Page 165 - Analytical method for food addtives

110 Analytical methods for food additives

(Note 1: It is essential to obtain a clear filtrate within the time specified. For this
purpose, it may be necessary to use a larger volume of each precipitation reagent
(for example, if well-matured cheeses are analysed). Reduce the volume of the
preheated water accordingly to maintain the volume of filtrate at about 200 mL.
Note 2: The total volume of filtrate approximates to about 200 mL. This is
regarded as such in the calculation.)

Reduction of nitrate to nitrite
Pipette 20 mL of the filtrate into the reservoir on top of the prepared reduction
column. Add 5 mL of buffer solution to the contents of the reservoir. Mix by
stirring with a small glass rod. Pass the contents of the reservoir through the
column at a flow rate not exceeding 6 mL/min. Collect the eluate in a 100 mL
volumetric flask. When the reservoir of the reduction column is nearly empty,
wash the walls of the reservoir with about 15 mL of water. After the water has run
off, repeat the same washing with another 15 mL of water. After the second
washing has run into the column, completely fill the reservoir with water. Pass the
complete content of the reservoir through the column at maximum flow rate.
Collect nearly 100 mL of the eluate in the 100 mL volumetric flask. Remove the
100 mL volumetric flask. Dilute its contents to the 100 mL mark with water and
mix well.

Colour development and measurement
Pipette equal volumes (for example 25 mL) of the filtrate obtained (from extraction
and deproteination) and of the eluate into separate 100 mL volumetric flasks. Add
water to each of the flasks to obtain a volume of about 60 mL and mix. Add to the
solution 6 mL of solution I (450 mL concentrated hydrochloric acid diluted to
1000 ml with water) and then 5 mL of solution II (0.5 g of sulphanilamide
dissolved in 75 mL water and 5 mL concentrated hydrochloric acid in a 100 mL
volumetric flask with heating on a water bath. Cool to room temperature and dilute
to the 100 mL mark with water and mix. The solution is filtered if necessary). Mix
carefully and leave the solution protected from direct sunlight at room temperature
for 5 min. Add to the solution 2 mL of solution III (0.1 g N-1-naphthyl ethylenedi-
amine dihydrochloride dissolved in water in a 100 mL volumetric flask and diluted
to the 100 mL mark with water and mixed. The solution is filtered if necessary).
Mix carefully and leave the solution protected from direct sunlight at room
temperature for 5 min. Dilute its contents to the 100 mL mark with water and mix.
Measure the absorbance of the solution III. Mix carefully and leave the solution
protected from direct sunlight at room temperature for 5 min. Dilute its contents to
the 100 mL mark with water and mix. Measure the absorbance of the solution
obtained above against that of the blank test at a wavelength of 538 nm within
15 min.
Blank test
Carry out a blank test in parallel with the determination but omitting the test
portion.