Attempts were made to improve the method for measuring K value, a fish freshness index which is defined as the ratio of the sum of inosine (H
XR) and hypoxanthine (H
X) to that ATP and its autodegradation products in
postmortem fish muscle.
Ion exchange resin (Bio-Rad, AG 1-X4, 100-200 mesh, Cl
- type) was washed with acetone and a ctivated with 1 N HCl. A column (0.6×5 cm) of the resin was washed with 10 ml of solution A (1.9 g sodium tetraborate, 1.2 g NaCl, 22.5 ml 1 N HCl, 10 ml 1 M ethylenediamine per 1 l, adjusted to pH 7.5).
The muscle was minced, extracted with cold perchloric acid neutralized to pH 6.5-6.8 with caustic potassium solution. The column was charged with the extract equivalent to 0.1-0.2 g muscle, and washed with deionized water. HxR and Hx were eluted with solution A (elute A) while IMP, AMP, ADP and ATP were eluted with solution B (17.4 g NaCl and 150 ml 1 N HCl per 1 l) (elute B).
Elutes A and B were made up to 50 ml respectively and measured for absorbance at 250 nm to calculate K value by the following formula: K(%)=E
250nmA/E
250nmA+E
250nmB×1/2×100
This simple method was satisfactorily applied to squid muscle as judged by the standard deviation of K value, 0.12-0.16.
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