Improvement and validation of the method to determine neutral detergent fiber in feed.

To improve the performance of the analytical method for neutral detergent fiber in feed with heat-stable α-amylase treatment (aNDFom), the process of adding heat-stable α-amylase, as well as other analytical conditions, were examined. In this new process, the starch in the samples was removed by adding amylase to neutral detergent (ND) solution twice, just after the start of heating and immediately after refluxing. We also examined the effects of the use of sodium sulfite, and drying and ashing conditions for aNDFom analysis by this modified amylase addition method. A collaborative study to validate this new method was carried out with 15 laboratories. These laboratories analyzed two samples, alfalfa pellet and dairy mixed feed, with blind duplicates. Ten laboratories used a conventional apparatus and five used a Fibertec(®) type apparatus. There were no significant differences in aNDFom values between these two refluxing apparatuses. The aNDFom values in alfalfa pellet and dairy mixed feed were 388 g/kg and 145 g/kg, the coefficients of variation for the repeatability and reproducibility (CV(r) and CV(R) ) were 1.3% and 2.9%, and the HorRat values were 0.8 and 1.1, respectively. This new method was validated with 5.8% uncertainty (k = 2) from the collaborative study.