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PRACTICAL 2: DETERMINATION OF FAT

INTRODUCTION
Lipids are one of the major constituents of foods, and are important in our
diet for a number of reasons. They are a major source of energy and provide
essential lipid nutrients. Nevertheless, over-consumption of certain lipid
components can be detrimental to our health, e.g. cholesterol and saturated fats.
In many foods the lipid component plays a major role in determining the overall
physical characteristics, such as flavour, texture, mouth feel and appearance. For
this reason, it is difficult to develop low-fat alternatives of many foods, because
once the fat is removed some of the most important physical characteristics are
lost. Lipids are usually defined as those components that are soluble in organic
solvents (such as ether, hexane or chloroform), but are insoluble in water. This
group of substances includes triacylglycerols,
diacylglycercols, monoacylglycercols, free fatty acids, phospholipids,
sterols, caretonoids and vitamins A and D. The lipid fraction of a fatty food
therefore contains a complex mixture of different types of molecule.
It is important to be able to accurately determine the total fat content in
foodsfor a number of reasons such as economic which avoid to give away
expensive ingredients in the food products. Besides that, the quality and food
properties of the product depend on the lipid content on it. It is also important for
legal conformation to standards of identity and nutritional labelling laws and
health. A variety of techniques are available to perform quantitative analysis.
The sample material, the time frame and the level of accuracy are the deciding
factors to select the suitable technique. Among of the techniques are Goldfisch
Fat Extractor (Continnuous), Soxhlet method (Semi-continuous) and Mojonnier
method (Discontinuous). In this experiment, we will only do Soxhlet method and
Mojonnier Method.
Soxhlet Extraction method is a continuous solvent method that is usually
used to remove fats from the food sample. Soxhlet also can be used to
determine crude fat content. Crude fat content is determined by extracting the
fat from the sample using a solvent, and then weigh the fat extracted. Mojonnier
Method is a discontinuous solvent extraction method and does not require
removal of moisture from the sample.

SOXHLET EXTRACTION METHOD


OBJECTIVE
To determine the fat content of selected food samples using Soxhlet method.

PRINCIPLE
Fat is extracted semi continuously with an organic solvent. Solvent is heated and
volatilized, then is condensed above the sample. Solvent drips onto the sample
and soaks it to extract the fat. At 15-20 minutes intervals, the solvent is
siphoned to the heating flask, to start the process again. Fat content is measured
by weight of fat removed.

APPARATUS

Analytical balance
Dessicator
Soxhlet apparatus
Thimbles
Round bottom flask 250 mL
Heating mantle
Water bath
Graduated cylinder 500 mL
Beaker 250 mL
Mortar and pestle
Spatula
Oven

CHEMICAL
Pet ether (boiling point: 40-60 )

PROCEDURE
1. About 1 g of dried sample was weighed accurately into an extraction thimble
or a piece of filter paper.
2. The opening of the thimble was loosely plugged with cotton or fold the filter
and plug with cotton.

3. The thimble or paper and content was placed into a Soxhlet extractor.
4. A dried round bottom flask was weighed accurately and 150 mL petroleum
ether was added.
5. The apparatus was connected to the condenser. The water was turned on and
extracted for 2 hours on an electrothermal extraction unit.
6. After the extraction has completed, the flask containing the petroleum ether
extract was removed.
7. The petroleum ether was evaporated on a boiling water bath.
8. The flask was transferred into an oven at 105

for one hour to dry the

extract.
9. The flask was immediately transferred into a dessicator to cool and weigh.

MOJONNIER METHOD
OBJECTIVE
To determine the fat content of selected food samples using Mojonnier method.

PRINCIPLE
Fat is extracted with a mixture of ethyl ether and petroleum ether, and the
extracted fat is dried to constant weight and expressed as percent fat by weight.
The assay uses not only diethyl ether and petroleum ether, but also ammonia
and ethanol.

APPARATUS

Mojonnier extraction flask with stopper


Evaporating dish
Analytical balance
Oven
Dessicator
Graduated cylinder 25mL

CHEMICALS

Ammonium hydroxide
Ethanol
Diethyl ether

Petroleum ether (boiling point: 40 60

PROCEDURE
FIRST EXTRACTION
1.
2.
3.
4.
5.
6.

10 g milk was weighed to the nearest 0.1 mg into a Mojonnier fat extraction
flask.
1.5 mL NH4OH was added and was shake vigorously.
10 mL of 95% ethanol was added and was shake for 90 seconds.
25 mL of diethyl ether was added and was shake for 90 seconds.
25 mL of petroleum ether was added and was shake for 90 seconds.
Ether solution was decanted fromthe Mojonnier flaskinto the previously
weighed evaporating dish.

SECOND EXTRACTION
1.
2.
3.
4.

5 mL of 95% ethanol was added and shake vigorously for 15 seconds.


15 mL of diethyl ether was added and shake for 60 seconds.
15 mL petroleum ether was added shake for 60 seconds.
Solution was decanted into the same evaporating dish.

THIRD EXTRACTION
1. 15 mL of diethyl ether was added and shake for 60 seconds.
2. 15 mL of petroleum ether was added and shake 60 seconds.
3. Solution was decanted into the same evaporating dish.

4. The solvent was distilled from the flask, the fat was dried at 105
hour, then it was cooled and weighed.

CALCULATION
Trial 1:
Weight of fat in sample = 124.1217 g 123.7954 g
= 0.3263 g
% Fat in Sample =

0.3263
1.0131

g 100

= 32.21%

Trial 2:
Weight of fat in sample = 124.8453 g 124.4432 g

for 1

= 0.4021 g
% Fat in sample =

0.4021
1.0004

g 100

= 40.19%

Trial 3:
Weight of fat in sample = 124. 5480 g 124.1451 g
= 0.4029 g
% Fat in sample =

0.4029
1.0057

g 100

= 40.06%

Mean of weight of sample = 1.0131 g + 1.0004 g + 1.0057 g


3
= 1.0064 g
Mean of weight of round bottom flask (g) = 123.7954 + 124.4432 + 124.1451
3
= 124.1279 g
Mean of weight of round bottom flask + fat (g) = 124.1217 + 124.8453 +
124.5480
3
= 124.505 g
Mean of % Fat = 32.21 + 40.19 + 40.06
3
= 37.49%

DISCUSSION
In Soxhlet method, the fat is extracted semi continuously with an organic
solvent. The solvent will be heated and volatilized, then condensed above the
sample. Solvent drips onto the sample and soaks it to extract the fat. Fat content
is measured by weighed of fat removed. The errors that occur during this
experiment is the extraction take too much time than the actual time stated. This
is because the preparation of sample is influenced which is the solvent is poured
too much. The temperature also must be different and the sample does not have

accurate temperature during the water bath and drying. The weight may also be
the cause by not weighing accurately. To overcome this, the temperature of
water bath and oven must be set up first with the correct temperature. While
doing the weighing, weigh it correctly with 3 times to get constant reading. The
petroleum ether also need to be measure correctly.
The Mojonnier method, the fat is extracted with a mixture of ethyl ether
and petroleum ether, and the extracted fat is dried to constant weight.
Ammonium hydroxide and ethanol was also use as the reagent. The errors that
occur during the experiment are the time taken to shake the Mojonnier flask was
not constant which is more time or less time. The volume of the reagent that has
been put in the coconut milk also does not measured correctly. During the drying
process, the fat content cannot be weighed because the reagent does not react
or extracted correctly. To prevent this from happening, time taken and shaken
must be set first and the temperature need to be the same. So the weight of fat
content can be calculated.

CONCLUSION
The objective of this experiment is to determine the fat content of selected food
samples using Soxhlet method and to determine the fat content of selected food
sample using Mojonnier method. In conclusion, we know how to determine fat
content of selected food samples using Soxhlet method and Mojonnier method.
As a result, biscuits contain 37.49% of fat. So the objective was achieved.

QUESTIONS
1. Draw and label the main components of Soxhlet apparatus.

2. Describe the function of the various reagents used in each method.


Petroleum ether: Removes moisture from the ethyl ether extract
anddissolve non-polar lipid (reduce proportion of water and non-fatty
solublesubstances)
Ethanol: Precipitates protein and prevents possible gel formation.
Diethyl ether: Dissolved the lipids
Ammonia: To neutralizes acidic samples and dissolve protein
3. Which is the best extraction procedure:
a) One extraction with a large volume of solvent OR
b) Several extractions with smaller volumes but the same total volume?
Explain
The best extraction method is several extractions with smaller
volumes but the same total volume because the steps are simple
and reading is more accurate compared with large volume of
solvent.

REFERENCES

https://www.scribd.com/doc/203587765/Lab-Food-Analaysis-fat
https://en.wikipedia.org/wiki/Fat
http://pearl1.lanl.gov/external/ccde/scf/pubs/king/102_determination_of_fat_content.pdf