Microscopy is a powerful research tool in food science, although a number of difficulties in sample preparation may discourage its use. Investigation at structure and ultrastructure level helps to understand changes and interactions the raw material components undergo when processed into food. We have adopted various microscopy techniques to study the nature and origin of different types of crystals and spots originating in hard cheeses during ripening. Although not directly affecting the flavor, in this type of cheese they are considered a desired attribute. Compositional, biochemical and microbiological data were obtained on the same samples to support the microscopy study.
In hard cheeses upon ripening, protein is progressively degraded into free amino acids. After 10-12 month ripening, free amino acids represent more than 20% of the cheese protein. This fact largely contributes to increase the concentration of solutes in cheese water phase, where sodium chloride, calcium, phosphates, lactate and other soluble molecules are already present. Crystals of tyrosine, calcium lactate and calcium phosphate are already reported to occur in some cheese varieties, such as Cheddar, Gouda, Emmental, Grana Padano and Parmigiano-Reggiano, due to the decreased solubility as the cheese water content decreases. In addition, non-crystalline spherical spots are reported to occur in the last two cheeses, sometimes named as \u201cpearls\u201d and whose origin is not yet understood. We have focused our attention on those pearls and investigated their structure and ultrastructure for the first time. The matrix, as observed by both optical and confocal microscopy after suitable staining procedures, appeared to be rather homogeneous but more compact with respect to the surrounding cheese portion from which the pearl is clearly distinguishable, with several crystals embedded. By TEM of the resin embedded material, the crystals showed a star-shaped core surrounded by a thick layer of dense material. The nature of the different components of the crystals was further investigated by confocal microscopy, confocal Raman microscopy and compositional data, and a possible role of some free amino acids as seeding components was hypothesized
