Noncasein nitrogen analysis of ultrafiltration and microfiltration retentate

Previous research has suggested that the standard noncasein nitrogen (NCN) measurement method for milk overestimates the NCN content of microfiltration (MF) retentate. The objective of this study was to develop a modified method to more accurately measure the NCN content of ultrafiltration and MF retentate products. The standard method is based on precipitation of casein micelles at their isoelectric point (4.6) with acetic acid. In the standard method, a 10-mL milk sample and 75 mL of 38°C water are placed in a 100-mL volumetric flask. One milliliter of 10% acetic acid solution is added and the flask is incubated at 38°C for 10 min. Subsequently, 1 mL of 1N sodium acetate solution is added and mixed. After cooling the contents to 20°C, the flask is made up to 100mL with water, mixed, and then filtered (Whatman No. 1 filter paper). The N content of the filtrate is then determined by Kjeldahl analysis and referred to as NCN. A method was developed that used a 50-mL centrifugal tube instead of a volumetric flask. This modification facilitated measurement of the pH after addition of acetic acid. Subsequently, the sample was centrifuged (800×g at 25°C) for 10 min to facilitate filtration with a smaller pore size filter paper (Whatman no. 6). In this study, we evaluated the effect of pH after addition of 1% acetic acid and pH of the final filtrate on NCN analysis. Four pH levels after acetic acid addition (4.0, 4.2, 4.4, and 4.6) and 2 pH levels after sodium acetate addition (4.6 and 4.8) were evaluated. As the pH after acetic acid addition was increased from 4.0 to 4.6, the NCN content significantly decreased. Sodium dodecyl sulfate PAGE results also indicated that the casein fractions present in the filtrate were significantly decreased when the pH was increased from 4.0 to 4.6. The NCN content slightly decreased but the difference was not significant when the final pH of the filtrate was increased from 4.6 to 4.8. Subsequently, the NCN contents of several ultrafiltration and MF samples were determined using the standard method and modified method. The modified method gave significantly lower NCN values for most samples as compared with the standard method.