related enzymes are catalase, n-Acetyl-β D-glucosaminidase, and acid phosphatase.

5. Antimicrobial activity.

6. Commercial source of ribonuclease and lactoperoxidase

Common Milk Enzymes and their Role in Dairy Industry

The salts of the milk include those constituents that are present as ions or in equilibrium with ions. The main salts of milk are phosphates, citrates, chlorides, sulfates, carbonates, and bicarbonates of sodium, potassium, calcium, and magnesium. Approximately 20 other elements are found in milk in trace quantities including copper, iron, lead, boron, manganese, zinc, iodine, etc. These ions are more or less associated with themselves and with proteins. Some metals act as catalysts in the oxidation of milk fat leading to oxidative rancidity and certain metals like Ca, P, Mg, Mo, Co, Fe, etc. are components of metalloproteins such as lactoferrin, lactoperoxidase, xanthine oxidase, and parts of phospholipids.

Distribution of Salts Between Casein Micelles and Serum

The phosphorus is present as orthophosphate, but part of it is bound to organic components like serine and threonine residues of casein, molecules of hexoses and glycerol, phospholipids, etc. Phosphorus is also found in the hydroxyapatite structure of the colloidal calcium phosphate in the micellar structure of casein. The sulfur content of milk is about 0.36 g/kg, but most of it is in the amino acid residues methionine and cysteine of the proteins. About 10% is present as inorganic sulfate.

Physical Equilibria Among Salts

Milk contains several elements but all of them are not entirely in a soluble state. Some minerals exist in colloidal and ionic states at the normal pH of milk. The compounds that are in soluble condition play an important role in keeping various milk constituents in stable condition. A balance exists between the components that are in a soluble state and those which are in colloidal state.

Milk lipids or milk fats exist as an emulsion of small spherical droplets called milk fat globules. These globules are very small in size but very large in number. The diameter of the milk fat globules ranges between 0.1 and 22 µm and the number varies between 1.5 and 3.0 billion globules/mL. The average size of milk fat globules ranges from 2 to 5 µm. The average fat globule size in cow and buffalo milk are 3.4 and 4.5 µm, respectively. The surface of the milk fat globules is coated with an adsorbed layer of material known as the milk fat globule membrane (MFGM). MFGM contains phospholipids and proteins in a complex form. MFGM has two main functions (a) fat globule stabilization and (b) preservation of the fat globule’s identity.

General Composition

Triacylglycerols (triglycerides) represent 97–98% of the total lipids in the milk of most species. The diglycerides probably represent incompletely synthesized lipids in most cases. Although phospholipids represent less than 1% of total lipids, they play an important role in being present mainly in the MFGM and other membranous material in milk. The principal phospholipids are phosphatidylcholine, phosphatidylethanolamine, and sphingomyelin. Trace amounts of other polar lipids, including ceramides, cerebrosides, and gangliosides, are also present. Phospholipids represent a considerable percentage of the total lipid of buttermilk and skim milk reflecting the presence of proportionately larger amounts of membrane material in these products.

Carbohydrates are polyhydroxy derivatives of aldehydes or ketones and undergo all the organic reactions common to the carbonyl groups and hydroxyl groups. The principal milk carbohydrate is lactose. It is a disaccharide. Hydrolysis of lactose with enzyme β-galactosidase yields glucose and galactose.

Lactose is the major carbohydrate present in all types of milk. The first record of isolation of lactose was in 1663, by Bartolettus by evaporation of whey. Milk contains only trace amounts of other sugars, including glucose, fructose, glucosamine, galactosamine, neuraminic acid, and neutral and acidic oligosaccharides.

Lactose plays a significant role in milk and milk products that are

• It plays a major role in the production of fermented dairy products.

• It contributes to the nutritive value of milk and milk products

• It affects the texture of certain concentrated and frozen products;

• It is involved in heat-induced changes in the color and flavor of highly heated milk products.

Status of Lactose

Lactose content varies extensively in the milk of different species. Breed of animal, individuality factors, udder infection, and especially the stage of lactation regulates the lactose content in milk. The lactose concentration in milk decreases gradually and significantly during lactation; this behavior contrasts with the lactational trends for lipids and proteins, which, after a decrease during early lactation, increase significantly during the second half of lactation. Lactose synthesis is hampered during mastitis. Lactose, along with sodium, potassium, and chloride ions, plays the principal role in controlling the osmotic pressure of the mammary system. Thus, any change in lactose content is compensated by a change in the soluble salt constituents. The osmotic relationship between lactose and soluble salts during the secretion of milk partly explains the presence of high lactose in milk with a reduced ash content and vice versa (Table 1).

Bovine milk contains an average of 3.5% protein. The concentration of protein changes significantly during lactation. The first few days of lactation exhibit the main change in protein concentration. The main changes occur in whey protein fraction.

Distribution of Protein in Milk

80% casein and 20% whey protein (β lactoglobulin- 10%, α- lactalbumin- 5%, serum albumin, and immunoglobulin- 5%).

Fractionation of Milk Proteins Fractionation of milk protein is mainly done by acidification at pH 4.6 around 30 °C.

Almost 80% of the bovine milk protein precipitates at pH4.6, and this fraction yielded by precipitation is called casein.

The remaining soluble part of the protein is referred to as whey protein/serum protein/ non- casein protein. Wide variation is observed in the casein- whey protein ratio in different types of milk(protein synthesis is independent of animal diet)

Significant Difference Between Casein and Whey Protein

1. Casein precipitates at pH 4.6 but does not precipitate whey protein at that pH. This property of milk protein is commercially exploited for the preparation of industrial casein and certain varieties of cheese.

2. Chymosin or other proteolytic enzymes produce specific changes in casein resulting in coagulation in the presence of calcium. Whey protein does not undergo this type of coagulation.

3. Casein is very stable to high temperatures. They can withstand heating up to 140 °C for 20 minutes but whey protein is relatively heat labile. They are completely denatured by heating at 90 °C for 10 min.

4. Casein is a phosphoprotein containing an average of 0.85% phosphorous while whey protein does not contain any phosphorous. The phosphate of casein is an important contributor to remarkably high heat stability and calcium- induced coagulation of rennet- altered casein.

5. Casein contains very low sulfur (0.8%) whereas whey protein is relatively rich in sulfur (1%). The principle casein contains only methionine but whey protein contains a significant amount of cystine, cysteine, and methionine. These sulfur- containing amino acids are responsible for many changes in milk on heating.

6. Casein is synthesized only in the mammary gland and no other sources in nature. Some of the whey proteins β lactoglobulin, and α- lactalbumin are synthesized in the mammary gland but bovine serum albumin and immunoglobulin are not synthesized in the mammary gland but derived from blood.

7. The whey protein is dispersed in solution and has a simple quaternary structure. Casein has a complex quaternary structure and exists in milk as large colloidal aggregates.

Introduction

The mammary glands of female mammals secrete milk, a necessary food that is liquid and meets the newborn’s nutritional needs. All mammals consider milk to be a complete food for their newborns. It is considered a necessary food for humans and can be consumed as a product or as liquid milk. It contains almost all essential nutrients except iron, which is very low in milk. In essence, milk is an emulsion of protein and fat in water, supplemented with carbohydrates, vitamins, and minerals.

Definition of Milk

• Milk is a complex biological fluid that differs from species to species in both composition and physical properties. It is the full lacteal secretion of a mammary gland that is obtained by milking mammals for a minimum of 72 h after calving or until the milk is free from colostrum.

• FSSAI defines milk as the normal mammary secretion derived from the complete milking of healthy milch animals. It should be free from colostrum.

From a chemical perspective, milk is a dynamically balanced mixture that comes in different forms such as emulsion, colloidal suspension, and true solution. It also contains varying amounts of proteins, fats, carbohydrates, salts, and water.

Gross Composition of Milk

Milk composition varies among different mammals. The average chemical composition of milk in various species is given in Table 1. Buffalo milk and sheep milk contain higher amounts of fat, while human milk comparatively has the highest lactose content. Human milk has lower protein content as compared to milk of other species.

Introduction

Wastes produced during fish processing operations can be solid or liquid. Solid wastes consist of skin, viscera, fish heads, fish bones, and scraps of f lesh. Liquid wastes are the blood water and brine from drained storage tanks, water discharges from washing and cleaning, blood and soluble substances from entrails, and detergents and other cleaning agents. The effluent from the processing of fatty fish contains very high levels of oil. Overall, the wastewater from fish processing operation contains an undefined mixture of substances, mostly organic in nature.

13.1. Physicochemical Parameters of the Wastewater

13.1.1. pH

The pH is an important parameter that reveals contamination level before biological treatment. The effluents from fish processing plants are usually close to 7 or alkaline, due to decomposition of the proteinaceous matter and emission of ammonia compounds.

13.1.2. Solids Content

Solids may be present either in dissolved or suspended form. Suspended solids are objectionable as they settle in the wastewater ducts reducing their capacity or settle in the waterbody affecting bottom - dwelling flora or sometimes float reducing the light entering from the surface or remain suspended reducing the amount of light that enters the water. The settling solids are measured using an Imhoff cone. If the wastewaters contain solids that settle after 10 min, their discharge is usually not permitted. The suspended solids are measured by passing a sample through a fiberglass filter; and determining the quantity by gravimetry, after evaporation of a known volume of the filtrate.

Introduction

Seafood provides about 25% of the global intake of animal protein and fetches a huge economy for developing nations. Many challenges concerning food safety and quality exist due to globalization and liberalization of the global f ish trade. International food safety regulatory bodies such as WTO and CAC through SPS and TBT Agreements work on ensuring food safety and quality. Adoption of risk-based analysis and implementation of the HACCP system are encouraged to maintain high seafood safety standards. International regulatory bodies set maximum residual limits for various environmental pollutants and contaminants to ensure consumer safety. Consumer education programs play important role to enhance awareness and improve transparency in the seafood supply chain. Internationally traded seafood products therefore need to meet the standards prescribed for food safety and quality requirements. In case of non-conformance, they could be detained, rejected and destroyed.

The global demand on seafood quality and safety requirement and the complexities of seafood supply chain make difficulty to ensure authenticity and traceability of seaood products in the market. Most seafood is exported from the country where it is caught while part of it via. a third country. Seafood pass-through complex supply chains involving, capture/culture, transshipment, processing, packaging, wholesale, retailing, and consumer. The global trade of seafood has led to a significant concern associated with seafood fraud.

12.1. Seafood Fraud

Seafood fraud is the practice of providing misleading information to the consumers about their product for financial gain. There are nine types of seafood fraud, which include species substitution, fishery substitution, IUU substitution, species adulteration, chain of custody abuse, catch method fraud, undeclared product extension, modern day slavery, and animal welfare

Introduction

Nutrition is the study of how food and nutrients affect the body’s health and well-being. It encompasses all aspects of food, from the food production, processing, consumption and digestion by the body. A balanced diet included all the essential nutrients such as carbohydrates, protein, fat, vitamins and minerals for maintaining good health. The recommended daily intake of nutrients varies depending on the age, gender and physical activity of a person. The nutritional need also varies depending on the lifestyle and medical condition of a person. The nutrition facts are based on recommended daily intakes for various nutrients, which vary based on factors such as age, sex, and physical activity. These facts are given on food packaging or label, providing important information on the serving size, calories, macronutrients and micronutrients, and other ingredients.

Nutrition labels are mainly intended to guide the consumer in food selection. Nutrition information provided on labels must be true and should not mislead consumers. Nutrition labelling regulations are provided by countries to help manufacturers. The Codex guidelines on Nutrition Labelling provide guidance to member countries to develop or update their national regulations and to encourage harmonization of national standards with international standards. According to it, no food should be described or presented in false, misleading or deceptive way. Nutrition labelling cannot solve nutrition problems, instead it is one of the elements of nutrition policy.

11.1. Nutrition Facts

Nutrition Facts means the nutritional information required for most of the packaged food in many countries giving the energy value and nutrients contained in the food. Nutrition facts labels are required by regulation similar to other types of food labels. It provides detailed information on the nutrient content of the food such as the amount of fat, sugar, sodium and dietary fiber.

Introduction

The international fish trade increases day by day reaching to a tune of USD 151 billion in 2020 (FAO, 2022). Many developing countries like India gain benefits from this trade as income through net foreign exchange reached to a level of 8.09 billion USD in 2019-20. Seafood supplied by developing countries contribute to 77% of fish consumption worldwide. The European Union (EU) is the single largest market for fish and fishery products to a tune of 91.74 billion USD in 2022 as a consequence of an increased per capita consumption among its Member States. Spain, Italy, Germany and the United Kingdom are the largest importer of fish and fishery products.

To protect consumer health and to protect the territory from the introduction of animal and plant diseases, the European Community (EC) has laid down joint conditions for imports of foodstuffs. The Directorate General for Health and Consumer Protection (DG SANCO) is responsible for food safety in the EU. The import rules for fishery products are framed of high standards with respect to hygiene, consumer safety and animal health status. It is important that importing countries and food business operators (FBOs) must understand the fundamental principles and requirements of the European Food Law to ensure efficient and smooth imports. The EU system relies on Government to Government assurances without any intervention of any private type certification or standards. The Competent Authority (CA) in the country of origin is responsible for giving the official guarantees to the EU regarding compliance and conformity to all requirements of its legislation. The Export Inspection Agency (EIA) is the Competent Authority in India for issuing the required official guarantees for export of fish and fishery products. The EIA maintains a complete list of all establishments under its control.

Introduction

Export of quality seafood products is important to have a success in international marketing. Exporters must be quality conscious and the Governments must insist on quality control and inspection. The seafood export industry in India is over 50 years old. Seafood contributes 10% of the total exports and nearly 20% of all agricultural exports. In the year 2022-23, India exported 17,35,286 MT seafood to more than 100 countries worth US $8.09 billion equivalent to Rs. 63,969.14 crore. There are over 150 fish processing facilities approved by the European Union out of the existing 350 processing facilities. The Export Inspection Council and the Marine Products Export Development Authority are working very closely with the fish processing industries for the growth of seafood exports.

The Government of India enacted the Export (Quality Control and Inspection) Act, 1963 for the development of export trade through quality control and inspection. It empowers the Government to notify the products intended for compulsory quality control and inspection prior to their export. It specifies the type of quality control required for the notified products. It recognizes standard specifications for a notified product. It prohibits the export of notified products that are not complying with export conditions. It creates a machinery for quality control and take measures to protect markets for Indian commodities. As the first step, Export Inspection Council (the Council) was established on 1st January 1964 for implementation of various provisions of the Export (Quality Control & Inspection) Act, 1963.

Introduction

In all countries, food is governed by a complexity of laws and regulations, which set out the Government requirements to be met by food chain operators to ensure food safety and quality. Food laws and regulations also govern standards that ensure the safety, quality, composition and suitability of food for consumers. Food laws and regulations of any nation depends on whether a nation adopts International norms developed by the Codex Alimentarius Commission (CAC) of the Food and Agriculture Organization (FAO) of the United Nations as well as the World Health Organization (WHO).

8.1. Food Laws

Food law means legislation that regulates the production, trade and handling of food. It covers the regulation of food control, food safety, quality and relevant aspects of food trade across the entire food chain, from the production to the consumer. Food laws protect the consumers against unsafe, adulterated and misbranded food and facilitate the movement of food across borders to sustain a fair trade.

Each nation administers its own laws, regulations and standards. It is also not static always and is constantly revised and updated in all nations. The food business operators (FBO) must obtain up-to-date information on any changes made in legislation, that are likely to affect the product standards. The regulations of all nations shall be obtained from their respective websites of the national ministries.

8.1.1. International Legislations

8.1.1.1. United States of America (USA)

In USA, the Food and Drug Act (1906) and the Federal Meat Inspection Act (1906) are the first federal food laws passed to protect American consumers. Later, Food, Drug and Cosmetic Act (1938) regulated the meat and poultry products. At present, Food Safety Modernization Act (FSMA), 2011 regulates all food products. https://www.fda.gov/food/guidance- regulation- food- and dietary- supplements/ food- safety- modernization- act- fsma. Manufacturers play important role in preventing food borne illness. The Act gave the Food and Drug Administration (FDA) the authority to oversee and enforce supply chains. The Act shifts the focus towards prevention of food borne illness and insists every FBO to register with the FDA.

Introduction

The risk assessment is gaining importance as the scientifically-based approach for the development of food safety and quality standards. It is a logical extension of HACCP revolution. In hazard analysis, first the hazards likely to occur in foods are identified, then an assessment is made of the severity of each hazard, and an evaluation is done of its likelihood to occur. Severity and likelihood are the two factors involved in risk assessment. Second, the increase in international trade has raised new safety and quality challenges. New approaches address the risk of cross-border transmission of infectious and hazardous agents with emerging food-borne diseases and quality problems. This has led to the development of a new safety and quality regulatory framework in 1995, the Sanitary and Phytosanitary (SPS) and the Technical Barriers to Trade (TBT) Agreements of the World Trade Organization (WTO). In these two Agreements, two provisions are of importance to fish safety and quality:

• National SPS and quality requirements should reflect standards agreed on in the international standards setting bodies i.e. Codex Alimentarius for food quality and safety.

• Domestic standards can be developed based on scientific risk assessment.

Risk assessment of microbiological hazards is identified as a priority area by the Codex Alimentarius Commission (CAC). In 1999, the Codex Committee on Food Hygiene (CCFH) in the 32nd session, identified a list of 21 pathogen commodity combinations that require expert risk assessment advice. FAO and WHO jointly launched a programme with the objective to provide an expert advice on risk assessment of microbiological hazards in foods to their member countries and to the CAC. Under which, expert groups are established to examine four priority pathogens: product pairings, e.g. Listeria monocytogenes in ready-to-eat food; Salmonella in eggs and broiler chickens; Campylobacter spp. in broiler chickens; and Vibrio spp. in seafoods. Risk assessment is therefore important throughout all aspects of the seafood industry - for companies, national governments and for international regulators.

Application of HACCP concept in surveillance & quality assurance programs for raw, frozen, canned, cured, irradiated, cooked, chilled, MAP & freeze dried products.

HACCP

Hazard analysis and critical control point (HACCP) is a system that identify, evaluate and control hazards that are significant for food safety (CAC, 2003). It is a science based systematic approach that assesses hazards and establishes control systems to focus on prevention rather than end product testing. It enhances the safety of the product from biological, chemical, and physical hazards in a production process. It provides way to demonstrate competence to consumers and compliance with legislative requirements.

6.1. History

HACCP was originally developed by Pillsbury Company in 1959 to produce zero defect food for astronauts of National Aeronautics Space Administration (NASA) in USA. In 1971, the HACCP concept was presented at the United States National Conference on Food Protection. The first major document on HACCP was approved by the National Advisory Committee on Microbiological Criteria for Foods (NACMCF) in 1989. The Codex then issued the first HACCP guidelines in 1993, which was adopted by the FAO/WHO Codex Alimentarius Commission (CAC). The USFDA’s Seafood HAACP program became mandatory in USA since 1997. The European Union, in 2004, laid down the general hygiene requirements of all food business operators (EC Regulation 852/2004) and the additional specific requirements for foods of animal origin, including live bivalve mollusks and fishery products which (Regulation EC 853/2004). The United States Food Safety Modernization Act 2011 became the latest enforcement of food safety measures to protect public health. Thus, HACCP has become an International reference system for food safety assurance.

Introduction

Quality is very important in any production and operation system. It ensures whether the products produced or the services provided are of the right quality in right quantity at right time. Quality management must ensure proper quality of the product to survive in the market and expand its market. Quality of the product must be given importance to avoid quality failures that cause serious human inconveniences and wastage of money.

5.1. Definitions and Terminologies

5.1.1. Quality

We say a product is of satisfactory quality, if it satisfies the consumers. According to C. D. Lewis, a British writer, quality is “an asset which is offered to the potential customer of that product or service”. The International Organization for Standardization (ISO) has put up a more comprehensive definition of quality as “the degree to which a set of inherent characteristics fulfils requirements” (ISO 9000: 2005 – 3.1.1). In general terms, ‘quality’ is defined as the composite of characteristics that differentiate individual units of a product to determine its acceptability by the customer. Quality characteristics are nothing but elements that makes a product fit for use. Quality characteristics are further classified into categories called as parameters.

5.1.2. Total Quality Management

Total Quality Management (TQM) is an organization’s management approach, centered on quality. It improves the effectiveness and flexibility of a company or an organization. TQM is defined as “the system of establishing defect prevention actions and attitudes with a company or an organization on a permanent basis for the purpose of assuring conforming products or services directed at customer satisfaction”. TQM is based on the participation of all its members in decision making, which is aimed for long-term success, through customer satisfaction and benefits to the members of the company or organization and to the society.

Introduction

“Quality” refers to the aesthetic appearance and freshness of fish. It also involves safety aspects such as being free from harmful bacteria, parasites or chemicals. The best quality fish is the one consumed within the first few hours after postmortem. We studied in our previous chapters that in some instances, even very fresh fish in rigor mortis are unsuitable for filleting. So, slightly older fish in post-rigor are more desirable for the processors.

The assessment methods of fish quality are divided into sensory, instrumental, chemical and microbiological. Sensory method is superior and all other methods must correlate with the results of sensory evaluation. So, sensory method must be done more scientifically under controlled conditions without the effects of test environment, personal bias, etc.

4.1. Sensory Methods

Sensory evaluation is defined as the scientific discipline used to evoke, measure, analyze and interpret reactions characteristics of fish, as perceived through the senses of sight, smell, taste, touch and hearing. There are few instruments capable of measuring sensory profile viz. Instron for texture, electronic nose for odour, microscopic imaging for structure, etc.

In sensory evaluation, appearance, odour, flavour and texture of fish are evaluated by the human senses. The process involves three steps: (1) detection of a stimulus by the human sense organs, (2) evaluation and interpretation by a mental process; and (3) the response of the assessor to the stimuli. The response of stimuli varies among individuals e.g. some people cannot taste rancid flavour and some have low response to cold-storage flavour.

Introduction

Seafood undergoes rapid spoilage after capture or harvest due to their highly perishable nature. Spoilage refers to any change that renders seafood unacceptable for human consumption. Spoilage occurs as a consequence of various microbial, biochemical and chemical breakdown processes. Spoilage leads to quality changes in seafood. Quality change refers to any change that occur after harvest due to unpredictable catch, uncontrolled harvest, inadequate handling, processing and storage facilities. The initial quality loss is mainly due to the post-mortem autolytic activity (enzymatic process) and chemical degradation process (lipid oxidation); and later by microbial degradation. The quality problems occur as a result of poor-quality raw material, ice and water, microbiological contamination, presence of heavy metals and other storage aspects during processing.

Processing of fish means preparing raw fish into a marketable form. Pre processing is done prior to final processing. Pre-processing is often done on board fishing vessel or in a shore based plant. It includes operation such as washing, sorting, grading, butchering, etc of the harvested fish. The butchering involves removal of non-edible portions such as viscera, head, tails and fins from fish. The yield of fish meat varies from 30 to 70% depending on the butchering process. Butchering process includes beheading, gutting, filleting, peeling, deveining, shucking, etc. The discarded portion of fish meat are fish wastes, often used for animal feed or for conversion to bioactive functional foods. Final processing of fish includes processes such as heating, freezing, drying, salting, marinading, irradiation, or addition of chemicals. The processed seafood is categorized into five major groups – chilled, frozen, canned, cured and other value-added products in this chapter.

Introduction

Quality of fish is affected by various factors, which are classified into two categories: intrinsic factors and extrinsic factors. Intrinsic factors are the chemical and physical properties inherent to fish. Extrinsic factors are treatments given to fish before and after catch until it reaches the consumer, which includes the method of catch, cross contamination, processing, preservation, and product storage environment (temperature, humidity, packaging). The processing methods such as drying, smoking, chilling, and salting greatly help to minimize the spoilage. The quality of fish is also dependent on cold chain, in which the temperature is managed and monitored at every stage of production, transport, storage and sale.

2.1. Pre-Harvest Factors

Pre-harvest factors relates to the intrinsic factors affecting the quality of fish, as they are beyond the direct control of human. Various intrinsic factors affecting quality are fish species, size, sex, physiological state, parasites, naturally toxic fish, pollutants, and occasional peculiarities. Additionally, some of the extrinsic factors like method of catch and feeding interruption for starvation are also considered as pre-harvest factors affecting the quality of fish.

2.1.1. Species Characteristics

The quality of fish is related to species. For instance, flat fish keep better than round fish. Post-mortem pH varies between species. For example, long rigor period and low pH (5.4-5.6) of very large flatfish and halibut give relatively long iced storage life of 21 days, while mackerel with low post-mortem pH has no effect on shelf life. Some fish are rated as good quality as they are expensive, example, seer fish and pomfret. Fatty fish are rejected sensorial before lean fish due to the appearance of oxidative rancidity. So, lean fish keep longer than fatty fish under aerobic storage. Bony fish are edible longer than cartilaginous fish. Skin of fatty pelagic fish is very thin and contribute to faster

spoilage rate, as it allows enzymes and bacteria to penetrate more rapidly. Thick skin and slime of flatfish contribute for better keepability, as slime contains bacteriolytic enzymes, antibodies and other antibacterial substances.

In this chapter, the importance of seafood in terms of their nutrition, and different methods of processing are dealt. Seafood quality and safety terminologies are explained briefly. The quality parameters affected by rigor mortis in raw, iced and cooked fish are given. Phenomenon of gaping described with reference to rigor mortis. Quality dimension of seafood is explained with intrinsic and extrinsic parameters. Various intrinsic factors affecting seafood quality such as f ish species, seasonal changes, parasite infestation and capture conditions are discussed in detail. The next part elucidates the sensory parameters affecting the seafood quality describing the objective and subjective method of quality assessments. Objective method covers both descriptive and discriminative tests. The final part mentions the Codex guidelines available for the conduction of seafood sensory assessment. At the end, the reader gains the knowledge on the basic information about seafood, role of rigor mortis in affecting seafood quality and the sensory quality assessment methods.

Objective

To learn the nutritional importance of seafood and their processing; rigor mortis and their role in seafood quality, intrinsic and extrinsic factors affecting seafood quality and sensory quality assessment methods.

1.1. Importance of Seafood

Seafood has been traditionally a part of diet in many parts of the world. In some countries like India, it constitutes the main supply of animal protein among coastal population. Seafood is a low-calorie food and easily digestible compared to other protein-rich meat and poultry products and hence, it is a good choice for elderly persons to obtain their daily protein requirements. Seafood has high-quality quality enriched with all the essential amino acids required for human health. It provides about 1/3rd of the average recommended daily allowance (RDA) of protein.

A laboratory is essential for research work in science and development. Most of the research institutes have dedicated laboratories for researchers and students to conduct experiments, perform research work, as well as to provide hands-on learning/training to students. There are certain rules and regulations laid down by the laboratory in-charge which need to be followed strictly for safety purposes. Some of the laboratories keep ‘safety manual’ for researchers to read before entering into the laboratory to conduct experiments. Whenever safety is compromised in a laboratory, there is increase in risk of unwanted incidents in the laboratory. Storing food/beverages in the same refrigerator where laboratory chemicals are kept may lead to disastrous effects on health. This chapter aims to provide instructions in brief on the things to be performed (Do’s) before and after entering onto a laboratory as well as the things never to be done (Don’ts) in a laboratory for the safety reasons.

Good Laboratory Practices (Do’s)

• P Wear apron/labcoat while working in the laboratory.
• P Use gloves/respiratory mask/fumehood while handling toxic/volatile/inflammable chemicals/reagents.
• P Use pippet or pippetor for safety in transfer of solutions.
• P Protect eyes, face and naked parts of the body from UV radiations. Use safety goggles or face mask/apron.
• P Keep your eyes away from Liquid Nitrogen and use cryogloves while handling it.
• P Arrange all the necessary materials/solutions before starting the experiment.
• P Periodically fumigate/ozonize culture and inoculation rooms.
• P Autoclave all contaminated materials before discarding.
• P Pre-treat gel or solution containing Ethidium Bromide (EtBr) before discarding into laboratory waste-disposal bin, and use gloves while handling them.
• P Burn all the transgenic plant materials in an incinerator after taking necessary observations.
• P Make entry in the designated log-book and clean the instrument after using it properly.
• P Switch-off the instrument after use or when not in use.
• P Wash hands with soap and remove the labcoat before leaving/going out of the laboratory7.
• P Ensure that switches for fans/lights, and water taps are off while leaving the laboratory.
• P Follow the biosafety guidelines suggested for handling and disposal of the hazardous chemical.

Introduction

Several hazardous chemicals, microbes, and techniques like recombinant DNA technology are used for research and development work in research Institutes and universities. Therefore, the researchers, technical, helping and supporting staffs as well as the laboratory outputs must be kept free from contaminations. Thus, biosafety is necessary to prevent unintentional exposure/ release of hazardous chemicals, microbes/pathogens, and toxins to the laboratory staff and environment. Following the biosafety guidelines and ethical considerations are essential to maintain safety standards for the researchers as well as the environment. This chapter deals with various biosafety measures and ethical issues to be strictly followed while working in a research laboratory. The required safety level needs to be determined as per the chemical/biological materials are used in the laboratory to keep the researcher/environment safe for sustainable research and developmental activities.

Biosafety

The term biosafety is used to describe the procedure and policies needed to ensure environmental and personal safety. Biosafety refers to the containment principles, technologies and practices that are implemented to prevent unintentional exposure to pathogens, toxins, or their accidental release into the environment. A fundamental objective of any biosafety program is the containment of potentially harmful chemical/biological agents as precautionary safety measures.

Why Biosafety?

With the increasing number of countries adopting molecular biology tools and techniques in life science research and development activities, especially in the areas of agriculture and medicine, the biosafety issues are gaining importance to ensure biological safety for the public and the environment. Conducting research in safe and sustainable manner is not only a personal requirement but essential collective efforts to ensure biological safety for a clean and safe environment. This requires rules, regulations, monitoring bodies, and awareness among the workers as well as the public. Therefore, biosafety per se is an integral part of laboratory research and requires awareness among the researchers so that biological safety can be well taken care from the grassroot level.

For a researcher, laboratory is another home where he/she spends much time every day. As we take care of cleanliness, safety, and design of our home, we must take care on these aspects of laboratory too. It is the responsibility of the laboratory personnel to ensure cleanliness/safety of the laboratory. Since every laboratory chemical and equipment have the potential to cause harms if not used with appropriate precautions. Hence, as a laboratory user, it is our responsibility to follow the basic safety/biosafety rules and regulations set up by the Principal Investigator of the laboratory, Institutional Biosafety Committee, as well as by other regulatory authorities, if any. We must be aware of the general/personal protective measures, cleanliness and laboratory waste disposal prior to joining a research laboratory. In this chapter we will briefly discuss about personal safety, chemical safety, cleanliness, waste disposal, etc.

A. Personal Safety

1. Personal Protective Equipment (PPE)

Personal protective equipment (PPE) is special arrangement to protect the worker from specific hazards because of using a hazardous substance. PPE does not reduce or eliminate the hazard, but protects the wearer from exposure to hazardous material. PPE should not be reused or brought outside the laboratory as these might increase the risk of spread hazardous material or contamination. Used PPE must be kept inside the laboratory in a designated waste storage area adjacent to the work area.

Introduction

Proper electrical wiring and earthing are crucial for laboratory safety and functionality. A laboratory houses several installations/equipment that demand stable/continuous power supply. Inappropriate or faulty wiring and earthing may lead to significant risks, including electrical shocks, damage to equipment, and even incidence of fire. Therefore, understanding the requirements of electrical wiring and earthing is essential for designing, construction, and maintenance of laboratory. This chapter aims to provide a basic knowledge of electrical wiring and earthing necessary for a laboratory. Importance of earthing in mitigating electrical hazards and ensuring integrity of electrical installations are also described.

Basics of Electrical Wiring

Basic understanding of electrical wiring is necessary for laboratory safety and efficient electrical functioning. The potential difference between two electrical points (negative and positive) is known as Voltage, which drives the flow of electric current. This is measured as volt (V). The rate of flow of electricity in a circuit is known as Current (I), which is measured as ampere (A). Current can be either direct (DC) or alternating (AC). Opposition/resistance experienced in the flow of current in circuit is known as Resistance (R) and it is measured in ohms (Ω). Higher the resistance means lesser the current flows at a given voltage. The rate at which electrical energy is transferred/utilized in an electric circuit is known as Power (P), which is measured in watt (W). The Power can be calculated as the product of voltage and current (P = V × I).

Introduction

We come across a variety of chemicals every day in a laboratory. While some of them are relatively safer, others are extremely hazardous. Thus, it is very important to know the potential hazards that can be caused by the chemicals present in the laboratory and the safety measures needed to be followed to avoid the hazards. Additionally, we should also be aware of the steps necessary to be taken on such hazardous situations. According to IUPAC, a hazardous chemical mean the chemical which has the potential to cause damage in various ways. In a laboratory, there are chemicals that might be irritant, carcinogen, respiratory fibrogen, systemic poison, asphyxiant, flammable, etc. which have the potential to cause hazard to the workers or environment. Therefore, it is important to use appropriate procedure to reduce the risk and protect the health/safety of laboratory personnel, the public and the environment.

Types of Hazards Associated with Chemicals

Chemical hazards: The hazards caused by chemicals can be broadly classified into three categories.

1. Health hazard: This is also known as toxic effects of chemical. The risk associated with a chemical in laboratory depends on the extent of exposure and the inherent toxicity of the chemical. Many chemical reactions form the product that might be much more toxic than the reacting chemicals. For example, inadvertent mixing of formaldehyde (a common tissue fixative) and hydrogen chloride results in the generation of bis(chloromethyl) ether, a potent human carcinogen. For most of the chemicals, the route of exposure (through skin, eyes, gastrointestinal tract, or respiratory tract) is more important (Figure 17.1) that must be considered during risk assessment.

Introduction

Flowering plants (Angiosperms) are identified by their morphological characteristics like flower, fruit and seeds that help their taxonomic characterization into different species, genera, families and orders. The floral characters which are used for taxonomic characterization of plants include the type of inflorescences, perianth structure, floral symmetry, arrangement of floral leaves, type of androecium, stamens, gynoecium, carpel, ovule, etc. The type of fruit and seed also play important roles in taxonomic classification. In this chapter we will briefly discuss about various morphological features of flowering plants (in botanical terms) that are useful in taxonomy.

Nomenclature of Plant

Nomenclature of plant is the formal and scientific naming of plants. It is related to but distinct from taxonomy for grouping and classifying plants and provide botanical name.

The scientific/botanical nomenclature started with Species Plantarum by Carolus Linnaeus in 1753. Botanical nomenclature is governed by the International Code of Nomenclature for algae, fungi, and plants (ICN), which replaces the International Code of Botanical Nomenclature (ICBN).

ICBN and ICN

International Code of Botanical Nomenclature, (ICBN) was adopted by the VIIth International Botanical Congress held in Stockholm, July1950. While the International Code of Nomenclature for algae, fungi, and plants (ICN) was adopted by the XVIIIth International Botanical Congress held in Melbourne, Australia in July 2011.

Introduction

Test of pollen viability (extent of pollen viability) is important in pollination experiments to ensure fertility/seed setting. It is also important in monitoring state of pollens during storage in ecological/ taxonomic studies as well as during research on pollen biochemistry, genetics, stigma interactions, and incompatibility studies. Pollen viability refers to the ability of pollen to accomplish post-pollination events to achieve fertilization. Pollen viability test has gained importance particularly in plant breeding and hybrid seed production experiments/ practices. In this chapter, you will be acquainted with the basics of pollen viability test. However, for more professional knowledge on the topic readers are suggested to refer specialized books.

Pollen Viability Test

It is important to know the extent of viability of pollens used in pollination experiment, in finding fertility of a plant of interest, to monitor state of pollen on storage, in ecological/taxonomic studies, as well as in research on biochemistry, genetics, stigma interaction, incompatibility assay, etc. of pollen. Several pollen viability tests have been developed time-to-time. It is recommended to initially use different tests for a given pollen type to find out the most appropriate method that provides reliable result on pollen viability. It is important to note that every procedure requires standardization of protocol according to the sample used. Following different tests are used for assessing pollen viability.

1. In-vitro Pollen Germination Test

It is a simple, speedy, and quantitative test for assessing pollen viability. In this method, a collection/sample of pollen is germinated in vitro and observed under microscope. The percentage of germinating pollens (producing pollen tube) after appropriate treatment and a given period of time is determined. The percentage provides an index of viability of the pollen sample.

Introduction

Seed is a living entity (mature ovule) consisting of a tiny plant in embryonic stage along with the stored food protected by an outer layer known as seed coat. Seed is an important biological input in agriculture, which is economically important for successful crop husbandry. Cotyledons serve as the source of nutrients (proteins, carbohydrates, and oils) that not only nurture the plant at the early developmental (seedling) stages of plant but also the vegetarian (heterotroph) organisms. Being one of the important inputs in agriculture, viability of seed needs to be assessed to know the germination percentage of seeds used to raise a crop. Since viability of seeds may deteriorate on storage depending on the storage conditions. Therefore, often it becomes important to test viability/germinability and vigour of seeds before being used for raising a crop or conducting an experiment. In this chapter, we briefly discuss about the different methods which can be used for testing the viability of seeds.

Seed Viability

Seed viability can be defined as the capability of the seed to germinate and produce a normal seedling within a specified period. In other words, seed viability indicates how many seeds in a lot are alive and could develop into plants under appropriate field conditions. It is very important that most of the stored seeds are capable of germinating when sown in a field. They must possess higher viability at the beginning of storage and maintain it over the storage. A seed with high initial viability survives longer in storage; however, the viability deteriorates slowly at first and then rapidly as the storage prolongs. It is important to know the duration of storage after which a considerable decline in viability occurs under given storage conditions so that to take necessary action to revive the seeds. Excessive deterioration in vianility of seeds may lead to loss of the seed/material/ germplasm.

Introduction

Tissue culture refers to the technique of growing cells, tissues, or organs under controlled, artificial conditions. This allows propagation of tissues from a small sample of cells/tissues under sterile conditions. Plant tissue culture is widely used for various purposes, including the production of disease-free plants, rapid multiplication of plants with desirable traits, genetic modification, conservation of rare or endangered species, etc. Tissue culture is a valuable technique because it enables consistent and large-scale production of plants, ensures genetic uniformity, and allows manipulation of plant biology at cellular level. In this chapter, we will discuss about the major types of tissue culture techniques and their advantages, disadvantages, as well as applications in brief.

Different Types of Tissue Culture

Callus culture involves induction and growing of unorganized, undifferentiated cell masses from an explant on a nutrient/culture medium. The part of plant used to induce callus is called explant which can be tissue, organ or part of a plant.

Organ culture is used to maintain or grow an entire organ or a part of organ in vitro. It preserves the structure and function of the organ for studies or transplantation.

Embryo culture involves isolating and growing embryos from seeds or ovules in a nutrient medium. It is used to bypass seed dormancy or rescue embryos from hybrid crosses that may not develop naturally.

Cell suspension culture involves growing dispersed cells or small cell clusters in a liquid medium under continuous shaking. This is used for large-scale production of cells, secondary metabolites, and studying cell biology..

Introduction

Different types of culture media are used for culturing microbes and plant tissues. Preparing as well as sterilizing them properly following appropriate method with safety for the desired purpose are important parts of research work. In this chapter, we will discuss briefly about different types of media, their composition, preparation and sterilization in laboratory for microbial or plant tissue culture.

Culture Medium

Ingredients mixed to prepare solid, liquid or semi-solid substance to support the growth of microorganism or plant tissue in vitro (outside the body) is called culture medium. Frequently, it becomes important to grow microorganism or plant/animal tissue in vitro for any or all of the following purposes:

• To study characteristic features or properties of microorganisms.

• To identify the causal organism(s) of infection in a clinical sample, so that appropriate remedy can be determined.

• To culture/multiply microorganism or plant/animal tissues in vitro for research or commercial purposes.

• To prepare/produce biological molecules, vaccines, antigens, proteins, metabolites, etc. in vitro for research or industrial purposes.

Introduction

Use of acid/base is common in laboratories for experiments like titration and proper safety in their handling laboratory is necessary. In this chapter we will briefly discuss about the safety measures necessary to be adopted while using acids/bases in laboratory for any experiment.

Handling of Strong Acid/Base

There are four major points that need to be taken care of while working with strong acids or bases that might be quite harmful if any accident happens.

1. Protect Yourself

Wear a lab-coat and acid-resistant gloves: Always use lab-coat/apron and make it sure that the sleeves properly cover your arms. More importantly, use gloves made of acid-resistant materials and also wear shoes to protect your feet while wearing long-pants/jeans. Female with long hair, must tie her hair properly (with rubber band) to avoid any disturbance caused by appearance in between.

Wear safety goggles: It is important to wear safety goggles to protect your eyes using larger and fitting goggles that cover front as well as sides of your eyes. Always use properly fitting and adjustable goggles because smaller/ loose goggles may not protect the areas around your eyes against accidental acid spills and it may fall down from your face, if it is too loose.

Locate emergency wash area: There must be an emergency washing area nearby the work place or laboratory containing lab spill management kits. All the workers in the laboratory must be aware of using the kits and emergency plans.

Neutralize acid: Don’t panic even if an acid or chemical spill happens. Stay calm and act as quickly as possible. In case an acid spills, neutralize it by using sodium bicarbonate so that they can be cleaned and disposed safely. If acid comes in contact of skin, wash it off under running tap water but do not use/ apply sodium bicarbonate on skin.

Introduction

Measurement of pH of a solution and preparation of different types/strength of buffers are common practices in a laboratory, particularly in the Biochemistry and Molecular Biology laboratories. Therefore, in this chapter you will be acquainted with the procedures of measuring pH of a solution as well as preparation of some of the commonly used buffers in a laboratory.

Potential or Power of Hydrogen Ion (pH)

Measurement of pH of a solution is measuring the molar concentration of hydrogen ion [H+] in the solution, which is an indication of the acidity or basicity of the solution. The symbol pH stands for “power of hydrogen” and numerical value for pH is just the negative logarithm of the molar concentration of hydrogen ions [H+]. The ionic product of water (Kw ) is the basis for the pH scale. The total hydrogen ion concentration from all the sources and it can be experimentally measured.

The pH scale designates the concentration of H+ (and thus of OH−) in an aqueous solution, which ranges between 1.0 M H+ and 1.0 M OH−. The term pH is defined by the expression

Thus, the ionic product of water makes it possible to calculate the concentration of H+, with given concentration of OH−, and vice versa. The value of 7 for pH of a neutral solution is not an arbitrarily chosen figure; it is derived from the absolute value of the ionic product of water. A solution having pH greater than 7 is basic, and the concentration of OH− is greater than that of H+. A solution having a pH less than 7 is acidic.

Introduction

For agricultural applications as well as for research work, often we need to prepare solutions of different agrochemicals of varying strength, as per the requirements. Agrochemicals are used to help crop plants grow, produce, and protect them from biotic/abiotic stresses. Many of the agrochemicals (insecticides, herbicides, fungicides, rodenticides, molluscicides, nematicides, etc.) are used to protect the crop from insect pests and diseases. However, some of them (fertilizers and soil conditioners) are used to augment crop yield. While in the previous chapter (Chapter 8) we discussed about preparation of laboratory solutions, in this chapter we will briefly discuss about the preparation of different agrochemicals.

Agrochemical/agrichemical is a generalized term used for the chemicals applied in agriculture. In general, it also includes the feed-supplements used in f isheries and animal husbandry.

Agrochemicals

Fertilizers

Fertilizers are used to augment growth of crop plants and to mitigate nutrient deficiency in the soil. Fertilisers can be organic (containing natural substances, prepared through natural processes) and inorganic fertilisers (synthetic fertilizers, manufactured artificially using chemical processes using natural deposits, by chemical alterations).

Soil Conditioners

Soil conditioners include manure, livestock and leaf compost, peat that help making the soil in good condition. They are laid on the top soil and then mixed properly. They not only increase nutrient contents in the soil but also improve aeration and water holding capacity.