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Retention time of feed particles and liquids in thestomachs and intestines of dairy cows. Direct

measurement and calculations based on faecal collectionM Mambrini, Jl Peyraud

To cite this version:M Mambrini, Jl Peyraud. Retention time of feed particles and liquids in the stomachs and intestines ofdairy cows. Direct measurement and calculations based on faecal collection. Reproduction NutritionDevelopment, EDP Sciences, 1997, 37 (4), pp.427-442. �hal-00900041�

Original article

Retention time of feed particles and liquidsin the stomachs and intestines of dairy cows.

Direct measurement and calculationsbased on faecal collection

M Mambrini JL Peyraud

Station de recherches sur la vache laitière, Inra, 35590 Saint-Gilles, France

(Received 24 April 1996; accepted 2 June 1997)

Summary ― To validate a method for analysing indigestible marker excretion patterns in terms ofdigesta passage, the mean retention time (MRT) of long hay, ground hay and concentrate, marked,respectively, with thulium, ytterbium and dysprosium was measured in the total digestive tract(TMRT) and in the stomachs (SMRT) of four cows fed on a diet of hay in the long form (17.7 kgDM/day). The MRT of the particulate and liquid phases in the intestines was obtained after faecal par-ticles labelled by Europium and Chromium-EDTA were pulse dosed through the duodenal cannula.Following test meals, total faecal collection and spot sampling of duodenal digesta were performedat fixed intervals. TMRT were 51.7, 45.6, 40.6 h and SMRT were 39.5, 31.9 and 28.0 h, respec-tively, for hay, ground hay and concentrate. The MRT of the liquids in the rumen (8.7 h) was shorterthan the SMRT of particles but there was no differential passage between liquids and particles afterthe duodenum. Intestinal MRT averaged 11 h and was partitioned into 7.5 and 3.5, respectively, forMRT in the tubular sections and the caecum-proximal colon. The compartmental analysis of thefaecal patterns of markers given during a test meal gives the following results. The time associatedwith the descending part of faecal kinetics (respectively, 25.3 and 22.9 h for hay and concentrate) isdirectly related to the escape of feed particles from the rumen. The delay of first appearance of mark-ers mostly reflects transit in the post duodenal tubular sections for the concentrate. The time associ-ated with the ascending part (respectively, 16.9 and 9.4 h for hay and concentrate) represents thetime required to reduce the size of the forage particles (7 h according to the difference betweenTMRT of long and ground hay direct measurements) and caecal mixing (3.5 h) as well as othercompartments or processes that are not clearly identified.

dairy cow / transit / particle / liquid / rumen / intestine

* Correspondence and reprintsTel (33) 02 99 28 51 00; fax (33) 02 99 28 51 O1; e-mail: peyraud@st-gilles.rennes.inra.fr

Résumé ― Temps de séjour des particules d’aliment et des liquides dans les estomacs et l’intestindes vaches laitières. Mesures directes et calcul à partir des cinétiques d’excrétion fécale desmarqueurs. Pour estimer à partir des cinétiques d’excrétion fécale des marqueurs le transit dans lesprincipaux compartiments digestifs, le temps de séjour moyen (MRT) de particules de foin long, defoin broyé et de concentré, marquées respectivement avec du thulium, de l’ytterbium et du dysprosium,a été mesuré dans l’ensemble du tube digestif (TMRT) et dans les estomacs (SMRT) sur quatrevaches laitières alimentées avec un régime à base de foin non haché (17,7 kg MS/j). Le MRT dans lesintestins (IMRT) des particules et des liquides a été mesuré à partir de l’excrétion fécale de particulesde fèces marquées avec de l’europium et du chrome-EDTA introduits par la canule du duodénum. Aprèsles repas test, les fèces ont été collectées en totalité et des prélèvements de contenus duodénaux ontété effectués à intervalles fixés. TMRT a été de 51,7 ; 45,6 ; 40,6 h et SMRT a été de 39,5 ; 31,9 ; 28,0 hrespectivement pour le foin, le foin broyé et le concentré. Le transit des liquides dans le rumen(8,7 h) a été beaucoup plus court que celui des particules mais il n’y a pas eu de passage différentieldes liquides et des particules dans les intestins. Le MRT dans les intestins a été de 11 h qui ont été répar-tis en 7,5 h de transit dans les compartiments tubulaires et 3,5 h dans le caecum-colon proximal. Ladurée associée à la phase décroissante des cinétiques d’excrétion fécale (25,3 et 22,9 h pour le foin etle concentré) traduit assez fidèlement la sortie des particules d’aliment du rumen. Le délai d’appari-tion des marqueurs (9,5 et 8,3 h pour le foin et le concentré) reflète principalement le transit dans lesportions tubulaires situées après le duodénum. Le temps associé à la phase croissante (16,9 et 9,4 hpour le foin et le concentré) représente le temps requis pour réduire la taille des particules de foin long(estimé à 7,0 h par la différence entre les TMRT des foins entier et broyé), le transit dans le caecum-colon proximal et d’autres phénomènes qui n’ont pu être identifiés clairement.

vache laitière / transit / particule / liquide / rumen / intestin

INTRODUCTION

In ruminants, the extent to which dietarycomponents are fermented in the reticulo-rumen is dependent on the mean retentiontime (MRT) of feed particles. Feed intakeand MRT are also closely related. MRT indifferent sections of the gastro intestinal(GI) tract can be estimated after killing ani-mals (O’Connor et al, 1984) but is moreoften derived from a single dose of a markerfollowed by time sequence samplings(Faichney, 1975a). Because of the difficul-ties in obtaining representative samples forparticulate markers in the rumen, there havebeen many attempts to estimate the rumi-nal MRT of feed from the mathematical

analysis of marker excretion in faeces. TheGI tract has been assigned to two (Grovumand Williams, 1973) or sometimes three(Faichney and Griffiths, 1978; Milne et al,1978) time-independent compartments sep-

arated by tubular segments. Dhanoa et al( 1985) proposed to describe the digesta flowalong the GI tract by a multi compartmentalprocess with two major compartments and aseries of segmental mixing compartmentseach having a very low MRT. A family ofmodels using gamma age dependency in thefirst compartment was also proposed (Pondet al, 1988; Quiroz et al, 1988) because itgenerally improved the quality of curve fit-ting (Quiroz et al, 1988).

However, difficulties have sometimesbeen encountered in fitting the compart-mental models (Ellis et al, 1979; Milne etal, 1978; Dhanoa et al, 1985; Mambrini,1990; Lall!s et al, 1991 All these modelsdiffer by the way they fit the ascending partof the curve, which is based only on a fewpoints. From a study of more than 60 pat-terns obtained in dairy cows, Mambrini(1990) showed that these curves are very

sensitive to the initial data and inferred thatno confidence could be placed in the esti-mations of the lower MRT because it variedwith the different models. The subsequentdifficulty is to attribute a biological signif-icance to the partial MRT obtained frommodelling. Despite the number of compre-hensive studies comparing partial MRT gen-erated by different models (Lalles et al,1991; Huhtanen and Kukkonen, 1995) thereare surprisingly few meaningful commentsupon the biological significance of the com-partments derived from mathematical anal-ysis. In the study of Grovum and Williams(1973) the MRT in the rumen was alwayslonger than the MRT in the caecum-proxi-mal colon (hereafter called caecum) and ithas become common practice to assume thatthe longer MRT represents rumen outflow.However in some circumstances, rumenMRT can be shorter than MRT in caecum

(Faichney and Boston, 1983). In their model,Pond et al (1988) considered that the reduc-tion of particle size within and outflowingfrom the rumen was the major factor butthey did not make reference to the anatom-ical compartments. Finally, all these modelsassume that the flow of digesta is continuousthroughout the GI tract, which is obviouslynot correct since defecation occurs at dis-crete times.

The present study was designed: 1 ) togain knowledge concerning the MRT offeed particles and liquids in the whole GItract or an anatomical section of it and onthe MRT required for forage particle sizereduction (Poppi et al, 1981); 2) to test asimple mathematical treatment of the fae-cal excretion curve that would be less sen-sitive to the initial data and to the disconti-

nuity of defecation than the models currentlyused; 3) to attribute physiological signifi-cance to the two compartments and to the

delay generated by calculations. Prelimi-nary results of this work have been pub-lished (Peyraud and Mambrini, 1992).

MATERIALS AND METHODS

Animals and feeding

Four Holstein cows (625 ± 30 kg body weight,20 ± 4 kg/day fat corrected milk, 36 weeks oflactation) with a ruminal and T piece duodenalcannula were housed in metabolic crates andmilked twice daily. They were fed a diet con-sisting of (in percent of total DM) 55 perennialrye grass hay (not chopped), 42 concentrate, 1

soya bean meal, 1.6 mineral mix, 0.4 urea adlibitum. The animals were allowed 10% refusals.The ingredients of the concentrate, chemicalcomposition and nutritive values of feeds aregiven in table I. The cows were fed two equalsmeals daily (8 h 00, 17 h 00) and they had freeaccess to water and to block salt.

Principles and organisationof measurements

The MRT within the GI tract (TMRT) and thestomachs (SMRT, reticulo-rumen, omasum andabomasum) of foodstuffs (concentrate and hay)were determined by following the excretion pat-terns of markers in the faeces and duodenumafter the cows were given a test meal. The MRTof the liquids and particles after the duodenumwere measured by introducing a solute markerand labelled faecal particles through the duode-nal cannula and following their faecal excretion.The MRT required for hay particle size reduc-tion was calculated by the difference betweenthe MRT of hay as fed (mean particlesize > 100 mm) and hay ground through a 2 mmscreen (mean particle size = 0.6 mm).

The experimental period consisted of 14 daysof adaptation to the diet and metabolic crates,followed by a 9 day collection phase. To evalu-ate the differential passage rates simultaneouslyon the same animal, six different markers wereused. Hays, concentrate and faecal particles werelabelled with rare earth metals. Fed hay wasmarked with thulium (Tm-H) or with ytterbiumbefore grinding (Yb-GH), concentrate waslabelled with dysprosium (Dy-C) and faecal par-ticles were labelled with europium (Eu-F).Chromium-EDTA (Cr-EDTA) was used for mea-suring the intestinal transit of liquids, andpolyethylene glycol (PEG) was used to estimatethe ruminal fractional outflow rate of liquids(FOR). The choice of rare earth elements was

justified because the MRT of feed particles isnot influenced by the nature of the rare earth ele-ment used (Goetsch and Galyean, 1983; Mooreet al, 1992) and also because non-radioactiveDy, Eu, Yb and Tm are easily determined byatomic absorption.

Marker preparations

Rare earth markers were bound to particles fol-lowing the competitive binding technique adaptedfrom Ellis and Beever (1984) and Poncet (perscomm). Feedstuffs and faecal particles wereboiled (80 °C for 1 h) in a commercial detergentwithout EDTA (Ergamatic AC, Societe

Chimiotechnique, Lyon, France) to remove thecell soluble. The cell wall residues were soakedat room temperature in a solution containing therare earth element (20 g/kg DM) together withcitric acid as a competitive ligand, and sufficientwater to cover the material. pH was brought to2.2-2.5 using 2N HCI. After 24 h of soaking withoccasional stirring, the labelled material wasstrained through cheesecloth and carefully rinsedwith tap water to remove any unbound markerand dried at 60 °C for 48 h. Faecal particles wereobtained by sampling the faeces excreted overa 24 h period from the four cows on day 10. Thefaeces were mixed, dried and labelled witheuropium in the same way as the feed particles.The rare earth concentrations in the labelled par-

ticles were 7.0, 7.0, I 1.2, 4.5 g/kg DM, respec-tively, for Tm-H, Yb-GH, Dy-Co and Eu-F.

Cr-EDTA solution was prepared accordingto Binnert et al ( 1968) and diluted, so that thefinal concentration was 1.5 g Cr/L. The solutionof PEG was obtained by dissolving 25 g PEG4000 per kg DM intake in 1 L of warm water.

Marker administration, duodenaldigesta and faeces sampling

On day 15 at 8 h 00, 500 g Tm-H and 300 g Dy-C were offered to each cow. Thirty minutes wereallowed for eating the marked feeds then, anyuneaten labelled feed was introduced into therumen after being soaked in warm (38 °C) arti-ficial saliva. The cows were then allowed to eatthe usual morning meal. At 10 h 00, 300 g Yb-GH were introduced into the middorsal to mid-ventral regions of the rumen. At 1 h h 00, 50 gof Eu-F and 250 mL of Cr-EDTA were intro-duced through the duodenum cannula after beingmixed in some heated duodenal contents (39 °C)that had been previously sampled on the sameanimals on day 10 and stored at 4 °C. On day 21 1at 8 h 00, PEG was pulse-dosed into the rumen(ventral sac).

The total faecal output was collected 22 timesover the 178 h post-dosing. The sampling timeswere 6, 10, 12, 16, 20, 24, 28, 32, 36, 48, 54,

60 h post-dosing and then every 12 h, the mid-point of the meal being considered as time 0. Ateach sampling time, faeces were weighed, thor-oughly mixed and approximately 500 g weredried (80 °C for 48 h). Faecal samples wereground through a 0.8 mm screen and analysedfor Tm, Yb, Eu, Dy and Cr. Spot samples of duo-denal digesta were collected over 144 h after thetest meal at 1.5 h intervals (from 0 to 20 h post-dosing), 3 h intervals (from 20 to 43 h post-dosing), 6 h intervals (from 43 to 67 h) and 12 h hintervals thereafter. They were freeze dried andground through a 0.8 mm screen before analy-sis of Tm, Yb and Dy. Rumen fluid was sampledat 10 h 00, 10 h 30 and 11 h 00 on days 21 and 22,strained through six layers of cheesecloth andstored at 4 °C until PEG analysis (on day 23).

Chemical analysis

Concentrations of PEG were analysed usingHyden’s turbidimetric method (1955) as modi-fied by Malawer and Powell (1967). Rare earthmetals and chromium were determined by atomicabsorption spectrophotometry (Varian, AA-20)using a nitrous oxide/acetylene flame. Chromiumwas extracted according to the method describedby Siddons et al (1985). For rare earth analysis,the samples were ashed (550 °C for 6 h) anddigested in a solution containing 2% nitric acidand 2 g/L of potassium chloride. Wavelengthschosen were 357.8, 371.8, 398.8, 421.2,459.4 nm, respectively, for chromium, Tm, Yb,Dy and Eu. Previous studies had indicated thatthere was no interaction when the four rare earthmetals and the chromium were present together(Mambrini, 1990).

Calculations

FOR was estimated as the slope of the naturallogarithm regression of PEG concentration vstime post-dosing. The total MRT within the entireGI tract (TMRT) of fed hay, ground hay and con-centrate and intestinal transit time (IMRT) offaecal particles and liquids were obtained, respec-tively, from the quantities of markers Tm-H, Yb-GH, Dy-C, Eu-F, Cr-EDTA excreted in faeces,using the equation:

where ti is the time elapsed between dosing andthe mid point of each interval i during which fae-ces were collected, and mi is the quantity ofmarker excreted during the ith interval. Thismethod, based on the quantitative recovery ofmarkers does not make any assumptions regard-ing the pattern of excretion.SMRT of fed hay, ground hay and concen-

trate were calculated using the equation:

where ci is the concentration of the marker in theduodenal digesta in the ith sample collected atthe time ti and dti is the time elapsed betweentwo successive sampling times. This methodassumes that the flow of digesta is continuous.The intestinal transit time for fed hay, groundhay and concentrate was calculated by subtract-ing SMRT from TMRT.

Compartmental analysis of faecal and duo-denal passage curves was performed by a simplegraphical method derived from that proposed byUden (1984). Natural logarithm of the markerconcentration was plotted against time. The con-centration curves were then divided up in a

descending part (compartment of greater resi-dence time, MRT1), an ascending part (corre-sponding to a second mixing compartment withMRT2 as mean retention time) and a delay (TT).For faecal kinetics, the delay (TTfec) was deter-mined by the time elapsed between dosing andthe mid point of the interval when the markerappeared for the first time. MRTlfec was calcu-lated as the reverse of the slope of the descend-ing part of the curve. MRT2r corresponding tothe ascending part of the curve was obtained bysubtracting MRTlfec and TTfec from TMRT.Similar calculations were performed for the duo-denal sampling site (MRTlduo, TTauo and

MRT2doo). ’

Statistical analysisTotal and partial retention times obtained fromthe duodenal and faecal excretions of Tm-H, Yb-GH and Dy-C were analysed according to a split-plot design using the following model: Y.-k =

Fi + C. + Sk + SkFl + SkCi + F!g +eijk where = =feed effect (i = 1 to 3); C. = cow effect (j = 1 to 4);Sk = sampling site effect (k = 1 to 2); SkF¡ and

SkCi = interaction site x feed and site x cow,

respectively, F!C! = interaction feed x cow and

e = error term with 6° of freedom. Feed and coweffects were tested with Fi x G as an error term.

Data obtained from the faecal excretion ofmarkers introduced into the duodenum were anal-

ysed according to the following equation: Y =

P. + C. + e.. where Pi = labelled phase (particle orliquid); Ci = cow effect and e!i = error term with3° of freedom.

Retention times of Tm-H, Yb-GH, Dy-Cwithin the intestine were compared to the IMRTof Eu-F and Cr-EDTA according to the follow-ing model: Y iik = S! + M ’.(S) + Ck + Sick + e¡jkwhere Si = site of dosing (i = I to 2);M = marker/feed effect (i = 1 to 5), Ck = cow

effect (k = 1 to 4); and e! = error term with 9° offreedom. Site effect was tested with Mi as an

error term. When the effect was significant,means were compared by the Newman andKeul’s test at the 0.05 significance level. All anal-yses were performed using the GLM procedure

of SAS (1987). Correlation was tested with theCORR procedure of SAS (1987).

RESULTS

Direct measurements

SMRT was always shorter (P < 0.001;table II) but highly correlated (r = 0.98;n = 12) with TMRT. SMRT accounted for76 and 70°70 of TMRT, respectively, for fedhay and concentrate. No interaction betweenfeed and site of sampling was detected.TMRT and SMRT of marked feed particlesdecreased in the order Tm-H > Yb-GH > Dy-C (P < 0.01; table II). In particu-lar, the difference between fed hay andground hay was similar for TMRT and

SMRT (6.1 vs 7.6; P > 0.10). These twoestimates of time for particle size reductionwere highly correlated (r = 0.94, n = 4) andthe slope of the relation did not differ from1 (P > 0.10). Estimated MRT within theintestines (TMRT-SMRT) did not differ(P > 0.10) among the three feeds (table III)and the mean value was 12.9 h. The IMRTof Eu-F and Cr-EDTA were similar

(table III) and averaged 10.5 h. This valuewas shorter (P < 0.03) than the time calcu-lated by the difference between TMRT andSMRT. FOR was 11.5%/h so that the liq-uid retention time in the rumen was 8.7 h, avalue which is much lower than the SMRTof the feeds. Variance analysis showed sig-nificant individual variations (sd of coweffect was 9.5 and 5.8 h, respectively, forTMRT and SMRT). Two cows showed amuch lower retention time (TMRT = 37.9 h,

SMRT = 25.5 h) than the other two animals(TMRT = 45.4 h, SMRT = 32. I h).

Compartmental analysis of faecaland duodenal patterns of markersgiven during the test meal

The shape of the marker concentrationcurves in the duodenal samples were not asregular as in the faecal samples (fig 1), pre-sumably due to the greater difficulty inobtaining representative samples throughthe duodenal cannula as compared to totalfaecal collection. Consequently, fitting thedescending part of the excretion curves ofTm-H, Yb-GH and Dy-C to an exponentialmodel explained 99% of the variance forfaecal curves with only one value lower than98% but it was less than 98% on five occa-sions for the duodenal curves. MRT1 gen-erated from the analysis of faecal markerconcentration was lower but highly corre-lated with estimates based on calculations

performed from the quantities of markersexcreted (24.3 vs 26.3 h; r = 0.91, n = 12,P < 0.05). The differences reached 4 h inthree kinetics but were lower than 1 h forthe other kinetics.

The MRT associated with the descendingpart of the curves (MRT1) were not affectedby ,the sampling site (P > 0.10; table II).MRT 1 fe! and MRT 1 duo were highly corre-lated (table V) but the slope of the relationwas significantly different from 1. Themarkers appeared in the faeces during thethird or the fourth sampling interval, whilein the duodenal contents, they appeared assoon as the first sample (fig 1 ) in 10 out ofthe 12 excretion kinetics. It was thus not

possible to estimate TTduo and MRT2d!oprecisely although the sum TTduo +

MRT2d!o, obtained by substractingMRTlduo from SMRT, was not biased. Insuch cases, TTd!o was determined as halfthe time elapsed between dosing and thefirst sampling time. MRT2duo was signifi-

cantly shorter (P < 0.001 ) when calculatedfrom duodenal rather than from faecal sam-

pling (P < 0.001). TTduo was on average7.5 h shorter than TTfc and MRT2duo was

4.4 h shorter than MRT2fec (table II).

There was no site x feed interaction forthe estimation of MRT1, MRT2 and TT.

MRTIfe! did not differ between Tm-H andYb-GH, and was slightly longer than forDy-C. MRT 1 d!o was slightly higher for Tm-H compared to Yb-GH and Dy-C. The delaywas not affected by the nature of the feed(P > 0.10; table II). On the contrary MRT2was markedly affected by the feeds, theselatter being in the order long hay > groundhay > concentrate (P < 0.001). In particu-lar, the differences in total transit timebetween fed and ground hays were mostlyattributable to MRT2 in the whole GI tract

(TMRT) as well as in the stomachs (SMRT).

The total time spent in the two compart-ments generated from the analysis of fae-cal excretion curves (TMRTI + TMRT2)for Tm-H, Yb-GH and Dy-C was 3.3 h hlonger but highly correlated with SMRT(table V). The slope of the relationship didnot differ from 1 (P > 0.10).

Compartmental analysis of faecalpattern of markers introducedinto the duodenum

A fairly similar pattern of Eu-F and Cr-EDTA excretions was observed (fig 2,table IV). After a delay, the marker con-centration reached a peak between two suc-cessive defecations and decreased there-after. However there were small differencesbetween Eu-F and Cr-EDTA in MRT1 andTT which tended to be significant. The delay

represented two thirds of IMRT. MRT1 Iaveraged 3.2 h. MRT2 was always veryshort and was most often not significantlydifferent from zero. Negative values forMRT2 were obtained on two occasions forCr-EDTA and this is biologically unrealis-tic. This compartment may thus be ignored.

DISCUSSION

Choice of markers

The migration of markers from labelled par-ticles was assumed to be of little effect andthe different rare earth elements wereassumed to behave similarly in interpretingdata on particle passage. Labelling tech-niques and choice of markers are crucial inestimating the MRT of particulate matter(Coleman et al, 1984; Faichney et al, 1989).Rare earth elements have been criticised

because they may migrate from the labelledfeed particles (Combs et al, 1992). In thisstudy, particles were labelled using a com-petitive procedure (Ellis and Beever, 1984).It could therefore be expected that markerswere selectively bound to a site on the feedparticles having an association constantgreater than that of citric acid (pK > 8.1).Because the dissociation of bound rare earthelements may also occur at acidic segmentssuch as the abomasum, the pH was broughtto a value lower than 2.5 when labelling thefeeds. These approaches for more tenaciousbinding of marker on feed particles wouldhave minimised the eventual migration ofthe marker.

The choice of rare earth elements wasalso justified by their specific properties.Goetsch and Galyean (1983) compared Dyand Yb and showed that the feeds MRTwere not influenced by the nature of the rareearth element used. Similarly, Moore et al

(1992) reported that Dy, erbium and Yb hadthe same excretion pattern in sheep whenthey were labelling either hay or concen-trate. Therefore it can be concluded that Yband Dy were useful for comparing the MRTof concentrate and ground hay in our study.Moreover, Tm and Yb were chosen forlabelling fed hay and ground hay, respec-tively, because these elements occupy relatedpositions in the periodic table. They wereassumed to exhibit similar affinities for haycell walls as was the case for EDTA andmetabolite molecules in the rumen (Allenand van Soest, 1984). As expected, rare earthelement concentrations in labelled long andground hay were identical (7.0 g/kg DM).Finally, because faecal particles have to beintroduced into the duodenum with Cr-

EDTA, Eu was chosen for labelling faecalparticles owing to its much lower associationconstant (pK = 17.4) with EDTA comparedto chromium and the three other rare earthelements (pK = 24.0, 19.5, 19.3 and 18.3respectively for Cr, Yb, Tm and Dy).

Passage of particulate matterand liquids in the stomachs

The TMRT of particles of fed hay and con-centrate within the GI tract as well as liq-uid rumen FOR agree with data previouslyobtained on dairy cows (Hartnell and Satter,1979; Uden, 1984). The results obtained inthis experiment confirm that particulate mat-ter is retained much longer in the stomachsof dairy cows than within the intestines(O’Connor et al, 1984). The MRT in thestomachs accounted for more than two thirdsof the total MRT of the feed particles. More-over, the differential passage of fed hay,ground hay and concentrate particles in theentire GI tract was fully explained by SMRTvariations and the MRT in the intestines didnot differ among feeds. SMRT can be con-sidered as being mainly located in the retic-ulo-rumen. Actually particulate matterTMRT in abomasum and omasum lasts less

than 4 h in sheep and cattle (Faichney andGriffiths, 1978; Huhtanen and Kukkonen,1995) and might be even lower in dairycows having a higher level of intake.

The longer MRT of Tm-H compared toYb-GH can be attributed to the time neces-

sary for particle size reduction. Estimatesof the time necessary to comminute coarse

particles did not differ between the two sam-pling sites and averaged 6.9 h. These dataagree well with the mean ruminating time indairy cows (6 to 8 h, Jarrige et al, 1995),which is a process accounting for more than80% of the comminution of large particlesentering the rumen (Kennedy, 1985). TheTMRT and SMRT of Yb-GH were on aver-

age 4.5 h longer than the TMRT and SMRTof Dy-C. The critical size theory (Poppi et al,1981) divides the rumen particles into twopools: a large particle pool, which cannotpass out of the rumen and a small particlepool, which can flow out of the rumen.When estimating the time necessary toreduce particle size from the differencebetween fed hay and ground hay TMRT,we assume that ground hay particles belongto the small particle pool and that theirbehaviour is similar to that of particles aris-ing from the mastication of the long markedhay. But a further size reduction for groundhay can not be excluded. Grenet (1970) andDixon and Milligan (1985) reported thatsmall particles (< 0.6 mm) accounted forhalf of the faecal particles in steers fed withlong hay. Shaver et al (1988) demonstratedthat ground hay was ruminated by dairycows at a rate of 20 min/kg DM, ie, 5.5 hfor an intake level similar to that of our ani-mals. In addition, specific gravity has beenshown to be an important physical criterioninfluencing passage (Ehle and Stem, 1986)and differences in functional specific grav-ity between ground hay and concentrateshould not be excluded. Because of a muchfaster rate of fermentation, the density ofparticles may increase more rapidly for con-centrate than for forage particles.

The MRT of liquids in the reticulo-rumenwas four times shorter than the MRT of par-ticulate matter. The processes taking place inthe reticulo-rumen are likely to be respon-sible for this differential passage rate, as

previously observed in dairy cows (Prangeet al, 1982). The observed differencebetween liquid and particulate transit islower than the one reported for sheep (1.5 to2.5 times; Faichney, 1975b; Poncet et al,1986). Sheep spend a longer time chewingthan do cows (Carle and Dulphy, 1980) andthe amount of small particles, which absorbwater, is greater in the swallowed bolus ofsheep (Michalet-Doreau et al, 1992). Therumen contents are consequently more strat-ified in cows than in sheep (Poppi et al,1981 ), explaining the relative independenceof liquid and solid phases in the rumen ofthe cow.

Passage of particulate matterand liquids in the intestines

The MRT of liquid and faecal particles intro-duced into the duodenum were similar, indi-cating that, on the contrary to what happensin the rumen, no differential passage of liq-uid and solid digesta occurs in the intestines.This is the reverse of what happens in thereticulo-rumen. These results agree withdata previously obtained in sheep (Faich-ney and Boston, 1983), cattle (Huhtanenand Kukkonen, 1995) and cow (VanBruchem et al, 1981). The value for intesti-nal MRT are within the limits reported byO’Connor et al (1984), Prange et al (1982)and Shaver et al (1988) for dairy cows.These values appear to be lower than thoseobtained for sheep (22 h on average;Coombe and Kay, 1965; Faichney, 1975b).This could be explained either by differ-ences in the gut architecture between thetwo species (namely the caecum) or by thehigher intake level of cows. It was shownin dairy cows that intestinal MRT decreasesfrom 20 h in cows given 6.5 kg DM (Pond et

al, 1988) to 8 h when DM intake increases to23.7 kg DM per day (Poore et al, 1991).

Intestinal MRT of particles was higherwhen estimated by the difference betweentotal MRT derived from faecal and duodenal

sampling than when calculated from the fae-cal excretion pattern of markers introducedinto the duodenum. It is unlikely that thefaecal particles have a longer MRT than hayparticles because the intestinal MRT did notdiffer between Tm-H, Yb-GH and Dy-C.The origin of this bias remains unclear.Nonetheless the difference was small and

may not be biologically significant. It canbe considered that I I are spent by feedparticles in the intestines. This low MRTimplies that the digestibility of fibre afterthe duodenum would be low and could not

compensate for a decrease in ruminaldigestibility at higher levels of intake orwith diets containing high amounts of starch.

Being able to follow faecal kinetics ofmarkers introduced through the duodenalcannula allowed the MRT of the intestineto be partitioned. Particle and liquid MRT inthe intestines can be described by a delayof 7.5 h and one mixing compartment of3.2 h on average. This compartment can beconsidered as located in the caecum. Thisestimate of caecal MRT is within the limits

reported in other studies when direct mea-surements were made after killing animals (2to 5 h, Makela, 1965; Miller et al, 1967).The delay was higher than direct estimatesof MRT in the small intestine (2 to 4 h,Mdkeld, 1965 ; Miller et al, 1967). Assum-ing that the delay is also associated with thetubular segments of the hindgut, the MRT insuch segments might last for 4 h.

Interpretation of the parametersderived from faecal excretion curves

The declining phase of the marker excre-tion pattern was similar whether described atthe duodenal or at the faecal level. This indi-

cates that the preduodenal segments are atthe origin of MRT1fec’ Moreover, because

MRTlfe! did not differ between fed hay andground hay on the one hand, and is corre-lated with liquid MRT in the reticulo-rumenon the other, MRTIfec may be allocated withconfidence to the reticulo-rumen and directlyrelated to the escape of dietary particles fromit. The anatomical and physiological iden-tifications of this mixing compartmentagrees with other studies (Grovum andWilliams, 1973; Ellis et al, 1979). In somecircumstances, namely in sheep, the rumenMRT can be shorter than the caecum MRTand in such a case, the MRT1 will reflectthis latter compartment (Faichney andBoston, 1983). This does not appear to bethe case for dairy cows (Mambrini, 1990;Mambrini and Peyraud, 1994). Betweenfeeds, the difference in MRTI calculatedfrom the faecal excretion of markers maybe biased, because the slope of the rela-tionship between MRT1 fec and MRTId!o(0.73) differs from 1. Faecal analysis mayhave underestimated the longer retentiontimes and overestimated the shorter ones.

However, the difficulty in obtaining repre-sentative samples of the duodenal contents(Faichney, 1975a) may explain this bias andthe fairly good agreement between MRTlfeccalculated from the analysis of the concen-tration curves or the amounts of markerexcreted supports the use of faecal samples.

Numerous comparisons have emphasisedthe difficulty to obtain, with models, accu-rate estimations of TT and partial MRTassigned to the second mixing compartment,ie, the delay and the ascending part of thecurve (Dhanoa et al, 1985; Lalles et al,1991). In the present experiment the delay offirst appearance of markers in faeces wasdetermined experimentally, it was only sub-ject to analytical errors. TT was 7.4 h longer(on average) when estimated from faecalsampling than from duodenal sampling.These results are consistent with the reporteddata (between 7 and 10 h; Prange et al, 1982;Pond et al, 1988). The difference between

TTfec and TTduo was numerically equiva-lent and highly correlated within the fourcows with the delay of first appearance inthe faeces of markers introduced into theduodenum (r = 0.94). This shows that TTfecreflects mostly transit in the tubular seg-ments after the duodenal cannula. It mayalso represent a delay occurring before theduodenum due to a slow mixing of markedfeeds in the rumen or during transit throughomasum. In the present experiment, thenature of foodstuff did not influence TTfec,although TTfec tended to be longer for haythan for concentrate. Indeed a greater dif-ference between forages and concentratesfor TTfec has been reported (Mambrini andPeyraud, 1994). Our results indicate thatthis could not be explained by a fasterintestinal transit of concentrate particles.These differences could therefore beattributed to a supplementary mixing delayin the case of forages. Actually the lowerdensity of forage particles could have beenresponsible for a slower mixing of themarked forage in the rumen (Martz andBelyea, 1986), or in the omasum or, asalready demonstrated by Welch and Palmer(1987), for the driving back of these particlesfrom omasum to reticulum. Concentrate

TTfec seems to be more clearly assigned totransit in the tubular segments than does theforage IT fee’

Differences between Tm-H and Yb-GHin TMRT and SMRT were mainly explainedby the variations in MRT2fec’ MRT2fecprobably represents the time required toreduce the size of forage particles. Reten-tion time in the second compartment wasaffected by sampling site. MRT2 was onaverage 4.4 h longer for faecal than for duo-denal sampling and the total time spent inthe two compartments derived from faecalexcretion curve (sum MRT 1 fee + MRT2fec)was on average 3.4 h longer than SMRT.Thus, MRT2fec does not represent processesconfined to the reticulo-rumen alone butalso reflects compartmental mixing in thepost-duodenal segments. This compartment

should be the caecum because these differ-ences were fairly similar to the values ofMRTlduo’ which is assumed to be the time

spent in the caecum. Assuming a MRT of3 h for the caecal mixing compartment and7 h for forage particle size reduction, thereare still 5 to 7 h that remains unexplainedfor the partial time assigned to the secondmixing compartment after faecal curve anal-ysis. This time period can be partitionedbetween other compartments or processesfor which no data are available in this study.It could be expected that the retention timein the abomasum and omasum MRT, whichaveraged 3 to 4 h (Faichney and Griffiths,1978), must also be assigned to MRT2fec!Nonetheless because the MRT of feed par-ticles did not differ after the reticulo-rumen,it is worthwhile noting that the differencein MRT2fec between forage and concentrategives an index of the time necessary toreduce the particle size of coarse forage.

CONCLUSIONS

In dairy cows, information on retention timein the reticulo-rumen can be deduced frommeasurements of the faecal excretion curvesfor ingested markers. Calculation of totalmean retention time from the quantities ofmarkers excreted gives relevant indicationsof the reticulo-rumen retention time varia-tions because the latter largely governs vari-ations in the former. The mathematicallysimple method presented here for analysingsuch data provides answers to questions per-taining to the passage of ingesta. MRTassigned to the decreasing part of the markerconcentration pattern represents the escapeof particles from the rumen (25 and 23 h forhay and concentrate, respectively). Delayfor first appearance of the markers (9 to 10 h hand 8 h for hay and concentrate, respec-tively) depends mainly on the transit in thetubular sections after the duodenum (7.5 hon average) and to a smaller extent beforethe duodenum for concentrates. Mixing of

the marked feeds in the rumen and somesmall indistinct phenomena for forages alsotakes longer. Indications on the time for for-age particle size reduction (eg, 7 h) may bededuced from the differences between MRT

assigned to the increasing part of the excre-tion pattern of labelled forage and concen-trate particles ( 17 and 9 h for hay and con-centrate, respectively). The remaining time(9 h) represents the retention time in cae-cum (3 h) and 5 to 7 h that could representthe retention time in the abomasum andother processes taking place in the reticulo-rumen.

ACKNOWLEDGMENTS

We wish to thank AG Deswysen, C Poncet andR Verite for their helpful criticism, H. H6taultand his crew for the care of the animal and tech-nical assistance, and A Brasseur for the chemicalanalysis.

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