Tocopherols Excipient »Tocopherols Excipient is a vegetable oil solution containing not less than 50.0percent of total tocopherols,of which not less than 80.0percent consists of varying amounts of beta,gamma,and delta tocopherols. Packaging and storage— Preserve in tight containers,protected from light.Protect with a blanket of an inert gas. Labeling— Label it to indicate the content,in mg per g,of total tocopherols and of the sum of beta,gamma,and delta tocopherols. USP Reference standards á11ñ USP Alpha Tocopherol RS. Identification— A: Dissolve about 50mg in 10mLof dehydrated alcohol,add,with swirling,2mLof nitric acid,and heat at about 75for 15minutes:a bright red or orange color develops. B: The retention time of the third major peak (i.e.,the peak occurring just before that of the internal standard)in the chromatogram of the Assay preparationcorresponds to that of the Standard preparation,both relative to that of the internal standard,as obtained in the Assay. Acidity— Dissolve 1.0g in 25mLof a mixture of equal volumes of alcohol and ether (which has been neutralized to phenolphthalein with 0.1Nsodium hydroxide),add 0.5mLof phenolphthalein TS,and titrate with 0.10Nsodium hydroxide until the solution remains faintly pink after being shaken for 30seconds:not more than 1.0mLof 0.10Nsodium hydroxide is required. Organic volatile impurities,Method IVá467ñ: meets the requirements. Assay — Internal standard solution— Transfer about 600mg of hexadecyl hexadecanoate,accurately weighed,to a 200-mLvolumetric flask,dissolve in a diluting solution containing 2volumes of pyridine and 1volume of propionic anhydride,dilute with the diluting solution to volume,and mix. Standard preparations— [NOTE—Use low-actinic glassware.]Transfer 12-,25-,37-,and 50-mg portions of USP Alpha Tocopherol RS,accurately weighed,to separate 50-mLconical flasks having 19/38standard-taper ground-glass necks.Pipet 25mLof the Internal standard solutioninto each flask,mix,and reflux for 10minutes under water-cooled condensers. Assay preparation— [NOTE—Use low-actinic glassware.]Transfer about 60mg of Tocopherols Excipient,accurately weighed,to a 50-mLconical flask similar to the flasks used in preparing the Standard preparations,add 10.0mLof Internal standard solution,mix,and reflux for 10minutes under a water-cooled condenser. Chromatographic system (see Chromatography á621ñ)—The gas chromatograph is equipped with a flame-ionization detector and contains a 4-mm ×2-m borosilicate glass column packed with 2%to 5%liquid phase G2on 80-to 100-mesh support S1AButilizing either a glass-lined sample introduction system or on-column injection.The column is maintained isothermally at a temperature between 245and 265,and the injection port and detector block temperatures are maintained at about 10higher than the column temperature.The flow rate of dry carrier gas is adjusted to obtain a hexadecyl hexadecanoate peak 30to 32minutes after sample introduction.[NOTE—Cure and condition the column as necessary.] System suitability— Chromatograph a sufficient number of injections of the Assay preparation,as directed under Calibration,to ensure that the resolution,R,between the major peaks occurring at retention times of approximately 0.50(delta tocopheryl propionate)and 0.63(beta plus gamma tocopheryl propionates),relative to hexadecyl hexadecanoate at 1.00,is not less than 2.5. Calibration— Chromatograph 2-to 5-µLportions of each Standard preparation,and record the peak areas as directed for Procedure.Calculate the relative response factor,F,for each concentration of the Standard preparationtaken by the formula: (AS/AD)(CD/CS), in which CDand CSare the concentrations,in mg per mL,of hexadecyl hexadecanoate and of USP Alpha Tocopherol RS,respectively,in the Standard preparation.Successively chromatograph a sufficient number of portions of each Standard preparationto ensure that the factor,F,is constant within a range of 2.0%.Prepare a relative response factor curve by plotting Fversus the alpha tocopheryl propionate peak area. Procedure— Inject a suitable portion (2µLto 5µL)of the Assay preparationinto the chromatograph,and record the chromatogram.Measure the areas under the 4major peaks occurring at relative retention times of approximately 0.50,0.63,0.76,and 1.00,and record the values as ad,abg,aa,and aD,corresponding to delta tocopheryl propionate,beta plus gamma tocopheryl propionates,alpha tocopheryl propionate,and hexadecyl hexadecanoate,respectively.Calculate the quantity,in mg,of each tocopherol form in the Tocopherols Excipient taken by the formulas: delta tocopherol =(10CD/F)(ad/aD); beta plus gamma tocopherols =(10CD/F)(abg/aD); alpha tocopherol =(10CD/F)(aa/aD), in which Fis obtained from the relative response factor curve (see Calibration)for each of the corresponding areas under the delta,beta plus gamma,and alpha tocopheryl propionate peaks produced by the Assay preparation.[NOTE—The relative response factor for delta tocopheryl propionate and for beta plus gamma tocopheryl propionates has been determined empirically to be the same as for alpha tocopheryl propionate.] Auxiliary Information— Staff Liaison:Ravi Ravichandran,Ph.D.,Senior Scientist Expert Committee:(EMC)Excipients:Monograph Content USP28–NF23Page 3098 Phone Number:1-301-816-8330