Digestibility - Department of Animal Sciences
Digestibility - Department of Animal Sciences
Digestibility - Department of Animal Sciences
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DIGESTIBILITY
Apparent v. true digestibility<br />
True digestibility involves correction for endogenous losses,<br />
apparent digestion does not.<br />
Endogenous losses<br />
– Include:<br />
• Sloughed <strong>of</strong>f intestinal cells<br />
• Digestive juices (enzymes)<br />
• Microbial matter<br />
– Quantified by measuring fecal output <strong>of</strong> fasted animals<br />
– Can be 9.8 to 12.9 % DMI<br />
– Should they be quantified?
In vivo digestibility methods<br />
Direct or total/complete collection<br />
Difference method<br />
Regression method<br />
Indirect method
1. Total collection
In vivo digestibility trials in<br />
metabolism crates
In vivo digestibility trials in pens
Total collection<br />
calculations<br />
<strong>Digestibility</strong> (g/kg) =<br />
Nutrient in feed - Nutrient in feces x 1000<br />
Nutrient in feed<br />
Dry matter digestibility (DMD, g/kg) =<br />
DM in feed - DM in feces x 1000<br />
DM in feed<br />
Organic matter digestibility (OMD, g/kg) =<br />
OM in feed - OM in feces x 1000<br />
OM in feed<br />
Can be expressed as a proportion, % or g/kg
<strong>Digestibility</strong> indices that estimate<br />
energy value<br />
Digestible organic matter content (DOMD) (g/kg DM)<br />
= OM in feed - OM in feces x 1000<br />
DM in feed<br />
TDN = DCP + DCF + DNFE + DEE(2.25)<br />
– DCP= Digestible Crude Protein<br />
– DCF= Digestible Crude Fiber<br />
– DNFE= Digestible Nitrogen-Free Extract<br />
– DEE= Digestible Ether Extract (2.25)
2. Difference method<br />
Allows digy calculation for 2 feeds fed simultaneously<br />
Assumptions<br />
– No interaction b/w the digy <strong>of</strong> the feeds<br />
– Must know digy & fecal DM output (DMO) <strong>of</strong> base<br />
feed<br />
Test feed DMD =<br />
Cons<br />
Test feed DMI – (Fecal DMO- Base feed DMO)<br />
Test feed DMI<br />
– Assumptions may be invalid
3. Regression method<br />
Schneider & Flatt (1975)<br />
Also allows digy. estimation for two feeds<br />
– Feed different ratios <strong>of</strong> the two feeds<br />
– Estimate digy <strong>of</strong> each <strong>of</strong> the ratios<br />
– Fit regression <strong>of</strong> test feed inclusion vs. digy<br />
– Extrapolate to estimate digy <strong>of</strong> test feed.<br />
Cons<br />
– Considerable expense and labor for estimating digy<br />
<strong>of</strong> one feed.
DMD (g/kg)<br />
Regression method<br />
800<br />
Base feed digy.<br />
600<br />
400<br />
Test feed digy.<br />
200<br />
20 40 60 80 100<br />
% inclusion <strong>of</strong> test feed in ration
Digy trial issues<br />
Changeover designs<br />
– necessary if period effects are an issue e.g.<br />
• <strong>Animal</strong> physiological changes<br />
• Forage physiological changes<br />
Adaptation period<br />
– Necessary to adapt the animals to<br />
• New feed (microbial population changes)<br />
• Strange equipment<br />
• Strange housing<br />
– 6 – 14 day period is the norm
Marker digestibility trials<br />
Particularly useful for grazing animals<br />
Procedure<br />
– Add indigestible marker to feed eg chromic oxide<br />
– Measure concentration in feed & feces<br />
– Estimate disappearance <strong>of</strong> marker from gut.<br />
E.g. if a feed contains 1% Cr 2 O 3 & feces contains 2%<br />
Cr 2 O3, diet digestibility = 50%<br />
– Since Cr 3 O 2 conc. has doubled, 50% <strong>of</strong> DM must have<br />
been digested
Marker trials contd.<br />
For the digy <strong>of</strong> a specific nutrient,<br />
must also know the % nutrient in feed & feces<br />
%Nutrient = 100 – 100 x % indicator feed X % nutrient feces<br />
<strong>Digestibility</strong> % indicator feces % nutrient feed<br />
Homework:<br />
If lambs are fed a bahia grass diet containing 7%<br />
protein & 1% chromic oxide, and their feces contains<br />
5% CP and 2% chromic oxide. Calculate CP digy.
Marker digestibility<br />
Pros<br />
– Total feces collection not necessary<br />
– Total intake determination not necessary<br />
– Easier, less labor<br />
Cons<br />
– Representative sampling essential<br />
– Accurate estimation <strong>of</strong> nutrient or marker conc.<br />
essential<br />
– Assumes complete excretion <strong>of</strong> marker hence<br />
Recovery <strong>of</strong> marker determines accuracy <strong>of</strong> digy
Marker types<br />
External<br />
– Chromic oxide<br />
– Dysporium<br />
– Polyamide<br />
Can contaminate<br />
forage<br />
Internal<br />
– Lignin<br />
– AIA<br />
– ADF<br />
– n-alkanes<br />
Easier, less labor
Marker issues<br />
Difficulty <strong>of</strong> mixing marker with forages<br />
– Dose cows instead- ( s handling)<br />
Marker migration<br />
– Must not affect feed digy<br />
External markers may contaminate forage
Problems with in vivo<br />
experiments<br />
<strong>Animal</strong> trials are:<br />
– Expensive<br />
– Protracted<br />
– Laborious<br />
– Public concerns<br />
– <strong>Animal</strong> stress ???<br />
Must estimate nutritive value with less animal<br />
dependent techniques
Ideal in vitro methods should be:<br />
– Rapid (one step) & routinely practicable<br />
– Accurate<br />
– Cheap & not laborious<br />
– Repeatable & robust<br />
– Biologically meaningful<br />
– Broad-based (apply to all forage types)<br />
– Handle large nos. <strong>of</strong> samples<br />
– Laboratory-based
Rumen fluid –pepsin in vitro<br />
digestibility (IVOMD)<br />
•Developed by Tilley & Terry<br />
(1967)<br />
•Measures apparent digy in rumen<br />
fluid (48 h) and acid pepsin (48 h)<br />
•Gives accurate predictions <strong>of</strong> in<br />
vivo digy for most forages
Prediction <strong>of</strong> silage OMD in vivo from<br />
different methods (g/kg DM)<br />
Method r 2 RSD<br />
KMnO 4 lignin 21.8 54.6<br />
ADF 32.1 50.9<br />
NDF 45.7 45.5<br />
(M) ADF 55.8 40.9<br />
IVOMD 74.1 33.6<br />
(Givens et al., 1989)
Rumen fluid problems<br />
Variation in Inoculum composition & activity due to<br />
– Host animal diet<br />
– <strong>Animal</strong> species<br />
– Collection time<br />
– Processing (blending vs. filtration)
Rumen fluid problems<br />
Analytical issues<br />
– Maintenance <strong>of</strong> anaerobic media; optimal pH, temp<br />
– High viscosity hinders filtration<br />
– Offensive odors<br />
– Hygiene – (Prevent pathogen infection)
In vivo DOMD<br />
Relationship between in vivo and<br />
in vitro DOMD <strong>of</strong> wheat silage (g/kg DM)<br />
690<br />
670<br />
650<br />
Year One<br />
Year Two<br />
630<br />
r 2 =0.24<br />
610<br />
590<br />
570<br />
550<br />
530<br />
530 580 630 680<br />
Rumen fluid-pepsin DOMD<br />
(Adesogan et al. 1998)
Rumen fluid technique -<br />
problems<br />
Standards needed to correct for variability in rumen<br />
fluid composition & activity<br />
Disregards / inappropriately represents:<br />
– Ruminal outflow (uses a batch process)<br />
– Digests maillard product not digested in vivo<br />
– Associative effects between feeds<br />
– Endogenous secretions<br />
– Post abomasal digestion
Alternatives to Tilley & Terry<br />
1. Rumen fluid – Neutral detergent (Van Soest, 1967)<br />
– More akin to true digestibility<br />
– Gives higher digy. values<br />
– Still requires rumen fluid<br />
2. Feces<br />
– Gives lower digestibility estimates<br />
3. Enzyme- based assays
Prediction <strong>of</strong> DMD in vivo from in vitro<br />
fecal liquor DMD<br />
Spp. <strong>of</strong> feces donor<br />
r 2 range<br />
Ovine 0.33 – 0.98<br />
Bovine 0.77 – 0.97<br />
Equine 0.90<br />
Caprine 0.96-0.97<br />
(Ohmed et al., 2001)
Cell-free enzyme in vitro digestibility<br />
Examples <strong>of</strong> procedures used:<br />
1. Cellulase<br />
2. Neutral detergent- cellulase<br />
3. Neutral detergent-cellulase +gammanase<br />
4. Pepsin cellulase<br />
Amylase pre-treatment important for starch-rich feeds<br />
Gammanase for oil-rich feeds
Relationships between DMD in vivo and<br />
enzyme predicted DMD<br />
Method R 2<br />
Cellulase 0.83<br />
Neutral detergent cellulase 0.94<br />
Acid pepsin – cellulase 0.88<br />
Rumen fluid 0.83<br />
(Bughara & Sleper, 1986)
Prediction <strong>of</strong> in vivo OMD <strong>of</strong><br />
forages from different methods<br />
Method r RSD (%) AE(+)<br />
ND + cellulase 0.90 3.3 0.9<br />
Pepsin + cellulase 0.94 2.6 0.3<br />
(McLeod & Minson, 1982)<br />
Higher analytical error with ND – cellulase technique<br />
may outweigh shorter processing time
Prediction <strong>of</strong> in vivo OMD <strong>of</strong> spring<br />
grass from different methods<br />
Method r 2 RSD<br />
ND + cellulase 76.6 27.1<br />
Pepsin + cellulase 75.9 28.8<br />
Rumen fluid-pepsin 67.0 33.2<br />
(M) ADF 66.9 33.3<br />
Poorer relationships found for autumn grass (r 2 = 13- 20)<br />
(Givens et al., 1990)
Effect <strong>of</strong> enzyme source on cellulase<br />
activity<br />
% DM solubilized<br />
Fungi Herbage Cellulose paper<br />
Trichoderma spp. 57 69<br />
Basidiomycete 48 20<br />
Aspergillus niger 45 10<br />
Rhizopus spp. 35 7<br />
(Jones & Hayward, 1975)
14<br />
C-Casein hydrolysis (mg/ml)<br />
0.5 Co-culture<br />
0.0 0.25<br />
S. bovis<br />
0.0<br />
S. ruminantium<br />
10 20<br />
Time (h)<br />
Commercial enzymes don’t fully simulate microbial<br />
activity <strong>of</strong> mixed rumen microbes
Enzyme method problems<br />
Equations are species-specific<br />
Represent effect <strong>of</strong> a few enzymes<br />
Variability in enzyme activity<br />
– Due to enzyme source & batch
The ANKOM equipment
tube<br />
tube<br />
Ankom digestibility validation<br />
Prediction <strong>of</strong> tube app. DOMD from bag app. DOMD<br />
Prediction <strong>of</strong> tube true DOMD from bag true DOMD<br />
80<br />
y = 0.87x + 4.25<br />
r 2 = 0.83; rsd = 4.04<br />
80<br />
y = 0.99x + 3.61<br />
70<br />
r 2 = 0.93; rsd=2.93<br />
70<br />
60<br />
50<br />
60<br />
40<br />
bag<br />
40 50 60 70 80<br />
50<br />
bag<br />
50 55 60 65 70 75 80 85
ANKOM pros & cons<br />
Pros<br />
– Simplifies filtration, incubation and mixing<br />
– Uses a batch process (& ash-free bags)<br />
Cons<br />
– Bag pore size may allow excess outflow or restrict<br />
microbial colonization<br />
– Bag material & pore size may affect results<br />
• Mon<strong>of</strong>ilamentous cloth – precise aperture<br />
• Multifilamentous cloth – pore size affected by stresses<br />
e.g. dacron
In vitro digestibility summary<br />
Pros<br />
– Predicts in vivo digy more accurately than NDF or<br />
lignin<br />
– Handles several samples & are biologically<br />
meaningful<br />
Cons<br />
– May require fistulated animals<br />
– Labor intensive & protracted<br />
– Plagued by variability in composition & activity <strong>of</strong><br />
inoculum/enzyme<br />
– Doesn’t indicate the kinetics <strong>of</strong> digestion
<strong>Digestibility</strong> references<br />
Chapters 6 – 8 In: D.I. Givens, E. Owen, R.F.E. Axford and H.M. Omed (Editors) 2000,<br />
Forage Evaluation in Ruminant Nutrition. CABI Publishing, Wallingford, UK, pp. 113-<br />
134.<br />
Adesogan, A.T, Givens D.I. and Owen. E. Measuring chemical composition and nutritive<br />
value in forages. Field and Laboratory methods for grassland and animal production<br />
research. CABI Publishing. P 263<br />
Tilley, J.M.A. and Terry, R.A., 1963. A two stage technique for the in vitro digestion <strong>of</strong><br />
forage crops. Journal <strong>of</strong> the British Grassland Society, 18: 104-111.<br />
Van Soest, P.J., Wine, R.H. and Moore, L.A., 1966. Estimation <strong>of</strong> the true digestibility <strong>of</strong><br />
forages by the in vitro digestion <strong>of</strong> cell walls. Proceedings <strong>of</strong> , The Xth International<br />
Grassland Congress, Helsinki. Finish Grassland Association., pp 438-441.<br />
Vogel, K.P., Pedersen, J.F., Masterson, S.D. and Toy, J.J., 1999. Evaluation <strong>of</strong> a filter bag<br />
system for NDF, ADF, and IVDMD forage analysis. Crop Science, 39: 276-279.<br />
Wilman, D. and Adesogan, A., 2000. A comparison <strong>of</strong> filter bag methods with conventional<br />
tube methods <strong>of</strong> determining the in vitro digestibility <strong>of</strong> forages. <strong>Animal</strong> Feed Science and<br />
Technology, 84: 33-47.