Standard preparations— Prepare a solution of freshly ignited zinc oxide in dilute hydrochloric acid (1in 60)to obtain the equivalent of 1.0mg of zinc per mL.Dilute quantitatively with water to obtain separate solutions containing the equivalent of 15and 30µg of zinc per mL.

Assay preparation— Transfer about 1.0g of Ointment,accurately weighed,to a 100-mLbeaker.Add 25mLof dilute hydrochloric acid (1in 20),swirl,and heat carefully until the mixture is liquefied.Cool,and transfer the mixture to a 250-mLseparator.Complete the transfer of the waxy residue by thoroughly rinsing the beaker with 50mLof water and two 50-mLportions of chloroform and adding the rinsings to the separator.Equilibrate the mixture,and transfer the chloroform extract to a 500-mLseparator.Extract the aqueous phase with another 100-mLportion of chloroform,combine the second chloroform extract with the main extract in the 500-mLseparator,and transfer the aqueous phase to a 200-mLvolumetric flask.Wash the combined chloroform extracts with three 25-mLportions of water,add the aqueous washings to the 200-mLvolumetric flask,dilute with water to volume,and mix to obtain a specimen stock solution.[NOTE—Retain the chloroform extract for the Assay for undecylenic acid.]Transfer 15.0mLor other suitable volume (see Procedure)of this specimen stock solution to a 100-mLvolumetric flask,dilute with water to volume,and mix.

Procedure— Aspirate each Standard preparationand the Assay preparationinto the flame of a suitable atomic absorption spectrophotometer,and determine the absorbances of the solutions at 214nm.Typically,an acetylene–air mixture is adjusted to obtain a blue flame about 7mm in height with a suitable burner that is rotated to a position perpendicular to the light path.[NOTE—If the absorbance of the Assay preparation is outside the central 70%of the range between the absorbances of the Standard preparations,discard the Assay preparationand prepare another by diluting the specimen stock solution quantitatively as necessary to obtain a suitable absorbance.]Calculate the percentage of zinc undecylenate in the Ointment taken by the formula: in which 431.94is the molecular weight of zinc undecylenate;65.39is the atomic weight of zinc;Wis the weight,in g,of Ointment taken;AU,AH,and ALare the absorbances of the Assay preparationand the high-and low-concentration Standard preparations,respectively;and CHand CLare the concentrations,in µg per mL,of the high-and low-concentration Standard preparations,respectively.

Internal standard solution— Prepare a solution in chloroform containing 10mg of tridecanoic acid in each mL.

Standard preparation— Dissolve an accurately weighed quantity of USP Undecylenic Acid RSin chloroform to obtain a solution having a known concentration of about 3.8mg per mL.Transfer 5.0mLof this solution to a 50-mLvolumetric flask,add 3.0mLof Internal standard solution,dilute with chloroform to volume,and mix.

Assay preparation— Pass the chloroform extract prepared from the Ointment as directed in Assay for zinc undecylenatethrough phase-separating filter paper into a 250-mLvolumetric flask.Rinse the separator with three 15-mLportions of chloroform,passing the rinsings through the filter and combining them with the main chloroform solution,add chloroform to volume,and mix.Transfer 20.0mLof this solution to a 50-mLvolumetric flask,add 3.0mLof Internal standard solution,dilute with chloroform to volume,and mix.

Chromatographic system— Under typical conditions,the gas chromatograph is equipped with a flame-ionization detector and contains a 2-mm ×1.8-m glass column packed with 3%liquid phase G1on 100-to 200-mesh support S1A.The column is maintained at a temperature of about 165.Dry helium is used as the carrier gas at a flow rate of about 30mLper minute.

System suitability— Chromatograph five injections of the silylated Standard preparation,and record peak responses as directed for Procedure:the resolution,R(see Chromatography á621ñ),is not less than 3.0.

Procedure— Transfer 1.0-mLportions of the Standard preparationand the Assay preparationto separate,stoppered test tubes.To each tube add 50µLof bis(trimethylsilyl)trifluoroacetamide,insert the stopper,mix,and allow to stand for 30minutes.Inject a suitable portion (2to 5µL)of the Standard preparationinto a suitable gas chromatograph,and record the chromatogram so as to obtain not less than 50%of maximum recorder response.Similarly inject a suitable portion of the Assay preparation,and record the chromatogram.Measure the peak responses for the first (undecylenic acid)and second (tridecanoic acid)peaks of the chromatograms.[NOTE—Relative retention times are,approximately,0.43for undecylenic acid and 1.0for tridecanoic acid.]Calculate the percentage of total undecylenic acid in the Ointment taken by the formula: in which Cis the concentration,in mg per mL,of USP Undecylenic Acid RSin the Standard preparation;Wis the weight,in g,of Ointment taken;and RUand RSare the ratios of the peak responses of undecylenic acid to those of tridecanoic acid from the Assay preparationand the Standard preparation,respectively.The difference between the total undecylenic acid and the undecylenic acid equivalent to the determined zinc undecylenate (the weight of zinc undecylenate multiplied by 0.8533gives the equivalent of undecylenic acid),both expressed as a percentage of the Ointment,gives the percentage of free undecylenic acid in the Ointment.