Metmyoglobin measurement

Four spin-labelled derivatives of sperm whale metmyoglobin were prepared by site-directed mutagenesis.101 Cyanide anion or imidazole was added to prepare low-spin Fe(III). Iron relaxation rates were measured by saturation recovery or inversion recovery between 5 and 17 K and by analysis of the temperature-dependent contribution to the CW line widths of the iron signal at 20 to 160 K. The nitroxyl 7i values in spin-labelled Zn-substituted myoglobin were measured to provide values in the absence of interaction with the more rapidly-relaxing Fe(III). The full shapes of the nitroxyl saturation-recovery curves for the spin-... 

There are several important points that need to be stressed. First, the estimation of discoloration in meat by measurement of percent metmyoglobin is useful in a relative sense only. Proper control samples (i.e., nontreated samples subjected to identical conditions, dilutions, and analyses, as treated samples) must be in place, and color measurement must occur in the same manner with both control and treated samples. Additionally, comparisons of values for percent metmyoglobin between different laboratories are only valid when investigators use the same measurement procedure. The use of three different formulas (i.e., from three different procedures) will yield different values for percent metmyoglobin. Again, it is the relative differences between treatments, or the changes over time, that become important, and... 

Meat discoloration studies typically involve a maximum of 5 days, with discoloration analy-sis being performed every day or on alternate days. The actual experimental time involved in the objective assessment of discoloration is not extensive and depends on the number of samples being analyzed. Colorimetric measurements with hand-held colorimeters are very rapid (three measurements per meat surface in < 1 min). Spectral scans of meat surfaces require 1 to 2 min. Extraction and analysis of ground meat products has the added step of homogenization and filtration prior to spectrophotometry, but relative to many laboratory procedures, this is relatively quick. Isolation and purification of preparative amounts of myoglobin requires only 2 to 3 days once appropriate preparations are made. Finally, metmyoglobin can be reduced to oxymyoglobin in 15 to 20 min. 

Figure 16.9 The ratio of the effective calorimetric enthalpy to the enthalpy calculated by the van t Hoff method from optical measurements, plotted against the temperature of denaturation. The symbols represent the following proteins , metmyoglobin A, ribonuclease O, cytochrome c O, a-chymotrypsin , lysozyme. Reproduced by permission from P. L. Privalov, Adv. Prot. Chem., 33, 167 (1979).

Fig. 9. Differential IR spectra of a 1% metmyoglobin-99% KBr pellet. The reference spectrum was measured with the non-irradiated pellet.

Eagerman et al. (1978) have followed the colour change with time in beef semimembranous muscle. Approximately 1 cm thick slices were plaeed against the port of a speetrocolorimeter and the reflectance spectra measured between 600 and 650 mn. The formation of a depression in the region of the speetral curve around 632 nm is an indieator of brown metmyoglobin pigment produetion. 

Figure . First derivative X-band high-spin Fe " ESR spectra from iron proteins (a) Frozen solution of metmyoglobin at 2°K and v = 8.9 GHz. Insert shows the high field peak expanded and enlarged. Reproduced with permission from G. Fehrer et al (1973). (b) Frozen solution of Pseudomonas oleovorans rubredoxin containing a single iron atom per molecule of protein. The spectrometer gains for the different traces have been adjusted arbitrarily. The effective g, alues for some of the spectral features are indicated on the spectra, and the temperature of the measurement is shown for each spectrum. Reproduced with permission from Blumberg and Peisach (1973).

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