Techniques - Undeclared Freezing (Changes in HADH Enzyme)

The method involves the determination of the enzyme HADH (ß-hydroxyacyl-CoA dehydrogenase), which is naturally present in muscle mitochondria, the energy-producing centres within cells. The method relies on the fact that when meat freezes, ice crystals form within the cells, eventually rupturing the membranes and releasing the soluble contents into the intracellular fluid. The juice pressed from meat that has been frozen and thawed will therefore exhibit higher HADH activity than meat which has not been previously frozen.

The method is a comparative one, where the HADH activity is determined on the sample as received (X₀), and then determined for a second time after a freezing and thawing cycle (X₁). The ratio of the HADH activities (X₀/X₁) is called the R₁ value. If the R₁ value is close to 1, then the meat or poultry is regarded as having been previously frozen since there is little or no change in HADH activity with subsequent freezing.

The liquid is extracted using a G-clamp press.

It is important that the clamp is at 90 degrees, as shown, so that the liquid is not reabsorbed by the meat.

The HADH activity is measured spectrometrically by determining the decrease in the absorption obtained from the following reaction, catalysed by HADH.:                                                         

 Acetoacetyl-coenzyme A +NADH+H⁺     ------>         ß-hydroxybutyryl-coenzyme A + NAD⁺_.

The method is widely used to check for prior freezing of meat and poultry sold as "fresh"

This is one of the few methods, other than detailed examination of cellular structure which requires interpretation by an expert microscopist, that can determine prior freezing.

Chill chains for modern meat supply are ofen kept only marginally above zero centrigrade, much colder than when the methods were originally developed and validated.  This can cause cellular ruptures on the surface of the meat, which may be falsely interpreted as evidence of prior freezing.

This is a screening test.  Results are not clear cut - they fall within uncertainty bands, and there may be overlap between HADH ratios for previously frozen vs not frozen meat.  Limits are usually set for what is "suspicious" e.g. a ratio of below 0.9 in the example of chicken, below, subjected to different chilling processes.

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There is little difference in R₁ values for fresh (i.e., chilled) and superchilled chicken breast. However, the original limit of 0.9 for frozen chicken is not appropriate for rapidly frozen chicken breast and is changed to 0.5.