manual of methods for determining micronutrients in fortified foods

7. Transfer both supernatants to a 100 mL-volumetric flask. The supernatant must be clear. Make up to
volume (100 mL) with deionized water.
8. Pipet 10.0 mL aliquots of sample solutions and standard solutions into different 25 mL volumetric
flasks.
9. Add 5.0mL acetate buffer and 4.0mL of 1,10-phenanthroline to each flask. Mixwell and color will
start developing.
10. Let stand it for 30 min and then make up to volume (25 mL) using deionized water.
11. Turn on the spectrophotometer and warm it up for 15-20 minutes prior reading the absorbance.
12. Adjust the wavelength to 510 nm. Set the mode to Absorbance.
13. Set the instrument to zero absorbance using deionized water.
14. Read the absorbance of the 0 mg/L standard solution (blank) and record the absorbance.
15. Read the absorbance for the standard solutions and flour sample solutions. The solutions will get
a red-orange color. The more intense of the color, the higher the concentration of iron in the sample.
16. If color intensity of the samples is too high, make appropriate dilution of the sample solutions and
record the absorbance again.
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