The roles of pH extraction and colloidal protein solubility in the optimization of spectrophotometric nitrite determination in meat products via response surface methodology

The influence of four critical factors such as sample weight/borax reagent ratio (B-R factor), ascorbic acid content (A(R) factor), neutralization with HCl 1 N (N-R factor) and stirring extraction time (S-ET factor), was investigate in order to find the best conditions (optimization) to develop the official ISO 2.918 spectrophotometric method to determine the residual nitrite content in meat products, using the response surface methodology (RSM) as optimization tool. The factors most strongly affecting nitrite determination in meat products are B-R, N-R and A(R), due to their respective effects on pH extraction parameters and on the amount of colloidal protein present in the sample extract. At pH <= 6, for example, the extract - though appearing clear and transparent to the analyst contains a considerable amount of hydrolyzed protein, which will severely interfere with measurements, generating false-positive results. The colloidal protein present in the extract (<= 20 mg/g, corresponding in these working conditions to an OD340 value of <= 0.600) will lead to the recording of nitrite values greater than those actually present in the sample. In order to avoid these drawbacks, this paper proposes that the amount of borax added (B-R) varies as a function of sample weight (W-S), using the ratio W-S/B-R = 1.11. In order to monitor the analytical method, it is further recommended that pH be adjusted to 6-7 (lower protein solubility) and that colloidal protein levels be >= 20 mg/g, as confirmed by an OD340 value of >= 0.600. (C) 2008 Elsevier Ltd. All rights reserved.