LA CREATIVIDAD EN CLASE.

8.3.1 Rihaakuru: manufacturing process, physiochemical analysis, nutritional composition and bacterial microflora

This study may help regulatory authorities in the Maldives create guidelines that will provide a more shelf-stable Rihaakuru. This could include standardizing the product and the manufacture of the product to inhibit bacterial growth.

It would be interesting to know the bacterial flora of the Rihaakuru raw material, which could be identified by extracting the DNA from the product and using cloning and PCR methods. By knowing the bacterial species, the bacteria responsible for histamine production can be identified.

8.3.2 Biogenic amines in Rihaakuru

It is important to regulate the maximum allowable level of the histamine in the Rihaakuru. It is suggested that regulatory authorities use these research findings as scientific evidence on the importance of setting a maximum allowable limit of histamine. This limit must be no more than 500 ppm although a limit of 100 ppm would be preferable as the product is consumed daily by children. Since a few of the products surveyed in this study did not contain any detectable histamine, Rihaakuru can be manufactured within such limits.

8.3.3 Long-term histamine stability in Rihaakuru

It is suggested that further studies on histamine stability in Rihaakuru be carried out and that the mechanism for the reduction in histamine observed in stored samples be determined. It will be useful for the regulatory authorities to know that there will be no increase histamine in Rihaakuru that is normally stored at 30°C.

8.3.4 Histamine degradation by bacteria and DAO

It is recommended that more bacterial isolates from Rihaakuru or the soup used for Rihaakuru manufacture be screened for histamine degradation. Histamine reduction using a bacterial inoculum would be cheaper than using DAO, although the bacteria may affect the sensory characteristics of the product.

8.3.5 Histamine degradation using DAO in a model system

It is important to determine DAO activity at histamine concentrations above 500 ppm and to identify the maximum concentration of histamine that can be inactivated by DAO. Schwelberger and Bodner (1997) observed that at concentrations greater than 0.5 mM of histamine, the degradation by DAO was inhibited. According to Mondovi et al. (1964) the optimum oxidation of histamine occurred at 1mM concentrations above which inhibition was observed though at 16.6 mM DAO was still slightly active. Guerrieri et al. (1976) have demonstrated 3mM histamine oxidation by DAO at pH 6.3.

In the current study 4.5 mM (500 ppm) histamine was successfully degraded by DAO. Histamine degradation by DAO with more than 4.5 mM histamine therefore needs to be investigated before application of this model.

Some early work to optimise histamine concentrations for DAO activity done in the early 90’s should be re-visited using the more recent sensitive methods to detect histamine.

8.3.6 Histamine degradation by DAO in tuna soup

Exponential decline and RSM models are suitable to use to predict histamine degradation in tuna soup by DAO within pH 6-7 and 1-3% salt. Manufacturers could use this as a tool to remove histamine in Rihaakuru to ensure the safety of the product. This would involve using the two model equations to obtain the desired predicted optimum rate and amount of histamine degradation by the enzyme.

Determining the sensory characteristics of Rihaakuru produced using DAO at a pH variation from 6 to 7 and salt concentration from 1 to 3% is required before the model is implemented. If the sensory characteristics did not change or improved then the present model could be applied between pH 6-7 and salt 1-3%.

Once the histamine is degraded from the soup, the soup has to be further processed into Rihaakuru without delay; if delays occur bacteria in the soup may produce more histamine or spoil the product. The effect of this DAO treatment of the soup may have some effect on the taste and functionality of the final Rihaakuru product particularly of leaving the soup at 37°C for 10 h.

To determine if the DAO technique for histamine control can be used without affecting the quality of the Rihaakuru, tuna soup treated with DAO under the conditions described will need to be taken through the final manufacturing steps to produce Rihaakuru.

Once the histamine in tuna soup is degraded by the DAO enzyme, a system to remove this enzyme from the soup may be needed if the enzyme imparts any undesirable sensory attributes to the final product.

The activity of DAO could be trialled in tuna soup at different water activity levels to determine the feasibility of treating tuna soup later in the manufacturing process. This, if effective, may reduce the risk of microbial growth in the soup during DAO treatment. The DAO enzyme is expensive (NZ $ 800/5g) therefore, if this is to be used as a practical control measure, a cheaper source of the enzyme will be needed. Alternatively a culture of the enzyme producing bacteria may be used, but this may affect the sensory characteristics of the product.

The first major impact of this research is likely to be regulatory authorities generating guidelines for the manufacture of Rihaakuru which ultimately will produce Rihaakuru containing lower histamine contents. Prior to this work, the guidelines were difficult to make due to popularity of the product and without scientific evidence to support the enforcement of guidelines.

Secondly, as consumers become more aware of the hazards they will demand safer production of the product which will pressure the manufactures to produce Rihaakuru in hygienic condition. Therefore due to this research, a safer Rihaakuru may be produced in the future. In cases where it is not possible to control temperature abuse of the fish (as low quality rejected fish are used in order not to throw away fish), this work provides an option for the manufactures of using the DAO technique to bring down the histamine to a safe level.

In summary, this work has described a unique fish paste, Rihaakuru from the Maldives. The nutritional composition of the product is now known and the food safety risk from histamine contamination has been confirmed. The traditional control of histamine is refrigeration to prevent its formation but this work provides an alternative control method which is to use DAO to degrade pre-formed histamine in the Rihaakuru under specific conditions.

8.4 References

Bardsley, W. G. (1973). Kinetics of the diamine oxidase reaction. Biochemical Journal, 131(3), 459-469.

Buffoni, F. (1966). Histaminase and related amine oxidases. Pharmacological Reviews, 18(4), 1163-1199.

Guerrieri, P., Finazzi-Agrò, A., Costa, M. T., Rotilio, G., & Mondovì, B. (1976). Salt effect on diamine oxidase activity. Italian Journal of Biochemistry, 25(2), 160-166.

Mondovì, B., Rotilio, G., Costa, M. T., Finazzi-Agrò, A., Chiancone, E., Hansen, R. E., & Beinert, H. (1967). Diamine oxidase from pig kidney. Journal of Biological Chemistry, 242(6), 1160-1167.

Mondovi, B., Rotilio, G., Finazzi, A., & Scioscia-Santoro, A. (1964). Purification of pig-kidney diamine oxidase and its identity with histaminase. Biochemical Journal, 91(2), 408-415.

Saeed, M. A. (2011). Rihaakuruge hageegai science verin furathama fahara hoadhaifi. Haveeru. Male’, Maldives: Haveeru Daily.

Schwelberger, H. G., & Bodner, E. (1997). Purification and characterization of diamine oxidase from porcine kidney and intestine. Biochimica et Biophysica Acta (BBA)/Protein Structure and Molecular Enzymology, 1340(1), 152-164.