379 / 696
Where Abs
MS
= absorbance of the standard measuring solution
100 = dilution factor for stock solution to standard measuring solution
1000 = factor for g to mg
E
1%,1cm
= extinction coefficient (from Craft and Soares, 1992)
Lutein in MTBE: 2589
β-carotene in MTBE: 2588
W = weight of reference standard, in mg
(2)
Chromatographic Purity (CP)
Inject lutein and β-carotene working solutions at least three times. The CP is calculated as:
CP = (area of the all-trans-carotenoid peak)/(sum of areas of all relevant peaks)
Relevant peaks include all peaks in the HPLC chromatogram with the exception of solvent peaks.
(3)
Reference Standard Purity (P)
Calculate the purity of each reference standard:
P(%) = SP x CP x 100
Where 100 is the conversion of decimal to percent
(b)
Calculate the concentration of each carotenoid analyte (e.g. lutein, C
L
) in the all-
trans
form, in
μg/100 mL, in the working solution:
C
L
= W
L
x P
L
x (1000/10,000)
Where W
L
= the weight of lutein used to make the stock solution, in mg
P
L
= the reference standard purity of all-
trans
-lutein calculated in section G.d. above
1000 = the conversion of mg to μg
10,000 = the factor for the dilution of 1 mL stock in 100 mL working solution and for the
conversion of percent to decimal
(c)
Calculate the concentration of the apocarotenal internal standard (C
A
), in μg/100 mL, in the
working solution:
C
A
= W
A
x (1000/100)
Where W
A
= the weight of apocarotenal used to make the stock solution, in mg
1000 = the conversion of mg to μg
100 = the factor for the dilution of 1 mL stock in 100 mL working solution
Carot-02 (February 2016)
FOR ERP USE ONLY
DO NOT DISTRIBUTE