engleski

Separation of Honey Colour-Contributing Molecules by Gel Filtration on Sephadex G-25

Honey colour is unique property of each honey, attributed to the presence of various endogenous molecules including melanoidins, Maillard products, proteins, polyphenols, flavonoids, furosine, free amino acids, minerals, vitamins, and sugars. However, which of aforementioned class of molecules plays major role in honey colour, and to what extent it contributes to colour, is still unknown. In this respect, we have separated 14 different honey samples (3 of sage, 3 of lime, 3 of black locust, 3 of honeydew honey, and 2 of chestnut honey) using gel filtration on Sephadex G-25. Honey samples dissolved in 100 mM phosphate buffer (pH = 6.1) at ratio 1.5 g per 3 mL of buffer, were separated on Sephadex G-25 column. Collected fractions (20 fractions, each containing 1 mL of eluate) were analysed for colour by spectrophotometric method, followed by protein, total polyphenol and reactive amino group content determination. Elution profiles of colour of examined honeys revealed that colour-contributing molecules were spread between third up to 15th fraction, with dominant sharp peak centred at fifth fraction, and additional wide spread peak of quite lower intensity between seventh up to fifteen fraction. On the basis of area under the curve calculation (AUC), and data on protein, total polyphenol and reactive amino group content in fractions, it can be concluded that the great amount of honey colour-contributing molecules belongs to group of molecules with molecular mass greater than 5000 Da, what includes proteins, melanoidins and Maillard products. However, honey colour seems also significantly affected by the amount of low molecular weight compounds such as polyphenols, flavonoids, and free amino acids, while the effect of mineral and vitamins also can not be neglected. Moreover, on the basis of AUC it can be concluded that majority of colour of light honeys, such as black locust, sage and lime honeys, can be attributed to protein, melanoidin and Maillard products fraction which comprise from 50 up to 79% of total honey colour. On the contrary, in the case of dark honeys, such as honeydew and chestnut honey, colour is more dependent on low molecular weight compounds such as polyphenols, flavonoids and free amino acids, where proteins, melanoidins and Maillard products comprise from 22 up to 43% of total honey colour.

honey colour, colour contributing molecules, size exclusion chromatography

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