INTRODUCTION

Lipids are relatively non-polar chemical substances found in plant bacteria and animal cells.

In the first phase of this experiment, a lipid extract of ground nutmeg was partially

crystallized by crystallization from acetone. Nutmeg is approximately 25% fat by weight; is

simple to extract. Nutmeg is composed nearly of a single type of fatty acid. Nutmeg is broken

down; the liquid fats can be isolated using filtration methods. The triglycerides obtained are

used in a variety of products in industries. Cooks often use these as fats, or oils if they are a

liquid, for making food product or to aid them in deep frying after further processing.

Nutmeg is a spice made from seed of a tree that grows in hot countries. Nutmeg is usually

used to flavor sweet food. It is also used as a preservative agent. Major producers of nutmeg

are Indonesia and Sri Lanka. The amount of essential oil found in the seed varies with origin,

climate, and soil. There is a wide range of essential oil content from 5 to 15 wt %. About 80%

of the essential oil is composed of terpenes. Other important components are safrol, elemicin

and myristicin, of which the last one is responsible for the characteristic aroma of nutmeg.

Nutmeg is applied in medicinal drug, perfume, and shampoo formulations.

The second phase of the experiment dealt with the determination of acid value of fats. The

acid value (AV) is a common parameter in the specification of fats and oils. It is defined as

the weight of KOH in mg needed to neutralize the organic acids present in 1g of fat and it is a

measure of the free fatty acids (FFA) in a sample of oil or fat with hydrolysis of triglycerides.

Such reaction occurs by the action of lipase enzyme, and it is and indicator of inadequate

processing and storage conditions (i.e., high temperature, and relative humidity, tissue

damage). Besides of free fatty acids, hydrolysis of triglycerides produces glycerol. RCOOH +

KOH → ROO- K+H2O. Free fatty acids are a source of flavours and aromas. On one side,

short chain of free fatty acids tend to be water soluble and volatile with characteristic smell.
On the other hand, we have long chain saturated and unsaturated fatty acid. The later are

more prone to oxidation in their free form and their breakdown products (aldehydes, ketones,

alcohols, and organic acids) provide characteristic flavours and aromas. In most cases, these

flavour and aromas are considered a defect in oils, fats, and foods that contain them.

However, there are instances where hydrolysis of triglycerides and oxidation of free fatty acid

are key in the development of desirable flavour and aroma in foods. This is the case of aged

cheeses and some processed meats.

There are standard methods for determining the acid number, such as ASTM D 974 and DIN

51558 (for mineral oils, biodiesel), or specifically for Biodiesel using the European Standard

EN 14104 and ASTM D664 are both widely utilised worldwide. Acid number (mg KOH/g

oil) for biodiesel should be lower than 0.50 mg KOH/g in both EN 14214 and ASTM D6751

standard fuels. This is since the FFA produced may corrode automotive parts and these limits

protect vehicle engines and fuel tanks. As oil-fats rancidify, triglycerides are converted into

fatty acids and glycerol, causing an increase in acid number. A similar observation is

observed with Biodiesel aging through analogous oxidation processes and when subjected to

prolonged high temperatures (ester thermolysis) or through exposure to acids or bases

(acid/base ester hydrolysis).

In addition, the fats often become rancid during storage and this rancidity is caused by

chemical and enzymatic hydrolysis of fats into free acids and glycerol. The amount of free

fatty acids can be determined volumetrically by titrating the sample with potassium

hydroxide. The acidity of fats and oils is expressed as its acid value or number which is

defined as mg KOH required to neutralize the free fatty acids present in 1g of fat or oil. The

number of free acids present, or acid value of fat is a useful parameter which gives an

indication about the age and extent of its deterioration.

The aim of the experiment is to extract a lipid and find the acid values of fats.
MATERIALS

Nutmeg, Hexane: Isopropanol (3:2), Acetone, Diethyl ether, Erlenmeyer flask, Extracted

lipid, Soya bean oil, Vegetable oil, Margarine, Fat solvent (95% v/v ethanol and diethyl

ether ), Phenolphthalein ( 10 g/I in alcohol ), Potassium hydroxide ( 0.1 mol/l ), Burettes ( 5

ml and 25 ml )

METHOD

To isolate and purify the unknown lipid, 6 grams of finely ground nutmeg was weighed into a

100 Erlenmeyer flask. 40 ml of extraction solvent was added and warmed on a water bath (45

to 50 degree Celsius) in a hood. The solution was mixed while being heated for 15 minutes.

Filtered through fluted filter paper into a pre-weighed beaker, the residue was rinsed with 50

ml extraction solvent. The solvent was removed from extract by evaporation using a boiling

water to obtain yellow or off- white solid. The crude extract was weighed.

To purify the extract, the crude extract was dissolved in about 40 ml of acetone on a steam

bath (in a fume hood). The solution was cooled slowly to room temperature and further

cooled on ice to allow extract to form. The mixture was then filtered, washed with acetone

(25 ml) and weighed.

In finding the acid value, the extracted lipid from previous experiment in addition to test

samples Vegetable oil, Soyabean oil, Olive oil, and margarine were used. 0.5 grams from the

extracted lipid was weighed and 1 gram which is equivalent to 32 drops of the test samples

were weighed into a 100 ml Erlenmeyer flask and about 10 ml fat solvent was added. The test

samples included rancid and fresh oils each. 1 to 3 drops phenolphthalein solution was added

and mixed thoroughly. Titration was done with 0.1 M Potassium hydroxide until the faint
pink color persisted for 20 to 30 seconds. The number of milligrams per standard alkali

required was noted and the acid value of the fat calculated.
RESULTS

TABLE 1.0 A TABLE OF RESULTS OF THE ACID VALUE OF FRESH AND RANCID

TEST SAMPLES.

SAMPLE RANCID OIL FRESH OIL MASS/ g ACID VALUES

TITRE TITRE RANCID FRESH

VALUE/ ml VALUE/ ml

Vegetable oil 1.0 0.1 1.0 5.6 0.6

Soyabean oil 1.3 0.2 1.0 7.2 1.1

Olive oil 0.1 0.1 1.0 0.6 0.6

Margarine 4.9 0.2 1.0 27.5 1.1

TABLE 2.0 A TABLE OF THE ACID VALUE OF THE EXTRACTED LIPID

SAMPLE TITRE / ml MASS ACID VALUE

Extracted Lipid 0.7 0.5

The table above shows each of the titre values that were obtained from titrating our samples

with 0.1 M Potassium hydroxide. The table also shows the masses of each sample used and

their corresponding acid values calculated in appendix 1.0.