50.1.
09 Teflon stopcock, titrate rapidly with indophenol standard solution
AOAC Official Method 985.33 until light but distinct rose-pink persists for 5 s. (Each titration
Vitamin C (Reduced Ascorbic Acid) should require ca 15 mL and titrations should check ±0.1 mL.) Ti-
in Ready-to-Feed Milk-Based Infant Formula trate 3 blanks composed of 7 mL precipitant solution plus 15 mL
2,6-Dichloroindophenol Titrimetric Method H2O. Average blank is 0.1 mL. Calculate dye equivalents: ascorbic
First Action 1985 acid equivalent to 1 mL indophenol standard solution = mg ascorbic
Final Action 1988 acid/(mL dye − blank titration) = 2 mg/(mL dye − blank titration).
A. Principle C. Preparation of Test Solution
Ascorbic acid is estimated by titration with colored oxida- Pipet 25–30 mL test portion and equal volume of precipitant solu-
tion-reduction indicator, 2,6,dichloroindophenol. EDTA is added as tion, (a), into 125 mL beaker. Designate total volume as V mL and
chelating agent to remove Fe and Cu interferences. volume of composite aliquoted as E mL. Filter through folded rapid
B. Reagents qualitative paper, 18.5 cm (Whatman No. 541, or equivalent). Des-
(a) Precipitant solution.—Dissolve with shaking 15 g glacial ignate filtrate as assay solution.
HPO3 pellets in 40 mL glacial CH3COOH and 150 mL H2O. Dilute D. Determination
to 250 mL with H2O and filter rapidly through folded qualitative pa- Pipet three 10 mL aliquots of test solution each into separate
per (rapid, 18.5 cm, Whatman No. 541 or equivalent) into 500 mL 50 mL Erlenmeyers and titrate with indophenol standard solution.
beaker. Similarly titrate 2 blanks composed of mL precipitant solution and
Dissolve with shaking 0.9 g EDTA in 200 mL H2O and dilute to H2O equivalent to their respective volumes in test solution aliquot ti-
250 mL. Mix equal volumes of HPO3 and EDTA solutions immedi- trated. Add volume of H2O equivalent to mL indophenol standard
ately before use. solution used in titration of assay solution. Titrate with indophenol
(b) Ascorbic acid standard solution.—1 mg/mL. Accurately standard solution to same color end point observed in titration of
weigh 50 mg USP Ascorbic Acid Reference Standard (stored in des- standard aliquot.
iccator away from sunlight). Transfer to 50 mL volumetric flask. Di-
lute to volume with precipitant solution, (a). Prepare immediately Ascorbic acid/L ready-to-feed formula, mg =
before use in standardization of indophenol standard solution. (X − B) × (F/E) × (V/Y) × 1000
(c) Indophenol standard solution.—Dissolve 0.0625 g
2,6-dichloroindophenol Na salt (stored in desiccator over soda lime) where X = average mL for test solution titration; B = average mL for
in 50 mL H2O in 250 mL volumetric flask to which has been added test solution blank titration; F = mg ascorbic acid equivalent to
0.0525 g reagent grade NaHCO3. Shake vigorously; when dye dis- 1.0 mL indophenol standard solution; E = volume composite
solves, dilute to 250 mL with H2O. Filter through rapid-flow folded aliquot; V = test solution volume; Y = volume test solution titrated =
paper into amber glass bottle. Store in refrigerator. 10 mL; 1000 = conversion of mL to L.
Transfer three 2.0 mL aliquots of ascorbic acid solution, (b), into
each of three 50 mL Erlenmeyers containing 5 mL precipitant solu- Reference: JAOAC 68, 514(1985).
tion, (a). Using 25 mL buret calibrated in 0.05 mL and with glass or CAS-50-81-7 (ascorbic acid)
© 2000 AOAC INTERNATIONAL
