SOP for ESTIMATION OF IODINE

PRINCIPLE:

Mercuric chloride added to aqueous elemental iodine solutions causes complete hydrolysis 

of iodine to give hypoiodus chloride. Leuco crystal violet reacts with hypoiodus acid to form 

crystal violet dye.

MINIMUM DETECTION LIMIT:  10 µg I as I₂ / l

REAGENTS:

1.    Stock Iodine solution: Prepare a saturated iodine solution by dissolving 20 g 

elemental iodine in 300 ml water. Let stand several hours. Decant iodine solution and dilute 

170 ml to 2000 ml. Standardize solution by titrating with standard sodium thiosulphate

 titrant. Calculate iodine concentration:

      mg I as I₂/ml = normality of iodine solution x 126.9

      Prepare a working solution of 10mg as I₂/ml by approximate dilution of the standardized 

stock solution.

2.    Citric buffer solution: pH 3.8

a)    Citric acid: Dissolve 192.2 g C₆H₈O₇ or 210.2 g C₆H₈O₇.H₂O and dilute to 1 liter

 with distilled water.

b)   Ammonium hydroxide (2N): Add 131 ml concentrated ammonium hydroxide to about 

700 ml water and dilute to 1 lit. Store in a polyethylene bottle.

c)    Final buffer solution: slowly add with mixing 350 ml 2 N ammonium hydroxide solution 

to 670 ml citric acid. Add 80 g ammonium dihydrogen phosphate and stir to dissolve.

3.    Leuco crystal violet indicator: Measure 200 ml water and 3.2 ml concentrated sulfuric 

acid into a brown glass container of at least 1 Lit capacity mix with magnetic stirrer at 

moderate speed. Add 1.5 g 4,4’,4” – methylidynetris (N, N dimethylaniline) and with a small

 amount of water wash down any reagent adhering to neck  or sides of container. Mix 

until dissolve.To 800 ml water add 2.5 g mercuric chloride and stir to dissolve. With mixing 

add mercuric chloride solution to leuco crystal violet solution. For maximum stability, adjust pH

 of final solution to 1.5 or less, adding, if necessary, concentrated sulfuric acid dropwise. Store

 in brown glass bottle away from sunlight. Discard after 6 months. Do not use a rubber stopper.

4.    Sodium thiosulphate solution: Dissolve 5 g sodium thiosulphate in water and dilute to 

1 lit.

PROCEDURE:

1] For accuracy standardize working solution immediately. Prepare standards in the range of 0.1 to 

6 mg I as I2/L by adding 1 to 60 ml working solution to 100 ml volumetric flask in increments of 1 ml 

or larger. Measure 50 ml of each diluted iodine working solution in to a 100 ml volumetric flask and 

add 1 ml citric buffer solution gently swirl to mix and let stand at least 30 sec. Add 1 ml leuco crystal 

violet indicator and swirl to develop colour. Dilute to 100 ml and mix. Measure the absorbance at 592

 nm.

2] Measure 50 ml of sample in 100 ml volumetric flask and treat as per temporary 

iodine standards.

3] For sample contain > 6.0 mg I as I2/L: Place approximately 25 ml H2O in a 100 ml 

volumetric flask. Add 1 ml citric buffer solution and a measured volume of 25 ml or less of

 sample mix and let it stand for 30 sec. Add 1 ml leuco crystal violet indicator mix and 

dilute to mark and read absorbance at 592 nm.

Select one of the following sample volumes to remain within optimum iodine range.

Iodine range (mg/L)                                         Sample volume required (ml)

6.0 – 12.0                                                             25.0

12.0 – 30.0                                                          10

30 – 60                                                                 5

4] Samples containing free and combined chlorine and iodine follow the procedure above [2]

 or [3]. But read the absorbance within 5 min after adding leuco crystal violet indicator.

5] Compensation for turbidity and color: compensation for natural colour or turbidity by 

adding 5 ml sodium thiosulphate to a 50 ml sample and proceed as above.

REFERENCE: APHA – 4500 I B – Leuco Crystal Violet Method