Richard Lab:ADF

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Fiber analysis is an established method of determining fiber contents and digestibility of animal feeds and forages. When the steps are performed in a sequential manner, fiber analysis also provides estimates of hemicellulose and cellulose contents. The steps are: (1) neutral detergent fiber (NDF), (2) acid detergent fiber (ADF), and acid detergent lignin (ADL).


  • acid detergent solution (stored in room 113B)
  • 2000 mL flask
  • acetone (in flammables cabinet below fume hood in room 113B)
  • 4 glass jars
  • microwave
  • pH paper
  • digestion apparatus (ANKOM fiber analyzer)
  • analytical balance
  • drying oven at 105°C
  • dessicator
  • zip-loc bags with dessicant pouches (recently recharged at 105°C)
  • hazardous waste containers


  1. Close valve on ANKOM fiber analyzer.
  2. Load filter bags into bag suspender, starting with bottom tray. Place three bags per tray and stack trays on center post with each level rotated 120 degrees. Insert bag suspender into fiber analyzer.
  3. Measure out approximately 2 L acid detergent solution into flask (this may need to be done 2 x 1 L, due to foam). Pour mixture into fiber analyzer. Place weight on bag suspender, turn on agitation and check liquid level.
  4. Close lid. Heat to 100°C and agitate for 60 min.
  5. After 45 min, fill 4 glass jars (to just below neck) with nanopure water. Microwave 1st jar for 11 min.
  6. After 60 min, turn off heat and agitation. Open valve and drain solution into hazardous waste container.
  7. Open lid. Close valve. Pour heated rinse water into fiber analyzer. Agitate for 5 minutes with lid open. Microwave 2nd jar of nanopure water for 11 min.
  8. Repeat rinses until water is neutral pH. Check this using pH paper and compare to nanopure water for reference. Should require 3-4 rinses.
  9. After final rinse is complete, remove bag suspender from fiber analyzer. Gently squeeze bags (4-6 at a time) to remove excess water.
  10. Place bags in 1 L beaker. Add acetone to cover (~400 mL) and let soak in fume hood for 3-5 min. Gently squeeze bags (4-6 at a time) to remove acetone. Let air-dry on clean tray in fume hood for 1 h (until acetone has evaporated). Pour used acetone into hazardous waste container.
  11. Place tray in 105°C oven to dry overnight.
  12. Remove bags from oven and place immediately in zip-loc bags (flatten to remove air) and into dessicator. After cool (~30 min), record ADF bag weight using appropriate analytical balance. Proceed with ADL when 2 batches are ready.



  1. Van Soest, P. J., Robertson, J. B., and Lewis, B. A. (1991). "Methods for dietary fiber, neutral detergent fiber, and nonstarch polysaccharides in relation to animal nutrition." Journal of Dairy Science, 74(10), 3583-3597.
  2. See ANKOM Technology website for procedures (


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