Study of melanoidins of the Maillard reaction in Dulce de Leche

The “Dulce de leche” (DL) is a dairy product elaborated by milk and sucrose concentrated in favorable conditions for the Maillard Reaction (MR). In this reaction organoleptic properties, as aroma and color are developed. It is a type of non-enzymatic browning which involves the reaction of carbonyl compounds, especially reducing sugars with compounds which possess a free amino group such as amino acids, amines and proteins.

The final stage of the chemical reaction is typified by the production of brown and nitrogen-containing polymers and co-polymers, the melanoidins. Recently research relate this products with antioxidant, antimicrobial, anti-inflammatory, antihypertensive or prebiotic activity.

In foods melanoidins are linked to proteins in a complex structure as melanoproteins which is more difficult to analyze so it is relevant the melanoidins separation and isolation from a complex matrix.

In this study, DL was elaborated in the laboratory with skim milk and sucrose, with initiated pH 7,6. Firstly, separate with dialysis and centrifugation to obtain melanoproteins. Moreover, the insoluble and colored fraction was digested by pronase enzyme. Nextly, the digested enzymatic product was fractionated by gel filtration (BioGel P2 1 a 1,8kDa). Finally, the melanoidins were separated by solid phase extraction (SPE) using a C18 cartridge with different mixtures of methanol and water. The fractions obtained were monitored by TLC and analyzed by mass spectrometry (LC-ESI-IT-MS²).

The TLC reveal with orcinol and ninhydrin, using phenylalanine, alanine and glucose as standards, indicate the presence of carbohydrates and amino acids. In addition, some compounds were fluorescent (detection at 365 nm), in agreement with reported compounds obtained in models of the MR. Mass spectrometry analysis shows compounds with typical structural motifs of melanoidins in a complex mixture with m/z less than 1200.