Practical Manual ---

Chemical Analysis and EXPERIMENT 26 DETERMINATION OF

Quality Assurance
ALLYL ISOTHIOCYANATE
IN MUSTARD OIL
Structure
26.0 Objectives
26.1 Introduction
26.2 Principle
26.3 Requirements
26.4 Procedure
26.5 Calculation
26.6 Results and Inference
26.7 Precautions

26.0 OBJECTIVES
After attending to this experiment, we shall be able to:

• learn to perform determination of allyl isothiocyanate in mustard oil.

26.1 INTRODUCTION
The oil obtained from mustard seeds and rape seeds contains sinigrin and myrosin
(glucosinolates), which are sources of goitrogens. As such they are relatively non-
toxic but their hydrolysis products include allyl isothiocyanates and oxazolidinethiones,
which are powerful goitrogens, producing varying manifestations of toxicity in non-
ruminants.

26.2 PRINCIPLE
The allyl isothiocyanate in the oil is steam distilled into a known excess of silver
nitrate solution, and the excess of silver nitrate solution is determined by titration with
standard ammonium thiocyanate solution.

26.3 REQUIREMENTS
Apparatus
Distillation Flask - 500 ml round-bottomed flask.
Any Efficient Reflux Condenser (~ 90 cm long).
Measuring Flask - 200 ml capacity.
Water-Bath
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Reagents Determination of Allyl
Isothiocyanate in
Ethyl Alcohol -95%, v/v neutral to phenolphthalein. Mustard Oil

Silver Nitrate Solution - 0.1 N.

Ammonium Hydroxide Solution - 10 % (w/v).
Conc. HNO3
Ferric ammonium sulphate Indicator - 0.1 % solution in water.
Standard Ammonium Thiocyanate (NH4SCN) Solution - 0.1 N.

26.4 PROCEDURE
Weigh accurately about 5 g of the material into a 500 ml distillation flask and add to
it 25 ml of ethyl alcohol, 250 ml of water and a few pieces of pumice stone. Distil
the mixture in steam and collect the distillate in a 200 ml measuring flask containing
exactly 25 ml of silver nitrate solution and 10 ml of ammonium hydroxide solution.
Collect as rapidly as possible about 150 ml of the distillate. Attach the reflux air
condenser to the measuring flask and heat the mixture for about one hour on a
boiling water-bath. Cool to room temperature, add water to make up to 200 ml and
filter the contents after shaking. Take 100 ml of the filtrate, add 6 ml of HNO3 and a
few drops of ferric ammonium sulphate indicator, and titrate with standard ammonium
thiocyanate solution until a permanent red colour is obtained. Carry out a blank test
simultaneously along with the sample.

26.5 CALCULATION
9.915 (B-S) × N
Allyl isothiocyanate, % by mass = ———————
W

where,
B = Volume, in ml, of NH4SCN solution required for blank,
S = Volume, in ml, of NH4SCN solution required for the sample,
N = Normality of NH4SCN solution, and
W = Weight, in g, of the sample taken for the test.

26.6 RESULTS AND INFERENCE

The difference between the results of two concurrent determinations carried out
simultaneously or in rapid succession by the same analyst (repeatability) shall not
exceed 0.02% by mass. The allyl isothiocyanate content in different varieties if
mustard varies from 0.1 to 0.7% by mass.

26.7 PRECAUTIONS
• Sample and blank test should be conducted simultaneously under similar
conditions.
• Standardized ammonium thiocyanate should be used.
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