1

Approximate
Analyses
of KLM
Golden
RHA Baldonasa,
KF Briones,
CaceresOats

Approximate Analyses of Oats

RB Hera Angelica Baldonasa
Kathleen Faith Briones
Kristhia Loren Maree Caceres

University of the Philippines Visayas

Miagao, Iloilo
Abstrac:t
Oats are generally considered "healthful", or a health food, being touted commercially as nutritious. A system of
analyses on the composition of food, called proximate analyses is performed in this sample which is Golden oats.
The objective of the experiment is to prove and perform analyses on parameters namely moisture, ash, protein, and
carbohydrates.
Proper moisture content is essential for maintaining fresh, healthy foods. The method used for moisture analysis in
this experiment is the Microwave Oven method. Weighed samples are placed in a microwave oven for a specified
time and power-level and were dried until they have reached a constant final mass.
Carbohydrates is known the source of energy. It can be digestible or indigestible. Indigestible carbohydrates is also
called fibers.
Human body needs fat in order to be healthy. Fat supply necessary acids and glycerol that the body can
only get from food. Percent fat was determined using the Ultrasonic extractor in separating fat from sample and
using Rotary evaporator in separating fat from the solvent
Ash is formed when a sample undergoes oxidation or ignition. Determining the % ash of a sample is essential
because it represents the total mineral content in food.
Nitrogen is one of the five major elements found in organic materials such as protein. Kjeldahl method is used since
it is considered to be the standard method of determining protein concentration. This method has three steps and
that includes digestion, distillation and titration.
As obtained in the experiment, the value for moisture is 14.01 0.405 , fat content is 6.557 0.2677, ash content is
2.557 0.0343 , carbohydrates is

I.

Introduction

Oats (Avena sativa L.) are distinct among cereals

due to their considerably higher protein
concentration. At the same time oats possess a protein
quality of high nutritional value and a special protein
composition. Amino acid composition of oats is
superior to that of other cereals due to the higher
amount of limiting amino acids like lysine and
threonine. During germination, total amino acid
analysis revealed an increase in essential amino acids

63.2112 and protein is 13.37 0.5701 .

like lysine and tryptophan, which leads to an

increased nutritional value of germinated oats. Oats
protein products including globulin have been
chemically modified by various methods to improve
their functional properties.
Proteins are polymers of amino acids.
Twenty different types of amino acids occur naturally
in proteins. Proteins differ from each other according
to the type, number and sequence of amino acids that
make up the polypeptide backbone. As a result they

have different molecular structures, nutritional

attributes and physiochemical properties. Proteins are
important constituents of foods for a number of
different reasons. They are a major source of energy,
as well as containing essential amino-acids, such as
lysine, tryptophan, methionine, leucine, isoleucine
and valine, which are essential to human health, but
which the body cannot synthesize. Proteins are also
the major structural components of many natural
foods, often determining their overall texture.
Isolated proteins are often used in foods as
ingredients because of their unique functional
properties, i.e. their ability to provide desirable
appearance, texture or stability. Typically, proteins
are used as gelling agents, emulsifiers, foaming
agents and thickeners. Many food proteins are
enzymes which are capable of enhancing the rate of
certain biochemical reactions. These reactions can
have either a favorable or detrimental effect on the
overall properties of foods. Food analysts are
interested in knowing the total concentration, type,
molecular structure and functional properties of the
proteins in foods.
The Kjeldahl method of nitrogen analysis is
the worldwide standard for calculating the protein
content in a wide variety of materials ranging from
human and animal food, fertilizer, waste water and
fossil fuels. The Kjeldahl method was developed in
1883 by a brewer called Johann Kjeldahl. A food is
digested with a strong acid so that it releases nitrogen
which can be determined by a suitable titration
technique. The amount of protein present is then
calculated from the nitrogen concentration of the
food. The same basic approach is still used today,
although a number of improvements have been made
to speed up the process and to obtain more accurate
measurements. It is usually considered to be the
standard
method
of
determining
protein
concentration. Because the Kjeldahl method does not
measure the protein content directly a conversion
factor (F) is needed to convert the measured nitrogen
concentration to a protein concentration. A
conversion factor of 6.25 (equivalent to 0.16 g
nitrogen per gram of protein) is used for many
applications, however, this is only an average value,
and each protein has a different conversion factor
depending on its amino-acid composition. The
Kjeldahl method can conveniently be divided into
three steps: digestion, neutralization and titration.

Food moisture analysis involves the whole

coverage of the food items in the world because
foods comprise a considerable amount of water rather
than other ingredients.
Moisture content of the food material is
important because it affects the physical, chemical
aspects of food which relates with the freshness and
stability for the storage of the food for a long period
of time and the moisture content determine the actual
quality of the food before consumption and to the
subsequent processing in the food sector by the food
producers. If a food is too moist or too dry, it may not
be suitable to eat and will not taste as good as it
would if it had the correct moisture content. Fresh,
ripe fruits and vegetables are moist to the touch. As
they age and begin to rot, some dry out and some
pick up excess moisture and begin to mold. Food
industries require a minimum or maximum
percentage of moisture on certain foods in order for
them to be packaged and labeled. If they don't fit to
these standards, the foods cannot be sold. In
processed foods, the percentage of water in a product
can determine its final price. Generally, a product
with more water will cost less. Biologists and
manufacturers need to know the moisture content of
food to ensure that it's processed and packaged in a
safe, stable way. Moisture content determines the
way most foods taste, feel and look. It is one of the
important ways to measure food quality.
Carbohydrates are one of the most important
component in many foods. It is important as a major
source of energy. In this experiment, carbohydrate
content will be determined by difference analysis
given that all components needed for the calculation
is dealt with.
Ash is formed when a sample undergoes
oxidation or ignition. Determining the % ash of a
sample is essential because it represents the total
mineral content in food. It is the inorganic fraction
which does not any specific information on any
nutrient or mineral and just mostly used in
determining the carbohydrate by difference. The
ashed sample can be further treated to determine 21
to 26 minerals element. It is done by incineration
using the muffle furnace at 550C.

Determining the %ash is not truly

representative of mineral levels in either quantitative
or qualitative terms. This is true for sulfur &
phosphorus either of which could be derived from the
organic or inorganic fractions.

Lipid refers to the group of compounds

that are sparingly soluble in water, but has
variable solubility in some organic solvent.
Lipids are one of the major constituents of foods
and of course important to our diet. They are the
source of energy and provide essential nutrients.
But too much consumption can lead to health
diseases. Lipid component plays a role in
determining the physical characteristics like
flavor, texture, and mouth feel and appearance
any lipid. Lipid content of a food determined
with another solvent different polarity.
Fat is composed of Carbon, Hydrogen and
Oxygen. Fats are essential for the proper
functioning of the body. Human body needs to
consume some fats to remain healthy. Fats
supply necessary fatty acids, acids and glycerol
that the body can only get from food such as
oats. Fats also serve as storage substance for the
bodys excess calories. When the body has
depleted its carbohydrate stores, it draws on fat.

Fat

Approximately 7.50 g of sample were weighed and

were wrapped in a filter paper tied in the sides like a
timble. It was then placed in 3 erlenmeyer flasks and
were added with 150 ml of petroleum ether as
solvent. The extraction of lipid through the Ultrasonic
bath was done for 30 minutes each. After 30 minutes,
the mixture was filtered. Extraction was repeated
twice when the solvent was already colorless. All the
extracts were combined in a 500 ml flask and
transferred to a 1L round bottom flask. The rotary
evaporator was then used to separate the petroleum
ether from the fat. The extracted fat was transferred
to the constant weighed vials and evaporated to
remove the solvent. Fat content was measured by
weight loss of the sample or by weight of fat
removed by subracting the weight of the vial from the
recorded weight.

Protein

Refer to AOAC Official Method No. 950.48

III.

Results and Discussion

Table 1. Parameter Values

To determine the fat content of the sample, fat is

extracted using petroleum ether as solvent in a
process called ultrasonic extraction. The fat
content is then calculated.
Determining the total lipid / fat content is one of
the many objectives of this experiment.
Table 2. Theoretical Values

II.

Methodology
Moisture

Refer to AOAC Official Method No. 925.40

Ash

Refer to AOAC Official Method No. 950.49

A. Fat

Petroleum ether was used as the solvent due to

its low boiling point of about 35 60C and is
non polar in nature. Lipids are relatively non
polar molecules. By using petroleum ether,
lipids are pulled out of the sample and only the
non polar molecules in the sample are
dissolved.
The sample with the solvent was transferred to
an ultrasonic cleaner that used ultrasound and
cavitation bubbles induced by the high pressure
sound waves to agitate the liquid. The agitation
produces high forces on contaminants like oil.
The solvent was separated using the rotary
evaporator. There is a water bath that can be
heated that keeps the solvent from freezing due
to evaporation process. Solvent is then collected
from the condenser to the receiving flask. A
cooling bath was placed on the receiver to
prevent volatile solvents to evaporate. The
theoretical value for fat as indicated in the
nutritional facts is 6.8% and the obtained
experimental value of 6.557 % giving a relative
error of 3.57%

Neutralization
After the digestion has been completed the digestion
flask is connected to a receiving flask by a tube. The
solution in the digestion flask is then made alkaline
by addition of sodium hydroxide, which converts the
ammonium sulfate into ammonia gas:
(NH4)2SO4 + 2 NaOH 2NH3 + 2H2O + Na2SO4
(2)
The ammonia gas formed is liberated from the
solution and moves out of the digestion flask and into
the receiving flask - which contains an excess of
hydrochloric acid. The low pH of the solution in the
receiving flask converts the ammonia gas into the
ammonium ion, and simultaneously converts the
boric acid to the chlorate ion:
NH3 + HCl NH4+ + Cl- (chlorate ion) (3)
Titration
The nitrogen content is then estimated by titration of
the ammonium borate formed with hydrochloric acid,
using phenolphthalein as indicator to determine the
end-point of the reaction.
Cl- + H+ HCl (4)
The concentration of hydrogen ions (in moles) required to
reach the end-point is equivalent to the concentration of
nitrogen that was in the original food sample (Eq3). The
following equation can be used to determine the nitrogen
concentration of a sample that weighs m grams using
a xM HCl acid solution for the titration:

B. Protein
Digestion
The process in which the food sample to be analyzed
is weighed into a digestion flask, heating it in the
presence of sulfuric acid, an oxidizing agent which
digests the food, anhydrous sodium sulfate (to speed
up the reaction by raising the boiling point) and a
HgO as a catalyst to speed up the reaction. Digestion
converts any nitrogen in the food (other than that
which is in the form of nitrates or nitrites) into
ammonia, and other organic matter to C02 and H20.
Ammonia gas is not liberated in an acid solution
because the ammonia is in the form of the ammonium
ion (NH4+) which binds to the sulfate ion (SO 42-)
and thus remains in solution:
N(food)(NH4)2SO4 (1)

(5)
Where vs and vb are the titration volumes of the
sample and blank, and 14g is the molecular weight of
nitrogen N. A blank sample is usually ran at the same
time as the material being analyzed to take into
account any residual nitrogen which may be in the
reagents used to carry out the analysis. Once the
nitrogen content has been determined it is converted
to a protein content using the appropriate conversion
factor:
%Protein = Factor* %N.
C. Moisture
Oven- drying method/microwave oven method is
widely used in determination of moisture. Water is
removed due to heating at 110C. Oats may
also contain volatile oil in addition to moisture and

these volatile oils also get loss during oven drying.

Weight loss due to the loss of volatile oils also gets
counted for moisture determination. This is the main
disadvantage of oven drying method. This method is
not suitable to determine the moisture content
of thermally unstable compounds and this
method removes only free water. Since oven
drying method is simple, low cost and easy, it is
widely used for routine analysis.
The accuracy of results of moisture
determination
is
affected
by
drying
temperature, relative humidity, particle size of
sample, handling method of sample, amount of
sample, type of evaporation dish, variation in
temperature inside the oven. To minimize
these errors various precautions should be
taken. Measures such as drying the sample in
crucibles, which do not decomposed during
heating. Three consecutive samples were
carried out to eliminate the errors of the handlers.
Approximately 2.00 g of ground sample was taken to
facilitate the drying, and since particle size is small,
evaporation is hastened.
D. Ash
The sample was pre-ashed with the use of a
Bunsen burner to avoid ignition inside the furnace.
When the sample was ignited, the water and other
volatile compounds are vaporized. Organic
compounds are burned with the presence of oxygen
in CO2 and oxides in N2. Most minerals are converted
into oxides, sulfates, chlorides, phosphates and
silicate.
Minerals are not decomposed when undergoing
heating. They have low volatility as compared to
other food components. However, not all minerals
have low volatility at high temperatures such that of
muffle furnace and may partially volatilize and be
lose. If the process of ashing is used for preliminary
elemental analysis, samples that contain Iron, Lead,
Mercury, Copper, Nickel, Zinc utilizes an alternative
procedure as these are examples of minerals that are
partially volatilized.
E. Carbohydrates
Total carbohydrate content of foods has, for many
years, been calculated by difference, rather than
analysed directly. Under this approach, the other
constituents in the food (protein, fat, water, alcohol,
ash) are determined individually, summed and
subtracted from the total weight of the food. This is

referred to as total carbohydrate by difference and is

calculated by the following formula:
100 - (weight in grams [protein + fat + water + ash +
alcohol] in 100 g of food)
It should be clear that carbohydrate estimated in this
fashion includes fibre, as well as some components
that are not strictly speaking carbohydrate, e.g.
organic acids (Merrill and Watt, 1973). Total
carbohydrate can also be calculated from the sum of
the weights of individual carbohydrates and fibre
after each has been directly analysed. Available
carbohydrate represents that fraction of carbohydrate
that can be digested by human enzymes, is absorbed
and enters into intermediary metabolism. (It does not
include dietary fibre, which can be a source of energy
only after fermentation - see the following
subsections.) Available carbohydrate can be arrived at
in two different ways: it can be estimated by
difference, or analysed directly.[6] To calculate
available carbohydrate by difference, the amount of
dietary fibre is analysed and subtracted from total
carbohydrate, thus:
100 - (weight in grams [protein + fat + water + ash +
alcohol + dietary fibre] in 100 g of food.)

IV.

Conclusion

Different analysis are performed in order to

determine the components in the sample oats.
Using certain methods of analysis, the values
moisture, ash, carbohydrates, fats and protein
were calculated. Using gravimetric analysis,
moisture is 14.01 0.405 ,Using simple
extraction with ether, fat content isfound to be
6.557 0.2677. Ash content is also obtained
which is equal to 2.557 0.0343representing the
inorganic substance present. Usingdifference
analysis
,
carbohydrates
is

computed be about 63.2112

using the

mean average of each component. P rotein,

using theuniversal standard method of analysis
which is Kjeldahl method is 13.37 0.5701 .

The values obtained is summarized in the

table above and its relative error. Possible factors
of error include inexact measurements, improper
handling of materials or uncalibrated equipment.
Also, further improvement in the analysis
method must still be considered

South Australian Resource and Development

Incorporation
Department of Agriculture and Food
FAO
Corporate
methodsoffoodanalysis

Document

Repository

People.umass.edu/mcclemen Carbohydrates

V.

Reference

Worlds healthiest food.org

Websites
Golden Nutritious Food Corporation

VI.
Appendixes
A. Moisture
Moisture
Trial
wt. crucible (g)
wt. wet sample (g)
wt. crucible + wet sample (g)
wt. crucible + dry sample (g)
wt. dry sample (g)

1
34.7506
2.0063
36.7569
36.5043
1.7537

2
29.3826
2.0035
31.3861
31.1397
1.7571

3
34.427
2.0098
36.4368
36.1963
1.7693

wt. water (g)

%moisture
Average %Moisture
Std. Dev

0.2526
14.40
14.01
0.4057

0.2464
14.02

0.2405
13.59

dry wt . sample (g)=( wt . crucible +dry sample ) (constant wt .crucible )

T1:

T3:

dry wt . sample ( g )=36.504334.7506

= 1.7537 g
T2:

dry wt . sample ( g )=31.139729.3826

= 1.7571 g

dry wt . sample ( g )=36.196334.4270

= 1.7693 g

wt . water ( g )=dry wt . samplewet wt . sample

T1:

= 0.2464 g

wt . water ( g )=1.75372.0063
= 0.2526 g

T3:

wt . water ( g )=1.76932.0098

T2:

= 0.2405 g

wt . water ( g )=1.75712.0035

%Moisture=

wt . water
100
wt . dry sample

T1:

T3:

%Moisture=

0.2526
100
1.7537

%Moisture=

= 14.40%
T2:

%Moisture=

0.2405
100
1.7693
= 13.59%

%Moisture ave =

%Mois 1+%Mois 2+%Mois 3

3

%Moisture ave =

14.40+14.02+13.59
3

0.2464
100
1.7571
= 14.02%

= 14.01 %
B. Ash
Ash
Trial
wt. crucible
wt. crucible + dry sample
wt. ash
wt. dry sample
%Ash

1
23.07
23.1147
0.0447
1.7537
2.549

2
18.8373
18.8817
0.0444
1.7571
2.527

3
23.3497
23.3956
0.0459
1.7693
2.594

Average % Ash
Std. Dev

2.557
0.0343

wt . ash ( g )= ( wt . crucible+ dry sample )wt . crucible

T1:

= 0.0444
T3:

wt . ash ( g )=23.114723.07

wt . ash ( g )=23.395623.3497

= 0.0447
T2:

= 0.0447

wt . ash ( g )=18.881718.8373

%Ash=

wt . ash
100
wt . dry sample

T1:

%Ash=

0.0447
100
1.7537

%Ash=

= 2.549 %

= 2.594 %

T2:

%Ash=

0.0459
100
1.7693

%Ashave =

%Ash 1+%Ash 2+%Ash 3

3

%Ashave =

2.549+2.527+2.594
3

0.0444
100
1.7571

= 2.527 %

= 2.557 %
T3:
C. Fat Content
Fat Content
Trial
wt. sample
wt. constant vial

1
7.5339
52.3008

2
7.5022
50.3177

3
7.5101
50.9275

wt. constant vial + oil

wt. oil
wt. dry sample
% fat
Average % Fat
Std. Dev.

52.4211
0.1203
1.7537
6.8598
6.557
0.267705632

50.4293
0.1116
1.7571
6.3514

51.0418
0.1143
1.7693
6.4602

wt . oil ( g )= ( wt . vial+ sample )wt . vial

T1:

T3:

wt . oil ( g )=52.421152.3008

wt . oil ( g )=51.041850.9275

= 0.1203

= 0.1143

T2:

wt . oil ( g )=50.429350.3177
= 0.1116

%Fat=

wt . oil
100
wt . dry sample

T1:

%Fat=

0.1203
100
1.7537

= 6.8598 %
T2:

%Fat=

%Fat=

0.1143
100
1.7537

= 6.4602%

%Fat ave=

%Fat 1+%Fat 2+%Fat 3

3

%Fat ave=

6.8598+6.3514 +6.4602
3

0.1116
100
1.7537

= 6.3514 %

= 6.557 %
T3:
D. Crude Protein

Crude Protein Determination

Trial
Standardization of NaOH
Mass of Std. KHP (g)
Corrected Mass of Std. KHP (g)
Volume NaOH used (ml)
Molarity of NaOH (mol/L)
Average Molarity of NaOH
Standardization of HCl
Volume of HCl (ml)
Volume NaOH used (ml)
Molarity of HCl (mol/L)
Average Molarity of HCl
Crude Protein Content
Mass of sample (g)
Mass of dry sample (g)
Volume of HCl (ml)
Volume NaOH added (ml)
Volume of HCl titrated (ml)
Amount HCl reacted (mmol)
Percent Nitrogen (%)
Percent Crude Protein (%)
Average Crude Protein (%)
Standard Deviation

0.7063
0.7056
36.20
0.0954
5
0.09463

0.7002
0.6995
36.40

22.90
20.00
0.1083
5
0.10520

22.40
20.00
0.1059
9

21.40
20.00
0.1012
6

2.0013
1.7553
50.00
36.40
8.75
2.74
2.18
12.73
13.37
0.5701

2.0079
1.7611
50.00
32.85
9.80
3.18
2.53
13.82

2.0089
1.7620
50.00
32.80
9.20
3.12
2.48
13.56

0.7019
0.7012
36.40
0.0943
0.09411
4

Amount HCl reacted=[ ( vol . HCl+ vol. HCl titrated )M HCl ] (Vol . NaOHM NaOH )
T1:

T3:

Amount of HCl reacted = 2.74

Amount of HCl reacted = 3.12

T2:
Amount of HCl reacted = 3.18

[(

amount HCl reacted )14.007

g
mol

1000
wt dry sample

%Nitrogen=

]
100

T1:

T3:

% Nitrogen = 2.18 %

% Nitrogen = 2.48 %

T2:
% Nitrogen = 2.53 %

%Crude Protein= Nitrogenjones factor (5.83)

T1:

%Crude protein ave=

% Crude Protein = 12.73 %

T2:

%Fat ave=

% Crude Protein = 13.82 %

T3:

12.73+13.82+13.56
3

= 13.37 %

% Crude Protein = 13.56 %

E. Carbohydrate
Carbohydrate
% Moisture
% Ash
% Fat
% Protein
% Carbohydrate

%Carbs=100%Mois%Ash%Fat%Protein
%Carbs=63.211

%CP 1+%CP2+%CP 3
3

14.01
2.549
6.8598
13.37
63.2112