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9
Table 8: Preparation of calibration standards of cystine
Solution
10 pmol/µL
5 pmol/µL
2.5 pmol/µL
1 pmol/µL
0.5 pmol/µL
0 pmol/µL
Cystine solution
200
a
100
a
50
a
200
b
100
b
0
b
1% DDP in 0.2 M
NaOH
600
600
600
600
600
600
0.1 % phenol in
12 M HCl
2500
2500
2500
2500
2500
2500
10 mM Nva in
0.1 M HCl
200
200
200
200
200
200
0.2 M HCl
600
600
600
600
600
600
H2O
900
1000
1050
900
1000
1100
a
10 mM cystine stock solution in 0.05 M NaOH.
b
1 mM cystine in 0.05 M NaOH.
Close the tubes with a screw cap and place them in an oven at 110°C for 24h.
5.3
Neutralization of infant formulas and cystine standards
Transfer 0.2 mL of the hydrolyzed sample mixture or the converted cystine standards into Eppendorf
tubes, add 0.2 mL of 6 M NaOH and then 0.4 mL of 0.2 M HCl. Filter through a 0.45 µm PVDF
membrane filter into another Eppendorf tube.
5.4
Preparation of amino acid calibration standards
Table 9 provides an example of how to prepare 0.5 mL of calibration standards containing 0 to 25
pmol/µL and 10 pmol Nva/µL (final concentration after derivatization). If quantification of taurine is
not needed, replace the taurine solution by water.
AMINO-01
FOR ERP/WORKING GROUP USE ONLY
DO NOT DISTRIBUTE