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Different enzymatic methods have been proposed as potential alternatives to rumen inoculum studies to determine dry matter digestibility and protein degradation. However, the type of enzyme used and the procedure adopted differ (Marten and Barnes, 1979). The enzyme methods offer distinct advantage under circumstances of the difficulties in maintaining rumen fistulated animals as a source of rumen inoculum. Since, the enzymes used in these techniques are commercially available enzyme preparations, and are not of rumen microbial origin, the digestibility obtained from these techniques are related to in vivo digestibility by means of mathematical equations. Therefore, equations developed to predict energy content using enzyme techniques differ with the type of feedstuff and type of enzymes used in the technique (Table 5.1.) (Martens and Barnes, 1979). The enzyme methods are popular among the feed compounding industries to evaluate raw materials and compound feeds.

The method used by DeBoever et al. (1986) to determine digestibility of mixed feeds is given below.

The main groups of some diagnostically important enzymes are as below:

1. Oxido Reductases
Glucose 6-phosphate dehydrogenase, Isocitrate Dehydrogenase, Lactate dehydrogenase, Malate dehydrogenase, Sorbitol dehydrogenase.

2. Transferases
Alanine transaminase (GPT)    
Aspartate transaminase (GOT)
Creatin binase
Hexose-1 phosphate uridyltransferase

3. Hydrolases
Acetylcholinesterase
phosphatases
Amylases
Pepsin
Trypsin
Lipase

Introduction

During last four decade poultry has taken giant step and become corporate sector with double digit growth. Especially in India poultry play pivotal role in fight against malnutrition, unemployment and also contributing in women empowerment. In poultry production the biggest single expense is of feed, which accounting for 70% of total production cost per bird (Kumar et al., 2011). So, obviously more Research and Development focus and new innovative technology need to be implemented in poultry nutrition area. Since last 40 years, feed enzymes have played a major part in helping radically to improve the efficiency of meat and egg production by changing the nutritional profile of feed ingredients. 

Introduction

Billions of chemical reactions are happening inside human body and that too with lightning speed. However, the same reactions if carried out in laboratory, they take a very long time. The reasons behind it are the biocatalysts, also known as enzymes, which are synthesized by living cells. Chemically, they are proteins(except ribonucleic acids: RNA), colloidal in nature, thermo-labile in character and very specific in their action. For example, hydrolysis of proteins by a strong acid at 100°C takes a couple of days; but same proteins can be digested within couple of hours inside human body at 37°C. Similarly, carbonic anhydrase is used as a catalyst by living cells to transfer carbon-di-oxide from cells to blood. This process is 107 times quicker than the same process if occurs without the help of enzymes. Therefore, enzymes can be defined as the biomolecules or biocatalysts which can speed up the rate of the chemical reactions. The substance upon which an enzyme acts is called as substrate and the one which is produced as the result of enzyme-substrate reaction is known as product.

Veterinary epidemiology is the study of disease within a livestock population, dealing with the investigation (including identification, quantification, and examination), of the direct and indirect determinants of disease distribution in animal populations. Any epidemiological investigation starts with whether there is an animal health problem in a population, which requires data collection on animal, management, economic, and environmental factors. These data are then analyzed descriptively to assess the frequency of disease occurrence. The results are then used to decide whether investigations of causal relationships between disease occurrence and potential risk factors are necessary for the development of control or prevention policies. Once the relationships are understood, necessary interventions for the disease are then planned and their impacts assessed. Hence, the goal of epidemiological investigations is the control and prevention of animal disease problems by integrating the available pieces of evidence into the decision-making process for disease control policy development.

Equines are herbivorous monogastric animals. Though, simple-stomached, they can digest considerable amount of fibrous feed in the caecum and are called as hindgut fermenters. Equine includes horse, donkey, zebra and mule which are used as work and pack animal. All three species of equine belongs to the family Equidae. The scientific name of Horse and Donkey is Equus caballus and Equus asinus, respectively. The horse was probably last farm animal domesticated by the man. Horses appear to have been domesticated first in central Asia or Persia, more than 3000 years ago.

Entrepreneurship theory has been evolving over the last 20 years and is ever growing. It is defined as a verifiable and logically coherent formulation of relationships, or underlying principles that either explain entrepreneurship, predict entrepreneurial activity or provide normative guidance (prescribing the right action in particular circumstance). Entreprenuership is interdisciplinary and contains various approaches that would increase one’s understanding of it. One way to examine these theories is with a ‘schools of thought’ approach that divides entrepreneurship into specific activities. These activities may be within a ‘macro view or a micro view’, but all address the conceptual nature of entrepreneurship.

Development of entrepreneurship

Creativity: Creativity and innovation are often used to mean the same thing, but each has a unique connotation. Creativity is the ability to bring something new into existence. Ideas usually evolve through a creative process whereby imaginative people bring them into existence, nurture them, and develop them successfully. The creative process for an idea contains five stages – germination, preparation, incubation, illumination, and verification.

Introduction

Public concern over use of antibiotics in livestock production has increased in recent years because of their possible contribution to emergence of antibiotic resistant bacteria, and their transmission from livestock to humans. Accordingly, ruminant microbiologists and nutritionists have been exploring alternative methods of favorably altering ruminal metabolism to improve feed efficiency and animal productivity. Plant extracts contain secondary metabolites, such as essential oils (EO) that have antimicrobial properties that make them potential alternatives to antibiotics to manipulate microbial activity in the rumen. Essential oils are naturally occurring volatile components responsible for giving plants and spices their characteristic essence and color. Over the last few years, a number of studies have examined effects of EO, and their active components, on rumen microbial fermentation. However, many of these studies are laboratory based (i.e., in vitro) and of a short-term nature. Nevertheless, results from in vitro batch culture studies provide evidence that EO and their components have the potential to improve N and/or energy utilization in ruminants. Effects of EO on ruminal N metabolism is more likely mediated by their impact on hyper-ammonia producing (HAP) bacteria resulting in reduced deamination of amino acids (AA) and production of ammonia N.

Extent of Passage
(Method of Krishna and Ekern, 1974D)
Definition of Terms
The term  extent of passage is used to describe how far a given ingested material travelled  or was allowed to pass. The point at which  undigested residue of the food comes to or has passed through the alimentary tract may be found by the use of markers without disturbing the subject. In some conditions of digestive system malfunction or disorder , the food residue may be blocked at point along with digestive tract.

Transit Time
This is the time it takes the digesta of a meal to pass through the alimentary tract or segments of it. This time also represents the  retention time in the tract or the particular segment. A simple way of calculating it is by recording the time of first or last appearance of the marked residue of a meal. Other more useful methods for calculating retention time include “mean retention time” (Castle 1956a), “mean time” (Blaxter et al., 1956) and “turnover time” (Hungate, 1968).

Principle

The portion of the total ash which is insoluble in dilute hydrochloric acid is called acid insoluble ash (AlA) and it is the part of ash which is having unavailable mineral matter like silica and sand. Silica is the part of mature plant materials which are highly lignified or woody like straw, husk, stems and stalks, while sand inters in feeds by faulty handling in the fields or as an adulterant.

Equipment/glass wares
    1.    Asbestos sheet
    2.     Filter paper (Whatman No.1)
    3.    Funnel stand
    4.    Glass funnel
    5.     Glass rod
    6.     Hot plate/heater/Bunsen flame
    7.     Metal tongs
    8.     Silica basin (crucible)
    9.     Volumetric flask
    10.     Wash bottle with distilled water

Nutritional significance of tannins in feeds and fodders

Tannins are widely distributed in many types of browse plants, tree leaves, forages and feed ingredients. The structure of tannins are not completely well defined but they are polymers of phenolic and aliphatic hydroxy compounds. Although tannins and related polyphenols are considered as protein binders and, protease and cellulolytic inhibitors, they can also exert beneficial effects on rumen fermentation. There are two forms of tannins viz., hydrolysable and condensed. Hydrolysable tannins are considered to have less deleterious effects on protein digestion since these types of tannins might get hydrolysed under acidic gastric condition and release bound proteins. Attempts have been made to utilize this to advantage in protection of proteins from rumen degradation.The condensed tannins are resistant to hydrolytic degradation. The reduction in digestibility due to condensed tannins is not clear whether it is due to specific interference with cellulases or precipitation of proteins.

Certain feed ingredients have been incorporated as a source of tannins, in the diets of ruminants to benefit ruminal fermentation and performance. Inclusion of Bengal gram husk as a source of tannins in the diet of sheep prevented potentially digestible carbohydrate in the rumen, but digested post ruminally (Sreerangaraju et al., 2000).

Neem seed cake (NSC) despite of being rich in protein is not used as animal feed due to the presence of large number of toxic/bitter compounds in it. Studies have shown that NSC can be safely included the ration of animals after water washing / alkali/ urea treatment. These treatment methods appear to remove / reduce or largely inactivate the toxic principles present in NSC. In order to assess the quality/safety of the treated product, quantification of the residual toxic compounds is essential. One such compound is azadirachtin. It has been reported to be biologically most active.

One of the best techniques for the estimation of azadirachtin is by using high performance liquid chromatography (HPLC).

Estimation of calcium and phosphorus in feeds: Calcium is precipitated as calcium oxalate by adding ammonium oxalate to acid soluble extract prepared after running a sample for ash. Precipitates of calcium oxalate are dissolved in sulfuric acid and titrated with potassium permanganate to know the content of calcium.
Using N1V1 ≡ N2V2, 1.0 ml of 0.1 KMnO4 = 2.004 mg Calcium.
Calcium, g % = Burette reading difference X 2.004 X dilution factor X 100 / weight of the sample used for preparation of acid soluble mineral extract X 1000.
[dilution factor, (for example) = 250 / 25 (ml aliquot taken) = 10]
Phosphorous present in the sample is precipitated with ammonium molybdate and the precipitates are dissolved in NaOH solution. Excess sodium hydroxide is titrated with nitric acid to calculate the actual volume of NaOH used up which is direct proportion to the Phosphorous content.
1 ml of 0.1 N NaOH = 0.1347 mg Phosphorus
Phosphorus % = Actual volume of 0.1N NaOH solution used to dissolve precipitate X 0.1347 X dilution factor X100 / weight of sample used for acid soluble extract X 1000
[Here, dilution factor is 250 / 50 (aliquot taken) = 5]

Principle

When a solution containing calcium is treated with ammonium oxalate, all the calcium present is precipitated as calcium oxalate. The precipitate when dissolves in sulphuric acid, forming calcium sulphate and liberating free oxalic acid which is quantitatively estimated by titration against standard N/10 KMnO₄ to arrive at the calcium content present in the given solution. Following chemical reactions takes place in the estimation of calcium:

I. Titrimetric Method of Estimating Salt in Fish Meal
(Modified AOAC, 1965 method)
Principle

The salt content is calculated as sodium chloride from the content of water solubles chlorides. The chlorides are dissolved in water and directly precipitated by titration with a silver nitrate solution using potassium chromate as an indicator.

Principle

When a sample of moisture and fat free feedstuff is boiled with dilute acid and then with dilute alkali, the soluble carbohydrates and proteins go into the solution and are extracted. The undissolved residue left behind represents cellulose, hemicellulose, lignin and mineral matter. The cellulose, hemicellulose and lignin put together are called crude fibre (CF). When the residue after drying and weighing is ignited, the fibre being organic in nature is burnt whereas the ash or mineral matter is left over. The fibre is obtained by deducting the weight of ash from the weight of the dried residue. In simple way it can also be explained that the crude fibre is determined gravimetrically as part of the substance which is left when protein, soluble carbohydrate, fat, ash and water have been removed.

In feeding of simple stomached animals, the analysis of crude fibre content is very essential because the amount of crude fibre predicts the digestibility of the feedstuff. Higher the fibre content lesser will be the digestibility.

Principle

Crude protein constitutes a mixture of true protein and non protein nitrogenous substances and is determined by estimating nitrogen and then multiplying the nitrogen content of feed by the factor 6.25.

The factor 6.25 is derived on the basis that all proteins contain 16% nitrogen. The total nitrogen content in the feedstuff is determined by Kjeldahl method which generally consists of three steps, i.e. digestion, distillation and titration. The sample is digested with an excess quantity of concentrated sulphuric acid which hydrolyses the protein, decomposes to ammonia and finally the ammonia combines with the excess of sulphuric acid forming ammonium sulphate. The acid digest is cooled, diluted with distilled water and distilled with strong alkali (sodium hydroxide, 40 per cent), the ammonia released during distillation is taken up into a boric acid solution forming ammonium borate complex and such ammonia is titrated with standard sulphuric acid. While estimating the nitrogen content, following reactions take place:

(Method of Visek et al., 1953; Lueker and Lofgreen, 1961)

By using isotope dilution technique, It is possible to separate faecal calcium into endogenous and exogenous fractions. The true digestibility can be measured independently by making correction for faecal endogenous loss. The isotope dilution method described by Visek et al., (1953) involves daily intravenous injection of 45Ca for 15 days, followed by a collection of the serum and the faeces for 15 days. However, these numerous veinpunctures result in considerable discomfort to the animal. Lueker and Lofgreen (1961) modified this method in sheep by giving a single subcutaneous injection of 45Ca, followed by a collection of blood and the faeces for 7 days only.

The endogenous faecal calcium and true digestibility of calcium is calculated by the formula of Lofgreen and Kleiber (1954) as given below:

Principle
When a sample of feed, which is free from moisture, is extracted continuously with fat solvent like petroleum ether, the vapour from the ether boiling in the flask passes into the condenser where it condenses and drops back on the sample which dissolves ether soluble materials such as fat, sterols, lecithin, resins and volatile oils in the extraction flask. The extract is termed as crude fat or ether extract (EE) because it also contains other fat like substances in addition to true fat.

Equipment/glass wares
    1.     Analytical balance
    2.     Beaker
    3.     Cotton wool
    4.     Dessicator
    5.     Extraction thimble
    6.     Hot air oven
    7.     Oil flask (150-250 ml)

Principle
If a sample of feed is heated and dried in a hot air oven at the temperature of boiling water (100°C ± 2°C), the water present in feed sample escapes as vapour. The loss in weight is considered as moisture and the residue is termed as dry matter (DM).

Requirement
    1.    Aluminium/steel moisture cup or Iron trey
    2.     Analytical balance
    3.     Dessicator
    4.     Hot air oven
    5.     Metal tongs
    6.     Spatula