The rumen: a feed-deconstructing reactor fueling
milk production and superintending its composition
D. P. Morgavi, M. Doreau, A. Cornu, B. Graulet
INRA, Herbivores Unit, Theix, France
9th International Symposium on Milk Genomics and Human Health
October 8-10, 2012 Wageningen, the Netherlands
Presentation plan
The rumen and its importance in ruminant nutrition
Feed-deconstruction reactor
Effect on milk composition (and production) – CLA
– Polyphenolic compounds
– Vitamins
– Mycotoxins
Milk metabolites as biomarkers of rumen function – Odd- and branched-chain fatty acids (OBCFA)
Final remarks
Ruminant production
Capacity to use forage feeds by ruminants
The ruminant super-organism
Gastrointestinal symbionts
PROVIDE
Energy VFA can provide up to 80% of the energy needs
Protein microbes convert NPN into high quality protein
Vitamins synthesis of B-complex and K vitamins
Detoxifying functions
PRODUCE
Toxic products
Methane
Presentation plan
The rumen and its importance in ruminant nutrition
Feed-deconstruction reactor
Glucose (1 mole)
Fibre Hydrolysis
step
Starch Cellulose hemicellulose
~ 7% energy losses
Feed carbohydrates
Pyruvate (2 moles)
(+ 4H)
Butyrate (1 mole)
Propionate (2 moles)
Acetate (2 moles)
CH4 (2 moles)
CO2 (2 moles)
CO2 (1 mole) (+ 4H)
(- 8H) (+ 0H)
(- 8H) Fermentation step
~70% energy
Jouany & Morgavi, 2007
Nitrogen metabolism in the rumen
Protein N
Polypeptides
Di- and Tripeptides
proteases
peptidases
Amino acids
peptidases
Undegraded dietary protein
Non-protein N
N recycling
NH3
absorption
Urinary N
deaminases
Microbial protein
Jouany & Morgavi, 2007
2-20 kg DM 20-80 L water 50-150 L saliva
VFA 2.0-6.0 kg 0.75-2.0 kg microbial cells
140-600 L CH4
260-560 L CO2
Presentation plan
The rumen and its importance in ruminant nutrition
Feed-deconstruction reactor
Effect on milk composition (and production)
Rumen effect on milk
Milk yield
Milk composition
Feed – microbiota interaction
Interest in manipulating composition
– Nutritional value for human consumption
– Essential micronutrients and bioactive compounds
– Processing and transformation
1970 1980 1990 2000 2010
decrease saturated C14 C16
increase ratio n-3/n-6
decrease saturated FA
increase polyunsaturated FA
increase CLA
decrease some C18:1 trans
decrease C18:1 trans
Manipulation of milk FA to increase nutritional value
Mechanisms of lipolysis and biohydrogenation
Lipolysis
End products → free fatty acids
No mono- and diglycerides
Lipases: mainly of bacterial origin
– Anaerovibrio lipolytica: triglycerides
– Butyrivibrio sp: phospholipids and galactolipids (Lourenço et al, 2010)
Plant lipases
Triglycerides Phospholipids
Glycolipids
Microbes
RUMEN
SMALL
INTESTINE
Absorption Fatty acids
Undigested fatty acids
Hydrogenated fatty acids
Fatty acids
Lipolysis
Hydrogenation
Synthesis
Uptake
Bauchart et al., 1990
Triglycerides (TG) lipolysis in the rumen (%), in vivo experiment
Control 10% rapeseed 10% tallow 85 97 97
Lipolysis
Rapid
almost complete even if large amounts of lipids are fed
Mechanisms of lipolysis and biohydrogenation
Lipolysis
End products → free fatty acids
No mono- and diglycerides
Lipases: mainly of bacterial origin
– Anaerovibrio lipolytica: triglycerides
– Butyrivibrio sp: phospholipids and galactolipids (Lourenço et al, 2010)
Plant lipases
Biohydrogenation
The process which leads to saturation of unsaturated FA
Succession of isomerizations and saturations
Linoleic acid cis-9 cis-12 C18:2
CLA cis-9 trans-11 C18:2
Vaccenic acid trans-11 C18:1
Stearic acid
C18:0
RUMEN
Isomerase
Reductase
Reductase
c9c12-18:2
c9t11
t11-18:1
18:0
t10-18:1
10,12 CLA
c9c12c15-18:3
c9t11c15-18:3
t11c15 11,13 CLA other
9,11 CLA c9t13 8,10 CLA
t13+14-18:1 t15+c15-18:1 c12-18:1 t4-t9-18:1
other 9,12 18:2
(Chilliard et al, 2007)
Interconversion between all 18:1 isomers
From oleic acid (Mosley et al., 2002) and vaccenic acid (Laverroux et al., 2011)
(Doreau and Laverroux, unpublished)
18:2 t11c15
0 2 4 6
0 5 10 15 20 time (h)
%
18:1 t10 + t11
10
15
20
0 5 10 15 20 time (h)
%
18:3 n - 3
0
2
4
0 5 10 15 20 time (h)
%
Rumen hydrogenation
is rapid
Concentration of linolenic acid and hydrogenation derivatives in the rumen
0
20
40
60
80
100
0 50 100 150
FA intake (g/kg DMI)
Linoleic acid
(n=135)
Linolenic acid
%
75%
83%
Doreau and Ueda, unpubl.
Ruminal biohydrogenation
Linseed Sunflower Fish
0
2
4
6
8
c9,t11 c9,c11 t10,c12 t11,t13 t,t Total
Less CLA for fish oil
trans 10 cis 12 produced with sunflower
CLA isomers produced in cow’s rumen
0
50
100
150
200
250
300
trans10 trans11 trans13+14 Total
Linseed Sunflower Fish
Less trans 10 and more trans 13+14 with linseeds
Loor et al, 2005
trans-18:1 isomers produced in cow’s rumen
Linoleic acid cis-9 cis-12 C18:2
CLA cis-9 trans-11 C18:2
Vaccenic acid trans-11 C18:1
Stearic acid
C18:0
RUMEN
Isomerase
Reductase
Reductase
UDDER
9- desaturase
9- desaturase
Oleic acid cis-9 C18:1
Linoleic acid cis-9 cis-12 C18:2
CLA cis-9 trans-11 C18:2
Vaccenic acid trans-11 C18:1
Stearic acid
C18:0
MILK
0
2
% total FA
CLA c9 t 11
RUMEN OUTFLOW
hay
Concentrate + linseed oil
0
10
18:1 t 11
% total FA
Loor et al., 2005
- Post-absorptive origin of CLA - CLA and vaccenic acid strongly correlated
CLA-trans 18:1 relationship
Fatty acids characteristics in ruminant GIT
Biohydrogenation of PUFA is extensive
Presence of 18:2 and 18:1 cis and trans isomers in duodenal flow (≠ monogastrics)
composition in trans 18:1 FA is diet dependent
t 10 18:1 concentrate & PUFA in diet
duodenal t11 18:1 → milk c9 t11 CLA
Polyphenol classification Simple phenols
phenol
pyrocatechol
o-cresol
Phenolic acids
cinnamic acid
benzoic acid
ferulic acid
gallic acid
Stilbenes and Lignans
Resveratrol
Secoisolariciresinol
Flavonoids
Kaempferol
Flavonols
Apigenine
Flavones
Naringenine
Flavanones
Cyanidine
Anthocyanidines
Daidzeine
Isoflavones Flavanols
Epicatechine
Phenolics, secondary metabolites in vegetals
Condensed Tannins Hydrolyzable Tannins
ose + phenols
Polymers
Proanthocyanidins (flavanol polymers)
Ferulic ac.
Caffeic ac.
Phloretic ac.
3-phenyl-propionic ac.
(50-80 % of rumen aromatics)
3-Hydroxy-phenyl
propionic ac.
4-ethyl-phénol
4-Vinyl-phenol
4-Vinyl-guaiacol
Coumaryl-esters
r-coumaric ac.
Rumen
4-hydroxy-benzoic ac.
r-Cresol
Phenylacetic ac. (13-50 % of rumen
aromatics)
Benzoic ac. (< 2 % of rumen
aromatics )
Homovanillic ac.
Vanillic ac.
Protocatechuic ac.
catechol
Phenol
Cinnamic ac. (0-7 % of rumen
aromatics)
Besle et al., 1995
Ferruloyl-esters
Phenolic acids from plant cell wall
What we know about phenolics
behavior in the rumen
Phytoestrogens
Dickinson et al, 1988
equol
daidzein
formononetin
Milk
What we know about phenolics
behavior in the rumen
Bioconversion of isoflavones from clover
Equol up to ~300 g/L Daidzein up to ~5 g/L
Phenolics in milk : a tool for feed traceability ?
Pattern of UV-visible components of milk is characteristic of diet
20 40 60 80 100 0
Natural mountain prairie
Natural mountain hay (87 % DMI)
Ray grass hay (90% DMI)
Ray grass silage (86 % DMI)
Maize silage (86 % DMI)
Concentrate + hay (66 % and 34 % DMI)
Besle et al., 2010
Phytoestrogens
Milk
Main origins of cow milk vitamins Ration
Forages
Concentrates
MVC
Vitamins A, E, D3
Vitamins E, D3, K1
(Vitamins A and E)
beta-carotene
Vitamins C, B1 to B9
Vitamins C, B1 to B9
Rumen
Vitamin K2
Vitamins B1 to B12
Intestine
beta-carotene →Vitamin A
Liver
Vitamin C
Vitamin B3
Vitamin D3
Skin
Udder
2000
2400
2800
3200
3600
4000
pg/g
Grass Mountain
Grass Lowland
Maize Mountain
Maize Lowland
No variation significant according to the period of the year
Highest vitamin B12 concentrations in milk from cows in Maize Lowland production system (exhibit the highest % of maize in the diet)
System Period Syst X Period
** NS NS
Vitamin B12 content in cow milk varies according to diet
Chassaing et al., 2011
Vitamin B9 content in cow milk varies also according to diet
Grass Mountain
Grass Lowland
Maize Mountain
Maize Lowland
Variations according to the period, the system, and interaction
System Period Syst X Period
** *** ***
Chassainget al., 2011
80
90
100
110
120
ng/g
Highest in grass-based system
Lowest in May Hay % ?
Ruminal bioconversion and carry over of mycotoxins to milk
Mycotoxin Main product of
rumen metabolism Reduction of
biological potency Estimated carry-over
rates
Aflatoxin B1 aflatoxicol minor n.d.b 0–12.4 μg l−1
aflatoxin M1 minor 2.0–6.2%
Cyclopiazonic acid unchanged unchanged n.d. 6.4–0.7 μg l−1
Fumonisin B1 unchanged unchanged 0–0.05%
Ochratoxin A ochratoxin-α significant n.d.
T-2 toxin various significant 0.05–2%
DON (and related trichothecenes)
de-epoxy-DON (DOM) significant DON: 0.0001–0.0002 DOM: 0.0004–0.0024
Zearalenone α-zearalenol none 0.06–0.08%
Patulin unchanged unchanged n.d.
Ergovalin unchanged unchanged n.d.
Lolitrem unchanged unchanged n.d.
Fink-Gremmels, 2008; Yannikouris & Jouany, 2002
Ruminal bioconversion and carry over of mycotoxins to milk
Mycotoxin Main product of
rumen metabolism Reduction of
biological potency Estimated carry-over
rates
Aflatoxin B1 aflatoxicol minor n.d. 0–12.4 μg l−1
aflatoxin M1 minor 2.0–6.2%
Cyclopiazonic acid unchanged unchanged n.d. 6.4–0.7 μg l−1
Fumonisin B1 unchanged unchanged 0–0.05%
Ochratoxin A ochratoxin-α significant 0.02-0.04%
T-2 toxin various significant 0.05–2%
DON (and related trichothecenes)
de-epoxy-DON (DOM) significant DON: 0.0001–0.0002 DOM: 0.0004–0.0024
Zearalenone α-zearalenol none 0.06–0.08%
Patulini unchanged unchanged n.d.
Ergovalin unchanged unchanged n.d.
Lolitrem unchanged unchanged n.d.
Modif. from Fink-Gremmels, 2008; Yannikouris & Jouany, 2002
Presentation plan
The rumen and its importance in ruminant nutrition
Feed-deconstruction reactor
Effect on milk composition (and production) – CLA
– Polyphenolic compounds
– Vitamins
– Mycotoxins
Milk metabolites as biomarkers of rumen function – Odd- and branched-chain fatty acids (OBCFA)
Volatile FA
De novo FA synthesis
Saturated FA including odd- and branched-chain FA
Monounsaturated FA
Microbial FA synthetases
FA synthesis in bacteria
15:0 17:0
Vlaeminck et al., 2006
Straight chain FA are synthesized from propionate or valerate
CH3 – CH – (CH2)11 - COOH
CH3
CH3 – CH – (CH2)11 - COOH
CH3
CH3 – CH2 – CH – (CH2)10 - COOH
CH3
CH3 – CH2 – CH – (CH2)10 - COOH
CH3
iso-13:0 iso-14:0 iso-15:0 iso-16:0 Iso-17:0 anteiso 15:0 anteiso 17:0
Branched-chain FA are synthesized from branched-chain amino acids and iso-VFA
Odd and branched-chain FA
Fievez et al., 2012
Odd and branched-chain FA
Relationship concentration rumen milk
Rumen fermentation
– VFA (and methane)
iso-14:0 (+) acetate, methane (-) 15:0
iso-15:0 (-) propionate (+) 17:0
Microbial protein flow
Acidosis
(+) 17:0; 17:1 c9 (-) iso-14:0
Presentation plan
The rumen and its importance in ruminant nutrition
Feed-deconstruction reactor
Effect on milk composition (and production) – CLA
– Polyphenolic compounds
– Vitamins
– Mycotoxins
Milk metabolites as biomarkers of rumen function – Odd- and branched-chain fatty acids (OBCFA)
Final remarks
Milk yield
Milk composition
The ruminant super-organism
Is there a need to know more about the rumen and its microbiota?
Nutritional value for human consumption
Environmental footprint
Meet challenges ruminant production YES
Production as usual NO
Microbiome characterization
Catalogue
microbes
genes
Interactions
Change
Time
Space Who they are?
What they do?
Genetically homogeneous animals can have different phenotypes because of differences in their microbiome
Respond differently to diet, drugs and feeding practices
Nicholson et al., 2005
Homogeneous microbiome Heterogeneous microbiome Different microbiomes
Thank you for your attention
INRA – F. Glasser
– A. Ferlay
– Y. Chilliard
J. Loor U. Illinois, USA
K. Ueda U. Sapporo, Japan
