ABSTRACT
The determination of the heme and non-heme iron fractions in raw and cooked beef steak by using spectrophotometric methods and high-performance liquid chromatography coupled to a double-focusing sector field inductively coupled plasma mass spectrometer (HPLC-SF-ICPMS) is reported. Size exclusion chromatography coupled to SF-ICPMS was used to measure the iron-containing biomolecules in the samples. This approach allowed for the direct on-line detection of the most abundant iron isotope 56Fe without interference from 40Ar16O. The HPLC-ICPMS results for the iron speciation analysis of a raw beef steak, used as an analytical quality control (AQC) sample, showed that the main iron biomolecule present was the heme iron-containing protein myoglobin. For the AQC sample, the agreement among the HPLC-ICPMS method, the non-heme iron spectrophotometric method, and the total iron concentration showed 100% recovery of iron. The sum of the different iron-containing compounds determined using the developed HPLC-ICPMS method accounted for all the iron-containing compounds extracted. The analysis of myoglobin in steak by HPLC-ICPMS showed that on cooking the concentration was reduced by 85%. However, a spectrophotometric method specific for heme iron showed that it was still intact, even after heating to 80 degrees C. The measurement of the total iron in the cooked steak and the HPLC extracts by inductively coupled plasma optical emission spectroscopy (ICP-OES) indicated that the extraction method for the HPLC analysis was no longer applicable and that loss of the heme group from the protein rendered it incompatible with the size exclusion separation. The detection limit (concentration equivalent to 3 times the baseline for a blank injection) of the HPLC-ICPMS method was 2.4 ng as iron. The results demonstrate that a combination of analytical methods can be used to provide valuable information about dietary levels of nutritionally important metal-containing compounds as well as the efficiency of established extraction methods for raw and cooked meat samples.
