Dilute nitric acid— Bubble air through nitric acid until the acid is colorless, then mix 1 volume of the acid with 4 volumes of water.

Standard phenol solution— Dissolve about 1 g of phenol in about 100 mL of water, and determine the actual C6H6O concentration as follows. Pipet 4 mL of the solution into an iodine flask, add 30.0 mL of 0.1 N bromine VS, then add 5 mL of hydrochloric acid, and immediately insert the stopper. Shake the flask repeatedly during 30 minutes, allow to stand for 15 minutes, add quickly 5 mL of potassium iodide solution (1 in 5), taking precautions to prevent the escape of bromine vapor, and at once insert the stopper into the flask. Shake thoroughly, remove the stopper, and rinse it and the neck of the flask with a small quantity of water so that the washings flow into the flask. Add 1 mL chloroform, shake the mixture, and titrate the liberated iodine with 0.1 N sodium thiosulfate VS, adding 3 mL of starch TS as the endpoint is approached. Perform a blank determination. Each mL of 0.1 N bromine is equivalent to 1.569 mg of C6H6O. Dilute a suitable volume of the solution with water to obtain a concentration of 250 µg of C6H6O per mL.

Procedure— Place about 2.5 g of Cresol, accurately weighed, in a 250-mL volumetric flask, add 10 mL of sodium hydroxide solution (1 in 10), dilute with water to volume, and mix. Pipet 5 mL of this solution into a 200-mL volumetric flask, add 45 mL of water and 1 drop of methyl orange TS, neutralize with Dilute nitric acid, added dropwise, then dilute with water to volume, and mix. Pipet 5 mL of the neutralized solution into each of two 20- × 180-mm test tubes, graduated at the 25-mL mark, and pipet 5.0 mL of Standard phenol solution into each of two similar test tubes. To the contents of each tube add 5 mL of Millon's Reagent, allowing it to flow down the inner wall of the tube, mix, place the tubes simultaneously in a boiling water bath provided with a rack so that the tubes do not touch the bottom of the bath, and maintain the bath at boiling temperature for 30 minutes, accurately timed. At once remove the tubes from the bath, cool them immediately and thoroughly by placing them in a bath of cold water for not less than 10 minutes, add 5 mL of Dilute nitric acid to each tube, and mix. Add 3 mL of a mixture of 1 volume of formaldehyde solution and 50 volumes of water to one of each pair of tubes, add water to fill all tubes to volume, shake thoroughly, and allow to stand for 16 hours, during which time the added formaldehyde imparts a yellow color while the contents of the other 2 tubes acquire an orange-red color.

5V/W