(toe kof' er ols ex sip' ee ent).
» Tocopherols Excipient is a vegetable oil solution containing not less than 50.0 percent of total tocopherols, of which not less than 80.0 percent 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 11
A: Dissolve about 50 mg in 10 mL of dehydrated alcohol, add, with swirling, 2 mL of nitric acid, and heat at about 75 for 15 minutes: 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 preparation corresponds to that of the Standard preparation, both relative to that of the internal standard, as obtained in the Assay.
Acidity Dissolve 1.0 g in 25 mL of a mixture of equal volumes of alcohol and ether (which has been neutralized to phenolphthalein with 0.1 N sodium hydroxide), add 0.5 mL of phenolphthalein TS, and titrate with 0.10 N sodium hydroxide until the solution remains faintly pink after being shaken for 30 seconds: not more than 1.0 mL of 0.10 N sodium hydroxide is required.
Internal standard solution Transfer about 600 mg of hexadecyl hexadecanoate, accurately weighed, to a 200-mL volumetric flask, dissolve in a diluting solution containing 2 volumes of pyridine and 1 volume of propionic anhydride, dilute with the diluting solution to volume, and mix.
Standard preparations [noteUse low-actinic glassware. ] Transfer 12-, 25-, 37-, and 50-mg portions of USP Alpha Tocopherol RS, accurately weighed, to separate 50-mL conical flasks having 19/38 standard-taper ground-glass necks. Pipet 25 mL of the Internal standard solution into each flask, mix, and reflux for 10 minutes under water-cooled condensers.
Assay preparation [noteUse low-actinic glassware. ] Transfer about 60 mg of Tocopherols Excipient, accurately weighed, to a 50-mL conical flask similar to the flasks used in preparing the Standard preparations, add 10.0 mL of Internal standard solution, mix, and reflux for 10 minutes under a water-cooled condenser.
Chromatographic system (see Chromatography 621)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 G2 on 80- to 100-mesh support S1AB utilizing either a glass-lined sample introduction system or on-column injection. The column is maintained isothermally at a temperature between 245 and 265, and the injection port and detector block temperatures are maintained at about 10 higher than the column temperature. The flow rate of dry carrier gas is adjusted to obtain a hexadecyl hexadecanoate peak 30 to 32 minutes after sample introduction. [noteCure 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-µL portions 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 preparation taken by the formula:
(AS / AD)(CD / CS)in which CD and CS are 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 preparation to ensure that the factor, F, is constant within a range of 2.0%. Prepare a relative response factor curve by plotting F versus the alpha tocopheryl propionate peak area.
Procedure Inject a suitable portion (2 µL to 5 µL) of the Assay preparation into the chromatograph, and record the chromatogram. Measure the areas under the 4 major peaks occurring at relative retention times of approximately 0.50, 0.63, 0.76, and 1.00, and record the values as a, a, a, 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)(a / aD);
beta plus gamma tocopherols = (10 CD / F)(a / aD);
alpha tocopherol = (10CD / F)(a / aD)in which F is 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. [noteThe 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. ]
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