Tocopherols Excipient

This article is compiled based on the United States Pharmacopeia (USP) – 2025 Edition

Issued and maintained by the United States Pharmacopeial Convention (USP)

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1 DEFINITION 

Tocopherols Excipient is a vegetable oil solution containing NLT 50.0% of total tocopherols, of which NLT 80.0% consists of varying amounts of beta, gamma, and delta tocopherols. 

2 IDENTIFICATION 

A. 

Sample solution: 50 mg of Tocopherols Excipient in 10 mL of dehydrated alcohol 

Analysis: To the Sample solution add with swirling 2 mL of nitric acid, and heat at about 75° for 15 min. 

Acceptance criteria: 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) of the Sample solution corresponds to that of the Standard solution, both relative to that of the internal standard, as obtained in the Assay. 

3 ASSAY 

Procedure 

Solution A: Pyridine and propionic anhydride (2:1) 

Internal standard solution: 3 mg/mL of hexadecyl hexadecanoate in Solution A 

Standard solutions: Using low-actinic glassware, add 12-, 25-, 37-, and 50-mg portions of USP Alpha Tocopherol RS 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, and reflux for 10 min under water-cooled condensers. 

Sample solution: Using low-actinic glassware, add 60 mg of Tocopherols Excipient to a 50-mL conical flask similar to the flasks used in preparing the Standard solutions. Add 10.0 mL of Internal standard solution, and reflux for 10 min under a water-cooled condenser. Chromatographic system 

(See Chromatography 〈621〉, System Suitability.) 

Mode: GC 

Detector: Flame ionization 

Column: 4-mm × 2-m borosilicate glass; packed with 2%–5% liquid phase G2 on 80- to 100-mesh support S1AB using either a glass-lined sample introduction system or on-column injection 

Temperatures 

Column: 245°–265°, maintained isothermally 

Injection port: 10° higher than the Column temperature 

Detector: 10° higher than the Column temperature 

Flow rate: Dry carrier gas is adjusted to obtain a hexadecyl hexadecanoate peak 30–32 min after sample introduction. [Note—Cure and condition the column as necessary.] 

Injection volume: 2–5 µL 

System suitability 

Sample: Sample solution 

[Note—The relative retention times for delta tocopheryl propionate, beta plus gamma tocopheryl propionate, and hexadecyl hexadecanoate are about 0.50, 0.63, and 1.00, respectively.] 

Suitability requirements 

Resolution: Chromatograph a sufficient number of injections to ensure that a resolution of NLT 2.5 between delta tocopheryl propionate and beta plus gamma tocopheryl propionate relative to hexadecyl hexadecanoate is met. 

Analysis 

Samples: Standard solutions and Sample solution 

Calibration: Chromatograph each Standard solution, and calculate the relative response factor, F, for each concentration of the Standard solution taken: 

F = (r_S/r_D) × (C_D/C_S) 

r_S = peak response of alpha tocopherol in the Standard solution 

r_D = peak response of hexadecyl hexadecanoate in the Standard solution 

C_D = concentration of hexadecyl hexadecanoate in the Standard solution (mg/mL) 

C_S = concentration of USP Alpha Tocopherol RS in the Standard solution (mg/mL) 

Chromatograph a sufficient number of injections of each Standard solution to ensure that F is constant within a range of 2.0%. Prepare a relative response factor curve by plotting F versus the alpha tocopheryl propionate peak response. 

Inject the Sample solution, and measure the responses for the four major peaks occurring at relative retention times of approximately 0.50, 0.63, 0.76, and 1.00, and record the values as a_δ, a_βγ, a_α, and a_D, corresponding to delta tocopheryl propionate, beta plus gamma δ

tocopheryl propionates, alpha tocopheryl propionate, and hexadecyl hexadecanoate, respectively. 

Calculate the quantity of each tocopherol form in the Tocopherols Excipient taken: 

delta tocopherol = (V × C_D/F) × (a_δ/a_D) 

beta plus gamma tocopherols = (V × C_D/F) × (a_βγ/a_D) 

alpha tocopherol = (V × C_D/F) × (a_α/a_D) 

V = volume of Internal standard solution used in the Sample solution (mL) 

F = obtained from the relative response factor curve (see Calibration) for each of the corresponding responses for the delta, beta plus gamma, and alpha tocopheryl propionate peaks produced by the Sample solution 

[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.] 

Acceptance criteria: NLT 50.0% of total tocopherols, of which NLT 80.0% consists of varying amounts of beta, gamma, and delta tocopherols 

4 SPECIFIC TESTS 

Acidity 

Solution A: Alcohol and ether (50%:50%). Neutralize to phenolphthalein with 0.1 N sodium hydroxide. 

Sample solution: Dissolve 1.0 g of Tocopherols Excipient in 25 mL of Solution A. 

Analysis: To the Sample solution 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 s. 

Acceptance criteria: NMT 1.0 mL of 0.10 N sodium hydroxide is required. 

5 ADDITIONAL REQUIREMENTS 

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/g, of total tocopherols and of the sum of beta, gamma, and delta tocopherols. 

USP Reference Standards 〈11〉 

USP Alpha Tocopherol RS