The following parameters were determined on each milk sample: lactose and fat by means of the infrared analysis with Milko-Scan (Foss Electric, Hillerød, Denmark), pH by a potentiometer with a specific electrode (Crison Instruments, Barcelona, Spain), and somatic cells and total bacterial count by fluoro-opto-electronic and flow cytometry methods with Fossomatic 250 and BactoScan FC, respectively (Foss Electric, Hillerød, Denmark). Dry matter and ash were determined by drying at 102 °C for 7 hours and calcination in the muffle at 530 °C, respectively, and from ash solubilized in HCl (2N). The total contents of Ca, Mg, Na, K, Zn, Fe and Cu were assessed by Atomic Absorption Spectrometry (AAS) and phosphorus (P) was assessed with the colorimetric method, according to Malacarne et al. [12 (link)]. Total nitrogen (TN) on whole milk, non-casein nitrogen (NCN) on milk acid whey at pH 4.6, and non-protein nitrogen (NPN) on trichloroacetic acid (TCA) filtrated and obtained from milk after treatment with TCA (120 g/L) were determined with the Kjeldahl method. Proteose-peptone N (PPN) was measured on acid whey obtained, according to van Boeckel and Crijns [13 ]. From these nitrogen fractions, the following parameters were calculated: casein nitrogen (CN = TN – NCN), true protein nitrogen (TPN = TN−NPN), crude protein (TN × 6.38), casein (CN × 6.38), true protein (TPN × 6.38), and proteose-peptones (PPN × 6.38). The Kjeldahl method was performed using a DK6 Digestor and UDK126A Distiller (VELP Scientifica, Usmate, Italy), according to the Association of Official Analytical Chemists (AOAC) standards [14 ,15 ,16 ].
Chloride (Cl−) was assessed by titration with silver nitrate (AgNO3) using the volumetric method of Charpentier-Volhard.
All glasses and polyethylene tubes used for collection, storage, and analysis of samples were previously washed with 2N hydrochloric acid (Carlo Erba reagent, Milano, Italy) solution. All solutions were prepared using two-time distilled water (conductivity < 4.3 µS/cm).
Chloride (Cl−) was assessed by titration with silver nitrate (AgNO3) using the volumetric method of Charpentier-Volhard.
All glasses and polyethylene tubes used for collection, storage, and analysis of samples were previously washed with 2N hydrochloric acid (Carlo Erba reagent, Milano, Italy) solution. All solutions were prepared using two-time distilled water (conductivity < 4.3 µS/cm).
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