SIMULTANEOUS DETERMINATION OF 13-CIS AND ALL-TRANS VITAMIN A PALMITATE (RETINYL
PALMITATE), VITAMIN A ACETATE (RETINYL ACETATE), AND TOTAL VITAMIN E (Α-TOCOPHEROL
AND DL-Α-TOCOPHEROL ACETATE) IN INFANT FORMULA AND ADULT NUTRITIONALS BY
NORMAL PHASE HPLC
CODE:
DATE:
PAGE:
VER:
Purpose:
AOAC 2012.10
Jan 2014
1 of 11
04
AOAC MLT Study
Page: 8 of 11
12.0
1.5
5
95
14.0
1.5
5
95
15.0
1.5
95
5
20.0
1.5
95
5
Note
: The gradient given can be altered as required to maximize the analytical
separation and avoid interferences. This should only be done by an authorized
analyst.
N. SYSTEM SUITABILITY
a.
The correlation coefficient of each calibration should be not less than 0.995.
b.
The resolution between
cis
and
trans
forms of Vitamin A Palmitate and 13-
cis
and
trans
forms of Vitamin A Acetate should be not less than 1.5.
Resolution of a minor cis
peak (9 or 11-cis) between the 13-cis and trans peak gives confidence that the
resolution is sufficient.
c.
When stable baseline is obtained, inject a medium reference standard three times in
succession and determine the %RSD, The %RSD should be not more than 2%.
d.
The appropriate number of samples between successive calibrations is typically 6
samples.
e.
The target limits of tailing factor for meeting the system suitability are 0.8 to 1.2.
O. CALCULATIONS
The HPLC software will automatically calculate the concentration of the sample in µg/100g for
Vitamin A and mg/100g for Vitamin E, (powder or liquid), provided the concentration of the working
standards in µg/mL (Vitamin A) or mg/mL (Vitamin E), sample weight / powder equivalent (g),
dilution (10mL) and conversion to 100g are entered correctly.
s
S
m
S
I A
w
100
10
)
(
Where:
A
S
is the peak area or height of the Vit A or Vit E in the test sample solution
I
is the intercept of the calibration curve
S
is the slope of the calibration curve
10
is the volume (mL) of isooctane used
100 is the factor to convert in 100 g basis
m
S
is the sample weight (liquid samples) / powder equivalent in grams (powder samples)
Vitamin A & E - MLT
For ERP Use Only
DO NOT DISTRIBUTE
Page 8
