1

CHAPTER I

INTRODUCTION

Nature and Importance of the Study

In Philippines, swine industry contributes a major economic impact on agriculture.

In fact, according to DOST-PCAARD, the swine industry is the second largest contributor

to the country’s agriculture coming in second to rice— contributing a significant 2.0% to

2.8% of the total value of the national GDP. The total inventory of swine as of July 2019

was estimated at 12.70 million heads according to Philippine Statistics Authority. With this

huge number of pigs, the constant need of inexpensive and efficient feeds has been

underscored as key aspect in selecting feeds on an array of feed brands in the country. By

that virtue, companies constantly formulate nutritional diets through extensive research

which goal is to seek the proper and most ideal nutritional value for pigs could have — to

give the animal a healthy growth which maximize the profit of the company. Evaluation of

the stated nutritional contents of the feed products can be done through proximate analysis.

Proximate analysis is a series of nutritional analyses that performed to seek the estimate

percentage of the different components of feeds such as moisture, ash, crude fat, crude

protein and crude fiber. It is of crucial commercial concern as food-manufacturing

companies need to ensure that their products meet the appropriate laws and legal

declaration requirements as well as the safety aspects of the end products when released to

the end consumer.

The moisture content analysis is one of the most important and most widely used

measurements in the processing and testing of foods (Bradley,2010). Since the amount of
 2

dry matter in a food is inversely related to the amount of moisture it contains, moisture

content is of direct economic importance to the processor and the consumer. Of even

greater significance, however, is the effect of moisture on the stability and quality of foods.

Feeds that contains too much water is subjected to rapid deterioration from mold growth,

heating, insect damage, and sprouting (Henderson, 1985).

The analysis of ash content in foods is simply the burning away of organic content,

leaving inorganic minerals (Sluiter et al., 2005). This helps determine the amount and type

of minerals in feed; important because the amount of minerals can determine

physiochemical properties of foods, as well as retard the growth of microorganisms

(Marshall, 2010). Therefore, mineral content is a vital component in a food’s nutrition,

quality and, like water, microbial viability.

Soxhlet extraction is one of the most commonly used methods for determination of

total fat. This is mainly because it is simple to use and is the officially recognized method

for a wide range of fat content determinations (AOAC, 1990). Fats or lipids (crude fat) are

a very diverse group of chemicals, but they have one similarity in that they dissolve in non-

polar solvents and are not soluble in polar solvents such as water (Ellefson & Min, 2010).

Foods contain many types of lipids, but those which tend to be of greatest importance are

the triacylglycerols and the phospholipids. Dietary lipid has two main functions - as a

source of energy and as a source of its component fatty acids, some of which are essential

(i.e., cannot be synthesized by the animal itself) dietary components for the growth and

survival of the recipient animal (Ellefson & Min, 2010).

The Kjeldahl method for determining the nitrogen in organic compound is used

commonly in the analysis of crude protein (Saez-Plaza et al., 2013). Proteins have a major
 3

role in the growth and maintenance of the human body (Chang, 2010) and are, along with

carbohydrates and lipids, the energy giving nutrients in the diet (Dalheim et al., 2018). In

addition, proteins also pose a wide range of other functions in the body, such as enzymatic

activity and transport of nutrients and other biochemical compounds across cellular

membranes. Inadequate intake of dietary proteins containing essential amino acids results

in increased turnover of muscular proteins, leading to reduced growth and loss of muscle

mass. Impaired immunity, as well as reduced hormonal and enzymatic activity may

subsequently follow (Dalheim et al., 2018).

The crude fiber method was developed to estimate indigestible carbohydrate in

animal feeds (Möller, 2014). Crude fiber is a measure of the quantity of indigestible

cellulose, pentosans, lignin, and other components of this type in present foods. Adequate

consumption of dietary fiber from a variety of foods will help protect against colon cancer

and also help to keep blood lipids within the normal range, thereby reducing the risk of

obesity, hypertension, and cardiovascular disease in general (BeMiller, 2010).

Objectives of the Study

The objective of the analysis was to evaluate the nutritional content of the Nutri-

Start Pellet feed sample such as moisture, ash, crude fat, crude protein and crude fiber

content using proximate analyses. The experiment also aimed to compare the results of

proximate analyses to the literature values from the manufacturer.

 4

Time and Place of the Study

The proximate analysis was conducted on different time and date. The moisture

content determination was conducted on September 2, 2019 at 7 to 10 am. The ash content

determination was conducted on September 9, 2019 at same time. Crude fat determination

was conducted on September 16, 2019 at 7 to 10 am but resumption of analysis was on the

following day. The crude protein and crude fiber analysis were on October 3, 2019 at 7 to

10 am. Both experiments approximately finished at the same time for about 3 more days

after. All the analyses were conducted in Visayas State University, Baybay City, Leyte,

Philippines.

Scope and Limitation of the Study

The purpose of the analysis was mainly focused on the investigation and evaluation

of the nutritional content of feeds such as moisture, ash, crude fat, crude protein and crude

fiber content of the Nutri-Start Pellet. Hence, whatever the result, the analysis does not

intend to sully the name of Universal Feed Mill Corporation or any of the related

organizational body.
 5

CHAPTER II

REVIEW OF RELATED LITERATURE

Proximate Analysis

Proximate analysis in food products is the analysis of the major constituents which

together comprise nearly 100% of the food composition. This generally includes water,

ash, total protein, and carbohydrates. Proximate analysis is the first approach for food

product characterization and control, and it is not dependent on expensive or sophisticated

equipment. It is easily carried out by quality control laboratories (Trugo et al., 2003).

A. Moisture Content

According to Bradley (2010) on the book “Food Analysis: Fourth Edition”,

moisture assays can be one of the most important analyses performed on a food product

and yet one of the most difficult from which to obtain accurate and precise data. The

approximate, expected moisture content of a food can affect the choice of the method of

measurement. It can also guide the analyst in determining the practical level of accuracy

required when measuring moisture content, relative to other food constituents.

The ease of water removal from food depends on how it exists in the food product.

The three states of water in food products are (Bradley, 2010):

1. Free water: This water retains its physical properties and thus acts as the

dispersing agent for colloids and the solvent for salts.

2. Adsorbed water: This water is held tightly or is occluded in cell walls or

protoplasm and is held tightly to proteins.

 6

3. Water of hydration: This water is bound chemically, for example, lactose

monohydrate; also, some salts such as Na2SO4·10H2O.

The moisture content – equilibrium relative humidity relationship of animal

feedstuffs is important, because it affects the storage potential according to Henderson

(1985) on the article “The Relationship Between Moisture Content and Equilibrium

Relative Humidity of Pig Feed”. Storage characteristics such as flow properties are related

to moisture content and the attack of feeds by microorganisms such as Salmonellae is also

influenced by moisture content – equilibrium relative humidity relationship and

temperature.

B. Ash Content

Ash content, as defined by Ismail (2017) on “Food Analysis Laboratory Manual”,

refers to the inorganic residue remaining after either ignition or complete oxidation of

organic matter in a foodstuff. The inorganic residue consists mainly of the minerals present

in the food sample. A basic knowledge of the characteristics of various ashing procedures

and types of equipment is essential to ensure reliable results. Two major types of ashing

are used: dry ashing, primarily for proximate composition and for some types of specific

mineral analyses; wet ashing (oxidation), as a preparation for the analysis of certain

minerals

Moreover, according to Marshall (2010) on the book “Food Analysis: Fourth

Edition”, dry ashing refers to the use of a muffle furnace capable of maintaining

temperatures of 500–600◦C. Water and volatiles are vaporized, and organic substances are

burned in the presence of oxygen in air to CO2 and oxides of N2. Most minerals are
 7

converted to oxides, sulfates, phosphates, chlorides, and silicates. Elements such as Fe, Se,

Pb, and Hg may partially volatilize with this procedure, so other methods must be used if

ashing is a preliminary step for specific elemental analysis. Wet ashing is a procedure for

oxidizing organic substances by using acids and oxidizing agents or their combinations.

Minerals are solubilized without volatilization. Wet ashing often is preferable to dry ashing

as a preparation for specific elemental analysis. Wet ashing often uses a combination of

acids and requires a special perchloric acid hood if that acid is used.

C. Crude Fat Content

According to Ellefson and Min (2010) in the book “Food Analysis: Fourth

Edition”, an accurate and precise quantitative and qualitative analysis of lipids in foods is

important for accurate nutritional labeling, determination of whether the food meets the

standard of identity, and to ensure that the product meets manufacturing specifications.

Inaccuracies in analysis may prove costly for manufacturers and could result in a product

of undesirable quality and functionality.

Moreover, in the article authored by Anderson, Thiex, and Gildemeister (2003)

titled “Crude Fat, Hexanes Extraction, in Feed, Cereal Grain, and Forage

(Randall/Soxtec/Submersion Method): Interlaboratory Study, ISO/IUPAC/AOAC

Harmonized Protocol”, a method for determining crude fat in animal feed, cereal grain and

forage (plant tissue) was collaboratively considered. The use of hexanes provided an

alternative to diethyl ether for fat extractions. The proposed submersion method

considerably decreased the extraction time required to complete a batch of samples

compared to the Soxhlet method. The increase in throughput provided faster turnaround

times, greater efficiency in the use of labor, and reclamation of the solvent. The submersion
 8

method for fat extraction using petroleum ether as the extractant was previously studied

for meat and meat products and was accepted as AOAC Official Method 991.36. Petroleum

ether (pet ether) is a commonly used solvent due to its relatively low cost compared to

other organic solvents. It is less hygroscopic than diethyl ether, is less flammable than

diethyl ether, and is more selective for hydrophobic lipids than diethyl ether. Many labs

use petroleum ether as a single solvent while other labs use a specific blend (ratio) of

petroleum ether and diethyl ether for lipid extraction. Petroleum ether generally dissolves

more non-polar lipids than diethyl ether and has less potential for peroxide formation.

Furthermore, according to Limsuwan et al. (1984) on the article “Determination of

Crude Fat in Fish Feeds”, traditional solvent extraction procedures have yielded crude fat

values lower than anticipated, based on fat contents of composing ingredients, in extrusion

processed fish feeds but provided reasonable values in steam pelleted feeds. Acid

hydrolysis before solvent extraction increased the crude fat values in extrusion processed

fish feeds by two and one‐half times. Acid hydrolysis pretreatment offered no advantage

over solvent (ether) extraction alone for crude fat determination in steam pelleted fish

feeds, fishmeal, catfish waste meal, or yellow corn meal. Acid hydrolysis pretreatment

doubled the yield of crude fat in commercial, solvent‐extracted soybean meal.

D. Crude Protein Content

According to Dalheim et al. (2018), on the article “Protein Determination – Method

Matter”, stated that the reported protein content of foods depends on the analytical method

used for determination, making a direct comparison between studies difficult. Some of

these methods require extraction preceding analysis. The efficacy of protein extraction
 9

differs depending on food matrices and thus extraction yield was determined. Overall, most

analytical methods overestimated the protein contents. The inaccuracies were linked to

indirect measurements, i.e., nitrogen determination and subsequent conversion to protein,

or interference from other chemical substances. Amino acid analysis is the only protein

analysis method where interfering substances do not affect the results.

On the other hand, as stated on the article “An Overview of the Kjeldahl Method of

Nitrogen Determination. Part II. Sample Preparation, Working Scale, Instrumental Finish,

and Quality Control” authored by Saez-Plaza et al. (2013), the Kjeldahl method has been

validated and standardized for total (crude) protein estimation for a wide variety of food

matrices, indirectly determined by their nitrogen content, and is the reference method

adopted by many international organizations. It consists of three main steps: sample

digestion, distillation, and ammonia determination (titration being the primary method).

The Kjeldahl method uses sulfuric acid, a variety of catalysts, and salts to convert

organically bound nitrogen in samples to ammonium with its subsequent measurement.

However, according to Chang (2010) on the book “Food Analysis: Fourth Edition”,

the analysis of proteins is complicated by the fact that some food components possess

similar physicochemical properties. Nonprotein nitrogen could come from free amino

acids, small peptides, nucleic acids, phospholipids, amino sugars, porphyrin, and some

vitamins, alkaloids, uric acid, urea, and ammonium ions. Therefore, the total organic

nitrogen in foods would represent nitrogen primarily from proteins and to a lesser extent

from all organic nitrogen-containing nonprotein substances. Depending upon

methodology, other major food components, including lipids and carbohydrates, may

interfere physically with analysis of food proteins. Numerous methods have been
 10

developed to measure protein content. The basic principles of these methods include the

determinations of nitrogen, peptide bonds, aromatic amino acids, dye-binding capacity,

ultraviolet absorptivity of proteins, and light scattering properties

E. Crude Fiber Content

According to Pigden et al. (1979) on the book “Standardization of Analytical

Methodology for Feeds”, crude fiber has been and remains a common means of evaluating

fibrous feeds. It is, however, grossly misleading and involves large errors based on

nutritional and biochemical criteria. In searching for a replacement there is a conflict

among criteria that evaluate the analytical parameters as indicators of nutritive value. The

favored criterion is that which evaluates on the basis of recovery of refractory and

indigestible residues. Another important consideration is economics of the methodology,

which needs to be competitive with proximate analysis in terms of laboratory cost and

technician time.

Also, according to BeMiller (2010) on the book “Food Analysis: Fourth Edition”,

the measurement of insoluble fiber is important not only in its own right, but also for

calculating the caloric content of a food. According to nutrition labeling regulations, one

method allowed to calculate calories involves subtracting the amount of insoluble dietary

fiber from the value for total carbohydrate, before calculating the calories based on protein,

fat, and carbohydrate content (approximately 4, 9, and 4 Calories per gram, respectively).

This method ignores the fact that soluble fiber, like insoluble fiber, is also essentially

noncaloric. Fiber components can contribute calories via absorption of products of

fermentation (mostly short-chain fatty acids) from the colon.

 11

CHAPTER III

METHODOLOGY

The methods for the determination of moisture, ash, crude fat, crude protein and

crude fiber content of the sample was patterned from the experimental procedures given

by the instructor, Prof. Allan A. Ramal. The feed that was being analyzed was the Nutri-

Start Pellet, a hog starter feed formulated by the Universal Feed Mill Corporation in Cebu

City which prior purpose was to give nutrient content such as vitamins, minerals and trace

elements for growth, strength and energy sources needed by 2 – 3 months old pigs.

I. Moisture Content Determination

In the moisture content determination, 15 grams of well mixed finely macerated

feed sample was weighed into a previously tared moisture dish. Three replicates were

prepared. The sample was then placed in the oven maintained at 105˚ C for at least five

hours. The feed contains water which is diffused on the mass in a relatively low quantity.

Since the water, or the moisture was part of the mass of the sample, the sample will

continually increase or decrease in weight because of hygroscopic action. Hygroscopic

action is the amount of moisture a material will absorb relative to ambient temperature and

humidity conditions (Reeb, et. al., 1999). The introduction of the very high temperature

declines the hygroscopic action of the feed and eventually evaporating the water that was

absorbed by the feeds, that is why the weight of the sample decreased. After drying in oven,

the sample was then cooled to room temperature in the desiccator. Afterwards, the dried

sample was the weighed. The sample was reheated again for 30 minutes and reweighed
 12

until there is a difference of not more than 0.001 g in weight. The percent moisture was

calculated using the Equation 1.

𝑤𝑡. 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 𝑏𝑒𝑓𝑜𝑟𝑒 𝑑𝑟𝑦𝑖𝑛𝑔 − 𝑤𝑡 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 𝑎𝑓𝑡𝑒𝑟 𝑑𝑟𝑦𝑖𝑛𝑔

% 𝑀𝑜𝑖𝑠𝑡𝑢𝑟𝑒 =
𝑤𝑡. 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 𝑏𝑒𝑓𝑜𝑟𝑒 𝑑𝑟𝑦𝑖𝑛𝑔

× 100 (𝐸𝑞. 1)

II. Ash Content Determination

A 0.1 gram (100 mg) of the desiccated feed sample (from the previous experiment)

was weighed and placed into a glazed crucible. Three replicate samples were prepared. The

sample was ignited in muffle furnace at extreme temperature of 550˚C until the sample

particle turned into light gray ash. The weight of the sample significantly decreased after

the ignition at 550˚C; the residue that remained was the ash. The application of very high

temperature to the sample evaporates the components with high volatility, and the fact that

inorganic compound such as minerals are not destroyed by heating, and that they have low

volatility so it would naturally remain as residue of the feed (Ismail, 2017). Afterwards,

the sample was cooled to room temperature in the desiccator and was weighed. The ash

percent of the sample was the calculated using the Equation 2 below.

𝑤𝑒𝑖𝑔ℎ𝑡 𝑜𝑓 𝑎𝑠ℎ (𝑔)

% 𝐴𝑠ℎ = × 100 (𝐸𝑞. 2)
𝑤𝑒𝑖𝑔ℎ𝑡 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 (𝑔)

Where: 𝑤𝑒𝑖𝑔ℎ𝑡 𝑜𝑓 𝑎𝑠ℎ = 𝑤𝑡 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 + 𝑐𝑟𝑢𝑐𝑖𝑏𝑙𝑒 𝑎𝑓𝑡𝑒𝑟 𝑖𝑔𝑛𝑖𝑡𝑖𝑜𝑛 − 𝑤𝑡 𝑜𝑓 𝑐𝑟𝑢𝑐𝑖𝑏𝑙𝑒

 13

III. Crude Fat Determination

One gram of freshly dried sample was weighed and was placed in a pre-weighed

filter paper pocket which both ends was secured by taping the folds. Three replicate

samples were prepared. The sample was the defatted in Soxhlet extractor using petroleum

ether solvent. The analyte was the fats or lipid such as monoglycerides, diglycerides,

triglycerides, sterol, free fatty acid, phospholipid, carotenoids, cholesterols and fat-soluble

vitamins, were all non-polar, which in turn, soluble to non-polar organic solvents like

benzene, chloroform, hexane, methanol and diethyl ether. This makes the petroleum ether

a good solvent for it is highly non-polar and could dissolves the fat and related substances

that is present in the feed sample. Since the only measurable components of the analysis

was the defatted sample, then it is intuitive that the mass lost after the extraction was the

crude fat contents of the feed samples because ether could only dissolve nonpolar crude fat

components leaving the polar intact. The length of time of defatting depends on the material

as specified. After the extraction, the Soxhlet extractor was removed and the air filter paper

pocket plus the sample under the hood until they were ether free. The staples of the folds

were then removed. Afterwards, the samples were then heated for 1 hour in oven at

maintained temperature of 105˚ C and then desiccated for another 1 hour. The defatted

samples plus paper were then weighed. The percent fat was then calculated using the

formula below.

𝑤𝑡. 𝑜𝑓 𝑓𝑎𝑡
% 𝐹𝑎𝑡 = × 100 (𝐸𝑞. 3)
𝑤𝑡. 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒

Where:𝑤𝑡. 𝑜𝑓 𝑓𝑎𝑡 = 𝑤𝑡 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 + 𝑝𝑎𝑝𝑒𝑟 𝑏𝑒𝑓𝑜𝑟𝑒 − 𝑤𝑡. 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 𝑎𝑛𝑑 𝑝𝑎𝑝𝑒𝑟 𝑎𝑓𝑡𝑒𝑟
 14

IV. Crude Protein Determination

One gram of the desiccated sample was weighed and placed in Kjeldahl tube.

Fifteen (15) mL of concentrated sulfuric acid and a tablet of Kjeldahl of catalyst was then

added in each tube. Four tubes were prepared; three for the replicate sample and one for

blank determination. Blank determination is a procedure which follows all steps of analysis

but in the absence of a sample, and its main purpose is for detection and compensation of

systematic analysis mistakes. The tube was then placed in the digester and the start button

was pressed to run the instrument. The feed samples were digested with concentrated

sulfuric acid to produce ammonium sulfate. After digestion, the tubes were cooled for at

least 5 minutes before removing. After removing, the samples were proceeded to

distillation by using the Foss automated distillation instrument. The operating procedure of

the instrument were followed. Eight drops of methyl red and bromocresol green indicator

were placed in 250 mL Erlenmeyer flask. Four Erlenmeyer flask was prepared. In

distillation, the ammonium was liberated to the Erlenmeyer flask and raising the pH of the

solution. 0.1 N HCl was titrated to silver gray endpoint. The method indirectly quantifies

the total protein content of foods by direct nitrogen measurement and subsequent

multiplication by a conversion factor (Persson et al, 2013). The general conversion factor

6.25 (i.e., 100/16) is used for most foods because their nonprotein nitrogen content is

negligible (Mariotti et al., 2008). The % N of the sample was calculated using Equation 4

and the % Crude Protein of the was calculated by using the equation 5.

(𝑚𝐿 𝑡𝑖𝑡𝑟𝑎𝑡𝑒𝑑 𝑎𝑐𝑖𝑑 − 𝑚𝐿 𝑏𝑙𝑎𝑛𝑘) × 𝑁 𝑜𝑓 𝑎𝑐𝑖𝑑 × 1.4007

%𝑁 = × 100 (𝐸𝑞. 4)
𝑤𝑡. 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒

% 𝐶𝑟𝑢𝑑𝑒 𝑃𝑟𝑜𝑡𝑒𝑖𝑛 = (% 𝑁) × 6.25 (𝐸𝑞. 5)

 15

V. Crude Fiber Analysis

A 0.1 grams of defatted feed sample (residue from Crude Fat Analysis) was

weighed and then placed in centrifuge tube. Three replicates were prepared. The sample

was added with a drop of octanol and 15 mL of boiling sulfuric acid (H2SO4) solution. The

resulting solution was then mixed gently to wet all powder and the tubes were covered with

glass marble. Sulfuric acid was added to extract the components that can be extracted

trough acid such as the sugar and starch of the feed. The tubes were then transferred to

boiling water bath and boil for 30 minutes. The tubes were then centrifuge at 5,000 x g for

10 minutes. The extraction process was fastened by subjecting the centrifuge tube to

centrifugation. Centrifugation is the technique for separation of particles from a solution

according to their size, shape, density, viscosity of the medium and rotor speed. The

supernatant was discarded, and the residue was washed with boiling distilled water and

centrifuged again. The washing with distilled hot water and centrifugation was repeated

until the residue is acid-free. The residue was digested in 15 mL boiling NaOH solution.

Sodium hydroxide was used to remove proteins and some hemi-cellulose and lignin. The

resulting solution was then centrifuged and washed with boiling water until the residue is

alkali-free. The residue was transferred to the prepared gooch crucible and was filtered

through suction washing the residue with distilled water. The residue was finally rinsed

with about 20 mL of 95% ethanol. The suction was removed, and the sample was dried in

the oven at 105˚C overnight. After drying, it was cooled to room temperature in the

desiccator and weighed. Afterwards, the sample was ignited in the muffle furnace at 550˚C

for an hour or longer, and then cooled in desiccator and weighed. The loss of weight was

the crude fiber, and the percent crude fiber was calculated using equation 6 below.
 16

𝑤𝑡. 𝑜𝑓 𝐶 + 𝑆 𝑎𝑓𝑡𝑒𝑟 𝑜𝑣𝑒𝑛 𝑑𝑟𝑦𝑖𝑛𝑔 − 𝑤𝑡. 𝑜𝑓 𝐶 + 𝑆 𝑎𝑓𝑡𝑒𝑟 𝑖𝑔𝑛𝑖𝑡𝑖𝑜𝑛

% 𝐶𝑟𝑢𝑑𝑒 𝐹𝑖𝑏𝑒𝑟 =
𝑤𝑡. 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒

× 100 (𝐸𝑞. 6)

Where: 𝐶 + 𝑆 = 𝑐𝑟𝑢𝑐𝑖𝑏𝑙𝑒 + 𝑠𝑎𝑚𝑝𝑙𝑒

 17

CHAPTER IV

RESULTS AND DISCUSSION

Table 1. The Proximate Analysis on Nutri-Start Pellet Feed Sample

Animal Feed Replicate Replicate Replicate Literature

Mean(±SD)
Component 1 2 3 Value

% Moisture 8.7931 % 11.0724 % 5.7874% 8.5359(±2.6296) % 12% max

% Ash 4.0143 % 3.8033 % 3.4387 % 3.7521(±0.2912) % > 1.65% est

% Crude Fat 6.1197 % 7.6496 % 7.2692 % 7.0128(±0.7965) % 3% min

% Crude Protein 17.3550 % 17.4338 % 17.1975 % 17.3289(±0.1203) % 18% min

% Crude Fiber 2.0731 % 4.1071 % 3.7250 % 3.3017(±1.081) % 5% max

% Carbohydrate 60.0686 %

Table 1 shows the data gathered from the proximate analysis of UNIFEEDS Nutri-

Start Pellet feed sample — measuring the moisture content, ash content, crude fat content,

crude protein content and crude fiber content the feed sample has.

Percent Moisture in Feed Sample

It was shown on the Table 1 that the sample 1 has 8.7931 % moisture, sample 2 has

11.0274 % moisture and sample 3 has 5.7874 % moisture. With this, it was calculated that

the mean % moisture of the sample was 8.5359 % with a standard deviation of 2.6296 %.

It is noticeable that the % moisture of each sample deviates largely from the mean value

which reflects on its large standard deviation. Even though the percentage deviates
 18

unprecedently does not mean the data was erroneous because it was natural for a bulk of

feeds to have different percent moisture depending on circumstances such as uneven

exposure to air and many more. As stated by the feed description, the maximum % moisture

that the Nutri-Start pellet would have is 12%, and since the experimentally determined

average % moisture was 8.5359 %, it implies that the data was satisfactory, and the

moisture analysis was successful.

As stated, the 8.5359% of the sample was merely water and the percentage was

inside the range for correct percent moisture for the Nutri-Start pellet. Moisture function

as a vital component in the food-chemical changes that takes place between ingredients

during processing, and therefore has a determining effect on the quality and nutritional

value of a final product. If the product would have too low percent moisture content, some

nutrients would lose so it would be less palatable. On the other hand, feeds with high

moisture content are liable to spoil quickly, mainly from fungal contamination. The molds

and more particularly the toxins that are produced by many molds make the feed

unpalatable and can cause illness or even death to the animals.

Percent Ash in Feed Sample

For the analysis of ash content of Nutri-Start Pellet, it turns out that the % ash of

the sample 1 was 4.0143 %, sample 2 was 3.8033 %, and sample 3 was 3.4387 %. With

this, the mean value was calculated to be 3.7521 (±0.2912) %. The feed does not provide

any description for ash content, but it does provide the percentage for calcium and

phosphorus content. Calcium and phosphorus are not the only minerals can be found in

feed ash since some oxides, silicates and sulfates together with elemental Na, Fe, K

(Marshall, 2010) and many more is also present in residual ash. Knowing the relative
 19

percentage of calcium and phosphorus would give an excellent estimate on the ash content

of the sample feed— the percent ash of the feed would not go lesser than the added

percentage of Ca and P. Since the approximate percentage of calcium and phosphorus is

1.65 %, which is lesser than the experimental ash content and the fact that the data has high

precision with a standard deviation of ±0.2912 %, then it is safe to assume that the % ash

of the feed was inside the small range of the data (3.7521±0.2912 %) and the true value of

% ash was very close to the experimental % ash value.

Among the mineral present in ash, calcium and phosphorus are the two most vital

for young pigs because these two elements are important in skeletal structure development.

About 99 % of the calcium and 80 % of the phosphorus in the body are found in the skeleton

and teeth. Therefore, deficiency of calcium and phosphorus will result in impaired bone

mineralization, reduced bone strength, and poor growth on the consumer swine.

Percent Crude Fat in Feed Sample

As what have been calculated based on the data of the experiments, the sample 1,

2 and 3, have a % crude fat content of 6.1197 %, 7.6496 % and 7.2692 %, respectively—

averaging of 7.0128 (±0.7965) % which was stated on Table 1. The individual percent fat

of the sample correlates with each other with high precision, which can be seen on its very

small standard deviation. Nutri-Start pellet describes that the feed would have a minimum

crude fat content of 3%, and since the experimentally determined average % crude fat was

7.0128 % which was clearly above the minimum crude fat content the feed describes, then

it was valid and the analysis precisely determined the crude fat content of the at-hand feed

sample.
 20

The measurement of crude fat was an important part of the historical method of

proximate analysis where it represented feed components with a caloric value 2.25 times

(Western Diary Inc, 2004) that of protein and carbohydrates. Fat, which contains 2.25 times

the amount of energy as cereal grains, is often used to increase the energy density of swine

diets. It helps the weanling pigs to promote efficient growth, improves the lean tissue

deposition and helps maintain the performance during heat stress.

Percent Crude Protein in Feed Sample

The average % crude protein of the feed sample was determined to be 17.3289

(±0.1203) %— calculated from the data on Table 1. The UNIFEEDS describes that the

Nutri-Start pellet has a minimum % crude protein of 18 %. The experimental result shows

to be not compatible with the described percentage because 17.3289% was below the

minimum value of the % crude protein nutritional content of the said feed. The

experimental % crude protein value of the three replicates was very close to each other

which reflects to its very small standard deviation of only 0.1203%, thus the data correlate

with each other with precision and the error was minimized. Considering the 18% percent

as the true value of the % crude protein, then the Relative Error of the experimental result

was 3.7283% — quite small and negligible to some measurement. And if 18% was an

assumption, then the experimental average % crude protein was the precise % crude protein

value of the sample feed due to the fact that the individual % crude protein of the sample

correlate to each other with precision, and the Relative Error value was very small.

Similarly, there is no machine that could formulate feed with exact the same value of

percent of nutrients present in every making, all values are fall under certain range.
 21

Therefore, the experimental average % crude protein was acceptable, and the analysis was

successful in other perspective.

Protein is a macronutrient that is made up of amino acids; it is very essential to

building muscle mass. It is commonly found in animal products, though is also present in

other sources, such as nuts and legumes. It is one of the three nutrients that provides calorie

and energy, the other two were fats and carbohydrates. A protein that contains the perfect

balance of the individual essential amino acids combined with adequate amounts of non-

essential amino acids is referred to as "ideal protein." The protein most useful in pig feeds

are those high in lysine, methionine and threonine. Similarly, protein is a very important in

the human body. It is responsible for the growth, cell maintenance, repair of tissues,

production of hormones and production of antibodies for the immune system.

Percent Crude Fiber in Feed Sample

Lastly, the % crude fiber was the calculated using the Equation 6; it turned out that

the % crude fiber of each sample was 2.0731 %, 4.1071 % and 3.7250 %, respectively from

sample 1 to 3, which gives an average % crude fiber of 3.3017(±1.0810) %. The

UNIFEEDS describes that the Nutri-Start pellet has a maximum crude fiber content of 5%

and since the experimental average crude fiber content of the sample was 3.3017% which

below the threshold, then implies that the data was valid, and the analysis was a success.

Fiber is very essential as nutrients. High fiber content normalizes bowel

movement— it increases the weight and size your stool and softens it. A bulky stool is

easier to pass, decreasing the chance of constipation. Fiber also helps maintain bowel health

for it lower your risk of developing hemorrhoids and small pouches in the colon. Aside
 22

from its effect on digestive health, it also helps lower cholesterol level and control sugar

level on the body.

 23

CHAPTER V

SUMMARY, CONCLUSION AND RECOMMENDATIONS

Summary

The study was conducted to quantitatively determine the proximate percent

composition of moisture, ash, crude fat, crude protein and crude fiber content of

UNIFEEDS Nutri-Start Pellet, and to compare the experimental result with the literature

value of the Nutri-Start Pellet provided by the manufacturer. The moisture content of the

feed sample was determined through oven drying method and it was determined that the

percent moisture of the sample was 8.5359(±2.6296) %. On the other hand, the ash content

of the Nutri-Start Pellet was determined through incineration of the sample at high

temperature in muffle furnace. With that, it was determined that the percent ash of the

sample was 3.7521(±0.2912) %. Furthermore, for the determination of crude fat, the feed

sample was subjected to Soxhlet extraction method. Based on the findings, it was

determined that the percent crude fat of the Nutri-Start Pellet was 7.0128(±0.7965) %. The

resulting defatted sample was then used for crude fiber determination wherein the defatted

sample was subjected to sequential acid and alkali digestion, drying and ignition. The loss

of weight before and after ignition was the amount of crude fiber, which accommodate to

about 3.3017(±1.081) % of the Nutri-Start Pellet sample. Lastly, the crude protein content

of the sample was determined through Kjeldahl Method. Kjeldahl Method assessed the

percent nitrogen of the sample and multiplying it with 6.25 gave the percent crude protein

which for the sample was 17.3289(±0.1203) %. Comparing with the literature value, the

experimental results shows minimal deviation, but the values agrees on the specific ranges
 24

of moisture, ash, crude fat, crude protein and crude fiber content of Nutri-Start Pellet.

Therefore, the analyses were successful.

Conclusion

It was concluded from the proximate analysis that the UNIFEEDS Nutri-Start Pellet

has 8.5359 % average moisture content, 3.7521 % average ash content, 7.0128 % average

crude fat content, 17.3289 % average crude protein content and, 3.3017 % average crude fiber

content. Based on the findings, it was also concluded that the experimentally determined

moisture, ash, crude fat, crude protein and crude fiber content correlates to the literature

value of the said nutrients provided by the Universal Feed Mill Corporation to UNIFEEDS

Nutri- Start Pellet. Therefore, based on the results, the relative value of the of the primary

nutrient composition of the feed was enough to give nutrient content such as vitamins,

minerals and trace elements for growth, strength and energy sources needed by 2 – 3

months old pigs.

Recommendation

For the improvement of the study, the following are needed to consider:

1. For the crude fat extraction, select the best organic solvent for extraction of fats and

fat-like compounds such as petroleum ether in the experiment. Hexane and other

suitable solvent can be also used.

 25

2. Measuring of the weights of the samples on the analysis that depends on the loss of

weight must be done in an air conditioned, vibration-free and close room. The

balance must also calibrate periodically.

3. For the next proximate analysis, the student must devote themselves on doing the

experiment for dedication does a lot on the outcome of the analysis.

 26

LITERATURE CITED

Ayuba V. O., E. K. (2010). PROXIMATE COMPOSITION OF SOME COMMERCIAL

FISH FEEDS SOLD IN NIGERIA. Fisheries Society of Nigeria.

BeMiller, J. N. (2010). Carbohydrate Analysis. In Food Analysis: Fourth Edition (pp.

165-166). West Lafayette, IN 47907–2009, USA.

Chang, S. K. (2010). Protein Analysis. In Food Analysis: Fourth Edition (pp. 133-136).

Fargo, ND 58105, USA : Springer Science+Business Media.

E. T. Kornegay, H. R. (1974). Evaluation of Protein Levels and Milk Products for Pigs

Starter Diet. Journal of Animal Science, Vol. 39, Issue 3.

E. W. Crampton, L. A. (1938). The Relation of Cellulose and Lignin Content to the

Nutritive Value of Animal Feeds. The Journal of Nutrition, Volume 15, Issue 4,

383-395.

Hanne K. Mæhre, L. D.-J. (2018, January 1). Protein Determination—Method Matters.

Norwegian College of Fishery Science, Faculty of Biosciences, Fisheries and

Economics, UIT The Arctic University of Norway, Tromsø, Norway.

Henderson, S. (1985). The Relationship Between Moisture Content and Equilibrium

Relative Humidity of Pig. Pergamon Press Ltd .

Ismail, B. P. (2017). Ash Content Determination. In Food Analysis Laboratory Manual

(pp. 117-119). St. Paul, MN, USA : Springer International Publishing.

Jan-Åke Persson, P. (2008, February). Handbook for Kjeldahl Digestion. FOSS, DK-

3400 Hilleroed, Denmark.

 27

Jim Reeb, M. M. (1999). Moisture Content by the Oven-Dry Method for Industrial

Testing. Corvallis, OR, Oregon State University.

Marshall, M. R. (2010). Ash Analysis. In Food Analysis: Fourth Edition (pp. 105-108).

Gainesville, FL 32611-0370, USA : Springer Science+Business Media.

Min, D. B. (2010). Fat Analysis. In Food Analysis: Fourth Edition (pp. 117-120).

Columbus, OH 43210, USA : Springer Science+Business Media.

Möller, J. (2014, February). Comparing methods for fibre determination in food and feed

Nancy J. Thiex, S. A. (2003). Crude Fat, Hexanes Extraction, in Feed, Cereal Grain, and

Forage (Randall/Soxtec/Submersion Method): Interlaboratory Study,

ISO/IUPAC/AOAC Harmonized Protocol. Journal of AOAC International Vol.

86.

Robert L. Bradley, J. (2010). Moisture and Total Solids Analysis. In Food Analysis:

Fourth Edition (pp. 85-88). Madison, WI 53706, USA : Springer

Science+Business Media.

https://psa.gov.ph/livestock-poultry-iprs/swine/inventory

http://www.pcaarrd.dost.gov.ph/home/momentum/swine/index.php/industry-status
 28

APPENDICES

Appendix Tables
Moisture Content Analysis
Appendix Table 1. Mass of Container (Moisture Analysis)

Before drying in oven First Drying in oven Second Drying in

Container No.
(g) (g) oven (g)
1 0.8535 0.8535 0.8535
2 0.4833 0.4833 0.4832
3 0.8330 0.8329 0.8328

Appendix Table 2. Mass of Sample (Before and After Heating) and Moisture

Parameter Sample 1 (g) Sample 2 (g) Sample 3 (g)

Sample not heated 15.0903 15.0471 15.0413
Sample after heating 13.7634 13.3878 14.1708
Moisture 1.3269 1.6593 0.8705
Mean(±SD)
1.2856 (±0.3960)
Moisture

Appendix Table 3. The Percent Moisture of each sample

Replicates % Moisture
Sample 1 8.7931 %
Sample 2 11.0274 %
Sample 3 5.7874 %
Mean 8.5359 %
Standard Deviation 2.6296 %
 29

Ash Content Analysis

Appendix Table 4. Mass of Crucible (Ash Analysis)

Crucible No. 1st Drying (g) 2nd Drying (g) 3rd Drying (g)
1 24.8080 24.8079 24.8079
2 22.2295 22.2301 22.2301
3 21.0552 21.0547 21.0546

Appendix Table 5. Mass of Crucible + Sample and Sample Before and After Ignition

Crucible No. Mass of Crucible + Sample (g) Mass of Sample (g)

Before Ignition After Ignition Before Ignition After Ignition
1 24.9200 24.8124 0.1121 0.1076
2 22.3379 22.2342 0.1078 0.1037
3 21.1622 21.0583 0.1076 0.1039

Appendix Table 6. The Percent Ash of each sample

Crucible No. % Ash (%)

1 4.0143
2 3.8033
3 3.4387
Mean 3.7521
Standard Deviation 0.2912

Crude Fat Analysis

Appendix Table 7. Initial Weights of Filter Paper and Sample

Parameter
Tea bag 1 (g) Tea bag 2 (g) Tea bag 3 (g)

Filter paper 1.2230 1.2582 1.3204

Sample 1.0507 1.0079 1.0799
Filter paper +
2.2737 2.2661 2.4003
sample
 30

Appendix Table 8. Weight of the Sample and Crude fat After Defatting

Parameter Tea bag 1 (g) Tea bag 2 (g) Tea bag 3 (g)
Filter paper +
2.2094 2.1890 2.3218
sample
Defatted sample 0.9864 0.9308 1.0014
Crude Fat 0.0643 0.0771 0.0785
% Fat 6.1197 % 7.6496 % 7.2692 %
Mean 7.0128 %
Standard Deviation 0.7965 %

Crude Protein Analysis

Appendix Table 9. Crude Protein Determination Data

Parameter Sample 1 Sample 2 Sample 3 Blank

Mass of foil 0.6012 g 0.5637 g 0.6193 g 0.5795 g
Mass of Sample 1.0759 g 1.0080 g 1.0134 g X
Volume
22.4 mL 22.5 mL 22.2 mL 0.4 mL
(endpoint)
N% 2.7768 % 2.7894 % 2.7516 % X
Crude Protein % 17.3550 % 17.4338 % 17.1975 % X
Mean Crude
17.3289 % X
Protein %
Standard
0.1203 % X
Deviation
 31

Crude Fiber Analysis

Appendix Table 10. Crude Fiber Determination Data

Parameter Sample 1 Sample 2 Sample 3

Mass of sample 0.1013 g 0.1120 g 0.1047 g
Mass of crucible +
21.0803 g 19.9129 g 19.0498 g
sample after drying
Mass of crucible +
sample after 21.0782 g 19.9036 g 19.0459 g
ignition
% Crude Fiber 2.0731 % 4.1071 % 3.7250 %
Mean 3.3017 %
Standard Deviation 1.0810 %

Calculations

A. Percent Moisture

𝑤𝑡. 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 𝑏𝑒𝑓𝑜𝑟𝑒 𝑑𝑟𝑦𝑖𝑛𝑔 − 𝑤𝑡 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 𝑎𝑓𝑡𝑒𝑟 𝑑𝑟𝑦𝑖𝑛𝑔

% 𝑀𝑜𝑖𝑠𝑡𝑢𝑟𝑒 = × 100
𝑤𝑡. 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 𝑏𝑒𝑓𝑜𝑟𝑒 𝑑𝑟𝑦𝑖𝑛𝑔

Replicate 1:

% 𝑀𝑜𝑖𝑠𝑡𝑢𝑟𝑒 = (15.0903 𝑔 − 13.7634 𝑔) × 100

% 𝑀𝑜𝑖𝑠𝑡𝑢𝑟𝑒 = 8.7931 %

B. Percent Ash

𝑤𝑒𝑖𝑔ℎ𝑡 𝑜𝑓 𝑎𝑠ℎ (𝑔)

% 𝐴𝑠ℎ = × 100
𝑤𝑒𝑖𝑔ℎ𝑡 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 (𝑔)

Replicate 1:

0.0045 𝑔
% 𝐴𝑠ℎ = × 100
0.1121 𝑔

% 𝐴𝑠ℎ = 4.0143 %
 32

C. Percent Crude Fat

𝑤𝑡. 𝑜𝑓 𝑓𝑎𝑡
% 𝐹𝑎𝑡 = × 100
𝑤𝑡. 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒
Replicate 1:
0.0643 𝑔
% 𝐶𝑟𝑢𝑑𝑒 𝐹𝑎𝑡 = × 100
1.0507 𝑔
% 𝐶𝑟𝑢𝑑𝑒 𝐹𝑎𝑡 = 6.1197 %
D. Percent Crude Protein

(𝑚𝐿 𝑡𝑖𝑡𝑟𝑎𝑡𝑒𝑑 𝑎𝑐𝑖𝑑 − 𝑚𝐿 𝑏𝑙𝑎𝑛𝑘) × 𝑁 𝑜𝑓 𝑎𝑐𝑖𝑑 × 1.4007

%𝑁 = × 100
𝑤𝑡. 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒

% 𝐶𝑟𝑢𝑑𝑒 𝑃𝑟𝑜𝑡𝑒𝑖𝑛 = (% 𝑁) × 6.25

Replicate 1:

(22.4 𝑚𝐿 − 0.4 𝑚𝐿) × 0.965 𝑀 × 1.4007

%𝑁 =
1.0759

% 𝑁 = 2.7768 %

For % Crude Protein

% 𝐶𝑟𝑢𝑑𝑒 𝑃𝑟𝑜𝑡𝑒𝑖𝑛 = 2.7768 % × 6.25

% 𝐶𝑟𝑢𝑑𝑒 𝑃𝑟𝑜𝑡𝑒𝑖𝑛 = 17.3550 %

E. Percent Crude Fiber

𝑤𝑡. 𝑜𝑓 𝐶 + 𝑆 𝑎𝑓𝑡𝑒𝑟 𝑜𝑣𝑒𝑛 𝑑𝑟𝑦𝑖𝑛𝑔 − 𝑤𝑡. 𝑜𝑓 𝐶 + 𝑆 𝑎𝑓𝑡𝑒𝑟 𝑖𝑔𝑛𝑖𝑡𝑖𝑜𝑛

% 𝐶𝑟𝑢𝑑𝑒 𝐹𝑖𝑏𝑒𝑟 = × 100
𝑤𝑡. 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒

Where: 𝐶 + 𝑆 = 𝑐𝑟𝑢𝑐𝑖𝑏𝑙𝑒 + 𝑠𝑎𝑚𝑝𝑙𝑒

Replicate 1:

21.0803 𝑔 − 21.0782 𝑔
% 𝐶𝑟𝑢𝑑𝑒 𝐹𝑖𝑏𝑒𝑟 = × 100
0.1013 𝑔

% 𝐶𝑟𝑢𝑑𝑒 𝐹𝑖𝑏𝑒𝑟 = 2.0731 %