A: Infrared Absorption 197F.

B: Thin-Layer Chromatographic Identification Test 201—

Caution—Ethylene oxide is toxic and flammable. Prepare all solutions in a well-ventilated hood. The operator must protect hands and face by wearing polyethylene protective gloves and an appropriate face mask. Store all solutions in hermetic containers, and refrigerate at a temperature between 4 and 8.

note—Perform all determinations three times.

Ethylene oxide stock solution— Into a dry, clean test tube, cooled in a mixture of 1 part of sodium chloride and 3 parts of crushed ice, introduce a slow current of ethylene oxide gas, allowing condensation onto the inner wall of the test tube. Using a glass syringe, previously cooled to 10, transfer about 300 µL of liquid ethylene oxide, equivalent to about 0.25 g, to 50 mL of polyethylene glycol 200. Determine the absorbed quantity of ethylene oxide by weighing before and after absorption. Dilute with polyethylene glycol 200 to 100.0 mL, and mix. This is the Ethylene oxide stock solution.

Ethylene oxide solution— Quantitatively dilute a volume of Ethylene oxide stock solution, accurately measured, with polyethylene glycol 200 to obtain a solution containing about 50 µg of ethylene oxide per g. Dilute 1.0 mL of this solution with water to 5.0 mL to obtain a solution having a known concentration of about 10 µg of ethylene oxide per mL. [note—Prepare immediately before use.]

Dioxane stock solution— Dissolve about 1.00 g of dioxane, accurately weighed, in water, and dilute quantitatively, and stepwise if necessary, with water to obtain a solution having a known concentration of about 1.0 mg per mL.

Dioxane solution— Quantitatively dilute a volume of the Dioxane stock solution, accurately measured, with water to obtain a solution having a known concentration of about 0.5 mg of dioxane per mL.

Standard solution 1— Transfer about 1.0 g of the substance under test, accurately weighed, to a 10-mL vial, and add 1.0 mL of N,N-dimethylacetamide, 0.1 mL of Ethylene oxide solution, and 0.1 mL of Dioxane solution. Close the vial, and mix to obtain a homogenous solution. Allow to stand at 90 for 45 minutes.

Standard solution 2— Transfer 0.1 mL of Ethylene oxide solution to a 10-mL vial, add 0.1 mL of a freshly prepared solution of acetaldehyde, containing about 10 mg of acetaldehyde per L, and add 0.1 mL of Dioxane solution. Close the vial, and mix to obtain a homogenous solution.

Test solution— Transfer about 1.0 g of the substance under test, accurately weighed, to a 10-mL vial, and add 1.0 mL of N,N-dimethylacetamide and 0.2 mL of water. Close the vial, and mix to obtain a homogenous solution. Allow to stand at 90 for 45 minutes.

Chromatographic system (see Chromatography 621)— [note—Headspace apparatus that automatically transfers a measured amount of headspace may be used.] The gas chromatograph is equipped with a flame-ionization detector and contains a 0.32-mm × 30-m glass or quartz capillary column bonded with a 1.0-µm layer of phase G1. The carrier gas is helium, flowing at a rate of about 1 mL per minute. The detector and injection port temperatures are maintained at 250 and 150, respectively. The column temperature is programmed as follows. Initially it is maintained at 50 for 5 minutes after injection, then increased to 180 at a rate of 5 per minute, further increased to 230 at a rate of 30 per minute, and maintained at this temperature for 5 minutes. Chromatograph the gaseous phase of Standard solution 2, and record the peak responses as directed for Procedure, adjusting the sensitivity of the system so that the peak heights of the two principal peaks in the chromatogram are not less than 15% of the full scale of the recorder: the relative retention times are about 0.94 for acetaldehyde and 1.0 for ethylene oxide; the resolution, R, between acetaldehyde and ethylene oxide is not less than 2.0; and the relative standard deviation for replicate injections is not more than 15.0%.

Procedure— Using a heated, gas-tight gas chromatographic syringe, separately inject equal volumes (about 1 mL) of the gaseous headspace of Standard solution 1 and the Test solution into the chromatograph, record the chromatograms, and measure the peak responses: the mean areas of the ethylene oxide and dioxane peaks in the chromatogram obtained from the Test solution are not greater than half the mean areas of the corresponding peaks in the chromatogram obtained from Standard solution 1, equivalent to about 1 µg of ethylene oxide per g and 50 µg of dioxane per g. Calculate the concentration of ethylene oxide, in µg per g, in the test specimen taken by the formula: in which C is the concentration, in µg per mL, of ethylene oxide in Standard solution 1; rU and rS are the peak responses of ethylene oxide obtained from the Test solution and Standard solution 1, respectively; and MU and MS are the quantities, in g, of the substance under test taken to prepare the Test solution and Standard solution 1, respectively: not more than 1 µg per g is found. Calculate the concentration of dioxane, in µg per g, in the test specimen taken by the formula: in which CD is the concentration, in µg, of dioxane in Standard solution 1; dU and dS are the peak responses of dioxane obtained from the Test solution and Standard solution 1, respectively; and MU and MS are as defined above: not more than 10 µg per g is found.