Factors involved in the conservation of honey enzymatic activity in frozen and dehydrated systems.

SANTAGAPITA, PATRICIO R.; GUTIERREZ, NORA E; MAZZOBRE, MARÍA F.; BUERA, MARÍA DEL P.

Lubiana

Encuentro; 3rd INTERNATIONAL FORUM ON APITHERAPY AND 2nd INTERNATIONAL FORUM ON APIQUALITY.; 2010

Many health-promoting and curative properties attributed to honey are related to the enzymatic systems that it contains. Strategies for the long-term conservation of those enzymatic systems is thus necessary when pharmaceutical or nutraceutical products based on honey bioactive components are to be developed. The stabilisation of labile biomolecules is generally performed in dehydrated or frozen media. However, enzymes are sensitive to the local environmental changes derived from the water removal, and trehalose is often employed as protein protectant. One of less stable enzymatic systems in honey is amylase. Thus, it could be considered a suitable marker of the damage occurring in proteins during dehydration or freezing, which has been poorly investigated. The objective of the present work was to analyse the enzyme stabilization in freeze-dried or dehydrated honey and its relationship to physical properties of systems containing trehalose and salts. For the cryo-concentrated systems, solutions containing a multifloral honey (from Chaco, Argentina) in the presence of 20% (w/v) of trehalose or sugar-salt mixtures (chlorides, citrates and acetates of sodium and potassium), were stored at −26° C up to 60 days. Amorphous dehydrated systems were obtained by freeze-drying the solutions employed in the cryo-concentrated systems. The freeze-dried samples were re-humidified at relative humidities from 11 to 43 % at 25°C. After storage at 55°C, enzyme activity was measured by the Tietz method. A differential scanning calorimeter was used to determine glass transition temperatures (Tg), enthalpies  and temperatures of sugar or water crystallization and melting. In frozen systems enzyme stability impaired when salts were added by the increased molecular mobility (due to the increased amount of water in the amorphous frozen matrix). In the more diluted honey systems, the higher amount of crystal surfaces had a deleterious effect on protein stability. In dehydrated systems, as trehalose crystallinity increased, the remaining enzymic activity decreased. Some salts improved the efficiency of trehalose as an enzyme protectant by delaying its crystallization, but when about 50% of trehalose crystallised, the protective effect was lost, and the enzyme quickly became inactivated in salt containing systems. In order to develop formulations for the long-term conservation of honey enzymes the presence of trehalose and of electrolytes should be considered as critical factors.