Rev 9/20/18 pg. 89
Appendix A – Reagents (AOAC 991.43, 985.29, 2001.03)
Use Distilled Water throughout.
Best practices for solution preparation
The best practice for preparation of solutions is to make them fresh and use them within 2 days. This is especially true of the
MES-TRIS buffer and Phosphate buffer solutions which can readily support microbial growth. Good laboratory cleaning
practices are critical to ensure against contamination and microbial growth. Failure to maintain necessary cleanliness and
prevent contamination can dramatically affect shelf life of solutions.
Where high volume usage makes daily solution preparation less practical, the following guidelines can be used.
2 days at room temperature
Store refrigerated at 4˚C
(evidence suggests refrigeration extends shelf life –
user to determine best practice)
Phosphate Buffer solution
2 days at room temperature
Store refrigerated at 4˚C
(evidence suggests refrigeration extends shelf life –
user to determine best practice)
Enzymes, diluted
(Amylase, Protease, Amyloglucosidase as
purchased from ANKOM Technology)
Insufficient data to make
recommendation
Store refrigerated at 4˚C
(not to exceed expiration date on original packaging)
Enzyme solutions for AOAC 991.43, 985.29, and 2001.03 methods
With no specific enzyme activity standards referred to in the dietary fiber methods, ANKOM has chosen to use the activity
numbers specified by Megazyme based on their extensive research. As always, refer to the specific method for Enzyme purity
information, and run specified test samples as appropriate.
(a) Enzyme solutions—To make your enzyme solutions for use in the ANKOM
TDF
Dietary Fiber Analyzer, dilute
ANKOM Concentrate enzymes (TDF80/TDF81, TDF82/TDF83, TDF84/TDF85) with H
2
O according to the Enzyme
Dilution Table below.
α-Amylase (ANKOM Concentrate)
Dilute 5 ml of amylase to 25 ml
Protease (ANKOM Concentrate)
Dilute 5 ml of protease to 25 ml
Amyloglucosidase (ANKOM Concentrate)
Dilute 5 ml of Amyloglucosidase to 25 ml
The ANKOM
TDF
Dietary Fiber Analyzer delivers 1 ml of each enzyme solution to
each of the six stations per run (6 ml of each enzyme solution). If you are NOT
using the ANKOM Concentrate enzymes, you must prepare enzyme solutions that
include 0.02% w/v of Sodium Azide (to prevent microbial growth) and the
following enzyme concentrations:
150 Ceralpha Units per ml
The 2001.03 method calls for 10% of the Protease used in the 985.29 method.
Therefore, when running the AOAC 2001.03 method in the ANKOM
TDF
Dietary
Fiber Analyzer, take the ANKOM TDF82/TDF83 Protease you diluted above and
further dilute it from 5ml to 50ml using H
2
O.
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